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fl

108 PRODUCTS OF THE DECOMPOSITION OF HAEMOGLOBIN. [BOOK I.

which can afterwards be obtained from it by distillation after acidu-
lating with sulphuric acid. It is to be noted that the spectrum
of the supposed hydrocyanic compound is identical with that of
oxy-haemoglobin, and that the behaviour of the solution to reducing
agents is absolutely the same as that of a solution of oxy-haemoglobin.

Those who advocate the existence of the compound however rely
somewhat upon the fact that blood to which hydrocyanic acid has been
added shews the bands of oxy-haemoglobin, or bands identical with
them, for a much longer time than normal blood — a fact which
they explain by supposing that the hydrocyanic compound is some-
what more stable than oxy-haemoglobin.

It appears to the Author that all proofs of the existence of such
a compound are wanting. That some hydrocyanic acid should adhere
to the haemoglobin as it crystallizes out is quite in accordance with
a variety of experiences of a similar kind, and can by itself afford
no evidence of an actual compound existing. The resistance of
hydrocyanic blood to decomposition can on the other hand be easily
explained by the unquestionable arrest or slowing *'of the process
of putrefaction in the presence of hydrocyanic acid ; it is undoubtedly
the products of putrefaction which are the causes of the spontaneous
reduction of the oxy-haemoglobin of blood confined in a vessel which
has no access to air, so that an agent which will inhibit putrefaction
and at the same time not decompose oxy-haemoglobin would be
expected to act as hydrocyanic acid acts and cause the persistence of
the oxy-haemoglobin bands.

Products of the decomposition of Haemoglobin.

When subjected to the action of various reagents, especially to that
of acids and of salts having an acid reaction, the molecule of haemo-
globin undergoes a profound decomposition, the ultimate products of
which are, amongst others, a proteid substance or substances, and a
body called HAEMATIN, which contains all the iron originally con-
tained in the blood- colouring matter. The formation of haematin
is, according to Hoppe-Seyler, necessarily dependent upon the
presence of oxygen, in the absence of which the process of decom-
position yields a proteid and a body to which he has given the name
of HAEMOCHROMOGEN ; the latter may by oxidation pass subsequently
into haematin. Haematin is an interesting body which forms definite,
well crystallized, compounds with hydrochloric, and apparently also
with hydriodic acid.

Before describing the various bodies which are the products of
a profoundly decomposing action exerted upon haemoglobin, it is
essential to refer to a modification of haemoglobin which is brought
about by the action of various agents, and concerning which very
much difference of opinion still lingers, viz. methaemoglobin.

CHAP. II.] THE BLOOD. 109

Methaemoglobin.

spectrum When a solution of haemoglobin is exposed to air

of Methaemo- for some time it loses its blood-red colour, assumes a
brownish tinge, presents an acid reaction, is precipi-
tated by solutions of basic lead acetate, and on examining its
spectrum it is found that the two bands of oxy-haemoglobin have
become faint, and that a new band has appeared in the red near C ;
this line occupies nearly though by no means exactly the position of a
similar band in the spectrum of acid haematin. On now rendering
the solution alkaline by the addition of ammonia, the band in the red
disappears, and is replaced by a faint absorption band immediately
near D.

The most remarkable phenomenon, however, relates to the action
of reducing agents.

If to a solution which exhibits the last - mentioned spectrum
there be added some sulphide of ammonium, it is observed that it
manifests the spectrum of reduced haemoglobin. On shaking the so-
lution containing the latter with air, oxy-haemoglobin is again formed.

Production The peculiar and remarkable properties above mentioned

of methaemo- were described by the Author in 18671 and more fully in
S«bii,™!! 1868> as developed by the action of nitrites on solutions of
haemoglobin and upon blood. It was shewn that besides
presenting the remarkable optical properties and reactions
previously referred to, as a result of the action of nitrites, the respiratory-
. oxygen of haemoglobin had become irremovable by carbonic oxide or in a
Torricellian vacuum, but that after undergoing the change the haemoglobin
could be crystallized repeatedly, the body thus produced only differing from
oxy-haernoglobin by its colour and its spectrum. On analysis it was found that
the crystalline compound always retained some of the nitrite used, and the
view was therefore expressed that in all probability the action exerted by ni-
trites consisted in the formation of a compound of those bodies with oxy-hae-
moglobin, which compound was decomposed by the reducing agent employed.
It was subsequently observed by Sorby2, Lank ester3, and Jaderholm4
that Gamgee's nitrite-haemoglobin spectra coincided with those of methae-
moglobin prepared by the action of potassium permanganate, and the
presumption has been established that his bodies really consisted of
methaemoglobin generated by the action of nitrites. This change in the
view as to the nature of the bodies produced under the influence of
nitrites does not however affect the facts established by the researches
above referred to. According to Sorby, however, methaemoglobin would
be a per-oxy-haemoglobin, i.e. a more highly oxygenized haemoglobin,

1 Gamgee, "Note on the action of nitric oxide, nitrous acid and nitrites on
Haemoglobin." Proceedings of the Royal Society of Edinburgh, 1867, p. 168. " On
the action of nitrites on blood." Philosophical Transactions, 1868, pp. 589 — 625.

2 Sorby, Quarterly Journ. of Micros. Sc. 1870, p. 400.

8 Lankester, " Abstract of a Keport on the Spectroscopic Examination of certain
animal substances." Journal of Anat. and Phys. Vol. iv. p. 123.

4 Jaderholm, " Untersuchungen uber den Blutfarbstoff und dessen Zersetzungs
producte," Abstracted from the original Swedish by Hammarsten in Maly's Jahres-
berichtt Vol. vi. p. 85.

A TEXT-BOOK

OF THE

PHYSIOLOGICAL CHEMISTKY

OF THE

ANIMAL BODY.

A TEXT-BOOK

OF THE

PHYSIOLOGICAL CHEMISTEY

OF THE

ANIMAL BODY

INCLUDING AN ACCOUNT OF THE CHEMICAL CHANGES
OCCURRING IN DISEASE

BY

ARTHUR GAMGEE, M.D., F.R.S.

\\

PROFESSOR IN THE VICTORIA UNIVERSITY, MANCHESTER; BRACKENBURY
PROFESSOR OF PHYSIOLOGY IN THE OWENS COLLEGE.

WITH ILLUSTRATIONS.

L I B L> A

VOL. L I U]STI

MACMILLAN AND CO.

1880

[The Right of Translation is reserved.]

BIOLOGT

LIBRARY

G

(flamforrtjge:

PRINTED BY C. J. CLAY, M.A.
AT THE UNIVERSITY PRESS.

TO

SIB EGBERT CHRISTISON, BART., M.D., LL.D., D.C.L.

EMEEITUS PEOFESSOE OF MATEEIA MEDICA IN THE
UNIVEESITY OF EDINBUEGH.

SIR,

In the preparation of this work the intention which I
had formed from the first of dedicating it to you has always
been present with me, and whilst the thought has added pleasure

to what has truly been a congenial task, it has incited me to

-.4; <^ *\"C»
make every effort in my power to render the book worthy of

your acceptance.

In offering you this tribute of admiration and regard, I
would express my sense of the signal benefits which, by your
labours as a teacher and investigator and by your personal
example, you have conferred upon the great University of
which you were so long an ornament, no less than upon the
whole medical profession in this country.

ARTHUR GAMGEE.

MANCHESTER, July 19, 1830.

G,

PREFACE.

IT has been my desire in the preparation of this work to
consider the subject of Physiological Chemistry from the
point of view of the biologist and the physician rather than
from that of the chemist, and, accordingly, I have adopted a
classification of the subject based entirely on morphological
or physiological considerations. Whilst I have, however,
given special prominence to all those facts which offer at
present the greatest interest to the biologist, and have kept
in the back ground such as only possess interest to the pure
chemist, because involving some doubtful question of consti-
tution, I have, nevertheless, taken care that no chemical
fact and even that no chemical speculation should be
omitted which appeared likely to throw light upon a bio-
logical question.

In the present volume the chemical composition of, and
the chemical processes relating to, the elementary tissues of
the body are treated of, the blood, lymph, and chyle being-
included in that classification. This volume forms a com-
plete and independent work, though it is intended that it
shall, within twelve months, be followed by a second volume,
in which the chemistry of the chief animal functions will be
treated of.

Some may be inclined to remark that I have introduced
into this work too large a reference to the sciences of

62

viii PREFACE.

anatomy and physiology and to practical medicine, and
that I have not always been consistent in the extent of
these references. The attentive reader will however dis-
cover, I trust, that I have proceeded with great deliberation,
and that if in certain cases I have made greater digressions
into the provinces of the cognate sciences than in others,
it has been because I considered that I was called upon to
do so in the interest of the particular subject, and therefore
in the interest of the reader. Thus, in the chapter on the
' Contractile Tissues' the histological descriptions are far more
detailed and the general review of known physiological facts
much more complete than in the case of the nervous tissues,
and the reason is obvious. It would have been unsatis-
factory to discuss the chemical processes of muscle without
considering, in some cases in considerable detail, the results
of the work of the histologist and of the experimental
physiologist. On the other hand, in dealing with the
scanty facts yet known to us concerning the chemical
history of the nervous tissues, only the barest outline of
the histology of the nervous system is essential.

Although this volume, in the main, deals with the
chemistry of the elementary tissues and not with the
processes which are characteristic of the complex organs of
the body, for the sake of convenience some exceptions have
been made. Thus the chemistry of the organs of sense
has been made to follow the chapter on the chemistry of the
nervous tissues, because this seemed the most convenient
place for introducing a systematic account of any facts
relating to them.

It has been a constant object with me to give the reader
a very full and, so far as possible, independent account of
the state of knowledge on the subjects discussed, and I trust

PREFACE. IX

I may with complete truthfulness say that this work is
based upon a study of original memoirs rather than upon a
study of text books. In the interest of the student nearly
all papers are quoted by their full titles and few have been
quoted which have not been read throughout and studied.
Whenever quotations have been made at second hand the
fact is stated.

Another feature which I have desired to render promi-
nent in this work is the description of the methods which
have been followed in important and, to borrow a con-
venient Germanism, 'epoch-making7 researches. It seemed
the more important to do this as I desired to write in the
interest of the truly scientific student, anxious not merely
to learn what has been already acquired to science, but
wishful himself to extend her boundaries.

I have, so far as possible, tried all the experimental
processes mentioned in this work, and throughout it I have
incorporated the results of my own independent researches
which in many cases have not yet been published else-
where.

Thus much as to the plan of this book : 1 know only too
well its deficiencies. I trust, however, that notwithstanding

', ' o

these it may assist the progress of science, and whilst I
plead for it the indulgence of my scientific brethren, I would
beg of them to aid me by communicating to me any errors
which they may discover, or any suggestions for a better
exposition of the subjects discussed.

In the discharge of my very arduous work I have been
greatly helped by many friends. In the first place I have to
express my unbounded acknowledgments to my friend and
former pupil Mr John Priestley, who has, with the exception
of some comparatively unimportant sections, written the

X PREFACE.

very important chapter on the ' Contractile Tissues/ and in
such a manner as will, I feel sure, attract the good opinion
of physiologists. Mr Priestley had, without any intention
of writing on the subject, made himself so thoroughly
master of all that had been written on the subject of the
physiology of muscle, that in the best interests of my
readers I asked him to assist me in dealing with this
subject. Although any credit which it may merit is due to
Mr Priestley for the greater part of this chapter, I must in
justice to myself say that every section and almost every
sentence in it have been the subjects of discussion be-
tween us.

I have been helped by Mr William Dodgson in the
reduction of the valuable tables of blood-spectra of Pro-
fessor Preyer to a scale of wave-lengths, and in the
actual drawing of the scale attached to the spectra of
haemoglobin and its derivatives. I may here incidentally
remark that in the description of spectra of any import-
ance I have referred all measurements to wave-lengths,
taking care to check the reduced observations of others by
measurements made with the help of one of Herr Zeiss's
beautiful spectroscopes furnished with a scale of wave-
lengths.

Lastly, I have to express my deep obligations to Dr
Alfred Young, to Mr Marcus Hartog, M.A., and to my
pupils Messrs Larmuth, Reynolds, and William Thorburn
for much useful help. Upon the first of these gentlemen
devolved the greater part of the labour of preparing a full
and accurate index.

ARTHUR GAMGEE.

Manchester,

July, 1880.

CONTENTS.

BOOK I.

THE PROTEIDS.
THE ELEMENTARY TISSUES OF THE ORGANISM.

CHAPTER I.

THE PROTEIDS. PAGE

SECT. 1. GENERAL CHARACTERS or THE PROTEIDS ...... 4

SECT. 2. CHEMICAL REACTIONS CHARACTERISTIC OF THE PROTEIDS ... 13
Methods of completely separating proteids from solutions containing

them 34

Determination of the temperature at which the proteids coagulate . 14

SECT. 3. SYNOPSIS OF THE CHIEF PROTEID BODIES 16

SECT. 4. PRODUCTS OF DECOMPOSITION OF THE PROTEIDS 18

SECT. 5. THEORETICAL VIEWS AS TO THE CONSTITUTION OF THE PROTEIDS . • 20

CHAPTER II.
THE BLOOD.

SECT. 1. THE PHYSICAL CHARACTERS OF THE BLOOD .23

SECT. 2. THE LIQUOR SANGUINIS. FIBRIN AND ITS SUPPOSED PRECURSORS . . 31

Fibrin 34

The Assumed Precursors of Fibrin in the Liquor Sanguinis :

1. Serum-Globulin or Paraglobulin (Schmidt's fibrinoplastic
substance) 37

2. Fibrinogen 40

Xll CONTENTS.

PAGE

Theories of Coagulation 42

The Fibrin-ferment 48

The origin of Fibrin-ferment 49

The researches of Hammarsten 51

The influence of salts on coagulation 53

Non-coagulation of salts within the living blood-vessels ... 54

SECT. 3. THE SERUM AND THE CONSTITUENTS OF THE LIQUOR SANGUINIS WHICH

REMAIN IN IT 57

The Proteids of the Serum 60

1. Serum-globulin or Paraglobulin 60

2. Serum-Albumin 61

The Extractive Matters of the Plasma and Serum .... 64

The Salts of the Plasma and Serum 66

The Gases of the Plasma and Serum 70

SECT. 4. THE COLOURED CORPUSCLES OF THE BLOOD . . . . . 71

Enumeration of the corpuscles ........ 74

Summary of the composition of the coloured corpuscles ... 79

The Stroma and the proteids associated with it .... 80

The Nuclei of the Bed Corpuscles 82

Fatty matters containing Phosphorus (Lecithin, Protagon) . . 83

Cholesterin 84

Oxy-Haemoglobin 84

Action of certain gases which displace the oxygen of Oxy-haemoglobin 104

Products of the decomposition of Haemoglobin . . • . . . . 108

Methaemoglobin 109

The proteid matter derived from the decomposition of Haemoglobin . 112

Haematin 113

Hydrochlorate of Haematin — Haemin 115

Haematoporphyrin . 117

Haemochromogen 118

Haematoidin 120

The mineral constituents of the red corpuscles . . . . . 121
The gaseous constituents of the coloured corpuscles . . . .123

SECT. 5. THE COLOURLESS CORPUSCLES OF THE BLOOD 123

SECT. 6. THE GASES OF THE BLOOD AS A WHOLE 12G

SUMMAKY OF THE QUANTITATIVE COMPOSITION OF THE BLOOD . 126

SECT. 7. CHARACTERS PRESENTED BY THE BLOOD OF INVERTEBRATE ANIMALS . 129
Distribution of Haemoglobin through the vascular liquids of various

groups of Invertebrata 130

On the green blood of certain Annelids. Chlorocruorin . . . 131

On the blue blood of certain of the Mollusca and Molluscoida . . 132

The blue blood of the Octopus. Haemocyanin .... 133
On certain coloured corpuscles found in the perivisceral fluid of

certain Sea-urchins and Holothurians 134

CONTENTS. Xlii

CHAPTER III.
CHANGES WHICH THE BLOOD UNDERGOES IN DISEASE.

PAGE

INTRODUCTION 136

SECT. 1. VAEIATIONS IN THE PROPORTION OF THE PRINCIPAL CONSTITUENTS OP

THE BC,OOD IN DISEASES IN GENERAL 139

SECT. 2. THE CHANGES WHICH THE BLOOD UNDERGOES IN PARTICULAR DISEASES 145

A. The Blood in Disorders of Nutrition :

Anaemia 145

Leucocythaemia (Leukaemia) 152

Progressive Pernicious Anaemia 154

Scurvy 156

. Purpura Hemorrhagica and Haemophilia 157

Gout 157

Articular Rheumatism and Rheumatoid Arthritis .... 158

Rickets and Osteomalacia 158

B. The Blood in Fevers :

Febricula or Ephemeral Fever 159

Typhus Fever 159

Typhoid Fever 159

Relapsing Fever * ... 159

Splenic Fever (of Cattle) 161

Intermittent Fevers 162

Scarlet Fever, Measles, Small-pox, Erysipelas ^_ . . 163

The Blood hi Cholera 163

C. The Blood in Diseases of the Heart 164

D. The Blood hi Diseases of the Lungs 167

E. The Blood in Diseases of the Liver 167

F. The Blood in Diabetes Mellitus .168

G. The Blood in Diseases of the Kidney 172

CHAPTER IV.
THE BLOOD (continued).
DESCRIPTION OF CERTAIN METHODS OF RESEARCH.

Determination of the Specific Gravity of the blood .... 174

Determination of the Reaction of blood 176

Determination of the Water, Total Solids and Ash of the blood . 177

Determination of the amount of Fibrin yielded by the blood . . 180

Determination of Haemoglobin in the blood 182

Determination of Cholesterin, Lecithin, and Fats in blood . . 187

Determination of Water, Total Solids, and Salts of Serum . ' . 188
Determination of the total amount of Proteids contained in the Serum

and of the Serum-albumin . 188 •

XIV CONTENTS.

PAGE

Determination of the amount of Fibrinogen contained in the liquor

Banguinis 189

Determination of the amount of Serum-globulin in the serum . . 189
Determination of the presence and quantity of Urea in the blood . 190
Determination of the amount of Uric Acid in the blood . . .193
Determination of the amount of Sugar hi the blood . . . .194
Determination of the weight of the Moist Corpuscles contained in the

blood 195

Separation and determination of the Gases of the blood . . . 196
Collection of the blood for the determination of gases . . . 197

Mercurial Pumps 198

Analysis of the gases of the blood 206

Description of the methods of Frankland for the analysis of gases . 207
Description of more simple methods of gas analysis .... 213
Determination of the total quantity of blood contained in an animal's

body ....... 215

Medico-legal detection of Blood-stains 217

Medico-legal detection of Carbonic Oxide in blood . . . .219

CHAPTER V.

THE LYMPH AND CHYLE. THE SO-CALLED TRANS UDATIONS, NORMAL
AND PATHOLOGICAL.

SECT. 1. THE LYMPH (INCLUDING THE CHYLE) 220

Preliminary observations 220

Physical characters of the lymph . . . . . . 221

The Proteids of the lymph . .' 223

The Fats of the lymph and chyle 223

The Extractive matters of lymph . . . . . . 223

The Salts of the lymph 224

The Gases of the lymph .* 225

SECT. 2. THE LIQUIDS CONTAINED IN THE HEALTHY SEROUS SACS. — SYNOVIA. —

THE CEREBRO- SPINAL LIQUID 229

SECT. 3. THE LIQUID IN DROPSIES 231

Preliminary remarks on the mode of production of Dropsies . . 231

General Characters of Dropsical Fluids 233

Characters of particular Transudations ...... 235

SECT. 4. METHODS OF ANALYSING LYMPH, CHYLE, AND OTHER TRANSUDATIONS,

NORMAL AND PATHOLOGICAL 236

CHAPTER VI.
PUS.

SECT. 1, INTRODUCTORY REMARKS ON THE PHYSICAL PROPERTIES OF Pus AND ON

THE NATURE OF Pus . . 238

CONTENTS. XV

PAGB

SECT. 2. THE Pus SERUM 239

SECT. 3. THE Pus CORPUSCLES . 241

The Proteids present in the Cell-protoplasm ..... 241

The matter of the nuclei. Nuclein ....... 241

The extractive matters of pus cells soluble in water .... 243

SECT. 4. COLOURING MATTERS FOUND IN Pus.

Pyocyanin 245

Pyoxanthose . . 246

SECT. 5. THE GASES OF Pus 246

SECT. 6. DIRECTIONS FOR THE QUANTITATIVE ANALYSIS OF Pus 248

CHAPTER VII.
THE CONNECTIVE TISSUES.

INTRODUCTION 249

SECT. 1. CONNECTIVE TISSUE PROPER 250

Connective Tissue Cells 251

The White Fibres of Connective Tissue. Collagen and Gelatin . 252

The Elastic Fibres of Connective Tissue. Elastin .... 255

Connecting or Ground substance of Connective Tissue . . . 256

SECT. 2. ADIPOSE TISSUE 259

Stearin 262

Palmitin .... . ^ . . . 262

Olein 263

Glycerin 263

Fatty matters found in the adipose tissue of certain of the lower

animals 264

Analysis of the Fats 265

SECT. 3. CARTILAGE ............ 268

General Composition of Cartilage ....... 268

Chondrigen . 269

Chondrin 269

SECT. 4. OSSEOUS TISSUE OR BONE 272

The Water found in bone 273

The Animal or Organic basis of bone ...... 273

The Mineral matters of bone ........ 274

The composition of the Marrow of bone 277

THE CHANGES WHICH BONE UNDEEGOES IN DISEASE.

Osteomalacia 280

Eachitis . 281

Caries .......••••• 284

Necrosis . • •' • 284

XVI CONTENTS.

METHODS FOLLOWED IN THE QUANTITATIVE ANALYSIS OF BONE.

PAGE

Determination of the quantity of Fat in bone 285

Determination of the total quantity of Ash in bone .... 285

Determination of the quantity of Chlorine in the ash of bone . . 285

Determination of the amount of Calcium in bone .... 286

Determination of the Magnesium in bone 286

Determination of Phosphoric acid in bone 286

Determination of Carbonic acid in bone 287

Determination of Fluorine in bone 288

SECT. 5. TOOTH 289

Dentine 289

Enamel . . . 291

Crusia Petrosa 293

CHAPTER VIII.

EPITHELIAL TISSUES OE EPITHELIUM. KEEATIN. CHITIN. PIG-
MENTS DEPOSITED IN THE EPITHELIAL STBUCTUEES.

CEETAIN OTHEE ANIMAL PIGMENTS 295

SECT. 1. EPIBLASTIC KERATIN-PRODUCING EPITHELIAL TISSUES. THE HORNY

SUBSTANCE OF CUTICLE, NAILS, HORN, HAIR, AND FEATHERS . . 296

Horny Substance or Keratin 297

Inorganic Matters contained in the Horny Tissues .... 298

SECT. 2. TISSUES WHICH YIELD CHITIN, SPONQIN, TUNICIN, AND HYALIN . 299

Chitin 299

Glycosamine 301

Conchiolin 301

Spongin 302

Hyalin 302

Tunicin or Animal Cellulose 302

SECT. 3. ON CERTAIN COLOURING MATTERS OF THE EPITHELIAL TISSUES OF

VERTEBRATES 303

Brown and black Pigments. Melanin 303

Pigments of the Feathers of Birds. Turacin 304

SECT. 4. CERTAIN OTHER COLOURING MATTERS OCCURRING IN THE ANIMAL

KINGDOM 305

Blue Stentorin « . . 307

Actinioohrome 307

Bonellein 307

Carminic acid 303

Tyrian Purple 309

Chlorophylloid Colouring Matters 309

CONTENTS. xvii

CHAPTER IX.
THE CONTEACTILE TISSUES.

PAGE

SECT. 1. THE STRUCTURE OF MUSCLE AND THE CONSTITUENTS OF NORMAL LIVING

MUSCLE 310

The Structure of the Contractile Tissues 310

Structure of Unstriped Involuntary muscle 311

Structure of Voluntary muscle 313

The Structure of the Muscular Substance of the Heart . . . 318

Terminations of Nerves in muscle , 318

CHEMICAL CONSTITUTION OF NORMAL LIVING MUSCLE, so FAR AS IT CAN BE

KNOWN OR INFERRED 319

On the distribution of liquid and solid parts in a voluntary muscular

fibre 319

Chemical characters of the Sarcolemma 321

Chemical nature of the doubly-refracting elements of voluntary

muscle 321

The Muscle Plasma 322

Myosin 323

Muscle Serum 324

The Haemoglobin of muscles . . 325

NITROGENOUS (NON-PROTEID) ORGANIC CONSTITUENTS OF MUSCLE . 325

Creatine 326

Creatinine 329

Hypoxanthine or Sarcine 329

Xanthine 330

Carnine ~~T . , , 332

Uric acid « 333

Urea ... 333

Inosinic acid 333

Taurine 333

NON-NITROGENOUS ORGANIC CONSTITUENTS OF MUSCLE . . . 333

Fats 334

Glycogen 334

Dextrin 336

Fermentable Sugar . 336

Inosit 336

The Ferments present in muscle 338

THE INORGANIC CONSTITUENTS OF MUSCLE 338

SECT. 2. GENERAL PHENOMENA OF LIVING MUSCLE 339

Muscle in a state of rest ......... 839

Muscle in action 341

Eigor Mortis 347

SECT. 3. SPBCIAL STUDY OF THE CHEMICAL CHANGES OF LIVING MUSCLE . 348

THE CHEMICAL CHANGES OF CONTRACTION AND EIGOR . . . 349

A. Changes in the composition of muscle itself .... 349

Changes in the gaseous constituents . . . . . . 349

Changes in the non-gaseous constituents 359

xviii CONTENTS.

PAGE

1. Change in reaction and its causes 359

2. Changes in the proportion of water 364

3. Changes in the water and alcohol extractives . . . 364

4. Changes in the proteids 364

5. Changes in the amount of Creatin ..... 364

6. Changes in the proportion of glycogen and sugar . . . 365

7. Changes in the amount of fat and volatile fatty acids . . 365

8. Oxidizing and reducing properties of Muscle during rest

and tetanus 365

B. Changes in the chemical composition of the medium surrounding

Muscle 365

a. When Muscle is exposed to the air ..... 365

/3. When Muscle is still in the body 373

Changes of the medium surrounding muscle as shewn on analysis of

the blood of Muscle 375

Changes in the medium surrounding Muscle as shewn in the analyses

of the general excreta of the body 381

THE CHEMICAL CHANGES OF LIVING MUSCLE WHEN AT BEST . . 401

SECT. 4. FATIGUE, EXHAUSTION AND KEVIVAL 404

SECT. 5. THE THEORY OF MUSCULAR ACTIVITY . . . . . . 406

CHAPTER X.
THE NEBVOUS TISSUES.

SECT. 1, INTRODUCTORY 420

Nerve-cells 420

Nerve-fibres 421

SECT. 2. THE PROTEIDS FOUND IN THE NERVOUS TISSUES .... 423

SECT. 3. NEUROKERATIN AND NUCLEIN . . 423

SECT. 4. THE PHOSPHORIZED CONSTITUENTS FOUND IN NERVOUS TISSUES . 425

Protagon 425

Lecithin 430

Description of some of the products of decomposition of lecithin

and protagon . 433

Phosphorized principles other than protagon and lecithin . . 437
SECT. 5. NON -PHOSPHORIZED NITROGENOUS BODIES OF UNKNOWN CONSTI-
TUTION . • . . . 439

Cerebrin? orCerebrins? .' 439

SECT. 6. CHOLESTERIN ..'.." 442

SECT. 7. EXTRACTIVE MATTER COMMON TO THE NERVOUS AND OTHER TISSUES 444

SECT. 8. THE INORGANIC CONSTITUENTS OF THE NERVOUS TISSUES . . 445
SECT. 9. GENERAL SUMMARY SHEWING THE RESULTS OF QUANTITATIVE ANALYSES

OF BRAIN, SPINAL CORD, AND NERVES 445

SECT. 10. THE CHEMICAL PROCESSES CONNECTED WITH THE ACTIVITY AND

DEATH OF THE NERVOUS TISSUES . . . 446

CONTENTS. xix

CHAPTER XI.

CHEMICAL HISTOEY OF CEETAIN OF THE PEEIPHEEAL TEEMI-
NATIONB OF THE NEEVOUS SYSTEM AND OF THE AC-
CESSOEY STEUCTUEES CONNECTED WITH THEM, THE
TISSUES AND MEDIA OF THE EAE, THE TISSUES AND
MEDIA OF THE EYE.

PAGE

SECT. 1. THE TISSUES AND MEDIA OF THE EAR 448

Perilymph and Endolyrnph . . . . . . . 449

Otoliths, Lapilli, or Otoconia 449

The membranous Labyrinth 450

SECT. 2. THE TISSUES AND MEDIA OF THE EYE 450

The Cornea 450

Sclerotic 451

Aqueous Humor 452

Crystalline Lens . . • 452

The Choroid 454

THE EETINA.

Introductory 454

Chemical composition of the Eetina as a whole . . . 459

General chemical facts relating to Eods and Cones .... 459

Colouring matters associated with the Cones (Chromophanes) . . 460
Colouring matters associated with the Eods (Visual Purple or Eho-

dopsin) . . 461

Chemical facts relating to the Eetinal Epithelium .... 468

Action of light upon the Visual Purple of the Living Eye . . 469

Eegeneration of Visual Purple 469

INDEX 471

BOOK I.

THE PROTEIDS,
THE ELEMENTARY TISSUES OF THE ORGANISM.

V OF

'vLlFUMN! A.

CHAPTER I.

THE PROTEIDS.

AMONGST the organic proximate principles which enter into the
composition of the tissues and organs of living beings, those belonging
to the class of proteid or albuminous bodies occupy quite a peculiar
place and require an exceptional treatment, for they alone are never
absent from the active living cells, which we recognize as the primor-
dial structures of animal and vegetable organisms. In the plant,
whilst we recognize the wide distiibution of such constituents as
cellulose and chlorophyl, and acknowledge their remarkable physio-
logical importance, we at the same time are forced to admit that
they occupy altogether a different position from that of the proteids
of the protoplasm out of which they were evolved. We may have a
plant without chlorophyl and a vegetable cell without a cellulose
wall, but our very conception of a living, functionally active, cell,
whether vegetable or animal, is necessarily associated with the
integrity of its protoplasm, of which the invariable organic constitu-
ents are proteids.

In the animal, the proteids claim even more strikingly our
attention than in the vegetable, in that they form a very much larger
proportion of the whole organism, and of each of its tissues and organs.
We may indeed say that the material substratum of the animal
organism is proteid, and that it is through the agency of structures
essentially proteid in nature that the chemical and mechanical
processes of the body are effected. It is true that the proteids are
not the only organic constituents of the tissues and organs, and
that there are others, present in minute quantities, which probably
are almost as widely distributed, such as for instance phosphorus-
containing fatty bodies, and glycogen, yet avowedly we can (at the
most) only say probably, and cannot, in reference to these, affirm
that which we may confidently affirm of the proteids — that they are
indispensable constituents of every living, active, animal tissue, and
indissolubly connected with every manifestation of animal activity.

There are then, it will be admitted, good reasons why a general
sketch of the proteid bodies should be the proper introduction to a
treatise on physiological chemistry, in which the classification is

1—2

4

DISTRIBUTION OF THE PEOTEIDS.

[BOOK i.

intended to be as much as possible one based upon physiological
considerations ; and the reader will not find it inconsistent that
whilst a systematic account of these bodies is given in the first
place, apart from any special tissue or organ, in the case of other
proximate principles their description and consideration is incorpo-
rated in the account of the organ or tissue with which they appear
to have the closest connection.

SEC. 1. GENERAL CHARACTERS OF THE PROTEIDS.

The bodies included under this category are highly complex, (for
the most part) non-crystallizable, compounds of carbon, hydrogen,
oxygen, nitrogen and sulphur, occurring in a solid viscous condition,
or in solution, in nearly all the solids and liquids of the organism.
The different members of the group present differences in physical
and, to a certain extent, even in chemical properties ; they all possess,
however, certain common chemical reactions, and are united by a
close genetic relationship.

The following table exhibits the proportions of proteids, or their
immediate derivatives, contained in the various liquids and solids of the
body (Gorup-Besanez1).

A. LIQUIDS.

Cerebro-spinal liquid contains

Aqueous humour

Liquor Amnii

Intestinal juice

Liquor Pericardii

Lymph

Pancreatic juice

Synovia

Milk .

Chyle .

Blood .

0-09 per cent, of Proteids.
0-H
0-70
0-95
2-36
2-46
3-33
3-91
3-94
4-09
19-56

B. SOLID TISSUES AND ORGANS.

Spinal Cord contains

Brain. ....

Liver ....

Thymus (of Calf) .

Muscles ....

Tunica media of Arteries

Crystalline lens .

The proteids of the animal body are all derived, directly or indi-
rectly, from vegetable organisms, which possess the power of con-
structing them out of the comparatively simple chemical compounds
which serve as their food. Such a synthesis never takes place in the

1 Vide Gorup-Besanez, Lehrbuch der physiologischen Cliemie, 4te Auflage (1878), p. 128.

7-49 per cent, of Proteids.

8-63 „ „

H-74
12-29
16-18
27-33
38-30

CHAP. I.] THE PROTEIDS. 5

animal body, though the latter possesses the power of converting any
vegetable or animal proteid into the various proteids which are
characteristic of its solids and liquids. By the action of certain fer-
ments present in the alimentary juices, all proteids are capable of
being converted into closely allied bodies called peptones, which
after absorption are capable of reconversion into proteids. In the
organism the proteids thus introduced, after forming part of the
circulating blood, are partly employed in the reconstruction of slowly
wasting proteid tissues and organs ; for the most part, however, they
are subjected to a rapid series of decompositions, of which presumably
the most important take place in the liver, and which finally result in
the formation of carbonic acid, water and various imperfectly oxidized
organic bodies which contain all the nitrogen originally present in
the proteid; of those bodies the most abundant by far is carbamide

or urea, C^ JNH*

To the assemblage of chemical processes, or rather to the assemblage
of transformations, which a constituent of the organism, such as a proteid,
undergoes in its passage through the body, the term metabolism has been
applied, and we shall frequently employ it in this sense, the processes
themselves being designated when convenient metabolic processes.

In the processes of metabolism to which the proteids are subjected
and which result in the formation of C02, H20 and urea, there are
formed intermediate bodies, such as glycogen and fats, which play an
important part in the economy of the body.

It is further unquestionable that within the animal body certain
remarkable synthetic processes occur, by which proteids are built up
into bodies of a yet more complex structure, such for instance as the
blood colouring matter, Haemoglobin.

Percentage The various Proteids differ somewhat in elemen-

S^£?wSi0f tary comP°sition> within the limits of the following
numbers1 :

C H N S O

From 51-5 6'9 152 (V3 209

to 54-5 to 7'3 to 17'0 to 2'0 to 23'5.
In addition to these essential constituents, the proteids, however
carefully they may have been purified, usually leave when ignited a
small quantity of ash, the composition of which varies in different
cases, chlorides and phosphates of the alkaline metals being the pre-
dominant constituents.

Proteidsfor Certain of the proteids exist in a state of solution

the most part in the liquids of the organism; others are present in
soluble. the same state in the tissues; all may be dissolved by

certain reagents, though in some cases riot without suffering radical
changes.

1 Hoppe-Seyler, Handbuck d. pkys.- und path.-chem. Analyse, 4te Aufl. p. 223.

6 COLLOIDAL CHARACTER OF PROTEIDS. [BOOK I.

When solutions of the proteids are dried at a gentle heat so as to
drive off the water in which they are dissolved, or with which they
are combined, they appear as translucent and perfectly amorphous
solids, which break with a vitreous fracture, and furnish, when
triturated, a yellowish-white or white powder. Unless it has been
subjected to a high temperature, the powder thus obtained by
evaporating watery solutions of proteids, is found to be again soluble
in water. By exposure to too high a temperature the body may be
rendered insoluble.

Proteids Solutions of all proteids are found to be non-dif-

are Colloids, fusible through parchment-paper, and this property

i.e. non-dif- allows us in certain cases to separate proteids from

other matters with which they are mixed, and in some

cases even to separate one proteid from another.

Thus the chief proteid constituent of the blood is a body termed
serum-albumin. If this body, which is soluble in water, be pre-
sent in a solution which contains saline ingredients and diffusible
organic bodies, such for instance as sugar or urea, we can effect
the separation of the albumin by taking advantage of its properties
as a colloid. If we place the solution in a dialyser (Fig. 1 and
Fig. 2), i.e. in a suitable vessel where it may be in contact with
one side of a surface of parchment-paper, the other side of which
is immersed in pure water, which is frequently renewed, the diffu-
sible or so-called crystalloid constituents, such as the soluble salts,
the sugar and the urea, will pass through the parchment-paper into
the water, and there will be ultimately left within the dialyser a
solution of pure serum-albumin; if there be present in the original
solution not only albumin which is soluble per se in water, but such
a proteid as paraglobulin, which is held in solution by the water in
virtue of the salts which may be present, as these diffuse out it is
precipitated, so that 'by the process of dialysis alone we may succeed
in separating not only the proteids from diffusible admixtures, but, in
certain cases, to separate partially one proteid from another.

The process of dialysis is one which is frequently of great use in
physiological chemistry. Various methods of carrying on the process are
employed. In some cases the dialyser is made by stretching and tying a
sheet of moist parchment-paper over a Loop of gutta percha ; the liquid
to be dialysed is then placed in this dialyser, which is immersed in a
larger vessel containing water (Fig. 1). A convenient form is made of glass
of the shape shewn in Fig. 2, the parchment-paper being tied across the
wide open mouth of a bell of glass, which is suspended iu water by its
narrower neck.

Of late, hollow tubes of parchment-paper have been sold for the manu-
facture of sausages, and these serve admirably as dialysers ; the fluid to
be dialysed being placed within the tube, which is suspended in water.
In this case, as also in using the instruments shewn in Fig. 1 and Fig. 2,
it is often advisable to arrange for a constant influx and efflux of water
from the vessel in which the dialyser is immersed.

CHAP. I.]

THE PROTEIDS.

In all experiments on dialysis care has to be taken, before an experi-
ment is commenced, to ascertain that the parchment-paper is quite free

Fig. l.

HOOP DIALYSER. Fig. 2. BULB DIALYSER.

DlALYSERS DESCRIBED IN THE TEXT.

Proteids all
rotate the
plane of po-
larized light
to the left.
Determina-
tion of their
Specific Ro-

from even the minutest holes. These are readily detected if the outer
surface of the dialyser (i. e. the surface which during the actual experiment
is to be immersed in water) be dried and placed upon a sheet of filtering
paper, and then water poured into the interior ; a leak being evidenced
by the appearance of moisture on the outside.

Amongst the organic constituents of the animal
body a large number when dissolved possess the power
of rotating the plane of polarized light ; as for instance
the proteids, the sugars, the bile acids, &c. The deter-
mination of the fact that a solution of a body rotates
the plane of polarized light in a definite direction and
to a definite extent is sometimes of great service in

tatory power, aiding its identification, and in enabling its amount

to be determined.

As the rotation exerted by an active body dissolved in an inactive
liquid is dependent upon the molecules of the active body existing in
solution, the degree of rotation will in the case of any particular substance
be proportional to the number of active molecules traversed by the light,
and therefore proportional to the length of the column of liquid traversed,
and to the degree of concentration, of the solution. If, for instance, a
column of solution of any active substance, say of cane-sugar, of any given
length, rotate the plane of polarized light x degrees, then if the column be
doubled the rotation will amount to 2x degrees; or the double rotation will
be observed if instead of doubling the length of the tube the amount of
active substance in a given volume of liquid be exactly doubled. It can
be shewn that any active body rotates to different degrees the plane of
polarization of light of different colours. In determining, therefore, the

8 SPECIFIC ROTATORY POWER . [BOOK I.

rotatory power exerted by different bodies, care must be taken that the
nature of the light is the same. The light obtained by volatilizing sodium
compounds in a colourless gas flame affords an admirable source of light of
one uniform wave-length.

The expression * specific rotatory power ' or * specific rotation ' is used
to designate the rotation (expressed in degrees) of the plane of polarized
light, produced by 1 gramme of substance dissolved in 1 cubic centimetre
of liquid when examined in a column 1 decimetre thick.

Let a be the rotation observed, and p the weight in grammes of the
active substance contained in 1 cubic centimetre, and let I be the length
of the tube in decimetres, then if we designate by (a)D the specific rotation
for light having a wave-length corresponding to D,

In this formula the sign + indicates that the substance is dextrogyrous,
the sign — that it is Icevogyrous. In some cases the rotation is determined
for mean yellow light and not for Z>, and is expressed by (a)jf the value
of which is always somewhat different from that of (a)D.

Various instruments have been devised and much employed in the
determination of rotation of the plane of polarization, especially in the
estimation of sugar, and are known by the terms Saccharimeters, Polari-
meters, and Polaristrobometers. One of the most convenient and most
widely employed is the saccharimeter of Soleil, which as modified by
Yentke and Hoppe-Seyler, enables the percentage of serum-albumin and
of glucose present in a liquid to be directly read from a scale attached
to the instrument. In this instrument the rotation is determined for
the mean yellow.

The instrument of Soleil1 has however been of late years sur-
passed by others, especially by those invented by Wild, Jellett, and
Laurent. A description of the latter instrument will alone be
given.

Laurent's This instrument is shewn in Figs. 3 and 4. A Vis a

Poiarimetre a Bun sen lamp. A (Fig. 3) is a small spoon of platinum
Penombres. gauze with the tip turned upwards, and in this is placed
a small quantity of common salt. The tip of the spoon is placed
in the outer flame, and when the salt is volatilized an extremely
brilliant sodium flame is produced. At B is a cell containing potas-
sium bichromate, which cuts off all but the yellow rays. To the
lever /is attached a double refracting prism which polarizes the light,
and at D (Fig. 4.) is a diaphragm of which one half is covered by a
plate of quartz. This serves to modify the light in a manner
explained in the account of the theory of the instrument. The

1 For the description of the Soleil- Ventke Saccharimeter, and of Wild's Polari-
strobometer, the reader is referred to Hoppe-Seyler 'a Handbuch der physiologisch-u.
pathologisch-chemischen Analyse, and for a fuller description of these instruments,
as well as for a discussion of the whole subject of rotatory polarization, to Professor
Llandolt's recent work entitled Das optische Drehungsvenuogen organischer Snb-
stanzen und die practischen Anwendungen desselben. Braunschweig, Vieweg und Sohn,
1879, p. 237.

CHAP. I.]

THE PROTEIDS.

eye-piece tube 0 contains a Nicol's prism as analyser at K (Fig. 3),
and the whole tube, with the vernier and reading lens L attached^ can

Fig. 3. LAURENT'S POLARIMETRE.

be rotated by the screw 6r, or the eye-piece and vernier remaining fixed
the analyser can be rotated independently by the tangent screw F.
The vernier moves against the circle c, of which the rim is gradu-
ated. When the rotatory power of any substance is to be deter-
mined, a tube containing water is first placed in the position T (Fig. 4)
and by means of the screw G (Fig. 3) the zero of the vernier is brought
to coincide with that of the scale. When the eye- piece has been ad-
justed so that the line dividing the two halves of the field is perfectly
clear and sharply defined, these two halves are brought to the same in-
tensity by means of the screw F, the scale still reading zero. Should
the illumination of the field be too faint it may be increased by
moving the lever J (Figs. 3 and 6) slightly, though it is preferable to
work with the instrument when the lever is in such a position that
almost all the light is cut off. The water tube is now replaced by that
containing the substance to be tested. If it is active the two halves
of the field will at once be seen to be of unequal intensities. The

10

LAURENT'S POLARIMETRE.

[BOOK i.

screw G is then turned till the equality is restored and the reading
of the circle at once gives the rotation due to the substance, right-
or left-handed, according as the vernier is to the right or left of the
zero on the scale. The following is an example of the determination
of the rotatory power of a solution of sodium glycocholate in alcohol.
The solution in a tube 2 decimetres long gave a rotation of
+ 1° 40' or r-b'66. On evaporation, 10 c.c. of the solution gave 0'322
grm. of dry residue, or 1 c.c. contained '0322 grm. of the salt. Now
the specific rotation aD being denned as that due to a column of
liquid 1 decimetre long and containing 1 grm. of salt per 1 c.c., we have

+ 1°'666 = * x 2 x -0322

or

Theory of Laurent's Polarimetre1. The light from the sodium flame A
(Fig. 4) is deprived of all traces of blue or violet rays by the potassium
bichromate solution in the cell B. It then passes to the doubly refracting
prism P, whence half of it emerges polarized in one plane, the other half,
polarized in a perpendicular plane, being refracted away from the axis and
stopped by a diaphragm.

Fig. 4. DIAGRAM OF LAURENT'S POLARIMETRE.

At D is a diaphragm of which one half is covered by a plate of quartz
cut with the axis in the surface and parallel to the edge2. To understand
the effect of this crystal let Fig. 5(1) represent the diaphragm, the shaded
part being the quartz plate. Let OB be the direction of vibration of
the light after polarization by the prism. This will still continue to be the
direction of vibration of the light which goes through the right half of the
diaphragm, but a ray vibrating parallel to OB will on entering the quartz
on the left be resolved into two rays, one vibrating parallel to the axis
OA, which we represent by Oy, the other perpendicular to the axis, which
we represent by Ox. These two rays will travel at different rates through

1 For this account of the theory of Laurent's Polarimetre, I am indebted to my
fiicnd Mr J. H. Poynting, M.A., Fellow of Trinity College, Cambridge.

2 When cut in this manner quartz has no rotatory power but behaves just as any
other uniaxal crystal.

CHAP. I.] THE PROTEIDS. 11

the crystal, which is cut of such a thickness that one ray is retarded in
its passage just half a wave-length of sodium light behind the other, or
what amounts to the same, executes half a vibration more than the other
while in the crystal. On emergence then, while one vibration is from
0 toy the other instead of being from 0 to x is from 0 to x' in the opposite
direction, and the two now unite to form a resultant vibration OB' equal
to OB but at an angle AOB' equal to AOB on the other side of OA.

Now if the tube T (Fig. 4) only contain water or some non-rotating
liquid, the two rays will pass through it with their directions of vibration
OB, OB' unaltered to the analysing NicoFs prism N. This will only allow
rays to pass through it which vibrate parallel to a particular direction. If
the prism be turned so that this direction SP (Fig. 5, (2)) is perpendicular
to OB, the right-hand ray having no component parallel to SP is
extinguished, while the left-hand ray will have a more or less considerable
component in that direction and the left-hand side of the diaphragm D will
alone be visible in the telescope OH (Fig. 4).

So if the prism be turned round till SP is perpendicular to OB'
(as in Fig. 5, (3)) only the right-hand side of the diaphragm is visible.

But if SP be turned so as to be perpendicular to OA, vibrations
parallel to OB, OB' have equal components parallel to SP, and the two

\ *'k-S

(1) (2) (3) (4)

Fig. 5.

halves of the diaphragm appear equally illuminated (as in Fig. 5, (4)). In
this position of the analyser the instrument should read 0°.

It is possible to adjust SP perpendicular to OA with very great accuracy,
for when OB, OJ? make small angles with OA a very small rotation of the
analyser makes a great difference in the relative illumination of the two
halves of the field1.

When $P is thus adjusted perpendicular to OA and the instrument reads
0°, let a liquid possessing the power of rotating the plane of polarization be

1 This will be seen at once from the mathematical expression for the intensity of
the component parallel to SP.

Let AOB Fig. 5 (2) =a, BOP — 90-0, where 0 = a in the position of equality of
illumination. Let the intensity of the resolved part of the ray OB parallel to SP = I.

Then 1= OB2 cos2 BOP = OB* sin2 0, and ~ = 20 & sin 0 cos 0.

da

Therefore i ~ = 2 cot 0.
I dO

This, which expresses the proportion between the change of intensity and the original
intensity, is greatest when 0 is least, and therefore o should be as small as possible.

12

SPECIFIC ROTATION OF VARIOUS PROTEIDS. [BOOK I.

placed in the tube T. Both the directions of vibration OS, Off will be turned
through equal angles in the same direction, and their components along SP
will be no longer equal, and one half of the field will appear brighter

Fig. 6.

than the other (Fig. 6, B or C). If the prism be now turned round
by means of the screw G till we again have equal illuminations in the
two halves of the field (Fig. 6, A), SP has evidently been turned through
the same angle and in the same direction as that through which the liquid
has rotated the planes of polarization OB, OK, and the reading of the
instrument in its new position at once gives us the angle of rotation.

TABLE EXHIBITING THE SPECIFIC ROTATION OF SOME OF THE CHIEF
PROTEID BODIES FOR THE YELLOW LINE D. (COMPILED FROM THE
OBSERVATIONS OF HOPPE-SEYLER1 AND HAAS2.)

Proteid.

Observer.

Value of (a) U

Serum-albumin.

Hoppe-Seyler.

-56°

Egg-albumin.

Hoppe-Seyler.
Haas.

- 33°-5

- 38°-08

Paraglobulin obtained from ascitic fluid

Haas.

-59°-75

by dilution of CO2.

Sodium-albuminate prepared from pure

Haas.

-62°-20

egg-albumin.

Acid-albuminate (Syntonin) prepared from

Haas.

-63°-12

pure egg-albumin by action of acetic acid.

Syntonin prepared from myosin by solu-

Hoppe-Seyler.

-72°

tion of that body in very dilute hydro-

chloric acid.

Casein, dissolved in solution of magnesium

Hoppe-Seyler.

-80°

sulphate.

1 Hoppe-Seyler, Zeitschrift f. Chem. u. Pharm. 1864, p. 737.

2 Haas, "Ueber das optische und chemische Verhalten einiger Eiweisssubstanzen,
inabesondere der dialysirten Albumine." Pfluger's Archiv, vol. xn. pp. 378--410.

CHAP. I.] THE PROTEIDS. 13

SEC. 2. CHEMICAL REACTIONS CHARACTERISTIC OF THE
PROTEIDS \

Only certain of the proteids are soluble in water; they are all
soluble however, especially with the aid of heat, in concentrated acetic
acid, and in solutions of the caustic alkalies; they are insoluble in
cold absolute alcohol and in ether.

Solutions of the proteids are precipitated by the following
reagents: —

1. By strong mineral acids added in sufficient quantities.

2. By acetic acid and potassium ferrocyanide.

3. By acetic acid and a large addition of concentrated solutions
of neutral salts of the alkalies or alkaline earths.

4. By basic lead acetate. * £<,£,, JU.*^. »/ '-,

5. By mercuric chloride. ~t n.J .., ..^ q^^J&ft .

6. By tannic acid.

7. By powdered potassium carbonate added to the solution until
it is nearly saturated.

8. The majority of the proteids are completely precipitated from
their solutions by alcohol, though in the presence of free alkali they
are slightly soluble in hot alcohol.

Detection When proteids are present in a solution the following

of Proteids reactions are employed in their detection : —
in solution. j ^ Uquid ig boiled and nitric ^ ^^ g() ag

to produce a strong acid reaction. The occurrence of a precipitate on
boiling, which is undissolved by nitric acid, and the immediate pro-
duction of a precipitate by nitric acid indicates the presence of a
proteid, to be confirmed by other tests.

2. The liquid is rendered strongly acid with acetic acid, and
solution of potassium ferrocyanide added; all proteids are thrown
down in the form of a white flocculent precipitate.

3. The liquid is rendered strongly acid with acetic acid, and is
boiled with its own volume of a saturated solution of sodium sulphate,
which will precipitate any proteid present.

The above tests are very satisfactory except in the case of only
slight traces of proteids being present ; under any circumstances it is
desirable to obtain confirmatory evidence ; the following methods are
then useful : —

4. Millon's reaction. When a strongly acid solution of mercuric
nitrate, made according to the directions to be afterwards given, is

1 In preparing a part of this section the author has availed himself greatly and
followed very closely, in some sentences almost literally, §§ 135 and 136 of Professor
Hoppe-Seyler's Handbuch der physiologisch- und pathologisch-chemischen Analyse.
3rd Edit. 1870.

14 SEPARATION OF PROTEIDS. [BOOK I.

added to a solution containing even a trace of a proteid, and the
mixture heated, the liquid assumes a purple-red colour. This reaction
is common to all the proteids and to their immediate derivatives.

Millon's reagent is made by dissolving 1 part by weight of mercury in
2 parts of nitric acid of specific gravity 1*42 and after complete solution
diluting each volume of liquid with two volumes of water.

5. Xanthoproteic reaction. The liquid supposed to contain a
proteid is boiled for some time with concentrated nitric acid. If a
proteid be present the liquid assumes a yellow colour, which changes
to an amber-red when an excess of alkali is added to it.

Methods of completely separating proteids from solutions containing

them,.

It is often of great importance to remove all the proteids which
a liquid contains, so as to proceed to the search for other substances.
The following methods are available : —

1. The liquid is treated with several times its volume of absolute
alcohol, and acetic acid added until the reaction is acid. After 24
hours the fluid is filtered ; the proteids are contained in the insoluble
matters on the filter.

2. To the liquid rendered faintly acid and heated to boiling, and
from which all the proteids separable by mere boiling have been re-
moved, a solution of ferric acetate, made by saturating acetic acid with
recently precipitated ferric hydrate, is added. After boiling for a few
minutes a solution is obtained which contains neither proteids nor
iron.

3. In some cases when soluble proteids precipitable by boiling
are present, by merely boiling the liquid they are entirely separated ;
such is usually the case with albuminous urine. If the liquid have
an alkaline reaction, a little acetic acid should be added, in quantity
just sufficient to neutralize the free alkali. If the quantity of acid be
either too scanty or too great the separation is incomplete; under
these circumstances the addition of a few drops of the solution of
ferric acetate mentioned in the last paragraph brings about the com-
plete precipitation and separation.

Determination of the temperature at which the proteids coagulate.

As will be shewn in the sequel, two groups of proteid bodies (the
albumins and the globulins) are precipitated from their solutions when
they are heated, and the temperature at which coagulation occurs
is in some cases an important characteristic.

CHAP. I.]

THE PROTEIDS.

15

The method of determining the temperature of coagulation is
illustrated by Fig. 7. A glass beaker containing water is placed within
a second larger beaker also containing water, the two being separated
by a ring of cork. Into the water contained in the inner beaker
there is immersed a test-tube, in which is fixed an accurately graduated
thermometer, provided with a long narrow bulb. The solution of
proteid of which the temperature of coagulation is to be determined

Fig. 7- APPARATUS EMPLOYED IN DETERMINING THE TEMPERATURE OF COAGULATION.

is placed in the test-tube, the quantity being just sufficient to cover
the thermometer bulb.

The whole apparatus is then gradually heated. With the arrange-
ment described the rise in temperature of the contents of the test-
tube takes place very slowly and equably throughout. Care being
taken to have as good an illumination as possible (the best plan
being to place the apparatus between the operator and a well lighted
window) the experimenter notes the temperature at which the liquid
first shows signs of opalescence ; he afterwards notes again the tem-
perature at which a distinct separation of flocculent matter occurs.

16

TEMPERATURE OF COAGULATION OF PROTEIDS. [BOOK 1.

TABLE EXHIBITING THE TEMPERATURE AT WHICH SOLUTIONS OF
VARIOUS PBOTEIDS, BELONGING TO THE GROUP OF ALBUMINS AND
GLOBULINS, COAGULATE.

Temperature

Tempera-

Name of
Proteid.

Character of the Solution.

Observers.

at which
opalescence

ture of
Coagula-

first occurs.

tion.

Serum-Albu-
min.

Dissolved in blood-
serum, hydrocele
fluid, &c.

Hoppe-
Seyler1.

60°— 65°

72°— 73°

Egg-Albu-
min.

Dissolved in water.

Hoppe-
Seyler1.

72°— 73°

II.

Vitellin.

Dissolved in a weak
solution of NaCl.

Weyi2.

•
70°

75°

Myosin.

Dissolved in a weak
solution of NaCl.

Kiihne3.
Weyl2.

55°— 60°

Fibrinogeii.

Dissolved in the liquor
sanguinis.

Frederique4.

56°
(55°-57°)

Paraglo-

Dissolved in solution

Hammar-

fiQO

7^0

bulin.

. ofNaOl.

sten6.

SEC. 3. SYNOPSIS OF THE CHIEF PROTEID BODIES.

The various proteid bodies occurring in the animal body will be
described in connection with the tissues of which each is most charac-
teristic; it will be convenient, however, to give a synopsis exhibiting
the principles upon which they have been classified.

CLASS I. Albumins: proteid bodies which are soluble in water
and which are not precipitated by alkaline carbonates, by sodium
chloride, or by very dilute acids. If dried at a temperature below
2 40°, they present the appearance of yellow transparent bodies, break-
ing with a vitreous fracture, which are soluble in water.

Their solutions are coagulated when heated to temperatures
varying between 65° and 73°.

(1) Serum-albumin. Specific rotation (a)0 = -56°. Not pre-
cipitated from its solutions when these are agitated with ether.

1 Hoppe-Seyler, Handbuch d. phys.- u. path.-chem. Analyse.

2 Weyl, "Beitrage zur Kenntniss thierischer und pflanzHcher Eiweisskorper." Zeit-
schrift f. physiol. Chem. , vol. i, p. 72.

3 Kiihne, Untersuchungen iiber das Protoplasma und die Contractilitat, Leipzig, 1864,
p. 317.

4 Frederique, "De 1'existence dans le plasma sanguin d'une substance albuminoide
se coagulant a + 56°." Annales de la Socie'te de Medecine de Gand, 1877.

Frederique, Recherches sur la constitution du Plasma sanguin, Grand, 1878, p. 25.
6 Hammarsten, "Ueber das Paraglobulin. Zweiter Abschnitt," Pfliiger's Archiv,
1878, vol. xviii. p. 67. According to the amount of salt present, and the greater or less

£

CHAP. I.] THE PROTEIDS. 17

(2) Egg-albumin. Specific rotation (a),, = -35° -5. Precipitated
from its solutions when these are agitated with ether.

CLASS II. Peptones: proteid bodies exceedingly soluble in water.
Solutions not coagulated by heat; not precipitated by sodium chloride,
nor by acids or alkalies. Precipitated by a large excess of absolute
alcohol and by tannic acid. In the presence of much caustic potash
or soda, a trace of solution of copper sulphate produces a beautiful
rose colour.

CLASS III. Globulins : proteid substances which are insoluble
in pure water, but soluble in dilute .solutions of sodium chloride;
their solutions are coagulated by heat; they are soluble in very dilute
hydrochloric acid, being converted into acid-albumins • they are
also readily converted by alkalies into alkali-albumins.

(1) Vitellin, not precipitated from its solutions when these are
saturated with common salt. Solutions coagulate at 70° — 75° C.

(2) Myosin, precipitated from its solutions in weak common
salt when these are saturated with sodium chloride. Solutions
coagulate at 55° — 60° C. Solutions in common salt not coagulated
by solution of fibrin-ferment.

(3) Fibrinogen, soluble in weak solutions of sodium chloride.
Precipitated from them completely by the addition of sodium chloride,
when this amounts to 12 or 16 per cent. Solutions coagulate on the
addition of fibrin ferment. Temperature of coagulation 56° C.

(4) Paraglobulin, soluble in weak solutions of sodium chloride.
From very weakly alkaline solutions paraglobulin is precipitated by
the addition of a very small quantity of common salt ; a further ad-
dition of this body leads to re-solution of the precipitate, which is
thrown down again when the amount of sodium chloride in solution
exceeds 20 p.c. -The precipitation of paraglobulin by sodium chloride
is never complete. Paraglobulin is completely precipitated when its
solutions are saturated with magnesium sulphate. Solutions not
coagulated by addition of fibrin-ferment. Temperature of coagulation
varies (according to amount of salts present and mode of heating)
between 68°— 80° C.; on-an average 75° C.

CLASS IY. Derived Albumins1 : proteid bodies insoluble in pure water
and iu solutions of common salt, but readily soluble in dilute hydrochloric
acid and in dilute alkalies. Solutions not coagulated by heat.

(1) Acid-albumins : obtained by the action of dilute acids (preferably
dilute hydrochloric acid) upon solutions of proteids, by action of strong
acids upon the solid proteids, and as first products in the action of gastric
juice upon proteids. On neutralizing solutions of acid-albumins, they are
precipitated even in the presence of alkaline phosphates. NaCl, added to
saturation, also precipitates them.

(2) a. Alkali-albumins or alkaline albuminates : obtained by the
action of dilute alkalies upon the proteids. Possess the properties of sub-
class 1, with the exception that in the presence of alkaline phosphates

rapidity of heating, the coagulation temperature varies, according to Hammarsten,
between 68° and 80°. , . ,

1 This convenient designation I borrow from Dr Michael Foster. See Text-book of
Physiology, Appendix.

o. : 2

18 PRODUCTS OF DECOMPOSITION OF PROTEIDS. [BOOK I.

the solutions are not precipitated by neutralization. When heated with
strong solution of caustic potash potassium sulphide is not formed.

/?. Casein, the chief proteid constituent of milk. Same properties as a,
but when heated with strong solution of caustic potash, potassium sulphide
is formed. In milk is coagulated by rennet.

CLASS V. Fibrin : Insoluble in water and in weak solutions of sodium
chloride. White elastic solid, usually exhibiting fibrillation when examined
under a high magnifying power; swells up in cold hydrochloric acid of *1 per
cent., but does not dissolve; when thus swollen dissolves with ease when a
solution of pepsin is poured over it. When heated for a great many hours
at 40° in dilute hydrochloric acid, it dissolves and the solution contains
acid-albumin.

CLASS VI. Coagulated Proteids : Insoluble in water, dilute acids
and alkalies. Give Millon's reaction. Are dissolved when digested at
350 — 40°, in artificial gastric or pancreatic juice, giving rise to peptones.

CLASS VII. Lardacein, so-called amyloid substance : Insoluble in
water, in dilute acids, in alkaline carbonates ; not dissolved by gastric juice
at the temperature of the body. Coloured brownish-red or violet by iodine.

SEC. 4. PRODUCTS OF DECOMPOSITION OF PROTEIDS.

The methods which the chemist follows in arriving at a know-
ledge of the constitution of a body are various ; his chief information
is derived from a careful study of the way in which the body is
decomposed under various circumstances, and of the structure and
amounts of the various products thus obtained ; subsidiary informa-
tion is derived from a consideration of physical properties, which
sometimes suggest analogies which otherwise would pass undetected.
The correctness of any view as to the structure of a body will be
tested by its being able, or not, to account for all known reactions,
and it will receive singular confirmation if it enable the experimenter
to effect the synthesis of the subject of speculation.

Great though the progress of organic chemistry has been, and
remarkable the development of our knowledge of the constitution
of bodies, we are yet far from being able to unravel the constitution
of such complex bodies as the proteids. We can therefore merely
record the results of laborious experiments which shew the pro-
ducts, or rather the classes of products, yielded by the proteids, and
scarcely venture to surmise what the exact constitution of the
proteids may be.

In the animal body, the proteids are ultimately subjected to
processes of oxidation of which the chief ultimate results are water,
carbon dioxide and urea ; what all the intermediate substances may
be we do not exactly know, though it is certain that glycine, leucine,
tyrosine and some other bodies are formed ; moreover it is certain that
substances destitute of nitrogen, such as carbohydrates, and also fats,
may take their origin in the decomposition of the proteids. Can

CHAP. I.] THE PROTEIDS. 19

these processes be imitated in the laboratory ? Only in part, indeed, for,
in spite of certain statements to the contrary, no one has given valid
proof of having, by an artificial oxidation, obtained urea.

The following are the chief facts which have been discovered in
reference to the decomposition of proteids ; after quoting these we shall
refer to some of the theoretical views to which they have given rise.

1. Action of water. When heated with water in sealed tubes at a
temperature of 100° C., the proteids are in part dissolved, the solution
afterwards undergoes decomposition, it being found to contain sulphur-
etted hydrogen, and a number of complex bodies of which some are
soluble in alcohol and ether (Gautier).

2. Action of heat. When subjected to dry distillation, the proteids
furnish the oily liquid long known as Dippel's oil, which contains
(1) ammoniacal salts of the fatty acids, as ammonium butyrate, valerate
and caproate; (2) amines, derived from the monatomic alcohols, viz.
methylamine, propylamin^ butylamine ; (3) aromatic compounds, as ben-
zine, aniline, phenol ; (4) picoline and lutidine, which are bases which
combine with the iodides of alcohol radicals to form compound ammonium
iodides.

3. Action of putrefaction. When exposed to the combined influences
of air and moisture, especially at a high temperature, the proteids yield
ammonia, ammonium sulphide, carbon dioxide, volatile fatty acids, lactic
acid, amines, leucine and tyrosine. Uncjer certain circumstances indol may
be formed.

4. Action of strong mineral acids and of caustic alkalies. Prolonged
boiling with sulphuric and hydrochloric acid and fusion with caustic
alkalies gives rise to products of which the chief are the same in the two
cases, viz. leucine, tyrosine, aspartic acid and glutamic acid.

When proteid bodies are treated with dilute acids they undergo hydro-
lytic decomposition, and certain definite compounds may be extracted from
the resulting mass. Their proportion is however small in comparison with
the by-products which we have no means of investigating.

The term "liydrolytic decompositions" has been applied by Hermann1 to designate
decompositions in which a body splits up after combining with the elements of water;
thus under various circumstances the neutral fats combine with the elements of water
and decompose into a fatty acid and glycerin, as shewn in the case of stearin by the
following equation:

^HiioO^ + 3H2 O = JJC18 H^ + CsH^V

Stearin. Water. Stearic acid. Glycerin.

Nasse first observed that the nitrogen in proteids appears to exist in two
conditions, as evidenced by the fact that a certain fraction of it is much
more unstable, apparently more feebly combined, than the rest. Schutzen-
berger has fully confirmed these observations. He heated proteids with
caustic baryta, in aqueous solution, up to 100° and collected the ammonia
given off in sulphuric acid. There separated a good deal of granular
matter, which increased as the reaction proceeded and which was found to
consist of carbonate, a little sulphate, oxalate, and phosphate, of barium.

1 Hermann, Elements of Human Physiology, 2nd English ed., p. 2. Smith, Elder
and Co., 1878.

2—2

20 PRODUCTS OF DECOMPOSITION OF PROTEIDS. [BOOK I.

The ammonia evolved, as well as the CO2 in combination with barium,
were estimated and found to be in the same ratio as would result if urea
were treated in a similar manner. Although the boiling was continued
for some time, the decomposition still progressed and only terminated
when the liquid was heated in sealed tubes up to 150° C. The relative
amounts of CO2 and NH3 still remained the same.

The substance thus treated did not give out any more ammonia, even
when heated to 200°. Nearly the whole of the resulting mass could be
got into a crystalline form, and Schiitzenberger was able to identify the
following substances1: — the elements of urea, (CO2 and NH3) : traces of
CO2, H2S, oxalic and acetic acid, tyrosine: amido-acids of the series
SH^ + NO2n corresponding to the fatty series Cn H2n O2, from amido-
aenanthylic to amido-propionic acid : leucine, butalanine, and amido-butyric
acid predominated. There were also obtained one or two acids nearly
allied to aspartic and glutamic acid, and one or two related to Bitthausen's
leguminic acid; furthermore a small quantity of a substance resembling
dextrin.

5. Action of hydrochloric acid and stannous chloride. When heated
with these reagents there are formed ammonia, aspartic acid, glutamic
acid, leucine and tyrosine.

6. Action of various oxidizing agents, a. When oxidized by means of
manganese dioxide and sulphuric acid, or potassium bichromate and
sulphuric acid, the proteids furnish bodies belonging to the aromatic and
fatty groups. Amongst others the following: benzoic aldehyde, propionic
aldehyde, propyl cyanide, benzoic acid, valerianic aldehyde, butyl cyanide,
hydrocyanic acid, acetic acid, propionic acid, valerianic acid.

b. By the action of nitric acid, there is first produced a yellow in-
soluble body (xanthoproteic acid) which dissolves on further action,
paroxybenzoic and oxybenzoic acids being ultimately formed.

c. When oxidized by means of chlorine, the proteids yield, amongst
other products, fiimaric acid, oxalic acid and chlorazol.

d. When heated with bromine and water, under pressure, there are
formed carbon dioxide, oxalic acid, aspartic acid, leucine, leucimide,
bromacetic acid, bromoform, bromanil and amidotribromobenzoic acid.

SEC. 5. THEORETICAL VIEWS AS TO THE CONSTITUTION OF THE

PROTEIDS.

" Under the most diverse influences : — action of water and strong
acids, action of bases, oxidations, putrefactions... &c. — the proteid
bodies when decomposed yield : firstly amides, such as glycocine and
leucine, containing radicals derived from fatty acids, or from the
homologues of lactic acid, as well as more complex amides, such as
aspartic acid, C4H7N04, the amide of malic acid, and glutamic acid,

1 Sohutzenberger, Bulletin de la Soc. Chimique, 15 F6vrier, 5 Mars, et 15 Mars,
1875.

CHAP. I.] THE PROTEIDS. 21

C5 H9 N04, which is a homologue of the preceding ; secondly, amides
having aromatic nuclei, such as tyrosine ; thirdly, amides containing
sulphur, such as cystine : fourthly, acids and aldehydes corresponding
to the radicals of the amides before mentioned. The proteid sub-
stances behave as amides containing both radicals of the higher homo-
logues of lactic and tartaric acids and residues of aromatic acids.
Hence" it follows that when the proteids are oxidized there is simul-
taneous production of fatty acids, of aromatic compounds and doubt-
less also of bodies analogous to urea.

" Though all the proteids when they are decomposed or oxidized
nearly always furnish the same products, they yet do not furnish
them in the same proportions. It must therefore follow that the
different radicals which they contain differ, not only in their arrange-
ments, but in their relative proportion, and in some cases even in
their nature1."

The views ^ is beyond the scope of this work to discuss

of Schtitzen- hypotheses as to the constitution of bodies unless these
berger2. appear to possess a legitimate interest to the biologist

or the physician. The speculations of Schiitzenberger can therefore
only be summarized in a few words. From the products obtained
by the action of caustic baryta upon the proteids (see pp. 19 and 20),
this author looks upon the proteids as complex ureids, i.e. as resulting
from combination in different proportions of urea with amido-acids,
some of which belong to the leucine series, others to the aspartic
series, whilst the more complex products of decomposition allied
to leguminic acid must be considered as resulting from complete
decomposition. Tyrosine represents the aromatic group, and is the
source of benzoic acid found amongst the products of the putrefactive
decomposition of proteids. When decomposed by means of caustic
baryta, he assumes that the molecule of albumin, which he represents
by the empirical formula C72 Hm N18 Ow S, yields, in addition to urea,
acetic acid, and some sulphur-containing body, a substance to which
he ascribes the formula C^ H132 N14 034 and which he admits may be
split up in various ways. Useful, nay indispensable as are such
hypotheses as suggesting lines of research to the actual chemical
worker, they possess no interest as yet to the biologist.

. ,3 It is in the cells of the organism that the processes

views^reTative take Place whose results are the external activities
to the consti- which it manifests; it is within the cells that the
tution of tiie oxidation processes of the economy have their seat.
Proteids. There is nothing more striking than the wide con-

trast which exists between the non-living proteid matter, say that of

1 Gautier, Chimie appliquee a la Physiologic, a la PatJwlogie et a VHygiene, tome
premier, p. 251.

2 Schiitzenberger, " Kecherches sur 1'Albumine et les maticres albummoides,
Bulletin de La Soc. Chimique, v. 23 and 24. ,,

3 Pfliiger, " Ueber die physiologische Verbrennung in den lebendigen Orgamsmen,
Pfliiger's Arcliiv, Vol. x1., p. 251.

22 PFLUGER'S VIEWS ON THE PROTEIDS. [BK. i., CH. i.

white of egg, and that which forms part of the living cell. The
former may be kept for years, the latter is continually decomposing
without any influence from without being necessarily exerted upon it.

The proteids which we consume as food are indifferent to neutral
oxygen ; so soon as they are taken up by organized cells they change
their character, by changing the structure of their molecules, and are
now subject to the influence of oxygen. The molecule of albumin
begins to live by breathing oxygen.

How thoroughly independent of an immediate supply of oxygen
very complex animal processes may be, which are essentially asso-
ciated with the metabolism of cell protoplasm, is, however, shewn by
certain remarkable experiments in which Pfluger introduced living
frogs into chambers containing no oxygen, and the temperature of
which was kept low, and observed that for many hours all the processes
of the organism continued to be performed.

How can we explain the immensely increased instability of
the living protoplasmic proteid matter as contrasted with non-living
proteid matter?

The assimilation of proteid matter is looked upon by Pfluger as
due to the formation of ether-like combinations between the proteid
of the cell protoplasm, and the proteid which serves as its food, water
being eliminated. In this process a living proteid molecule may
bind to itself a non-living, but isomeric, proteid molecule, and this
process of polymerism may be conceived to go on almost indefinitely,
so that a large and heavy mass may be produced out of, and yet
continue to exist as, a simple molecule1.

Pfluger inclines to the belief that in this process of assimilation by
the cell, proteid matter undergoes a change in its constitution, the
nitrogen passing from the state in which it exists in amides to the
more unstable condition in which it exists in cyanogen and its
compounds. In this way Pfluger explains why it is that in uric acid,
as in many other products of proteid metabolism — creatine, guanine,
&c. — cyanogen radicals are contained, whilst none of these decompo-
sition products are to be obtained from non-living proteids.

Bodies (so -"-n concluding this sketch of the proteids, it must be
called Albu- mentioned that there occur in the epithelial and con-
minoid) re- nective tissues of the organism certain bodies which
lated to the have somewhat close relationship to the proteids, but
which are nevertheless distinct from them ; these are
chondrin, collagen and gelatin, mucin, elastin, keratin. They will be
considered in detail in future sections of this work.

1 The Author understaiids Pfluger to say that the same constituent atoms or groups
of atoms (radicals) mhst be present in different proteids : that the difference is caused
either by the final molecule being a different multiple of the same group or groups of
atoms (polymerism) or by the oxygen or nitrogen occupying different relative positions
with respect to groups of atoms which they serve to link together (metamerism), or by
differences in the relative position of groups of atoms or their constituent parts with
respect to one another (general isomerism).

CHAPTER IT.

THE BLOOD.

SEC. 1. THE PHYSICAL CHARACTERS OF THE BLOOD.

Physical THE blood as it circulates in the vessels of man
and vertebrates generally is a viscous, and to the naked
eye homogeneous liquid of red colour : the blood of the pul-
monary veins, of the left side of the heart, and of the systemic
arteries being normally of a bright scarlet hue, and the blood
of the right side of the heart, of the systemic veins, and of the
pulmonary artery being of a brownish-red colour. On exposure to
air or to oxygen the brown-red colour of venous blood soon changes
to scarlet, and this change takes place most rapidly when the blood
and gas are shaken up together.

In order to collect for purposes of analysis or demonstration pure arterial
or venous blood, or both, so as to avoid contact with air, the following
apparatus or some modification of it may be employed : —

A and B are two glass tubes of about 100 c.c. capacity, which at their
lower extremities are connected by means of elastic tubing with a forked
tube 0, to which is attached the elastic tube D, which at its other end is
connected with the glass bulb 7?, having a capacity of about 250 c.c. At
their upper extremities, A and B have connected with them two glass stop-
cocks, the tubes leading from which are of narrow diameter ; it is convenient
that these tubes should be of such a size that india-rubber tubing of
narrow diameter can easily be attached to them. The tubes are fixed in
two separate iron clamps such as are shewn in the drawing, attached to a
firm upright rod of iron with a firm stand. The reservoir R is also
held by a similar clamp, which can easily be attached either to the top or
to the bottom part of the upright rod, so as to place it above or below any
given level in relation to the tubes A and B.

The reservoir being, say, in the lower position indicated in the figure,
mercury is poured into it so as to fill it. It is next undamped and raised

24

MODE OF COLLECTING BLOOD.

[BOOK i.

so that its lower part is above the level of the stop-cocks of A and B.
These are now opened, mercury rises into the tubes, driving the air
which they previously contained before it; when the tubes are filled and
a stream of mercury is issuing from them, the stop-cocks are closed. In
order to determine whether the stop-cocks do not leak, the reservoir R
may now be held in the hands of the experimenter at thirty-five or forty
inches below the stop-cocks of A and B. The mercury in these tubes will
naturally fall at first and then remain steady: on raising the reservoir
cautiously the metal should however rise and fill the tubes completely.

Fm. 8. APPARATUS FOR COLLECTING BLOOD OVER MERCURY WITHOUT ALLOWING IT

TO COME IN CONTACT WITH AIR.

In order to collect apart arterial and venous blood, glass cannulae,
to which are attached narrow elastic tubes of considerable length, are in-

CHAP. II.] THE BLOOD. 25

serted into the artery and vein of the animals to be experimented upon ',
which should be deeply anaesthetized2* The clips which control the entrance
of blood from the cardiac side of the arterial and the distal side of the venous
cannula having been removed or opened, blood is allowed completely to fill
the elastic tubes attached to the cannulae, which are held at a fairly high
level so as to allow the blood to rise and expel the air before it. The
instant the tubes are filled their open ends are slipped over the ends
of A and B.

The reservoir R having been placed in its lower position, the stop-cocks
of A and B are opened ; blood will then flow from the artery and vein into
the respective tubes. As soon as enough has been obtained the stop-cocks
are closed, and the tubes are simultaneously shaken by assistants so
as to defibrinate their contents. On placing the two tubes side by side
the contrast between the colour of arterial and venous blood will appear
most striking.

A detailed description of this procedure has been given as, mutatis
mutandis, it illustrates the method in which blood can be obtained from
blood-vessels without being brought in contact with air, not only for
purposes of class demonstration, but also in researches on the gases of the
blood..

Where it is required to keep the blood for some hours, as for example
in order to make repeated analyses, one or both tubes may be taken out of
their respective clamps and laid in troughs containing broken ice. In some
cases it is desirable to obtain two separate samples of the same blood ; in
such cases the free upper ends of A and B have attached to them a T tube,
to which is connected the elastic tube leading to the artery or vein. The
blood-stream will then divide itself equally between the two "tubes.

Although to the naked eye the blood appears to be a homo-
geneous red liquid, it is found on microscopic examination to
consist of a colourless fluid — the so-called liquor sanguinis, or
plasma of the blood — holding in suspension large numbers of solid
bodies, the coloured and colourless corpuscles of the blood. It is
the former of these which preponderate very greatly over the latter,
and which by the colouring matter, haemoglobin, of which tbey
mainly consist, confer upon the blood its red colour ; the shade
of this at any time depends, as will be shewn in the sequel, chiefly

1 Handbook for Physiological Laboratory, p. 212.

2 The Author would very strongly recommend all experimenters who have occasion
to perform experiments upon the lower animals, and especially dogs, to employ as
the chief means of producing insensibility to pain, subcutaneous injections of morphia.
Solutions of bimeconate of morphia may be obtained which contain as much as
two grains in half a drachm. As large a dose as two grains of the bimecoiiate
may with perfect safety be injected under the skin of a dog of medium size; the
injection is followed in about half an hour by salivation and by a staggering gait, and
then by deep somnolence. In this state the animal is quite passive, and may without
a struggle and without any fear being evinced on its part, be properly fixed, and then
rendered completely insensitive to pain by the administration of ether or chloroform ;
as was pointed out by Claude Bernard, under these circumstances chloroform anaesthesia
is induced with remarkable ease, and persists for a long time. This method not only
abolishes the fear which often must constitute the most important part of the pain in-
flicted by a physiological experiment, but in those rare cases where the animal must be
allowed to recover after the experimental proceeding has been carried out, the long period
of narcotism which succeeds it secures the absolute and beneficial rest of the animal.

26 SPECIFIC GRAVITY AND REACTION OF BLOOD. [BOOK I.

upon the chemical relations of the colouring matter to oxygen, though
in part also upon the shape of the coloured blood corpuscles, which is
subject to various physical influences.

The specific gravity of the living blood cannot for obvious reasons
be ascertained ; that of defibrinated human blood drawn from
healthy subjects has been found to vary between 1045 and 1062 *,
the average being 1055 ; greater variations than are indicated by
the above numbers are however consistent with health, the widest
limits being probably indicated by the numbers 1045 — 1075.

The mean specific gravity of the blood of the dog was found by
Pfluger to be 10602, and by Nasse to be 10593 ; that of the blood of
the rabbit was found by Gscheidlen to vary in three cases between
1042 and 1052.

As blood is drawn from a vessel it is found to vary slightly in
density, that drawn first having a somewhat higher specific gravity
than that which follows, owing to the quantity of water of the blood
increasing as a result of haemorrhage4.

Reaction. Blood always possesses a feebly alkaline reaction, which
rapidly diminishes from the time of its being shed to the time of its
coagulation.

The red colouring matter of the blood interferes with the ready determina-
tion of the reaction as by simply immersing ordinary test-papers into the
fluid, and therefore one or other of the three following methods may be
employed, of which the second and third, b and c, are to be preferred.

(a) Kuhne's Method5 consists in placing a drop of blood in a
specially constructed tiny dialyzer of parchment-paper; this is then immersed
in a drop of water contained in a watch-glass. After a short interval the
reaction of the water is determined by means of litmus paper.

(b) Liebreich's Method0. Plaster of Paris absolutely free from
alkaline reaction is cast into thin slabs, which are then dried, and
afterwards coloured by dropping upon them a perfectly neutral solution of
litmus. When a droplet of blood is allowed to fall upon the coloured slab,
the fluid of the drop is soon absorbed by the porous gypsum whilst
the corpuscles are left. On placing the spot under a stream of water,
the corpuscles are washed away and the colour of the slab at the
site of the blood spot is found to be a more or less deep blue.

(c) Zuntz's Method7. This method rests upon the fact that the

1 Becquerel et Rodier, Recherches sur les alterations du sang. Paris, 1844.
3 Pfluger, "Ueber die Ursache der Athembewegungen, sowie der Dyspnoe und
Apnoe." Archiv d. gesammten Physiologic. Bd. i. (1868) p. 75.

3 Nasse, Haematologische Mittheilungen. Quoted by Gscheidlen, Physiologische
HethodiTc, p. 328.

4 Becquerel et Rodier, Traite de Chimie Paihologique, appliquee d la Medecine pra-
tique. Paris, 1854, p. 41 et seq.

& Kuhne, "Ein einfaches Verfahren, die Reaction hamoglobinhaltiger Fliissig-
keiten zu priifen." Virchow's Archiv, vol. xxxui. (1865), p. 95.

6 Liebreich, " Eine Methode zur Priifung der Reaction thierischer Gewebe. " Berichte
d. deutschen c/iem. Gesellsch. zu Berlin, 1868, p. 48.

7 Zuntz, Centralblatt, 1867, ' No. 34. See also Adam Schulte, Ueber den Einflms
des Chinin auf einen Oxydatioiuprocess im Blute. Inaugural Dissertation. Bonn, 1870.
p. 9 et seq.

CHAP. II.] THE BLOOD. 27

blood colouring matter does not diffuse out of the blood corpuscles into
solutions of common salt of considerable strength. Litmus paper is
moistened with a strong solution of salt and a drop of the blood to be
tested is placed upon it ; after a few seconds a drop of the same salt
solution is placed over the drop of blood ; the liquid is then sucked up by
means of filter paper. By following this method the blood can be so
removed from the test-paper that the colour of the latter may be readily
observed. The litmus paper to be used for this purpose must be highly
glazed and the tincture of litmus used in its preparation must have been
neutralized with acid until its colour is violet.

By adding standard solutions of acids to blood, and employing the
above method for ascertaining when the reaction became faintly acid,
Zuntz determined the previously mentioned diminution of alkalinity
of blood removed from the body.

The Pheno- As it circulates in the blood-vessels of the living body,

agnation C°~ tne blood consists, as we have said, of a liquid, the so-
called liquor s. plasma sanguinis (often designated
blood-plasma, or more shortly the plasma), holding in suspension the
blood corpuscles. Within a short time of its being shed — usually
between two and six minutes — the process known as coagulation
commences — a process in which the blood passes first into the state
of a soft red jelly, which gradually acquires greater consistence,
and which, by a contraction of one of its constituents, expresses a
fluid — the serum, which surrounds the clot, and in which the latter
often ultimately floats.

If we desired to ascertain the exact time when this coagulation
commenced in a sample of blood, we should collect it in a watch-glass
and at very short intervals pass a needle through the liquid ; as soon
as coagulation had set in the needle would, in its passage through
the fluid, entangle itself in the newly formed jelly, which would then
be apparent on drawing the needle out.

When blood coagulates, the process usually commences on the
surface of the liquid and then near the sides of the vessel which
contains it, the newly formed coagulum having in the former case the
appearance of a pellicle. Very rapidly, however, the process invades
the whole mass of the blood, which then presents the appearance of a
soft, easily broken, jelly. Soon this acquires greater consistence, so
that the blood has, as it were, taken a cast of the vessel which
contained it, adhering closely to its sides and permitting of the vessel
being inverted without any escape of the contents; at the same time
drops of serum begin to transude from the clot. This transudation
of serum is brought about by the contraction of the clot and continues
for a time varying between ten and forty-eight hours, at the end of
which the clot is found to be surrounded by serum. According to
Nasse, the first stage of coagulation (characterized by the formation
of a pellicle) commences in the blood of men in about 3 minutes
45 seconds, in that of women in 2 minutes 50 seconds; the second
stage, in which not only the surface but the portions of blood next to

28 THE PHENOMENA OF COAGULATION. [BOOK I.

the walls of the vessel have become gelatinized, occurs on an average
in the blood of men in 5 minutes 52 seconds, and in that of women
in 5 minutes 12 seconds; the third stage, in which the blood has been
converted throughout into a soft jelly, is usually developed in the
blood of men in 9 minutes 5 seconds, and in that of women in
7 minutes 40 seconds; the fourth stage, of complete solidification
with obvious commencement of transudation of serum from the clot,
occurs in the blood of men in about 11 minutes 45 seconds, and in
that of women in 9 minutes 5 seconds1.

This process of coagulation is due to the separation from the
plasma of a body called Fibrin, which entangles in its meshes the
corpuscles of the blood, the mechanical interlocking of the corpuscles
by the threads of fibrin giving rise to the crassamentum or blood clot.

The blood of certain animals coagulates more rapidly than that
of others : we might with fair accuracy arrange the blood of various
common domestic animals in the following order, according to the
rapidity of coagulation, the first-named coagulating most rapidly —
rabbit, sheep, dog, ox, horse ; in the latter animal coagulation com-
mences usually between five and ten minutes after the blood is shed.
If human blood were included in the above list it would immediately
precede that of the ox.

When the commencement of coagulation is delayed for several
minutes — as it normally is in horse's blood, and as it usually is in
the blood of men and other animals when suffering from inflammatory
diseases — the blood corpuscles, being specifically heavier than the
plasma, have time to subside partially before coagulation commences,
so that the uppermost layers of such blood if undisturbed are
nearly free from coloured corpuscles ; subsequently when the blood
coagulates, the clot exhibits the phenomenon of the huffy-coat,
'inflammatory crust/ or crusta phlogistica, i.e. the upper part of the
clot is of a yellowish colour ; in the lower strata of the buffy-coat are
found large numbers of colourless corpuscles, which being specifically
lighter than the red have not time to sink as far as the latter before
coagulation occurs. The formation of the buffy-coat, though in part
due to slow coagulation, is dependent greatly upon the blood cor-
puscles aggregating so as to form little clumps, which more readily
overcome the resistance offered by the fluid and therefore sink more
readily than individual corpuscles.

If instead of allowing blood to coagulate undisturbed, it be
stirred or whipped with twigs immediately after it is shed, the process
of coagulation is modified. The fibrin generators unite to form fibrin,
but this does not entangle the blood corpuscles ; it separates as a
stringy mass, which adheres to the instruments which have been
used to stir the blood, whilst the blood corpuscles remain suspended
in the serum, the mixture being designated defibrinated blood.
Defibrinated blood differs from the living blood which has yielded it,

-• Nasse, Article Blut, Wagner's Handtvorterluch d. Physiologic, Vol. i. pp. 102, 103.

CHAP. II.] THE BLOOD. 29

merely in having lost the fibrin-generators, which have united to form
fibrin.

circum- ^he following circumstances hasten or promote co-

stances which agulation :

hasten Co- a. Exposure to a temperature higher than that

agulation. of tlie living body (Hewson1, Hunter2, Thackrah3,

Scudarnore4, Davy5, Gulliver6), but probably not exceeding 52° C. or
54° C.

b. Contact with foreign matter: thus the time of coagula-
tion will be affected by the shape of the vessel in which blood is
collected, the process occurring sooner where a large surface of blood
is in contact with the vessel, as for example when it is allowed to
flow into a wide shallow vessel. The influence of foreign matter ia
promoting coagulation will be again referred to.

c. Closely connected with b. is the effect of agitation, which,
as Hewson7 and John Hunter8 shewed, and as has been fully con-
firmed, hastens coagulation.

d. The dilution of blood with not more than twice its volume
of water (J. Hunter9, Prater10).

e. The addition of minute quantities of sodium chloride,
sodium sulphate or other neutral salt (Ancell11).

Conditions The following circumstances hinder or suspend co-

which retard agulation .__

or suspend ^ -, ,

Coagulation. a- Exposure to a low temperature.

Blood which is rapidly reduced to the temperature of
melting ice does not coagulate (Davy once kept blood fluid for one hour
at 0° C.) : it may be frozen and remain in a frozen condition for hours
without losing its power, of coagulating when thawed (Hunter12,
Hewson13, Davy14). It may be frozen and thawed several times in
succession without coagulating or losing its property of coagulating
(Davy).

1 Hewson, Properties of the Blood, p. 3. The Works of William Hewson, F.E.S.
edited with an introduction and notes by George Gulliver, F.K.S. London, printed for
the Sydenham Society, 1846.

2 Works, edited by Palmer, iii. 26, 110.

8 Thackrah On the Blood, ed. 1834. Exp. 44, 45, 50, 51, 52, 56.
4 Scudamore On the 'Blood, p. 20. 8vo. London, 1824.

3 Davy, Researches, Physiological and Anatomical. London, 1859, Vol. 2, p. 78.
0 Gulliver, Hewson 's Works, p. 4. Note in.

7 Hewson, op. cit., p. 15.

8 Hunter, Works, ed. by Palmer, Vol. in. 31.

9 John Hunter, General Principles of the Blood, at p. 135 of Vol. in. of Palmer's
edition, of The Works of John Hunter.

10 Prater, Experimental Inquiries in Chemical Physiology, p. 81. Part I. 'On the
Blood.' London, 1832.

11 Ancell, Course of Lectures on the Physiology and Pathology of the Blood, &c.
Lecture VII. Lancet, 1839-40, p. 522.

12 Hunter, Works of, by Palmer, Vol. in., p. 67.

13 Hewson, op. cit., p. 17.

14 Dr John Davy, op. cit., Vol. n., p. 75.

30 CIRCUMSTANCES WHICH RETARD COAGULATION. [BOOK T.

The following is the best method of exhibiting this fact for purposes
of class demonstration :

A small platinum crucible, or still better, as permitting more easily of an
examination of its contents, a small platinum basin is immersed in a vessel
containing a mixture of ice and salt ; a frog is then decapitated, and the
blood is allowed to flow into the frozen vessel, where it instantly congeals.
The platinum vessel can be taken out of the ice and held up so as to shew
the hard frozen drops of blood. The experimenter then places the vessel on
the palm of his hand, the heat of which almost instantly thaws the blood,
which can then be dropped into a watch-glass. The platinum vessel
is again placed on the ice and the thawed blood transferred to it, to be
frozen a second time. This freezing, thawing, and transference from one
vessel to another may be repeated several times ; at last the blood is
allowed to remain in the watch-glass, when after a few minutes it sets into
a firm jelly.

b. Contact with the living tissues.

If a vein be exposed and ligatures be applied to it so as to
confine a quantity of blood within it, and it be then cut out of the
body, it will be found that on opening the vein after an hour the
blood will still be fluid, though after contact with foreign matter it
will coagulate in a few minutes (Hunter1, Hewson2). For some hours
after somatic death the blood remains fluid in all vessels except the
heart and principal trunks, provided that the vessels have been pre-
viously healthy. Blood will remain fluid for hours in a vein after
being exposed with the utmost freedom to the air by being poured
in a thin stream from one vein to another (Lister3).

c. The addition of a sufficient quantity of sodium chloride,
sodium sulphate, potassium nitrate or some other neutral salts
(Hewson4, Davy5), will prevent coagulation, which will however
occur subsequently if a sufficient quantity of water be added.

Thus to quote Hewson's own words, " if six ounces of human blood
are received from a vein upon half an ounce of true Glauber's salt
reduced to a powder, and the mixture agitated so as to cause the
salt to be dissolved, that blood will not coagulate on being exposed
to the air, as it would have done without the salt ; but if to this
mixture about twice its quantity of water be added, in a short time
the whole will be jellied or coagulated, and on shaking the jelly,
the coagulum will be broken, and the part so coagulated can now be
separated as it falls to the bottom and proves to be lymph" (i.e. fibrin).

1 Hunter's Works, by Palmer, Vol. in., p. 29.

2 Hewson's Works, p. 22.

3 Lister, "On the Coagulation of the Blood ;" the Croonian Lecture for 1863. Pro-
ceedings of the Royal Society, Vol. xn. p. 580.

4 Hewson, op. cit., p. 11 et seq.

* Davy, Researches, Vol. n. 101-2.

CHAP. II."| THE BLOOD. 31

SEC. 2. THE LIQUOR SANGUINIS. FIBRIN AND ITS SUPPOSED

PRECURSORS.

The Liquor Sanguinis.

Methods of It has already been stated that in the living

obtaining Li- blood the corpuscies float in a fluid termed the liquor
ouor San- •• » 11 i 111 i •

guinis. sangmms or plasma, and that when blood coagulates it

does so in consequence of the separation from the
plasma of a proteid substance termed fibrin. We have now to
describe the mode of obtaining liquor sanguinis, to describe fibrin,
to examine the bodies which the plasma contains, and to examine
the facts which relate to the separation from it of fibrin.

Almost as soon as the liquor sanguinis is withdrawn from the
living vessels, it undergoes that change which results in the separa-
tion of fibrin and serum. The change may however be hindered by
various methods, which may be employed to furnish us with plasma
for examination.

1. In order to obtain plasma in a state of great purity, blood
must be rapidly cooled to a temperature approaching that of melting
ice, at which temperature its coagulation is, as has been already
stated, deferred.

The blood of most animals coagulates so rapidly that it is difficult
to cool any considerable quantity of blood to a temperature at which
coagulation would be long deferred, before the process has actually
occurred. The blood of the horse or donkey, however, usually
coagulates so slowly that with the aid of suitable contrivances con-
siderable quantities may be cooled to near 0° C. before coagulation has
had time to occur ; and once at that temperature the process of co-
agulation may be long postponed.

Under these circumstances the corpuscles sink pretty rapidly,
tending to form a sediment at the bottom of the vessel in which the
blood was received, and leaving an upper stratum of liquor sanguinis
perfectly free from red colour. The liquor sanguinis, decanted from
the corpuscles and exposed to a temperature favourable to coagula-
tion, exhibits the phenomena which have been described as character-
izing the coagulation of the blood, save that the coagulum is
colourless. If the fluid be stirred with twigs there will separate
from it stringy fibrin exactly similar to that obtained by similar
treatment from blood, save in the absence of colour derived from
entangled blood corpuscles.

A convenient contrivance for collecting considerable quantities of plasma
from the blood of the horse is shewn in the annexed figure, and was sug
gested by Dr Burdon Sanderson ' . The apparatus consists of a vessel .with

i Handbook for the Physiological Laboratory, p. 168.

32

METHODS OF SEPARATING LIQUOR SANGUIN1S. [BOOK I.

three concentric compartments. Into the central and external of these
are placed small lumps of ice, whilst into the intermediate compartment
blood is received as it issues from the vessels of the animal. The middle
compartment being very narrow (its width, not exceeding half an inch) the
whole of the liquid, which it contains, is rapidly reduced to the temperature
of melting ice. In the course of about two hours the corpuscles have
subsided to the lower part of the partition containing the blood, and
considerable quantities of pure plasma may be drawn off, with the aid of a
syphon or pipette.

--3/4

FIG. 9. DK SANDERSON'S APPARATUS FOR COLLECTING LIQUOR SANGUINIS. (Hand-
book for the Physiological Laboratory.}

2. Plasma may be much more easily obtained, though mixed
with water and saline matters, by mixing blood, immediately on its
being shed, with solutions of certain neutral salts of sodium, potas-
sium or magnesium, or by dissolving suitable quantities of such
salts in the blood before coagulation has occurred. From such
mixtures of blood and neutral salts the corpuscles separate by sub-
sidence, and the plasma may be obtained by decantation or filtration.
The following are the proportions in which sodium sulphate arid
magnesium sulphate, which are the salts chiefly employed, should
be added to blood in order to prevent coagulation and lead to the
separation of the liquor sanguinis.

a. One part of finely powdered sodium sulphate is added to
12 parts of blood and the powder is gently stirred with the blood to
hasten its solution. Instead of employing the solid salt in the

CHAP. II.] THE BLOOD. 33

above proportions (Hewson's method1), it is more usual to mix the
blood with a saturated solution of the salt; the blood is received
directly into a vessel, which contains }th of its volume of a saturated
solution of sodium sulphate2, and the two liquids are gently mixed.

b. Magnesium sulphate, as has been shewn by Schmidt3,
Semmer4 and by Hammarsten, is decidedly preferable to sodium
sulphate for hindering the coagulation of the blood and for yielding
a plasma suitable for experimental researches on the formation of
fibrin. According to Semmer four parts of blood are mixed with
one part of a solution of magnesium sulphate containing 25 p. c. of
the salt. According to Hammarsten5 the blood is mixed in the
same proportion with a saturated solution of magnesium sulphate6.

In addition to the substances, which, when added in suitable
proportions, prevent the coagulation of the blood, there are others
which merely postpone its occurrence and facilitate the separation
of blood corpuscles from the plasma. Thus when frog's blood is
mixed with its own volume of a ^p.c. solution of cane-sugar, the
corpuscles may be separated from it by filtration, and there passes
through the filter-paper a clear fluid which consists of plasma
diluted with solution of sugar, which coagulates after a short
interval. This method of separating the blood corpuscles from the
plasma was suggested by Johannes Muller7.

In relation to the action of neutral salts in hindering the coagula-
tion of the plasma it must be remembered that these substances
only exert their action when present in certain proportions ; if added
in too small quantities to blood, coagulation occurs, and if sufficient
water be added to blood or plasma which has been kept from
coagulating, the process sets in. Thus, as Hewson shewed, if to blood
which has been maintained in a fluid state by the addition of solid
sodium sulphate in the proportions previously mentioned, there be
added twice its volume of water, in a short time the whole will
coagulate.

Properties Plasma, obtained by subjecting blood to a low

temperature, is a viscous liquid possessing the same
colour as the serum which separates from the blood
of the same animal after coagulation ; if kept at a temperature below
5° C. it may be filtered from any colourless corpuscles floating in it8.

Hewson's Works, p. 11.
Denis, Memoire sur I'e sang, 1859, p. 31.
A. Schmidt, Haematologische Studien. Dorpat, 1865, p. 44.
Semmer, quoted by Gscheidlen, Physiologische Methodik, p. 342.
Hammarsten, "Zur Lehre von der Faserstoffgerinnung," Pfliiger's Archiv, Vol.
xiv. (1877) p. 220.

6 Many other neutral salts may be employed instead of those previously mentioned,
as was shewn by Hewson, Gulliver and Davy. The reader will find much valuable
information on this subject in Gulliver's edition of Hewson's Works, p. 12, and in
Davy's Researches, Vol. n., p. 101.

7 Joh. Muller, "Beobachtungen,zur Analyse der Lymphe, des Blutes und des Chylus."
Poggendorff 's Annalen, Vol. xxv. (1832) p. 540.

' Alex. Schmidt, Pfliiger's Archiv, Vol. xi. (1875) p. 318.

G. 3

.34 PROPERTIES OF LIQUOR SANGUINIS. FIBRIN. [BOOK I.

The specific gravity of plasma doubtless differs imperceptibly
from that of the serum which separates from it, and which in the
case of man varies between 1026 and 1029. It is stated by Gautier
that the density of human plasma varies between 1027 and 1028,
though no authority for the statement is given1.

The reaction of the plasma is, like that of the blood, and of the
serum which separates from it after coagulation, alkaline.

The coagulation of the liquor sanguinis, which may be readily
watched by allowing the temperature of the fluid separated from
horse's blood at 0° C. to rise slightly, follows exactly the same course
as the coagulation of the blood. The process commences on the
surface and sides of the liquid and then extends throughout the whole
mass, which assumes the appearance of a colourless trembling jelly ;
the surface of this jelly is from the first seen to be somewhat
depressed, and from it there exude droplets of clear serum. After
some hours the coagulum is found to have contracted and floats
in serum exactly as does a blood clot under similar circumstances; in
the case of the coagulation of plasma, however, the coagulum, as it does
not entangle blood corpuscles, is colourless and comparatively small.

The serum is found to be more alkaline than the plasma from
which it has separated.

The plasma, it has already been remarked, differs from the serum
in its containing the body or bodies which, separating from it, form
fibrin. It will be convenient therefore to examine first of all the
properties of fibrin and then to consider the facts which relate to
the assumed precursors of fibrin in the plasma.

Fibrin.

Micro- When a drop of freshly drawn blood is examined under

scopic ob- the microscope in the usual way, filaments are often ob-

onThe OI served to stretch across the preparation ; these are usually

character onty seen under tolerably high powers and by careful focus-

and ar- ing ; the filaments consist of the newly formed fibrin. If

range- a pretty thick stratum of frog's blood be mounted for

JS?*? °f microscopic examination in the usual way, the edges of the
Fibrin in .r , . , . . . d __ J>

blood clot, preparation being touched with paraffin to prevent evapo-
ration, after some hours the coloured corpuscles are seen
to have arranged themselves into patches, the corpuscles in each
patch appearing to radiate from a centre, at which are seen minute
granulations. Under a sufficiently high power each individual blood
corpuscle is seen to have assumed a pear shape. The appearances
alluded to, which have been admirably described by Ranvier2, are
due to the contraction of filaments of fibrin, which have the afore-
mentioned granulations for their centre. The actual arrangement of

1 Gautier, Chimie applique'e a la Physiologic, 1874, Vol. i., p. 489.

2 Ranvier, Traite technique d'Histologie, p. 214 et seq.

CHAP. II.]

THE BLOOD.

35

fibrin in the clot of human blood can be admirably and easily shewn
by- following the method also described by Ranvier. A pretty large
drop of human blood (obtained by pricking the finger) is treated as
was mentioned in the case of frog's blood. After some hours, the
paraffin is scraped off, the cover-glass is lifted, and the coagulum of
blood which adheres to the slide or cover-glass, or to both, is subjected
to the action of a gentle stream of water. Ranvier allows the water
to flow out of a pipette, but the Author finds that a very small stream
at very low pressure from a water tap is even preferable. After all
the red colour has disappeared, a drop of a strong solution of magenta
is placed upon the site of the former blood clot ; this is then covered
with a covering glass and examined. The preparation is then, seen to
be covered by reticula, each of which appears to radiate from a cen-
tral granulation. The granulations as well as the fibres are stained
by magenta and by solution of iodine, but not by carmine or picro-
carmine. These granulations will be further referred to in con-
nection with the part which the formed elements of the blood play
in its coagulation.

Fro. 10. EETICULUM OF FIBRIN FROM: THE BLOOD OF MAN. 500 diain. (Eanvier.)

Mode of Fibrin may be obtained either from blood or from

separating liquor sanguinis, either by allowing these fluids to coagu-
Fibrin for late at rest, or by stirring them with twigs, or by agitating
chemical them with small pieces of metal or glass. The fibrin, obtained
examination. by stirring blood, adheres to the instrument employed; it
is at first deeply stained with blood, but by washing in a stream of
water it gradually loses its red colour and presents the appearance of
a white, stringy, elastic body.

When obtained by the first method from plasma, the coagulum
at first presents a gelatinous appearance ; if, however, the coagulum
be placed in a cloth and be kneaded with water, as the serum is
squeezed out, there is left fibrin in the form of a white stringy solid.

3-2

36 PROPERTIES OF FIBRIN. [BOOK I.

It is in the latter form that fibrin always separates from blood when
it is stirred or shaken with foreign matters. When dried, fibrin
presents the appearance of a greyish white solid. In order to purify
fibrin it is carefully dried at a temperature not exceeding 110°C.,
and is then reduced to powder; the powder is successively and
repeatedly treated with water holding hydrochloric acid in solution,
with alcohol and with ether. However carefully the process of
purification may be carried out, fibrin always retains a small quantity
of inorganic salts amounting to about 0*9 in 100 parts.

Properties Fresh fibrin is an elastic substance, as evidenced by the
of Fibrin. way jn which serum is squeezed out of the clot which forms
in plasma or blood.

Fibrin belongs to the group of proteid or albuminous substances,
from the majority of which it differs in that once formed it is in-
soluble in pure water, though it has not been subjected to the action
of heat OF acids or metallic salts.

Fibrin has the following elementary composition ; C, 52'6 : H, 7'0 :
N, 17'4 : S, 1-2 : 0, 21'8.

Freshly prepared moist fibrin is soluble in a 6 per cent, solution
of potassium nitrate, if digested with it for some time at a tempe-
rature of 30° or 40°. It is similarly soluble in solutions of sodium
chloride, and in a 10 per cent, solution of magnesium sulphate. The
solutions of fibrin in the neutral salts are coagulated by heating to
60° or 65°, by the addition of acids and of alcohol, and by the addition
of powdered magnesium sulphate.

Denis asserted that fibrin obtained from arterial blood is not
soluble in 10 per cent, solutions of the neutral salts, whilst that
obtained by stirring venous blood is soluble in the same solutions.

When placed in water containing about 5 parts of hydrochloric
acid in 1000, moist fibrin swells into a transparent jelly, which does
not dissolve. In water containing 1 part of hydrochloric acid per
1000, fibrin dissolves in a few hours, at a temperature of 40° C. The
fibrin is in this process converted into so-called acid-albumin or
syntonin. Solutions of syntonin are not precipitated when they are
boiled ; when they are carefully neutralized, the proteid which had
been dissolved is thrown down in the form of gelatinous flakes which
are insoluble in water, but are readily soluble in dilute solutions of
acids, of alkalies and alkaline carbonates. Acetic and phosphoric acids
exert a similar action to hydrochloric acid. From the acetic solution
of fibrin, potassium ferrocyanide throws down a white precipitate.

When digested at the temperature of the animal body in dilute
solutions of ammonia, or of potassium or sodium hydrate, fibrin
dissolves, and the solutions are not coagulated by heat, but are pre-
cipitated by mercuric chloride, lead acetate, and copper sulphate.

Fibrin possesses the power of decomposing solutions of hydric
peroxide, H20a, which enter into effervescence, owing to the libera-
tion of oxygen ; if it be first immersed in a tincture of guaiacum
and afterwards in a solution of hydric peroxide or in a mixture of

CHAP. II.] THE BLOOD. 37

the two reagents, it assumes an intensely blue colour. This is due to
the oxidation of the resin of guaiacum by the oxygen which the
fibrin has liberated from the peroxide.

Quantity of Human venous blood in health yields from 2 '2 to

fibrin in the 2 '8 parts of fibrin per 1000, and it is said that arterial
blood- yields somewhat more than venous blood.

The Assumed Precursors of Fibrin in the Liquor Sanguinis.

1. Serum-Globulin or Paraglobulin. (Schmidt's fibrinoplastic

substance.)
i

Schmidt'a When plasma is diluted with ten or fifteen times its

methods of volume of ice-cold water and subjected to the action of
Paraglobulin a stream of carbon dioxide1, or when it is carefully
neutralized with acetic acid, the liquid soon becomes
turbid, and deposits after some time a proteid substance to which the
above terms have been applied, of which the first indicates its resem-
blance to a proteid contained in the crystalline lens to which the
name of globulin was long ago ascribed; and the second the property
which has been ascribed to it of inducing, under certain circumstances,
the separation of fibrin from solutions containing fibrinogen.

The quantity of dilute acetic acid (25 per cent.) to be added is
4 drops for every 10 c.c. of serum diluted with 150 c.c. of H20.

As the body which is precipitated under these circumstances is
not only contained in the plasma but also exists in the serum, the
latter much more readily available fluid may be employed for its
preparation.

The same substance it is2, which is precipitated when blood serum
is subjected to dialysis (see p. 6), a process which may be employed
for the quantitative estimation of paraglobulin. With this object a
known weight or volume of serum is dialysed for 24 — 36 hours ; at
the end of this time the contents of the dialyser have become turbid,
and they are subjected to a current of C02 ; the precipitate produced
is collected on a filter, washed with water and alcohol and dried.
Following this method Schmidt found that 100 c.c. of the serum of
ox's blood yielded on an average 0'887 grammes of dry paraglobulin.

Hammar- ft has however been shewn by Hammarsten3 that

sten's meth neither by acetic acid, nor by dialysis and carbonic

serum^giob^ acid> is paraglobulin fully precipitated : indeed these

lin. reagents only throw down a small fraction of the

paraglobulin contained in the serum or plasma. Having discovered

1 A. Schmidt, " Weiteres iiber den Faserstoff und die Ursachen seiner Gerinnung.
1. Die fibrinoplastische Substanz." Archiv f. Anatomic u. Phijs., 1862, p. 429^et seq.?

2 A. Schmidt, " Untersuchung des Eiereiweisses und des Blutserum durch Dialyse.
Beitrdge zur Anatomic und Physiologie, als Festgabe Carl Ludwig geividmet. Leipzig,
1875. Part I., p. 101.

3 Hammarsten, "Ueber das Paraglobulin," Erster Abschnitt. Pfliiger's Archiv,
Vol. xvn. (1878) p. 447 et seq.

SERUM-GLOBULIN.

[BOOK i.

that magnesium sulphate, added to complete saturation, precipitates
every trace of paraglobulin present in a solution, whilst it has no
action on serum-albumin, Hammarsten has by its aid determined
how much paraglobulin the blood serum contains. His determina-
tions would appear to leave no doubt that paraglobulin is in many
cases the chief proteid of the serum, as can be seen by studying the
accompanying table : —

Variety of Serum.

Total Solids
in 100 pts.

Total Proteids
in 100 pts.

Sermn-
globulin
in 100 pts.

Sernm-
albumin
in 100 pts.

Lecithin,
fat,
salts, «&c.
in 100 pts.

Serum-
globulin.
Serum -
albumin.

From blood of horse

» » » ox
„ „ „ man
„ „ „ rabbit

8-597
8-965
9-207
7-525

7-257
7-499
7-619
6-225

4-565
4-169
3-103

1-788

2-677
3-329
4-516
4-436

1-340
1-466
1-587
1-299

1

0-591
1

0-842
1

1-511
1
T5"

Properties Serum-globulin precipitated by any of tbe methods

buST"11"510" Described is found to be soluble in water holding C02
in solution, in water holding oxygen in solution, in
very weak aqueous solutions of the alkalies, in lime water, in weak
solutions of neutral alkaline salts, in solution of sodium phosphate
and of the carbonates of the alkalies.

When considerable quantities of serum-globulin are dissolved in
very weak solutions of the alkalies, perfectly neutral solutions are
obtained which are not coagulated by heat, but which are so when
very cautiously treated with acetic, hydrochloric, nitric, or sulphuric
acids, the precipitate being readily dissolved by an excess of the
reagent; such weak alkaline solutions are precipitated by the addition
of a large quantity of alcohol.

Serum-globulin is in great part, though by no means completely,
precipitated when sodium chloride is dissolved to saturation in its
solution ; the precipitated serum-globulin is found to be soluble in
weak solutions of sodium chloride.

It was stated by A. Schmidt that paraglobulin is completely precipitated
by the addition of powdered NaCl to its solutions, but Eichwald and
Hammarsten, and especially the latter, have shewn conclusively that
Schmidt was in error. On the other hand, the body to be next described,
viz. Fibrinogen, is completely precipitated when treated in the same manner
by NaCl.

According to Hammarsten1, if a very small quantity of common

1 Hammarsten, "Ueber das Paraglobulin," Zweiter Abschnitt. Pfliiger's Archiv,
Vol. xvni. (1878) p. 39 et seq.

CHAP. II.] THE BLOOD. 39

salt (from 0*03 to 0'5 or 07 p. c.) be added to a very feebly alkaline
solution of paraglobulin this body is precipitated, but on a further
addition of salt the precipitate re-dissolves, only to be again
precipitated when the amount of sodium chloride exceeds about
20 p. c.

Solutions of paraglobulin (as for example in NaCl) coagulate at
temperatures varying between 68° and 80° C., most commonly at
75° C., the variations being due to the amount of sodium chloride
present, to the duration of the process of heating, and perhaps to
other circumstances. (Weyl1, Harnmarsten2, Frederique3.)

Moderately concentrated solutions of paraglobulin are not pre-
cipitated by the addition to them of 16 — 20 p.c. of NaCl (Ham-
marsten4).

Serum-globulin is said to diffuse with considerable ease through
animal membranes. On the other hand, it is absolutely unable to
pass through parchment paper.

The term paraglobulin sufficiently indicates that this body belongs
to that group of proteids of which the first well-known member was
the proteid constituent of the crystalline lens to which the name of
Globulin was given.

Because of its assumed co-operation in the formation of fibrin, the
term fibrinoplastic substance was ascribed to it by A. Schmidt, but,
as will be shewn in the section on coagulation, there are no longer
grounds for ascribing this function to pure paraglobulin.

Paraglobulin is not only found in the plasma and in the serum,
but it is a constituent of the colourless and coloured (?) blood-
corpuscles, of the lymph, chyle, &c.

According to A. Schmidt's more recent views, the paraglobulin
of the serum is derived from the colourless corpuscles of the liquor
sanguinis, which in breaking down liberate this constituent, and the
body known as the fibrin-ferment. Hammaisten, whilst not denying
that a portion of the paraglobulin of serum may be derived from
the colourless corpuscles, does not believe that it all takes its origin
in this manner, for he has found the plasma to contain large
quantities of paraglobulin; he is moreover inclined to think that
some portion of the globulin found in serum may be derived from
the decomposition of fibrinogen. The Swedish observer has found
that when a solution of pure fibrinogen coagulates, besides fibrin,
there is formed a soluble proteid which belongs to the group of
globulins, and which therefore, if present in the serum, would be
reckoned as paraglobulin.

The view has been held by Brlicke and Heynsius, that para-

1 Weyl, "Beitrage zur Konntniss thierischer und pflanzlicher Eiweisskorper,"
Pfluger's Archiv, Vol. xn. p. 635 — 638.
1 Hammarsten, loc. cit., p. 64.

3 Frederique, Recherchcs sur la constitution du Plasma Sanynin. Gand, 1878.

4 Hammarsten, "Zur Lehre von der Faserstoffgerinnung," Pfluger's Archiv, Vol.
xiv. (1877) p. 224.

40 FIBRINOGEN. [BOOK I.

globulin is an alkaline albuminate1. According to Hammarsten
paraglobulin would be a proteid having the characters of a weak
acid2.

2. Fibrinogen.
Schmidt's When plasma which has been diluted with ten or

fifteen times its volume of ice-cold water, and has
keen freed from paraglobulin by the action of a long-
continued stream of CO2, is still further diluted, and
again subjected to G02, there separates a second precipitate which
is found to consist of a body very closely resembling paraglobulin, but
yet possessing certain marked distinctions. This body is denomi-
nated fibrinogerij a term which sufficiently indicates that it is pre-
sumed to be one, at least, of the precursors of fibrin.

Unlike paraglobulin, fibrinogen does not exist in the serum
which separates from blood olot, but it is present in the liquid found
in many serous cavities, as in the pericardium, the peritoneum, the
pleurae ; also in the liquid of hydrocele.

From all these liquids fibrinogen may be separated by the method
previously referred to, viz. by dilution with water, and the subsequent
action of CO, — or instead of passing C02, the liquids may be cautiously
neutralized with acetic acid. Fibrinogen may also be precipitated
from liquids which hold it in solution by adding common salt.

Like paraglobulin, fibrinogen is insoluble in pure water, but
soluble in water which holds oxygen in solution ; it is soluble in
weak solutions of the alkalies, and in solutions of many neutral salts,
as in weak solutions of sodium chloride.

Hammar- The behaviour of fibrinogen to solutions of common

salt has been studied with care by Eichwald and Ham-
marsten8, and is so important as to deserve careful
consideration, for upon it is based a method of sepa-
rating this substance from paraglobulin, and obtaining it in a pure
condition from the fluids which contain it.

Both fibrinogen and paraglobulin are soluble in solutions of sodium
chloride which contain 5 — 8 per cent, of the salt. When however
the quantity of salt attains 12 — 16p.c., fibrinogen is precipitated whilst
paraglobulin remains in solution ; the quantity of salt must amount
to more than 20 p.c. before any appreciable quantity of paraglobulin is
thrown down.

In order to obtain pure fibrinogen Hammarsten proceeds as
follows : —

The blood of the horse is mixed on its issue from the blood-

1 Heynsius, "Ueber die Eiweissverbindungen des Blutserams und des Huhnerei-
weisses," Pfluger's Arcldv, Vol. ix. 514—552.

2 Hammarsten, "Ueber das Paraglobulin," Erster Absclmitt. Pfluger's Archiv,
Vol. xvn. (1878) p. 466.

3 Hammarsten, " Untersuchungen iiber die Faserstoffgerinnung. § 5, Ueber eine
neue Methode zur Bemdarstelhmg des Fibrinogens aus dem Blutplasma". Nova Acta,
Regiae Societatis Scientiarum Upsalensis. Ser. in., Vol. x. 1, p. 31, Separatabdruck.

CHAP. II.] THE BLOOD. 41

vessels with one-third of its volume of a saturated solution of mag-
nesium sulphate. The mixture is then subjected to nitration in
order to obtain salted plasma free from corpuscles. As filtration is,
however, often very difficult from clogging of the filter, and at all
times very slow, I have, in repeating Hammarsten's experiments,
subjected the mixture of blood and magnesium sulphate to the
action of the centrifugal machine (see p. 58); in this way, in about
half an hour, perfectly clear salted plasma may be obtained in con-
siderable quantities.

To the salted plasma there is now added an equal volume of a
saturated solution of common salt; the fluid instantly becomes turbid,
and in two or three minutes an abundant flaky precipitate forms.

From this point the process may be conveniently modified as follows : —
The liquid with the suspended precipitate is carefully stirred, whereby
the precipitate usually floats to the surface and forms a thick dense layer
on the top of the liquid, which is then syphoned off. The precipitate is
now well mixed with a solution made by diluting saturated solution of
common salt with an equal volume of water, the quantity of the half
saturated solution of salt being equal to that of MgSO4 plasma which was
employed in the process. The precipitate floats up to the surface, the
NaCl solution is syphoned oft', and a fresh quantity of the same added; the
process of washing and syphoning being repeated not less often than six
times. The fibrinogen is then collected on a separate funnel, pressed
between folds of filtering paper, suspended in water and the solution
filtered. The whole process can be completed in from 2J — 3 hours1.

This is separated by filtration, and may be washed with saturated
solutions of Nad. The precipitate is freed from much adhering
moisture by pressing between folds of blotting paper, and is then
mixed with a solution of common salt containing 6 — 8 p.c. of the
salt, and in this it soon dissolves. The solution is filtered, and to it
is added an equal volume of saturated solution of NaCl, which again
throws down fibrinogen, but in a purer condition than at first, in
the form of gelatinous flakes. The precipitate may be again
dissolved in the weak solution of sodium chloride and precipitated a
third time. It may then be assumed to be pure ; it is at least free
from the minutest traces of paraglobulin and of serum-albumin. In
consequence of the common salt which adheres to it, the precipitate
is found to be soluble in pure water. A solution of fibrinogen thus
obtained is found not to be spontaneously coagulable, but to yield
fibrin when mixed with serum or other solutions possessing the
peculiar ferment action to be subsequently referred to when speaking
of Theories of coagulation.

Solutions of fibrinogen containing 1 — 5 p.c. of NaCl coagulate at
52° C. — 55°C. (Hammarsten, Frederique).

Solutions of fibrinogen coagulate at 56° C. according to Frederique,

1 Hammarsten, "Ueber das Fibrinogen." Pfliiger's Archiv, Vol. xix. (1879) p. 563,
et seq.

42 THEORIES OF COAGULATION. [BOOK I.

with whose observations agree those made on the same subject by
Weyl and Hammarsten. Frederique has shewn that if an excised
jugular vein of the horse, tied so as to confine blood within it, be
heated to 56° C., a proteid matter separates, and the plasma is there-
after found to be uncoagulable. No better proof than this could be
given to shew that fibrinogen is really contained as such within the
living blood. Frederique has made use of the low temperature at
which fibrinogen coagulates to separate this body from paraglobulin
and determine its amount. He thus determined 100 grammes of
the plasma of the horse (in one experiment) to contain 0'4299 of
fibrinogen and to yield 0'375 grms. of fibrin.

Theories of Coagulation.

The views The first step in the accurate study of the nature of

of the An- fae coagulation of the blood was made when it was
positively determined that coagulation is due to the
separation of a solid constituent from the liquor sanguinis, and this
fact was assuredly first determined with certainty by Hewson. It is
true that, as previously mentioned, Borelli had expressed himself
with correctness in the same sense ; still he did not adduce evidence
which can be considered to furnish full proof of his position1 .

Even before the discovery of this fact the cause of the coagulation
had been sought for, and various views had been expressed, none of
which, as even Hewson shewed, were at all capable of accounting for
the phenomenon. Thus it had been assumed by some that the
blood is maintained in a liquid condition in the living body by the
continual movement to which it is subjected (Borelli2, Lower3); by
others, that coagulation was due to the action of air upon the blood ;
by a third set, that coagulation was due to the cooling of the blood
on its withdrawal from the vessels ; by a fourth, that the coagulation
of the blood was an act of life and connected with the vitality of the
blood (Hunter). The first of these views is contradicted by the

1 See Borelli, De Motu Animalium. Opus posth., pars altera, 4to. Eoraae, 1681. Under
tlie heading "Analysis sanguinis in suas partes integrales, et forma compositionis ejus
inquiritur" (Prop, cxxxn.) Borelli says, "Deinde sicut in lacte adest succus con-
crescibilis in caseum, sic in sanguine reperitur succus viscosus, et glutinosus, qui post-
modum facta concretione, abit in fibras, vel membranas reticulares; quodque tales
fibrae sic condensatae non praeextiterint intra vasa animalis viventis, facile suadetur ex
eo, quod tales fibrae, et membranae albae sanguineae microscopio inspectae, crassiores
sunt vasis sanguineis capillaribus, et ideo neque excipi, neque effiuere in iis possint,
cum saltern longitudine filarnentorum, et latitudine membranarum, vias illas angus-
tissimas obstruerent. Ideo fatendum est, gluten album sanguiiieum lubricani et
nuidarn consistentiam retinere chim in animali viventi movetur."

2 Borelli, op. cit., Vol. n. p. 266.

3 Lower. The only passage in Lower's works which appears to the Author to indicate
that he entertained this opinion (which has been attributed to him) is the following :
"In cordis systole, qua liquor sanguinis conquassatur usque et ad ventriculi latera et
vasorum parietes alliditur, paululum diutius elanguescat: succus ejus nutritius in
partes secedere, grumescere, et gelatinae in modum incrassari, tandemque intra ribras
cordis hinc inde pendentes implicari, et ipsis ventriculorum parietibus accrescere, et a
cordis aestu indurari incipit, &c." Lower, De Motu Cordis.

CHAP. II.] THE BLOOD. 43

fact that the blood retains its fluidity within the healthy and yet
living blood-vessels even though the circulation have ceased; the
second is disproved by the fact that blood retained in vessels which
contain no air and are shut off from air, coagulates with readiness ;
the third is summarily and conclusively disproved by the facts that
whilst a low temperature hinders coagulation instead of hastening
it, a temperature such as that of the body of warm-blooded animals
is specially favourable to its occurrence. The fourth view is set
aside by the fact that the coagulation of the blood can be post-
poned almost indefinitely by exposure to a sufficiently low tem-
perature or by the addition to it of certain salts, and that after long
periods have passed, the experimenter may, by altering the con-
ditions, induce the previously inhibited coagulation, as for instance
by suitably diluting blood of which the coagulation has been
prevented by the addition of large quantities of neutral salts.

If coagulation were a vital act, the results of the above experi-
ments would, as Gulliver remarked1, be equivalent to a demonstra-
tion that we can pickle the life of the blood, that it is preserved
after repeated freezing and thawing, and that the blood may remain
alive many hours after the death of the body, when the muscular
fibre has lost its irritability, the limbs have stiffened, and even
partial decomposition has begun.

In considering the progress of research and the succession of
doctrines relating to coagulation, it is well to remember that the
following facts amongst many others were demonstrated by Hewson,
and were published by him in the year 1772 : Firstly, that the coagu-
lation of the blood is due to the coagulation of the liquor sanguinis,
a fact which he proved (a) by skimming off the liquor sanguinis
of the slowly coagulating blood of inflammatory diseases after
the corpuscles had subsided, and determining that it coagulated,
(6) by ligaturing a vein so as to include fluid blood within it, and
opening it after the corpuscles had subsided, and drawing off the clear
liquor sanguinis, which then coagulated. Secondly, that the coagula-
tion of the blood drawn from the body cannot be explained as due to
loss of heat, to arrest of motion, or exposure to air. Thirdly, that
coagulation may be restrained by cold and by the addition of neutral
salts to blood, the process setting in when the conditions are
modified. Fourthly, that the walls of the living blood-vessels exert
a remarkable influence in restraining coagulation.

Discoveries The serous sacs of the body, even in health, contain

of Buchanan. small quantities of liquid which at first sight appears
closely to resemble the serum of blood, but which is similar to that
found in the lymphatic vessels, viz. lymph. Of such serous sacs the
pericardium is the one which invariably contains after death more or
less liquid, which has received the name of liquor pericardii. In
disease, the fluid contents of the serous sacs may however increase

1 Hewson's Works, note 12, p. 21.

44 BUCHANAN'S VIEWS ON COAGULATION. [BOOK i.

very materially, and sacs which normally contain no appreciable quan-
tity of liquid may contain large amounts ; this is, for instance, true of
the tunica vaginalis testis, the serous sac which envelopes the testis,
which is liable to become distended with liquid, the condition being
denominated hydrocele.

The liquor pericardii of man after it has remained for some
hours after death in the pericardium, and the liquid of hydrocele,
if removed without any admixture of blood, do not coagulate spon-
taneously, and they differ in that respect from the liquor sanguinis1.

It was however shewn by Dr Andrew Buchanan of Glasgow in
18312, that on adding to ascitic fluid, to serum from the chest, and to
hydrocele fluid the liquid obtained by pressing a blood clot in linen
cloth, there was produced a coagulum similar to that which separates
spontaneously from blood.

At first Dr Buchanan believed that the blood-colouring matter
was the agent present in the squeezed clot, which conferred upon
these transudations the property of coagulating. On mixing, how-
ever, some peritoneal fluid with the serum of blood, a coagulum was
obtained. On subsequently mixing perfectly clear blood-serum
with peritoneal fluid and with the fluid of hydrocele, removed after
death from the body of the same man, a beautiful pellucid and pretty
firm coagulum was obtained. Dr Buchanan remarked, "I repeated
the experiment very frequently with serum obtained from the
serous cavities of the testis, from the peritoneum, from the cavities
of the pleura, and from the pericardium. The result has generally
been as I have just described, but not always so."

These observations of Dr Buchanan on the coagulation of the
fluids of serous cavities with other most interesting facts and
generalizations were published in 1845 3.

"The opinions commonly entertained by physiologists and chemists
to which allusion has just been made, are that fibrin has a spontaneous
tendency to coagulate; that this spontaneous coagulability is a characteristic
property of fibrin, by which it is distinguished from albumin and casein ;
and that the coagulation of the blood and of various animal fluids depends
on the spontaneous coagulation of the fibrin which they contain. My
experiments, on the other hand, shew that fibrin has not the least tendency
to deposit itself spontaneously in the form of a coagulum : that, like
albumin and casein, fibrin often coagulates under the influence of suitable

1 The liquor pericardii of the dog and of the horse does not coagulate spontaneously ;
that of the rabbit coagulates, however, with readiness.

2 "Contributions to the Physiology and Pathology of the Animal Fluids, containing
Experiments and Observations on the effects of certain substances upon the blood;
on the coagulation of the blood ; on the difference between membranous and sanguineous
serum; on the formation of the buffy or inflammatory crust; on the formation of pus;
and on the process of sanguification, by Andrew Buchanan, M.D., Junior, Surgeon to
the Glasgow Infirmary." London Medical Gazette, vol. xvm. (2nd vol. for session 1835
—36), p. 50.

a "On the Coagulation of the Blood and other fibriniferous liquids, "London Medical
Gazette, 1845, Vol. i. (New Series) p. 617. (Communicated to the Glasgow Philosophical
Society, 'Feb. 19, 1845.) Reprinted in the Journal of Physiology, 1879.

CHAP. II.] THE BLOOD. 45

reagents : and that the blood and most other liquids of the body which
appear to coagulate spontaneously, only do so in consequence of their
containing at once fibrin and substances capable of reacting upon it and
so occasioning coagulation."

Dr Buchanan then announced that he had found that the ad-
dition of that which he designated washed blood clot was most efficient
in inducing the coagulation of such liquids as do not coagulate spon-
taneously, but do so on the addition of blood. The 'washed blood
clot' he obtained by mixing one part of liquid blood with from six to
ten parts of water, and stirring carefully for five minutes. After the
mixture had stood for twelve or twenty-four hours, it was filtered
through a coarse linen cloth, and the substance left in the cloth
washed with water.

When a small portion of this washed clot was reduced to frag-
ments and diffused through the liquid of hydrocele, coagulation
ensued, in many cases as rapidly as in the blood itself. The
washed coagulum retains, according to Buchanan, its coagulating
power for a long period, and with the addition of a little spirit of
wine may be kept for many months with its activity un-
impaired.

"The power," Buchanan remarked, "which the washed clot has of
coagulating fibrin, is not less remarkable than that of rennet in
coagulating milk, to which indeed ifc may be aptly compared."

The 'washed clot' of Buchanan is a mechanical mixture of
fibrin with colourless corpuscles. Upon which of these constituents
did its coagulant power depend? Buchanan concluded, from many
considerations, that this was seated in the colourless corpuscles.
He found that the buffy-coat of the blood of the horse, which is
exceedingly rich in colourless corpuscles, possessed a much greater
power of inducing coagulation, and preserved that power after being
kept for months and pulverized (from which statement we must con-
clude that the substance was dried). Moreover that the upper layers
of red clot which are comparatively rich in colourless corpuscles have
a stronger coagulating power than the lower layers. Furthermore
Buchanan found that many tissues of the body, muscle, connective
tissue and central nerve-organs possess, though in a much less degree,
the coagulant power, and he leant to the opinion that their influence
is seated in their cellular elements ('primary cells or vesicles').

To recapitulate: — Buchanan held that the coagulation of the blood
is due to the conversion of a soluble constituent of the liquor san-
guinis into fibrin by an action exerted probably by the colourless
corpuscles and comparable to the action which rennet exerts in
effecting the coagulation of milk. Furthermore, that the liquid
which accumulates in certain serous sacs may be made to yield a
coagulum of fibrin when subjected to the action of liquids or solids
rich in the cellular elements with which the coagulant action ap-
peared to be associated.

Although not altogether forgotten by a few individuals in England,

46 DENIS' PLASMINE. SCHMIDT'S VIEWS. [BOOK i.

these most interesting results of Professor Buchanan have not formed
part of the common stock of scientific knowledge, and are generally
known only as re-discovered and greatly added to by Professor Alex-
ander Schmidt of Dorpat.

Denis' Although Buchanan believed in the existence of

fibrin in solution in the liquor sanguinis he had no idea

tion to Fibrin °^ separating the dissolved substance. Denis in 1859

announced1 the separation from the plasma of aproteid

body to which he gave the name of Plasmine and which yields

fibrin as a product of decomposition.

Denis commences by mixing uncoagulated blood with one-seventh
its volume of a saturated solution of sodium sulphate. After the
corpuscles have subsided, the supernatant mixture of liquor sanguinis
and solution of sodium sulphate is decanted and sodium chloride
is added little by little as long as it is dissolved. The solution be-
comes turbid and soon acquires a creamy consistence, from the separa-
tion of a bulky flocculent precipitate. The fluid is thrown upon a.
filter and washed with a saturated solution of sodium chloride.
The matter which remains undissolved is the plasmine of Denis.
Of this plasmine Denis obtained 14*59 grammes from 1000 grammes
of human blood.

If plasmine, thus precipitated through the agency of sodium
chloride, be placed in water, the solution, in the course of a few
minutes, undergoes spontaneous coagulation; the coagulum consists
of fibrin similar to that obtained directly from blood, and the amount
yielded by the plasmine also corresponds with that which would
have been obtained directly from blood. In addition, however, to
the insoluble fibrin which separates, there is found to be present in
the solution a proteid substance to which Denis gives the name of
'fibrine soluble' to distinguish it from the first 'fibrine concrete' or
'fibrine ordinaire/

Denis therefore believed that the precursor of fibrin in the blood
is a complex body, plasmine, which at the moment of coagulation
splits up into two proteids, of which the one separates in the form of
the insoluble fibrin and the other dissolves in the serum. These
views of Denis will be again referred to when speaking of the more
recent investigations of Hammarsten.

The disco- The fundamental fact discovered by A. Schmidt was

veries and the very same which it has been shewn was clearly
hypotheses of described long before him by Dr Andrew Buchanan, viz.
A. Schmidt. that there occur animal fluids from which fibrin does
not separate spontaneously but only after the addition of blood or of
blood-serum, or certain of their constituents2.

1 Denis, Nemoire sur le sang, 1859, p. 32.

2 A. Schmidt, "Ueber den Faserstoff und die Ursachen seiner Gerimmng." Archiv
f, Anat. u. Physiolog., 1861, p. 545.

CHAP. II.] THE BLOOD. 47

Schmidt however soon proceeded a step further1. He studied the
effect of dilution upon, and the passage of carbon dioxide through,
liquor sanguinis and serum, and shewed how to obtain in this way,
though certainly not in a state of purity, the bodies which have been
described as paraglobulin and fibrinogen. He discovered that when
these bodies in a separate condition exist in solution and the
solutions are mixed, if. circumstances be favourable, coagulation
occurs sooner or later.

These facts he explained by supposing that the formation of
fibrin is due to the inter-action of the two closely allied proteids,
of which the one, fibrinogen, is often present without the other,
paraglobulin; and to designate the property which the latter

sesses of leading to the formation of fibrin from fibrinogen,
hmidt applied to it the name of the fibrinoplastic substance.

Schmidt at first supposed that the plasma contained both fibrin-
generators in solution, there being, however, an excess of the fibrino-
plastic substance. When blood or plasma coagulates, he supposed
the whole of the fibrinogen to be used up, whilst the paraglobulin
over and above the quantity which had taken part in the formation
of fibrin, remained in solution in the serum, whence it could be
separated by dilution and neutralizing either with CO2 or acetic acid.
Fluids which, like hydrocele, do not coagulate spontaneously, but only
after the addition of paraglobulin, he supposed to be wanting in this
body, which he regarded as one of the two essential fibrin-
generators.

There are many ways of repeating Schmidt's observations on the
coagulating influence of paraglobulin on nbrinogen. One of the most
convenient is the following : the serum of blood is diluted, precipitated
by dilute acetic acid (10 c.c. of serum being diluted with 150 c.c. of
water and treated with four drops of 25 p. c. acetic acid). The precipitate
is washed with water. Eibrinogen is then precipitated (in an impure
condition) by saturating any fluid which contains it, e. g. hydrocele fluid,
with sodium chloride. The precipitate is collected on a filter, and after
the filtrate has passed through, the filter is filled up with water, which
dissolves the precipitated fibrinogen, in virtue of the sodium chloride
adhering to it. To this solution of impure fibrinogen the previously
precipitated paraglobulin is added, when coagulation sometimes occurs.

Amongst the facts which were adduced by Schmidt and which
appeared to give great support to his views was this one : that if
from diluted plasma, the paraglobulin is precipitated by dilution of
water and passage of a stream of carbon dioxide, the power of
spontaneous coagulability is unquestionably destroyed, whilst it may
be occasionally restored by the restoration of the removed paraglobu-
lin to the fibrinogenous liquid.

It is to be remarked that Schmidt never committed himself to a

1 A. Schmidt, " Weiteres iiber den Faserstoff und die Ursachen seiner Gerimmng."
Archivf. Anat. u. Phijs., 1862, pp. 428—469 and 533—564.

48 RESEARCHES OF A. SCHMIDT. THE FIBRIN-FERMENT. [BOOK I.

statement of the way in which the two bodies which he believed to
be fibrin-generators, associated themselves in the formation of fibrin.
He however believed that he had proved the actual co-operation of
paraglobulin in the formation of fibrin by shewing that the amount of
fibrin which separates from a solution containing paraglobulin is to a
certain extent influenced by the amount of paraglobulin added to
that fluid.

The Fibrin-fmnent.

Such were the principal facts published by A. Schmidt anterior
to 1872, and the views which he based upon them. It will be seen
how widely these views differed from those of Buchanan and of Denis,
each of whom was acquainted with many of the most important facts
independently discovered by the Dorpat professor. But in their turn
the views of Schmidt soon received from their author most important
modifications.

Schmidt's theory of coagulation postulated that when a fluid
containing fibrinogen did not coagulate spontaneously, this was due
to an absence of the fibrinoplastic substance. But he discovered
that the two fibrin-generators may be present in the same fluid and
yet coagulation not occur. Hydrocele fluid is for instance by no
means free from paraglobulin and may sometimes contain considerable
quantities of that body, without coagulating spontaneously, though the
addition of blood or of blood serum will lead to its coagulation. Does
blood or blood serum then contain some constituent other than
paraglobulin which exerts a fibrinoplastic action ?

It appears so, and this body Schmidt believes to be of the nature of
a ferment which is liberated after the blood is removed from the blood-
vessels, and which in an impure condition he prepares as follows1:

Schmidt's Blood or, still better, serum separated from the clot of
coagulated blood, is treated with twenty times its volume
soTutto^of °^ alcon°l a]Qd the mixture set aside in a stoppered bottle
Fibrin-fer- f°r at least a fortnight, but preferably for a period of three
ment. months. The alcohol coagulates the proteid matters of the

plasma and corpuscles as well as the haemoglobin contained in the
latter, and by the prolonged action of alcohol these various matters are
for the most part rendered insoluble in water. The insoluble matter
is then collected on a filter and dried over sulphuric acid, and, when
dry, finely pulverized. The powder is treated with water ; the aqueous
solution is found to contain the so-called fibrin-ferment.

Such a solution when added to a liquid which contains fibrinogen
and paraglobulin but which does not coagulate spontaneously, often
rapidly gives rise to a coagulum. The amount of fibrin which
separates is, according to Schmidt, in no respect influenced by the

1 A. Schmidt, "Neue Untersuchungen iiber die Faserstoffgerinnung." Pfliiger's
Archiv, Vol. vi. (1872) p. 445.

CHAP. II.] THE BLOOD. 49

amount of the ferment, but the rapidity of coagulation is so influenced.
The influence of solutions of the fibrin-ferment may be well seen by
adding it to dilute solutions of salted plasma. It has been said that
blood or plasma which has been prevented from coagulating by the
addition of a neutral salt, such as sodium or magnesium sulphate,
will coagulate if a sufficient quantity of water be added. The
coagulation is, however, under the circumstances not an immediate
one. But if to a slowly coagulating mixture of plasma, water, and
neutral salt, there be added some of Schmidt's solution of fibrin-ferment
the process may be remarkably hastened.

The Au- In narrating the discoveries of I)r Andrew Buchanan

thor's Method attention was called to the action of the so-called ' washed
solution^ g a blood clot' of that author, in bringing about the coagu-
Fibrin-fer- lation of certain fluids ; washed blood clot being really
ment1. fibriii obtained by washing the coagulum which separates

from blood when, at the time of being shed, that fluid is mixed with
about 10 times its volume of water. As Buchanan pointed out, such
fibrin possesses remarkable coagulant power, and, if preserved in weak
spirit, will retain that power for many months.

By digesting Buchanan's washed blood clot in an 8 p.c. solution of
common salt, a solution is obtained which possesses in a very intense
degree the properties of Schmidt's solution of fibrin-ferment. This
solution contains a proteid in solution which possesses all the reactions
of a globulin; it is rendered inactive by exposure to temperature
of 56° — 58° C., and when it is saturated with powdered magnesium
sulphate.

The origin of the Fibrin-ferment.

After the discovery of the so-called fibrin-ferment, Schmidt's
views might be stated as follows, though not in his words : — In cases
where a fluid coagulates spontaneously with the formation of fibrin
there must be present the two fibrin-generators and a yet unknown
body, the fibrin-ferment, whose presence is, however, essential in order
that the two bodies shall associate themselves.

Where again a liquid does not coagulate spontaneously but does so
on the addition of blood or of serum, the absence of coagulation may
be due to the absence of ferment, the two fibrin-factor-s being present ;
or it may be, and sometimes is, due to the absence of paraglobulin. In
the first case coagulation will be induced by the addition of , fibrin-
ferment alone, in the latter not until the previous addition of para-
globulin. The interaction of the fibrin-factors necessitates, hoivever, the
presence' of certain quantities of salts, and especially of sodium chloride.

According to Schmidt, then, the formation of fibrin is due to the
interaction of two bodies under the influence of a ferment.

1 A. Gamgee, "Some old and new experiments on the Fibrin-ferment." Journal
of Physiology, 1879. No. n.

50 SCHMIDT'S RESEARCHES ON THE FIBRIN-FERMENT. [LOOK i.

But whence comes the ferment? Schmidt received the blood
as it flowed from the blood-vessels of a living animal directly into
absolute alcohol and then subjected the product to the process
followed in the separation of the fibrin-ferment, and found that the
solution obtained under these circumstances was free from any ferment
action, and he therefore concluded that the ferment is generated in
the blood after it is withdrawn from the blood-vessels. But how
generated? Many facts conspired to connect the formation of
ferment with the colourless corpuscles of the blood.

Schmidt found that liquids coagulate more or less rapidly, very
much according as they contain many or few colourless corpuscles; he
found that horse plasma, diluted with ice-cold water and filtered
from all corpuscles, coagulates not only much more slowly but also
much more feebly than the same plasma unfiltered; that in cooled
horse plasma from which the corpuscles have subsided, the upper lay-
ers, most free from corpuscles, coagulate more imperfectly, yielding
actually less fibrin than the lower, richer in corpuscles, and that such
plasma free from corpuscles, when subjected to the process for
separating fibrin-ferment, yields a solution comparatively inactive,
when compared with a solution prepared from plasma rich in
corpuscles. Moreover Schmidt found that by adding paraglobulin to
the above plasma the yield of fibrin was increased.

Furthermore Schmidt thinks he has proved that in the short
interval which, at ordinary temperatures, intervenes between the
shedding and coagulation of the blood there is a rapid breaking down
of colourless cells and of cells which appear in some way intermediate
between the colourless and coloured cells, which are nucleated like
colourless cells, but whose protoplasm is tinged with haemoglobin.
He therefore has come to the conclusion that the coagulation of the
blood is due to the union of fibrinogen, which exists preformed in the
plasma, with paraglobulin derived from the colourless corpuscles — a
union which takes place under the influence of a ferment-like body
which also arises in the same cells, and which like paraglobulin is de-
rived from them in the short interval which elapses before coagulation.

In their latest developments the views of Schmidt approach
much more closely to those of the man wrhose facts and theories have
both been buried in oblivion, Dr Buchanan. Both observers look
upon coagulation as due to a ferment-like action, exerted upon a
constituent of the plasma, which is, in the living body, dissolved in
that fluid ; both connect that ferment action with the colourless cells
of the blood, and Schmidt adds definiteness to the older views of
Buchanan by connecting the ferment action with the actual breaking
down of those bodies.

The chief point of divergence — the one element in Schmidt's
theory which had no place in Buchanan's — relates to the accessory
body paraglobulin, whose existence he did not even surmise, much less
consider to be essential to the formation of fibrin. But is it essential?

CHAP. II.] THE BLOOD. 51

The Researches of Hammarsten1.

In describing paraglobulin and fibrinogen it has been stated that
the researches of Eichwald and Hammarsten, and especially of the
latter, have shewn that the behaviour of the two bodies which, according
to Schmidt, are the fibrin-factors, in respect to sodium chloride is
exceedingly diverse. Both bodies are precipitated from their solutions
when these are saturated with sodium chloride, though fibrinogen
alone is completely precipitated. Fibrinogen is precipitated from its
solutions when these contain 13 p. c, of sodium chloride or more,
whilst paraglobulin only becomes insoluble when the solution contains
about 20 p. c. or more of sodium chloride.

Making use of these reactions and following the method which
has been described when speaking of fibrinogen, Hammarsten has
separated fibrinogen which is free from all traces of serum-albumin and
of paraglobulin, and has found that such fibrinogen dissolved in weak
solutions of sodium chloride may be kept indefinitely without under-
going coagulation. When, however, there is added to it serum of
blood, or a solution of fibrin-firment prepared according to the
directions of Schmidt or by improved methods, coagulation occurs
with great rapidity.

Fibrinogen According to Hammarsten, then, the coagulation of

the one pre- the blood depends upon the production of fibrin from
cursor of one body, fibrinogen, existing in solution in the liqu'or

sanguinis, under the influence of that yet non-isolated
body, the fibrin-ferment. Although provisionally em-
ploying the term fibrin- ferment, Hammarsten, like Schmidt, does not
commit himself to the view that this body is really of the nature of a
ferment.

The grounds upon which Hammarsten has corne to the conclu-
sion that paraglobulin is not indispensable to the formation of fibrin
are the following : — 1st. The fibrinoplastic action is not a specific
property of paraglobulin, but is 'exerted by some other substances,
such as calcium chloride and impure casein. 2nd. The fibrinoplastic
activity does not belong to pure paraglobulin, but only to that
substance when precipitated from serum and certain other fluids.
In accordance with this statement Hammarsten has obtained from
hydrocele fluids, which were quite free from ferment, a pure para-
globulin, which possessed all the typical properties of that body, but
exerted no fibrinoplastic activity. 3rd. The chief proof in support
of Schmidt's hypothesis is based upon the surmise that those
hydrocele fluids which do not coagulate when treated with ferment
alone, but only after the addition of paraglobulin, either do not

1 Hammarsten : " Untersucliungen iiber die Faserstoffgerinmmg." Nov. Acta Reg.
Soc. Scientiar. UpsaL, Ser. x. Vol. x. Separatabdruck, Upsala, 1878.—" Zur Lehre von
der Faserstoffgerinnung." Pfliiger's Archiv, Vol. xiv. (1877) pp. 211— 274.— "Ueber das
Paraglobulin." PflUger's Archie, Vol. xvn. pp. 413— 468.— "Ueber das Paraglobuliu,
zweiter Abschnitt." Pfliiger's Archiv, Vol. xvm. pp. 38—116.— "Ueber das Fibrinogen.
Pniiger?s Archiv, Vol. xix. pp. 563—622.

4—2

52 THE RESEARCHES OF HAMMARSTEN. [BOOK I.

contain paraglobulin or at most mere traces of it. This surmise is
however thoroughly incorrect, as Hammarsten's quantitative analyses
have shewn that such fluids contain, on the contrary, very consider-
able quantities of paraglobulin. This paraglobulin possesses, how-
ever, no fibrinoplastic activity, affording another proof that the fibrino-
plastic property is to be ascribed to some contaminating substance.
4th. The most weighty fact in opposition to Schmidt's hypothesis is
however the possibility of obtaining solutions of nbrinogen which
are free from paraglobulin, and which, when treated with ferment
solutions which are free from paraglobulin, yield typical fibrin.

The observations of Hammarsten corroborate those of Schmidt
in reference to the living plasma containing less paraglobulin than
serum, and he believes with Schmidt that some of the paraglobulin
is derived from the colourless corpuscles ; he does not however,
as has been said in speaking of paraglobulin, ascribe the origin of
this body entirely to this source ; much is doubtless present in solu-
tion in the living liquor sanguinis, and some may perhaps originate as
a product in the decomposition which gives rise to fibrin, for even
Hammarsten was at first inclined to view coagulation very much
as Denis did, viz. as being a process in which a complex body decom-
poses with the formation of simpler products, of which fibrin is one.

Hammarsten corroborates Schmidt also in his statement that
the addition of paraglobulin to scantily coagulating plasma or to a
transudation which will not coagulate in the presence of ferment,
may in the first case lead to an increase of the fibrin produced and
in the second to the production of a coagulum. But Hammarsten
shews that many substances besides paraglobulin will under the same
circumstances exert the same fibrinoplastic influence. The addition
for example of calcium chloride, CaCl8, to some specimens of hydrocele
fluids, which will not coagulate on the addition of Schmidt's fibrin-
ferment, produces the same effect as the addition of paraglobulin.

If paraglobulin were specifically one of the fibrin-factors, it would
not, presumedly, be replaceable by any other proteid substance. Ham-
marsten having, however, by a process for which the original must
be consulted, prepared casein which was readily soluble in solutions
of sodium chloride, found that the addition of its solution to transu-
dations led not merely to an acceleration of the process of coagula-
tion, but to a remarkable increase in the amount of fibrin formed.

In other experiments he found that the mere neutralization of
a transudation, which does not coagulate spontaneously, will often
lead to coagulation setting in. Furthermore, Hammarsten has
found that from some hydrocele fluids, which will not coagulate on
the addition of fibrin-ferment, it is possible to separate, by his
process, fibrinogen, which when dissolved and treated with the same
fibrin-ferment, will yield a coagulum of fibrin.

It is obvious, then, that in a fluid there may exist substances
which either hinder the formation of fibrin, or prevent its precipita-
tion when formed. We know, for instance, that such substances

CHAP. II.] THE BLOOD. 53

as free alkalies or their carbonates, and some salts, will actually exert
such an influence. The addition of any body which will combine with,
or neutralize, the substances which prevent the separation of fibrin
will naturally lead to its formation. If, for example, a hydrocele
fluid, which contains but a small quantity of fibrinogen and which
will not coagulate on the addition of ferment, be treated with solu-
tion of calcium chloride, coagulation will sometimes occur. Now it is
conceivable that in this case the chloride acts by decomposing the
alkaline carbonate which hinders the precipitation of the fibrin, for
were sodium carbonate and calcium chloride to come in contact the
reaction would be as follows: Na2CO3 + CaCl^ = 2NaCl + OaGO3.
Paraglobulin may, perhaps, act in a similar manner, by combining
with substances which hinder the precipitation of fibrin.

Schmidt had discovered that the addition of serum which has
been freed from paraglobulin (by dilution with water, passage of CO2
and concentration in vacua) to hydrocele fluid does not cause the
latter to coagulate, and Hammarsten confirms the statement in
reference to hydrocele fluid or to solutions which are as poor in
fibrinogen as that fluid. If, however, such serum, free from para-
globulin, be added to a strong solution of fibrinogen, the latter will
coagulate well. Hammarsten's explanation is the following : — A
solution of fibrinogen, prepared according to the method of Schmidt,
contains more free alkali than the original fluid did whilst it con-
tained paraglobulin. The former contains therefore a larger propor-
tion of substances capable of dissolving fibrin, and when it is mixed
with a liquid containing only a small quantity of fibrinogen, a larger
portion of the resulting fibrin, or it may be the whole of it; may be
held in solution. When, however, the same solution is added to
a fluid containing an abundance of fibrinogen, the substances capable
of dissolving fibrin are no longer capable of holding in solution all the
fibrin which is formed ; and in this case coagulation occurs.

Keviewing all the facts which have been recorded in the preceding
pages, it would appear that, on the whole, the evidence is decidedly in
favour of the view that the coagulation of the blood is dependent
upon the presence in the plasma of a proteid body, fibrinogen, which
under favourable circumstances undergoes conversion or perhaps
decomposition into fibrin. The conversion of fibrinogen into fibrin
outside of the body appears to be connected with the action of a
ferment produced in the colourless cells of the blood and probably
only set free when these cells break down.

The influence of salts on coagulation.

In the course of his researches Schmidt discovered that salts exert a
remarkable action in furthering the spontaneous coagulation of liquids
containing the various fibrin-factors.

If from two liquids which yield when mixed a coagulum of fibrin,
the salts be separated by dialysis, and the proteids which are precipitated

54 INFLUENCE OF SALTS AND LIVING WALLS OF VESSELS. [BOOK T.

in the process (the supposed fibrin-generators) be dissolved in weak solu-
tions of sodium hydrate and mixed, no coagulation will occur, unless there
be added to the mixture the dialysate from the two operations, reduced by
evaporation to a small volume, or unless sodium chloride be added until
it amount to 1 per cent, of the mixture; then, however, coagulation does
occur. The quantity of salt which is needed to bring about coagulation
increases with the volume of the solution of the fibrin-factors, a circum-
stance which fully explains why by largely diluting a spontaneously
coagulating fluid, a slowing of the process of coagulation, and a diminution
in the quantity of fibrin produced, are always brought about.

Non-coagulation of the blood within living blood-vessels.

Any theory of the coagulation of the blood which would lay claim
to truth or completeness should be adequate to explain the remark-
able circumstances that the blood does not coagulate as long as it is
contained within the living uninjured vessels, but that it does coagulate
when the vessel is injured or dies.

Let us examine the facts which we at present possess in reference
to this matter.

(1). So long as the vessels are uninjured and alive, the
blood which circulates within them does not coagulate. When a
foreign- body is however introduced into the vessels, as when a silver
needle is made to transfix an artery, a coagulum of fibrin forms
around the metal, although it be in the stream of living blood.

(2) If, however, the coats of an artery be diseased or injured
in such a way that the endothelial coat which lines it ceases to be
intact, coagulation will occur, giving rise to a solid plug or 'thrombus/
the latter term being applied specifically to the coagulation which
occurs in a vessel during life. Perhaps the most common example
of a thrombus is that which is occasioned by the application of a
ligature to an artery; in this case both the middle and internal coats
are usually severely injured, the continuity of the endothelial lining
of the internal coat being certainly affected, and, almost immediately,
there results coagulation.

: Another common example of the production of thrombus during
life is afforded by the occurrence of the process in aneurisms, in which,
amongst other lesions of the arterial walls, a direct breach in the
continuity of the endothelium certainly often exists.

(3) But not only does thrombosis occur where a direct break
in the continuity of endothelium can be distinctly proved to exist,
but also where an injury of any kind is inflicted upon an artery.
The process has been studied with great minuteness by Zahn in the
case of the arteries of the frog and deserves particular attention in
reference to the doctrines of coagulation.

Zahn1 has observed that when a crystal of sodium chloride is

| : Zahn, Virchow's Arclilv, Vol. LXII. p. 81. See Cohuheim, Vorlcsungen iiber allgemeine
Pathologic, 1877, Vol. i. p. 150 et seq.

CHAP. II.] THE BLOOD. 55

thrust deeply into the tissue of the tongue or mesentery of the frog
so as to be in close proximity to an artery or a vein, the inner wall of
the vessel at the point which corresponds to the crystal becomes
covered by colourless blood corpuscles, whose number continually
increases ; soon there are three or four layers of colourless corpuscles
closely pressed against the wall, whilst the heap grows ever larger
and larger as the blood which flows past continually brings fresh
white corpuscles to add to it. Soon the vessel becomes completely
plugged by this agglomeration of colourless corpuscles. The subse-
quent progress of such a thrombus may be various. In some cases
however the following process may distinctly be observed: — The
Y\rhole mass of cells undergo a fine granulation and the contours
of individual cells become less distinct. Then the contours of the cells
become lost altogether, and a feebly refracting finely granulated mass
results, which is said to be not unlike a mass of fibrin.

(4) When blood is occluded by ligatures within a living vein, it
will be found to remain uncoagulated for many hours, providing the
vitality of the vein persist. This remarkable experiment was first
performed by Hewson, and was subjected to a careful study by
Professor Lister, and more lately by Frederique. If, however,
the vitality of the vein be destroyed by the application of caustic
ammonia to its exterior, coagulation will soon result (Lister). The
experiment is best performed with the jugular veins of horses. The
animal having, as is usually done in slaughtering horses, been struck
down insensible by a blow on the head, the jugular vein or veins are
exposed, and two ligatures are applied to the vein at a distance of
several inches apart, so as to include the blood contained within this
portion of the vein in a tube with venous walls. The vein may then
be dissected out without allowing its contents to escape. Such a
vein may be kept for many hours, and on being opened the blood
will be found still fluid within it, coagulating however when allowed
to flow into any ordinary vessel. After an interval of many hours,
however, the vitality of the vein being destroyed, the blood coagulates.
This experiment we owe to Hewson. Reasoning from it, it might be
surmised that the cause of the coagulation was the opening of the vein
and the exposure of its contents to air; that such an explanation is
entirely erroneous was shewn by Professor Lister, who determined that
blood would remain fluid for hours in a vein after being exposed
with the utmost freedom to the air by being poured in thin streams
from one venous capsule to another.

The observation of Lister might lead one to the conclusion
which Professor Briicke arrived at from his experiments. That
eminent observer, extending the observations of Hewson, shewed that
blood injected into the separated, but yet living, contracting, heart
of a turtle, would preserve its fluidity for days, and came to the
conclusion that the walls of the vascular system possess a power
of restraining coagulation — a view which was assuredly shared by
Hewson, but which in this case appeared to find its most striking

5() NON-COAGULATION OF BLOOD IN LIVING VESSELS. [BOOK I.

proof. On returning, however, to other ohservations of Lister we are
warned to pause before we draw the above conclusion. The author
quotes Professor Lister's description of one of his experiments conveyed
to him in a private communication:

" The manner in which I did succeed in my experiments long ago on
the coagulation of the blood in maintaining its fluidity outside the living
body seems to me, if I may venture to say so, deserving of more attention
than I think it has received. Having ascertained that the blood remains
fluid for many hours after somatic death in all vessels except the heart
and principal trunks provided that the vessels have been previously
healthy, I removed a portion of the jugular vein of an ox, after tying it
in two places to retain the blood ; and then, holding the portion of vein
vertical and opening the upper end with scrupulous care that the instru-
ments employed should not touch the blood, I slipped down with the
utmost steadiness a piece of glass tube nearly as large in calibre as the
vein, the lower end of the tube being of full width and smooth while the
upper end was drawn out and connected by an india-rubber tube with a
stop-cock for closing it. The blood having filled the large part of the tube
and passed on into the narrow part till it escaped at the stop-cock, the
stop-cock was turned to close ifc, after which the whole apparatus was
rapidly inverted so that the blood was now in the glass vessel with
its mouth covered with the vein as a cap. The vein was next care-
fully withdrawn and a cap of gutta-percha tissue was tied over the
mouth of the tube to prevent evaporation. The blood was now in a
vessel composed entirely of ordinary solid matter, as distinguished from
living tissue, but with the peculiarity as compared with blood shed into
a basin that only the circumferential parts of the mass of blood had been
exposed to the influence of the ordinary solid. The result was that
after 24 hours, or in one experiment 48 hours, the blood was found still
fluid except a crust of clot in contact with the containing vessel, the fluid
blood coagulating at once on being poured upon a plate. I had previously
ascertained that blood would remain fluid for hours in a vein after being
exposed with the utmost freedom to the air by being poured in thin
streams from one venous capsule (if I may so speak) to another ; while,
on the contrary, want of steadiness in pushing down the glass tube into
the vein and consequent admixture of the circumferential parts which
had touched the glass with the rest would, like a stir with a stirring rod,
have made the whole coagulate.

"Thus by this simple experiment was demonstrated incontrovertibly
the fact that healthy blood has no spontaneous tendency to coagulate and
therefore that Briicke's idea of the fluidity of the blood being due to an
action of the walls of the vessels upon it was erroneous. At the same
time was illustrated the truth, which, indeed, ought to have been apparent
enough from the results of every vivisection wound, that a perfectly
undisturbing coagulum resembles healthy living tissue in failing to induce
coagulation in its vicinity."

The difference between Lister's and Briicke's explanation of the
above facts will be perhaps more apparent by the following categori-
cal statement. Briicke explains the non- coagulation of the blood
contained in the uninjured and yet living jugular vein by sup-

CHAP. II. J THE BLOOD. 57

posing that blood does possess a strong tendency to coagulate and
that the tendency which the blood has to coagulate is inhibited by a
peculiar influence exerted by the living vascular walls. Lister on
the contrary maintains that blood possesses no spontaneous tendency
to coagulate and only does so when brought in contact with any
foreign body ; it coagulates within a blood-vessel when the latter dies
because then its walls become as all other extraneous matter, but not
because there is any cessation of an action previously exerted.

After all, there appears to be less difference between the views of
Lister and Briicke than would at first appear to be the case. Let us
examine however which of their views appears most probable in the
light afforded by recent discoveries.

Of all the facts which have, thanks to the labours of Buchanan,
Schmidt, and Hammarsten, been collected, in reference to the exact
mode of origin and nature of coagulation, none appear to be so
consistent and satisfactory as those which connect the colourless cells
of the blood with the developement of a ferment-like body which, once
liberated, soon converts soluble into insoluble proteid matter; the
developement of ferment being apparently connected with a disinte-
gration of certain of the colourless cells.

As this disintegration has a tendency to occur whenever the blood
removed from the living blood-vessels is kept at temperature above
0° C., we can scarcely agree in the proposition of Professor Lister
that the blood has of itself no tendency to coagulate, and we should
rather be inclined to say that inasmuch as it contains colourless
corpuscles within it, it does contain the elements for its future
coagulation.

The remarkable phenomena of the non-coagulation of blood with-
in the yet living venous walls is probably connected with a persistence
in an intact condition of the colourless cells, or rather of those cells
in which the fibrin-ferment originates, and not as might have been
supposed, upon the destruction of the fibrin-ferment by the vascular
walls at the moment of its liberation. But it is yet impossible to
conceive why the colourless corpuscles should not break down under
the circumstances of Lister's experiments.

SEC. 3. THE SERUM AND THE CONSTITUENTS OF THE LIQUOR
SANGUINIS WHICH REMAIN IN IT.

Modes of The serum is the liquor sanguinis from which fibrin

obtaining se- has separated; it differs from that fluid in having
nun- lost its fibrinogen and perhaps in having gained some

paraglobulin.

In order to obtain perfectly pure serum when horse's blood is
available, liquor sanguinis may be first separated by subjecting the
blood to a lower temperature in the apparatus described at page 32,
and the plasma allowed to coagulate.

58

CENTRIFUGAL APPARATUS.

[BOOK i.

B

FIG. 11. PLAN AND SECTION OP THE CENTRIFUGAL MACHINE IN THE PHYSIOLOGICAL
LABORATORY or OWENS COLLEGE.

A. An iron socket secured to top of table B.

C. A steel spindle carrying the turn-table D, and turning freely in A.

E. A flange round turn-table D.

F. F. Shallow grooves on face of D, in which the test-tubes are fixed by clamps

G. G.

H. A pulley fixed to end of spindle C and turned by the cord K.
1. 1. Are two guide pulleys for cord K.

CHAP. II.] THE BLOOD. 59

It is more usual however to obtain serum by allowing blood (prefer-
ably arterial blood) to coagulate, when after some hours serum will
separate and can be decanted.

The process of separation of serum is immensely facilitated and
the resulting serum is obtained most completely free from suspended
blood cells by subjecting recently coagulated blood to the action of a
centrifugal machine, such as is represented in the accompanying
figure.

The blood as it flows from the blood-vessel is collected in stout
test-tubes provided with india-rubber stoppers. When the blood has
coagulated the tubes are fixed to the turn-table so that the stoppers
are directed centrally. The turn-table is then made to revolve with
great velocity for about half an hour, after which time the clot is
found to have retracted itself to the peripheral end of the tube, leaving
a large quantity of clear serum occupying the ends of the tube directed
towards the centre of the rotating disk.

The centrifugal machine enables us to obtain in a short. time
considerable quantities of perfectly clear serum, which is thus obtained
before any putrefactive change can have affected its composition.
When serum has been merely decanted from the clot it is generally
more or less reddish from the presence of suspended corpuscles. From
such reddish serum, serum quite free from corpuscles can be obtained
by subjecting it to rotation in the centrifugal machine for about half
an hour.

Description The serum which separates from the blood of a

of physical healthy man, whilst fasting, is a liquid of a transparent
characters of n J , ' V1 ,. -, , •, °' . ^ . . n r , , e

serum. yellow colour like light sherry wine, varying in depth of

colour but always perfectly clear. In the lower animals
the colour of the serum differs somewhat, being colourless in the
rabbit, amber coloured in the horse, of a very red amber tint in the ox,
and in the dog somewhat yellowish, nearly identical with that of man.
After a full meal the serum ceases to be transparent and becomes more
or less milky in appearance ; this phenomenon is usually described as
occurring only after an abundant fatty diet, but although seen to
greatest advantage after such a diet it constantly occurs after a full
meal of meat.

The observations of Dr Andrew Buchanan1 on this matter are of
great interest, and two of them are quoted as illustrating the above
statement : —

"A vigorous man of about 35 years of age, after fasting 19 hours,
had for dinner, twenty ounces of beef-steak, sixteen liquid ounces of brown
soup and eight ounces of bread. He was bled immediately before his
ifieal and three times after it, two ounces of blood being taken away each
time. The serum obtained from the first bleeding before the meal was
perfectly limpid; the serum from the second bleeding, three hours and

1 Buchanan, "On. the white or opaque serum of blood," Proceedings of the Philo-
sophical Society of Glasgow, Vol. i. (1841—4), p. 226.

60 PHYSICAL CHARACTERS OF SERUM. SERUM-GLOBULFN. [BOOK I.

fifteen minutes after the meal, was turbid; the serum from the third
bleeding, eight hours and fifteen minutes after the meal, was still thicker ;
while that from the last bleeding eighteen hours after the meal,, was again
limpid, although some supper had been eaten in the interval.

" The young man first mentioned, after fasting eighteen hours,, dined
upon sixteen ounces of brown soup, four ounces of bread, eight ounces
of potatoes, twenty ounces of beef-steak, and sixteen ounces of London
porter, and fasted eighteen hours after the meal. He had blood taken
from his arm four times to the extent of two ounces each time. The
serum of the blood first taken, immediately before the meal, was of an
amber yellow and quite transparent ; the serum from the second bleeding,
two hours and ten minutes after the meal, was turbid ; the serum from
the third bleeding, eight hours after the meal, was exactly of the colour
of water gruel and quite opaque ; the serum of the blood last taken,
eighteen hours after the meal, was still turbid, its limpidity not having
been, as after his usual fare, restored by an eighteen hours fast."

The milkiness of such blood is due to finely divided fat which often
may be observed to float to the surface and presents the appearance
of oil globules or drops.

The specific gravity of the serum obtained from human blood
varies between 1027 and 1032, but is on an average 1028. Its re-
action is alkaline, and its alkalinity is greater than that of the plasma.

1000 grammes of blood yield between 440 and 525 grammes of
serum (Gautier).

Serum contains roughly about 10 per cent, of solid matters in
solution ; of these the most abundant are proteid in nature, the
chief being serum-albumin ; in addition to the proteids, the serum
holds in solution small quantities of nitrogenous matters soluble
in alcohol, which are technically grouped under the term extractives or
extractive matters, fats, sugar, inorganic salts and certain gases. These
various constituents will now be discussed in detail.

TH-E PROTEIDS OF THE SERUM.
1. Serum-globulin or Paraglobulin.

This constituent has been already discussed at considerable length
in relation to the subject of coagulation, and the reader is referred to
page 37 for the method of obtaining it from serum, as well as for
a discussion of the views which have been held as to its origin.

It was formerly held that serum-globulin was present in much
smaller quantities in the serum than serum-albumin. According
to Hammarsten, however, the older methods employed in the
separation of this substance were insufficient. He has discovered
that magnesium sulphate effects the complete precipitation of serum-
globulin, and therefore admits of the accurate determination of
its amount. In the following table are shewn the results of analyses

CHAP. It.]

THE BLOOD.

61

in which he determined the total quantity of proteids in the serum
and also the amount of serum-globulin; the quantity of serum-albumin
being found by subtracting the second result from the first. It will
be seen that according to Hammarsten the proportion of serum-
globulin to serum-albumin varies remarkably, in some cases (horse and
ox) the former being the more abundant constituent, in others (dog
and rabbit) the latter.

TABLE SHEWING THE AMOUNT OF SOLIDS, PROTEIDS, AND ALSO THE
RELATIVE PROPORTIONS OF SERUM-GLOBULIN AND SERUM-ALBU-
MIN IN 100 PARTS OF THE SERUM OBTAINED FROM THE BLOOD
OF VARIOUS ANIMALS1.

Solids.

Total
Proteids.

Serum-
globulin.

Serum-
albumin.

Serum-globulin

Serum-alb umi a '

Serum from horse

8-597

7-257

4-565

2-677

1

0-591

Serum from ox

8-965

7-499

4-169

3-329

1

0-842

0-907

7-filQ

^•103

4.^16

1

1-511

Serum from rabbit

7-525

6-225

1-788

4-436

1
2-5

By the term serum-casein some authors have designated the proteid
matter which is obtained from serum by adding a small quantity of acetic
acid to it after paraglobulin lias been precipitated by diluting and subjecting
to a stream of C02. We now know, however, that dilution with water and
the action of CO2 are not sufficient to precipitate all the serum-globulin
contained in the serum, and we cannot doubt that Panum's serum-casein
is merely serum-globulin which has escaped precipitation by CO2.3

2. Serum-albumin.

Having separated from the serum the serum-globulin which
it contains, there still remains in solution the most important and
usually the most abundant of its constituents, viz. serum-albumin.

In consequence of the serum-albumin which it contains, when
serum is heated to about 60° C. it becomes slightly opaque, full
coagulation occurring at 75°, the separation of the albumin being
accompanied by an increase in the alkaline reaction of the liquid.

1 Hammarsten, "Ueber das Paraglobulin." PMger's Archiv, 1878.

2 The reader who wishes to acquaint himself with the older statements as to serum -
casein is referred to Kiihne, Lehrbuch, p. 175, .and to .Gorup-Besanez, Lehrbuch d.
pkysiolog. Chemie, 1878, p. 119.

C2 METHODS OF PREPARING SERUM-ALBUMIN. [BOOK I,

When alcohol is added to serum in considerable excess, as in the
proportion of two volumes of absolute alcohol to one of serum, the
albumin is precipitated : at first the precipitate can be redissolved in
distilled water; by prolonged contact with the alcohol it becomes
almost absolutely insoluble. In order to secure the latter result as
perfectly as possible, as for instance in the preparation of solutions of
fibrin-ferment, the quantity of alcohol added must be much larger
than that indicated above, even 15 or 20 times as much alcohol as
serum being used, and the action of the former upon the latter
being continued for about three months.

The albumin recently precipitated by alcohol from serum, when
it is redissolved in water, yields a faintly opalescent liquid.

Prepara- Various methods have been suggested for the

tion of Serum- preparation of pure serum-albumin; they all yield a
albumin. substance, which can only be regarded as approximately

pure, inasmuch as they fail in separating inorganic salts which, more
or less, always continue to adhere to the substance and to modify
its physical properties.

Hoppe-Seyler's method.

Blood serum is treated with dilute acetic acid, as for the prepara-
tion of paraglobulin, and the clear liquid is filtered from the latter
body. The liquid is then concentrated by evaporation in shallow
basins at a temperature which must not exceed 40° C.

The concentrated liquid is neutralized with sodium carbonate and
is then placed in a dialyser suspended in distilled water, which
must be very frequently renewed. The dialysate is tested from
time to time with solution of silver nitrate; when this reagent no
longer produces a marked opalescence it is concluded that all dif-
fusible impurities, of which sodium chloride is the chief, have been
removed; the contents of the dialyser are then emptied into a flat
capsule and evaporated at a temperature not exceeding 40° C.

Prepared by this process, serum-albumin still contains from
0*5 to TO per cent, of salt, and is obtained in the form of a trans-
parent, yellowish, brittle solid, which breaks with a glassy fracture,
and which furnishes, when pulverised, a yellowish white powder.
It is soluble in distilled water, the solutions being slightly opalescent
and, when concentrated, viscous. Solutions of serum-albumin deviate
the plane of polarization to the left ; (A)j = — 56°. The specific
rotatory power is remarkably little affected by the presence of salts
or by the degree of dilution.

When dry, solid, soluble albumin, prepared by the previously
described method, is heated to 100° C. it is, after a considerable time
has elapsed, rendered insoluble in water.

Solutions of serum-albumin are not precipitated by carbon-
dioxide, by acetic or by orthophosphoric acid. They are precipi-

CHAP. II.] THE BLOOD. 63

tated by mineral acids, and especially by nitric acid; they are like-
wise precipitated by tannic acid and by metaphosphoric acid.

When heated to 60°C. solutions of serum-albumin usually become
opalescent, and at temperatures between 72° — 75° the albumin
separates in a flocculent form. Solutions of albumin which have
been long dialysed, but are not free from salts, are exceptions
to these statements. (See Schmidt's and Aronstein's pure albumin.)

Most metallic salts, such as mercuric chloride, copper sulphate,
lead acetate, at once precipitate ordinary serum-albumin.

Ether does not precipitate serum-albumin, whilst it does precipi-
tate

Schmidt's and Aronstein's pure serum-albumin and its reactions.

It was asserted by Graham1 tliat by mixing egg-albumin with acetic
acid and placing the mixture in a dialyser, all the acid, together with the
alkaline and earthy salts, diffused out, leaving the albumin pure, so that the
dried substance, on being ignited, left no ash. This albumin was found
by Graham to have a slightly acid reaction. Kiihne and Hoppe-Seyler were
unable to confirm the statement of Graham.

Arotistein2, working under the direction of A. Schmidt, asserted that
if serum be subjected to long continued dialysis, the whole of the para-
globulin is precipitated and the whole of the salts are removed from
the albumin, which when burned leaves no ash. He asserted that such
albumin when dissolved in water is not coagulated by boiling, and is not
precipitated by alcohol. The addition of small quantities of common salt
leads, according to Aronstein, to the albumin being again coagulable
by heat and by alcohol. These observations of Aronstein received the
full confirmation of A. Schmidt3, who, in addition, asserted that dilute
solutions of pure albumin, obtained by dialysis, gave no precipitate with
copper and zinc sulphate, with neutral lead acetate, with mercuric chloride
and many other salts which precipitate ordinary albumin. Platinum
tetrachloride, nitric acid, tannic acid, ferrocyanide of potassium and acetic
acid, were stated to be the reagents which most easily precipitate pure
albumin from its solutions.

The observations of Aronstein and Schmidt have not however been
confirmed.

Heynsius4 found it impossible to obtain any serum-albumin (by
Aronstein and Schmidt's process) free from ash, and attributed the non-
coagulation. in Aronstein's experiments to the presence of a slight alkaline
residue. Similarly Winogradoff5 repeating Aronstein and Schmidt's

1 Graham, " Liquid diffusion applied to analysis." Philosoph. Transact., 1861.

2 Aronstein, " Ueber die Darstellung salzfreier Album inlosungen vermittelst der
Diffusion." Pfliiger's Archiv, 1873, Vol. vm. p. 75.

3 A. Schmidt : " Untersuchung des Eiereiweisses und Blutserums durch Dialyse."
Ludwig's Festgabe, 1874, pp. 94—115. "Weitere Untersuchungen des Blutserums,
Eiereiweisses und der Milch durch Dialyse mittelst geleimten Papiers," Pfliiger's Archiv,
Vol. ii. pp. 1—52.

4 Heynsius, " Ueber die Eiweissverbindungen des Blutscrums und des HiOiner-
eiweisses." Pfliiger's Archiv, Vol. ix. pp. 514 — 552.

5 Winogradoff, "Darstellung und Eigenschaften salzfreier Eiweisslcsungeu."
Pfliiger's Archiv, Vol. n. p. 605.

64 THE EXTRACTIVE MATTERS OF THE LIQUOR SAXGUINIS. [BOOK I.

experiments, under the direction of Salkowski, failed to obtain albumin
free from ash. Huizinga1 by continued dialysis found that serum-albumin
contained from 0'36 to 0'56 p. c. of ash. Haas2 was able from the fluid
of ascites to obtain serum-albumin containing only 0'3 p. c. of ash; he
found that its solutions were precipitated by alcohol and by ether;
when boiled, the solutions always became opalescent, and often yielded
precipitates.

Are pep- It has been surmised that peptones, &c. which are

tones present formed in such large quantities in the alimentary
in the serum? canal, and which are doubtless absorbed into the
blood, may be present as peptones in that fluid. The most recent
research, of Drosdoff, shews that the presence of peptones cannot be
demonstrated with certainty even in the blood of the portal vein
taken whilst absorption is progressing3.

THE EXTRACTIVE- MATTERS OF THE PLASMA AND SERUM.

By the term extractive matters, physiological chemists formerly desig-
nated organic substances present in very small quantities in the various
solids and liquids of the body, and extracted from them by various liquids,
especially by alcohol, but which could not be obtained in a sufficiently
pure condition to admit of their identification as definite proximate
principles.

The progress of research has, to a very great extent, enabled us to resolve
the group of 'extractive matters,' obtained from most liquids and solids, into
its components ; still the term remains as a convenient one for the purpose
of grouping the organic constituents present in small quantities, and
capable of extraction by various liquids.

The extractive matters present in the plasma all pass into the serum.
These bodies, although present in small quantities in these fluids, are yet
possessed of the highest physiological importance. It is more in accordance
with the plan of the present work to consider at length the individual
extractive matters of the blood in connection with the functions of the
body with which they are most closely related, and the author therefore
limits himself in this place to little more than an enumeration. As
indicating the method of treatment which has been adopted, the reader
is informed that the fats of the serum will be considered under digestion
and in connection with the chemistry of the nervous organs, sugar when
discussing the functions of the liver, urea and uric acid in connection
with the secretion of urine, creatine and creatinine when treating of
muscle.

1 Huizinga, "Zur Darstellung des dialysirten Eiweisses." Pfliiger's Archiv, n.
pp. 392—402.

2 Haas : " Ueber das optische und chemische Verhalten einiger Eiweisssubstanzen,
insbeeondere <fer dialysirten Albumine." Pfliiger's Archiv, Vol. xn. pp. 378 — 410.
"Ueber die Eigenschaften des salzarmen Albumins," Prager medic. Wochenschrift,
1876, Nos. -84—36 ; also Maly's Jahresbericht, Vol. vi. (1877), p. 5.

3 Drosdoff, "Resorption der Peptone, des Rohrzuckers und der Indigoschwefelsaure
vom Darmcanal aus und ihr Nachweis im Blute der Vena portae." ZeitscJirift f. physiol.
Chem., i. 210—232.

CHAP. II.] THE BLOOD. 65

Neutral When serum is evaporated to dryness and the

ies11111 res^ue ^s powdered and boiled with ether, this liquid
terin extracts all the above bodies. Especially is this the

case when the serum presents, as it does some hours
after food, the milky appearance which has already been described.
The serum of fasting animals contains on an average about 0'2 per
cent, of fats and cholesterin, that of animals in digestion may
contain from 04 to 0'6 p.c. The fats present in the serum are
triolein, tristearin, and tripalmitin. It was formerly believed that in
addition to these fats, soaps, i.e. alkaline salts of the fatty acids, were
present in the blood. The incorrectness of this surmise has been
demonstrated by Rohrig1, who has shewn that soluble soaps cannot
exist in the blood.

Cholesterin, which will be studied in connection with the
chemistry of the nervous organs, may constitute 10 p.c. of the ether
extract of blood. It has been found by Hoppe-Seyler2 to vary in
the serum of the blood of fattened geese from 0*019 to 0'314 per cent.
Besides the fats and cholesterin the serum of blood always. contains,
according to Hoppe-Seyler, some lecithin; this ^substance will be
treated of under ' brain.'

Sugar. Glucose is a normal constituent of the blood, and is

contained in the serum which separates from it after coagulation. Its
amount appears, according to the most recent researches. (Abeles3,
Pavy4, v. Mehring5), to be nearly the same in the blood of all the
vessels with the one exception of the blood of the portal vein, which
contains an excess of sugar after the ingestion of a saccharine diet.

The quantity of glucose present in the blood of the dog was found
by Pavy to vary between 0*81 and 1*231 parts per 1000.

In the experiments of v. Mehring, the amount of sugar in the
serum of dogs was found to vary between 0'125 and 0'330 p.c. This
matter will be fully treated of under 'liver' and 'nutrition' (see also
Chapter IV.).

Creatine, creatinine, urea, carbamic acid, xanthine,
acid^Crea- ' hypoxanthine, uric acid and hippuric acid are found in
tine,' &c. the serum. The amount of urea present in the normal

blood of man varies between 0 02 and 0'04 p.c.

A yellow pigment is found dissolved in the serum of the blood of
man and most animals, although certairj. animals (e. g. the rabbit)
have colourless serum.

1 Rohrig, "Ueber die Zusammensetzung und das Schicksal der in das Blut eingetre-
tenen Nahrfette." Ludwig's Arbeiten, 1874.

2 Hoppe-Seyler, "Ueber das Vorkommen von Cholesterin und Protagon und ihre
Betheiligung bei der Bildung des Stroma der rothen Biutkorperchen." Med. Chem.
Untersuchungen, p. 145.

3 Abeles, " Der physiologische Zuckergehalt des Blutes." Med. Jahrbucher, Heft
in., 1875 ; also MaJj's Jahresbericht, Vol. vi. p. 95.

4 Pavy, " The Croonian Lectures on certain points connected with Diabetes.'
London, 1878.

5 v. Mehring, " Ueber die Abzugswege des Zuckers aus der Darmhohle." Archiv f.
Anat. u. Physiol. 1877, pp. 380—415.

a. 5

66 THE SALTS OF THE PLASMA AND SERUM. [BOOK I.

THE SALTS OF THE PLASMA AND SERUM.

When an organic liquid such as the plasma, the serum, or the
blood, is evaporated to dryness, and the dry residue is exposed to a red
heat in a crucible, the organic matters are oxidized and the products
of oxidation escape, leaving the inorganic or mineral matters behind.
We cannot, however, suppose that even the whole of the inorganic
matters originally present in the liquid can be obtained in this
way, for, however carefully we may proceed, there will always be more
or less volatilization of certain saline constituents as, for instance, of
sodium chloride. Still less are we justified in concluding from the
inorganic compounds left in the ash after the most cautious ignition,
that the same compounds were originally present in the liquid, for
they may only have been produced under the action of heat, and
at the expense of some constituents of organic bodies. We must
bear these considerations in mind in our judgment of the results
of analyses of the ashes of plasma and serum.

Much more reliable, however, is the information furnished us by
the direct precipitation of the inorganic constituents of the serum by
the addition of certain reagents to it. Under the direction of Ludwig,
Pribram1 and Gerlach2 have directly determined the amount of
calcium, magnesium, and phosphoric acid in the serum by a method
which will be found described in Chapter IV., and it is upon
their results that are alone based any accurate conclusions as to these
three constituents of the serum.

The following facts may be taken as resting upon a firm foundation.

1. The serum contains a somewhat smaller proportion of
inorganic salts than the liquor sanguinis, some being carried down
with, or perhaps more intimately associated with, the fibrin when it
separates.

2. The amount of inorganic matter left on the cautious ignition
of serum amounts to from 0'7 to 0'9 per cent.

3. The principal inorganic constituent of the ash of plasma and
serum, and not only of the ash, but of the liquids themselves, is
common salt, sodium chloride, NaCl; the amount present in the
liquid being about 0'5 per cent.

This salt constitutes, according to Lehmann, 61 per cent, and
according to Schmidt 65*2 per cent, of the ash left by serum. That
it is present before ignition is easily proved by the fact that crystals
of salt separate on concentrating serum.

4. In addition to sodium chloride, the ash of plasma and serum
contains, as its next most abundant ingredient, sodium carbonate.

It appears probable that the plasma and the serum before
ignition contain not sodium carbonate Na2CO3, but sodium hydric

1 Pribram, "Eine neue Methode zur Bestimmung des Kalkes und der Phosphorsaure
im Blutserum." Ludwig's Arbeiten, 1871.

2 Gerlaeh, "Ueber die Bestimmung der Minerale des Blutserums durch directe
Fallung." Ludwig's Arbeiten, 1872.

CHAP. II.] THE BLOOD. 67

carbonate NaHC03. The grounds for this supposition are (a) that
the plasma and serum contain considerable quantities of carbon
dioxide held partly in simple solution and partly in a state of feeble
combination as it is in sodium acid carbonate : (6) that when the
proteid matters are separated from the serum by adding to it a large
quantity of alcohol, and then solution of mercuric chloride is added,
there is produced a brown crystalline precipitate of oxychloride of
mercury (probably HgCl2, 4HgO), such as separates when mercuric
chloride is added to solutions of sodium acid carbonate. If the
sodium existed as normal sodium carbonate, a yellow precipitate of
HgO would be thrown down instead. (Liebig.)

5. The ash of plasma and of serum contains about 4 per cent, of
potassium chloride which, there is every reason to believe, exists as
such in these fluids before they are subjected to chemical treatment.

The great preponderance of salts of sodium as contrasted with
salts of potassium in the plasma and serum is one of the in-
controvertible and most interesting facts relating to the saline
constitution of these liquids.

Salkowsky found, in two cases, in the serum obtained from the
blood of healthy men, that the ratio of potassium to the sum of
potassium and sodium in the ash was respectively as 13 9 : 100
and 10'4 : 100, and A. Schmidt found the proportion to be in two
cases 7'6 : 100 and 8'6 : 100.

6. In addition to the inorganic constituents referred to, the ash of
plasma and serum is found to contain sulphuric and phosphoric acids and
magnesium and calcium ; and arranging the results of the analyses in
accordance with the rules followed in such cases it would result that
the serum contains ortho-phosphates of calcium, magnesium and
sodium, as well as a small quantity of potassium sulphate.

We can, however, have no certainty from the results of such
analyses as to the constitution of phosphates existing in the unaltered
liquids. Admitting for instance that sodium compounds of orthophos-
phoric acid (H3PO4) exist in the plasma and serum, analysis in no way
allows us to decide whether the compound present is the neutral tri-
sodium phosphate Na3P04 or the alkaline hydrogen di-sodium phos-
phate HNa2P04 or H2NaP04, of which the second, for other reasons,
has been supposed to exist in the blood and probably actually does
so. Again, the serum contains appreciable quantities of lecithin
or some other derivative of glycerin-phosphoric acid. When that
body is ignited it leaves metaphosphoric acid,which reacting upon alka-
line and earthy carbonates would produce salts which in the analyses
would be reckoned as phosphates. The question therefore arises whether
the phosphoric acid which is found as a constituent of the ash really
exists as such in the liquor sanguinis or whether it is there present as
one of the products of the oxidation of such an organic body as lecithin.

The observations of Pribram and Gerlach allow u-s to decide the
question. These observers have proved that calcium, magnesium

5—2

68

THE SALTS OF THE PLASMA AND SERUM. [BOOK I.

and phosphoric acid may by suitable methods be directly precipitated
from, and correctly estimated in, the serum. They find that whilst
the amount of calcium and magnesium estimated by direct precipita-
tion agrees exactly with that determined by analysis of the ash of the
same quantity of serum, the phosphoric acid does not so agree; in
other words the amount of phosphoric acid found in the ash of serum
is much larger than that directly precipitated from it. Furthermore
Pribram, and after him Gerlach, found that after precipitating directly
all the phosphoric acid in the serum, and precipitating all proteids
by absolute alcohol, on igniting the filtrate its ash contained phosphoric
acid ; the amount thus found added to that directly precipitated
agreed very closely with the total quantity of phosphoric acid deter-
mined in the ash of serum. Thus in one experiment,

100 c.c. of serum yielded on direct ignition 0*056 p,c. PS06
„ „ by direct precipitation 00108 „

„ „ in the alcohol extract 0'0325 „

„ „ by ignition of the ex-

tracted residue O'OOGO „

0-0493 p.c.

From this experiment, which may be taken as a type of many
others, it results that serum contains a much smaller quantity of
phosphoric acid than is present in its ash and that the greater part of
the phosphoric acid in the latter is derived from an organic compound
soluble in alcohol. The latter result had to a considerable extent
been anticipated by Sertoli1.

The phosphoric acid obtained on direct precipitation of the serum
doubtless exists as an inorganic compound, but we cannot assert its precise
condition ; probably it does not exist as calcium phosphate. At any rate
the whole of the phosphoric acid existing in the serum is not sufficient to
combine with the calcium of that fluid to form the compound Ca32(PO4),
as will be learnt by a study of the following results obtained by Pribram.

RESULTS OF SEVEN DETERMINATIONS OF THE PHOSPHORIC ACID
(CALCULATED 'AS P206) AND LIME (CaO) IN 100 c.c. OF SERUM.

By direct

Determined in

precipitation.

Ash.

CaO

PA

CaO P2O5

A

0-0174

0-0124

0-0177

0-0387

B

0-0216

0-0121

0-0230

0-0448

C

0-0150

0-0106

0-0170

0-0320

D

0-0173

0-0171

E

0-0200

0-0195

F

0-0188

0-0200

G

0-0155

0-0108

0-0170

0-0559

1 Sertoli, " Ueber die Bindung der Kohlensaure im Blute und ihre Ausscheidung in
der Lunge." Medicinisch-Chemische Untersuchungen von Hoppe-Seyler. Heft in. (1868)
p. 350 et seq.

CHAP. II.]

THE BLOOD.

If we calculate what quantity of P2O5 would have been found had the
compound Ca32(P04) existed in the serum we obtain the following results.

A would have contained 0-0146 P2O5
B „ „ 0-0182 „

C „ „ 0-0127 „

G 0-0131

The following tabular views represent the constitution of the saline
constituents of the liquor sanguinis and serum as derived from the most
reliable researches. It is to be remembered that in reference to the amount
of phosphoric acid the older analyses of Lehmann and Schmidt are un-
reliable, and that for this the results of Pribram and Gerlach are to be
taken.

I. COMPOSITION OF ASHES OF SEKUM (LEHMANN).

100 parts contain

Sodium chloride ....

61-087

Potassium chloride ....

4-085

Sodium carbonate . . .

28-880

Sodium phosphate ....

3-195

Potassium sulphate ....

2-784

100-031

II. COMPOSITION OF THE SALTS OF THE PLASMA (SCHMIDT).

1000 parts of plasma yield

Sodium chloride ' 5-546

Sodium phosphate calculated as Na3P04 . 0-271
Sodium in other states of combination calculated

asNa2O 1'532

Potassium chloride ....
Potassium sulphate ....
Calcium phosphate .....
Magnesium „ .

8-505

70 THE SALTS AND GASES OF THE SERUM AND PLASMA. [BOOK I.

III. COMPOSITION OF THE SOLUBLE SALTS YIELDED BY
SEBUM OF OX'S BLOOD (SEBTOLI).

1000 grammes of serum yielded

Cl 3-270

Na calculated as in combination with Cl . 2-120
!NTa calculated as Na,.2O but existing as chloride 1-291

K calculated as K2O 0'224

P2O5 representing the phosphoric acid actually

existing in the liquid . . . . 0'025
SO3 representing the sulphuric acid existing in the

liquid 0-305

Sertoli arranges the results of the above analysis
as follows, so as to represent the probable constitution
of the soluble salts of the serum : —

1000 parts of serum yield

Sodium chloride . . . . 5*39 grms.

Sodium sulphate . . . . 0-24 „

Disodium hydric phosphate (ISra2HPO4) 0-05 „
Na, calculated as NaaO (existing as carbonate

or bicarbonate) . . . . 1-16 „

Potassium sulphate . . . . 0'414 „ •

IV. RESULT OF THE ANALYSES OF PELBBAM OF THE CALCIUM AND
PHOSPHOBIC ACID EXISTING IN SEBUM (DETEBMINED DIBECTLY).

1000 grammes of serum yield

Phosphoric acid corresponding to . 0-179 grms. P2O6
Calcium corresponding to . . 0'173ofCaO

V. RESULTS OF THE ANALYSES OF GEBLACH OF THE
MAGNESIUM EXISTING IN SERUM.

1000 grms. of serum yielded

(1) Magnesium corresponding to . 0-025 grms. of MgO

(2) „ „ to . 0-027 „

THE GASES OF THE PLASMA AND SERUM.

It will be convenient to postpone a lengthened consideration
of this subject to the section which treats of the gases of the blood
as a whole, and to the chapter on Respiration. In this place it will
suffice to make the following remarks :

When the blood is heated in a Torricellian vacuum it gives up
more than half its volume of a mixture of gases, composed of oxygen,
carbonic acid, and nitrogen, which in the blood itself were contained

CHAP. II. J THE BLOOD. 71

partly in a state of solution and partly in the form of feeble chemical
compounds.

Whilst the oxygen which is contained in the mixed gases is derived
wholly from the decomposition of the oxy-haemoglobin of the blood
corpuscles, the greater part of the carbonic acid and the whole of the
nitrogen are derived from the plasma, in which they exist mainly in a
state of simple solution, though the carbonic acid is in part in a state
of feeble combination, probably in the form of sodium hydric carbo-
nate, NaHCO,.

SECT. 4. THE COLOURED CORPUSCLES OF THE BLOOD.

Shape, size, The red colour of the blood of vertebrates is due to

&c. of the suspension in a colourless or slightly coloured liquid

coloured Of large numbers of solid bodies, of which the principal

sorpuscies. solid ingredient is a red colouring matter, haemoglobin.

In the blood of man and the mammalia generally1, the coloured
blood corpuscles are non-nucleated biconcave disks, whilst in the blood
of birds, reptiles, and most fishes they are nucleated, elliptical, bicon-
vex bodies.

The size of the coloured blood corpuscles of the various orders of
mammals varies somewhat, though with some exceptions not within
very wide limits. The red blood corpuscles of man are amongst the
largest, being for instance larger than those of any domestic animal
inhabiting Europe.

The diameter of the average red blood corpuscles of human blood
is about the y^th of a millimetre, or about 7'9/£2, and the thickness
about 1'8/Aj expressed in English measurements the average coloured
blood corpuscle measures about s^^th °f an inch in diameter and
about y^oijth °f an mcn in thickness (Gulliver)3.

By means of a method which could only yield rough ap-
proximations to the truth, Welcker4 determined the approximate

MM. Cube

volume of a human coloured blood corpuscle to be 0-000,000,072 or
seven ten-millionths of a cubic millimetre, and the approximate
superficial area to be about 0'000128 or rather more than one ten-
thousandth of a square millimetre.

1 In the blood of the Camelidae the red corpuscles are oval.

2 The Greek letter /* is now employed to represent the micro-millimetre, or 1000th
part of a millimetre, which is taken as a convenient unit for microscopic measurement.
The micro-millimetre corresponds to about T^mnnr^8 of an English inch or more
accurately to 0-00003937 of an inch.

3 By far the most complete set of measurements of the corpuscles of the verte-
brata was made by Mr Gulliver, and the results of his researches were collected
together and published as a note (cxvin) to his edition of Hewson's works. They are
transcribed, with some additions, by Milne-Edwards, Lemons sur la Physiologie, Vol. i.
p. 84 et seq.

4 Welcker : for an account of his method followed see Strieker's Human and Com-
parative Histology, Vol. i., Article "Blood," by Rollett, p. 383.

72 THE STRUCTURE OF THE RED BLOOD CORPUSCLES. [BOOK I.

The views The structure of the red blood cell has been a matter of

of observers the greatest interest to histologists and physiologists, chiefly,
•espectingtne it must be confe^a, On account of the important part the
cotoured6 ° red blood corPuscle has played in the Theory of Cells. A
blood corpus- brief review of the opinions which have been held will
des. here be attempted.

One view of the structure of the red disk in man is to
be mentioned rather because it is curious than because it is important.
Delia Torre1 sought to explain the well-known optical characters of the
centre of the disk by supposing that the red corpuscles were in reality little
rings. Excluding this view, the remaining discussions of the structure of
red blood corpuscles may be grouped under two main questions and a sub-
sidiary one. Has the red corpuscle of man a mwleus ? Has the red cor-
puscle generally a cell-membrane ? And the corollary question to the
latter, Does the red colouring matter reside in the contents of the vesicle or
in the membrane ?

A nucleus was early detected by Hewson in the red blood corpuscles
of the frog, and for some time it was tacitly assumed that a similar struc-
ture was to be found in the corresponding corpuscles of mammalia. But
it should be clearly kept in mind that, from the first, it was analogy rather
than direct observation which supported this view. Accordingly we find
the red corpuscles of man playing the part of a non-nucleated cell in the
reform of the old Cell Theory which was consummated in 1861. In more
recent days the only supporter which the original doctrine has found has
been Bottcher2. For the present, therefore, we may regard the red cor-
puscles of mammals as non-nucleated.

The question of the membrane was longer and more hotly debated
than that of the nucleus. As early as 1685 the red corpuscles were spoken
of as vesicles3; and by many subsequent writers, among whom we may
mention Hewson and Wells4, the same doctrine is openly held. Wells
is said to have been the first to discuss the question systematically, bringing
in support of his view the facts of the action of water and saline solutions
upon the red cell. To the names of Hewson and of Wells we must add
that of C. H. Schultz, who was believed by Schwann5 to have been the
first to demonstrate the vesicular nature of the blood cells. Schwann
himself (fee. cit.), as is well known, maintained the same view; and for
many years afterwards it was the prevailing doctrine. Excluding the
botanical prejudices of Schwaim's Cell Theory, the only definite grounds
for the belief in the vesicular nature of the red cell were the appearance
of the cells when irrigated with water and solutions of salts. In the
case of the former, the cells swell up and become globular, with a diameter
less than the long, and greater than the short, diameter of the original
flattened disk. In the case of saline solutions of greater density than
the normal blood plasma the outer surface of the corpuscles assumes a

1 Delia Torre, Nuove osservazioni microscopiche. Naples, 1776. Milne-Edwards,
Le$ons, Vol. i. p. 63.

2 Bottcher, Virchow's Archiv, Bd. xxxvi. and xxxix. (See Article "Blood Corpuscles"
in Strieker's Handbook.) Quarterly Journal of Hicroscop. Science. N. S., Vol. xvn.
1877, p. 377.

3 Bidloo, Anatomia humani corporis, 1685. Milne-Edwards, Legons, Vol. i. p. 66.

4 Wells, " On the colour of the blood." Phil. Trans. 1797, p. 429. Milne-
Edwards, Lemons, Vol. i. p. 66.

5 Schwann, "Microscop. Researches." Syd. Soc. 1847, p. 67.

CHAP. II.] THE BLOOD. 73

folded or creased appearance like art ill-fitting glove. It cannot be denied
that these appearances are, at first sight, strikingly suggestive of a mem-
branous envelope ; but how little they prove the existence of such an
envelope the reader will find in Briicke's criticism of the question, too
long to be reproduced or even epitomized in this place1. With this under-
mining of the main support of the vesicular theory, its essential weakness
became evident. In the first place it is almost inconceivable that a fluid-
filled vesicle with walls which may collapse, should maintain the shape
of a biconcave or biconvex disk. In the second place, notwithstanding
the frequency of the search, no one has yet detected a structure at all
resembling the empty husk or skin of a red blood corpuscle. Indeed,
by alternately freezing and thawing blood, the coloured contents may be
extracted from blood corpuscles, leaving a colourless structure often exactly
resembling in shape and elastic property its red original. Of similar
import is the observation that the red corpuscles of amphibia may be cut
with a fine razor without the escape of any coloured contents2; as well
as the observations that the corpuscles may be eviscerated of their nuclei,
becoming non-nucleated, but still coloured, spheroids; and that two coloured
cells may actually become fused into one3. When to these considerations
we add that no one has ever observed a double contour around the red
blood cells, even when they are swollen and spherical under the influence
of water, and that the attention of Schwann himself was arrested by its
absence4, we may acknowledge how very doubtful the alleged membrane
of red blood cells has always been.

Most observers who adopted the view of the vesicular nature of red
blood corpuscles believed that the envelope included the coloured contents.
But the membrane was acknowledged to be slightly tinted red by Schwann5,
who remarked that, were it not so, the biconcave centre of the red cell
of man would appear colourless. On the other hand Prevost and Dumas6,
who succeeded in rupturing the corpuscles so as to permit the escape of
the nucleus, advanced the opinion that the colouring matter was not in
the contents, but in the skin.

While, however, the prevailing idea of the red cell was that of a vesicle,
there were not wanting other ideas of it which approached the modern
one. Blumenbach7, in 1797, taught that the globules were small semi-solid
or gelatinous, lenticular masses, as did also de Blainville8; and, later,
Donne9 adopted a similar view.

1 E. Briicke, "Die Elementarorganismen." Sitzungsber. d. k. Akad. Wien,Vol.
XLIV. Abth. ii. p. 387.

2 Krause, Menschliche Anatomie, 1876. 3rd edit. Vol. i. p. 328.

3 Article "Blood," by Bollett in Strieker's Handbook. Syd. Soc. Trans. Vol. i.
p. 391.

4 Schwann, loc. cit. p. 69.

5 Loc. cit.

6 Prevost and Dumas, " Examen du sang et de son action dans les divers phe'no-
menes de la vie." Biblioth. univer. de Geneve, xvn. pi. in. fig. 3. Milne-Edwards,
Legons, Vol. i. p. 67.

7 Blumenbach, Institutions physiologiques, traduit par Paget, 1797, p. 9. Milne-
Edwards, Legons, Vol. i. p. 67.

8 de Blainville, Cours de phijsiologie, i. p. 214. Milne-Edwards, Legons, Vol. i.
p. 67.

9 Donne, These sur les globules du sang, 1831, p. 13. Milne-Edwards, Legons,
Vol. i. p. 67.

74 COLOURED CORPUSCLES. THEIR ENUMERATION. [BOOK I.

To day, this simple notion is enlarged into the doctrine of the 'stroma.'
The red blood cell consists of a cavernous mass or 'stroma,' denser at
the periphery than at the centre, whose external limit or boundary appears
as a sharp contour. It is colourless and highly elastic : it is albuminous
in substance, and generally admitted to be non-contractile. In the central
trabeculae of the mass the nucleus is embedded, in those red corpuscles
which are nucleated. The interstices are quite filled by the coloured
substance of the corpuscle, which, under certain conditions (e.g. cautious
irrigation with water, or with boracic acid of 2 p.c.), retreats from the edge
upon the centre in a more or less regular manner. The stroma has
been called by Briicke the oekoid, and the contained coloured matter the
zooid. The special appearances upon which this view is founded have
already been stated. The view is not inconsistent with any of the known
reactions of blood corpuscles ; and it is especially adapted to interpret the
concentration of the zooid in the interior of the oekoid l.

Enumeration of the corpuscles.

Principle It might at first appear hopeless to attempt to

^th^110^3^ count the number of the blood corpuscles in the
enumeration blood, especially when we mention at the outset that
are based. one cubic millimetre of blood is estimated to contain

about 5 millions of corpuscles. But the possibility of
carrying out the process so as to permit of a fair approximation
becomes evident so soon as the principle is grasped, upon which all
the methods are based, all being but modifications of the method
suggested by Vierordt and first carried out in all detail by that
observer and by Welcker, whose results have received full confirma-
tion by the numerous researches carried on by the aid of the more
easy methods of Malassez, Gowers, &c.

The principle, then, is to dilute a known volume of blood with a
sufficient but definite and known quantity (say 100 times its volume)
of some colourless transparent solution which will not destroy the
blood corpuscles, and which will affect their shape as little as possible ;
thereafter to take a known and very minute volume of the diluted
blood, and with the aid of suitable micrometric arrangements to
count the corpuscles in it.

The methods will now be described in detail.

Vierordt "A measured volume of blood is diffused as equably

and Welcker's as possible in a thousand times its volume of an indifferent
method . fluid (six grammes of NaCl in one litre of water, according

to Welcker). A small quantity of the mixture is taken up in a capillary
tube of known calibre, and the length of the thread of fluid is estimated
under the microscope by means of a micrometer. When the contents
of the tubule have thus been ascertained, they are quickly distributed

1 For fuller information the student is referred to the article on "Blood" by Pro-
fessor Rollett, in Strieker's Handbook.

2 Welcker, " Grosse, Zahl, Volum, Oberflache, und Farbe der Blutkorperchen bei
Menschen und bei Thieren." Henle u. Pfeufer's Zeitochrift fur rat. Medicin, Dritte
Reihe, VoL xx. Heft 1 and 2, page 257.

CHAP. II.]

THE BLOOD.

75

with a little solution of gum upon a slide, and the whole is allowed
to dry. The preparation is covered with a micrometer divided into
squares, and the corpuscles in the several squares can then be successively
counted1."

Method of In this case, as in all other methods, the blood, which is

Malassez2. generally obtained by pricking the finger, is mixed with a

known volume of a suitable solution. This is effected by means of the little
pipette A, which is specially constructed for the purpose; the longer
portion of the tube through which fluid is sucked is of capillary diameter
and bears the mark 1 near the bulb.

On the shorter tube of the pipette, to which an indiarubber tube
is attached for facilitating aspiration, is marked the letter C. The bulb
of the pipette contains a glass bead.

Blood is now drawn into the pipette to the mark 1, then a solution
of sodium sulphate or Potain's solution is aspirated until it reaches the
level of C ; the blood and its diluent are then mixed by agitating
the pipette, the bead facilitating the process. The mixture is next
introduced into a flattened capillary tube which is connected to a glass
slide (B) and which has been accurately calibrated so as to determine the
cubic capacity of various lengths of the tube in fractions of a cubic milli-
metre. The results of the calibration are engraved on the glass slide. Thus in

A.

B.

FIG. 12. MALASSEZ' APPARATUS FOB THE ENUMERATION OF BLOOD CORPUSCLES.

A. Pipette in which blood is measured and diluted (Melangeur).

B. Flattened capillary tube fixed to slide, and calibrated.

1 Strieker's Human and Comparative Histology, Art. "Blood," by Eollett, Vol. i.
p. 383 ; see also for a description of Vierordt's original method, Gscheidlen's Physio-
logische Methodik, p. 378.

2 Malassez, " De la numeration des globules rouges du sang chez les mammiferes,
les oiseaux et les poissons." Comptes Rendus des Seances de VAcad. des Sciences.
2 Decembre, 1872.

76

ENUMERATION OF BLOOD CORPUSCLES.

[BOOK i.

the diagram those to the left indicate lengths in micro-millimetres, those
to the right capacities in fractions of the cubic millimetre.

Thus the numbers in the first row indicate that 500 /A of the tube contain

r-^- of a cubic mm., again the numbers in the second row indicate that
iou

450 /x contain ^-^ of a cubic mm.
lob

The number of corpuscles in a known length of the tube has now
to be counted. In order to do so the slide B is placed upon the stage
of a microscope whose eye-piece is provided with a micrometer ruled in
squares. As a preliminary, however, observations must be made to find
the value of the squares of the eye-piece micrometer in terms of a stage
micrometer divided into millimetres. By drawing out or in the draw-
tube of the microscope the side of the large square is made to correspond
exactly with that of the number of micro-millimetres of the stage micro-
meter, placed on the slide.

For instance one side of the large square (which is divided into
100 smaller squares) is made to correspond to 500 or say 450 /u,. The slide
with the capillary tube is now substituted for the stage micrometer and
the number of corpuscles contained in a certain number of the smaller
squares is counted.

FIG. 13. THE CAPILLARY TUBE OF MALASSEZ' APPARATUS. Viewed in a microscope
provided with an eye-piece micrometer ruled in squares. Magnifying power 100
diameters. (Copied from Banvier.)

Knowing first the length of the tube covered by the squares, second
the capacity of this length of the tube, we can by an easy calculation
ascertain the number of corpuscles in a cubic millimetre of the diluted, and
therefore also of the undiluted, blood. According to Malassez his method

CHAP. II.]

THE BLOOD.

77

occupies from first to last not longer than ten minutes and the mean possible
errors amount to 2 or 3 per 100.

Method of This method is almost exactly similar to that to be

Hayem and next described at length as Dr Gower's. The diluted blood
Nachet. js introduced into a cell of exactly known depth, and the

number of corpuscles is counted by means of an eye-piece micrometer
similar to that used in the method of Malassez.

Method of Dr Gowers has modified the instrument of MM. Hayem

Dr Gowers. and Nachet and to it has given the name of the Haema-
(The Haema- Cy tometer l.

cytometer.) r^^ following description of his' method is taken from an

article by Dr Gowers in the Lancet for December 1, 1877 : —

"The Haemacy tometer consists of (1) A small pipette, which, when
filled to the mark on its stem, holds exactly 995 cubic millimetres. It is
furnished with an indiarubber tube and mouthpiece to facilitate filling
and emptying. (2) A capillary tube marked to contain exactly 5 cubic
millimetres, with indiarubber tube for filling, &c. (3) A small glass
jar in which the dilution is made. (4) A glass stirrer for mixing the blood

FIG. 14.

A. Pipette for measuring the diluting solution.

B. Capillary tube for measuring the blood.

C. Cell with divisions on the floor, mounted on a slide, to which springs are fixed
to secure the cover-glass.

D. Vessel in which the solution is made.

E. Spud for mixing the blood and solution.

F. Guarded spear-pointed needle.

1 The Haemacytometer is made and sold by Mr Hawksley, Surgical Instrument
Maker, 300, Oxford Street, London, W.

78 ENUMERATION OF BLOOD CORPUSCLES. [BOOK I.

and solution in the glass jar. (5) A brass stage plate, carrying a glass
slip, on which is a cell, i of a millimetre deep. The bottom of this is
divided into -f^ millimetre squares. Upon the top of the cell rests the
cover-glass, which is kept in its place by the pressure of two springs
proceeding from the ends of the stage plate.

" Various diluting fluids l have been recommended in order to change
as little as possible the aspect of the corpuscles. It is not well, however,
to observe the characters of the corpuscles during the numeration. What-
ever solution be employed, the corpuscles are more or less changed by it.
One which answers very well is a solution of sulphate of soda in distilled
water, of a specific gravity of 1025.

"The mode of proceeding is extremely simple. 995 cubic millimetres of
the solution are placed in the mixing jar ; 5 cubic millimetres of blood are
drawn into the capillary tube from a puncture in the finger, and then blown
into the solution. The two fluids are well mixed by rotating the stirrer
between the thumb and finger, and a small drop of this dilution is placed
in the centre of the cell, the covering glass gently put upon the cell,
and secured by the two springs, and the plate placed upon the stage of the
microscope. The lens is then focussed for the squares. In a few minutes
the corpuscles have sunk to the bottom of the cell, and are seen at rest on
the squares. The number in ten squares is then counted, and this multiplied
by 10,000 gives the number in a cubic millimetre of blood.

" The average of healthy blood was decided by Vierordt and Welcker
to be 5,000,000 per cubic millimetres, and later results agree with this
sufficiently nearly to justify the adoption of this number as the standard, it
being remembered that in a healthy adult man the number may be a little
higher, in a woman a little lower."

By employing the methods previously described the following
results have been obtained :

Welcker found a cubic millimetre of the healthy blood of man to
contain 5,000,000 corpuscles. Malassez found the number to vary
between 4,000,000 and 4,600,000, the average being about 4,500,000.

Malassez has determined the total number of corpuscles in the
blood of a variety of animals, and he has determined for each
the number of corpuscles corresponding to a unit weight (one
gramme) of the body. This number he proposes to designate as
the ' corpuscular capacity ' (' capacite globulaire ') of the blood. In
the case of man the corpuscular capacity amounts to 341,000,000,

1 The following diluting solutions have been employed : —

a. A solution of sulphate of soda and distilled water of specific gravity 1025.

b. Potain's Solution : — Solution of gum acacia sp. gr. 1020. One volume.
Equal parts of sulphate of soda and chloride of sodium, in solution of specific

gravity 1020. Three volumes. Mix.

c. Keyes' Solution: —

" Take urine slightly phosphatic, easily obtainable after a meal, about 1020 sp. gr.,
and make of it a saturated solution of borax. Clouds of earthy phosphates are thrown
down. Filtration yields a clear alkaline fluid of sp. gr. about 1030. Add water
enough to reduce the specific gravity to 1020, and the fluid is ready for use."

CHAP. II.] THE BLOOD. 79

whilst the total number of corpuscles amounts to about 22,500
milliards1. Admitting the superficies of each blood corpuscle to be
j^^ths of a millimetre square, then the total superficies of the blood
corpuscles of men would amount to about 2880 square metres, i.e.
to the area of a square, each of whose sides is about 53 '66 metres
long.

Density and weight of the coloured corpuscles. According to C.
Schmidt the coloured corpuscles of the blood have a specific gravity
of 1089, and according to Welcker of 1105.

The student may feel some curiosity to know the method
which was followed in making these determinations, and it may be
said at once that the above results were obtained by calculation, and
that their correctness depends upon the reliability of several data.
Assuming that we know 1st the weight of the moist corpuscles in
a known volume of blood, 2nd the specific gravity of the same
blood defibrinated, 3rd the specific gravity of the serum, and if
we further assume that the specific gravity of the serum does not
differ sensibly from that of the plasma, we have all the data required.
Thus to take an example from Lehmann's Physiological Chemistry :
If we assume, for instance, that a specimen of blood contains 496 parts
of moist cells per 1000, besides 4 parts of fibrin, that the specific gravity
of the serum is 1028, and that of the defibrinated blood 1057*4,
then we may very readily determine the density of the blood cells by
the following considerations :

1000 — 4 = 996 parts of defibrinated blood occupy the space of

941'93 parts of water,

500 parts of serum ... „ 486'38

hence 496 parts of blood cells ... „ 45 5 '5 5 parts of water.

455 55 : 496 :: 1000 : 1088'8.

The density of the blood cells in this specimen of blood must there-
fore be 1088'82. It must be remarked, that the weight of the moist
blood corpuscles can only be determined in an approximately accurate
manner.

Welcker has calculated the approximate weight of the coloured
blood corpuscle to be O'OOOOS or TGiwinr ths of a milligramme3.

Summary of the' composition of the coloured corpuscles.

Before describing the principal organic and mineral constituents of
the red blood corpuscles it will be convenient to place before the reader
the following analytical data.

1 A milliard (Fr.) is one thousand millions.

2 Lehmann, Physiological Chemistry, Vol. n. p. 163.

3 Welcker, loc. cit. p. 274.

80 THE CHEMICAL CONSTITUENTS OF THE STROMA. [BOOK I.

I. According to Carl Schmidt 1000 parts of moist blood corpuscles
contain —

Water 688 parts.

Solid constituents &F"™ 30^* »

(Mineral . 8'12 „

II. According to Hoppe-Seyler and Jiidell1

100 parts of dried corpuscles contain —

Human Blood.^ Blood of Dog< Blood of ^^

Proteids . 12-24 . 5-10 . 12-55 . 3641
Haemoglobin 86*79 . 94-30 . 86'50 . 62-65
Lecithin . 0«72 . 0-35 . 0«59 . 0-46

Cholesterin . 0'25 . 0'25 . 0'36 . 0'48

We shall now more in detail examine the various proximate con-
stituents of the corpuscles.

The Stroma and the proteids associated with it.

Mode of I* nas been asserted (p. 74) that histologists have

obtaining the abandoned the view that the coloured corpuscle is a
stroma for vesicular body possessed of a cell-wall enclosing more
microscopic or }ess 1™^ contents, and have come to consider the
examination. -. , * -, •, . . TIP i />

coloured corpuscle as being a viscous solid formed ot

a stroma or framework in which are imbedded the other proximate
principles.

In order to demonstrate the existence of the stroma, defibrinated
blood is allowed to flow drop by drop into a platinum or silver dish,
which is cooled to — 13° C., by immersion in a freezing mixture, care
being taken that the blood contained in the capsule is frozen before
more is added. The frozen blood is then thawed and heated to 20° C.
The process of freezing and thawing may with advantage be repeated
several times. The blood will be then found to have lost its opaque
red colour and to present the appearance of a transparent lake-
coloured fluid. On microscopic examination the non-nucleated
coloured blood corpuscles are found deprived of all colour, sometimes
retaining their original shape, but more frequently either more globular
or more shrivelled than normal. The stromata retain, according to
Rollett, the extensibility and the elasticity of the original blood
corpuscles. Under the influence of the changes of temperature the
haemoglobin has entirely dissolved in the serum, leaving the colour-
less stroma in which it had been deposited.

The stroma is insoluble in serum, dilute solution of salt and of sugar,
and in distilled water at a temperature below 60° C., but readily
soluble in serum containing alcohol, ether, or chloroform : in solutions
of caustic alkalies: and in solutions of alkaline salts of the bile acids
(Kiihne).

1 Jiidell, " Zur Blutanalyse." Hoppe-Seyler's Med. Chem. Untersuchungen. Heft
in. p. 386.

CHAP. II.] THE BLOOD. 81

Mode of When defibrinated mammalian blood is mixed with

separating ten times its volume of a solution of sodium chloride

the proteids (made by mixing 1 volume of a saturated solution of
of the stroma.

the blood corpuscles are, for the most part, deposited as a slimy
precipitate. The fluid is decanted from the precipitate, which is
again treated with the same weak solution of common salt and
set aside for a day, when, after decanting the fluid, the corpuscles are
obtained almost absolutely free from adhering serum. By employing
the centrifugal machine in effecting this separation the whole process
from first to last may occupy only a few hours. If the precipitate
obtained in this way is now treated with water without being disturbed,
the haemoglobin contained in the corpuscles is dissolved arid there
remains behind a gelatinous mass, which may be shaken with water
and ether, and then separated by filtration. The body thus obtained is
insoluble in water, soluble for the most part in a very weak solution of
sodium chloride, and in water which contains 0*1 p.c. of HC1, and in
weak solutions of alkalies. This body possesses all the characters of
the globulins, and is said by Kiihne to act fibrinoplastically1 ; he
considers it to be paraglobulin. Instead of employing the above
method, which we owe to Hoppe-Seyler2, we may adopt a simpler
method recommended by Kiihne3, and having separated the corpuscles
as completely as possible from the serum (in this case too the cen-
trifugal machine should if possible be used) these are treated with a
large quantity of water. The solution is then subjected, to a stream
of 002 as long as white flakes continue to separate. The portion of
this precipitate which is soluble in water holding oxygen in solution
is composed of paraglobulin.

According to Kiihne the red blood corpuscles were to be looked
upon as the chief source of the paraglobulin of the blood, and this
view was at one time shared by A. Schmidt. This author now,
however, refers all the paraglobulin of the serum to the breaking
down of the colourless corpuscles4.

Peculiar! ^ ^ie ^lood of the newt or frog be placed in a

ties of the microscopic gas chamber5 and subjected to the action

stroma of the of a stream of CO2, the nucleus, which was at first
nucleated scarcely, if at all, visible, becomes beautifully distinct

coloured an(j somewnat granular ; if a stream of oxygen or atmo-

spheric air be then substituted for the C02 the nucleus

1 We have seen that, according to Hammarsten, there is reason to doubt the
existence of any specific fibrinoplastic substance, the separation of fibrin being brought
about under certain circumstances by other bodies than paraglobulin. See p. 51 et seq.

2 Hoppe-Seyler, Handbuch d. physiologisch- und pathologisch-chemischen Analyse.
3te Auflage, Berlin, 1870, p. 318.

3 Kiihne, Lehrbuch dcr physioL Chemie, p. 193.

4 "Ueber die Beziehungen des Faserstoffes zu den farblosen und den rothen Blut-
korperchen und iiber die Enstehung der letzteren." Pfluger's Archiv, Vol. rx., p.
353—358. Maly's Jahresbericht, Vol. iv., p. 122.

5 See "Blood Corpuscles," by Dr Klein, Handbook for the Physiological Laboratory,
p. 17.

6

82 THE CHEMICAL CONSTITUENTS OF THE NUCLEI. [BOOK I.

disappears. Occasionally this appearance and disappearance may be
observed to occur many times in succession. It is pretty obvious that
this phenomenon is due to the coagulation by the C02 of a proteid
existing around the nucleus, and which is probably identical with
paraglobulin, the re-solution under the influence of oxygen agreeing
with the known characters of that body.

The Nuclei of the Red Corpuscles.

If we except the blood of adult mammals that of all other vertebrates
contains red corpuscles possessed of a nucleus. This may, whilst
the corpuscle is living and unaltered, be scarcely if at all perceptible,
but readily comes into view when weak acids or carbon dioxide exert
their action.

Composed ID Order to investigate the chemical composition

according to of the nuclei of coloured blood corpuscles, the blood of
Brunton of a birds (and also of snakes) has been employed. In
mucin-iike j^g researches, carried on under the direction of Kuhne,
Dr Lauder Brunton1 followed the following process.
Defibrinated blood from the bird was treated with ten or twelve
times its volume of 3 per cent. NaCl solution, and the corpuscles
separated by nitration and decantation. On shaking the residual
mass of corpuscles with water and ether, the nuclei of the corpuscles
are set free from the stroma, and float at the junction between
the water and ether.

In order further to separate the nuclei from adhering stroma
and colouring matter, the agitation with ether and water may
be repeated several times and the residual matter washed with
dilute hydrochloric acid, hot alcohol and ether2. From his obser-
vations Brunton came to the conclusion which Kiihne had previously
arrived at, viz. that the nuclei of the blood corpuscles are composed
of a substance closely resembling, if not actually identical with
mucin. He found that they were insoluble in HC1 of 01 to 1
per cent., but soluble in solutions of the alkalies, the solutions
thus obtained being precipitated by the addition of mineral acids,
the precipitate being redissolved by an excess of acid. The solutions
were precipitated by acetic acid, the precipitate not being soluble in
excess, but were not precipitated by solution of mercuric chloride.

Nuclei of Plosz, repeating these experiments of Brunton, con-

blood cor- firms the statement that the body composing the nuclei

puscies said resembles mucin in its properties ; on subjecting it to
analysis, however, he found that it contained phos-
phorus, and he therefore considers it to be identical

1 Brunton, "On the chemical composition of the nuclei of Blood-corpuscles."
Journal of Anatomy and Physiology. Second series. Vol. in., p. 91.

* Plosz, "Ueber das chemische Verhalten der Kerne der Vogel- und Schlangen-
blutkorperchen," Hoppe-Seyler, Med. Chem. Untersuchungen, Heft iv. (1871) p. 460.

CHAP. II.] THE BLOOD. 83

with the body separated by Miescher1 from the nuclei of pus-corpuscles
and by him termed NUCLEIN. This body, which will be treated of
fully under ' pus,' is unacted upon by gastric juice, so that bodies
composed of it (e.g. the nuclei of the red blood corpuscles) may be
purified by subjecting them to artificial digestion.

The Nuclein of Miescher contains 9 '59 p. c. of P, and to it the
formula C^H49N9P3O22 has been ascribed. This formula must be
received with great caution.

Fatty matters containing Phosphorus (Lecithin, Protagon?).

Berzelius and Lehmann were aware that the corpuscles contained
a fatty body or bodies containing phosphorus, and the second of
these observers determined that the ash of the blood corpuscles
contains phosphoric acid and has an acid reaction. A closer study
of the phosphorized proximate principle of the coloured corpuscles
was, however, made by Gobley2, and afterwards by Hermann3 and
Hoppe-Seyler4.

Having dissolved the blood corpuscles in water, Hermann agitated
the solution repeatedly with ether. The ethereal solution was decanted
and evaporated, when it was found to leave a crystalline deposit
consisting of cholesterin and tufts of a body which Hermann considered
identical with the substance shortly before separated by Liebreich
from the brain and called by him PKOTAGON.

In order to purify this substance Hermann added water to the
mixed crystalline deposit left by the ether ; the effect of the water is
to cause the protagon to swell and to become less soluble in ether ;
by the latter reagent the substances soluble in ether are then
separated. The residue is dissolved in alcohol heated to 50°; and
from the alcoholic solution protagon is obtained in a crystalline form.

Of late years Hoppe-Seyler and, after him, nearly all physiological
chemists have come to consider protagon as not being a definite
proximate principle but as a mixture of a phosphorized body called
lecithin C^H^NPO^ with a body termed cerebrin C31H33NO3, and
it is the former substance which, according to Hoppe-Seyler, is con-
tained in the red blood corpuscles. These surmises in reference to the
non-existence of protagon have however been disproved by the author,
who has shewn that protagon is a perfectly definite proximate
principle. The observations of Hoppe-Seyler and Jiidell, however,
would appear to be irreconcilable with the view that the coloured
corpuscles contain protagon5.

1 Miescher, "Ueber die chemische Zusammensetzung der Eiterzellen." Hoppe-
Seyler, Med. Chem. Untersuchungen, Heft iv. (1871) p. 441.

2 Gobley, Journ. de Pharm. et de Chemie, Ser. in., T. xxi., p. 250.

3 Hermann, Archiv f. Anat. u. PhysioL, 1866, p. 33.

4 Hoppe-Seyler, " Ueber das Vorkommen von Cholesterin und Protagon und ihre
Betheiligung bei der Bildung des Stroma der rothen Blutkorperchen." Med. Chem.
Untersuchungen, Heft i. (1866) p. 140. Also Gustav Jttdell : "Zur Blutanalyse,"
Hoppe-Seyler's Med. Chem. Untersuchungen, Heft in. (1868) p. 386.

5 Gamgee and Blankenhorn, "On Protagon." Journal of Physiology, 1879.

6—2

84 CHOLESTERIN. OXY-HAEMOGLOBIN. [BOOK I.

According to Jiidell, who worked under Hoppe-Seyler's direction,
100 parts of the dried blood corpuscles of man contained (1) 0*35 and
(2) 072 of lecithin ; 100 parts of the dried corpuscles of the dog
contain 0*59, 100 parts of the dried corpuscles of the goose 0'46 of
lecithin.

Cholesterin.

This body, which will be treated of fully under ' nervous tissue,'
is an invariable constituent of the red blood corpuscles and can be
separated from them by ether. For the method to be followed the
reader is referred to the Appendix. According to Jiidell1, 100
parts of the dried blood corpuscles of man contain 0'25 of cholesterin.
In the dried corpuscles of the goose the cholesterin attained the amount
of 0*48 per cent.

It was formerly supposed that the neutral fats were contained in
the coloured corpuscles. Hoppe-Seyler2 has however found that such
is not the case.

OXY-HAEMOGLOBIN.

For a long time the opinions of chemists and physiologists were
divided as to the nature of the colouring matter of the red blood cor-
puscles, and for the most part this was supposed to consist of a body
termed HAEMATIN, which, as we now know, is but a product of
decomposition of the true blood-colouring matter — HAEMOGLOBIN, or
as we now term it when loosely combined, as it always is in the
blood, with a certain quantity of oxygen, Oxy -Haemoglobin.

Crystals of a beautiful red colour had under certain circumstances
been observed to separate from the blood of different animals by
Leidig3, Reichert4, and Kolliker5, and had been afterwards more care-
fully described by Funke, Kunde, and Lehmann.

The researches of several observers, but especially those of Hoppe-
Seyler, soon proved that the blood crystals are in reality crystals of
the true blood-colouring matter, which forms the chief part of the solid
constituents of the red corpuscles, and methods were soon found for
obtaining them in large quantities and in a very pure condition.
Thanks to these and to the application of varied methods of physical
and chemical research, we now have more definite knowledge as to the
part played by the blood-colouring matter in the processes of the
economy than we possess with regard to any other of the proximate
principles of its tissues and organs.

1 Jiidell, loc. cit.

2 Hoppe-Seyler, Handbuch d. physiologisch- u. pathologisch-chemischen Analyse.
Dritte Auflage (1870), p. 318, note.

3 Leidig, Zeitschrift fur wiss. Zoologie. Bd. i. (1849) p. 116.

4 Eeichert, Miiller's Archiv (1849), p. 197.

5 Kolliker, Zeitschrift fur u-iss. Zoologie. Bd. i. (1849) p. 216.

CHAP. II.] THE BLOOD. 85

Methods of It must be stated in limine that great difference

preparation exists in the ease with which this body can be obtained

mogiobm6" in an odiously Pure condition from the blood of

different animals. By obviously pure condition we

mean to indicate in the form of well-defined crystals.

The principle upon which nearly all methods of separating oxy-
haemoglobin is based is the following : to effect the solution of the
haemoglobin of the red corpuscles either in the serum or in water
added to the separated corpuscles, and then either by the addition of
alcohol or by the agency of cold, or of both conjointly, to cause the
oxy-haemoglobin (which is sparingly soluble in dilute alcohol and at
low temperatures) to crystallize out.

From the blood of some animals, and especially of the rat, oxy-
haemoglobin can be obtained for microscopic examination in two or
three minutes by receiving a drop of blood on a glass slide, adding to
it a drop of distilled water, and after mixing the two together cover-
ing with a microscopic covering-glass. Needle-shaped crystals form
almost at once. In order to separate considerable quantities of
oxy-haemoglobin or even to obtain large crystals for microscopic
observation it is advisable to follow one or other of the following
methods, of which the fifth and seventh are those which are most
easily carried out and most uniformly successful1.

I. Blood is allowed to coagulate and the clot is allowed to contract so
as to separate the serum as completely as possible. (This end is naturally
most readily attained by employing a centrifugal apparatus.) The clot is
finely divided and then squeezed in a cloth; in this way the corpuscles
are separated from the fibrin of the clot.

Water is added to the expressed grumous liquid (cruor) in quantity
equal to one or one and a half times its volume. A stream of oxygen gas is
now passed through the liquid for half an hour, and then a stream of C02
for ten minutes. After about five minutes a turbidity appears, crystals
commence to form, a large quantity separating out in the course of two
hours. By this method crystals are obtained only from the blood of the
guinea-pig, the rat, and the mouse. In order to obtain them from
the blood of the dog and other animals, before and during the passage of the
gases, dilute alcohol is added in small quantities to the fluid, which then
often yields a magma of crystals. Crystals thus obtained are, however,
not pure, and in order to separate them from adhering impurities they
must be washed with distilled water, or water holding a little alcohol in
solution, until the nitrate is no longer precipitated by solutions of silver
nitrate or of mercuric chloride2.

Preyer has found that by merely passing air free from carbonic acid
through the defibrinated blood of the dog crystallization ensues, even
though the temperature of the blood be as high as 35° — 38° C.

1 The description of the first six methods of preparing oxy-haemoglobin is based
upon that given by Preyer (in his admirable work entitled Die Blutkrystalle, Jena, 1871)
as abridged in Maly's Jahresbericht, Vol. i. (1873) p. 57; the seventh the Author
learned from Professor Kiihne ; he can highly recommend it.

2 Lehmann, Ber. d. konigl. sachs. Ges. d. Wiss. zu Leipzig, 1853. Also Physio-
logical Chemistry. Translation by Day. Cavendish Society, 1854. Vol. in., p. 489
et seq.

8G METHODS OF PREPARING OXY-HAEMOGLOBIN. [BOOK I.

II. A platinum capsule is placed in a freezing mixture and then
freshly defibrinated blood is poured into it, so as to convert it into a red
lump of ice. After being in this freezing mixture for half an hour, the
blood is allowed to thaw gradually, and the contents of the basin are then
poured into a glass vessel of such dimensions that the bottom is covered by
the lake-coloured blood to a depth of 1 5 millimetres ; the glass vessel is
then set aside in a cool place. In a short time the blood of guinea-pigs and
of squirrels furnishes by this method well-formed crystals. According to
Rollett, cat's blood is next in the order of facility of crystallization. Then
follow dog's blood, human blood, and the blood of rabbits. The blood
of the pig and of the frog yield by this method no crystals, though their
oxy-haemoglobin is crystallizable. In order to obtain crystals from the
blood of these animals, the process of freezing and thawing must be fre-
quently repeated \

This method is, accord ing to Preyer, very convenient in winter, especially
when comparative crystallographic and optical investigations of the oxy-
haemoglobin of the blood of many different animals have to be carried on.

III. In this method, the stroma of the coloured corpuscles is dissolved
by the addition to the blood corpuscles of a watery solution of crystallized
bile (a mixture of sodium glycocholate and taurocholate).

A. The blood of the horse is collected in a cylinder and at once
cooled. As soon as the plasma and subjacent stratum of colourless cor-
puscles have separated, these are separated from the red corpuscles, and the
mass of residual red corpuscles is treated with a 0*5 per cent, watery
solution of crystallized bile. Then the mixture is allowed to coagulate.
The fibrin as it separates encloses the yet undissolved corpuscles, so that the
surrounding deeply lake-coloured fluid is entirely free from them. To the
fluid, which is kept continually stirred, there is then added 90 p.c. alcohol
containing a trace of acetic acid, as long as the precipitate which is pro-
duced continues to redissolve. After some hours the fluid is converted into
a magma of crystals which are collected on a filter and washed, first with
diluted alcohol and then with iced water. Instead of this method we
may use :

B. 100 c.c. of dog's blood is allowed to coagulate in a shallow basin ; the
clot is then separated from the sides of the vessel and set aside for 24 hours.
(The centrifugal apparatus might be used with advantage.) The serum
is then decanted and the clot washed with water; it is then finely
divided and diffused by the help of a syringe through 50 c.c. of water, and
after standing for 24 hours is filtered through linen and the residual fibrin
washed with 10 c.c. of water. The mixture thus obtained of diluted serum
and blood corpuscles is treated with 2 c.c. of a syrupy solution made by
dissolving 1 part of crystallized bile in 3 parts of water ; after 24 hours
every blood corpuscle has disappeared. Nevertheless the fluid filters
very slowly. On adding 20 c.c. of 90 p.c. alcohol for every 100 c.c. of the
nitrate, the latter is converted into a magma of crystals which are washed
first in dilute alcohol and then in iced water.

1 Rollett, "Versuche und Beobachtungen am Blute." Sitzungsber. d. math,
naturw. Classe der kaiser. Akad. d. Wissenschaft. Vol. XLVI. (1863). Abth. n., p. 77.

CHAP. II.] THE BLOOD. 87

These methods are not to be recommended.

IV. The defibrinated blood of the dog is mixed with its own volume
of distilled water and the diluted fluid is treated with one fourth of its
volume of alcohol. The mixture is kept for 24 hours at a temperature of
0°C. or below. The crystals which separate are dissolved in as small a
quantity as possible of water at 25° to 30° C., and the solution being cooled
to 0°0. a fourth of its volume of alcohol is added.

It is better to place the fluid in a freezing mixture at a temperature
of- 10° to - 20° for 24 hours. The whole fluid then becomes filled with a
magma of crystals. The process of recrystallization may be several times
repeated.

V. Blood is collected in a capsule and, having coagulated, is allowed to
stand undisturbed for some hours or, better still, for a whole day. The
serum is then decanted, the clot washed with water and cut into small
pieces, and these also are repeatedly washed with distilled water. When the
washings are no longer strongly precipitated by solution of mercuric chloride,
the pieces of clot are treated with water heated to 30° — 40° 0., and the fluid
is filtered, the filtrate being collected in a cylinder surrounded by ice. A
known fraction of the red solution is then treated little by little with alcohol
(poured out of a burette), the fluid being continually stirred, until a slight
precipitate is formed. In this way is ascertained how much alcohol may be
added without a precipitate resulting. Having thus found out how much
alcohol would have to be added to the whole quantity of filtrate, a some-
what smaller quantity is actually added, and the fluid is placed in a freezing
mixture. After some hours crystals separate abundantly. As much water
has been employed in the process, the crystals can easily be filtered.
These are washed, first with water holding alcohol in solution and after-
wards with iced water. The crystals thus obtained may either be at once
used or be purified by further crystallization. At a temperature below
0° C. they can be dried in the air without decomposition.

VI. Defibrinated blood is mixed with ten times its volume of a
solution of sodium chloride (made by diluting 1 volume of saturated
solution of NaCl with from 9 — 19 volumes of water), and allowed
to stand for one or two days in a cool place so as to allow of the
greater part of the blood corpuscles to settle. The supernatant liquid
is decanted and the corpuscles are placed with a little water in a
flask and then ether is added. After repeated agitation, the ether is
decanted, and the fluid is filtered through a plaited filter as rapidly
as possible. The filtrate is cooled to 0° C. and treated with £ its volume of
spirits of wine ; the mixture is then maintained for some days at — 5° or
- 10° C. The crystals which separate may be purified by recrystallization1.

VII. 500 c.c. defibrinated dog's blood are treated with 31 c.c. of ether
and the mixture shaken for some minutes. It is then set aside in a
cool place. After a period varying from 24 hours to 3 days the
liquid has become converted into a thick magma of crystals. These
may be separated by placing in tubes and using the centrifugal apparatus.

1 Hoppe-Seyler, Handbuch der physiologisch- und pathologiscTi-chemisclwn Analyse.
3* Aufl., 1870, p. 215.

88

ELEMENTARY COMPOSITION OF OXY-HAEMOGLOBIN. [BOOK I.

The cakes of crystals thus obtained are mixed with water holding
one-fourth of its volume of alcohol in solution and again centrifugalized.
By repeating this process the crystals are obtained free from serum-
albumin. If requisite the crystals are dissolved in water and recrystallized
by the method mentioned under IV.

VIII. In order to obtain very large crystals of oxy-haemoglobin for
microscopic examination, Gscheidlen1 seals defibrinated dog's blood which
has stood in the air for 24 hours in narrow glass tubes (vaccine tubes
answer well), and keeps the tubes for some days at a temperature of
37°C. On opening these tubes and emptying their contents into a
watch-glass, and allowing some time for evaporation to take place, there
are formed crystals of extraordinary size.

Elementary The analyses of Carl Schmidt and Hoppe-Seyler

composition have shewn that crystallized oxy-haemoglobin is a body
of oxy-hae- Of perfectly constant composition. Unlike any other of
the proximate constituents of the animal body it con-
tains the element iron.

The following table exhibits the mean results of the analyses of
Hoppe-Seyler of oxy-haemoglobin from various animals and from the
horse. The former were published in 18682, the latter in 1878s;
the latter are so different from the former as to be not above
suspicion, especially as they were not actually obtained by Professor
Hoppe-Seyler but by an assistant.

PEK-CENTAGE COMPOSITION OF CRYSTALLIZED OXY-HAEMOGLOBIN
DRIED AT 100° C.

Water of

Crystal-

C

H

N

0

s

F

lization.

Crystals from dog's blood

3—4

53-85

7-32

16-17

21-84

0-39

0

„ goose's blood

7

54-26

7-10

16-21

20-69

0-54

0

„ guinea-pig's blood

6

54-12

7-36

16-78

20-68

058

0

„ squirrel's blood

9

54-09

7-39

16-09

21-44

0-40

0

„ horse's blood

54-87

6-97

17-31

19-73

065

0

From the analyses of Hoppe-Seyler (excluding that of horse's
blood, which is more recent) and of C. Schmidt, Preyer deduced the
following as the mean per-centage composition of oxy-haemoglobin :

1 Gscheidlen, "Darstellung von Hamoglobin Krystallen zu mikroscopischer Beo-
bachtung." Physiologische Methodik, p. 361.

2 Hoppe-Seyler, "Beitrage zur Kenntniss des Blutes des Menschen und der
Wirbelthiere." Med. Chem. Untersuchungen, Heft in. (1868) p. 370.

3 Hoppe-Seyler, " Weitere Mittheilungen iiber die Eigenschaften des Blutfarbstoffs."
Zeitschriftf. phys. Chemie, Vol. n., p. 150.

CHAP. II.] THE BLOOD. 89

In 100 parts.

C 5400

H 7-25

N 16-25

Fe 0-42

S 0-63

O 2145

100-00

and assuming that the molecule contains one atom of iron the follow-
ing would be the empirical formula :

C«H960N154FeS30179.

Crystalline
form.

Oxy-haemoglobin obtained from the blood of man
and the majority of the lower animals, crystallizes in
prisms or rhombic plates of a beautiful blood-red colour, which belong
to the rhombic system ; the oxy-haemoglobin of the squirrel crystal-
lizes in six-sided plates which belong to the hexagonal system. The

FIG. 15. CRYSTALS OF OXY-HAEMOGLOBIN.

a, b, c and e illustrate the forms in which haemoglobin separates from the blood of
man and the majority of mammals, d are tetrahedral crystals from the blood of the
guinea-pig. / are hexagonal crystals from squirrel's blood.

90 PROPERTIES OF OXY-HAEMOGLOB1N. [BOOK I.

oxy-haemoglobin of the- guinea-pig crystallizes in the form of tetra-
hedra or of tetrahedra with truncated edges and angles, which were
at one time supposed to belong to the regular system ; they have been
proved by Lang1 to belong to the rhombic system.

Crystals of oxy-haemoglobin, of whatever form, are doubly re-
fracting and pleochromatic ; when examined in polarized light the
crystals, according to the position of their axes in reference to the
observer, appear of a dark reddish-blue or of a bright scarlet
colour.

Certain Oxy-haemoglobin, as obtained by any of the pro-

chemical cesses above described, presents when moist the appear-

reactions of ance of a pasty mass of a cinnabar-red colour ; it may be
oxy-haemo- dried in vacuo over sulphuric acid at temperatures
giobin. below 0° C. without undergoing decomposition, and the

dried crystals thus obtained are found to be perfectly soluble in
water, yielding a solution which presents the optical properties after-
wards to be described. The crystals of oxy-haemoglobin dried in
vacuo still retain 3 or 4 per cent, of water of crystallization, which is
driven off by heating to 110° or 120° C. If the crystals of oxy-haemo-
globin have been thoroughly dried at a temperature below 0° C., the
dried substance may be heated to 100° without undergoing decom-
position; the slightest trace of moisture suffices, however, to effect
decomposition at much lower temperatures — a decomposition evidenced
by the assumption of a brown colour, by the substance becoming in-
completely soluble in water, and by a modification of the optical pro-
perties (formation of methaemoglobin).

Solutions of oxy-haemoglobin in distilled water if sealed in vessels
with no perceptible air-space may be kept for many months, or perhaps
years, without undergoing any further change than the reduction or
loss of oxygen to be afterwards referred to. The fact is one of
importance practically, as enabling standard solutions of haemoglobin
to be preserved almost indefinitely2.

Oxy-haemoglobin obtained from different animals differs in its
solubility. That obtained from the guinea-pig is comparatively little
soluble, whilst that of bullock's and pig's blood is very soluble.
Gautier gives the following order of solubility of the haemoglobin
obtained from several animals — cat, dog, horse, man : the degree of
solubility increasing according to the order named.

Haemoglobin is readily soluble without decomposition in very
weak solutions of the caustic alkalies or of the corresponding
carbonates; an excess of alkali, however, very readily induces de-
composition.

All acids and salts having an acid reaction decompose haemoglobin
with the formation of haematin.

1 Sitzungsber. d. Wiener Akad. Vol. XLVI., p. 85.

a Hoppe-Seyler, " Weitere Mittheilungen iiber die Eigenschaften des Blutfarbstoffs.
2. Ueber die Fahigkeit des Hamoglobins der Faulniss sowie der Einwirkung des
Pankreasferments zu widerstehen. " Zei tschrift f. physiol. Chem., p. 125, et seq.

CHAP. II.] THE BLOOD. 91

Potassium carbonate added to solutions of haemoglobin precipitates
the body without decomposing it, if the temperature be low.

Solutions of haemoglobin are not precipitated by solutions of lead
acetate even after the addition of ammonia, nor by silver nitrate,
though these reagents soon lead to its decomposition.

Alcohol precipitates haemoglobin, the precipitate having at first a
red colour, but soon changing to brown, indicating that decomposition
has taken place.

When heated to 70° or 80°, dilute solutions of oxy-haemoglobin
undergo, for some time, no decomposition ; soon however the liquid
becomes turbid and brown, in consequence of the decomposition of the
oxy-haemoglobin and the separation of insoluble products.

These reactions will however be studied with greater advantage after
a careful investigation of the optical properties of oxy-haemoglobin,
as revealed by an examination of the spectrum of light which has
traversed crystals of oxy-haemoglobin, solutions of the body, or which
has merely been passed through dilute blood.

We have used the term oxy-haemoglobin to denote the colouring
matter as it exists in the living blood or as it is obtained by the
processes we have described : viz. under circumstances in which it
exists in combination with a very small proportion of oxygen —
oxygen which is linked to it by ties so easily broken that it can be
transferred to other easily oxidizable bodies existing by its side, that
it can be given up when its solutions are gently heated in a Torricellian
vacuum, or are agitated at moderate temperatures with large quantities
of inactive gases such as nitrogen or hydrogen — oxygen which may
with appropriateness be spoken of as the respiratory oxygen of
haemoglobin.

Tke ab_ It has long been known that if homogeneous white

sorption light be passed through certain coloured gases, liquids or

spectrum of solids, and then through a prism, the spectrum instead
oxy-haemo- Of being continuous, is seen to be intersected by dark
lines or bands which are termed absorption bands, the
spectrum which manifests such bands being designated an absorption
spectrum. The situation of such absorption bands, being perfectly
constant, often affords a valuable means of identification and a
ready means of determining the occurrence and course of changes
in composition effected in the body which exhibits them.

The blood was shewn by Hoppe-Seyler to exhibit when white
light is passed through it a very characteristic absorption spectrum,
which he was able to shew is identical with the spectrum of pure
oxy-haemoglobin, supplying by this discovery the absolute proof that
the blood crystals which had by many observers been suspected to
be the pure colouring matter of the corpuscles, actually did consist
of that substance.

In examining the absorption spectra of blood or any other solution it is
convenient to dilute the liquid sufficiently and then to pour it into a glass
vessel with parallel faces, which are a definite width apart. Snch

92

METHODS OF OBSERVING ABSORPTION SPECTRA. [BOOK I.

vessels are made, after the plan of Hoppe-Seyler, for the purpose of
the physiological chemist, and sold under the name of Haematinometers1;
the glass plates are exactly one centimetre apart, so that when the
apparatus is filled with liquid, the observer knows that he is examining
a stratum 1 cm. broad. Instead of such a vessel the Haematoscope or
Haemoscope of Hermann2, shewn in the accompanying woodcut, may
be employed. F is a plate of glass, and the piston G is a metallic tube
closed at its inner end by a plate of glass. Ity sliding the piston C in and
out of the tube D the capacity of the vessel DFB and the depth of a
stratum of fluid contained between the two glass plates may be modified at
will within wide limits. The depth of the stratum is read off by means of
a millimetre scale engraved 011 the sliding tube (7.

FIG. 16. THE HAEMATINOMETER.

FIG. 17. HAEMATOSCOPE.

Whichever the exact form of vessel containing the blood to be
examined, it is interposed between a source of light and a suitable
spectroscope.

Various forms of spectroscope may be employed in these researches.
Any ordinary spectroscope adapted to the requirements of the chemiht
will answer ; it is advisable, however, that the instrument shall be
provided with an arrangement for observing simultaneously two spectra,
and with a scale.

1 These are sold by Schmidt and Haensch, Berlin.

2 Hermann, "Notizen fur Vorlesungs- und andere Versuche." Pfliiger's Archiv,
Vol. iv. (1871) p. 209.

CHAP. II.]

THE BLOOD.

lu the annexed drawing the arrangement of the whole apparatus is
shewn.

FIG. 18.

1. At A is a tube bearing at its distal end a slit which may be made
narrow or wide at will and which is provided with a reflecting prism by
means of which the spectrum of light from two sources may be simul-
taneously observed ; at its proximal end it is furnished with an achromatic
lens. 2. In the centre of the instrument is a flint glass prism which
receives the parallel rays which have passed through the slit and collimating
lens. 3. At B is a telescope into which penetrate the rays which have
been dispersed, by the prism. 4. At 0 is shewn a tube bearing at its
distal end a scale photographed upon glass and which is illuminated by a
lamp as shewn in the engraving.

In actually working with such an instrument the observer, having
thrown a dark cloth over the prism, commences by adjusting the lights so as
to illuminate the slit and the scale, and by adjusting the slit at the end of
tube A, and focusing the scale in tube C and the telescope B, he endeavours
to get a sharply defined spectrum, and immediately above or below it a well-
illuminated image of the scale.

In working with the spectroscope it is of great importance to be able
to fix more or less precisely the locality of any line or band which has been
observed, and in order to do so various plans may be followed. One most
commonly followed is to examine very carefully the spectrum of sunlight
and to determine the position of the principal Fraueuhofer lines in reference
to the scale of the instrument. The observations are tabulated, or a
map drawn shewing the position of these lines, which are to serve as land-

94 METHODS OF OBSERVING ABSORPTION SPECTRA. [BOOK I

marks for future observations. A very useful method of recording the
position of lines and bands in the spectrum, lately suggested by Dr
MacMunn1, is thus described in his own words :

"The slit of the spectroscope being illuminated by some light, it
is sufficiently narrowed, and the eye-piece focussed, till the Frauenhofer lines
are seen distinctly; a camera-lucida is then slipped over the eye-piece, and a
point marked — on a piece of paper placed beneath the camera — just beyond
the extreme red, and another beyond the extreme violet. A number of
blank spectrum maps are then made of this length, and again brought beneath
the camera ; the position of the Frauenhofer lines is marked on the top one,
and afterwards on all the others. In this way a number of solar
maps are made, from which any required number can afterwards be
copied.

" When an absorption spectrum has to be mapped, a test-tube containing
the solution, illuminated by means of a strong light, is placed before
the slit, the right-angled reflecting prism is made to cover half the slit, and
a Bunsen burner, with a salt of sodium introduced into its flame, is placed
so that its light shall fall upon the right-angled prism. On looking
into the instrument two spectra are seen, one the absorption spectrum,
the other the spectrum of sodium — a yellow line on a dark background.
The camera lucida is then slipped over the eye-piece, two maps with
the Frauenhofer lines marked on them brought beneath it, and the paper
shifted till the bright-yellow sodium line covers the D line on the
maps ; with a lead pencil the position of the bands and the amount of
shading is marked on the maps, care being taken to keep the paper
from slipping. It must be remembered that the maps have to be
turned upside down while being made, otherwise the A line would be on the
right-hand side and the H on the left in the solar maps, and the
absorption bands in the wrong place in the others."

Spectro- For some time past physicists have been in the habit

scopes with of recording the position of bright or dark lines observed in
the spectrum by stating the wave-length of the region
in which they occur. Usually the observations have been
made with instruments furnished with an arbitrary scale
only. Having determined the position of certain lines on the solar
spectrum (of which the wave-length is precisely known) in reference to
the arbitrary scale, data are obtained for constructing, by an easy geo-
metrical process, a curve which represents the relation of any point on the
arbitrary scale to a scale of wave-lengths. The observations which have
been made with the arbitrary scale are . then reduced to wave-lengths.
Though the reduction is somewhat troublesome the observer knows that,
when made, his observations have acquired a definiteness which they other-
wise would never have possessed. Usually wave-lengths are now express-
ed in 10-millionths of a millimetre, but other units of measurement may
be employed.

Recently Herr Carl Zeiss, the eminent optician of Jena, has, at the
suggestion of Professor Abbe, constructed spectroscopes provided with an
illuminated scale, which is divided and numbered so as to permit of the

1 MacMunn, Studies in Medical Spectroscopy. Keprinted from the Dublin Journal
of Med. Sc., June, 1877.

CHAP. II.]

THE BLOOD.

direct determination of the wave-length of any region in the visible
spectrum.

The scale is similar to that shewn below, except that the position of the

? *

C

1

J>

F

b

1

i

<

f *

I

i

i

5 70

65

€

0

5

5

5

0

4

5

4(

i

i

i

1

1

1

Fm. 19.

SCALE OP WAVE-LENGTHS, SIMILAB TO THAT IN ZEISS'S SPECTROSCOPES.

lines of Frauenhofer is not marked on the scale. The numbers attached to
the divisions on the scale indicate wave-lengths expressed in 100,000ths of
a millimetre; and each division indicates a difference in wave-length equal to
one hundred thousandth of a millimetre (0*00001 mm.). By the eye, the
position of any line situated between two divisions of the scale can be
estimated to one-tenth of a division, so that its wave-length can be ex-
pressed in millionths of a millimetre. In using Zeiss's instruments, the
observer commences by causing Frauenhofer's line D, or the sodium line,
to coincide exactly with that part of the scale which expresses its
wave-length, that is to say to correspond to division 58*9 of the scale
(which expresses a wave-length of 589 millionths of a millimetre or
0*000589). Having done this the scale is accurately set for all other
points.

Let us suppose that the observer wishes to determine the wave-
length of Frauenhofer's line E. He will at once see that the line is placed
between divisions 52 and 53 and he would determine its position between
two divisions to within one-tenth of a division, but probably much nearer.
The reading would probably be 52*7, which would give for the wave-
length of E 527 millionths of a millimetre, a result which is only
three ten-millionths below the value deduced from the observations
of Angstrom1. As a result of many experiments with one of Zeiss's
spectroscopes the Author has found that the mean error in his case is not
greater than ±0*000001 mm.

Printed blank maps accompany Zeiss's instruments, which correspond
exactly to the scale of the spectroscope. There is therefore not the slightest

1 The following are the wave-lengths corresponding to Franenhofer's lines A, B, C,
D, E, F, G-, according to the most recent measurements, expressed in millionths of a
letre :

A 760-4
B 687*4
C 656-7
D 589-4
E 527*3
F 486-5
G 431-0
H 396-8

96

METHODS OF OBSERVING ABSORPTION SPECTRA. [BOOK I.

difficulty in drawing up a map which shall represent the relative and
absolute position of any lines or bands observed in a given spectrum.

The absorption bands which form the characteristic features in the
spectra of blood and certain other animal liquids do not admit of having
their limits determined with the same sharpness and precision as is possible
in the case of the bright lines in the spectra of incandescent metals or in
that of the lines of Frauenhofer in the solar spectrum. It would therefore
be mere pedantry to express their position or extent on a wave-length scale
to one ten-millionth of a millimetre. In this work all drawings of
spectra will be accompanied by a scale of wave-lengths, and the position
and extent of bands usually expressed in millionths of a millimetre.

Micro- spec- Where very small quantities of a solution are to be

troscopes. examined these may be introduced into small cells made by

cementing sections of barometer tubing of various lengths and diameters
to glass slides. Such a cell may be made which only requires two or
three drops of fluid in order to fill it. Instead of employing an ordinary
spectroscope we may in this case with advantage employ some form of
micro-spectroscope.

FIG. 20.

ZEISS'S MJCRO-SPECTROSCOPE.

The instrument consists of a drum A (Fig. 20) interposed between the field-lens and
eye-lens of an eye-piece. Within the drum there is a slit which by means of screws
H and F (Fig. 21) can be lengthened or shortened and made wider or narrower ;
it also contains a prism wherehy light coming from an aperture in a stage at the
side of the drum is totally reflected in the direction of the optic-axis of the eye-
piece. Over the eye-lens of the eye-piece is situated the combination of prisms
with the measuring apparatus ; this, which is the spectroscope proper, revolves
around the eccentric K (Fig. 20) : it can either be moved away from the eye-lens or
brought over it, and retained there by the catch L. At N is placed the scale of
wave-lengths (see Fig. 20), which is illuminated by the mirror 0. The screw P
and the spring Q are employed to alter the relation of the scale to the spectrum.
The former is always set by the observer so that Frauenhofer 's line D corresponds
to division 58-9.

CHAP. II.] THE BLOOD. 97

The first to apply a spectroscope to the microscope was Mr Sorby1, and
very numerous modifications of his original micro-spectroscopes have been
made. In all cases the micro-spectroscope consists of a modified microscopic
eye-piece which has superadded to it a direct- vision prism, an arrangement
of slits for allowing definite quantities of light to reach the prism, usually
arrangements for comparing two different spectra, and finally some micro-
metric arrangement. In consequence of the admirable nature of the micro-
metric arrangement we give the preference to the instrument made by Zeiss
and of which a vertical and horizontal section are given in Figs. 20 and 21.

Being provided, then, with one or other of the spectroscopes
piously described, or with a similar instrument, let the observer

previous

interpose between it and some source of light a solution of blood, say
made by diluting defibrinated blood with ten times its volume of
distilled water contained in a haematinometer (Fig. 16, p. 92) 1 cen-
timetre wide. It will then be found that the whole of the more
refrangible portion of the spectrum has been cut off but that the red
end of the spectrum remains visible, or rather, those rays having a wave-
length greater than about 600 millionths of a millimetre.

If now a stream of hydrogen or nitrogen be passed for a consider-
able time through the diluted blood it will be observed that the absorp-
tion is least between Frauenhofer's line a (W. L. 718) and Frauenhofer's
line B (W.L. 6867), but that the rest of the spectrum is less bright than
before the gas was passed. The effect of the N or H has been to drive
more or less of the respiratory oxygen from the haemoglobin, and in
consequence there is more light absorbed ; this difference in the spec-
trum corresponds to the change which the blood undergoes from a
bright vermilion colour to a brown-red when it passes from the arterial
to the venous condition, in other words from a condition in which its
haemoglobin is nearly saturated with its respiratory oxygen, to one in
which a portion of that oxygen has been given up.

If now the blood solution be rendered much more dilute so as to
contain -8 p.c. of haemoglobin, on examining a stratum 1 centi-
metre wide the spectrum becomes distinct up to Frauenhofer's line D
(W.L. 589), i.e. the red, orange and yellow are seen, and in addition
also a portion of the green between b and F. Immediately beyond D
and between it and b, however (between W. L. 595 and 518), the ab-
sorption is intense. (See Fig. 22, 4.) On diluting still further, what
appeared one wide black band between D and E is seen to resolve
itself into two beautifully distinct absorption bands separated by a
green interspace (Fig. 22, 3). Of these absorption bands, the
one next to D is narrower than its fellow; it has more sharply
denned edges and is undoubtedly blacker; its centre corresponds
with wave-length 579, and it may conveniently be distinguished as
the absorption band a in the spectrum of oxy-haemoglobin. The
second of these absorption bands, i.e. the one next to E, which we
shall designate /3, is broader, has less sharply defined edges, and is
not so dark as a. Its centre corresponds approximately to W. L. 553'8.
1 Sorby, Quarterly Journal of Science, 1865, xi. p. 198.

98

ABSORPTION SPECTRA OF OXY-HAEMOGLOBIN, &C. [BOOK I.

CHAP. II.] THE BLOOD. 99

FIGURE 22.

Spectrum 1. (Preyer, Plate 1, sp. 2 modified.} Solution of oxy-haemo-
globin containing less than O'Ol p. c. In this as in every other case, a
stratum 1 centimetre thick was examined. One distinct, though faint,
absorption band (a) between W. L. 583 and 575'1. According to Preyer1 there
is no trace of the band ft seen ; the Author finds, however, that whenever a
is visible he perceives a faint shadow in the position of wave-lengths 538 —
550. There is no absorption at either violet or red end of the spectrum.

Spectrum 2. (Preyer, PI. 1, sp. 4.) The solution contains 0'09 p. c. of
oxy-haemoglobin, a extends from 583 — 571 and ft from 550 — 532. The
violet end is absorbed to about 428. The red end is scarcely affected.

Spectrum 3. (Preyer, PI. 1, sp. 6.) The solution contains 0-37 p. c.
of oxy-haemoglobin, a extends from 589 — 567, and ft from 553 — 523.
The red end of the spectrum is perceptibly shortened. The violet is entirely,
and the blue partly, absorbed, to about 453.

/Spectrum 4. (Preyer, PI. 1, sp. 8.) The solution contains 0-8 p. c. of
oxy-haemoglobin. The two absorption bands have amalgamated and one
broad band is seen extending from 595 to 518. The green is just visible
between 518 and 498 ; the slightest increase in the strength of the solution
causes the green to disappear.

Spectrum 5. (Preyer, PI. 1, sp. 9.) Solution of haemoglobin (Syn.
reduced haemoglobin) of about 0'2 p. c. A single broad band with diffuse
edges, between 595 and 538; the band is darkest at about 550. Both
ends of the spectrum are more absorbed than by a solution of oxy-haemo-
globin having the same degree of concentration.

Spectrum 6. (Preyer, PJ. 1, sp. 14.) Solution of carbonic oxide hae-
moglobin. Two absorption bands very similar to those of oxy-haeino-
globin, but moved somewhat nearer to E. a extends from 587 to 564 and
ft from 547 to 529. The blue and violet are less strongly absorbed than
by a solution of oxy-haemoglobin of the same strength.

On diluting very largely with water nearly the whole of the
spectrum appears beautifully clear except where the two absorption
bands are situated. If dilution be pursued far enough even these
disappear; before they disappear they look like faint shadows
obscuring the limited part of the spectrum which they occupy. The
last to disappear is the band a.

The two absorption bands are seen most distinctly when a stratum
1 cm. thick of a solution containing 1 part of haemoglobin in
1000 is examined; they are still perceptible when the solution
contains only 1 part of haemoglobin in 10000 of water.

Haemogio- The spectrum of oxy-haemoglobin had been de-

scribed bJ Hoppe-Seyler when Professor Stokes made
the remarkable discovery that when diluted blood is
treated with certain reducing agents the colour of the liquid and
its spectrum undergo remarkable changes ; the former loses its bright
red and acquires a brown colour, whilst the green interspace which

1 Die Blutkrystalle : Untersuchungen von W. Preyer; mit drei farbigen Tafeln,
Jena, 1871.

7—2

100 HAEMOGLOBIN OR REDUCED HAEMOGLOBIN. [BOOK I.

had existed between the absorption bands a and $ of oxy-haemo-
globin disappears, and instead of the two bands there appears a
single one, less deeply shaded and with less finely denned edges,
extending between D and E. This band we may -distinguish as
absorption band 7. (See Fig. 22, Spect. 5 for description.)

Hoppe-Seyler1, who has described the spectrum of reduced haemo-
globin with great care, remarks that when a strong solution of that body
is diluted with water, avoiding at the same time the access of oxygen,
before the distinct absorption band which we designate 7 comes into
distinctness there is seen some green light between b and F. As the
solution is diluted, the dark absorption band, which extends at first
from D to b, diminishes in width, and the blue rays of the spectrum
become more distinctly visible.

On further diluting, the single absorption band is observed not to
exhibit any trace of subdivision, but to dimmish rapidly in intensity,
so that in a solution of such concentration that both absorption
bands of oxy-haemoglobin would be quite distinct, the single band 7 of
reduced haemoglobin has disappeared from view. Further, reduced
haemoglobin existing in solution is distinguished from oxy-haemo-
globin by its stronger absorption of the light between C and D, as
well as by its weaker absorption of the blue light above F.

If now the solution which presents this spectrum be shaken
with air or oxygen, the single band at once gives place to the two
original bands, whilst the liquid loses its brown and reacquires more or
less of its florid red colour. The process of reduction and subsequent
oxygenation may be repeated many times in succession.

From his experiments Stokes concluded that "the colouring matter
of bloody like indigo, is capable of existing in two states of oxidation,
distinguishable by a difference of colour and a fundamental difference
in the action on the spectrum. It may be made to pass from the more
to the less oxidized state by the action of suitable reducing agents, and
recovers its oxygen by absorption from the air*." This surmise has
been proved to have been perfectly correct, and to blood- colouring
matter after it has lost the oxygen which it can give up to reducing
agents, the name of reduced haemoglobin is given. By many, as by
Hoppe-Seyler, it is termed simply haemoglobin, to distinguish it from
the body as it exists combined with its respiratory oxygen and which
is then termed oxy-haemoglobin.

Methods Before proceeding further, it is advisable to consider

of reducing jlow faQ blood or a solution of ^xy-haemoglobin may be

gio^ixrto'bae- reduced so as to exhibit the spectrum of haemoglobin,
moglobin. The following are the methods which may be followed :

1 Hoppe-Seyler, "Beitrage zur Kenntniss des Blutes des Menschen und der
Wirbelthiere. Das reducirte Hamoglobin oder der venose Blutfarbstoff." Med. Ghem.
Untersuchungcn, Heft in. (1868) at p. 374 et seq.

2 Stokes, "On the reduction and oxidation of the colouring matter of the blood,"
by Professor Stokes, F.E.S. Proceedings of the Royal Society of London, Vol. xm.
(1864) p. 357, paragraph 8. Also Philosophical Magazine, 1864, p. 391.

CHAP. II.] THE BLOOD. 101

1. To a solution of a ferrous salt, as for instance of Fe SO4 + 7H2O, a
small quantity of tartaric or citric acid is added, and then ammonia until
the reaction is alkaline. In consequence of the presence of the vegetable
acid, ammonia does not throw down a precipitate of ferrous hydrate, but
a clear light-green solution is obtained which readily darkens by absorption
of oxygen from the air. Such a solution when freshly prepared exerts a
powerfully reducing action upon oxy-haemoglobin. When added in small
quantities to a solution of this body or to blood, the colour and spectrum
change almost instantly, to be restored again on agitation with air. Often
we may observe that on shaking up the reduced solution with air the
spectrum of oxy-haemoglobin is restored, though on leaving the solution a
moment or two at rest the two bands again disappear, and the single band
of reduced haemoglobin appears, proving that when existing side by side
with a ferrous salt, reduced haemoglobin can more readily take possession
of oxygen than that substance, to which however it afterwards cedes it.

2. Instead of ferrous sulphate we may employ a solution of a stannous
salt prepared after the same fashion, by mixing a solution of stannous
chloride, Sn C12, with tartaric acid and then adding ammonia to neutralization.
In this case, as in 1, by rendering the liquid alkaline we prevent it pro-
foundly decomposing the blood-colouring matter, whilst its oxidizing power is
increased. The tin presents the advantage over the iron solution of not
becoming deeply coloured as it absorbs oxygen, and therefore not absorbing
light passed through it.

3. The blood or solution of oxy-haemoglobin is boiled at a temperature
of 40° C., in a vessel in which a Torricellian vacuum is established by means
of a mercurial pump. Very shortly the colour of the liquid and the
change in spectrum evidence the complete removal of oxygen.

4. The blood or solution of haemoglobin is subjected for a long-con-
tinued period to the action of a stream, of washed hydrogen or nitrogen.
The same apparatus may be employed for this experiment as is used in
preparing Haemochromogen.

Whilst oxy-haemoglobin or its solutions very rapidly undergo
change at temperatures above 0°C. this is not the case with reduced
haemoglobin. Hoppe-Seyler has discovered that when a solution of
pure oxy-haemoglobin is sealed up in a glass tube (care being taken to
include very little air) after undergoing reduction, as exhibited by its
change of colour and spectrum, it suffers no further change and may
be kept for years. When such a solution is brought in contact with
oxygen oxy-haemoglobin is again formed and may even be crystal-
lized. This discovery of Hoppe-Seyler's is of great practical importance
to the physiological chemist, as it enables him to prepare standard
solutions of oxy-haemoglobin, when temperature and other circum-
stances are favourable, and to keep them indefinitely for subsequent use.

Hoppe-Seyler has also shewn that reduced haemoglobin resists tbe
action of pancreatic ferment1.

' l Hoppe-Seyler, "Ueber die Fahigkeit des Hamoglobins der Faulniss sowie der
Einwirkung des Pankreasferments zu widerstreben." Zeitschrift f. phys. Chemie,
Vol. i. p. 125.

102 COMBINATION OF O2 WITH HAEMOGLOBIN. [BOOK I.

The facts which have been narrated above supply the chief
materials for forming an opinion in reference to the nature of the
compound of haemoglobin with oxygen. From them it would appear
that this compound is of so remarkable a nature that it may be
formed with exceptional facility by the mere contact with atmospheric
oxygen, and that it is one which readily undergoes dissociation — the
decomposition being one in which the molecule of haemoglobin is left
intact and ready to combine again with fresh molecules of oxygen.

What is the quantity of oxygen which reduced
of the'reap*- haemoglobin can link to it as respiratory oxygen ?
ratory or Preyer1 as a result of three determinations found

loosely com- that 1 gramme of haemoglobin can link to itself 1'27
Dined oxygen cu^ cents, of oxygen measured at 0° C. and 1 metre
mog^obin6' pressure (or 1*671 c. c. measured, as is more usual in
England and France, at 0° C. and 760 mm. pressure),
and more recently Hufner2 has determined the amount again by a
different method and has obtained a result almost identical with that of
Preyer. According to Hufner and as the mean of ten separate deter-
minations, 1 gramme of haemoglobin fully saturated with oxygen is
associated with T28 c.c. of oxygen gas (measured at 0°C. and 1
metre pressure.)

Dissocla- Oxy-haemoglobin is one of those compounds which at

tion-tension particular temperatures and pressures undergo dissociation.
of the respi- At 40° C. the dissociation-tension is equal to about 30 mm.
ratory oxygen of mercury3. The Author attempted some time ago to ascer-
of oxy-hae- tain the dissociation-tensions of oxy-haemoglobin for various
moglobin. temperatures, but the results which he obtained were not

sufficiently accordant to allow of conclusions being drawn from them. The
subject will be discussed again under ' Respiration.'

Before leaving this division of our subject we have
oxy^aemo- *° re^er to a reaction which is possessed by oxy-
giobin upon haemoglobin and by some of its derivatives, though not
the resin of by reduced haemoglobin, and to which at one time con-
Guaiacum. siderable theoretical importance was attached, and which
still is of great practical value inasmuch as it affords us the most
delicate, though by itself not a conclusive, test for detecting exceedingly
minute quantities of these bodies.

It was found by A. Schmidt that when diluted blood is dropped
upon a filter paper which has been moistened with tincture of
guaiacum and then dried spontaneously in the air, a blue ring forms
at the edge of the drop ; it is best in this experiment to use blood
diluted with 20 times its volume of water, and it may be well to
remember that the reaction is one which is not produced by all

1 Preyer, Die Blutkrystalle : Untersuchungen von W. P., Jena, 1871, p. 134.

2 Hufner, " Ueber die Quantitat Sauerstoff welche 1 Gramm Hamoglobin zu binden
vermag." Zeitschriftf. physiologischen Chem. Vol. i. p. 317.

3 Worm Miiller, "Ueber die Spannung des Sauerstoffs derBlutscheiben." Ludwig's
Arbeiten, 1870, p. 119.

CHAP. II.] THE BLOOD. 103

specimens of tincture of guaiacum. This blueing of the resin of
guaiacum is due to its oxidation and is also observed when ozone acts
upon it, but not when common oxygen does so.

When the respiratory oxygen of haemoglobin has been expelled
from blood by the action of carbonic oxide, as will be afterwards
described at length, it no longer possesses (in the absence of oxygen) the
power of blueing guaiacum. If atmospheric oxygen, however, comes
in contact with the drop of CO blood and guaiacum, the blue ring
appears.

Oxy-haemoglobin shares with many other organic bodies and also
with many inorganic bodies, such as spongy platinum, the power of de-
composing hydrogen peroxide, H2O2, as is proved by the effervescence
produced in a solution of the latter by the addition of a few drops
of blood or of a solution of haemoglobin ; if to a mixture of blood and
tincture of guaiacum some solution of H2O2 be added, the fluid
assumes a blueish tint.

Does the These facts were formerly explained by A. Schmidt

oxygen of on *he hypothesis that haemoglobin possesses in an

oxy-haemo- intense degree the power of ozonizing oxygen and of
giobin possess rendering it therefore infinitely more active than atmo-
spheric oxygen. Against this view Pfliiger1 has raised
the most serious, and it appears to us the most reason-
able objections, which will have to be considered in detail in another
section. According to Pfliiger when blood is poured upon filter paper,
as in the guaiacum experiment previously referred to, the haemoglobin
almost instantly undergoes decomposition, and it is the products of
decomposition which occasion the reaction. According to Pfliiger
haemoglobin in no way modifies the properties of the oxygen which
it links to itself.

Proportion In former times, when blood-letting was highly

ofhaemogio- prevalent, a large number of analyses of blood were
bin in the made by competent observers who had no means, such

blood of man. ag we now pOSSesSj of determining directly the amount
of haemoglobin, but who ascertained the amount of iron contained in
the blood. Since we now know the exact proportion of iron which
haemoglobin contains, we may calculate the amount of this substance
found by the older observers. Preyer2 has taken the trouble to do
this in the case of a large number of the most reliable analyses, and
from his tables we take the following extract: —

QUANTITY OF IEON AND HAEMOGLOBIN COEEESPONDING TO IT
CONTAINED IN 100 GEMS. OF BLOOD.

A. Blood of woman (in health).

Iron. Hamoglobin.

Minimum . . . OO48 gnu. ll'57grm.

Maximum . . . 0-057 „ 13'69 „

1 E. Pfliiger, " Kritik iiber die Angaben vom Ozon im Thierkorper/' Pfluger's
Archiv, Vol. x. p. 252.

3 Preyer, Die Blutkrystalle, p. 117, et seq.

104 AMOUNT OF HAEMOGLOBIN IN BLOOD CORPUSCLES. [BOOK 1.

B. Blood of man (in health).

Iron. Hamoglobin.

Minimum . . . 0'0508 grm. 12-09 grm.
Average of 11 cases . 0056 „ 1345 „

Maximum . . 0-063 „ 15*07 „

The variations which the amount of haemoglobin undergoes in disease will be
considered in a future chapter.

By employing methods which will be subsequently

Relation of described, it is possible to determine with comparative
haemoglobin ,. ,, , , . ,

to the number readiness not only the number ot corpuscles contained

of the blood in a certain volume of blood, but also the amount of
corpuscles. haemoglobin, and the relation between the weight
of haemoglobin and the number of the blood corpuscles. Thus
Malassez found the number of red corpuscles in a cubic millimetre of
the blood of healthy men to vary between 4,000,000 and 4,600,000,
and the amount of haemoglobin between 0'125 and 0134 of a
milligramme1. Malassez has actually expressed the mean amount of
haemoglobin in each blood corpuscle of man in billionths of a
gramme (the billionth of a gramme he represents by the letters
yu-yLtgr.); his estimate is that each corpuscle contains on an average
30

By /x cub. Malassez2 designates the 1000th part of a cubic millimetre ; he
takes this as the unit of cubic capacity of the matter of red blood corpuscles,
and expresses the amount of haemoglobin in billionths of a gramme (/x/xgr.)
contained in one /x cub. of corpuscles of various animals, as is shewn below —

Volume of each corpuscle Haemoglobin contained

according to Welcker. in one /* cub. of corpuscles.

Man . . . 72 /x cub. 0'416/x/xgr.

Dove . .125 „ 0-416 „

Lacerta agilis .201 „ 0-348 „

Rana fusca . . 629 „ 0-343 „

Proteus . . . 9200 „ 0-115 „

These numbers must, however, be received with the greatest caution,
and as being very crude approximations to the truth, as will be obvious
when we consider that the number of corpuscles found in the healthy blood
of man by Malassez differs very notably from that found by other equally
competent observers, whose methods were probably more accurate.

Action of certain gases which displace the Oxygen of
Oxy-haemoglobin.

Carbonic It had been observed by Claude Bernard that

oxide, CO. foQ blood of animals poisoned with carbonic oxide

uniformly becomes of an intensely florid arterial hue, arid that this
differs from the normal colour of arterial blood by its persistence.

1 L. Malassez, " Sur les di verses methodes de dosage de 1'hemoglobine et BUT un
nouveau colorimetre." Archives de Physiologie, 2 ser. vol. iv.

8 Malassez, "Sur la richesse en hemoglobine des globules rouges du sang."
Gaz. med. de Paris, p. 534.

CHAP. IL] THE BLOOD. 105

He demonstrated, further, that when blood is shaken up with carbonic
oxide, not only does it become florid, but an exchange of gases takes
place, the loosely combined oxygen of the blood being expelled from
it, and its place taken by an equal volume of carbonic oxide.

After the discovery by Hoppe-Seyler and Stokes of the remarkable
spectroscopic properties of the blood-colouring matter, attention was
paid to blood which had been treated with CO, and it was found
that whilst the spectrum of such blood is almost identical with that
of oxy-haemoglobin, it possesses the property of resisting the action
of reducing agents.

Subsequently, Hoppe-Seyler found that after passing a stream of
CO through a solution of oxy-haemoglobin, and then adding alcohol,
on exposing the mixture to cold, crystals separated which were
identical in form with those of oxy-haemoglobin, but the solution of
which was unacted upon by the agents which reduce oxy-haemoglobin.

From all these observations it resulted that carbon mon-oxide
possesses the power of displacing the respiratory oxygen which
exists in a state of loose chemical combination with haemoglobin, and
of forming a compound possessed of nearly the same physical
properties but differing from it in being much more stable ; further
from the fact that, in the formation of this compound, one volume of
oxygen is exactly replaced by one volume of carbon mon-oxide, it
follows that a molecule of the latter takes the place of a molecule of
the former.

Although the spectrum of CO-haemoglobin very much resembles
that of oxy-haemoglobin, there are minute differences which are
shewn by comparing the spectra of the two bodies existing in a
solution of the same strength, and examined under precisely similar
circumstances. It will be seen (Fig. 22, spect. 6 compared with
spect. 2), that in the CO-haemoglobin both the bands a. and /3
are moved very slightly nearer the violet end of the spectrum.
Amongst other points of difference between the CO- and 0- com-
pound, we have to mention that the crystals and solutions of the
former have a tinge of blue which is wanting in the latter, and
that the crystals of CO-haemoglobin are slightly less soluble than
those of 02-Hb.

So far as the Author is aware, Jaderholm1 and Sorby are the only observers
who have stated the position of the bands of oxy-haemoglobin and of CO-hae-
moglobin in wave-lengths. According to Jaderholm the centre of oxy-haemo-
globin a corresponds to W. L. 5730, of ft to 5370. On the other hand the
centre of CO-haemoglobin -a corresponds to W. L. 5690 and of ft to 5340.
These determinations do not agree with those of Preyer, nor with indepen-
dent observations of the Author. In the first place the centre of these
bands is not constant for solutions of different strengths, for it will be found
that the band j3 extends more towards the blue than the green as the
concentration of the solution increases. According to Sorby the centre

1 Jaderholm; see abstract by Hammarsten in Maly's Jahresbericht, vol. iv. p. 106.

106 COMPOUNDS OF HAEMOGLOBIN WITH CO AND NO. [BOOK I.

of oxy-haemoglobin a corresponds to W. L. 5830, of (3 to 5450 ; of
CO-Hb a corresponds to W. L. 5755 and ft to 5420.

From our own observations we conclude in the case of the band a of oxy-
haemoglobin that its centre certainly corresponds almost exactly with W. L.
5780 (expressed in 10 millionths of a mm. for comparison with Jaderholm).
The band a of CO-haemoglobin corresponds, on the other hand, approxi-
mately to wave-length 5720. The centre of CO-haemoglobin (3 is from 5340
to 5380 according to concentration. It will be seen that these determi-
nations differ very materially from those of Jaderholm and Sorby.

It has been shewn by the researches of the Author1, of Bonders2,
and of Zuntz3, that although the compound of CO and haemoglobin is
much more stable than that of 0, it yet can be decomposed, and CO
expelled. By passing a neutral gas, or air, through solutions of
CO-haemoglobin for long periods the gas may be gradually driven
out, and the haemoglobin again becomes reducible. The same
happens if the blood be boiled in the mercurial pump.

The great stability of CO-haemoglobin enables us to detect it in the
blood of animals poisoned by this gas or by gaseous mixtures which
contain it.

The blood in these cases presents sometimes an unusually and per-
sistently florid colour ; whether it does so or not, it however is in great
part irreducible, i.e. after acting upon it with reducing agents two bands
yet remain in its spectrum.

It has recently been shewn by Hoppe-Seyler4 that the CO-haemoglobin
resists putrefaction for very long periods of time, so that two bands
remain visible for months and even years, whilst when normal blood
putrefies, the reduction of its O2-Hb to Hb takes place at once.
According to Hoppe-Seyler the fact that long-kept blood exhibits two
bands is a proof by itself that its haemoglobin has been combined
with CO.

In a later section of this chapter it will be mentioned that a useful test
for CO-blood is the production of a cinnabar-red precipitate on the
addition of caustic soda; this is believed by Jaderholm5 to be due to the
formation of a compound of CO with haematin.

Nitric So great is the affinity of this gas for oxygen

oxide, NO. ^nat ^ moment ft comes in contact with it, deep red

fumes of nitrogen peroxide, N02, are formed, and when these meet

water the decomposition indicated in the following equation results :

3N02 + H20 = 2HNO3 + NO.

. As has been previously said, all free acids, and salts with acid
reaction, ipso facto decompose the colouring matter of the blood, and

1 Gamgee, Journal of Anatomy and Physiology, vol. i. (1867) p. 346.

2 Donders, "Der Chemismus der Athmung, ein Dissociations-Process." Pfliiger's
Archiv, v. 20—26.

3 Zuntz, "1st Kohlenoxydhaemoglobin eine feste Verbindung? " Pfluger's Archiv,
v. 584—588.

4 Hoppe-Seyler, " Unveranderlichkeit des Kohlenoxyd-Hamoglobin bei Einwirkung
von Faulniss oder Pankreasferment ; Werth dieses Verhaltens fur den Nachweis der
Kohlenoxydvergiftung." Zeitschriftf.phys. Chem., Vol. n. p. 131.

5 Jaderholm, loc. cit.

CHAP. II.] THE BLOOD. 107

therefore in investigating whether NO can form a compound with
haemoglobin, analogous to the oxygen and carbonic oxide compounds,
certain precautions had to be taken; for firstly, by combining with the
respiratory O of haemoglobin, NO? would be formed, and next, by the
reaction of water upon this body, nitric acid would result, which would
immediately decompose the haemoglobin.

Hermann added ammonia to blood and then passed a stream of
NO through it; all the acid generated in the reaction, at the expense
of the oxygen of haemoglobin, was neutralised by the ammonia, and
thereafter a compound of NO with haemoglobin was formed.

Again when CO-haemoglobin was placed in a vessel from which
the air had been expelled and then a stream of NO was passed
through the liquid, this gas displaced the CO, and combined with the
haemoglobin in its stead.

Hermann found that the body thus produced yielded crystals
isomorphous with those of the oxygen and carbonic oxide compounds,
and that its spectrum presented a spectrum closely resembling theirs,
though like that of the CO-haemoglobin undergoing no change after
the addition of reducing agents.

We are therefore acquainted with three compounds of haemo-
globin with gases which are isomorphous, and in which presumably 1
molecule of haemoglobin is linked with 1 molecule of the gas. The
least stable of these compounds is that with oxygen, for it can be
decomposed by CO, which then takes its place, forming a compound
of intermediate stability, which in its turn can be decomposed
by NO. That in each case a molecule of the gas takes part in the
reaction is argued from the facts that CO displaces an equal
volume of 0 (O2 occupying the same volume as CO) and that the
three compounds are isomorphous, so that the constitution of the NO-
compound will almost certainly be similar to that of the CO body1.
Acetylene, In the case of both CO and NO we have unsaturated bodies

C2H2- which presumably satisfy their free affinities by linking them-

selves to the complex molecule of haemoglobin, and it is quite conceivable
that other similarly constituted bodies might exert a similar action. It has
indeed been surmised2 that Acetylene or Ethine, C2H2, actually does so form
a very unstable compound with haemoglobin, easily reducible by ammonium
sulphide or reducing tin solutions. An investigation made with a view
of testing the results in Hermann's laboratory has not confirmed the
existence of this acetylene compound.

Assumed Upon very slender evidence it has been advanced 3

compound of tkat hydrocyanic acid forms an easily broken up corn-
Hydrocyanic 1 •,! 1 11'

acid with hae- pound with haemoglobin.

mogiobin. If the acid be added to a solution of haemoglobin, on

crystallizing out the latter it retains some of the hydrocyanic acid,

1 See Hermann, "Ueber die Wirkungen des Stickstoffoxydgas auf das Blut."
Beichert und Du-Bois-Keymond's Archiv, 1865, p. 469.

2 Bistrow u. Liebreich, Ber. d. deutsch. chem. Gesellschaft Berlin, 1868,^p. 220.

3 Hoppe-Seyler, "Cyanwasserstoffhaemoglobinverbindungen." Med. Chem. Unter-
mchungen. Heft n. (1867) p. 207.

108 PRODUCTS OF THE DECOMPOSITION OF HAEMOGLOBIN. [BOOK I.

which can afterwards be obtained from it by distillation after acidu-
lating with sulphuric acid. It is to be noted that the spectrum
of the supposed hydrocyanic compound is identical with that of
oxy-haemoglobin, and that the behaviour of the solution to reducing
agents is absolutely the same as that of a solution of oxy-haemoglobin.

Those who advocate the existence of the compound however rely
somewhat upon the fact that blood to which hydrocyanic acid has been
added shews the bands of oxy-haemoglobin, or bands identical with
them, for a much longer time than normal blood — a fact which
they explain by supposing that the hydrocyanic compound is some-
what more stable than oxy-haemoglobin.

It appears to the Author that all proofs of the existence of such
a compound are wanting. That some hydrocyanic acid should adhere
to the haemoglobin as it crystallizes out is quite in accordance with
a variety of experiences of a similar kind, and can by itself afford
no evidence of an actual compound existing. The resistance of
hydrocyanic blood to decomposition can on the other hand be easily
explained by the unquestionable arrest or slowing 'of the process
of putrefaction in the presence of hydrocyanic acid ; it is undoubtedly
the products of putrefaction which are the causes of the spontaneous
reduction of the oxy-haemoglobin of blood confined in a vessel which
has no access to air, so that an agent which will inhibit putrefaction
and at the same time not decompose oxy-haemoglobin would be
expected to act as hydrocyanic acid acts and cause the persistence of
the oxy-haemoglobin bands.

Products of the decomposition of Haemoglobin.

When subjected to the action of various reagents, especially to that
of acids and of salts having an acid reaction, the molecule of haemo-
globin undergoes a profound decomposition, the ultimate products of
which are, amongst others, a proteid substance or substances, and a
body called HAEMATIN, which contains all the iron originally con-
tained in the blood-colouring matter. The formation of haematin
is, according to Hoppe-Seyler, necessarily dependent upon the
presence of oxygen, in the absence of which the process of decom-
position yields a proteid and a body to which he has given the name
of HAEMOCHROMOGEN ; the latter may by oxidation pass subsequently
into haematin. Haematin is an interesting body which forms definite,
well crystallized, compounds with hydrochloric, and apparently also
with hydriodic acid.

Before describing the various bodies which are the products of
a profoundly decomposing action exerted upon haemoglobin, it is
essential to refer to a modification of haemoglobin which is brought
about by the action of various agents, and concerning which very
much difference of opinion still lingers, viz. methaemoglobin.

CHAP. II.] THE BLOOD. 109

Methaemoglobin.

spectrum When a solution of haemoglobin is exposed to air

of Methaemo- for some time jt joges jts blood-red colour, assumes a
brownish tinge, presents an acid reaction, is precipi-
tated by solutions of basic lead acetate, and on examining its
spectrum it is found that the two bands of oxy-haemoglobin have
become faint, and that a new band has appeared in the red near C ;
this line occupies nearly though by no means exactly the position of a
similar band in the spectrum of acid haematin. On now rendering
the solution alkaline by the addition of ammonia, the band in the red
disappears, and is replaced by a faint absorption band immediately
near D.

The most remarkable phenomenon, however, relates to the action
of reducing agents.

If to a solution which exhibits the last - mentioned spectrum
there be added some sulphide of ammonium, it is observed that it
manifests the spectrum of reduced haemoglobin. On shaking the so-
lution containing the latter with air, oxy-haemoglobin is again formed.

Production The peculiar and remarkable properties above mentioned

of methaemo- were described by the Author in 18671 and more fully in

1868; as devel°Ped by the action °f nitrites on solutions of
haemoglobin and upon blood. It was shewn that besides
presenting the remarkable optical properties and reactions
previously referred to, as a result of the action of nitrites, the respiratory
, oxygen of haemoglobin had become irremovable by carbonic oxide or in a
Torricellian vacuum, but that after undergoing the change the haemoglobin
could be crystallized repeatedly, the body thus produced only differing from
oxy-haemoglobin by its colour and its spectrum. On analysis it was found that
the crystalline compound always retained some of the nitrite used, and the
view was therefore expressed that in all probability the action exerted by ni-
trites consisted in the formation of a compound of those bodies with oxy-hae-
moglobin, which compound was decomposed by the reducing agent employed.
It \vas subsequently observed by Sorby2, Lank ester3, and Jaderholm4
that Gamgee's nitrite-haemoglobin spectra coincided with those of methar-
moglobin prepared by the action of potassium permanganate, and the
presumption has been established that his bodies really consisted of
methaemoglohin generated by the action of nitrites. This change in the
view as to the nature of the bodies produced under the influence of
nitrites does not however affect the facts established by the researches
above referred to. According to Sorby, however, methaemoglobin would
be a per-oxy-haemoglobin, i.e. a more highly oxygenized haemoglobin,

1 Gamgee, "Note on the action of nitric oxide, nitrous acid and nitrites on
Haemoglobin." Proceedings of the Royal Society of Edinburgh, 1867, p. 168. " On
the action of nitrites on blood." Philosophical Transactions, 1868, pp. 589—625.

2 Sorby, Quarterly Journ. of Micros. Sc. 1870, p. 400.

3 Lankester, " Abstract of a Eeport on the Spectroscopic Examination of certain
animal substances." Journal of Anat. and Phys. Vol. iv. p. 123.

4 Jaderholm, " Untersuchungen iiber den Blutfarbstoff und dessen Zersetzungs
producte." Abstracted from the original Swedish by Hammarsten in Maly's Jahres-
bericht, Vol. vr. p. 85.

110 PRODUCTS OF THE DECOMPOSITION OF HAEMOGLOBIN. [BOOK I.

CHAP. II.] THE BLOOD. HI

in which the oxygen has become irremovable by a vacuum, but which
is decomposed at once by reducing agents, which first liberate oxy-haemo-
globin and subsequently form reduced haemoglobin. This view has
lately received the support of Jaderholm.

Taking all the facts in consideration we must admit that under the
innuence of various agents the loosely combined oxygen of haemoglobin
becomes irremovable by CO, and by a vacuum, whilst the new compound
still preserves the crystalline form of oxy-haemoglobin, and the capability
of being recrystallized. In this condition the body, which appears per-
fectly stable, can again be made to furnish haemoglobin. It is certainly
convenient at present to retain for this body the term Methaemoglobin.

Hoppe-Seyler, who was the first1 to describe briefly and to name
methaemoglobm, long ago arrived at the conclusion that probably no
definite body, such as is implied by the possession of a special name, exists,
but that it represents an intermediate stage in the decomposition of
haemoglobin into haematin and a proteid8.

FIGURE 23.

Spectrum 1. (Preyer, PI. 2, sp. 9.) Haematin in alkaline solution.
A single absorption band between C and D, from wave-length 618 to wave-
length 587. Strong absorption of the blue end

Spectrum 2. (Preyer, PL 2, sp. 10.) The same as 1, but more concen-
trated. As the concentration of the solution increases the band extends
more towards the red than the green. The red end of the spectrum is
much absorbed.

Spectrum 3. (Preyer, PI. 1, sp. 11.) Haemochromogen in alkaline
solution (Stokes' reduced haematin). The spectrum is distinguished from
all others by the extraordinary intensity and sharpness of the absorption
band nearest to D. This extends from wave-length 567 to 547. The
second absorption band, which is very much less intense and has less sharply
defined borders, extends from about wave-length 532 to 518. The solution,
even when concentrated, absorbs very little of the red. Violet and blue
are strongly absorbed.

Spectrum 4. Methaemoglobin. In weak solutions of certain strengths
four absorption bands may be made out. In a strong solution one is seen,
the centre of which, according to the Author's measurements, corresponds
to wave-length 632. According to Preyer this band would be a little nearer
to C, the centre corresponding to wave-length 634.

Spectrum 5. Diluted blood treated with acetic acid. An absorption
band in the red, the centre of which corresponds to wave-length 640.
According to Preyer the centre of this band corresponds to 65 6' 6.

Spectrum 6. Spectrum of acid haematin dissolved in ether. The
position of the three bands between B and E agrees with the observations
and drawings of Preyer. The centre of the band between b and F corre-
sponds to wave-length 502. According to Preyer its centre corresponds to
wave-length 505, i.e. it is somewhat nearer to b.

1 Hoppe-Seyler, Centralblatt f. d. med. Wissenschaften, 1864.

2 Hoppe-Seyler, Med. Chem. Untersuchungen, Heft in. (1868) p. 378.

112 PRODUCTS OF THE DECOMPOSITION OF HAEMOGLOBIN. [BOOK I.

Seyler's re- More recently Hoppe-Seyler has published fresh re-

searches and searches on the subject1. He opposes vehemently the view
views on that methaemoglobin is to be looked upon as a peroxidized

methaemo- oxy -haemoglobin, resting his opposition very much on the

globin. facts (a) that when a solution of oxy-haemoglobin is intro-

duced into the vacuum of the mercurial pump, so as to remove a part of its
respiratory oxygen, and then is left at the temperature of the room, the
fluid is found to contain a mixture of methaemoglobin and reduced
haemoglobin, (b) that when a piece of palladium saturated with hydrogen
is introduced into a flask filled with a saturated solution of oxy-haemo-
globin, the whole of the colouring matter is very quickly converted into
methaemoglobin, unless the quantity of the oxy-haemoglobin present was very
large. In these two experiments conditions existed for removing a great
part at least of the oxygen of the oxy-haemoglobin, and how therefore could
a per-oxy-haemoglobin be formed 1

Hoppe-Seyler has himself added lately the strongest proof of the possibi-
lity of reconverting methaemoglobin into oxy-haemoglobin by shewing that
when a solution of methaemoglobin is allowed to decompose in sealed glass
tubes, the band in the red of that body disappears and the spectrum of
reduced haemoglobin appears. When some months have elapsed and
the change has been completed, the tube is cooled to 0° until ice begins
to form, then opened, and alcohol is added to the extent of ^ of the volume
of the solution; on afterwards lowering the temperature to - 7° 0. or — 10°C.
crystals of oxy-haemoglobin separate,

It is now admitted by Hoppe-Seyler that this possibility of reconver-
sion into haemoglobin distinguishes methaemoglobin from haematin. Ac-
cording to this author methaemoglobin contains more oxygen than haemo-
globin but less than oxy-haemoglobin, and this oxygen is in a more stable
combination than in the latter body.

The Proteid matter derived from the decomposition of Haemoglobin.

When a solution of haemoglobin is boiled, the liquid becomes in-
tensely turbid and a coagulum soon separates which possesses a dirty
reddish-brown colour. Under the influence of heat the haemoglobin,
has been decomposed, and has yielded two substances insoluble in water,
the first of which is a proteid, and the second is the body already re-
ferred to as haematin.

The same decomposition takes place when strong acids, or when
large quantities of alcohol, act upon haemoglobin, though the rate
at which it proceeds varies in these different cases.

Very little information is possessed concerning the proteid matter
which results from this decomposition. According to Hoppe-Seyler it
behaves as a normal proteid in reference to bases and acids, .yielding
alkali- and acid-albumins.

Preyer has described the proteid substance under the term of
Globin, as a body which is free from all inorganic matter, which is
insoluble in water, which swells in solutions of sodium chloride

1 Hoppe-Seyler, "Die Zusammensetzung des Methamoglobin und seine Umwandlung
zu Oxyhamoglobin." Zeitschriftf. physiolog. Chemie, Vol. u. (1878) p. 150.

CHAP. II.] THE BLOOD. 113

and of sodium hydrate without dissolving. We agree with Kiihne
in holding that from the action of reagents one would conclude that
a mixture of proteids, rather than a single proteid, results from the
decomposition of haemoglobin.

Haematin.

When blood is treated with acetic acid it soon undergoes a
change of colour, from red to brown, which indicates the decomposition
of haemoglobin and the formation of haematin. If now the mixture
of blood and acetic acid be shaken up with ether, the latter dissolves
out a colouring matter, and on allowing the mixture to rest, the
coloured ether may be decanted.

On examining the ethereal solution it is seen to present the
spectrum represented in Fig. 23. 6, in which four separate absorption
bands are to be observed. Firstly an absorption band in the red between
C and D and corresponding to a wave-length of about 636, and secondly
a very faint and narrow band, close to D, with an approximate wave-
length of about 585, thirdly two much broader bands, one between D
and E, and another nearly midway between b and F, the centres of
which correspond approximately with wave-lengths 540 and 502 re-
spectively. Of all these bands the one in the red is by far the most
distinct.

If instead of experimenting in this way with ether holding acid
haematin in solution we merely add acetic acid to a haemoglobin
solution, we observe that as the liquid becomes brown in colour, the
band in the red developes (Fig. 23. 5) ; the other absorption bauds not
being obvious. If we render the liquid alkaline by the addition
of ammonia a single absorption band is seen, but much nearer to
D, its centre corresponding to about 636 or 640. A marked shading
of the blue end of the spectrum is noticed in addition. If now a re-
ducing solution as of ferrous tartrate (Stokes' reagent) be added to
the liquid, a spectrum is obtained which is marked by two bands
which at first sight appear to the tyro to be identical with the bands
of oxy-haemoglobin, but which are distinct from these ; they will be
found to be nearer the blue than are the bands of 02-Hb. (See
Fig. 23. 3.)

The first spectrum described is supposed to be that of haematin
in acid solution, the second haematin in alkaline solution, and the
third that of reduced haematin (Hoppe-Seyler's Haemochromogen).
That the last is a less oxygenized product than the second is proved,
not only by the fact that it is produced by the action of reducing agents,
but likewise by the fact that on shaking the two-banded spectrum of
reduced haematin with air or oxygen the two bands disappear and
are replaced by the single bands of alkaline haematin.

As will be more fully stated when discussing haemochromogen,
haematin is, according to Hoppe-Seyler, an oxidation product of
haemoglobin ; and it differs from haemochromogen, in that the latter

G. 8

114 PEEPAKATION AND PKOPERTIES OF HAEMATIN. [BOOK I.

is a simple product of decomposition, which can be formed from re-
duced haemoglobin in the absence of oxygen.

Methods of I. Blood (defibrinated) is mixed with ether and then a

preparation large quantity of strong acetic acid is added ; the two
ofHaematin. liquids are thoroughly shaken, and thereafter the dark-
brown ethereal solution is decanted, filtered and set aside. The deposit
which separates is washed with ether, alcohol, and water.

II. Blood is coagulated by the addition of an excess of cold alcohol ;
the precipitate is separated and boiled with alcohol holding sulphuric acid
in solution. The hot filtered solution is set aside, and the matter which
separates and adheres to the glass is washed with water and then with
alcohol, and ether.

Although the above methods may yield haematin with which some
qualitative experiments may be tried, we must employ the next process
if it be desired to obtain the pure substance, viz. : —

III. Crystals of Hydrochlorate of Haematin or Haemin are dissolved
in exceedingly dilute solution of pure potassium hydrate; the filtered solution
is neutralized with hydrochloric acid, which throws down haematin in the
form of a flocculent brown precipitate, which is washed with boiling water,
until the washings are no longer rendered turbid by solution of silver
nitrate. The haematin is then collected and dried, first at a gentle heat,
and then at 120°— 150° C. (Hoppe-Seyler1.)

Properties Haematin, obtained by the method last mentioned,

of fcaematin. ^as a blue-black colour and a decided metallic lustre; it
is free from crystalline structure, and when pulverized yields a dark-
brown powder.

It can be heated to 180°C. without undergoing decomposition, but
when heated more strongly it burns, evolving hydrocyanic acid, and
leaving an ash which consists of pure oxide of iron, amounting to
12 -6 per cent.

The following is the mean percentage composition of pure hae-
matin, as determined by Hoppe-Seyler: —

Carbon ........................ 64'30

Hydrogen ..................... 5*50

Nitrogen ..................... 9'06

Iron ........................... 8-82

Oxygen ........................ 12-32

100-00

These numbers agree well with the formula

Haematin is insoluble in water, alcohol, and ether, easily soluble
in solutions of the caustic alkalies, if these are not too concentrated,
insoluble in diluted acids, and soluble with difficulty in hot alcohol
holding sulphuric acid in solution.

Watery or alcoholic alkaline solutions of haematin when examined
in thin layers by reflected light possess an olive-green colour; deeper

1 Hoppe-Seyler, "Beitrage zur Kenntniss des Blutes des Menscheu und der
Wirbelthiere. Das Haematin." Med. Chem. Untersuchungen, Heft iv. 1871, p. 523. .

CHAP. II.]

THE BLOOD.

115

layers possess a fine red colour and absorb strongly, as is proved by
spectrum analysis, not only the violet rays, but also the yellow
between Frauenhofer's lines C and D, especially near the latter. If
alkaline solutions of haematin of sufficient dilution be examined, a
distinct absorption band, the centre of which corresponds approxi-
mately to wave-length 603, is observed.

Haematin dissolves sparingly in alcohol holding sulphuric acid in
solution, the solution assuming a dark-brown colouration.

Action of When heated with fuming hydrochloric acid to 160°C.,

hot HC1 on the iron which haematin contains is removed from it, and

is found in the solution as a ferrous salt, whilst a body free
from, iron, termed Haematoporphyrin, is formed. Alkaline solutions of
haematin, if pure, are not attacked by reducing agents. If, however,
organic matters, such as proteids, be present, haemochromogen (syn. reduced
haematin) is formed l. Haematin is scarcely, if at all, affected by putrefactive
processes. (Hoppe-Seyler.)

When potassium cyanide is added to an ammoniacal
solution of pure haematin, or to a solution of oxy-haemo-
globin, a broad band somewhat resembling that of reduced
haemoglobin, though by no means identical with it, is pro-
duced. This band extends from D to E. On adding reducing agents a
spectrum with two well-marked absorption bands is obtained. These optical
characters are supposed to depend upon the production of a compound
of haematiii and the cyanide employed, which has been denominated cyan-
haematin. We are, however, merely acquainted with the spectroscopic
characters of the supposed compound.

Action of
Potassium
Cyanide on

haematin.

Mode of
preparing
crystals of
Haemin for
microscopic
examination.

Hydrochlorate of Haematin — Haemin.

When a small drop of blood is boiled with a few drops
of glacial acetic acid, the red colour almost instantly gives
place to a brownish colouration. On evaporating down
the fluid a residue is obtained, which on microscopic
examination is found to be composed of reddish-brown

Fm. 24. CRYSTALS OF HAEMIN. (Frey.)

1 Hoppe-Seyler, " Weitere Mittheilungen iiber die Eigenschaften des BlutfarbstofFs."
Zeitschrift f. phys. Chemie. Vol. n. (1878) p. 154.

8—2

116 HAEMIN. [BOOK i.

prismatic crystals1. Such crystals can be obtained from any blood
stain, as on cloth or linen, by cutting out the stained tissue and
heating it with glacial acetic acid, taking care to add a small crystal
of sodium chloride. The evaporated residue contains the crystals.

Properties These crystals are of a dark brown and sometimes

of-naemin. Of nearly a black colour, and present the form of
rhombic plates sometimes arranged in radiating bundles.

Haemin is insoluble in water, alcohol, ether, chloroform, and in
cold dilute acetic and hydrochloric acids. It is however soluble in
caustic alkalies, in alcoholic solution of potassium carbonate, and in
boiling acetic and hydrochloric acids. It dissolves in concentrated
sulphuric acid, forming a violet-red liquid, which evolves hydrochloric
acid when heated.

Hoppe-Seyler has prepared this body by a method to be afterwards
referred to, and he considers it to be hydrochlorate of haematin and
ascribes to it the formula CggH^NgFe/^SHCl;, he found the com*
pound to contain 5'18 per cent, of chlorine.

It is held by Thudichum that haemin contains no chlorine, and
he therefore looks upon it as crystallized haematin. Hoppe-Seyler
however asserts that he has never obtained haemin crystals which
were free from chlorine, and the statement agrees with the original
observations of Teichmann who held the presence of chlorine to be
indispensable to their formation.

Prepara- Whilst it is very easy to prepare in a few minutes

tion of hae- microscopic crystals of haemin, the difficulties attending
min in large the preparation of considerable quantities in a pure con-
dition are considerable ; the following method has been
followed by Hoppe-Seyler: —

Defibrinated blood is mixed with a large excess of a solution of
sodium chloride, containing ^th its volume of saturated solution of
NaCl, and set aside in a cool place so as to allow the corpuscles to
subside; the clear supernatant fluid is decanted and the magma of
corpuscles is mixed with some water, placed in a flask, and shaken up
with ether ; the ethereal solution is decanted, the solution of colouring
matter is filtered and evaporated to dryness in shallow basins. The
residue can be readily pulverized. The powder is passed through a
sieve and then weighed. It is then mixed with glacial acetic acid in
a mortar, the mass is washed into a basin by the aid of more glacial
acetic acid, which is then added in such quantities that two litres of
glacial acetic acid are employed altogether for every 100 grammes of
the powder. The mixture, which has been mixed as well as possible,
is then heated on the water bath, the temperature of which is allowed
gradually to rise; the process of stirring is carried on from time to
time and the mixture is allowed to remain for some hours at 100°C.
Crystals soon commence to form, though long heating is required

1 Teichmann, Zeitschrift f. rat. Medizin f. Henle und Pfeuffer, 1853, Vol. m.
p. 375 and Vol. vm. p. 141.

CHAP. II.] THE BLOOD. 117

for the complete precipitation of the crystals and the solution of
the proteids. The whole mixture is then poured into a large beaker
and treated with many times its volume of water and set aside for
many days. The magma of crystals which has then fallen to the
bottom is washed many times in succession with water, and boiled
with strong acetic acid, as long as the crystals appear to be mixed
with masses of proteid matter ; they are then washed with water, and
collected on a filter and treated, first with alcohol, and then with
ether. Haemin crystals may also be obtained by adding water and
NaCl to a solution of haematin in alcohol which has been acidified
with sulphuric acid and then heating. A method has been suggested
by Gosdew for recrystallizing haemin, but it is not recommended by
Hoppe-Seyler, as he found it to yield a mixture of haemin with
haematin.

Haematoporphyrin.

Mode of When haematin is thoroughly mixed with concentrated

preparation. sulphuric acid, the substance dissolves, and, after filtering
through asbestos, a clear and beautifully purple-red solution is obtained.
When this solution is treated with a large quantity of water, the greater
part of the dissolved coloured body is precipitated in the form of a brown
flocculent precipitate, the quantity of which increases if alkalies be added so
as fully to neutralize the acid. In this operation the acid separates the
whole of the iron from the haematin, and it is found in the solution in the
state of a ferrous salt. In the process of decomposition of haematin by
sulphuric acid there is no evolution of hydrogen gas.

Properties. The precipitate which is thrown down by water from

the sulphuric acid solution is insoluble in concentrated solution of potassium
sulphate, but soluble in water, and the watery solution possesses the same
optical properties as the solution in sulphuric acid. It is also soluble in
alkaline leys, yielding solutions possessed of a reddish-brown colour ; in
undergoing solution the substance appears to undergo some decomposition.

Both the original sulphuric acid solution and the dilute alkaline solu-
tions of the body precipitated by water from it, possess characteristic and
different spectra.

The first (solution in strong sulphuric acid) exhibits a pretty dark
band immediately below D, and a very sharply defined band nearly inter-
mediate between D and E.

The second (.solution of precipitated body in alkaline leys) presents a
four-banded spectrum : to wit — a weak band midway between C and D, an
equally weak band between D and E but nearer to the former, a more strongly
marked band nearer to E, and lastly a fourth band, darkest of all, which
is not however very sharply defined, and extends through. -fths of the
space beween b and F.

To this iron-free body, obtained from haematin by the action of strong
sulphuric acid, Hoppe-Seyler attaches provisionally the term of Haemato-
porphyrin, and ascribes the formula C^H^NgO^.

\

118 HAEMATOLIN. HAEMOCHROMOGEN. [BOOK 1.

A second iron-free derivative from haematiii has been obtained by
Hoppe-Seyler, which differs from haeinatoporphyrin in being nearly in-
soluble in sulphuric acid and in caustic leys. To it he attaches the pro-
visional name of Haematolin, and the formula CegH^NgOy1,

Haemochromogen.

According to Hoppe-Seyler when reduced haemoglobin is decom-
posed in the absence of oxygen, instead of haematin, there is produced
a body to which he gives the name of haemochromogen, whose
solution presents a beautiful purple colour, but which is converted
almost instantly into haematin when oxygen comes in contact with it.
This body when in alkaline solution is, as proved by the most careful
measurement of its absorption bands, identical with the so-called
reduced haematin of Stokes.

The following are the two methods which at different times
Hoppe-Seyler has employed for the preparation of haemochromogen: —

I. In the Woulffs bottle A (see annexed woodcut) hydrogen is evolved
by the action of dilute hydrochloric acid upon zinc, and the gas is washed
by passing through the wash-bottle (7, which contains dilute solution of
caustic soda. In order that the acid which is to act upon the zinc shall be
free from oxygen, a piece of zinc is placed in the beaker B which contains
the acid.

First of all having opened the clips b and &', by aspirating at the a end
of the wash-bottle, a sufficient quantity of acid is made to flow out of the
beaker B to fill the tube f and then enter A where it evolves hydrogen,
which gradually expels all the air from the apparatus. The clip b is
then closed. After gas has been passing for about half an hour the
bulb-apparatus DEF is attached to the wash-bottle in the manner
represented in the diagram. This bulb-apparatus contains in the
division F concentrated solution of oxy-haemoglobin, and in the
division D alcoholic solution of sulphuric acid or potassium hydrate, or
instead of these an aqueous solution of potassium hydrate. A stream
of gas is now again allowed to pass through the apparatus by opening the
clip b and raising the vessel B so as to allow a fresh quantity of dilute acid
to enter A and act upon the zinc which it contains. (If A happens to be
already full, the solution of ZnCl2 which it contained might be removed by
depressing the vessel B and allowing it to be on a lower level than A.
The vessol A having been thus more or less completely emptied, and
the clip b closed, a fresh stock of dilute acid may be placed in BA,
and everything is ready for recommencing.) After a stream of H has
passed through the whole apparatus, including the bulbs, for some
considerable time (2 or 3 hours), the bulb-apparatus is sealed in a blowpipe
flame at d and at e. By means of the spectroscope the observer determines

1 The whole description of haematin and its derivatives is abridged from the
memoirs of Hoppe-Seyler, of which the most important relating to this subject
is the one entitled "Das Hamatin," under the general heading of "Beitrage zur
Kenntniss des Blutes des Menschen und der Wirbelthiere." Hoppe-Seyler's Med.
Cliem. Untersuchungen, Heft iv. (1871) p. 523. See also " XJeber die Zersetzungen der
Hamoglobine." Ibid, p. 377—385.

CHAP. II.]

THE BLOOD.

119

before sealing whether the froth which fills the division F exhibits, as it
ought to do, the spectrum of reduced haemoglobin; if it does so, 'after

FIG. 25. APPARATUS FOB THE PREPARATION OF HAEMOCHROMOGEN.

sealing, the fluids contained in the bulbs D and F are mixed by reversing
and shaking their contents together.

If in this way, in the complete absence of oxygen, acid alcohol
has been mixed with a little haemoglobin, a precipitate forms, which
soon loses its colour on being heated in the water bath, whilst the
liquid becomes coloured purple. The liquid then exhibits four absorption
bands, of which two are situated between C and D. A third absorption
band of greater degree of sharpness and darkness extends between D and E,
and a fourth is situated between b and F. The absorption band nearest C
is, if the oxygen has been thoroughly expelled, exceedingly weak and may
be due to a trace of haematin, as its position is identical with the band of
acid haematin.

If instead of sulphuric acid, alcohol holding caustic alkali in solution has
been employed, on mixing the fluids we obtain a rose-red or purple-
red precipitate and a solution having the same tints. This exhibits two
absorption bands which are identical . with those of Stokes' alkaline
haematin.

II. Lately Hoppe-Seyler has recommended the following method1.

A solution of oxy-haemoglobin is placed in a glass tube, and then a
smaller glass tube containing solution of dilute phosphoric or tartaric acid,
or solution of potassium hydrate, is introduced into the larger tube, the open
end of which is then drawn out and sealed ; the large tube with its
contained smaller tube is then heated gently for some time, care being
taken that the contents of the two tubes do not mix. The oxy-haemoglobin

1 Hoppe-Seyler, "Weitere Mittheilungen tiber die Eigenschaften des Blutfarb-
stoffs." Zeitschrift f. phys. Chem. Vol. i. p. 138.

120 HAEMATOIDIN. [BOOK I.

first becomes reduced, and thereafter the oxygen contained in the air
of the tube is removed by it. When many days have elapsed and
the whole of the haemoglobin is reduced, the tubes are reversed and their
contents mixed, when the optical properties of haemochromogen can
be satisfactorily observed.

According to Jaderholm1, Hoppe-Seyler's haemochromogen in alkaline
solution is identical with the reduced haematin of Stokes, and haemochro-
mogen in acid solution has a spectrum which is a combination of those of
acid haematin and haematoporphyrin. The former statement is indeed
admitted by Hoppe-Seyler, and is indisputable. Hoppe-Seyler urges, how-
ever, and as it appears to the Author, most correctly, that the term reduced
haematin is a misleading one, haemochromogen being a mere product of
decomposition of haemoglobin, whilst haematin is an oxidized product of
decomposition.

Haematoidin.

This name has been assigned by Virchow2 to a substance which
occurs in the form of yellow microscopic crystals in old extravasations
of blood, as for example in old apoplectic clots, and which was first
observed by Everard Home 3.

FIG. 26. CRYSTALS OF HAEMATOIDIN (AFTER FUNKE). (Frey.)

These crystals appear to be identical in form with those of Biliru-
bin, the chief colouring matter of human bile, and when treated with
fuming nitric acid give the same colour reaction (Gmelin's reaction).

Opinions have been divided on the question of the

identity identity or non-identity of haematoidin and bilirubin.

din and wii- On ^ne ground of different deportment towards solvents

rubin. Holm4 asserted that haematoidin, prepared from the

1 Jaderholm, " Untersuclmngen iiber den Blutfarbstoff und dessen Zersetzungspro-
ducte." Abstracted from the Swedish by Hammarsten in Maly's Jahresbericht, Vol.
vi. p. 85.

a Virchow, Archiv d. pathol. Anat. u. PhysioL Vol. 1 (1847), p. 383—443.

3 Sir Everard Home, A short tract on the Formation of Tumours, &c. London, 1830,
page 22. In Figs. 1, 2 and 3 of Plate I., crystals of haematoidin are admirably figured
as seen in an aneurismal coagulum. Home was, however, altogether ignorant of their
nature and referred to them as ' crystallized salts. '

•* Holm, "Haematoidin," Journ. f.prak. Chemie. Vol. c. p. 142.

.CHAP. II.] THE BLOOD. 121

corpora lutea of the cow, is not identical with bilirubin. Salkowski1,
on the other hand, found haematoidin prepared from the con-
tents of a strumous cyst to be identical in all respects with
bilirubin. Preyer2, relying mainly though not entirely upon the
spectra of the two bodies, denies the identity. According to this
observer bilirubin possesses no definite absorption-band, whilst solu-
tions of haematoidin when examined with the aid of magnesium light
present a well-marked absorption-band between b and F, and a
weaker one nearly midway between F and G.

The majority of physiological chemists are, however, now of the
opinion that haematoidin and bilirubin are identical. This matter will
be again referred to under 'bilirubin.'

THE MINERAL CONSTITUENTS OF THE RED CORPUSCLES.

It was pointed out in discussing the salts of the serum and plasma
that our information in reference to these was far from complete, in
consequence of the inherent difficulties which attach to the methods of
research. The same remark appears with still greater force to the
mineral matters of the corpuscles. It is possible to obtain plasma and
serum free from corpuscles (though certainly not free from all constitu-
ents of corpuscles, e.g. serum-globulin), but far from possible to obtain
corpuscles free from the liquids in which they float. Comparative
analyses, however, of the mineral matters of the serum and of the clot,
and of the blood as a whole, do lead to certain results which are to
be relied upon. They at once reveal, for instance, that the iron of
the blood is, with the exception of the minutest traces, contained in
the corpuscles, where we know it to exist as an essential constituent
of haemoglobin ; that the corpuscles are much richer in potassium
salts than the serum, and that the amount of chlorine is very much
greater in the latter than in the former. When, however, we enquire
whether phosphates and sulphates exist in the blood-corpuscles, or
whether these ingredients of the ash are not due to the oxidation of
organic constituents, we can merely say that the experimental data for
furnishing an answer to the question fail, though from the fact that
the blood-corpuscles are rich in lecithin we cannot doubt that nearly
the whole, if not the whole, of the phosphoric acid found in the ash,
is derived from the oxidation of that body.

The ana- In order to impress upon the reader the difference between

lysesof tne minerai constituents of the blood-corpuscles and the

plasma, the results of C. Schmidt's analyses of both are
here appended : —

1 Salkowski, " Zur Frage iiber die Identitat des Hamatoidin und Bilirubin,"
Hoppe-Seyler's Med. Chem. Untersuchungen, in. p. 436.

2 Preyer, Die Blutkrystalle, p. 187.

122 MINERAL CONSTITUENTS OF BLOOD CORPUSCLES. [BOOK I.

1000 parts of plasma yield :

Mineral matters
CHLORINE .
Sulphuric anhydride .
Phosphorus pentoxide
Potassium .
SODIUM .
Calcium Phosphate
Magnesium Phosphate

8-550

3'640

0-115
0-191
0-323

3-341

0-311
0-222

1000 parts of moist corpuscles yield :
Mineral matters (exclusive

of Iron) . . . 8-120
Chlorine .... 1-686
Sulphuric anhydride . . 0'066
PHOSPHORUS PENTOXIDE 1134
POTASSIUM . . . 3'328
Sodium .... 1-052
Calcium Phosphate . . 0*114
Magnesium . . . . 0'073

One would be inclined to attribute too great an importance to the
remarkable difference in the distribution of potassium and sodium in
the blood corpuscles of man if one were in ignorance of the undoubted
fact that this difference does not hold in the case of most animals.

Thus if we glance at the subjoined tabular view which contains
the results of the analyses of Schmidt of the inorganic matters yielded
by the blood cells and plasma of several animals, we come to the
conclusion that the proportions of sodium and potassium in the
corpuscles may vary within wide limits, and that in most animals the
salts of sodium preponderate greatly over those of potassium.

TABLE SHEWING THE AMOUNT OF POTASSIUM, SODIUM AND CHLOKINE
PEE SENT IN 100 PARTS OF THE INORGANIC MATTERS OF BLOOD
CELLS AND PLASMA1.

Blood Cells.

• i
Liquor Sanguinis.

K

Na

Cl

K

Na

Cl

Man (mean of 8 experiments)

-L'Og » 55 )) 5)

Cat ,, ,7 ,, 5,
Sheep „ „ „ ,,
Goat „ ,, ,, „

40-89
6-07
7-85
14-57
37-41

9-71
36-17
35-02
38-07
14-98

21-00
24-88
27-59
27-21
31-73

5-19
3-25
5-17
6-56
3-55

37-74
39-68
37-64
38-56
37-89

4068
37-31
41-70
40-89
40-41
i

The much more recent researches of Bunge2, whilst they differ in
some respects materially from those of C. Schmidt, indicate that in
some animals potassium and in others sodium preponderates. Thus
Bunge found no sodium (!) in the blood corpuscles of the dog and of
the cat, whilst he found nearly three times as much sodium as
potassium in the blood of the ox. These differences perhaps will be
explained, as some have surmised, by further researches proving that
when considerable quantities of potassium salts are ingested, they
replace sodium in the corpuscles, though probably before being able to do
so the richness of the blood in potassium must attain a certain figure.

1 Lehmann, Physiological Chemistry, Vol. n. p. 189. This table, which the
Author has modified somewhat in form, is compiled from the observations of C. Schmidt.

2 Bunge, " Zur quantitativen Analyse des Blutes." Zeitschr. f. BioL, Vol. xn.
p. 191—216.

CHAP. II.] THE BLOOD. 123

THE GASEOUS CONSTITUENTS OF THE COLOURED CORPUSCLES.

In discussing the properties of oxy-haemoglobin we have studied
with considerable minuteness the nature of that compound, and have
shewn that it is produced by the union of oxygen from the air with the
complex molecule of haemoglobin. We have shewn that under
various circumstances oxygen can be expelled from its state of com-
bination, as when blood is introduced into a Torricellian vacuum,
when neutral gases such as H and N are passed through it, or when
CO or NO act upon it.

Now, although the oxygen removed by these various means is de-
rived from the oxy-haemoglobin of the corpuscles, in that body it exists
in a state of actual combination — in a state very different from that in
which a gas exists which is merely dissolved in a liquid or absorbed by
a solid body, so that strictly we have as little right to speak of the 0 of
the corpuscles as one of their gaseous constituents as we have so to
designate the H or N which are essential constituents of haemoglobin.

We may however state that which we shall in succeeding sections
comment upon at far greater length, viz. that of the mixed gases
which are given up by blood when it is heated in a Torricellian vacuum
and which consist of a mixture of O, CO2 and N, practically the
whole of the first is derived from the dissociation of oxy-haemoglobin,
of which each gramme can give up as much as T28 c.c. of O (at 0° C.
and 1 metre pressure). Of the carbonic acid thus obtained the greater
part is derived from the plasma in which it is partly dissolved and
partly loosely combined, a small quantity only being derived from the
blood corpuscles. Probably the whole of the nitrogen, obtained from
the blood, is held in solution in the liquor sanguinis.

In short, if we wish to be strict in our expressions, we should say that
probably the only gaseous constituent properly so called,i.e.gsis not exist-
ing in a state of chemical combination in the corpuscles, is carbonic acid.

SECT. 5. THE COLOURLESS CORPUSCLES OF THE BLOOD.

In addition to the red corpuscles, which have been already described, the
blood of vertebrate animals contains a number of globules and particles of
various sizes and characters, all included under the designations of white
corpuscles and intermediate corpuscles.

The members of the first class are readily defined. They are nucleated
masses of protoplasm destitute of any cell-membrane, and containing fine or
coarse granules. They were first discriminated from the red corpuscles by
Hewson : and they were for a long time spoken of as lymphatic
corpuscles. In man they have a diameter of about 10 /* (^-^j in.),
while in batrachians they are much larger. Their most important property
is, without question, that of amoeboid movement, which was first observed
by Wharton Jones1 in the blood of the skate. The recognition of the
power of amoeboid movement of white blood corpuscles was one of the most

1 Phil. Trans. 1846.

124 THE COLOURLESS CORPUSCLES OF THE BLOOD. [BOOK I.

important steps in establishing the analogy between the sarcode of the lowest
animals and the substance of the cells composing animals of higher grades.

Another interesting property of white blood corpuscles is that of
enveloping and absorbing small particles of colouring matter, such as carmine,
with which they are in contact.

The members of the second class, the intermediate corpuscles, are less
clearly denned than the amoeboid corpuscles ; and for a full description of
all their varieties, the reader is referred to the larger text-books and
memoirs on the Histology of the Blood. But among them must be
mentioned some which seem to have a great importance in the phenomenon
of coagulation. These are described by Semmer under the name of red
granular corpuscles (rothe Komerkugel1), and by Hayem2 under the name
of haematoblasts. According to Semmer, who examined the blood of the
horse and other mammals, the granular red intermediate corpuscles are
nucleated granular bodies, the granules largely obscuring the nucleus. They
have about the same specific gravity as the white corpuscles ; hence they
subside in the uncoagulated plasma more slowly than the common red
corpuscles. They possess the power of amoeboid movement.- They become
colourless and readily disintegrate during the act of coagulation ; and the
detritus appears to be soluble in the plasma. The disintegrating corpuscles
iu many cases form centres for the radiation of threads of fibrin through the
coagulating liquor sanguinis (refer to p. 35, and fig. 10).

The number of the white corpuscles, though less than that of the red,
varies with the many conditions of age, sex, period after food and region
from which the specimen of blood was taken. On an average there is one
white corpuscle to 330 or 350 red ones.

The proportion is — 3

In boys 1 to 226

„ girls 1 to 389

„ men 1 to 346

„ old men 1 to 381

„ menstruating women . . . 1 to 247

„ pregnant women . . . . 1 to 281

„ the morning fasting state . . 1 to 716

Half an hour after breakfast . . 1 to 347 f

Three hours after breakfast . . . 1 to 1514
In splenic vein . . . . . - 1 to 60

„ splenic artery . . . . . 1 to 2260

„ hepatic vein . . . . . 1 to 170

„ portal vein. . . . ... 1 to 740.

Our knowledge of the physical and chemical characters of the
colourless corpuscles is for obvious reasons very much more defective
than that of the coloured corpuscles.

1 Alex. Schmidt, "Ueber die Beziehung der Faserstoffgerinnung zu den korperlichen
Elementen des Blutes." Pt. 2. Pfliiger's Archiv /. d. ges. Physiol. Vol. xi. (1875) p. 560.

2 Georges Hayeni, Eecherches sur Vanatomie normals et patholoaique du sang. p. 108.
Paris, 1878.

3 The above figures are taken from Strieker's Handbook. Art. "Blood," by Alex.
Rollett.

CHAP. II.] THE BLOOD. 125

The colourless corpuscles are obviously much lighter than the
coloured, as is evidenced (1) by their always being found in greater
abundance near the upper surface of a blood clot; (2) by their
forming a separate white layer on the surface of the red corpuscles,
when horse's blood is cooled with the object of separating the corpuscles
from the liquor sanguinis.

The colourless corpuscles exhibit obvious adhesiveness even when
contained in the blood-vessels of the living body, an adhesiveness which
causes them to cling one to the other when they meet, and to foreign
bodies or blood clots which may happen to project into the blood stream.

The great mass of the protoplasm of the colourless corpuscle is

undoubtedly proteid in its nature, the proteid matter having associated
with it smaller quantities of other principles, and imprisoning the
nucleus or nuclei which we may provisionally assume to be composed
of that somewhat non-descript, phosphorus-containing, non-digestible,
mucin-like body, Nuclein (see p. 82).

The protoplasm of the colourless corpuscles appears to undergo, at
least partial, coagulation at 40° 0. It swells and becomes transparent
when treated with acetic acid, which renders the nuclei much more
sharply defined and distinct.

The protoplasm swells and ultimately dissolves in 10 p.c. solution
of NaCl, leaving the nuclei undissolved. The salt solution thus
obtained is precipitated by the addition of a large quantity of water,
is coagulated by heat and by mineral acids.

The colourless corpuscles sometimes contain within them minute
fat-granules.

Many of the white corpuscles of the blood present, when treated
with a solution of iodine in iodide of potassium and water, a reddish
mahogany colour, which is due to their containing Glycogen. The
solution recommended to be used is one made by dissolving
1 gramme of iodine and 2 grms. of potassium iodide in 100 c.c.
of water. "The main substance of the corpuscles is uniformly
stained of a deep yellow, but many contain groups of mahogany-
stained granules, and from others are seen to exude after a time
pellucid drops of varying size, which become tinted of a mahogany or
port wine colour, and no doubt contain glycogen1."

The average proportion of colourless to coloured corpuscles is
liable to considerable variations consistently with health. It under-
goes physiological fluctuations which are related to the process
of digestion, viz. the colourless corpuscles increase after the in-
gestion of food, and diminish during fasting, a fact explained
in great measure by the fact that in the former case a larger
influx of colourless cells takes place through the thoracic duct.
The origin and destination of the colourless corpuscles, though
perhaps beyond the scope of this work, will be shortly treated of
under l lymphatic glands.'

1 Schafer, A Course of Practical Histology. Smith, Elder and Co., 1877.

126 THE GASES OF THE BLOOD. [BOOK I.

SECT. 6. THE GASES OF THE BLOOD AS A WHOLE.

Under the head of 'The Gases of the Liquor Sanguinis' and 'The
Gaseous Constituents of the Coloured Corpuscles,' it has been shewn
that from each of these constituent parts of the blood, there can be
separated, by certain methods of treatment, gases, which are a mixture
of carbonic acid, oxygen and nitrogen. We shall give a description
of the methods employed in separating the gases of the blood in
Chapter IV., and postpone a lengthened theoretical treatment of the
gases of the blood to the chapter on Respiration. In this place it will
suffice if we make the following brief statements.

(1) The blood, when admitted into an empty space- and exposed to
the temperature of the body, readily gives up more than half its volume
of mixed gases, consisting of oxygen, carbon dioxide, and nitrogen.

(2) The first (oxygen) is present in much larger quantities than
could be held in simple solution by the water of the blood, and, as will
be afterwards proved, is mainly held in feeble combination by the
haemoglobin of the coloured blood corpuscles; only a trace of it is,
under ordinary circumstances, held in solution in the liquor sanguinis.

(3) The second (carbon dioxide), whilst not existing in larger
quantity in blood than it could do if simply dissolved by the water of
that fluid, is partly in a state of chemical combination but chiefly in
a state of simple solution. It is contained in great part in the liquor
sanguinis and serum, but in part also in the corpuscles.

(4) The nitrogen is held in a state of simple solution in the liquor
sanguinis.

(5) Arterial blood of the dog of mean composition yields for
every 100 volumes, 58'3 volumes of mixed gases (measured at 0° C.
and 760 mm.), composed of 22-2 volumes of O, 34'3 volumes of
C02, and 1*8 volumes of N, the maximum amount of oxygen observed
having been 25*4 volumes (Pfluger1).

(6) As venous blood differs in composition according to the
vascular area whence it is obtained, it is impossible to state the mean
composition of its gases ; the following facts are however correct : —
the nitrogen is present in the same proportion as in arterial blood,
the 0 is less in amount (from 8 to 12 volumes per 100 of blood) and
the C02 greater (from 40 — 50 volumes per 100 of blood).

Summary of the .Quantitative Composition of the Blood.

Having treated at length the properties of the individual constituents
of the blood, we shall here append tables exhibiting the results of
the elaborate researches of C. Schmidt and Lehmanu on the blood of man,
although some of the data have already been referred to in the preceding
pages.

1 Efliiger, "Die normalen Gasmengen des arteriellen Blutes nach verbesserten
Methoden." Centralblatt f. d. med. Wissenschaft, 1868.

CHAP. H.]

THE BLOOD.

127

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128 SCHMIDT'S AND LEHMANN'S ANALYSES OF BLOOD. [BOOK i.

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Magnesium

CHAP. II.] THE BLOOD. 129

SECT. 7. CHARACTERS PRESENTED BY THE BLOOD OF
INVERTEBRATE ANIMALS.

It has already been stated that with very few exceptions *, the
blood of all vertebrate animals is characterized by the possession of a
red colour which is due to the presence within it of coloured
corpuscles, which in all classes but one (that of the Mammalia) are
nucleated. In addition to the coloured corpuscles, we have seen that
the blood always contains a much smaller number of colourless cells,
consisting of nucleated masses of protoplasm, endowed with con-
tractility, and presenting many of the essential features of independent
elementary organisms, and it has been incidentally remarked that
there appears to be a much greater uniformity in the shape and size
of the colourless than of the coloured corpuscles of the blood of
different classes of Vertebrates.

When we pass from the vertebrate to the invertebrate sub-king-
doms we find that in all those organisms in which a differentiated
blood-vascular system exists, the contained liquid presents floating in it
nucleated masses of protoplasm closely resembling the colourless cells
of vertebrate blood, but is generally, though not invariably, free from
all representatives of the coloured corpuscles. In the immense majority
of invertebrate animals this intra-vascular liquid is colourless, or
presents a yellowish tint, though in a small minority it is coloured red,
or green, or blue. Generally, however, the colour is diffused through
the liquor sanguinis if it is not actually dissolved in it.

In the colourless liquid contained in the vascular system of most
Invertebrates, we have probably a liquid which discharges only one
half of the functions of the vertebrate blood — which serves merely as a
common medium, supplying liquid and solid matters to the various
tissues and organs, and washing away from them products of waste
and decay, which it discharges through the agency of, or at, the various
excretory organs. The other half of the functions of the vertebrate
blood, the respiratory,, are probably scarcely represented by the
colourless blood of Invertebrata.

Such blood possesses, probably, no arrangement whereby the
oxygen of the medium external to the body can be stored up by it,
at certain points, to be carried away to tissues and organs far removed
from that medium and then given up.

The respiratory exchanges in creatures provided with such blood
probably take place by processes of diffusion directly between the
tissues of the organism and the medium which it inhabits, and

1 It is a matter of dispute whether the blood corpuscles of Amphioxus contain
haemoglobin. According to Bay Lankester they do not. In Leptocephalm we have at
any rate a fish whose blood is certainly free from haemoglobin. (Lankester: "A
Contribution to the Knowledge of Haemoglobin. " Proceedings of Royal Society, Vol. xxi.
(1872) p. 71 et seq.

G. 9

1.30 DISTRIBUTION OF HAEMOGLOBIN IN INVERTEBRATA. [BOOK I.

without the intermediation of any special arrangement such as is
represented by the haemoglobin of the vertebrate coloured corpuscles.

In the Invertebrata whose blood is coloured, we have, however,
undoubtedly, a clear indication of the blood discharging respiratory
functions, for such blood, when red, contains oxy-haemoglobin,and when
of other colours, sometimes undoubtedly does contain matters which
are capable of acting as oxygen carriers.

The following are the most important facts which have been
discovered in reference to the chemical composition of the blood of
invertebrate animals :

Distribution of Haemoglobin through the vascular liquids of various groups

of Invertebrata.

Our knowledge of this subject is mainly derived from the researches of
Professor Ray Lankester1. The following are the chief conclusions to
which he has arrived.

Haemoglobin is contained — •

1. in special corpuscles :

a. In the perivisceral fluid of some species of the Vermian genera,
Glycera, Capitella and Phor&nis.

b. In the blood of the Lamellibranchiate Mollusk, Solen legumen.

2. Diffused in a vascular or ambient liquid :

a. In the peculiar vascular system of the Chaetopodous Annelids very
generally, but with apparently arbitrary exceptions.

b. In the vascular system (which represents a reduced perivisceral
cavity) of certain Leeehes, but not of all (Nephelis, Hirudo).

c. In the vascular system of certain Turbellarians as an exception
(Polia sanguirubra).

d. In a special vascular system (distinct from the general blood-system)
of a marine parasitic Crustacean (undescribed) observed by Pro-
fessor Eclouard van Beneden..

e. In the general blood-system of the larva of the Dipterous Insect
Cheironomus.

f. In the general blood-system of the Pulmonated Mollusk Planorbis.

g. In the general blood-systems of the Crustaceans Daphnia and
Cheirocephalus.

In reference to Planorbis, Mr H. C. Sorby has made observations which
lead him to doubt very strongly whether the red colouring matter be really
haemoglobin 2, Mr Sorby's doubts are based (1) upon the fact that
the measurements of the bands in the spectrum of the blood of Planorbis
differed slightly from those of oxy-haemoglobin ; (2) that the red colouring
matter in the blood of Planorbis seemed to resist the action of decomposing
reagents (such as acids) longer than haemoglobin. According to Sorby
the following are the centres of the bands of normal haemoglobin and of

1 Lankester, Op. cit.t p. 76.

. 2 H. C. Sorby, " On the Evolution of Haemoglobin." Quarterly Journal of
Microscopical Science. Vol. xvi. N. S. (1876) p. 76 et seq.

CHAP. II.] THE BLOOD. 131

the colouring matter of the blood of Planorbis, expressed in wave-lengths in
millionths of a millimetre.

Centres of Bands.

Normal oxy-haemoglobin 581 545
Planorbis 578 542-J.

According to the measurements of Preyer and the Author, the position of
the bands in Planorbis as stated above really coincides almost exactly with
that of the bands of oxy-haemoglobin.

It must not be concluded that all the red colouring matters found
in invertebrate animals are identical with haemoglobin. Thus the peri-
visceral cavity of Sipunculus nudus, which is abundant in the Gulf of
Naples, has a pale madder-like colour due to a large number of coloured
corpuscles, varying in size between -^--Vfrth and -^V^-th of an inch, in which
a pink colouring matter is deposited. This colouring matter, which is
found in other tissues of that creature, is quite distinct from haemoglobin1.
Whether certain crystals which are obtainable from the blood of insects
consist of haemoglobin or not has been disputed, and yet deserves further
investigation 2.

On the Green Blood of Certain Annelids. Chlorocruorin.

In 1838 Milne Edwards3 had discovered that in certain Annelids
of the genus Sabella, the blood possessed a green colour, and a
similar observation was made by M. de Quatrefages in the case of the
annelid Chloronema Edwardsi. Professor Ray Lankester4 some years
ago shewed that the green colour is due to a body to which he gave
the name of CHLOROCRUOEIN.

Lankester's researches were carried out on Sabella ventilabrum
and Siphonostoma.

He found that the blood yielded an absorption spectrum with two
distinct bands, viz. one between C and D, and a second much less
distinct band in the green, almost midway between D and E. On
reducing a solution of the blood by means of one of the reagents
used with a similar object in the case of haemoglobin, Lankester
found that the two bands were replaced by a single band having nearly
the same position as the darker of the two, though fainter than it.
On agitating with air the two bands returned.

The Author has reduced Professor Lankester's observations to a scale of
wave-lengths, and finds that the first band of oxy-chlorocruorin, as drawn
by Lankester, extends from wave-length 588 '5 to 617, its centre being,
therefore, 602*7. The second band extends from 560 to 570. The band
of (reduced) chlorocruorin extends from wave-length 588'5 to 611'3, and
its centre therefore corresponds to wave-length 600.

1 Lankester, Op. cit., p. 80.

2 Landois, Zeitschr. f. wiss. Zoologie, Vol. xiv. pp. 55— 70, Plates vn.— ix.,
(quoted by Preyer, Op. cit. p. 10). The Author has not seen th'e original paper.

3 Milne Edwards : " Eecherches pour servir a 1'histoire de la circulation chez les
Annelides." Ann. des Sciences Natur., 1838. 2me aerie. Vol. x. p. 190.

4 Lankester : Journal of Anatomy and Physiology, 1868, p. 114: ibid. 1870, p. 119.

C) 2

132 THE BLUE BLOOD OF MOLLUSCA AND MOLLUSCOIDA. [BOOK I.

To the green substance Lank ester applied the term Chlorocruorin,
and concluded that this body, like haemoglobin, was capable of existing
in two states of oxidation ; when oxygenized he proposed to designate
it oxy-chlorocruorin. Furthermore Lankester found that the action
of certain reagents upon chlorocruorin appeared to indicate that
when decomposed it yields products which have identical spectra to
those of certain haematin derivatives.

ON THE BLUE BLOOD OF CERTAIN OF THE MOLLUSCA AND MOLLUSCOIDA.

1. The blood of the mollusca has received considerable attention.
Usually the blood of animals belonging to this class presents a white
colour, but sometimes it is distinctly of a blueish tint. C. Schmidt
analysed the blood of the Pond-mussel (Anodonta cygnea) and found it to
be colourless and slightly alkaline. It deposited a pale fibrinous coagulum ;
it contained 0-854 p. c. of solid constituents, and of these there were 0-033
of a fibrin -like body, 0-565 of albumin, 0-189 of lime, 0-033 of sodium phos-
phate, sodium chloride, calcium sulphate, and 0-034 of calcium phosphate !.

2. The blood of the large shell-snail (Helix pomatia) was found by
Harless and von Bibra2 to contain S'393 p. c. of organic and 6'12 p. c. of
mineral matters, there being 0'055 of oxide of copper in the latter.

This blood acquired a blue colour on exposure to air which disappeared
under the influence of C02. Alcohol precipitated a colourless coagulum and
ammonia removed the blue colour, which reappeared on neutralizing the
solution with hydrochloric acid.

Harless and von Bibra stated that the blood of Helix pomatia
contained copper, but no iron, but v. Gorup-Besanez states that on having
these observations repeated under his direction, in addition to copper, iron
was also found in the ash3.

3. Harless and von Bibra also investigated the blood of certain Cepha-
lopods (Loligo and Eledone) and Ascidians, which they likewise found to
contain copper but no iron. They assert that this blood possesses altogether
opposite colour properties to that of Helix pomatia, i.e. that it is blue
when free from oxygen but becomes colourless when shaken with air, again
being bleached when oxygen is passed through it. v. Gorup-Besanez con-
siders that this statement requires further proof before it can be accepted4.

4. The whitish-blue blood of Limulus Cyclops was examined by A.
Genth5. A few seconds after this blood is shed a yellowish-white coagulum
separates from the liquid, which retains its blue colour. The latter is
destroyed by boiling and by putrefaction. Genth analysed the ash of the
blood of this creature and found it to contain in one case 0-081 p. c. of
oxide of iron, and 0'085 of cupric oxide; in another case only a trace of
iron, but 0*297 p.c. of metallic copper.

1 C. Schmidt : see Lehmann's Physiological Chemistry, Vol. in., p. 256.

2 Harless und von Bibra, Miiller's Archiv, 1847, pp. 148—157. " Ueber das blaue
Blut einiger wirbellosen Thiere und dessen Kupfergehalt."

3 Gorup-Besanez, Lehrbuch der physiologischen Chemie, p. 369.

4 Gorup-Besanez, Op. cit. p. 370.

6 " Ueber die Aschenbestandtheile des Blutes von Limulus Cyclops." Ann. d. Chem.
'«-- Ptiarm., LXXXI. (1852), p. 68.

CHAP. II.] THE BLOOD. 133

The Blue Blood of the Octopus. Haemocyanin.

However interesting the above facts may have been as
rendering it most probable that the blue colouring matter of the
blood of certain of the Mollusca is concerned in the function of
respiration, and suggestive of the possibility that other metals may
take the place of iron as constituents of the blood-colouring matter,
they are infinitely less important than the observations of Frederique
made upon the blood of the Octopus.

Kabuteau and Papillon1 had described the blood of the Octopus,
and had correctly pointed out that it becomes blue on exposure to air,
doubtless in consequence of the action of oxygen. Their researches
have been continued by Le*on Frederique 2 with the following most
interesting results : —

The blood of the Octopus has a specific gravity of 1047, and it
contains between 13 and 14 per cent, of solid matters. The blood
contained in the vessels going to the branchiae is colourless, whilst
the blood leaving them is of a deep blue colour. If a large artery
be exposed in a living octopus, whilst it is immersed in water, and
breathing freely, it will be seen to have a deep blue colour, due
to a substance dissolved in the plasma ; if the animal be now with-
drawn from the water, as the respiration becomes impaired, the colour of
the artery is seen to become lighter and lighter, its contents becoming
ultimately colourless.

The blue blood drawn from an artery, if placed in a closed vessel,
undergoes, after some hours, a process of bleaching, the change of
colour being analogous to the change of the tint of arterial blood
when it is similarly treated. When the blue blood is boiled in the
receiver of a mercurial pump the blue colour disappears. The same
result follows when it is subjected to a stream of H2S or C02.

The blue colour is due to a body to which Frederique has given
the name of HAEMOCYANIN.

This body, like haemoglobin, is allied to the proteids, but still
more complex, seeing that it yields a proteid substance as one of its
decomposition products, but in addition a colouring matter. In the
case of haemocyanin this colouring matter is blue, and contains
copper. Following the analogy of haemoglobin the blue compound
might be termed oxy-haemocyanin, and the colourless derivative
simply haemocyanin.

Solutions of oxy -haemocyanin when examined with the spectro-
scope do not present any 'definite absorption-bands. Solutions of
the body, when heated, exhibit slight opalescence at 65° C., and this

1 Kabuteau et Papillon, "Observations sur quelques liquides, &c." Comptes
Rendus, v. 77, (14 Juillet, 1873) p. 137.

2 L6on Frederique, " Sur 1'organisation et la physiologie du Poulpe." Extrait des
Bulletins de V Academic Roy ale de BeJgique. 2me serie, T. XLVI. N° 11 ; 1878.

HAEMOCYANIN. PERIVISCEEAL FLUID.

[BOOK i.

increases to 73° C. ; coagulation actually occurs at 74° C. They are
likewise coagulated by alcohol, ether, mineral acids, and glacial acetic
acid ; and give the general reactions of the proteids.

Haemocyanin is a colloid, non-crystallizable body ; in addition to
it there appears to be no proteid or proteid derivative in the blood.
When decomposed with mineral acids it yields a prismatic crystalline
body.

It was said that the blue colouring matter of the blood of the
Octopus was contained in the liquor sanguinis. The blood does
contain a small number of corpuscles, but these are colourless.

The following table, extracted from Frederique's memoir, exhibits the
results of the quantitative analyses hitherto made of the blood of
Cephalopoda.

Paul

Le"on

Earless

Bert

Schlossberger

Frederique

Eledone

Sepia

Sepia Octopus

Octopus

Solid matters in 100 parts

7-23

10-9

18—20

12-6

13-689

Salts ....

2-63

3-205

2-225

3-014

„ soluble .

1-975

2-7918

1-940

2-33

„ insoluble

0-655

0-414

0-284

0-684

Organic matters

4-6

j?

10-375

10-675

Proteids

»

3-4

5J

8-9

On certain coloured corpuscles found in the Perivisceral Fluid of
certain Sea-urchins and Holothurians.

The perivisceral fluid of Sea-urchins and Holothurians has a more or less
distinct reddish tinge, which is due to the admixture of a considerable
proportion of coloured corpuscles1. These are large nucleated amoeboid
cells, of which the fluid endosarc is filled with small highly refracting
spherules of a rich mahogany-brown colour. They abound in the water-
vascular system and in the intestinal blood-vessels of the Urchin, and
are also to be found scattered throughout all the tissues, more particularlv
the integument. The following observations have been made by Mr
Patrick Geddes and have been kindly communicated by him to the Author.

If an Urchin be divested of its spines and left exposed to the air,
its warm hue soon becomes dingy, and, in the course of a few hours

1 For descriptions and figures of these corpuscles see —

Erdl Wiegmann's Archiv, 1842.

Williams, On the Blood-proper and Chylaqueous Fluid of Invertebrate Animals.
Philosophical Transactions, 1852. Part n. p. 595.

Semper, Eeisen im Archipel der Philippinen, Bd. i. Taf. xxxui.

Hoffman, Niederl. Archiv, 1871.

Geddes, " Observations sur le fluide pe'riviscerale des Oursins." Archives de
Zoologie experimental, 1878.

CHAP. II.] THE BLOOD. 135

changes into a peculiar dark green. When a quantity of perivisceral fluid
containing corpuscles in the dingy brown state is placed in the vacuum
of the mercurial gas pump, it rapidly recovers its normal colour. Thus the
colouring matter of these corpuscles is readily oxidised and deoxidised, and
there is considerable probability that it may have a respiratory function.
However, on account of the small number of brown corpuscles in the fluids
of the Urchin, it is impossible to make satisfactory analyses of the evolved
gases by means of the blood pump, nor has any attempt to isolate the
pigment yet succeeded.

That this brown substance is nearly related to the purple colouring
matter of the shell of many urchins, as well as to the yellowish-brown
(biliary 1) pigment of the intestinal epithelium, is made evident by adding a
mineral acid to their alcoholic solutions. All three immediately assume a
green tint, very similar to that of the integument of the dead Urchin.
Moreover, when a morsel of any of the highly pigmented tissues of
Spatangus purpureus, for instance, the ovary, is slightly torn with needles,
purple spots appear at the injured points, and, under the microscope,
the brown corpuscles may be watched, one by one changing into purple.

Lemon-yellow amoeboid corpuscles are also found, though sparingly,
in the fluids of certain of the regular Sea-urchins (Dorocidaris, Arbocia), and
are exceedingly abundant in the perivisceral fluid of the Spatangoidea.

The greatest variety of colour is to be seen in the contents of the intes-
tinal vessels of /Spatangus, in a single preparation of which may be seen
brown, purple, green, lemon-yellow, and indigo-blue amoeboid corpuscles,
together with vast numbers of peculiar greyish vesicles of very variable size,
from that of a micrococcus up to more than that of a coloured corpuscle.

rjris

CHAPTEB III.

CHANGES WHICH THE BLOOD UNDERGOES IN
DISEASE.

INTRODUCTION.

THE blood may be looked upon as the internal medium whither
tends the stream of matter which flows from the external world
into the organism, and whence simpler combinations of matter,
which are the result of the chemical processes of the organism, leave
it to form again a part of the external medium. The blood
represents a common reservoir which is continually being drawn
upon by each tissue and organ for the materials which it needs, and
to which, in its turn, each tissue and organ contributes its quota of
useful manufactured products or of useless waste.

If we except the coloured corpuscles, whose function it is to act as
the internal oxygen-carriers of the body, and the colourless corpuscles,
which we have good reason to think are the precursors of the
coloured, the blood represents a solution of organic and inorganic
matters, which is continually being added to and taken from, in
different ways and degrees, by the different tissues and organs, and at
varying rates by each tissue or organ according to the degree of
its functional activity.

The ancients looked upon the blood as essentially representing
vitality : as that part of the matter of the body in which specially
resided the life, and hence arose the natural wish to connect all the
morbid processes of the body, processes tending towards death, with a
perversion of the life-giving or actually vital liquid — a wish which
found expression in the various phases of the humoral pathology
which under one form or another reigned more or less imperiously
over medicine until the fifth decade of the present century had
passed.

If, however, we look upon the blood very much as a fluid con-
tained in a reservoir which is contributed to by many sources, and
whence at many points, by a variety of chemical and physical
processes, matter is being continuously removed, we shall, naturally, be
forced to admit that any changes which the blood undergoes are, in all

CHAP. III.] THE BLOOD IN DISEASE. 137

probability, nearly always dependent upon some modification of the
organs which intervene between the external world and itself, of the
organs through which certain of its materials have to pass in order to
reach it, or of the organs through which other of its materials have to
pass before they can be eliminated.

The progress of biological research has tended more and more to
confirm this view of the relation of the blood to the organs of the
body, and to transfer the vital processes to those elements of the
various organs which we term cells, modified though these may be
from the ideal conception of the cell in its primordial condition, as
represented, for instance, by the mammalian ovum, or the cells of
embryonal connective tissue.

Apparently, it is in connection with those extra- vascular centres of
nutrition, the cells, that take place those chemical processes (nearly
all of which are associated with oxidation) which result either in
the assimilation of fresh matter for the body's use, or of elimina-
tion of waste matter which would accumulate to the body's detri-
ment, or which primarily have for their object the evolution of
the kinetic energy which the body needs, in order that it shall
perform its internal and external work ; so that the life of an
organ, as evidenced by its ability to perform those acts which
characterize it as alive, may be philosophically considered as the sum
of the life of its constituent living centres, the cells, and the life of the
organism as, in one sense, the sum of the life of all the constituent
living centres of its various organs.

If this view be correct, disease will, in all probability, depend
primarily upon modifications in the processes of cells, rather than of
the fluid whence cells obtain their nourishment, and we shall be quite
prepared to find (1) that a morbid process may seriously interfere
with organs whose functions are essential to life, without influencing
the composition of the blood in a manner perceptible by our methods
of chemical and microscopic analysis, however delicate these may be,
and (2) that when a marked change is revealed by these methods of
enquiry it must be a difficult matter to trace the component causes of
which the change is the resultant effect. The first proposition is
proved by the paucity of results which have been obtained in spite of
the assiduous labours of many scientific physicians, the second may be
well illustrated by taking as an example that change in the blood which
is better characterized than all others, viz. anaemia, or that condition
in which the relative and absolute number of the coloured corpuscles
of the blood is diminished.

It is a condition which may result from accidental losses of blood,
or from some process (for example, abundant suppuration) which tends
unnaturally to drain the blood of some of its constituents, or from a
deficiency of proper food, or from causes so complex that we willingly
hide our ignorance under the expression of disorders of nutrition.
Where the actual fault primarily lies can, in many cases, be not even
guessed at, and the physician knows little more than that the disorder

138 A HUMORAL PATHOLOGY NO LONGER TENABLE. [BOOK I.

of nutrition is one which is often successfully overcome by the
administration of iron, by fresh air, and an abundant diet.

Eesearches on the chemical changes which take place in the blood
in disease were not possible until the chief proximate constituents of
the blood had been studied, and methods devised for their repara-
tion. Amongst the researches which proved of the greatest value in
this respect were those of Berzelius1, of The'nard and then of
MM. Prevost and Dumas 2, which, by determining the mean composi-
tion of healthy human blood, first established a standard of comparison
which might be referred to by those studying the changes induced in
the blood by disease. Amongst the most complete of the systematic
investigations which were made in the latter subject were those of
Andral and Gavarret3, of Becquerel and Rodier4, of Simon5, whilst
the changes in particular diseases engaged the attention of certain
distinguished writers, as of Christison6, of Garrod7, of C. Schmidt8.

During the last thirty years comparatively little attention has
been paid to the condition of the blood in various diseases, a fact
which may be explained partly as due to the discontinuance of the
practice of venesection, which has deprived the physician of the
material required for these investigations, partly as a result of
the change of views which has beeaa explained at the commencement
of this section.

Of late, however, attention has again been enthusiastically directed
to the modifications which certain constituents of the blood undergo
in disease, notably to the variation in the relative number of coloured
and colourless corpuscles, and of the haemoglobin contained in the
former, and we may therefore expect rapid accessions to the exact
knowledge which we possess.

We shall in the first place consider categorically the changes
which the various normal constituents of the blood undergo in disease,
and then draw special attention to the results of investigations of the
changes in particular diseases.

1 Berzelius : see " General views of the Composition of Animal Fluids." Transac-
tions of Med.-Chir. Soc. of London, Vol. in. p. 198.

2 Pre" vost et Dumas, " Examen du sang et de son action dans les divers ph&iomenes
de la vie." Ann. de Chimie, 1821, T. xvin., p. 280. A second memoir, with the same
title, was published in the Annales de Chimie et de Physique, 1823, Vol. xxni. p. 50
and p. 90.

3 Andral et Gavarret, " Eecherches sur les modifications de proportion de quelques
principes du sang (fibrine, globules, mat6riaux solides du serum et eau) dans les mala-
dies." Annales de Chimie et de Physique, Tome LXXV., p. 225 — " Eecherches sur la com-
position du sang de quelques animaux domestiques dans 1'etat de saiit<£ et de maladie,'
(in conjunction with M. Delafond). Annales de Chimie et de Physique, 3me s6rie, Vol.
v. p. 304. Andral, Essai d'Hematologie Pathologique, Paris, 1843.

4 Becquerel et Kodier, Recherches sur les alterations du sang. Paris, 1844. — Traite
de Chimie Pathologique appliquee a la Medecine Pratique. Paris, 1854.

6 Simon, Animal Chemistry, translated by G. E. Day, M.D. Sydenham Society, 1845.

6 Christison, " On granular degeneration of the kidneys, and its connexion with
dropsy, inflammations and other diseases. 8vo. Edinburgh, Adam and Charles Black,
1839. 7 Garrod : see page 143.

8 C. Schmidt, Charalcteristik der epidemischen Cholera gegenuber Transudations-
anomalien. Leipzig u. Mitau, 1850.

CHAP. III.] THE BLOOD IN DISEASE. 139

SECT. 1. Variations in the proportion of the principal Constituents of
the Blood in Diseases in general.

Water I. Before considering the changes which the blood

undergoes in different diseases, it is well to insist upon
the fact that loss of blood very rapidly influences the composition of that
which remains in the vascular system. It has been shewn by the con-
cordant results of many trustworthy observers1 that when an animal is
bled, the portion of blood first obtained contains the largest quantity of
solid matter, and that this gradually diminishes, so that the blood obtained
at the commencement of a venesection has a slightly, but still perceptibly,
different composition from that obtained at its termination, unless, of
course, the total quantity of blood withdrawn be excessively small.

This diminution in the solid matter of the blood which is noticeable
even in the course of venesection is naturally much more perceptible in
cases of excessive and repeated accidental haemorrhages. The diminution
of solid matter is partly due to actual loss of solids, but in great part to
the blood becoming more rapidly diluted by lymph than in the normal
condition.

The normal quantity of water in the blood of man may be estimated
as varying between 780 and 800 parts per 1000 of blood. An increase
in the water of the blood is much more frequent than the converse ; this
increase may be only slight or it may be considerable.

A slight augmentation of the water of the blood, i. e. to between 800 and
820 parts per 1000, occurs as a result of a temporary abstinence from food,
in the early stages of nearly all acute diseases, and in the majority of
chronic diseases.

A more marked augmentation, the water amounting to between 820
and 880 parts per 1000 of blood, occurs in starvation: after considerable
haemorrhages ; in cases of abundant suppuration, or in which some other
considerable drain is taking place, as in chronic diarrhoea ; in the course
of malarial diseases ; in lead poisoning ; in chronic mercurial poisoning ;
in cancerous and tubercular affections : and we might add in anaemia,
if it were not more correct to characterize the latter as the condition which
really exists in all the morbid states just enumerated.

A decrease in the quantity of water of the blood has been observed in
articular rheumatism, in erysipelas, in puerperal fever, and especially in
cholera.

Coloured II. The coloured corpuscles are increased in the first

Corpuscles stages of cholera ; the increase is however riot an absolute

and Haemo- One, but merely dependent on a diminution of the water
globin. 0£ tjie blood. A diminution of the coloured corpuscles occurs

in the various forms of anaemia, including chlorosis ; in Bright's disease ;
as a result of prolonged diarrhoea and dysentery ; of continued and abundant
suppurative discharges; in scurvy; in leucocy thaemia ; in the advanced
stages of continued and of intermittent fevers; in chronic metallic poisoning ;
in cases of advanced heart disease; in chronic diseases generally.

1 Pre'vost and Dumas, Becquerel and Eoclier, Simon, and others.

140 PROPORTION OF HAEMOGLOBIN IN VARIOUS DISEASES. [BOOK I.

In health, the amount of haemoglobin in the blood appears to be pro-
portional to the number of corpuscles. This relation does not hold,
however, in disease, as will be particularly mentioned in discussing the.
phenomena of anaemia.

The largest number of determinations of the amount of haemoglobin
in the blood of various diseases has been carried out by Quincke1, who
made use of Preyer's method for the determination of haemoglobin. In
the annexed table may be seen the results which he obtained. The letters
V-S. in the second column indicate that the blood was obtained by vene-
section, and the letter H that it was obtained by Heurteloup's artificial leech.

Sex and
Age.

Method by
which
blood
obtained.

Specific
gravity.

Haemo-
globin in
100

grammes.

Disease.

)
Observations.

F. 35

VS.

1058

14-4

Angina pecto-

Otherwise healthy, well

ris.

nourished woman.

F. 60—

vs.

1060-6

14-1

Cerebral Apo-

Previously healthy and

70

plexy.

well nourished, V-S. two

hours after the attack.

M. 44

H.

1060-8

14-6

Scorbutus.

Purpura haemorrhagica on

lower extremities which

quickly disappeared by

rest in bed ; state of nu-

trition good.

M. 20

H.

1049-6

10-1

Cirrhosis of the

Pretty intense jaundice.

liver; haemo-

Frequent epistaxis, pro-

philia.

fuse bleeding from any

accidental wound.

F. 15

H.

1035-2

5-3

Chlorosis.

Well developed body ; no

complication. Date Nov.

14, 1869.

H-,

1049-1

9-92

j>

10 weeks later. Has been i

taking iron. Date Feb.!

3, 1870.

M. 45

H.

1044-3

5-80

Splenic leuco-

cythaemia.

F. 28

VS.

1050-5

10-30

Parenchyma-

Patient died of acute j

tons Nephritis.

oedema of the lungs.

M. 40

vs.

1047-3

10-70

Nephritis

Considerable general oe-

Uraemia.

dema. The patient died i

a few hours after.

M. 27

vs.

1048-7

11*40

Nephritis

Considerable oedema. Con-

Uraemia,

stitutional syphilis. V-S.

during a uraemic convul-

sion. Sp. gr. of the se-

rum, 1044.

1 Quincke : "Ueber den Hamoglobingehalt des Blutes in Krankheiten." Virchow's
Archiv, Vol. LIV. (1872), p. 537,

CHAP. III.]

THE BLOOD IN DISEASE.

141

Sex and
Age.

Method bj
which
blood
obtained.

1

Specific
gravity.

Haemo-
globin in
100
grammes

Disease.

Observations.

M. 43

H.

1047-0

10-60

Bright's diseas

Considerable oedema. Very

contracted

abundant urine of light

stage.

colour and low specific

i

gravity.

M. 24

H.

1041-1

8-5

Bright's diseas

Very considerable oedema.

contracted

Chronic uraemia. Post-

stage.

mortem examination re-

i

vealed highly contracted

kidneys.

M.

H.

1054-9

14-4

Diabetes Me

Appetite still very good.

litus.

Total quantity of urine in

24 hours, 10 litres. Sp.

gr. 1030.

M. 30

H.

1059-5

15-9

Diabetes Me'

Enormously fat person.

litus1.

Good appetite. Urine in

24 hours from 3 to 4

litres. Sp. gr. 1020.

M. 22

H.

1056-6

12-9

Typhoid fever

A somewhat cachectic

1st week.

individual.

M. 25

H.

1059-6

12-7

Typhoid fever

Moderately strong man.

1st week.

M. 25

H.

1062-1

14-6

Typhoid fever,

Moderately strong man,

1st week.

an attack of medium se-

verity.

H.

1054-4

12-6

typhoid fever.

4th week.

M.

H.

1056-4

144

Relapsing

Strong man.

ever, 5th day.

F. 50

VS.

1057-9

15-0

Cerebro-spinal

A strong person. Ap-

meningitis of

parently has been ill three

great acute-

days. Deepest coma.

ness.

Death on the 5th day.

M. 56

H.

1050-5

11-3

Pyaemia, 2nd

following a phlegmonous

or 3rd week.

abscess of the neck, there

occurred phlebitis of the

F. 20

VS.

1056-7

14-9

Phosphorus

jugular vein and pyaemia.
Patient had four days before

poisoning.

swallowed an infusion of

lucifer matches. Intense

icterus, enlargement and

tenderness of the liver.

)eath 12 hours after vene-

section.

1 This was probably a case which should have been termed glycosuria, rather than
diabetes mellitus. Sugar not unfrequently occurs in the urine of very obese persona who
present none of the other symptoms of diabetes. This statement the author makes upon the
authority of a verbal communication from Dr Lauder Brunton.

142 PROPORTION OF BLOOD CONSTITUENTS IN DISEASE. [BOOK I.

III. The quantity of fibrin which separates from the
blood during coagulation, and which normally amounts in
the case of man to about 2-5 parts per 1000, may increase in disease to as
much as 10 parts per 1000. This increase of fibrin is to a certain extent
characteristic of acute inflammatory affections; it is clearly not to be
ascribed to the pyrexia which is often a prominent feature of these diseases,
seeing that in the fevers the proportion of fibrin is diminished instead
of being increased.

According to Becquerel and Rodier the cases in which the amount of
fibrin is increased may be divided into two categories. In the first
category the augmentation is only slight, the amount of fibrin fluctuating
between 3 and 5 per 1000 of blood. In the second it is considerable and
is comprised between 5 and 10 parts per 1000 of blood.

A. Slight augmentation of fibrin occurs (1) in chlorosis; (2) in
pregnancy, especially towards the close of utero-gestation ; (3) in slight
inflammatory affections, if accompanied by pyrexia ; such as slight attack
of erysipelas of the face &c. ; (4) in certain cases of scorbutus.

B. Considerable augmentation of fibrin (amount varying between
5 and 10 per 1000 of blood) is characteristic of the more serious inflammatory
affections. It is most marked for instance in pneumonia, pleurisy, and
acute rheumatism. Whenever the parenchyma of organs is implicated in
the inflammatory process the fibrin of the blood appears to increase.
Whence comes the increase 1 Seeing that we are yet in ignorance as to
the origin of the fibrinogen of the blood plasma, a solution of the above
question is impossible. In the proliferation of cellular elements which
accompanies the process of inflammation we have however a cause which
will add to the number of colourless cells of the blood, and to the amount
of serum-globulin which will be available in the process of coagulation.
Whether we admit or deny Schmidt's theory there is no question as to the
influence which serum-globulin exerts in increasing the amount of fibrin,
and this is one way (though only one) in which we may conceive that
inflammatory diseases cause the proportion of fibrin to increase.

A diminution in the proportion of fibrin (so that it sinks to between
1 and 2 parts per 1000 of blood) has been observed in certain acute and
certain chronic diseases. Amongst the former are to be reckoned typhoid
fever, small pox, scarlet fever and measles ; amongst the latter, organic
affections of the heart in their last stage, certain malarial cachexiae,
chronic scurvy, and chronic mercurial poisoning.

Serum-Al- ' normal amount of serum-albumin in the

bumin serum of the blood of man amounts on an average to 80

parts per 1000, the limits varying between the numbers
70 and 90.

An augmentation of serum-albumin has been observed to occur in
cholera and after the use of hydragogue cathartics1. To a less extent in
acute rheumatism and in the early stages of some intermittents (?).

A diminution of serum-albumin occurs most markedly in Bright's
disease, anaemia, scurvy, dysentery, and generally in chronic diseases
which impair the general nutrition : for instance, in the advanced stages
of some cardiac affections.

1 C. Schmidt, Characteristic dcr Cholera.

CHAP. IIT.] THE BLOOD IN DISEASE. 143

Fatg V. The nor ma amount of fatty matters in healthy blood

varies, according to Becquerel and Rodier, between 1 and
3-3 parts in 1000. It is said that the fats of the blood are increased in
pneumonia, in alcoholism, in diabetes, in Bright's disease, in the hepatitis
of hot climates, in cases of chylous urine, in some cases of acute rheumatism,
and in many acute and chronic cases of poisoning1. The information on
many of these points is in the highest degree unsatisfactory.

VI. The amount of cholesterin in normal blood varies

and L°ecUh£n probably between 0-5 and 2-0 parts per 1000. According to

Becquerel and Rodier this constituent increases in quantity

in all acute febrile affections, in all acute inflammations, and especially in

cases of jaundice in which there is almost complete retention of bile.

We possess no information whatever as to the amount of lecithin
present in the blood in disease ; indeed our knowledge of the proportions
present in health only rest on a very few analyses by Judell and Hoppe-
Seyler.

Sugar. VII. Sugar is increased in the blood of diabetes, as will

be mentioned under that disease.

Urea, Uric VIII. Amongst the so-called extractive matters present

acidandotner jn the blood, urea, uric acid, and hypoxanthine require to be
mentioned as being affected in disease.

The amount of urea in the blood is largely increased in the various
forms of Bright's disease2, as was first shewn by Christison, in cholera3, and
in yellow fever. It has been said that this is the case also in diabetes and
febrile affections4.

Uric acid5, as will be more particularly referred to under Gout, is
markedly increased in the blood in acute and chronic cases of that disease.

Hypoxanthine has been found in considerable quantities in the blood
of leucocythaemia3; according to Salomon this body is a constituent of
healthy blood.

Saltg IX. The salts of the Hood, especially the alkaline salts,

undergo certain changes in disease, though our knowldge is
yet very imperfect on this matter. In cholera, the serum of blood, though
it contains less salts than normal, contains a larger quantity of salts of
potassium; in dysentery, the salts of the serum are said to be increased,
and the same holds in the case of Bright's disease.

Tne Gases ^* ^s ye* ^ew ^ac*s have been collected which throw

of the Blood. an7 light upon the proportion of the gases in the blood

in disease. From a knowledge of the changes which other

constituents undergo in certain diseases, or from a knowledge of the

1 Gautier, Chimie appliqute a la physiologic, a la pathologic et a Vhygiene. Vol. u.
p. 314.

2 Christison, On granular degeneration of the kidneys, &c. Edinburgh, 1839.

3 Scherer, Verh. d. physik.-mcd. Ges. zu Wiirzburg, Vol. n. pp. 321—325, and Vol.
VIT. pp. 123—126.

Picard, These de Strasbourg, 1856.

5 Garrod, A Treatise on Gout and Rheumatic Gout. Third ed., 1876, p. 84 et seq.
The first researches of this author on this subject were published in the Medico-
Chirurgical Transactions, Vol. xxxvu.

144 THE GASES OF THE BLOOD IN DISEASE. [BOOK I.

physical conditions of the patients, we can often surmise the way in which
the gaseous exchanges of the blood must be affected. Thus from the
amount of haemoglobin found in cases of anaemia and chlorosis, we can,
with considerable accuracy, calculate the maximum amount of oxygen
which such blood can contain, and we arrive at the conclusion that the
amount is much below the normal.

Thus a healthy man's blood contains on an average say 13*5 grammes
of haemoglobin in one hundred parts. Such blood in virtue of its haemo-
globin would, if saturated with oxygen, be capable of absorbing 22-55 c.c.
of oxygen measured at 0° C. and 760 mm. pressure.

On the other hand the blood in cases of chlorosis may contain as little
as 5 '3 grammes of haemoglobin per 100 of blood. Such blood could in
virtue of its haemoglobin only take up 8 '85 c.c. of oxygen if fully
saturated. We see therefore that the respiratory capacity of such blood
is reduced to less than one half that of healthy blood.

Again in cases where mechanical causes exist which interfere with the
due amount of the gaseous exchanges in the lung, the cyanosis and the
dyspnoea, sometimes culminating in asphyxia, point to a condition in which
the oxygen of the blood is greatly diminished and the carbonic acid greatly
increased. Actual determinations are, however, almost entirely wanting1.

Attempts have been made by certain observers to determine the
changes which the gases of the blood undergo in disease. Unfortunately
the methods employed have been such as to deprive the results of all
value. Thus Quinquaud determined the amount of oxygen in the blood
of various diseases by means of a standard solution of sodium hydrosulphite2.
The results obtained by this method are unfortunately in no way
comparable with those obtained by the mercurial pump. Again, Brouardel3
has published analyses of the gases of the blood in variola and scarlatina
which would appear to shew that in these diseases the proportion of oxygen
which the blood can absorb is very much diminished. As, however, the
amount of nitrogen found is much greater than could possibly have been
held in solution by the quantity of blood analysed, the legitimate conclusion
to be drawn is that the analyses possess no value. Regnard4 has attempted
to determine the so-called ' respiratory capacity' of blood in disease, i. e.
the maximum amount of oxygen which a given quantity of blood will
absorb. Blood is shaken with air and then subjected to analysis in the
mercurial pump. According to Regnard the respiratory capacity of blood
is not affected even by putrefaction ; i. e. blood which is decomposed can
absorb as much oxygen, as it did before the process of putrefaction
set in. Regnard's observations were all performed with blood taken
from the dead body, the clot being broken up artificially. They led
to the conclusion that in many diseases the respiratory capacity is immensely
diminished ; were the results reliable they would indicate that under the

1 In a case of cyanosis due to a cardiac lesion Lupine found that 100 c.c. of
venous blood contained 64 c.c. of C02. Gazette Medic, de Paris, 1873, p. 128.

2 Quinquaud, "Sur un proc6d6 de dosage de 1'he'moglobine dans le sang"
Comptes Rendus, Vol. LXXVI. p. 1489. " Sur les variations de 1'h^moglobine dans les
maladies. " Comptes Eendus, Vol. LXXVII. p. 447.

3 Brouardel: "Des gaz du sang dans diffe'rentes maladies." Societe medicale des
hopitaux, 1870, quoted by Bernard.

4 P. Bernard: Eecherches experimental sur les variations pathologiques des combus-
tions respiratoires. These pour le Doctorat en Mddecine. Paris, 1878 109 et seq.

CHAP. Ill ] 'THE BLOOD IN DISEASE. 145

influence of morbid processes the power which haemoglobin possesses of
linking oxygen to itself is more or less affected. The conditions under
which these observations were made appear, however, to the author, to
deprive them of any value whatever.

Legerot1 produced septicaemia in dogs by the injection of putrefied blood
and compared the respiratory capacity before and after the induction of
the morbid state. His results would appear to shew that an enormous
diminution (sometimes to more than one half) of the respiratory capacity
occurs.

SECT. 2. THE CHANGES WHICH THE BLOOD UNDERGOES IN
PARTICULAR DISEASES.

In the preceding section we have grouped together under each
principal constituent or group of constituents of the blood, the varia-
tions which have been observed in diseases generally.

We must now consider in detail the changes of the several
chief constituents of the blood in certain special diseases, which
have been particularly studied from this point of view.

A. THE BLOOD IN DISORDERS OF NUTRITION.

Anaemia.

It has long been known that in various forms of anaemia the
coloured corpuscles of the blood undergo a diminution, which to
a certain extent appears to be proportionate to the intensity of the
disease. The observations of the earlier French writers on this
subject were definite enough, and although made by methods which
did not furnish an absolutely correct estimate of the weight of the
dry corpuscles, and gave no indication of the weight of the moist
corpuscles, yielded results which might be compared one with the
other. Thus Becquerel and Rodier2 classified cases in which the
coloured corpuscles of the blood are deficient, into the three following
classes, each distinguished by a separate letter. We give, in the
first instance, their account, but slightly abridged.

Bacquerei Qass A. Slight diminution (weight of dry corpuscles
and Rodiers b t 10Q d 12Q 1QOO of blood)< Individuals
classification , , , . ,,. , ,, '. r ri
of cases of belonging to this class are pallid, there is some feeble-
Anaemia, ness ; sometimes, but by no means always, a blowing
murmur is heard with the first sound at the base, and a murmur in
the carotids.

1 Legerot, Etudes dhematologie pathologique lasees sur V extraction des gaz du sang.
Paris, 1874, quoted by Rdgnard, Op. cit. p. 121.

2 Bocquerel et Rodier, Traits de Chimie pathologique. Paris, 1854, p. 50 et seq.

G. > 10

146 THE BLOOD COKPUSCLES IN ANAEMIA. [BOOK I.

This degree of diminution of the coloured corpuscles occurs under
the following circumstances; — in feeble individuals of the so-called
lymphatic diathesis : under the influence of insufficient diet : in
persons inhabiting marshy districts : as the result of a copious blood-
letting : as a result of the persistent use of purgatives : in chronic
Bright's disease : after some days of an acute disease, such as a fever :
in the course of many chronic diseases, &c.

Class B. Medium diminution (weight of dry blood-corpuscles
between 80 and 100 per 1000 of blood).

This state of the blood is accompanied by a much more marked
debility of those subject to it. The skin is pale and slightly
yellowish. Bodily exertion is irksome. There exists palpitation,
and some dyspnoea may come on if the patient takes exercise.
There is a soft bellows murmur in the aorta and carotids, which is
rarely propagated along other arteries.

The causes enumerated under Class A may, if continuing in
operation, lead to the case being classed under B. As special
causes are to be mentioned ; — considerable losses of blood : continued
diarrhoea (or dysentery) : malarial cachexia : the cancerous cachexia :
lead poisoning: chronic Bright's disease : the last stage of chronic
diseases : lastly, and chiefly, chlorosis.

Class C. Great diminution (weight of dry blood-corpuscles varying
between 40 and 80 per 1000 of blood).

Cases belonging to this class are much rarer than the preceding.
The skin is pale, and may present a greenish hue, the strength is
diminished; sometimes the least movement occasions sensations of
painful weariness, muscular pains, dyspnoea and palpitation. Cephal-
algia, vertigo, tinnitus aurium and other nervous troubles appear,
in varying degrees of intensity. Syncope is readily induced; the
pulse is quick and dicrotic ; there is a murmur with the first sound
of the heart at the base. A very loud bellows murmur is heard in
the carotids, and usually there exists, especially in chlorosis, a more
or less loud venous murmur (bruit de diable).

The following causes specially lead to the condition observed in
this class; — great or frequently repeated hemorrhages: chlorosis:
malarial anaemia: the cancerous cachexia, especially where cancer
affects the stomach.

If to the causes producing a diminution of the blood-corpuscles
enumerated by Becquerel and Rodier, we add abundant and long-
continued suppuration, scurvy, leucocythaemia and the affection
designated by the term 'pernicious anaemia' we shall have before
us a pretty complete catalogue of the various morbid states con-
nected with a diminution of the blood-corpuscles.

As will be appreciated by the reader of the section in which the
determination of the weight of the coloured corpuscles is described,
the weight of the dry corpuscles as found by such a method as that
employed by Becquerel and Rodier does not admit of absolute accuracy.
For the purposes of the physician it would be better if we could express the

CHAP. III.] THE BLOOD IN DISEASE. 147

variation which the weight of the moist corpuscles undergoes in respect to
the weight of the liquor sanguinis, in various diseases. The methods
which we possess for effecting this determination with accuracy are,
however, so complex and so difficult, that no large collection of data
directly obtained by this method exists. We can, however, as was shewn
by the researches of C. Schmidt, obtain a very close approximation to the
true weight of the moist blood-corpuscles present in the blood, if we
multiply the results obtained by Prevost and Dumas' method (which
was employed by Becquerel and Rodier in their researches) by 4.

Since, however, methods have been devised (1) for the enumeration of
the blood-corpuscles contained in a known volume of blood and (2) for the
determination of the amount of haemoglobin, the physician has been placed in
possession of methods which have thrown great light upon some of the
diseases in which the blood-corpuscles are diminished — in which typically
the condition of anaemia exists.

By means of any of the methods described at pages 74 — 78 a close
approximation to the number of corpuscles contained in the blood may be
made in a few minutes, by employing a single drop of blood. Similarly
by methods as ready and as accurate, the amount of haemoglobin in the
blood may be determined.

As we have seen, haemoglobin constitutes by far the most abundant
constituent of the red blood-corpuscles, and it might be supposed that the
second of the above determinations might be sufficient for the purposes of
the physician ; the richness or poverty of the blood in coloured corpuscles
being judged of by its richness or poverty in haemoglobin. Such is
however not the case, as will be now briefly shewn.

Changes It resulted from the labours of Welcker, the dis-

coverer of all the fundamental facts concerning the
corpuscles , , . . , . , . ., P . & ,

undergo in relative number, weight, cubic capacity, superficies and

Anaemia. colouration of the blood-corpuscles, that in the physio-

logical condition the colour of the blood is proportionate to the
number of its coloured corpuscles — in other words, that in the
healthy state the amount of haemoglobin contained in the red
blood-corpuscles is constant. That Welcker was correct in his statement,
in so far as the healthy state is concerned, has been proved by the recent
researches of Worm-Muller1, and is, on the whole, confirmed by Malas-
sez2. In his remarkable researches on the changes which the blood
undergoes in cholera and some other diseases, C. Schmidt3 had how-
ever pointed out that the composition of the blood-corpuscles is liable
to vary in disease, and attention was still more forcibly drawn to
this interesting fact by Johann Duncan in 1867 4. This observer

1 Worm-Muller, "Ueber das Verhaltniss zwischen der Zahl der Blutkorperchen
und der Farbekraft des Blutes." Om Forholdet imellem Blodlegemernes Antal og Blodets
Faroekraft. Christiania, 1876. Abstracted in Maly's Jahresbericht, Vol. vii. (1878),
p. 102. *

3 Malassez, " Sur les diverges methodes de dosage de 1'he'moglobine et sur un
nouveau colori metre." Archives de Physiologic, Ser. n., Vol. iv. (1877), pp. 1 — 13.

3 C. Schmidt, Charakteristik der epidemischen Cholera, &c.

4 Duncan, " Beitrage zur Pathologic und Therapie der Chlorose." Sitzungsber. d.
kais. Akad. d. Wissenschaften zu Wien. Naturwissenschaft. Cl. 1867. 2 Abth., pp.
516—522.

10—2

148 THE BLOOD COKPUSCLES IN ANAEMIA. [BOOK I.

counted the corpuscles contained in a given volume of blood in three
cases of chlorosis, and compared the colouring power of a given
volume of the same blood with the colouring power of the same
volume of healthy blood. From his observations he concluded that
whilst the coloured corpuscles were nearly as numerous in his
chlorotic patients as in healthy women, the amount of colouring
matter was remarkably diminished, being about three times less in
amount. The more recent researches of MM. Hayem1 and Ma-
lassez2, but especially of the former, have brought out the interest-
ing fact that in the various forms of anaemia the anatomical
characters of the red blood-corpuscles are affected, and that the normal
relations between the haemoglobin and the other constituents of the
corpuscles are disturbed. The following is an epitome of the state-
ments of Hayem.

The changes which occur in the anatomical characters of the
coloured corpuscles in anaemia are appreciated if we compare suc-
cessively the diseased with the healthy corpuscles ; paying attention
to size, number, form and colouration.

1. Size. In normal human blood we may, according to
Hayem, conveniently classify the corpuscles into three orders, accord-
ing to size, viz. large, medium, and small ; the large blood-cor-
puscles having a mean diameter of 8*5 //., the medium 7'5//,, and the
small 6'5/*. Usually the proportions in which these various corpuscles
are present is the following: the medium-sized amount to 75, the
large to about 12 and the small also to about 12 per 100, so that the
mean size of the average blood-corpuscle is (according to Hayem) 7'5/Lt.

If we except acute cases where the disease is suddenly produced
by hemorrhage, in all forms of anaemia the size of the corpuscles
is modified. Firstly, the blood contains a certain proportion of
unusually small coloured corpuscles, which have a diameter varying
between 2 '2//. and 6/i<. Almost as frequently, the blood contains a
certain number of unusually large corpuscles, which we may term giant-
corpuscles, measuring 10/i,or 12//, or even 14/x. Secondly, the rela-
tion between the corpuscles of different sizes is disturbed, so that the
blood contains a much larger number of small corpuscles in relation
to other sizes than healthy blood.

In all cases of chronic anaemia of considerable intensity, the mean
diameter of the corpuscles is always below the normal. It may fall
to 7fj>, to 6'8/ji, to 6'5/z,, and even to 6 ft.

But this diminution in the mean diameter corresponds to a
diminution in the mean volumes of the corpuscles.

Thus the normal blood-corpuscle, having a mean diameter of
7'ojj,, has approximately the volume of 66/^c.c. (cubic micro-milli-

1 Hayem, Jtecherclies sur VAnatomie normale et pathologique du sang. Avec figures
et tableaux. Paris, 1878. Here will be found reprinted the various papers on these
subjects, elsewhere published by this author.

2 Malassez, "Sur les di verses niethodes de dosage de Themoglobine et sur un
nouveau colorimetre. " Archives de Physiologic, Ser. n.t Vol. iv. (1877), pp. 1—43.

CHAP. III.] THE BLOOD IN DISEASE. 149

metres). The corpuscle whose diameter is only 7yn has the volume
of 57/z.c.c. ; that of 6'5yLt has a volume of 49/>tc.c.

When in anaemia the mean diameter of the blood-corpuscles falls
to 7/A, 100 corpuscles correspond in volume to only 80 healthy
corpuscles ; when the mean diameter falls to 6/4, 100 corpuscles
correspond only to 65 healthy corpuscles.

Even assuming that the proportion of haemoglobin remained
constant in anaemia, it is obvious from the above considerations,
that important consequences must result from the diminution in
the size of the corpuscles, but as will be shewn subsequently, the
proportion of haemoglobin does not remain normal.

2. Number. Usually the number of the coloured corpuscles is
diminished in anaemia, but by no means constantly so. In the most
intense cases of anaemia the diminution is however always very
marked. In a case of malarial anaemia Hay em found 1,182,750
corpuscles in 1 cubic millimetre, and in a case of purpura hemorrhagica
only 1,000,000, i.e. a diminution of the blood-corpuscles to between
Jth and <Uh the normal amount.

3. Form. The corpuscles of anaemic blood assume modifications
of shape which would seem to depend upon a diminished consistence,
and which affect chiefly the medium-sized or small corpuscles, which
often assume an elongated oval form.

4. Colour. The most important character which distinguishes
the blood of anaemia from healthy blood is a comparatively pale colour
of its red corpuscles, which depends directly upon a diminution in the
amount of haemoglobin. If by 1 we represent the maximum amour t
of haemoglobin, as measured by its colorific intensity, present in the
unit volume of healthy blood, we shall find that the amount may be
as low as 0'85 without health being sensibly impaired; indeed the
unit volume of blood of mean composition contains a quantity of
haemoglobin which fluctuates between 0'85 and 0'90, as compared
with 1 (the maximum amount of haemoglobin contained in healthy
blood). In anaemia, the amount of haemoglobin diminishes so much
that the amount present in the unit volume may be represented by
fractions varying between the limits 0'66 and 0'125, so that in the
most intense anaemia the quantity of haemoglobin may sink to one-
eighth its normal amount. In cases of anaemia of moderate intensity
the amount varies between one-half and one-fourth.

We have seen that most interesting conclusions may be drawn
from a consideration of the size alone of the corpuscles in anaemia;
still more interesting are, however, the results of simultaneous enume-
ration of the corpuscles and determination of their colorific intensity.

Thus if we find the number of corpuscles in a cubic millimetre of
blood to be 4 millions and the colorific intensity to correspond only
to that of 2 million healthy corpuscles, we at once come to the
legitimate conclusion that, in respect to the amount of haemoglobin
which it contains, each corpuscle, in the case under investigation, is
equivalent to one-half a healthy corpuscle.

150 CHARACTERS OF THE BLOOD IN ANAEMIA. [BOOK I.

By pursuing such methods of investigation Hayem has obtained
results which have led him to recognize four classes of cases of
anaemia, as determined by changes in the blood.

Hayem's 1. Slight anaemia, characterized either by slight

classification changes in the corpuscles or by none. Corpuscles
Anaemia °f equivalent to between 4 and 3 millions of healthy cor-
puscles (the actual number may be actually as large
as in health). The individual value of the corpuscles varying
between 1 and 0*70 (1 expressing the normal as determined by
richness in colouring matter).

2. Anaemia of medium intensity, characterized by marked
changes in the corpuscles, with a diminution in the size of the
corpuscles, the total of which may be equivalent to between 3 and 2
millions of corpuscles in the cubic millimetre, though the actual
number may vary between 5,500,000 and 3,000,000 ; the individual
value of the corpuscles may vary between 0*30 and 0'80.

3. Intense anaemia, characterized by marked changes in the
corpuscles and especially by corpuscles of very unequal dimensions,
some being of very large size. The total number of corpuscles may
be equivalent to between 2 millions and 800,000 healthy corpuscles,
though the actual numbers may fluctuate between 2,800,000 and
1 million ; the individual value of the corpuscles may vary between
0'40 and 1.

4. Extreme anaemia, characterized by altered corpuscles, of
unequal dimensions, whose mean size approaches the normal, and
may even exceed it. The total number of corpuscles may be equivalent
to between 800,000 and 450,000 healthy corpuscles, and the actual
number of corpuscles may be very small, even smaller than corre-
sponds to the number of corpuscles, so that in these cases the amount
of colouring matter in each corpuscle may actually be higher than
the normal.

During the course of treatment of anaemia by preparations of iron
the progress of the case can be studied by comparative determinations
of the total number of corpuscles and their colorific intensity, and it
is sometimes found that as recovery advances the corpuscles actually
decrease in number whilst their richness in colouring matter augments.

The Blood An attempt has been made to establish a distinction

in cases of between the changes in the blood in chlorosis and other
Chlorosis. forms of anaemia. It appears to the Author however

that such a distinction is of the most artificial and useless kind.

Chlorosis may be defined as an intense form of anaemia occurring
in women (usually in adolescents), evidenced by a yellowish-green
tint of the pallid skin, often associated with marked nervous symptoms,
and usually, if not always, associated with disorder of the menstrual
function. It is, that is to say, a disorder of nutrition accompanied by
intense anaemia, which in some yet unintelligible manner is specially
connected with the menstrual function.

CHAP. III.] THE BLOOD IN DISEASE. 151

In man, unless suffering from grave organic disease, or subject to
obvious drain of blood constituents, the conditions rarely exist which
are requisite to induce so characteristic an anaemia; in him, indeed,
anaemia from hidden functional disorders of nutrition is rare, and,
from the less susceptible nervous organization of man, when occurring
in him, anaemia wears a somewhat different aspect from that which
it presents in the chlorotic girl. Nevertheless chlorosis may be taken
as the very type of anaemia depending upon a purely functional
disorder qf nutrition, and in discussing the changes which the con-
stituents of the blood undergo in anaemia, we may take as examples
cases of chlorosis, as they have been most frequently studied and
obviously present the least complicated instances of anaemia.

It has been generally observed that in chlorosis, the bulk of the
clot as compared with the volume of serum is markedly diminished
and that usually the clot presents a buffy-coat. This appearance has
by some been interpreted as indicating that the blood of chlorosis
contains an excess of fibrin, which is not however the case. Amongst
the causes which may induce the appearance of a buffy-coat is un-
doubtedly the increase in the relative quantity of liquor sanguinis as
compared with the corpuscles. Thus Buchanan shewed long ago that
when recently shed blood is diluted with serum, the clot which sepa-
rates always exhibits a buffy-coat.

If we except the diminution in the number of corpuscles and of
haemoglobin and the consequent increase of water, the blood in
chlorosis exhibits no other deviations perceptible by chemical analysis.

The following exhibits tlie maxima, minima and mean results of twelve
analyses of blood made by Andral and Gavarret in nine cases of chlorosis1: —

Composition of Blood in 1000 parts.

Solid
Water. Solid residue. Fibrin. Corpuscles. residue of

Serum.

Maximum 8687 .. 181*3 .. 3*6 . . 95-7 . . 100-9
Minimum 818-5 . . 131-5 . . 2-1 . . 38'7 . . 75-4
Mean 853-2 . . 146-8 . . 2-9 . . 56-7 . . 88'0

The following table exhibits the mean composition of the blood of six
chlorotic girls as determined by Becquerel and Rodier.

Density of defibrinated blood . . 1045*8

Density of serum . . . . 1028-1

Water . . . 828-1

Solid constituents . . . . 171 '8

Fibrin 3-4

Fat 1-5

Albumin . . . . . 72-1

Blood-corpuscles . . . . 86*0
Extractive matters and salts .

1 Simon's Animal Chemistry, Vol. i. p. 314.

152 THE BLOOD IN LEUCOCYTHAEMIA. [BOOK I.

The iron amounted to 0'319 parts in. 1000. This would correspond
to 74'1 parts of haemoglobin in 1000 of blood.

Changes Experience has long shewn that in the treatment

effected in of all forms of anaemia and especially of chlorosis, the

the Blood by administration of preparations of iron possesses astound-

use oTinm18'1 *n° em"cacy an(^ tnat tneJ are often essential to recovery.
The clinical fact is borne out in a striking manner by
the results of chemical analysis, no less than by the results obtained
by the methods of enumeration.

The following are the results of the analyses by Andral and Gavarret
of the blood in two cases of chlorosis, before and after the administration
of iron. The number of blood-corpuscles increased coincidently with the
improvement in the complexion and general condition of the patients.

1ST CASE. Previous to use After use of

of iron iron

Water in 1000 parts . 8667 . 818-5

Fibrin . . " . . 3-0 . 2'5

Blood- corpuscles . . 46'4 . 95*7

Residue of serum . . 83'9 . 83-3

2ND CASE. Previous to use After use of

of iron iron

Water in 1000 parts . 852'8 . 831-5

Fibrin ... 3-5 3-3

Blood-corpuscles . . 49 '7 . 64 -3

Residue of serum . . 94-0 . 1009

Leucocythaemia (Leukaemia).

In this disease the colourless corpuscles of the blood are enor-
mously increased in number, so that they may amount to one-sixth
or, it is said, even one-third the number of the coloured corpuscles.
The condition is most usually associated with great hypertrophy
of the spleen (splenic leukaemia), but more rarely with a general
hypertrophy of the lymphatic glands of the body (lymphatic leukaemia}.
Usually the colourless corpuscles do not differ in shape or size from
those in normal blood, though in certain cases (lymphatic leukaemia),
they appear to* be smaller in size.

In certain cases of leucocythaemia, nucleated coloured corpuscles
are found which are similar to the nucleated coloured corpuscles
which occur in the blood of the embryo. It has been shewn by
Neumann1 that when these cells occur the marrow of the bones is
affected (myelogenic leukaemia] ; there occur, indeed, cases of leuco-
cythaemia in which an affection of the marrow is the primary and
essential lesion, others, and these are probably the more numerous,
in which it occurs as a secondary phenomenon 2.

1 Neumann, "Ein Fall von Leukamie mit Erkrankung des Knochenmarkcs."
Archiv d. Heilkunde, VoL xi. (1870), p. 1—15.

3 Eichhorst, Die progressive perniz lose Anamie. Leipzig, 1878, p. 6.

CHAP. III.] THE BLOOD IN DISEASE. 153

According to Scherer1 the blood in leucocythaemia, besides being
poor in haemoglobin, contains considerable quantities of hypoxanthine,
of lactic acid, and of an albuminoid substance whose solutions possess
the power of gelatinizing, and which is therefore surmised to resemble,
if not to be identical with, gelatine. These results have been confirmed
by the more recent researches of v. Gorup-Besanez and Salomon.
The whitish-red blood clot obtained from the vessels after death,
was by the former observer boiled with water, and the aqueous
extract, when concentrated and then allowed to cool, gelatinized.
As the body when separated did not in the least deviate the plane
of polarization, v. Gorup-Besanez2 considered its identity with gelatine
to be disproved.

Salomon found 0*116 grms. of hypoxanthin in 1550 c.c. of blood
obtained after death ; the same quantity of blood yielded 1'5 grms. of
zinc lactate.

Neither hypoxanthine nor lactic acid can, however, according
to Salomon, be looked upon as characteristic of the blood of leuco-
cythaemia, as they probably occur in healthy blood and certainly in
that of other diseases 3.

Charcot's Charcot4 discovered in the blood, the spleen and the

liver of leucocythaemic patients, certain microscopic
colourless elongated crystals, insoluble in water, but soluble in acids
and alkalies, which he and Yulpian5 were inclined to consider as
consisting of a proteid body. Salkowski considered them to be com-
posed of a mucin-like body. According to Zenker6 these crystals
are often observed to form within the colourless corpuscles.

These crystals are never observed in the blood during life, or
immediately after it is drawn from the vessels ; they usually separate
in large numbers as the leucaemic blood decomposes.

They usually appear as very much elongated octohedra, or simply
as spindle-shaped bodies, which are transparent, colourless, insoluble
in ether, chloroform, and alcohol ; soluble with difficulty in cold
or hot water, but easily soluble in dilute acids and alkalies. They
appear to consist of the phosphate of a base which has been discovered
by Schreiner7 in semen and in the spirit in which anatomical
preparations have been kept. To the hydrochlorate of this base
this author provisionally ascribes the formula C2H5N. HC1.

1 Scherer, Verli. d. physik.-med. Gesell. zu Wiirzburg, Vol. n. pp. 321 — 325.

2 v. Gorup-Besanez, " Untersuchung des Blutes bei lienaler Leukaemie." Sitzung*-
berichte der physikal-medicin. Societat in Erlangen, Mai, 1873. Abstracted at con-
siderable length in Maly's Jahresbericht, Vol. iv. p. 126.

3 Salomon, " Zur Lehre von der Leukamie." Archivf. Anat. u. Phys., 1876, p. 762.

4 Charcot et Kobin, Comptes Rendus de la Soc. de Biologic, 1853.

5 Charcot et Vulpian, Gazette Hebdomadaire, 1860, p. 755.

6 Zenker, " Ueber die Charcot'schen Crystalle im Blute und Geweben leuka-
mischer und in den Sputis." Archiv f. klin. Medicin, xvui. 125 — 135.

7 Schreiner, "Eine ueue organische Basis in thierischen Organismen." Liebig's
Annalen, Vol. cxciv., p. 68.

154

THE BLOOD IN PERNICIOUS ANAEMIA.

[BOOK i.

Results of

Leucocy-
thaemia.

The following Table exhibits the results of six sys-
tematic analyses of the blood of leucocythaemia by
various observers1.

Authorities.

Density
of

Density
of

Fibrin.

Solid
Matters

nf

Dry

blood

r*riY

Total
solid
matters

I
Water.

Blood.

Serum.

•

Serum.

puscles.

of
Blood.

Robertson

1041-5

1026-5

6-0

72-0

67-5

145-5

854-5

)>

1036

1023

2-3

67-0

49-7

119-0

881-0

»

1049-5

1029

5-0

95-0

80-0

180-0

820-0

»

1043-5

1027

3-2

80-7

82-3

166-2

833-8

Robertson

1049-5

1029

5-0

95

80

180

820

Isambert

1-4

69

69-2

142

858

Progressive Pernicious Anaemia.

By the above term Biermer2 has designated a remarkable form
of anaemia which had already been recognized and graphically
described by Drs Addison and Samuel Wilks3.

Occurring more frequently in women than in men, in adult life
than in adolescence or old age, this disease seems frequently to
originate in pregnancy, or to have exhausting disease as a predisposing
cause. Cases, however, undoubtedly occur in which no predisposing
cause can be traced.

Commencing insidiously as one of the more ordinary forms of
anaemia, this disease is distinguished by the rapidity with which
all the phenomena of the most intense anaemia are developed —
such as intense pallor, dyspnoea, inability to undergo the slightest
exertion, tendency to syncope, dropsy. It differs from the ordinary
forms of anaemia by the occurrence of more or less pyrexia, by a
great proneness to retinal hemorrhages, but especially by the much
greater tendency towards a rapid fatal termination.

The disease is not associated with any essential lesion of any

1 This table is taken from Robin's Traiti des humeurs, 2nd edition, p. 272, to which we
were referred by Gautier, who also uses it. (Gautier, Chimie Appliquee, &c., Vol. n, p. 321).

2 Biermer, "Vorlaufige Mittheilung iiber fettige Degeneration des Herzens und
der Gefasse in Folge von Anamie;" Tageblatt d. 42. Versamml. deutsch. Naturforscher
u. Aerzte in Dresden, 1868. Deutsches Archiv f. Min. Med. vol. xm., p. 209.

3 Addison, On the constitutional and local effects of disease of the suprarenal
capsules. London, 1855. Collected Works, New Sydenham Soc. 1868, p. 211.

Samuel Wilks, "Cases of idiopathic fatty degeneration. With remarks on arcus
senilis." Guy's Hospital Reports, 1857, p. 203.

In reference to the claims of these two authors to the merit of having first
recognized the disease under discussion, the reader is referred to an interesting paper
by Dr Pye-Smith, entitled "Zwei Falle von Anaemia idiopathica perniciosa" (Virchow's
Archiv, Vol. LXV. (1875), p. 507).

CHAP. III.] THE BLOOD IN DISEASE. 155

important organ, though in many cases fatty degenerations of heart,
liver, and kidneys — changes which must be considered as mere
results of imperfect nutrition — have been observed.

The statements of authors who have described this remarkable
disease vary as to the changes which the blood undergoes.

In several cases where an enumeration of the coloured corpuscles
has been effected, a very remarkable diminution has been found.
Thus in a case described by Lepine the coloured corpuscles sank
on the day preceding the death of the patient to 378,750 in 1 cubic
millimetre. In a case described by Ferrand the number of coloured
corpuscles was found to be 500,000 in 1 cubic millimetre, and the
amount of haemoglobin estimated by the colorific intensity had
sunk to one-tenth that of normal blood.

Dr Hermann Eichhorst1 has studied the anatomical changes
which the blood undergoes in certain cases of progressive pernicious
anaemia, and the following is a brief summary of his observations :

1. The blood has a serous, amber-coloured appearance, with
scarcely a trace of red (?), and coagulates with difficulty.

2. The colourless protoplasmic granules, which are always to be
found more or less distributed throughout healthy blood, are. com-
pletely absent.

3. The colourless cells of the blood are extraordinarily few in
number.

4. The coloured corpuscles of the blood are diminished in number.
Those which retain the form of normal corpuscles are observed to
be increased in size, having a diameter of 8 — 9//,, some being as
large as 9*5 //, and very few having a smaller diameter than 8 p.

5. In addition to the ordinary corpuscles there occurs a second
class of corpuscles. These are much smaller than normal coloured
corpuscles, having a diameter varying between 3/u. and 3'5yLt or 4/<t;
they are not biconcave but globular, and are of a deeper red colour
than normal blood-corpuscles. In one case these smaller corpuscles
were in the ratio of 1 : 5 of the normal corpuscles.

That cases of progressive pernicious anaemia occur in which the
changes which Eichhorst has described are not seen appears certain
from the observations which have already been published. He
himself admits thai there may be, and doubtless are, several
classes of cases, which have been, and which may be, grouped
under the term of Progressive Pernicious Anaemia, but he argues
that during the progress of the case a diagnosis is alone possible
where the changes which he has described are found to exist.

The observations of Dr Byrom Bramwell2 on several cases of
pernicious anaemia, whilst they in the main confirm Eichhorst's
statements in reference to the presence in the blood of small non-

1 Eichhorst, Die progressive perniziose Andmie. Eine klinische und kritische
Untersuchung, p. 378. Leipzig, Verlag von Veit u. Comp. 1878.

2 Byrom Bramwell, "Idiopathic or Progressive Pernicious Anaemia, with cases."
Edinburgh Medical Journal, Vol. xxin. (1877), p. 408.

156 THE BLOOD IN SCURVY, GOUT, RHEUMATISM, &c. [BOOK I.

biconcave coloured corpuscles, seem to shew that, not unfrequently,
nucleated coloured corpuscles are present. In these cases it is to be
assumed that an affection of the marrow of the bones existed similar
to that which occurs in cases of myelogenic leukaemia. Professor
Grainger Stewart1 failed to observe Eichhorst's corpuscles in two fatal
cases of pernicious anaemia. There can be no doubt moreover that
cases of pernicious anaemia do occur which terminate in recovery.
On the other hand, these corpuscles have been observed in large
numbers in the blood of a case of leucocythaemia2.

Scurvy.

Very little reliable information is possessed in reference to the
alterations of the blood in this disease. When humoral doctrines pre-
vailed, scurvy was looked upon as preeminently a disease due to
marked changes within the blood, and attempts were made to explain
the hemorrhagic tendency by ascribing it to a definite change in
the composition of the blood, notably to a great diminution of
fibrin (Andral3.)

At the present time we are naturally more inclined to consider
the hemorrhages which occur in this disease as due to a morbid
change of the walls of the blood-vessels, and we are therefore not
surprised to find that in many cases where the blood of patients
affected with scurvy has been analysed the results have been purely
negative.

Becquerel and Rodier had the opportunity of analysing the
blood of five patients affected with scurvy. They found it to be
poor in solid constituents and to be decidedly poor in corpuscles.
In these five cases the proportion of haemoglobin (which we have
calculated from the iron which they determined) was respectively
121-33; 64-41; 90'9 ; 99'3; 67'4 grammes per 1000 of blood.
The amount of fibrin was found in no case to be below the normal,
whilst in three of the five it was above the normal.

Mr Busk analysed the blood in three cases of scurvy which
occurred in the Dreadnought Hospital Ship, and he found a great
diminution in the solid constituents of the blood as a whole, a
marked increase in the proportion of fibrin, albumin, and salts, with
a very great diminution in the proportion of blood-corpuscles4.

From the evidence which has been accumulated, it would therefore
appear, that in scurvy a condition of anaemia is a constant antecedent,
using that term to designate a diminution in the total corpuscular
richness of the blood. The increase in the quantity of fibrin which
sometimes, though by no means always is observed, probably depends

1 Grainger Stewart, "Note upon Professor Eichhorst's new pathognomonic symptom
of progressive pernicious Anaemia." British Medical Journal, July 8th, 1876, p. 40.

2 Heuck, Zwei Fdlle von Leukdmie mit eigenthiimlichem Blut- resp. Knochen-
marksbefund. Virchow's Archiv, Dec. 1879, p. 475.

3 Andral, Essai d' Hematoloyie pathologiqtie, Paris, 1843, p. 128.

4 The Author has failed in 'his attempts to discover the medium through which Mr
Busk's results on this subject were published. They are, therefore, quoted at second hand.

CHAP. III.] THE BLOOD IX DISEASE. 157

/

.

upon the intercurrent inflammatory lesions which so frequently occur.
As yet, however, facts are wanting to shew in what precise manner
the composition of blood is affected in scurvy.

Purpura Hemorrhagica and Haemophilia.

In these two diseases no constant alteration of the blood has been
discovered.

It was maintained by Becquerel and Eodier that in these diseases
as well as in scurvy, the proportion of fibrin was diminished, but such
is not actually the case. The colourless cells of the blood too have in
some cases been stated to be increased ; in others, on the other hand,
to be unaffected. We can scarcely doubt that these are affections
in which primarily the blood-vessels are more at fault than the blood.

Gout.

There are few diseases in which so notable a change in the com-
position of the blood can be observed as in gout. Whatever may be
its cause, gout appears to be preeminently a disorder of nutrition, in
which there is a great tendency to a peculiar inflammatory process,
affecting in the first instance and chiefly (though by no means
exclusively) certain joints. In this peculiar, inflammation crystalline
deposits of sodium urate are formed in the affected parts.

It is the great merit of Dr Garrod1 to have discovered and
demonstrated that during the attack of acute gout, as well as in
the course of chronic gout, the blood invariably contains an excess of
uric acid, so that from a comparatively small quantity of blood serum,
a characteristic microscopic crystallization of uric acid can be obtained.
This accumulation of uric acid in the blood is probably in some
measure due to its non- elimination by the kidneys, seeing that whilst
the attack of gout is at its height, though the relative proportion
of uric acid in the urine may be high, and an actual spontaneous
separation from it often occurs after emission, the actual amount
excreted is usually below the normal.

The non -elimination is doubtless connected with the kidney
affection which is an almost invariable concomitant of gout.

Garrod shewed, moreover, that in gout the blood contains oxalic
acid2, and it is very probable that it contains an excess of urea ; upon
this point, however, further research is greatly needed.

It appears to be indisputably proved that chronic lead poisoning
renders those affected with it specially liable to attacks of gout,
also that the ingestion of minimal doses of lead compounds will some-
times induce in the gouty an acute attack of the disease.

1 Garrod, "Observations on certain pathological conditions of the blood and urine
in gout, rheumatism and Bright's disease." Medico-Chirurgical Transactions, Vol. xxxi.
p. 83, (1848). See also A treatise on Gout and Rheumatic Gout, 1876; and Article
"Gout," Eeynolds's System of Medicine, 3rd ed. London, Vol. i. p. 810.

2 Medico-Chirurgical Transactions, Vol. xxxvn. (1854), p. 54.

158 THE BLOOD IN RHEUMATISM, &C. [BOOK I.

Articular Rheumatism and Rheumatoid Arthritis.

In acute rheumatism there occurs a specific inflammation of the
serous membrane of the joints, usually accompanied by considerable
effusion, but scarcely ever ending in suppuration. There is at the
same time a very great tendency to inflammatory changes of the
pericardium and endocardium.

Clearly separated from gout by the absence of an excess of uric acid
in the blood, and in the large majority of cases by that of lesions of the
kidney, acute articular rheumatism is a type of a disease in which
local inflammatory changes are accompanied by a high temperature.

As in all other diseases in which a high temperature accompanies
local inflammatory changes, the most marked modification in the
chemical composition of the blood consists in an augmentation of the
fibrin which separates on coagulation.

The blood is usually buffed and cupped ; the amount of fibrin
is said to bear some relation to the intensity of the febrile disturbance.

At first the increase of fibrin is the only change which chemical
analysis reveals ; in the course of the disease, however, anaemia
supervenes, and is evidenced by the aspect of the patient, no less
than by a diminution in the number of the corpuscles. The changes,
if any, which occur in the anatomical characters of the corpuscles
or in the proportion of haemoglobin which they contain, have not
yet been made out.

According to Becquerel and Rodier, in acute rheumatism, as
well as in all other acute inflammations, there occur (1) an increase in
the amount of fibrin ; (2) a diminution in the amount of blood-
corpuscles ; (3) a diminution in the serum-albumin ; (4) an increase
in the fatty matters; (5) a diminution in the soluble salts of the
blood. With the exception of the increase in the amount of fibrin,
the other changes are in all probability chiefly to be referred to the
disturbance between the balance of income and expenditure which
occurs in all acute diseases, and which leads to rapid loss of weight
and anaemia; these are, however, like the changes revealed by
analysis, but the effects of a morbid process of which the exact starting
point, no less indeed than the exact seat, is hidden from us.

In subacute rheumatism and in rheumatoid arthritis there are no
constant changes in the composition of the blood. It is, however,
of great importance, as constituting the widest difference between
.the so-called rheumatic gout (rheumatoid arthritis) and true gout,
whether acute or chronic, that in the first-named disease there is no
tendency whatever to accumulation of uric acid in the blood, and to
its subsequent deposition in the tissues of the body.

Rickets and Osteomalacia.

In these two diseases, in which grave disorders of nutrition exist,
which must necessarily affect the composition of the blood, no systematic
investigation, so far as we know, has been attempted.

CHAP. III.] THE BLOOD IN DISEASE. 159

B. THE BLOOD IN FEVERS.

Febricula or Ephemeral Fever.

Becquerel and Rodier * have made many analyses of the blood in
fehricula, which led them to the conclusion that none of the con-
stituents of the blood undergo any constant or perceptible change
in amount.

Typhus Fever.

So far as we are aware the only analyses of the blood of typhus
fever are those made by Dr Guennau de Mussy and M. Rodier when sent
in 1847 by the French government to report upon an outbreak
of typhus fever in Ireland. The observations published by other
continental writers, and which have been supposed to refer to
typhus, have, for the most part at least, referred to typhoid2. The
observations of Guennau de Mussy and Rodier shewed that in typhus
there is, in general, a diminution in the density of the blood, no
augmentation in the amount of fibrin, and a lowering of the specific
gravity of the serum.

Typhoid Fever.

Very thorough analyses of the blood in typhoid were performed by
both Andral and Gavarret, and Becquerel and Rodier. From their
researches the latter observers arrived at the following conclusions.
(1) That in typhoid fever, in its early stages, there is usually no
change in the composition of the blood. Occasionally, however,
in very acute cases accompanied by great prostration and by hemor-
rhages, there occurs a diminution of the chief elements of the blood,
and particularly of fibrin. (2) That in the later stages of the disease,
the corpuscles and serum-albumin diminish, under the influence of
the deficient nourishment and of the losses of liquid which the patient
is subject to. The amount of fibrin remains normal, or is diminished as
the disease advances or becomes more grave.

Relapsing Fever.

Though there can hardly be any doubt that all zymotic diseases
are produced by certain germs or elementary organisms, yet there
are only two diseases in which the presence of such germs in the blood
has been clearly demonstrated. These, two diseases are : Relapsing
fever, and Splenic fever.

Obermeier3 was the first to demonstrate in the blood taken from

1 Becquerel et Rodier, Traite de Chimie pathologique, p. 133.

2 This remark applies to the observations of Lehmann (Fhys. Chem., Vol. n.
pp. 265 and 266), which are referred to hy Dr Buchanan in his Article on •' Typhus
Fever" in Reynolds's System of Med., Vol. i. p. 549.

3 Centralbl. /. d. med. Wissensch., 1873, p. 145.

160

THE BLOOD IN RELAPSING FEVER.

[BOOK i.

patients suffering from relapsing fever the presence of small organisms,
which consisted of slender spiral filaments in a continual state of
lively motion. Obermeier further observed that these organisms, which
belong to the class of Schizomycetes and which resemble spermatozoa
or spirilla, were absent in the period of apyrexia which intervened
between the first and second period of fever, and that they reappeared
again with the onset of the second period of fever. Obermeier's
observations have been verified by all subsequent observers, notably
by Weigert, Lebert and Heydenreich2. According to these observa-
tions it is now clearly made out, that these organisms have exactly
the same appearance as the Spirochaeta described by Ehrenberg and
found in stagnant waters. They consist of slender spiral filaments
O'OOl mm. in diameter, 015 — 0'2mm. in length; they are homo-
geneous in structure ; and they are continually moving, their motion
being both rotatory and progressive ; they occur either singly or in large
groups, attached to the red and white corpuscles of the blood (fig. 27).

FIG. 27. THE 'SPIRILLUM' OF RELAPSING FEVER. (Heydenreich.)
a, b, c, d shew various conditions of aggregation. At / groups of filaments are
to entangle a colourless and several coloured corpuscles.

They are only found in the blood of patients suffering from
lapsing fever, and have not been found in any of the organs of such
patients after death. They are found several hours after the onset of
the febrile attack, and increase in quantity up to the termination of
the first pyrexial period ; they are absent in the interval and reappear
again in the blood several hours after the second pyretic period has set
in. When examined under the microscope their movements continue
for several hours at the ordinary temperature, but a low or a high
temperature and several reagents (alkalies, acids, etc.) stop their move-
ments at once.

About their life-history nothing definite is known1, nor have
observers been able to propagate them in cultivating fluids. In several
cases, however, inoculation with a small quantity of blood taken from
patients suffering from Relapsing fever has induced the disease.

1 Klinische und mikroscopische Untersuchungen uber die Parasiten des Ruckfalltyphus,
1877.

2 See an interesting paper "On the life-history of Spirillum," by Patrick Geddes
and J. Cossar-Ewart, Proceedings of the Royal Society, 1878, No. 188, p. 481, which
treats of the spirillum of stagnant water.

CHAP. III.] THE BLOOD IN DISEASE.

Besides the presence of spirillum, other changes have been noticed
in the blood in cases of relapsing fever, such as the disproportion
between white and red blood corpuscles and the presence of large
masses of protoplasm and of degenerated endothelial cells; these
changes are however found in many pathological conditions, especially
in fevers.

Splenic Fever (of Cattle}.

Splenic fever is a disease not uncommon amongst cattle,

a^gus and is both enzootic and contagious, i.e. the disease occurs

chiefly in certain districts where it is enzootic, but is also

propagated by contact. In the blood of animals suffering from splenic

disease, Davaine long ago found certain bacilli or rod-like bacteria, which

he believed to be the cause of splenic fever. It was however found that,

though inoculation with blood containing these bacilli produced splenic

disease, the blood of the animals thus inoculated did not contain any

bacilli, though highly active when inoculated.

Koch1 and after him Cossar-Ewart2 have studied the life-history of
these organisms and have cleared up the discrepancy. Koch found that
in the blood and juices of the living body these rod-like bacteria multiply
by fission ; after the death of the animal, or when brought into cultivating
fluids, these rods grow into long filaments or threads which produce spores-
provided always the surrounding temperature remains moderately high ;
eventually the threads disappear and nothing but spores remains behind.
These spores, which withstand decomposition for a long time (while the
bacilli are very perishable), have been found to develop again into bacilli,
when in contact with the air and at a moderate temperature.

The contagious character is due to the bacilli, but in the blood of the
slaughtered or dead animals the bacilli develop under favourable circum-
stances into spores, and these being much less perishable and easily wafted
about by currents of air are the cause of the epizootic character of the
disease.

The observations on the presence of germs in the blood in other infectious
and contagious diseases have as yet led to no perfectly definite results,
though bacteria and micrococci have been seen in the blood in Pyaemia,
Septicaemia, Diphtheria and Scarlatina3.

1 Cohn's Beitraye zur Biologie der Pftanzen, Band 1, Heft 3, p. 277. "Die Aetiologie
der Milzbrand-Krankheit, begriindet auf die Entwicklungsgeschichte des Bacillus An-
thracis." Cohn's Beitrcige zur Biologie der PJlanzen, Vol. n. 1876.

2 Cossar-Ewart, "On the life-history of Bacillus Anthracis." Quarterly Journal of
Microscopic Science, Vol. xvm. (New Ser.), p. 161.

3 While speaking of living organisms in the blood we may just mention the presence
of some more highly developed parasites found occasionally in the blood, thus: The Bil-
harzia Haematobia (Cobbold), or Distoma HaematoMum (Bilharz) inhabits the branches
of the portal system, but most commonly the small veins of the ureter, bladder and
pelvis of the kidney. It is endemic in Egypt where Griesmeyer found it in nearly
one-third of all autopsies. The full-grown animal, bisexual and belonging to the
trematodes, measures from 5 — 8 mm., and is soft-skinned.

This disease is recognized during life by the presence of the characteristic ova and
embryos in the urine, which invariably also contains some blood. The eggs are

G. 11

162 THE BLOOD IN INTEKMITTENT FEVERS. [BOOK I.

Intermittent Fevers.

There is primarily no chemical change in the blood in these fevers,
as is shewn by the observations of Andral and Gavarret and of MM.
Leonard and Foley, who studied intermittent fevers in Algeria1.
According to the latter observers, especially, whatever the type of the
intermittent, there is primarily no alteration in the blood, and no
perceptible difference between the condition of the blood taken during
an ague fit and that taken in the period of intermission. As the
disease advances, however, the patient becomes anaemic, the blood-
corpuscles and the solid matters of the serum diminishing in quantity.

Bacillus Klebs and Tommasi-Crudeli2 have made the re-

Maiariae. markable discovery that the soil in malarial districts

contains a bacillus to which they have given the name of Bacillus
Malariae, which when introduced into the system of rabbits induces
fever of an intermittent type. They have succeeded in cultivating
this organism, which by cultivation loses none of its specific power.
In the animals in which intermittent fever was thus artificially
induced, the spleen became enlarged and in its juice, as also in the
blood, were usually found the spores of B. Malariae, and sometimes
the rod-like forms of the mature organism.

Since the above observations were first published, Marchiafava3
has in three fatal cases of pernicious malarial fever succeeded in
discovering in the splenic pulp, in the blood, and in the marrow of the
bones, spores and filaments of B. Malariae.

Pigment in In severe forms of intermittent fever particles of

the blood. black pigment have been found in the blood, and

accumulations of such masses have been seen after death in and
around the smaller blood-vessels and capillaries in the brain, liver,
and spleen, and, according to Arnstein4, in the medulla of the bones.

The pigment occurs in the form of smaller or larger granules, and
is found either free in the blood or contained in the white blood-
corpuscles.

oval, boat-shaped, bodies with a spiny projection at the anterior end. The embryo
(as seen in the urine occasionally) is ciliated and flask-shaped.

Filaria sanguinis hominis (Lewis), a minute nematoid entozoon, is found in the
urine and also in the blood of . patients suffering from chyluria. The disease is
endemic in India, but some cases have also been observed by Dr Bancroft in
Australia.

The filaria is a long narrow worm about the breadth of a red blood-corpuscle and
about 0-033 mm. long, it has a hyaline tubular envelope closed at both ends ; under
a high power transverse striae and granular masses are seen. When examined in the
fresh state it shews active snake-like movements.

1 Leonard et Foley, quoted by Bequerel et Eodier, Op. cit., p. 132.

2 Edwin Klebs e Corrado Tommasi-Crudeli, "Studi sulla natura della Malaria."
JReale Accademia dei Lincei, Roma, 1879. (Memorie della classe di scienze fisiche,
matematiche e naturali. Serie 3». Vol. iv°. Seduta del 1 giugno 1879.)

3 Dr Ettore Marchiafava, quoted by Klebs and Tommasi-Crudeli, op. cit. p. 61.

4 Arnstein, "Bemerkungen tiber Melanamie und Melanose." Virchow's Arch. Vol.
LXI. p. 494.

CHAP. III.] THE BLOOD IN DISEASE. 163

The pigment is considered to be altered haematin, and, ac-
cording to the older view (Frerichs, Virchow), its presence in the
blood is due to a breaking up of the red blood-corpuscles in the
spleen and liver. According to Arnstein, however, the pigment
originates in the blood itself, from the destruction of the red blood-
corpuscles during the attack of fever, and penetrating into the
leucocytes of the circulating blood becomes deposited along with
them in the spleen, liver and medulla of bone1.

The formation of black pigment has lately been observed going
on within the coloured corpuscles by Marchiafava2, whose observations
have been confirmed by Klebs and Tommasi-Crudeli ; in the process,
iron is set free from the state in which it exists in haemoglobin, so
that when the corpuscles which have become black are treated with
dilute hydrochloric acid and potassium ferrocyanide they acquire a
blue colour.

Scarlet Fever, Measles, Small-pox, Erysipelas.

In the first of the above diseases there is primarily no alteration
in the proportion of blood constituents. In scarlet fever, however,
when kidney complications set in, there is a tendency to accumu-
lation of urea in the blood, which often attains a high degree, and
subsequently a great diminution of the proteids of the blood occurs.

In small-pox, coincident with the inflammatory changes in the
skin, there occurs a moderate increase in the amount of fibrin.

In erysipelas, the blood at first presents in a characteristic manner
the appearances which i't always assumes when an acute inflamma-
tory process involves an organ of any magnitude, t>r implicates any
considerable extent of one of the tissues ; the blood, in consequence,
yields much fibrin. There is, however, no other constant alteration,
unless there be any truth in the very doubtful statement of Schonlein3
that the serum which separates from the blood, in erysipelas, is always
tinged yellow by the colouring matter of bile.

The Blood in Cholera.

A very elaborate investigation into the changes which blood
undergoes in cholera was made by Professor Carl Schmidt4 during the
epidemic of that disease which ravaged Dorpat in the summer and
autumn of 1848.

In consequence of the very great transudation of water and
albumin from the alimentary canal, the blood in cholera becomes
excessively poor in water and relatively rich in solid constituents, so

1 Virch. Arch. Vol. LXI. p. 494.

2 Marchiafava, " Commentario clinico di Pisa. Fascicolo del gennaio 1879. Quoted
by Klebs and Tommasi-Crudeli, op. cit. p. 57.

3 Schonlein, quoted by Simon, Animal Chemistry, Vol. I., p. 278.

4 Carl Schmidt, " Charakteristik der epidemischen Cholera gegenuber verwandten
Transudationsanomalien." Leipzig und Mitau, 1850.

11-2

164 THE BLOOD IN DISEASES OF THE HEART. [BOOK I.

as to assume a viscid consistency. There thus appears to be an increase
in the number of the coloured blood-corpuscles and even of the
serum-albumin. Whilst the proper salts of the serum may fall to
one half, the blood-corpuscles also are robbed of their mineral con-
stituents, their potassium and phosphoric acid diminishing.

C. Schmidt pointed out that the blood in cholera also contains
urea; his method was however obviously not calculated to obtain
very accurate information on this point. Voit found as much as 2*43
grammes of urea in 1000 parts of the blood of a cholera patient,
and Chalvet as much as 3'60 grms in 1000 \

C. THE BLOOD IN DISEASES OF THE HEART.

Mode in Through the interference which is brought about

in the functions of other organs, especially of the

fluence the lungs, the liver, and the kidneys, diseases of the heart

composition often lead " in the end to marked alterations in the

of the Wood. quality of the blood. We have only to cite the cyanosis

which accompanies a patent foramen ovale or persistent

ductus arteriosus to remind the reader how a mechanical lesion of the

vascular system may interfere with the respiratory exchanges of the

blood so markedly as to require no elaborate investigation of the gases

of the blood to reveal it.

In lesions of the mitral valve particularly, conditions are es-
tablished which, by rendering the pulmonary circulation difficult,
bring about changes in the lungs which soon lead to deficient oxy-
genation of the blood and to its concomitant symptoms ; especially is
this the case in mitral stenosis. The difficulty which the left auricle
encounters in completely emptying itself of blood, leads first of all to
a rise of the blood-pressure in the pulmonary capillaries, and this in
its turn prevents the propulsion of more than a small amount of
blood from the right ventricle, which in its turn reacts upon the right
auricle and through it upon the whole venous system. The increased
pressure in the latter opposing a greater resistance than normal to the
passage of blood from the arteries back to the heart, there is set up
an engorgement of vessels which soon reveals itself by congested liver
and kidneys, and by functional disorders of the alimentary canal, and
by increased transudations, which give rise to anasarca and to dropsy
of the serous sacs.

Diseases of the heart lead therefore secondarily to changes in
various organs, which in their turn react upon the composition of the
blood, and the extent to which they do so depends upon the manner
or extent in which each organ is affected.

Thus any great impediment to the pulmonary circulation will lead
directly to non-elimination of carbonic acid, and a diminution of the
oxygen taken up, evidenced by the blue cyanotic appearance of the
lips and face. A congested liver will be accompanied by the passage

1 Quoted by Gautier, Chimie appliqute, &c. , Vol. IT. , p. 337.

CHAP. III.] THE BLOOD IN DISEASE. 165

of biliary ingredients into the blood ; congested kidneys will secrete a
urine more or less charged with albumin and probably deficient in
urea, and secondarily there may be set up the condition of uraemia.

So multifarious are the ways in which heart disease may modify
the condition of the blood, that it would be useless to attempt to
classify all the changes which are thus brought about.

TheAnae- Amongst the most interesting of the phenomena

mia of Heart induced by heart disease is the condition of anaemia,
disease. Cases of anaemia in connection with heart disease may

be arranged in two classes. In the first, the anaemia follows very
closely upon the establishment of the cardiac lesion, and is obviously
dependent upon the disturbance in the blood-pressure, which has
not yet been compensated for, as it subsequently is, by changes in
the circulatory apparatus. The establishment of anaemia in these
cases is clearly explained. A certain difference between the arterial
and venous pressure, and more than that, a certain value of the
arterial pressure, is absolutely necessary in order that the nutrition of
each organ of the body shall be efficiently maintained; when the
conditions for effecting this do not exist, an alteration of the blood
will be amongst the first evidences of impaired nutrition.

Thus, then, we explain the anaemia which occurs so often a few
months after the setting up of an organic lesion of the heart by the
endocarditis of rheumatism. In this class of cases, however, the dis-
order of nutrition is often only transitory, as the disturbances in the,
circulation which followed the sudden establishment of the cardiac,
lesion are compensated, more or less completely and more or less
durably, by changes brought about, somewhat gradually, in the
circulatory apparatus. The compensating hypertrophy of the left
ventricle in aortic regurgitation, and of the left auricle in mitral
stenosis, are the best illustrations of the changes referred to.

In a second class of cases, however, anaemia occurs, and is more
intense than in those already referred to ; these are cases in which
anaemia occurs in the last stages of heart disease. After existing
for many years in an apparently dormant condition, without in-
fluencing in any very obvious manner the health of the individual
subject to them, certain heart affections reveal themselves apparently
with great suddenness. The shortness of breath, the puffy swelling
of the feet, which had been scarcely noticed for a long time, are
followed by symptoms which indicate how profoundly the circulation
is interfered with. We have no longer evidences of a mere deficient
blood-supply to important organs, depressing their functional activity,
but the sure signs of a disturbance of the proper relations between
arterial and venous pressure, which no compensating changes can
overcome. Congestions of important organs occur, dropsical accumu-
lations in the serous sacs or the areolar tissue and, of necessity, if the
patient live long enough, anaemia.

This anaemia is not difficult to explain. It is dependent partly
upon the alimentary canal being unfit to digest enough food to make

166 THE BLOOD IN DISEASES OF THE HEART. [BOOK I.

up for the waste of the body ; partly upon the presence within the
blood of products of waste which the organs formerly charged with
their excretion can no longer get rid of; but doubtless in great measure
to the disturbance in the nutrition of all tissues and organs consequent
upon the altered relation between arterial and venous pressure ; the
arteries are never as full of blood as they normally should be, and,
as a mere consequence of this, nutrition must suffer, even were all
other conditions to remain normal. The increased pressure in the
venous system may, in addition, hinder, in some measure, the discharge
of lymph and chyle into the blood.

Becquerel Becquerel and Rodier made a very complete in-

and Roller's vestigation into the changes which the blood undergoes
classification -n fo^ disease ; they classified these cases into three
Anaemia in categories, in the first of which whilst a definite lesion
Heart disease, existed it had not made itself manifest by any pro-
nounced symptoms. In the second category the general
health had become impaired ; there was some anaemia, breathlessness,
and palpitation, and oedema had supervened ; whilst the cases in the
third category were accompanied by great dyspnoea, by abundant
dropsy and by a markedly cachectic pallid skin.

In heart diseases belonging to the first category, according to
Becquerel and Rodier, there is a slight increase in the water of the
blood, and a tendency to diminution of both blood-corpuscles and
albumin. In the second stage the corpuscles and the serum-albumin
continue to decrease, and consequently the mean density of both blood
and serum falls. The fibrin of the blood often increases in quantity
though there be no localized inflammatory lesion. The fall in the
number of the blood-corpuscles is evidenced by the anaemic look of the
patient, and the diminution in the amount of serum-albumin by the
dropsy which supervenes.

The following table exhibits the mean composition of tlie blood, with

the maxima and minima in 24 cases of heart disease in the third stage.

Of the 24, 16 were cases of auriculo-ventricular (presumedly mitral)

stenosis ; in 10 cases anaemia existed, and in 11 dropsy.

Analysis of 1000 parts of blood :

Mean. Maxima. Minima.

Density of the blood 1052-54 1066-86 1040-88
Water 801-96

Blood-corpuscles 117*05 149*42 54-00

Solids of the Serum 77'53 99-52 61-74

Fibrin 3-46 646 1-25

Density of Serum 1027'60 1035-10 1020-10

In the third stage, whilst the water of the blood increases, and
corpuscles undergo a further diminution, the chief changes are per-
ceived in the serum, in which, whilst the water increases, the amount
of albumin, fats and salts diminishes more and more.

The following table exhibits the mean composition of the blood,
with the maxima and minima, in 31 cases of heart disease in the third stage.

CHAP. III.] THE BLOOD IN DISEASE. 167

Of these 20 were cases of mitral stenosis, 4 of mitral regurgitation, 2 of
aortic stenosis, and 5 of aortic regurgitation. 29 of the 31 were affected
with dropsy, and 27 had a cachectic appearance, the skin being pallid,
yellowish and generally discoloured.
Analysis of 1000 parts of blood :

Mean. Maxima. Minima.

Density of the blood 1050-19
Water 815-82

Blood-corpuscles 110'03 148-55 73-50

Solids of the Serum 71*60 81-10 52-40

Fibrin 2-55 4*47 1-30

Density of the Serum 1025'02 1032-50 1022-30

D. THE BLOOD IN DISEASES OF THE LUNGS.

In acute inflammatory affections of these organs, the blood
exhibits in a highly characteristic manner the properties which it
acquires whenever a sufficiently extensive and acute inflammatory
change occurs.

In pneumonia, especially, the blood becomes 'buffed and cupped'
and from it very much larger quantities of fibrin may be obtained
than from normal blood.

The following are the mean results of the determination of the
fibrin in several forms of acute pulmonary affections made by
Becquerel and Rodier: —

Fibrin.

In acute bronchitis 1000 parts of blood yielded 4'8 grms.
„ acute pleurisy „ „ „ „ „ frl „

. (first blood-letting ,. 7'4 „
„ acute pneumonia { -, -,-, -, , ~.Q

(second blood-letting „ 68 „

In chronic lung diseases, especially in phthisis pulmonalis, there
is invariably a considerable degree of anaemia, with marked
diminution in the number of coloured blood-corpuscles. It is alleged
that in phthisis, the amount of fibrin is frequently very much in-
creased.

E. THE BLOOD IN DISEASES OF THE LIVEK.

In affections of the liver in which, through an obstruction to the
flow of bile, jaundice occurs, there is always an accumulation of bili-
rubin in the blood, and from their passage into the urine in some cases,
we may doubtless assume the frequent concomitant presence of salts
of the bile acids. In cases of acute yellow atrophy the blood contains
leucine and ty rosin e, and doubtless (though the fact has not been
directly ascertained) the amount of urea in the blood is diminished.

A diminution of the red blood-corpuscles occurs in the course of
organic diseases of the liver, as is evidenced by the pale sallow face
of patients affected with cirrhosis.

168

THE BLOOD IN DIABETES MELLITUS.

[BOOK I.

F. THE BLOOD IN DIABETES MELLITUS.

The constant and characteristic feature of the blood in this disease
is the presence of an excess of glucose.

There are not wanting facts which point to other less inves-
tigated changes in the blood which supervene in the course of
the disease, such as for instance a large increase of the fatty matter
of the blood and the formation of acetone.

increase in Whilst in health, the amount of glucose amounts on

the glucose of an average to about 0'9 parts per 1000 p.c. (Pavy), in dia-
tiie Blood. betes it may, according to the gravity of the case, be

several times as great. In those cases where the urine contains a
large percentage of sugar, the blood likewise is very rich in that
constituent, as will be seen by the accompanying table taken from
Dr Pavy1 .-—

COMPAKATIVE STATE OF BLOOD AND UEINE IN DIABETES.

Urine.

Blood. |

.

Sugar per

Quantity per
24 hour.

Specific
gravity.

Sugar per
1000
parts.

Sugar ex-
creted in 24
hours.

lOOOparts,
mean of
two ana-

lyses.

CASE 1.

Jan. 5, mixed

6608 c.c.

1040

109-91

751-6 grms.

5-763

diet.

CASE 2.

Jan. 8, mixed

6474 c.c.

1041

94-08

633-0 grms.

5-545

diet.

Jan. 28, restrict-

3407 c.c.

1031

61-34

245-2 grms.

2-625

ed diet.

CASE 3.

June 8, mixed

5878 c.c.

1036

93-39

567'7 grms.

4-970

diet.

July 20, restrict-

2470 c.c.

1033

45-49

115 "8 grms.

2-789

ed diet.

CASE 4.

March 9, par-

1704 c.c.

1036

48-11

2 1-81 grins.

1-848

tially restricted

- diet.

June 28, parti-

852 c.c.

1034

31-76

14-40 grms.

1-543

ally restricted

diet.

In the course of diabetes mellitus there is apt to
occur a peculiar group of symptoms (first alluded to by
Prout, though only carefully studied of late years by
Kussmaul, Petters, Kaulich, Sanders and Hamilton),

1 The Croonian Lectures on certain points connected with Diabetes delivered at the
Royal College of Physicians, London, 1878.

Formation
of Acetone in
Blood. Aceton-

aemia.

CHAP. III.] THE BLOOD IN DISEASE. 1G9

which are included under the term diabetic coma. In reality, the first
symptoms are those of a very remarkable dyspnoea, in which there is
equal exaggeration of inspiratory and expiratory movements ; usually
it is only after this has existed for some hours that the patient, who
has been becoming more and more prostrate, sinks into a state of
coma and dies.

The peculiar ethereal smell exhaled by the breath of diabetics
had long been noticed, but it was Fetters who first pointed out that
in certain cases of diabetic coma, the apartment in which the patient
is confined acquires a peculiar odour, and that on distilling the urine
and even the blood of the patient, there is obtained a distillate which
contains traces of acetone.

Fetters based on these facts the theory that the phenomena of
diabetic corna depend upon a disengagement of acetone in the living
blood, that they are the symptoms, indeed, of a poisoning by acetone,
Acetonaemia.

That the blood in these cases does evolve acetone in small quan-
tities is proved by the concurrent testimony of several observers,
though it is very probable that the body does not exist free in the
blood, but is derived from the splitting up of ethyl-diacetic acid1.

There are however some serious objections to accepting the
acetonaemic theory of diabetic coma. In the first place, diabetics
sometimes evolve the most marked acetone (?) smell, without any of
the symptoms of diabetic coma being present ; in the second place, the
administration of very large doses of acetone is required in order to
produce any marked physiological symptoms, which even when pro-
duced, are by no means identical with those of diabetic coma.

It has been averred that when acetone is added to blood, there is
produced a white creamy appearance exactly similar to that which
has been observed, especially of late, in certain fatal cases of diabetic
coma. As has been shewn by Sanders and Hamilton, this statement is
erroneous. Acetone dissolved in alcohol and added in very small quan-
tities to blood, leads to no morphological change ; if added in larger
quantities it produces a coagulation of the proteids of the serum and
a solution of the coloured corpuscles, as has been shewn by Rupstein.

In a case of diabetes, which ultimately ended by coma, which
came under the Author's notice, the patient for some time evolved an
intense ethereal smell, which attracted the attention of patients in the
ward ; during the diabetic coma which preceded death, the acetone (?)
smell had diminished and the blood had only a faint smell of acetone.

1 C6H9Na03 + 2H20 = C8H60 + C2H60 + NaHC03.

Sodium ethyl- Water Acetone Alcohol Sodium hydric

diacetate Carbonate.

This is the equation by which Eupstein (Centralblatt, 1874, No. 55) explains the forma-
tion of acetone in the system from ethyl-diacetic acid.

In support of this theory is the fact that the urine of diabetics gives with Fe2 C16 a
reddish brown colouration which disappears on the addition of HC1, or on boiling —
properties which are possessed by ethyl-diacetic acid.

This subject will be discussed again in connection with the urine in diabetes.

170 THE BLOOD IN DIABETES MELLITUS. [BOOK I.

Whatever may be the part played by the acetone-like body
in the production of the phenomena of diabetic coma, we may safely
assert that when a diabetic exhales large quantities of that body,
the prognosis is peculiarly grave, the probability of a rapid fatal
termination being considerable. It is to be noted that the symptoms
of diabetic coma may set in and afterwards subside — a statement
which the Author bases upon a case observed and recorded by
Quincke1, and upon a second case observed by his friend Dr Grahame
Steell in the Manchester Royal Infirmary.

Lipaemic -^ ^as been stated (p. 59) that in a perfectly physio-

condition of logical condition, the serum of blood often presents a
the Blood in milky appearance which is due to the suspension of
Diabetes. fatty matters. Some of the older writers noticed that

the blood in diabetes is specially characterized by this lactescent
appearance ; the observations of Dr Babbington on this matter being
very precise8. The fact was, however, lost sight of for a long time,
or explained on the theory that diabetics consume large quantities of
food, and that as a result, their blood presents the appearance which
is usual whilst a full meal is being digested3. Recently investigated
cases* have directed attention afresh to this lipaemic condition of the
blood. It was observed by Dr Balthazar Foster5 that the blood in
certain fatal cases of diabetes presented a milky appearance, and he
averred that this was similar to the appearance produced on adding
acetone to blood, ether having no solvent action on the fat-like matter.
Professor Sanders and Dr Hamilton, in cases which they observed,
noticed that the blood had a pink colour, and that there separated
from it milk- or cream-like serum ; they however quite correctly
remarked that the milk-like appearance " proved to be due to oil, both
by microscopic examination, and by the removal of the milky appear-
ance by the action of ether, as well as by staining with perosmic acid.
Nothing identical with this can be produced by adding acetone to blood."
The interest attached to this lipaemic condition of the blood
depends upon the fact that Sanders and Hamilton discovered, in one
case where death had resulted from diabetic coma and where the
blood was intensely lipaemic, fat emboli in the vessels of the lungs and
kidney, the appearances being exactly similar to those observed in
cases of fat embolism from fractured bone. The resemblance of the
symptoms which were observed by Czerny in a case of fat embolism
due to this cause to those of diabetic dyspnoea and coma led Sanders
and Hamilton to advance the theory that " the peculiar terminal
dyspnoea and coma of diabetes are due to lipaemia and fat embolism,
rather than to acetonaemia."

1 Quincke, "Ueber Coma diabeticum." Berliner Minische Wochenschrift, 1880, No.l.

2 See Article "Blood" by Babbington in Todd's Cyclopaedia of Anatomy and
Physiology, Vol. i. p. 422.

3 Pavy, Researches on the Nature and Treatment, of Diabetes, London, 1862, p. 1

4 Sanders and Hamilton, "Lipaemia and Fat Embolism in the fatal Dyspnoea
Coma of Diabetes." Edinburgh Medical Journal, July 1879, p. 47.

5 Foster, "Diabetic coma. Acetonaemia." British Me d. Journal, 1878, Vol. i. p. 78.

CHAP. III.] THE BLOOD IN DISEASE. 171

Two cases which the Author has had the opportunity of studying
do not, however, support this fascinating theory. In one of these
cases the amount of fat in the blood was exceedingly great, yet
a most scrupulous investigation of the lungs, the kidneys and the
brain, conducted by Dr Dreschfeld, led to the conclusion that no
emboli were present. In the second case the amount of fat in
the blood (or rather the amount of matters soluble in ether) was
not larger than usual, and in this case also a most painstaking search
shewed the absence of emboli.

Quincke, who has rejected the 'Acetonaemia theory' of diabetic
coma, is inclined to consider it as a condition which, like uraemia,
is probably induced by a combination of circumstances, and by
the toxic action of more than -one product of tissue metabolism,
amongst which may be the body which is excreted in the urine, and
is coloured red by perchloride of iron. Whatever may be the toxic
agent or agents, it is difficult to see cases of diabetic coma without
coming to the conclusion that the condition is one due to a toxic
action and not to a suddenly-developed nervous lesion. As bearing
upon this question it is worthy of rnention that in the first of the two
cases of which the notes are given below, the liver was found, after
death, to be the seat of intense fatty infiltration, similar to that
observed in cases of poisoning by phosphorus.

The following .are brief notes of two cases previously referred to, in
which the Author has had the opportunity of examining the blood of
patients suffering from diabetic dyspnoea and coma.

I. X, a man of about 35 years of age, a patient in the Manchester Royal
Infirmary, under the care of Dr Roberts, F. R. S., had been suffering from
diabetes of two and a half years duration. Since his admission into the
hospital his urine had amounted to 300 ounces per diem with a specific
gravity of 1030-1035. The patient exhaled a peculiar ethereal (acetone-
like1?) odour which pervaded the whole ward and attracted the attention
of the other patients. On 9th July, 1879, after returning from a walk, the
patient was seized with intense dyspnoea ; the exaggeration of inspiratory
and expiratory movements was equally marked ; there was no evidence of
venosity of blood ; the exaggerated respiratory movements continued uni-
formly without any rhythmical variation in intensity. At first the patient
was conscious, but he subsequently became comatose, and died 21 hours
after the commencement of the attack.

During the attack of coma the blood was examined microscopically
without any deviation from the normal appearance being noticed. Some
blood was drawn from tfye arm by venesection ; it coagulated normally,
and there separated from it a serum distinctly milky, though not more so
than is compatible with a physiological condition. At the post-mortem
examination a considerable quantity of blood was collected from the cavity
of the chest. The broken-up clot mixed with serum was placed in a
bottle. After some hours a creamy layer had floated to the surface of
the liquid, this layer being about one-sixth of the total volume of ^ the
liquid. The milkiness was found to depend upon oil globules of various sizes.

172 THE BLOOD IN DISEASES OF THE KIDNEY. [BOOK I,

Aii analysis of the blood drawn during life and of that collected after
death gave the following results : — •

Blood drawn Blood collected

during life. after death.

Water in 1000 parts 744-6 757-7

Total solids „ 2554 242-3

I -g (Neutral Fats l-ina 9'86)

I 1 ^Lecithin j L 1-55V 13-55
gfg (Cholesterin 1'96 2-14)

II. J, T., a man 32 years of age, was admitted into the Royal Infirmary
on 3rd Nov. 1879, suffering from diabetes. He appeared very ill and
exhaled a very intense ethereal odour. At this time there were no
symptoms of dyspnoea or coma. The urine contained sugar and was of
high specific gravity. On the night after his admission he was seized with
purging, and in the morning he appeared very ill; his breathing then
became laboured and had the characters of diabetic dyspnoea. In the
course of the day he became unconscious, and he died shortly after midnight
on the 5th Nov. The urine which was passed on the 4th contained, besides
sugar, some albumin and many hyaline casts.

At the post-mortem examination the blood was found to possess the
acetone (?) like odour. Some was collected and analysed with the following
results : —

Ethereal extract of 1 000 parts of Blood 1-88 parts.
Cholesterin contained in ethereal extract 0'642 „

The amount of the ethereal extract obtained from the blood in case I. was
much larger than has been found in any published analyses of human blood.
Thus the mean amount of fat (including under this term all the constituents
of the ethereal extract of blood, viz. neutral fats, cholesterin and lecithin)
found by Becquerel and Rodier in. their numerous analyses was 1 -6 parts,
the maximum being 3'25 and the minimum 1-00 per 1000 parts of blood.

In published analyses of the blood of diabetics by 0. Schmidt, the
amount of fat was respectively 1-82 and 2-13 per 1000; in these cases
however, there was no diabetic coma. Hoppe-Seyler l also mentions that he
found the proportion of fat materially increased in the blood in four cases
of diabetes.

G. THE BLOOD IN DISEASES OF THE KIDNEY.

There is probably no class of diseases in which a change in the
chemical composition of the blood is so soon induced as in Bright's
disease, or exerts a more marked influence upon the exchanges of the
matters of the organism.

The fundamental knowledge which we possess on this subject was
mainly acquired by the classical investigations of Christison a which

1 Hoppe-Seyler, Physiologische Chemie, p. 482.

3 Christison, "Observations on the variety of Dropsy which depends on diseased
kidney." Edinburgh Med. and Surg. Journal, Vol. 32 (1829), p. 262.

Christison, "On granular degeneration of the kidneys, and its connection with
Dropsy, Inflammation and other diseases." Edinburgh, 1839.

CHAP. III.] THE BLOOD IN DISEASE. 173

followed very closely the masterly memoir in which Dr Bright1 had
first announced the connexion between albuminuria and morbid
changes in the kidney.

Christison pointed out that in the early stages of kidney disease
the blood presents the following characters ; the density of the serum
is low (1020 or even 1019), the proportion of albumin diminishes,
the fibrin of the blood may be increased, the proportion of blood-
colouring matter is unaffected, but, above all, the serum frequently con-
tains urea. He shewed that as the disease became chronic some of
these changes in the blood became less distinct, e.g. the diminution
in the amount of albumin, and the presence of urea, but that a very
constant and considerable diminution of the blood-colouring matter
was a characteristic occurrence.

Subsequent researches have thoroughly confirmed the statements
of Christison as to the excess of urea which is present in the blood of
Bright's disease. Although there is no longer any difference of opinion
as to the accumulation of urea in the blood in cases of Bright's disease
in which there is a marked suppression of urine or a very obvious
deficiency in the elimination of urea, facts have hitherto been
wanting to decide whether after the establishment of any of the
chief lesions of the kidney there is a permanent impairment of the
normal power which the kidneys possess of eliminating urea. This
question has occupied the attention of the Author, and he is inclined
to believe that in most cases of chronic Bright's disease, even whilst
the patient is in the apparent enjoyment of fair health, there is a per-
sistent excess of urea in the blood.

The convulsions and coma which are apt to supervene when the
elimination of urea is defective have been designated as 'uraemic'
or as evidences of ' uraemic ' poisoning. At first it was held as
certain that these nervous phenomena were occasioned by the
accumulation of urea in the blood acting as a poison on the
great nerve centres. There can be no doubt, however, that this
simple explanation is not sufficient ; the condition of uraemia is
one which depends upon many factors. It must not be for-
gotten, that before the condition of uraemia is induced, the blood
has usually become rich in water, poor in albumin, poor in corpuscles,
and that in addition to an accumulation of urea and uric acid it
probably contains an excess of other proximate principles which may
exert a specially poisonous action.

It was suggested by FrerichsHhat uraemic phenomena are due to
the conversion of urea into ammonium carbonate in the blood,
but there is no ground for believing that such a conversion actually
does occur during life.

1 Bright, " Cases illustrative of some of the appearances observable on the examina-
tion of diseases terminating in dropsical effusion, — and first of the kidney." Bright's
Beporte. London, 1827.

2 Frerichs, Archiv f. phys. HeilJc., 1852, Vol. xi. p. 88.

CHAPTER IV.

THE BLOOD (continued).

DESCRIPTION OF CERTAIN METHODS OF EESEARCH.
Determination of the Specific Gravity of the Blood.

EXCEPT by operating with extreme expedition and at temperatures
below 0° C. it would be quite impossible to determine the specific
gravity of the uncoagulated blood. It is obvious, however, that the
specific gravity of defibrinated blood can only differ very slightly
from that of blood which has not yet coagulated. It is therefore
usual to take the specific gravity of defibrinated blood as sufficiently
representing that of the blood.

In the case of blood, this is best done with the aid of a specific
gravity bottle, of which two forms are represented in the annexed
woodcuts (Figs. 28 and 29).

The flask is first weighed when empty ; then when filled with
distilled water at a known temperature ; the distilled water being
then poured out and the flask dried, the bottle is filled with
defibrinated blood at the same temperature as the water, and again
weighed. By these operations we ascertain the weight of the water
and of the blood respectively which at a given temperature are con-
tained in the bottle.

Let a be the weight of the water contained in the flask, and b that
of the defibrinated blood ; then the specific gravity of the latter,
designated by d, will be

a '

It is however very inconvenient to be obliged to weigh liquids at one
particular temperature, and if we have at our disposal a specific gravity
bottle such as is represented in fig. 29, arid a table of the density of water
at various temperatures, we can readily ascertain the specific gravity of the
blood, though its temperature be not exactly the same as that of the water

CHAP. IV.] THE BLOOD. METHODS OF EESEARCH.

175

which was weighed. Let us assume that we have found our bottle to hold
25-6515 grams of water at 15°, and 2 7 '2 70 grams of defibrinated blood at
7° C. ; we must first calculate the weight of water which would be con-

FIG. 28. SPECIFIC GRAVITY BOTTLE,
consisting of a light flask with a well-
ground perforated stopper.

FIG. 29.

GEISSLER'S SPECIFIC GRAVITY
BOTTLE.

a ig & ^ flagt & a yery accurately

graduated thermometer, c is a tube con-
nected with a, through which fluid escapes
when the thermometer is inserted in the
bottle, d a cap which fits perfectly the
top of c and which is applied to it after it
is filled.

tained if the temperature had been 7° and not 15°. We find on looking at
the subjoined table of the specific gravities of water that whilst the specific
gravity of water at 15° is 0'99915, at 7° it is 0*99994, and we can get the
weight of water which our bottle would hold at 7° by the following
proportion :

0-99915 : 0-99994 :: 25-6515 : x
_ 0-99994x25-6515 _

0-99915 -^o'-

176

DETERMINATION OF WATER, SOLIDS, AND ASH. [BOOK I.

Knowing the weight of equal volumes of water and blood at the same
temperature we can at once get the density as before by dividing the
latter by the former :

TABLE OF THE DENSITY OF WATER AT TEMPERATURES BETWEEN

0° AND 30° C.

Temp.

Density

Temp.

Density

0°

0-99988

16°

0-99900

1°

0-99993

17°

0-99984

2°

0-99997

18°

0-99866

3°

0-99999

19°

0-99847

4°

1-00000

20°

0-99827

5°

0-99999

21°

0-99806

6°

0-99997

22°

0-99785

7°

0-99994

23°

0-99762

8°

0-99988

24°

0-99738

9°

0-99982

25°

0-99714

10°

0-99974

26°

0-99689

11°

0-99965

27°

0-99662

12°

0-99955

28°

0-99635

13°

0-99943

29°

0-99607

14°

0-99930

30°

0-99579

15°

0-99915

Determination of the Reaction of Blood.

As was stated at p. 26, the reaction of blood cannot be accurately
ascertained by immersing into it ordinary test-papers, but by following
one of the methods suggested by Kuhne, Zuntz, or Liebreich. With
the aid of one of these methods the amount of a standard acid re-

?uired to neutralize a given volume of blood may be determined,
t is essential, however, to employ a carefully prepared solution of
litmus, free from alkali. With this object 16 grammes of com-
mercial litmus are finely pulverized, and the powder is mixed in a
beaker with 120 c.c. of water and frequently stirred. After 24 hours
the solution, which contains nearly all the free alkali of the litmus, is
thrown away and the residual litmus is again treated with 120 c.c.
of water for 24 hours. The solution thus obtained is divided into
two equal portions ; the one is carefully treated with a little
very dilute acid, added by means of a glass rod, until a red tint
just appears, and then to this is added some of the other portion,
until a little of the fluid, when much diluted, presents a blue-
violet colour. .

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH.

177

If a plaster of Paris slab (see p. 26) be imbued with such a solution
of litmus, a drop of blood or of blood-serum will be surrounded
at its edges by a distinct blue ring. In order to determine the degree
of alkalinity, a standard solution of tartaric acid may be made
by dissolving 7'5 grammes of crystallized tartaric acid in 1000 c.c.
of water ; one cubic centimetre of this solution should exactly
neutralize 0'004 grm. of NaHO. The acid solution is added
from a burette to 50 or 100 c.c. of the serum or blood, a drop of
the mixture being placed from time to time upon the slab coloured
with litmus ; the addition of acid is continued until the reaction is
faintly acid. The alkalinity of the blood may then be expressed as
corresponding to x milligrammes of sodium hydrate per 100 c.c. of
blood.

Determination of the Water, Total Solids and Ash of the Blood.

A Berlin porcelain crucible, furnished with a cover and having
a capacity of about 20 c.c., is dried and then accurately weighed.
From 2'5 to 5 grammes of defibrinated blood are carefully weighed
out in the crucible, which is then placed in a hot- water oven
heated to 100° C. (Fig. 30), until an apparently dry residue is left ;
the crucible is then heated in a hot-air oven furnished with a
regulator (Fig. 31), and kept at a temperature of 110° C. After
some time the crucible is transferred to an exsiccator (Fig. 32 or
33), where it is allowed to remain for a few minutes to cool,

FIG. 30. HOT-WATER OVEN WITH ARRANGEMENT FOR KEEPING THE WATER AT A CONSTANT

LEVEL.

G. 12

178

DETERMINATION OF ASH OF BLOOD.

[BOOK i.

in dry air, it is then placed on the balance and weighed. The
weight having been noted, the crucible is again heated to 110° for some

FIG. 31. Hoi-AiR OVEN WITH BDNSEN'S BEGULATOR.

time, and again weighed as before, the process being repeated until
two successive weighings give the same result.

FIG. 32. AN EXSICCATOR.
A bell- jar 6, with ground rim, fits
air-tight over the plate a. c is a vessel
containing sulphuric acid or phosphoric
anhydride, d is a tray with circular holes
for crucibles and capsules.

FIG. 33.

A SMALL EXSICCATOR, SUITABLE

FOR A CRUCIBLE.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 179

The crucible is now uncovered and placed, upon a triangle, over a
Bunsen flame, so as to char its contents. At first the application of
heat is conducted with much caution, the flame being at a con-
siderable distance from the bottom of the crucible ; if this precaution
be not taken the contents are very apt to froth up and to be partially
lost. Soon, however, the heat may be increased by placing the
crucible so that it is surrounded by the flame, and it will prove
advantageous to place the crucible in the tilted position indicated

FIG. 34. ARRANGEMENT EMPLOYED IN DETERMINING THE AMOUNT OF ASH IN BLOOD.

in the annexed figure (Fig. 34). If conducted in a porcelain crucible
the process is a very slow one. When the carbon has been entirely
burnt away, the ash presenting a reddish-white colour without
intermixture of black, the crucible is cooled in the exsiccator and
weighed. This method of determining the ash does not possess
great value, for the reasons already referred to at considerable length
(see p. 66). The following process should be followed when it is
desired to attain as great accuracy as is compatible with the
method of incineration.

Rose's me- The dried residue of the blood is heated over a

Bunsen flame until it is thoroughly carbonized ; care is
however taken that the crucible does not become even faintly red.
Having been allowed to cool, the contents of the crucible are treated
with boiling distilled water and heated for some time ; the aqueous
solution is filtered through a small filter of Swedish filter paper,
and kept. The carbonized residue is treated again and again with
hot distilled water, to make sure of dissolving all soluble salts.

12-2

180 DETERMINATION OF FIBRIN. [BOOK I.

The insoluble matters together with the small filter previously
referred to, are now dried in the hot-air oven and then ignited
at a red heat ; when the whole of the carbon has been burned
away the crucible is cooled and the solution of the soluble salts
added to it; the contents are first evaporated to dryness in the
water and air ovens and then ignited at a barely perceptible red
heat; the crucible is then cooled and weighed, and thus the total
amount of ash found. Or, by weighing in separate crucibles the
aqueous solution and the ignited insoluble residue, the amount of
soluble and insoluble constituents of the ash is ascertained *.

Determination of the amount of Fibrin yielded by the Blood.

If it be desired to determine the amount of fibrin which will
separate from the blood, the best method is the following :

Hoppe- A beaker, of a capacity between 100 and 150 c.c., is

Seyier's fitted with a caoutchouc cap, provided with a single opening

method. in ^he centre through which is thrust a rod of vulcanite,

somewhat spatula-shaped at its lower end (Fig. 35). The weight of

FIG. 35. HOPPE-SEYLEP.'S APPABATUS FOB SEPARATING FIBRIN FROM THE BLOOD.

1 The most valuable determinations of the salts of the serum have been made by
the method of direct precipitation. It has been shewn that sulphuric and phosphoric
acids, calcium, and magnesium may be precipitated from serum, as from aqueou
solutions ; the precipitates are separated by subjecting the liquid to rapid rotation in th
centrifugal machine; with the aid of the latter they may be efficiently and rapidly
subjected to the process of 'washing by decantation ', and then treated according tc
the ordinary methods. The reader who wishes to pursue researches in this directic
should consult the account given of these methods by Pribram and Gerlach (see p. 6C

CHAP. IV.] THE BLOOD. METHODS OF HESEARCH. 181

the apparatus is determined when empty. The caoutchouc cap having
been momentarily withdrawn, 30 or 40 c.c. of uncoagulated blood are
allowed to flow into the beaker ; the cap is replaced and the blood is
stirred with the little spatula, until the fibrin has separated. The
apparatus is then weighed. This operation being completed, the
caoutchouc cap is removed, the beaker filled with distilled water,
and the contents stirred by the aid of the spatula which is left in situ.
When the fibrin has subsided the red supernatant liquid is decanted.
The beaker is then filled up again with a 1 — 3 per cent, solution of
common salt, the contents again stirred and allowed to subside.
These operations are repeated until the fibrin is almost colourless ;
the beaker is then filled up once or twice with distilled water,
the water decanted, and then the fibrin is transferred to a small
weighed filter, washed with boiling alcohol, and then dried in the hot-
water oven at 100° C., or preferably in the hot-air oven at 110° C.; the
filter and its contents are then placed between two, weighed, ground

FIG. 36. WATCH-GLASSES, WITH CLIP, IN WHICH FILTEKS ARE ALLOWED TO COOL AND

WEIGHED.

watch-glasses (Fig. 36) held together by a clip, and weighed. On
subtracting the weight of the watch-glasses, clip, and filter-paper from
the total weight found, the weight of the fibrin contained in the
amount of blood analysed is ascertained.

Determina- if it be required to determine the amount of fibrin

iltedBi^T1" in blood which has already coagulated, the total weight
of the blood having been determined by weighing, the
whole is thrown upon a filter made of well- washed calico. When
the serum has drained through, the filter is tied so as to enclose
the clot in a little bag. This is then kneaded between the fingers,
whilst a stream of water is allowed to play upon it continuously.
After long-continued washing, the whole, or nearly the whole, of the
coloured corpuscles having been removed from the clot, the cloth is
opened, when it is found to contain filaments of fibrin, which are
more or less completely decolorized ; these are collected by means
of a pair of forceps, and transferred to a small beaker, washed with
weak salt solution, and afterwards with water, and treated as in
Hoppe-Seyler's method.

182 DETEKMINATION OF HAEMOGLOBIN. [BOOK I.

Determination of Haemoglobin in the Blood.

i. Hoppe- This method consists in comparing the tint of the

blood diluted with a known volume of distilled water,

with the tint of a solution of pure haemoglobin of

known strength, and then adding water to the first until it assumes

exactly the colour of the second.

This method necessitates in the first place a solution of pure Oxy-
haemoglobin. Oxy-haemoglobin, prepared from the blood of the dog
and at least twice crystallized, is dissolved in water at 0° C., and the
saturated solution is filtered. 50 c.c. are measured out in a capsule
and evaporated to dryness, first over a water-bath and then over
sulphuric acid, in vacua. In this way the strength of the solution is
determined.

In addition to this solution there are needed two haematin-
.ometers1 (see Fig. 16, p. .92), and an accurate burette divided into
tenths of a cubic centimetre.

The two haematinometers being placed side by side, with a sheet
of white paper beneath them, and in such a position as to be
illuminated in exactly the same manner, 10 c.c. of the standard
solution, diluted with from 10 to 60 c.c. of water, are placed in the one
haematinometer.

A solution, of known strength, of the blood to be investigated is
now made, e.g. by diluting 5 grammes of blood to 100 c.c.; 10 c.c.
of the solution are placed in the second haematinometer.

The solution of blood will now be seen to be very much deeper
in tint than the solution of haemoglobin. Water is added to the
former, from a burette, until there is no perceptible difference in tint
between it and the standard solution. When this result has been
obtained, the amount of haemoglobin in the two solutions must
be equal.

The method and the calculation required will be understood from the
following experiment quoted from Hoppe-Seyler : — 20 '186 gnus, of denbrin-
ated blood were diluted with water to a volume of 400 c.c. 10 c.c. of
this solution were placed in a haematinometer and 38 c.c. of water had to
be added so as to produce a solution of the same shade as the standard
solution of haemoglobin which had been placed in a second haematinometer.
The volume of water which the whole quantity of the solution of blood
would have required to bring it to the standard tint is found by the
proportion

10 : 38 :: 400 : x
# = 1520 c.c.

1 Haematinometers similar to the one represented in Fig. 16 are constructed by
the opticians Schmidt u. Haensch, Stallschreiber-Strasse 4, Berlin ; they are sold in
pairs, at 30 marks (£1. 10s.) the pair.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 183

By adding then 1520 c.c. of water to 400 c.c. of the diluted blood we
should obtain 1920 c.c. of a solution equal in shade and intensity of colour
to the standard solution of haemoglobin. But on analysis the latter was
found to contain in 100 c.c. 0*145 grms. of haemoglobin; we now have the
data for determining the total quantity of haemoglobin in the diluted
blood :

100 : 0-145 :: 1920 : x

x — 2784 grms.

But as this quantity of haemoglobin was derived from 20*186 grms.
of blood, the amount contained in 100 parts is found thus:
20-186 : 2-784 :: 100 : x
x= 13*79 grins.

This method gives most accurate results ; its use was formerly
deprecated, inasmuch as it requires solution of pure oxy-haemoglobin,
which cannot easily be prepared except in the depth of winter, and
which when prepared will not keep more than a few days in the
open air. Hoppe-Seyler has however shewn that solutioDS of
pure haemoglobin in sealed glass tubes may be kept indefinitely
without the haemoglobin undergoing decomposition. Nothing can
therefore be easier than to make a stock of pure solution of oxy-4
haemoglobin in winter and store it in a large number of sealed
glass tubes for use during the succeeding year. The oxy-haemoglo-
bin is soon reduced to haemoglobin, but, after that, resists all further
change. When the tubes are opened the solution rapidly absorbs
oxjgen and a solution of oxy-haemoglobin is obtained.

Instead -of employing a standard solution of oxy-haemoglobin, we
may, as suggested by Kajewsky1, use, as a standard, a solution of
picro-carminate of ammonia, corresponding in tint to a solution of
haemoglobin of known strength. A solution of picro-carminate, if per-
fectly neutral, may be preserved in stoppered bottles for long periods ;
according to Malassez2 it may be kept indefinitely.

The following is the method of preparing a solution of picro-carminate of
ammonia.

Take 100 c.c. of a saturated solution of picric acid. Prepare an
ammoniacal solution of carmine by dissolving 1 grm. in a few c.c. of water,
with the aid of an excess of ammonia and heat. Boil the picric acid, and
when boiling add the carmine solution. Evaporate the mixture to dryness,
and dissolve the residue in 100 c.c. of water, and filter. A clear solution
ought to be obtained ; if not, add some more ammonia, evaporate, and then
dissolve as before3. This solution is now added, little by little, to a
mixture of equal parts of water holding a little phenol in solution, and of
glycerine, until the tint, observed in a haematinometer, is exactly similar to

1 A. Rajewsky, "Zur Frage iiber die quantitative Bestimmung des Hamoglobin-
gehaltes im Blut.'" Pfluger's Archiv, Vol. xn., p. 70.

2 Malassez, " Sur les diverges methodes de dosage de I'hemoglobine et sur un
nouveau colorimetre." Archives de Physiologic, 1877, pp. 1 — 13.

3 Rutherford, Outlines of Practical Histology, p. 173.

184 DETERMINATION OF HAEMOGLOBIN. [BOOK I.

that of a standard solution of haemoglobin. As the picro-carminate has a
yellower shade than blood, it is advisable to add to it a small quantity of a
perfectly neutral solution of carmine.

2. Preyer's This method requires (1) at least one haematino-

meter, (2) a spectroscope, (.3) a steady light, (4) a standard
solution of haemoglobin, (5) a finely-divided burette.

The haematinometer being placed between the luminous source
and the spectroscope, a strong solution of crystallized oxy-haemoglobin
in water is poured into it. If the solution be very strong, as was
mentioned at p. 97, only the red rays will pass. Water is now
added very gradually from the burette, until the first gleam of green
between E and F, and close to b, is perceived. In order to make the
appreciation of this more easy, the experiment is conducted in a
darkened room, and the lamp is furnished with a shade (as in Fig. 18)
which only allows the rays of light to proceed in the direction of the
spectroscope. The amount of haemoglobin in the standard solution
is now ascertained as in Hoppe-Seyler's method, by evaporating a
known volume to dryness. (Preyer has found that when examined in
a haematinometer of which the sides are 1 centimetre apart, a
solution of haemoglobin containing 0'8 grms. of the substance per
cent, just allows a narrow band of green close to b to appear.)

About 0'5 c.c. of the blood, of which the amount of haemoglobin is
to be determined, is now poured into a haematinometer, and water
added to it from a burette until, when examined under exactly the
same circumstances as the standard solution, the green close to b just
appears. The amount of water added must be very precisely
measured. The amount of haemoglobin contained in the blood is
then found by the following equation :

Hb(e + s)

where Hb is the weight of haemoglobin contained in 100 c.c. of the
standard solution, e the volume of water added to the blood analyzed,
and s the volume of the latter.

Dr cowers' The tint of the dilution of a given volume of blood

toe SJLfOI> with distilled water is taken as the index to the amount
estimation of °^ haemoglobin. The distilled water rapidly dissolves
Haemogio- out all the haemoglobin, as is shewn by the fact that
bin1. the tint of the dilution undergoes no change on

standing. The colour of a dilution of average normal blood one
hundred times is taken as the standard. The quantity of haemoglobin
is indicated by the amount of distilled water needed to obtain the
tint with the same volume of blood under examination as was taken
of the standard. On account of the instability of a standard dilution
of blood, tinted glycerine-jelly is employed instead. This is perfectly
stable, and by means of carmine and picro-carmine the exact tint of
diluted blood can be obtained.

1 See "Report of the Meeting of the Clinical Society," the Lancet n. 1878, p. 822.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 185

The apparatus consists of two glass tubes of exactly the same size.
One contains a standard of the tint of a dilution of 20 cubic mm. of
blood in 2 cubic centimetres of water (1 in 100).

The second tube is graduated, 100 degrees = two centimetres (100
times twenty cubic millimetres).

The twenty cubic millimetres of blood are measured by a capillary
pipette (similar to, but larger than that used for the haemacytometer).
The quantity of the blood to be tested is ejected into the bottom of
the tube, a few drops of distilled water being first placed in the
latter. The mixture is rapidly agitated to prevent the coagulation
of the blood. The distilled water is then added drop by drop (from
the pipette-stopper of a bottle supplied for that purpose) until the tint
of the dilution is the same as that of the standard, and the amount of
water which has been added (i.e. the degrees of dilution) indicates the
amount of haemoglobin.

Since average normal blood yields the tint of the standard at 100
degrees of dilution, the number of degrees of dilution necessary to
obtain the same tint with a given specimen of blood is the percentage
proportion of the haemoglobin contained in it, compared to the
normal.

For instance, the twenty cubic millimetres of blood from a patient
with anaemia gave the standard tint at 30 degrees of dilution. Hence
it contained only thirty per cent, of the normal quantity of haemo-
globin.

By ascertaining with the haemacytometer (p. 77) the corpuscular
richness of the blood, we are able to compare the relation between
the number of corpuscles and the amount of haemoglobin. A frac-
tion, of which the numerator is the percentage of haemoglobin, and
the denominator the percentage of corpuscles, gives at once the
average value per corpuscle. Thus the blood mentioned above,
containing thirty per cent, of haemoglobin, contained sixty per
cent, of corpuscles ; hence the average value of each corpuscle
was fj or i of the normal. Variations in the amount of haemo-
globin may be recorded on the same chart as that employed for
the corpuscles.

In using the instrument the tint may be estimated by holding the
tubes between the eye and a window, or by placing a piece of white
paper behind the tubes ; the former is perhaps the best. Care must
be taken that the tubes are always held in the line of light, not below
it. In the latter case some light is reflected from the suspended
corpuscles from which the haemoglobin has been dissolved. If the
value of the corpuscles is small, then a perceptibly paler tint is seen
when the tubes are held below the line of illumination. If all the
light is transmitted directly through the tubes the corpuscles do not
interfere with the tint.

In using the instrument it will be found that between six or
eight degrees of dilution it is difficult to distinguish a difference
between the tint of the tubes. It is therefore necessary to note the

18G

DETERMINATION OF HAEMOGLOBIN.

[BOOK I.

degree at which the colour of the dilution ceases to be deeper than
the standard, and also that at which it is distinctly paler. The degree
midway between these two will represent the haemoglobin percentage.
The instrument is only expected to yield approximate results,
accurate within two or three per cent. It has however been found of
much utility in clinical observations1.

FIG. 37. DK GOWERS' APPAKATUS FOE THE CLINICAL ESTIMATION OF HAEMOGLOBIN.

E, block of wood with two holes, to serve as a stand for the tubes C and D.
D, tube containing glycerine- jelly treated with picro-carmiue.

C, graduated tube in which the blood is diluted with water.

B, capillary pipette marked so as to allow of 20 cubic millimetres of blood being
measured.

F, lancet-shaped needle for puncturing the finger. The point of the needle may be
protruded to a greater or less extent, so as to produce a more or less deep puncture.

A, bottle for distilled water.

8. By the
determina-
tion of iron
contained in
the ash of the
blood.

As it is known that pure haemoglobin contains 0'43
per cent, of Fe, and as all the iron in the blood exists in
haemoglobin, we may calculate the amount of this con-
stituent if we know how much iron is contained in the
ashes of a known volume or weight of blood.
With this object about 100 grammes of blood are evaporated to
dryness in a platinum basin, and then ignited, care being taken not
to lose any portion of substance by incautiously heating, which would
cause the burning organic matter to froth over. The ash is cooled
and boiled with 10 or 20 c.c. of pure hydrochloric acid diluted with
its own volume of water, and the extraction is repeated. The solution
of ferric chloride in then reduced by the action of metallic zinc, and

1 Dr Gowers' apparatus is sold, under the name of Haemoglobinometer, by Mr
Hawkesley, Surgical Instrument Maker, 300, Oxford Street, London, W.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 187

the amount of iron determined with the aid of a standard solution of
potassium permanganate. By multiplying the amount of metallic
iron in 100 parts of blood by 100 and dividing by 0'43, we obtain the
percentage of haemoglobin.

For the details of the method of determining the iron volumetri-
cally we refer the reader to any systematic work on Quantitative
Analysis.

Determination of Cholesterin, Lecithin, and Fats in Blood (Hoppe-

Seyler).

20 to 50 c.c. of blood are treated with 3 or 4 times their volume
of absolute alcohol, set aside for a few hours, and then filtered.
The insoluble matter is washed, first with pure alcohol and then with
alcohol holding ether in solution.

The mixed alcoholic and ethereal solutions are evaporated to
dryness on the water-bath. The residue is dissolved in ether, filtered,
evaporated to dryness, and weighed.

In this way is obtained the combined weight of the cholesterin.
lecithin and neutral fats.

The residue, after being weighed, is then treated with alcohol,
and a little alcoholic solution of caustic potash added, and heated, in
a silver dish, for some hours in the water-bath, until the whole of the
alcohol is expelled. The residue contains cholesterin, soaps, glycerin,
caustic potash and products of the decomposition of lecithin, viz.
glycerin-phosphoric acid, neurin, &c.

The residue is then mixed with water and agitated repeatedly
with ether. The ethereal solution is evaporated to dryness, and
dissolved in absolute ether, which dissolves the cholesterin alone,
leaving undissolved traces of soaps, which were mixed with it. The
ethereal solution is evaporated at a low temperature, then dried below
80° C. and weighed.

The watery solution, from which the cholesterin has been removed
by ether, is evaporated to dryness in a silver dish and fused with
sodium hydrate and pure nitre. The fused mass is dissolved in water,
and treated with an excess of nitric acid.

The phosphoric acid is then precipitated by means of an acid
solution of ammonium molybdate. The precipitate, which has
separated after 12 hours, is dissolved in ammonia and the solution
precipitated by magnesia mixture, the precipitate being washed,
dried, ignited and weighed as magnesium pyrophosphate.

100 parts of magnesium pyrophosphate correspond to 764 '5 parts
of lecithin.

For full directions how to estimate the phosphoric acid in the
above case, the reader is referred to works on quantitative analysis.

188 DETERMINATION OF PROTEIDS OF BLOOD. [BOOK I.

Determination of Water, Total Solids, and Salts of the Serum.

As the processes are exactly similar to those mentioned at page
177, they require no further notice here.

Determination of the total amount of Proteids contained in the Serum,
and of the Serum-albumin.

Precipita- 5Q to 100 c.c. of water are boiled, and an accu-

tionbyheat. rately weighed amount of serum (about 15 or 20
grammes) poured in. The fluid is boiled for some minutes, a drop
or two of very dilute acetic acid cautiously added with a glass rod,
until the precipitate separates in flakes from a perfectly clear (i.e.
not opalescent) liquid ; the precipitate is collected on a weighed
filter, washed with water, then with boiling alcohol, dried at 110° C.
and weighed. By this process all the proteids of the serum are
precipitated together ; by subtracting the weight of paraglobulin as
determined by other methods, that of serum-albumin is found.

Precipita- a. Hop pe-Seyler's method \

tionby ^n accurately measured or weighed quantity of

alcohol. rtn J . - i • i i -,! ,1

serum, say 20 grammes, is mixed in a beaker with three

or four times its volume of spirits of wine, and set aside at the
ordinary temperature for some hours; the precipitate is then col-
lected on a weighed filter free from ash, and washed, first of all
with spirits, then with hot absolute alcohol, then with ether, and
lastly with warm water.

There are thus left on the filter only proteids and insoluble salts.
The filter is washed with spirit so as to displace the water, and is
then dried at 120°, allowed to cool in an exsiccator, weighed, &c.
The filter with its contents is then ignited, and the weight of ash
deducted from that of the proteids. This method is of universal
application to albuminous fluids, and is useful as it enables one to
obtain in one operation, not only the amount of proteids, but
alcoholic and ethereal extracts, in which other constituents may be
determined.

6. Schmidt's method 2.

A weighed portion of serum is neutralized with acetic acid, mixed
with ten times its volume of strong alcohol, set aside for 24 hours, and

1 Hoppe-Seyler, Handbuch der^physiologisch- und pathologisch-cliemisclien Analyse,
3e Aufl., p. 312.

2 A. Schmidt, "Weitere Untersuchungen des Blutserums, &c." Pfluger's Archiv,
Vol. xi. p. 10 (1876). This method has lately been subjected to examination by Prof.
F. A. Hoffmann of Dorpat, who found it reliable. (Virchow's Archiv, Nov. 1879,
p. 255.)

CHAP. IV.] THE BLOOD. METHODS OF KESEARCH. 189

boiled. The flocculent precipitate is then collected on a filter and
washed with a mixture of 10 parts of alcohol and 1 part of water,
then with absolute alcohol and ether. It is then treated as the
precipitate obtained by process a.

Determina- This can only be done in the case of the serum

being absolutely colourless and transparent.

^he amount of albumin per gramme of solution will
be found by the formula

a

"^Wxl

where a indicates the rotation observed and I the length of the tube.
The results are however only approximative.

Determination of the amount of Fibrinogen contained in Liquor
Sanguinis. (Frederique's method a.)

If it be desired to determine the amount of fibrinogen in liquor san-
guinis, the liquid, which has been kept from coagulating by exposure
to cold, or by the addition of magnesium sulphate, is heated to 60°;
the precipitate is allowed to subside, washed by decantation with a £
per cent, solution of sodium chloride, and then thrown on a small
weighed filter. The filter is washed with distilled water, then with
alcohol, dried at 110° C., cooled under an exsiccator, and weighed
between watch-glasses. The filter and contents may be afterwards
ignited and the weight of ash deducted.

Determination of the amount of Serum-globulin in the Serum.

Two methods may be employed, which give however entirely
different results; the second is alone reliable.-

Schmidt's a. Serum is placed in a dialyser, the water around

method. which is frequently renewed. After about 48 hours a

stream of C02 is passed through the contents of the dialyser. The
precipitate which has formed is collected on a weighed filter,
washed, dried, and weighed.

Hammar- ^ 5 C-C> of serum are diluted with 25 c.c. of satu-

iethod3 ratec* somtion of magnesium sulphate and then treated

with powdered magnesium sulphate. The fluid is stirred

from time to time and the salt added to saturation. After at least

24 hours, it is filtered ; the precipitate is collected on a filter

moistened with saturated solution of .magnesium sulphate; the

1 Hoppe-Seyler, " Bestimmung des Eiweissgehaltes an Blutserum verm. Polarisa-
tion." Yirchow's Archiv, Vol. xi. p. 547.

2 L. Frederique, "Becherches sur la constitution du Plasma sanguin." Travail
du Laboratoire de Physiologie de VUniversite de Gand et du Laboratoire de Physiologic
de la Faculte des Sciences de Paris, 1878.

3 Hammarsten, " Ueber das Paraglobulin. " Pfluger's Archiv, 1878, p. 413.

1 90 DETERMINATION OF UREA IN BLOOD. [BOOK I.

substance left on the filter is thoroughly washed with the same
solution and the funnel and filter are heated for some hours to 110° C.
At the end of that time the serum-globulin has become so insoluble
that it can be washed with boiling water without any risk of solu-
tion. It is then treated repeatedly with warm alcohol and ether,
dried, weighed, incinerated, and the weight of ash deducted.

By Hammarsten's method the amount of serum-globulin found in
serum is sometimes greater than that of serum- albumin. (Consult
p. 61.)

Determination of the presence and quantity of Urea in the Blood.

Hoard's A weighed quantity of blood is diluted with about

modmed to* ^our ^mes *ts vo^ume of water, acidulated with sulphuric
Meissner'^and acid, and boiled so as to free it from proteids.
Gscheidien3. The clear filtrate is concentrated, and treated with

a solution of barium hydrate, which precipitates sul-
phates and phosphates, and the excess of baryta is removed by the
cautious addition of sulphuric acid. The fluid is then evaporated to
a syrupy consistence and mixed with absolute alcohol. The alcoholic
solution is separated from the precipitate (chiefly composed of inor-
ganic salts) which forms, the alcohol evaporated, and the residue
dissolved in water. The solution, which is of a light or deep yellow
colour, possesses an acid reaction. Solution of mercuric nitrate
is added cautiously ; this solution may be obtained by diluting
Liebig's mercurial solution for the estimation of urea with an equal
volume of water. A copious precipitate falls, the fluid is filtered and
the filtrate is rendered alkaline by adding baryta water or a solution
of sodium carbonate, and then more mercurial solution is added, until
a drop of the mixture brought in contact with solution of sodium
carbonate gives a yellow precipitate.

Proceeding in this manner a white precipitate is obtained, which
is well washed, then diffused in water and subjected to a stream of
sulphuretted hydrogen, which precipitates the mercury as sulphide.

The filtrate from the precipitate of sulphide of mercury is

concentrated and treated with a little concentrated and perfectly
colourless nitric acid. After some time crystals of nitrate of urea
separate, which are collected on a filter, dried over sulphuric acid and
weighed.

Whilst acknowledging the value of this method as demonstrating
the presence of urea in the blood in the most conclusive manner,

1 J. Picard, "De la presence de 1'Ur^e dans le sang et de sa diffusion dans
1'organisme a I'e'tat physiologique et a l'e"tat pathologique." These de Strasbourg, 1856.

2 Meissner, " Beitrage zur Kenntniss des Stoffwechsels im thierischen Organismus.
Der Ursprung des Harnstoffs im Harn der Saugethiere." Henle u. Pfeufer's
Zcitschrift f. rat. Medizin. Dritte Keihe. Vol. xxxi. pp. 234—349.

3 Gscheidien, Studien iiber den ' Ursprung des Harmtoffs im Thierlcorpcr. Leipzig,
1871.

CHAP. IV.) THE BLOOD. METHODS OF RESEARCH. 191

it is perfectly obvious that it must furnish results which must be
decidedly too low.

Grehant's A carefully weighed quantity of defibrinated blood

is mixed with twice its volume of strong alcohol
and set aside from one day to the next. The alcohol is then
filtered off, the insoluble coagulum is squeezed in a press and
washed with alcohol, the alcoholic liquids are concentrated in a water
bath. The residue is dissolved in water and is then introduced
into the vacuum of a mercurial pump, where it is subsequently mixed
with Millon's reagent (solution of mercurous nitrate in nitric acid).
The urea undergoes decomposition, yielding equal volumes of carbonic
acid and nitrogen, mixed with much nitric oxide. The gases are col-
lected and analyzed, and from the results the amount of urea is calcu-
lated. The author has found this method very troublesome of
execution and by no means as accurate as has been maintained
by its advocates. The decomposition takes place during a con-
siderable period of time, and in consequence of the disengage-
ment of nitric oxide continuing almost indefinitely the operator
is never sure when the process should be considered at an end.
Moreover the volumes of carbonic acid and nitrogen evolved are not
strictly equal ; in the author's experiments the volume of carbonic
acid was always somewhat below that of the nitrogen.

By decom- This method of estimating urea, which will be

position with described at length under the head of Urine, has been
sodium hypo- employed by the author in the determination of the
bromite. amount of urea in the blood.

A weighed quantity of blood, usually about 50 grammes, is
mixed with twice its volume of absolute alcohol and set aside
in a stoppered bottle for about 24 hours. At the end of this time
the precipitate is collected on a linen filter, washed with absolute
alcohol and subjected to firm pressure in a screw press. The
alcoholic liquid is evaporated to dryness in the water-bath, and the
residue is taken up in absolute alcohol, filtered, evaporated to
dryness, dissolved in a little water, and the watery solution filtered.
The filtrate is now placed in the decomposing bottle of Dupre"s
apparatus (Fig. 38) and subjected to the action of sodium hypobromite.
The nitrogen evolved, instead of being collected in a wide tube
such as is shewn in the engraving, is, in the case of urea determina-
tions in blood, collected in a much narrower tube divided into tenths
of a cubic centimetre.

From the volume of nitrogen obtained, the urea can readily be
calculated1.

It may be objected to this process that in reality it only furnishes
us with the amount of nitrogen given off by the extractive matters
of blood when treated with alkaline hypobromites, and that as other

1 The reader is referred for a full description of tlie process of estimating urea by
solutions of hypobromite to the section on Urine.

102 DETERMINATION OF UREA IN BLOOD. [BOOK I.

nitrogenous proximate principles than urea evolve nitrogen under
these circumstances, it is unphilosophical and inaccurate to consider
the nitrogen as all derived from urea. There is doubtless much
force in this objection; nevertheless as unquestionably urea is

FIG. 38. DUPRE'S APPARATUS.

infinitely the most abundant of the nitrogenous extractive matters
of the urine and of the blood, and as it is the only body which
does yield nearly the whole of its nitrogen to hypobromite, the
estimate of urea based upon nitrogen evolved under these cir-
cumstances is very near the truth. The method is one, too,
which yields most concordant results.

Haycraft's This method, which was worked out by John Hay-

method1, craft, M.B., in Professor Ludwig's Laboratory, appears

to be likely to supersede all others, though it has not yet been

1 Communicated privately to the Author. A short account of this method has
already appeared in the Journ. f. pract. Chemie.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 193

employed in any series of researches. It may be described as
follows : —

Defibrinated blood, varying in quantity between 10 arid 20 c.c., is
placed in a dialyser, so as to form a layer on the parchment-paper not
deeper than 4 mm. The dialyser is placed in a vessel containing
a volume of absolute alcohol equal to twice that of the blood.

In a period varying between one and four hours the fluid part of
the blood, holding the urea in solution, has passed into the alcohol,
leaving a solid mass behind. This is removed from the parchment-
paper, mixed with a little water and again placed in the dialyser.
The process is repeated three or four times, so as to make sure
that all the urea has been extracted. The alcohol is then poured
into a shallow porcelain dish, and after acidifying with oxalic
acid, so as to convert all urea into oxalate, the fluid is evaporated to
dryness. In the residue, crystals of oxalate of urea may be seen
with the naked eye, mixed with some fat, colouring matter, and
crystals of common salt. The fat and colouring matter are in
great part removed by the aid of petroleum naphtha, which readily
dissolves those substances, whilst it leaves the whole of the
oxalate of urea undi.ssolved. This is then dissolved in water and
mixed with a little barium carbonate, and the mixture evaporated to
dryness. On boiling with alcohol this fluid extracts the urea, leaving
traces of proteids and common salt. On evaporating the alcoholic
solution, almost pure crystals of urea are left ; the amount of this
substance is determined by weighing, or by Bunsen's method (see
'Determination of Urea in Urine'), or by means of the hypobromite
method.

This method is so precise that Dr Hay craft has never failed, by
its aid, in preparing a naked-eye demonstration of urea from ten
cubic centimetres of normal blood. He has found that by this
method the maximum error does not exceed 6 p. c. of the total weight
of urea.

Determination of the amount of Uric Acid in the Blood.

The amount of uric acid in the healthy blood of man is so
small as to render its determination, and even its detection, impossible.
In the blood of Birds, whose urinary excretion is very rich in uric
acid, Meissner1 succeeded, by a process for which we refer the
reader to his original memoir, in determining the amount of uric
acid.

The following is the process employed by Dr Garrod in the
detection and estimation of uric acid in the blood of gouty patients.
" The serum of the blood is first dried over a water- bath, then

1 Meissner, "Der Ursprung der Harnsaure des Harns der Vogel." Zeitschr. /. rat.
Med., Dritte Reihe. Vol. xxxi. p. 144 et seq.

13

194 DETERMINATION OF SUGAR IN THE BLOOD. [BOOK f.

reduced to coarse powder, and treated with hot alcohol ; the spirit
being removed, the residue is afterwards to be digested for some
minutes in distilled water, and raised to the boiling point ; the
watery solution is then filtered and evaporated to a thin syrupy
consistence. A drop or two of the solution, when heated on a
piece of porce'ain, with nitric acid and ammonia afterwards added,
exhibits at once the murexide test. A small portion of the same
solution, if acidulated strongly with acetic acid, and allowed to evapo-
rate spontaneously, gives rise to the crystallization of uric acid, the
crystals exhibiting its characteristic forms ; and lastly, the syrupy
solution, if merely allowed to evaporate without the addition of
any acid, exhibits upon its surface, after a few hours, small white
tufts of acicular crystal of urate of soda; the nature of the base
being determined by the examination of the white alkaline ash left
after incineration ; the acid by the murexide and other tests." In
cases where the amount of uric acid wrhich separates on acidulating
the aqueous solution in the above process by acetic acid is consider-
able, it may be collected on a weighed filter, washed, dried, and weighed.
" In the clinical examination of the blood, this process would be
too elaborate and tedious; but a method which answers admirably for
practical purposes is, to put about two drachms of the serum in
a flat glass dish, somewhat larger than a watch-glass, acidulate
slightly with acetic acid, and having placed in the fluid an ultimate
fibre from a piece of linen cloth (unwashed huckaback answers
well) set it aside in a safe place until the evaporation has proceeded
sufficiently far to cause it to become of a gelatinous consistence.
If there is uric acid in any abnormal quantity in the serum,
the fibre becomes studded with crystals of uric acid, which can
be at once recognized by placing the glass under the microscope
with a low power, or by the use of a small magnifying glass.
I have never yet, after very numerous trials, failed to discover
uric acid in the blood of gouty patients by this method, and the
test has an especial advantage in only requiring the abstraction of
a very small quantity of so important a fluid1."

Determination of the amount of Sugar in the Blood.

The principle upon which all methods of estimating the amount of
sugar in the blood are based is to dilute the blood (sometimes only the
serum) and to coagulate the proteid matters and haemoglobin which it
contains ; thereafter to determine the sugar in the filtrate by one of
the methods to be described in detail under the head of Urine. The
following method of separating the proteid matters is that which has
been followed by Dr Pavy 2 in his researches.

1 Garrod, Article "Gout," Eeynolds's System of Medicine, Vol. i. p. 825 and 826.
See also Garrod, Med. Chir. Transactions, Vol. xxxvu. (1854) p. 826.

2 Pavy, "The Croonian Lectures, on certain points connected with Diabetes."
London, 1878.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 195

" Forty grammes of sulphate of soda in small crystals are weighed
out in a beaker of about 200 c.c. capacity. About 20 c.c. of the blood
intended for analysis are then poured upon the crystals, and the beaker
and its contents again carefully weighed. In this way, the precise
weight of the blood taken is ascertained. The blood and crystals
are well stirred together with a glass rod, and about 30 c.c. of a hot
concentrated solution of sulphate of soda added. The beaker is
placed over a flame guarded with wire gauze, and the contents heated
until a thoroughly formed coagulum is seen to be suspended in a
clear colourless liquid, to attain which actual boiling for a short time
is required. The liquid has now to be separated from the coagulum
and the latter washed to remove all the sugar. This is done by
first pouring off the liquid through a piece of muslin resting in a
funnel into another beaker of rather larger capacity. Some of the
hot concentrated solution of sulphate of soda is then poured on the
coagulum, well stirred up with it, and the whole thrown on the
piece of muslin. By squeezing, the liquid is expressed, and to secure
that no sugar is left behind, the coagukim is returned to the beaker
and the process of washing and squeezing is repeated.

" The liquid thus obtained may be fairly regarded as containing
all the sugar that existed in the blood. From the coarse kind of
filtration and squeezing employed, it is slightly turbid, and requires
to be thoroughly boiled to prepare it for filtration through ordinary
filter paper. A perfectly clear liquid runs through, and to complete
this part of the operation the beaker that has been used and the
filter paper are washed with some of the concentrated solution of
sulphate of soda before referred to." In the solution thus obtained
the sugar is determined by the amount of cupric oxide which it can
reduce, the copper being separated electrolytically.

v. Mering1 merely dilutes the serum with four or five times its
volume of water, boils and adds dilute acetic acid in sufficient quantity
to cause thorough separation of the proteids in a flocculent form ;
the filtrate is then concentrated, and the sugar in it determined
either by Fehling's solution or by Sachsse's method (see 'Urine').

Determination of the lueight of the Moist Corpuscles contained
in the Blood.

Various methods have been suggested for determining the weight
of the moist blood corpuscles, all of which are attended with con-
siderable practical difficulties. Fortunately, by a combination of the
processes of enumeration of the blood corpuscles and determination
of the amount of haemoglobin contained in the blood, information
is acquired which possesses as great value to the physician as

1 v. Mering, "Ueber die Ahzugswege des Zuckers aus der Dannhohle." Archiv f.
Anat. u. Phyniol. 1877, p. 379 et seq.

13—2

196 DETERMINATION OF THE GASES OF THE BLOOD. [BOOK T.

would attach to a knowledge of the actual weight of the blood
corpuscles. Determinations of the weight of the moist corpuscles
will probably in the future be rarely attempted. The following
very brief description of the one method which is to be recommended
above all others will suffice : it is based n^on finding the relative weight
of fibrin in the liquor sanguinis and in the blood (Hoppe-Seyler).

This method, which can only be carried out when all suitable
preparations can be made before the blood is removed from the living
body, is as follows :

Blood is received in a cylindrical (metallic) vessel which is sur-
rounded with Iceland at the same time another portion of 30 — 50 c.c.
of blood is collected and the fibrin determined in it by the proceeding
described at page 179.

After an interval of an hour or two, the corpuscles having had
time to subside from the liquor sanguinis in the sample of blood
first collected, from 30 to 50 c.c. of the clear liquid are drawn off
by means of a cooled pipette and placed in a second apparatus for the
extraction of fibrin, and the process carried out exactly as in
the first case. The amount of fibrin being known, the operator is in
possession of the data required to be known.

The calculation will be understood by quoting the following example
from v. Gorup-Besatiez.

(1) the weight of fibrin in 1000 grammes of blood

was found to be . . . . . 3 -95 grins.

(2) the weight of fibrin in 1000 grammes of liquor
sanguinis was found to be . . . .8*07 grms.

If 8 07 grms. of fibrin correspond to 1000 grms. of plasma, to how much
plasma will 3-95 grms. of fibrin correspond ?

8-07 r'lOOO :: 3'95 : x

Thus is found the weight of plasma in 1000 parts of blood, and the
weight of moist corpuscles is found, by subtraction, to amount to
1000-486-98 = 513-02.

Separation and Determination of the Gases of the Blood.

The methods now universally adopted and alone to be recom-
mended for the extraction of the gases of the blood consist in
introducing an accurately determined volume of blood into the
vacuum of a mercurial air-pump, exposing it to a temperature of
40° — 45° C., removing the gases pari passu with their disengagement,
collecting them over mercury, and then subjecting them to analysis.
Whilst the principles which guide these operations are very similar,

1 A vessel constructed on Dr Sanderson's plan (Fig. 9, p. 32) should be used for
this experiment.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH.

197

the actual form of mercurial pump employed and the details of the
different operations employed by different experimenters, and, indeed,
in different laboratories, vary very much.

We shall here describe (1) methods of collecting blood intended
for gas analysis ; (2) the various pumps which may be employed with
advantage ; and (3) the methods of determining the composition of
the separated gases.

Collection of Blood for the Determination of its Gases.

In all cases the blood to be investigated must be collected over
mercury in such a way as to avoid all access of air.

The apparatus (Fig. 39) is admirably adapted for this purpose

FIG. 39. APPARATUS FOR COLLECTING BLOOD OF WHICH THE GASES ARE TO BE DETERMINED,

198 COLLECTION OF BLOOD. [BOOK 1.

One or both the tubes having been filled with mercury and the
stop-cocks being shut, a narrow elastic tube leading from the
blood-vessel whence the blood is drawn, after being allowed to fill
with blood, is slipped over the free tube leading upwards from
the stop-cock and which is quite full of mercury. The filling bulb (R)
being in a suitable position, and the stop-cock opened, blood will
flow into the tube displacing the mercury which it previously con-
tained. When enough blood has been collected, the stop-cock is
closed, and a clip (Fig. 40) being applied to the india-rubber tube

FIG. 40. CLIP FOB COMPRESSING INDIA-RUBBER TUBES.

leading from the graduated tubes to the filling bulb, the tube is
released from the clamp which held it and shaken, or rather
inverted repeatedly, so as to defibrinate its contents. When fibrin
is separated by shaking blood and mercury in this way, it does
so in a state of very fine division. A suitable tube being now
attached to the constricted part of the graduated tube, in place of
that which served to conduct blood into it, and the tube having
been again fixed by its clamp to the stand, the mercury bulb is
raised, and the graduated tube^may be placed in communication with
the blood-receptacle of the mercurial pump, and any quantity of the
blood which it contains may be allowed to flow into the vacuum.
As the tube is graduated, the volume allowed to flow in can be
determined. If some time must intervene between the collection
of the gas and its analysis, the tube A may be removed from its clamp
and laid in a trough containing broken ice.

Although other methods of collecting and measuring the blood
which is introduced into mercurial pumps have been employed,
and will be referred to. in describing the various forms of mercurial
pump, those here given, which the Author is in the habit of employing
in his laboratory, will be found to meet all requirements.

Mercurial Pumps.

Ludwig's The first pump to be described is Lud wig's1, which

pump. wag £rst figureci an(j described in a memoir by his pupil,

1 The first pump to which the name of ' Ludwig ' could be applied was described by
his pupil Setschenow (Zeitsclir. f. rat. Med., 3rd ser., Vol. x. p. 112). The form
described and figured is that at present employed in the Leipzig laboratory.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH.

199

FIG. 41. LUDWIG'S MERCURIAL PUMP.

200 MERCURIAL PUMPS. [BOOK I.

Alexander Schmidt1. It consists in reality of a combination of
two mercurial pumps. The vessel containing the blood of which
the gases are to be determined, G, is connected with the bulb B.
When a complete vacuum has been made in B arid in C, the stop-
cock E is closed, 0 is opened and G is plunged into hot water.
The blood enters into ebullition and its gases pass into B, some
of the blood also passing into that vessel. By opening E and K,
the gases are collected in (7, and K being then turned so as to shut
C off from B but to place it in communication with D, the latter is
raised so as to compress the gas in C\ on now opening the stop-
cock F, the gas maybe made to pass through H into a gas jar standing
over mercury. This very brief description will be understood by
carefully examining the drawing, especially if the reader make him-
self acquainted with the construction of Pfliiger's or Alvergniat's
pumps as described in the succeeding paragraphs.

In using Ludwig's pump, the blood is always defibrinated before
analysis. The blood to be analysed is introduced, without coming in
contact with air, into the receptacle G, which has previously been
filled with mercury and detached from the pump. The bulb having
a known capacity the volume of the blood analysed is known.

Pfluger's The pump, of which one form is represented by

pump. y-g 4^ possesses arrangements whereby watery vapour

which is disengaged in vacuo is at once absorbed.

G, C, E, F, D, represent parts of the pump proper ; C is the barometric
chamber of about two litres capacity, provided at G with a three-way
cock, which enables the chamber to be shut off or placed in communication
either with the chambers to be exhausted, B, A, or with the open
air, or by means of the glass gas delivery tube H with a mercurial
pneumatic trough. D is a bulb larger than C, and communicating with it
by means of a stout caoutchouc tube covered externally with a stout woven
fabric, so as to enable it to resist considerable internal pressure without
dilating. D is contained in a box which may easily be wound up and
down by means of the ratchet-wheel L, and the band and pulley connected
with it.

Mercury is poured into the filling globe />, when the latter is in its
lowest position. By winding D up until its level is above that of C,
and placing the stop-cock G in such a position that C communicates with
the external air, the bulb C is filled with mercury. The stop-cock G is
then turned so as to shut off C completely from communication above.
On now bringing D down to its initial position, viz., about a metre
below (7, the mercury in the latter sinks until it stands at the height
of the barometer above the mercury in the reservoir D. There is then
a Torricellian vacuum in C. By a suitable turn of the three-way cock G,
the chamber C is now brought into communication with the apparatus
to be exhausted. After the gas contained in the latter has diffused

1 Alex. Schmidt, " Ueber die Kohlensaure in den Blutkorperchen." Erste Abhand-
lung. Ber. d. Tconigl. sachs. Gesellsch. d. Wissenschaft. zu Leipzig. Math.-phys. Classe.
Vol. xix. (1867) S. 33.

FIG. 42. PFLUGER'S MERCURIAL ATR-PUMP, WITH THE ARRANGEMENTS FOR SEPARATING
THE GASES OF THE BLOOD (AS MADE BY GEISSLER OF BERLIN).

202 PFLUEGER'S MERCURIAL PUMP. [BOOK i.

into the chamber, the stop-cock G is shut, the globe D is elevated,
and by a suitable movement of the stop-cock the imprisoned gas is allowed
to pass either into the air, or is collected through H over mercury in
the graduated tube K standing in the pneumatic trough /. By repeating
several times the series of operations described the amount of residual
gas in the apparatus sinks to an insignificant amount, and, without
great labour, a practically perfect vacuum is obtained.

The accessory apparatus shewn in the drawing requires description.
0 is a mercurial gauge, B is the drying chamber, composed of four
glass tubes communicating below with two small reservoirs. The tubes are
filled with pumice-storie or asbestos saturated with sulphuric acid, and the
bulbs also contain some of the same acid. The drying- chamber is in
communication with two large glass bulbs A, which are intended to arrest
the froth which arises from the boiling blood. To the 'froth-chamber'
is carefully attached a glass bulb M, into which the blood is placed.
This bulb has at its upper part a single-way stop-cock, but below it
is provided with a two-way cock. The plug of the stop-cock is, in the
drawing, shewn to be prolonged considerably beyond the socket into
which it fits. This plug is perforated in its long diameter by a canal
which passes through it obliquely, and is so arranged that the fluid passing
through the canal may be directed upwards into the 'blood-bulb,' or
downwards and outwards. (Fig. 43.)

A vacuum having been made in the 'drying-chamber,' the 'froth-
chamber,' and the 'blood-bulb,' the plug of the lowest stop-cock of the
blood-bulb (a, Fig. 43) has attached to it, by means of a piece of thoroughly
sound black elastic tube, a flexible metallic tube, which is connected peri-
pherally with a glass cannula which is tied into the blood-vessel whence the
blood is to be drawn, or preferably with a blood-measuring tube.

Blood is now allowed to flow through the elastic tube until the latter is
filled, the plug being placed in such a position that the displaced air
and the displacing blood flow at first not into the blood-bulb but outwards.
At a given moment the stop-cock is turned (in the position shewn in Fig. 43)
so as to open a communication between the blood-bulb and the blood-
measuring tube, or the blood-vessel: the blood flows into the vacuous
bulb, and immediately enters into ebullition. As soon as enough blood
has entered, the lower stop-cock is shut, and the stop-cocks which shut

FIG. 43 exhibits the construction of the two-way cock (.¥, Fig. 42) at the lower part
of the blood-bulb. When the plug is in the position shewn the tuhe a communi-
cates with the interior of the bulb. "When the position is reversed a communicates
with 6. In intermediate positions the bulb is shut off and the tubes a and b do not
communicate.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 203

off the upper part of the blood-bulb from the drying-chamber and the
barometric chamber of the pump are opened. The blood-bulb M is now
immersed in a vessel containing water at 40° C., when the blood enters
into violent ebullition : if arterial in colour before being introduced into
the bulb, it assumes almost instantly the cherry-red colour which is
characteristic of reduced haemoglobin; if the reddened walls of the
froth-chamber be viewed through a spectroscope the simple broad band of
reduced haemoglobin is then seen. After a few minutes the gases which
have been given off are collected over mercury in a tube filled with
mercury, the vacuum is renewed, and the process of ebullition continued.

Some observers who have used the pump shewn in Fig. 42, have
determined the amount of blood analysed, by actually weighing it. With
this object, the exhausted and empty blood-bulb is detached from the pump
and weighed ; thereafter the quantity of blood to be analysed is introduced
into it, in the manner previously mentioned ; the stop-cock M through
which blood has flowed is then rinsed, first with water, then with alcohol,
and rapidly dried, and the bulb is again weighed. The blood bulb is then
again joined to the bulbs A (Fig. 42), of which the stop-cock N has been
kept closed. The junction having been made, the small quantity of air
which intervened between the upper stop-cock of the blood-bulb and N
having been removed by a few strokes of the pump, the process of boiling
the bulb is commenced. This process appears to the Author to be tedious
and unsatisfactory in the extreme. It is always better to pass the blood
from an apparatus in which it is first measured to the blood-bulb ; it is
indeed quite practicable to measure the blood and pass it into the bulb,
before coagulation has had time to set in.

In the mercurial pumps made by Geissler of Bonn, and which are,
the Author believes, identical with those used by Professor Pfliiger himself,
the arrangement shewn in Fig. 44 is employed.

FIG. 44. LARGE RECEIVER or PFLUGER'S PUMP, AS MADE BY GEISSLER OF BONN,

INTO WHICH BLOOD IS INTRODUCED, AND BOILED FOR THE EXTRACTION OF ITS GASES.

The tube which conveys the blood to be analysed is slipped over A.
By suitable manipulation of the stop-cock, the blood is first made to expel
the air in the tube A, outwards through 0. The blood may then be directed

204 ALVERGNIAT'S MERCURIAL PUMP. [BOOK i.

into the lower of the two large bulbs, which are attached to the pump, and
which have been perfectly exhausted.

The quantity of blood analysed is determined after the gases have been
extracted by weighing the bulb apparatus (Fig. 44), and the sulphuric acid
drying apparatus, and subtracting the weight of the same, as determined
before the blood was boiled. It is obvious that with this pump also it is
easiest to measure the blood before it is introduced into the vacuum.

The special features of the process described above, and which
renders it preferable to some others employed for the same purpose,
are, (1) the blood may readily be brought directly (if desired) without
previous defibrination, from the blood-vessels into the apparatus
where its gases are separated : in this respect it differs, for instance,
from Lud wig's pump ; (2) the blood is at once introduced into a very
large vacuous space, so that the O-pressure outside the blood is always
very much below the dissociation -tension of the 0 of the blood,
the latter therefore escapes very rapidly; (3) the vacuum is main-
tained in a dry condition by the sulphuric acid in the drying chamber ;
this appears to have very great influence in facilitating the removal
of the gases from the blood. With such an arrangement it is possible,
for instance, in a very brief space of time rapidly to extract all the
carbonic acid of the blood without the necessity of adding a dilute acid.

Alver- This pump, constructed by MM. Alvergniat freres

gniat's pump. of pariSj was first employed in the investigation of the
gases of tbe blood by Orphan t and Bert1, and bas already proved most
useful. Being constructed exactly on the principle of Pfluger's pump,
it does not require a special description ; it will be observed that its
barometric chamber is very much smaller than that of the first-named
instrument, and that it is not, as sold, provided with any arrangement
for absorbing watery vapour which may be given off in vacuo, though
such an arrangement can be contrived and connected with it by the
operator. The special features of this pump are, 1st, that just above
the three-way cock is situated, permanently, a most convenient
small mercurial trough, 2ndly and chiefly, that the three-way cock is
immersed in an iron box which is filled with mercury, so that all
risk of leakage is avoided.

In extracting the gases of the blood with this small pump it is
usual to connect with it a long glass tube about 2 inches in diameter,
with a bulb blown at its lower end having a capacity of about one litre.
This bulb is closed by an india-rubber stopper which is perforated by a
thermometer tube of narrow bore. The junction between the tube
and the pump is made by means of india-rubber tube carefully wired,
and is protected by a water-joint. The bulb also is immersed into a
tin vessel containing water. The narrow thermometer tube has
attached to it a fine glass stop-cock with an almost capillary bore.
This stop-cock is also immersed in water. This system of protecting
every junction by surrounding it either with water or mercury is un-

1 Bert, Lemons sur la Respiration. Paris, 1870.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH.

205

doubtedlyan admirable one, and relieves the mind of the experimenter
from the fear of an accidental leak — an eventuality which, unfortu-
nately, does occur where many stop-cocks are freely surrounded by air.

FIG. 45. ALVEKGNIAT'S MERCURIAL PUMP, FITTED UP AS IN AN ACTUAL EXPERIMENT.

206 ANALYSIS OF THE GASES OF BLOOD. [BOOK I.

In Fig. 45 the pump is shewn as fitted up for an actual
experiment ; between the pump and the blood-receiver a wide
glass bottle containing sulphuric acid and asbestos is shewn. All the
connections are protected by water or mercury joints. Standing in
front of the pump, and held by an iron clamp, is a tube similar to the
one which is only partiaUy seen in the drawing of the pump and its
connections.

The Author's experiments with Alvergniat's pump have impressed
him most favourably. The smallness of the barometric chamber
naturally makes the process of exhausting the apparatus connected
with the pump a very tedious matter, unless the plan be adopted of
exhausting at first by means of an ordinary air-pump or with the aid
of a water aspirator, and towards the close of the exhaustion allowing
two or three cubic centimetres of boiled-out water to enter the
nearly empty bulb and heating the water which surrounds the bulb
so as to cause the contained water to boil. The steam which is
disengaged, very rapidly and perfectly expels the last traces of air.
Without this expedient the experimenter will almost despair to
obtain a good vacuum with Alvergniat's pump, when there are
connected with it vessels having a capacity of between 1500 and
2000 c.c.

The Author has found it convenient to interpose a small sulphuric
acid chamber between the pump and the blood-receptacle, the object
being to prevent the passage of water into the former and from
it into the tube in which the gas is collected. With this addi-
tion he can recommend Alvergniat's pump as adapted for researches
on the gases of the blood1. By its portableness, it lends itself
admirably to demonstrations in the lecture-room.

It will be found convenient to employ about 30 or 35 cubic
centimetres of blood for the determination of gases. The tem-
perature at which the process is best carried on is 45° C. By simply
heating in vacuo, the whole of the gases which are in a state of
solution or feebly combined may be removed ; the last portions of
carbonic acid are however more rapidly evolved by allowing a small
volume (one or two cubic centimetres) of a thoroughly boiled out
solution of phosphoric acid to enter the blood-receptacle near the close
of the operation. As has been shewn, however, by Pfliiger and his
pupils, the addition of an acid to blood before the oxygen has been
pumped out leads to a considerable diminution in the volume of
oxygen obtained, in consequence doubtless of the gas being used up
in processes of oxidation.

ANALYSIS OF THE GASES OF THE BLOOD.

It is not consistent with the object of this work to give detailed
descriptions of operations which belong to general chemistry, and

1 This pump is manufactured by MM. Alvergniat freres, Kue de la Sorbonne, Paris.
It costs only 160 francs ; the tube with bulb, &c. being sold separately.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 207

which may be learned by reference to systematic works on analysis.
Accordingly, the extensive subject of gas analysis will not be treated
•of with any pretence to completeness, the reader being referred
to other sources for information on the subject. The analysis of
gases is best carried out either by the methods suggested by Bunsen l
or with the aid of the most ingenious and accurate methods devised
by Professor Frankland.

?

Description of the methods of Frankland for the analysis of gases.

We quote the whole description of these methods from the
excellent account given by Professor Btirdon Sanderson2.

Frankland1 s " ^tn a yiew to the analysis of the gases of drinking

smaller appa- water, Frankland has introduced an apparatus of great
ratus for the simplicity (see Fig 46). the working of which will be readily
analysis of understood by the diagram. It consists of two parts, viz.,

gases by afo- a laboratory tube (k), in which the gas to be analysed is first
sorptiometric received, and a measuring apparatus, to which it can be trans-
ferred from the laboratory tube, in order that its volume may
be determined before and after each absorption. The measuring apparatus
.consists of two tubes (a, b), fixed vertically side by side in a stand, surrounded
by a chamber containing water (n). They communicate below both with
each other and (by the long flexible tube) with a mercury-holder (t), like that
of Alvergniat's pump. One of them can bo brought into communication by
the arm (y) with the laboratory tube; the other (b) is open at the top. A
scale of millimeters is engraved on it, the zero of which is opposite o. A
corresponding scale, starting from a zero at the same level, is engraved on
the measuring tube. The apparatus is filled with mercury by raising
the mercury-holder (t) to a sufficient height, the stop-cock (/) remaining
open; in doing which the surface of the mercury in t must not be more
than a few millimeters higher than the tap. As soon as mercury appears
at g, the stop-cock is closed. The next step is to fill the laboratory
tube. Having inverted it in the trough, which has been previously
raised to the proper height, the operator draws out most of the air by means
of a bent tube, the point of which rises to the top of the laboratory tube,
and shuts the stop-cock as soon as the mercury rises. The removal of the
air is completed by joining g and g' so as to connect the laboratory tube
with the measuring apparatus, and then causing the air contained in
the former to pass over into the latter, by depressing t. The stop-cock h
must now be closed and g and y' disconnected to allow of the expulsion of
the air from a. This having been accomplished, g and g' are again
brought together and carefully t-ecured. The whole apparatus is now full
of mercury ; as soon as it has been ascertained that the joint is air-tight at
all pressures, it is ready for use. Before proceeding further, however,
the measuring tube, which, as already stated, is graduated in millimetres
measured from an arbitrary zero line near the bottom, must be calibrated.
In other words, it must be ascertained as regards each principal mark of tho

1 Bunsen's Gasometry, translated by Eoscoe.

2 Handbook for the Physiological Laboratory, pp. 202— 20f.

208

FRANKLAND'S METHOD OF GAS ANALYSIS. [BOOK i.

graduation, what volume of air or water (as the case may be) the tube
contains, when the upper convex surface of the mercury stands exactly level
with it. For this purpose the orifice a is connected by means of an

FIG. 46. FRANKLAND'S SMALLER APPARATUS FOR THE ANALYSIS OF GASES BY THE
OF LIQUID REAGENTS. (From Sutton's Volumetric Analysis.)

-

india-rubber tube with a reservoir (a funnel) containing distilled water.
The mercurial column is then allowed to descend until it stands exactly at
zero. A weighed beaker having been then placed under a, water is expelled
till the column stands at a height of fifty millimetres, and the beaker again
weighed. In a similar manner the outflow of the water corresponding to a rise

i

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 209

of the mercurial column from fifty to one hundred millimetres is determined,
until the capacity which corresponds to each fifty millimetres of the scale is
ascertained. To ensure accuracy, the process must be repeated several
times. If the results, after correction for difference of temperature, are in
close accordance, the means may then be taken as expressing the capacities
required. In the upper part of the tube, calibration must be made
at shorter intervals. In calibrating, as in all subsequent measurements, the
height of the column must be read horizontally through a telescope, so
adjusted that its axis is at the same height as the surface of the mercury.
The temperature is read by a thermometer suspended in the cylinder of
water by which the barometer and measuring tube are surrounded.

"The measuring and laboratory tubes having been brought into connec-
tion in the manner described above, and both filled with mercury, the gas
to be analysed is introduced into the laboratory tube from the test-
tube in which it has been collected. It is then at once transferred to
the measuring tube by depressing t until the mercury rises in the laboratory
tube as far as the stop-cock g '. This done, the stop-cock g is closed,
and t raised or depressed till the column stands at one of the marks
of the graduation, in reference to which the capacity of the tube has been
determined. The temperature is then observed, and the pressure deter-
mined by adding the difference between the height of the column in
the measuring tube and that in the pressure tube, to the reading of
a barometer which stands by. A few drops of solution of caustic
potash having been introduced into the laboratory tube, the gas is returned
from the measuring tube. Absorption takes place rapidly. It is accelera-
ted by slightly agitating the trough, and by allowing the mercury to stream
into the laboratory tube after the gas has passed. The measurement of the
gas after absorption is performed in the same manner as before. About
half a centimetre of strong solution of pyrogallic acid is then introduced in
the same way as the potash, and the gas again returned. After absorption
of the oxygen, what remains is nitrogen. In analysis of blood gases, the
proportion of nitrogen is nearly constant, viz. about 2 '5 volumes in
100 volumes of blood. If a larger quantity is obtained, the fact indicates
that air has entered. Whatever method of analysis is employed, the
results must be reduced to 0° temperature and 760 millimetres pressure —
i.e. they must be expressed as if the measurements had been made
under those conditions. A further deduction must be made from
each measurement in respect of the aqueous vapour which the gas contains
(the measuring tube being always moist). This is accomplished by the
following well-known formula: —

v= V B'-f

l + £ 0-00367* 760 '

V denotes the corrected volume; V the volume read; t the temperature;
H' the observed pressure; and f the tension of aqueous vapour at the
temperature t. The values of 1 + 1 0'00367 and/ are always obtained from
tables. For these and many other important practical details relating
to the performance of gas analysis, the reader is referred to Mr Button's
'Volumetrical Analysis,' whom I have to thank for two of the woodcuts
with which this section is illustrated. To illustrate the application
of the method to the analysis of the gases of the blood, I give the following
example : —

G. 14

210

FRANKLAND'S METHOD OF GAS ANALYSIS.

[BOOK i.

ANALYSIS OF GASES OF ARTERIAL BLOOD OF DOG.

1st Measure-
ment. Total
Quantity of
Gas Ex-
tracted.

2nd Measure-
ment. After
Absorption
of Carbonic
Acid Gas.

3rd Measure-
ment. After
Absorption
of Oxygen.

Height of Column in Measuring-
, tube
„ „ Pressure-tube

230-0
312-8

270-0
369-0

450-0
320-0

Difference
Reading of Barometer

82-8
764-0

99-0
764-0

-130-0
764-0

H' =

Temperature = 1 9-8° C. - 1
Tension of Aqueous Vapours from
table = f =

846-8
17-2

863-0
17-2

634-0
17-2

H'-f=

Volume of Gas as measured in
cubic centimetres = V' =

829-6
11-822

845-8
3-865

616-8
0-562

l + t 0-00367 (from table) = 1-0725.
Hence from first measurement we have —
11-822 829-6
'~ =

From second measurement —

3^ 84^
1-0725' 760
From third measurement —

jr.. 0-562 616-8

1-0725 ' 760 ~

Thus the total volume of gases obtained as measured at 0° C. and 760
mm. was 12-030 cubic centimetres: of carbonic acid gas was 12*030
-4-010 = 8-02 c.c. ; of oxygen 4-010-0-425 = 3-585 c.c. ; and of nitrogen
0-425 c.c.

As the volume of blood employed was 20-266 cubic centimetres, we have
the following final result : —
In 100 volumes of blood —

Carbonic acid gas 39-585 volumes

Oxygen
Nitrogen
Total

17-695

2-09

59-370

/ G-02
\ 0-20266

3-585

0-20266

0-425

0-20266

12-030

20266

vols

vols.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 211

"In the preceding example such variations of temperature and barometric
pressure as may occur during the analysis are disregarded. The readings
are taken immediately after the absorption of the carbonic acid gas ; as the
time occupied in the analysis up to this point is very short, the error
arising from the variations in question is inconsiderable. As regards the
absorption of oxygen, the error might be of more consequence, were it riot
that the residue of nitrogen is so small. As it is, it can be easily sheM'n
that it would require a difference of pressure amounting to three
millimetres, and a difference of a degree of temperature, to make an error
of one-hundredth of a percentage in the result as regards nitrogen or
oxygen. Within these limits, therefore, the errors arising from this source
may be regarded as trivial.

Frankland's " Although determinations of oxygen made by absorption

larger appa- with hydrate of potash and pyrogallio acid are not entirely
ratus for the free from objection on the score of accuracy, the results ob-
tained by the method above described are quite accurate
Som^trLTas enough for most of the purposes of physiological research,
well as ab- ^or ^e small errors are practically inappreciable, as compared

sorptiometric with the variations in the proportion of oxygen contained in
methods. the blood to be analysed, produced by what might be regarded

as very trifling differences in the mode of collecting it. If it is desired to
have recourse to explosion with hydrogen, the best methods for the purpose
are those of Dr W. Russell arid of Frankland and Ward. The following
short description of the latter will be readily understood from what has pre-
ceded. The apparatus (Fig. 46a} consists of two parts, corresponding to the
laboratory tube arid measuring tube of the instrument previously described.
The measuring tube (F, Fig. 46ct) communicates, as in that instrument, with
a second tube (ff, Fig. 46a), containing a column of mercury, by the height
of which the pressure to which the gas to be measured is subjected can be
estimated. The chief difference is that, whereas in the former more simple
instrument the pressure tube is open at the top, so that if air is contained in
the measuring tube, and the stop-cock by which it communicates with the
laboratory tube is closed, the difference between the heights of the two
columns indicates the difference between the tension of the gas in the measur-
ing tube and that of the atmosphere — in the instrument now before us the
tube is closed and constitutes a barometer, so that the difference expresses the
actual tension of the gas in inches of mercury. In the horizontal channel, by
which the measuring tube and barometer communicate at the bottom, is a
three-way stop-cock (not shewn in the figure); by which they may be brought
into communication either with a vertical escape tube, the end of which
dips into a receptacle containing mercury several feet below, or with a tube
open at the top (G, the middle and longest in the figure), called the filling tube.
In this way the gas can be expanded or compressed at the will of the opera-
tor, and consequently can (in most analyses) be readily brought to the same
volume after each successive operation. The convenience of this is very
great, for obviously the tensions of different quantities of gas when expan-
ded to the same volume are proportional to the volumes they would
assume if they were all under the same pressure, so that the original
volume of gas to be analyzed being known, the relation between
that volume and the volume of the other quantities to be measured can
be readily calculated, the several volumes being proportional to the

14—2

Fin. 4t5a. FKANKLAND'S LARGER APPARATUS FOR THE ANALYSIS OF GASES.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH.

213

corresponding readings of the barometer. The original yolume of gas
to be analyzed is measured as before described, with this difference,
that the absolute pressure to which it is exposed is known without
reference to the barometric pressure outside at the time. The ex-
plosion is effected in the eudiometer, into the upper end of which
two platinum wires are fixed for the purpose; the arrangement of
these wires is the same as in Bunsen's eudiometer. As to the mode of pre-
paring and introducing pure hydrogen and of exploding the mixture,
the reader will find sufficient information in Roscoe's translation of
Bunsen's Gasometry."

Description of more simple methods of gas analysis.

For purposes of demonstration it is sometimes- convenient to employ the
following expeditious and far from inaccurate method.

The tubes for collecting the gases which are strongly recommended are
represented in Fig. 47. They are about 250 millimetres long, and are

FIG. 47. ABSOKPTION TUBE1, WITH

DOUBLE SCALE, AS MADE BY AjLVER-

GNIAT. (Scale about J.)

FIG. 48. IRON SPOON EMPLOYED
IN TRANSFERRING TUBES FROM ONE
MERCURIAL TROUGH TO ANOTHER.
(Scale about £.)

1 These tubes are constructed by MM. Alvergniat freres, 10 et 12Eue de la Sorbonne,
Paris. From personal observation, the Author can testify to their accurate calibration.

214

METHODS OF GAS ANALYSIS.

[BOOK i.

provided with two scales placed side by side, one of which indicates volumes in
tenths of a cubic centimetre, the other is divided into millimetres. Further,
these tubes are very much constricted at their upper part, so that exceedingly
minute quantities of gas can be very accurately measured; this device
renders the tubes of special value in the determination of the gases of the
blood, as the volume of nitrogen which has to be read off is always very
small.

We shall suppose, then, that the experimenter has, by employing
Alvergniat's pump, collected the gases given off from a known volume of
the blood which he is analyzing, in such a graduated tube, the walls of
which have been moistened by a drop of water. He now transfers the

FIG. 49. WELL-SHAPED PNEUMATIC
TROUGH FOB MERCURY. (Scale about

\J

FIG. 50. PIPETTE WITH BULB,
FOR INTRODUCING LIQUID REAGENTS
INTO ABSORPTION TUBES STANDING
OVER MERCURY. (Scale about ^.)

tube with its contents to the mercurial trough having the form shewn in
Pig. 49 ; the transference being effected by means of the iron spoon shewn
in Fig. 48. The tube is then fixed in a clamp and plunged into the mercury
so that the level of the metal inside and outside the tube is exactly the
same; it should be left for an hour, and a second observation made to see
whether the level is still the same. If any change has occurred the tube
is again adjusted and the volume of the gas is read off, either by the
unaided eye, or still better by means of a telescope magnifying a few-
diameters and situated at a distance of a few feet from the tube.

The observer then reads the thermometer and barometer, and thus
obtains the data for calculating the total quantity of gas given off by the
volume of blood which he has analyzed.

With the aid of a pipette such as is shewn in Fig. 50, the observer
now throws up into the tube about half a cubic centimetre of solution

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 215

of caustic potash of sp. gr. 1-2, taking care that not a trace of air be
introduced ; by unclampin& the tube and alternately raising and depressing
the tube in the mercury the absorption of CO3 by the caustic potash is much
facilitated. After absorption appears to be complete, the tube is again
adjusted, so that the level of mercury inside and outside is the same, and
the volume of gas determined as before. If we subtract this volume from
that of the original gas the amount of carbon dioxide is ascertained.

About half a c.c. of a strong solution of pyrogallic acid (1 part
of acid to 8 of water) is now introduced into the tube, and the process of
shaking, &c., repeated. After the oxygen is absorbed the absorption tube
is transferred to a vessel containing water, and the level of the liquid
inside and outside of the tube being the same, the volume of gas is read
as before; the gas consists entirely of nitrogen, and by subtracting its
amount from that of the gas remaining after the absorption of CO2, we
determine the quantity of oxygen which was present.

It will be apparent to the reader that carried out as above there are
certain inherent errors which it is not easy to eliminate ; the method may
be rendered more accurate, however, by absorbing the carbonic acid by a
ball of caustic potash fused on platinum wire, then, after determining
the volume, absorbing the oxygen by solution of caustic potash and
pyrogallic acid, and after complete absorption transferring the tube which
contains the residual gas to a trough containing water, and reading the
nitrogen over water; or, again, after absorption of the C02 by the ball
of caustic potash, the oxygen may be absorbed by a bullet of phosphorus
left in the gas for 24 hours ; in this case, however, before reading off the
volume, a bullet of caustic potash must be introduced into the gas and left
for some hours, and the residual gas read as dry.

Determination of the total quantity of blood contained in an
animal's body.

Weicker's A tube is tied into the carotid of the animal

whilst yet alive, and a few cubic centimetres of blood
collected, defibrinated, measured, and set aside (portion A). The
animal is then bled to death, the whole of the blood defibrinated and
kept (portion B). The blood-vessels are then washed out with
normal salt solution until the washings issue quite colourless ; these
are added to portion B, and tbe whole mixed and measured. Some
of the red solution thus obtained is placed in a liaematinometer,
which we shall designate as H.B. ; a small quantity of A is then
diluted with 10 times its volume of distilled water, and an accurately
measured volume (one or two cubic centimetres) is placed in a
second haematinometer (H.A.) placed by the side of the first and
illuminated in exactly the same manner.

Distilled water is now added from a burette to the contents of
H.A. until their tint is exactly equal to the fluid in H.B. ; when
equality is obtained, the volume of water added is read off, and thus
is found the volume of the solution of pure blood which was equal to
the previously unknown mixture of portion B and washings. By
simple calculations, of which the steps are perfectly obvious, we can

216

DETERMINATION OF TOTAL VOLUME OF BLOOD. [BOOK I.

then find the amount of blood which the washings contain, which
added to the volume of A gives the total volume of blood contained in
the body.

This method may be modified in various ways. Thus the amount
of haemoglobin in the fluid may be determined by Preyer's method ;
or the tissues and organs may be chopped up finely and treated with
water, and the fluid thus obtained after being filtered may be added
to the washings from the blood-vessels ; or carbonic oxide may be
passed through the blood and through the mixture of blood and water,
so as to secure a fluid of more uniform and more persistent tint.

The following determinations of the relation of volume of blood to
weight of body have been made by these methods.

VOLUME OF BLOOD, EXPEESSED AS A FRACTION OF THE BODY
WEIGHT, CONTAINED IN THE BODY OF VARIOUS ANIMALS.
(GSCHEIDLEN1.)

According to

Welcker.

Heidenhain.

Gscheidlen.

Panum.

Spiegelberg
and
Gscheidlen.

Guinea-pig .
Rabbit . .
Dog ...
Cat ...

T5

T* to iV

TT t0 A
TT *° 2T

1 fr> 1

TT to T¥

AtoA

Malassez'
method.

It is obvious that by the enumeration of blood
corpuscles in blood diluted to a known extent, and in
the mixture of blood and washings, the amount of blood contained in
an animal's body could also be ascertained, though doubtless not so
accurately as by Welcker's method.

Malassez 2 has actually attempted to determine the total mass of
the blood of a living man by the process of enumeration. The
number of blood corpuscles contained in a cubic millimetre of blood
obtained from the finger having been determined as exactly as
possible, 300 cubic centimetres of blood were removed by venesection.
Some hours afterwards, the corpuscles in blood again drawn from the

1 Gscheidlen, Physiologisches Methodik, Dritte Lieferung, p. 337. On this subject
consult also

Gscheidlen, " Studien iiber die Blutmenge." Untersuchungen aus dem physlolog.
Laboratorium zu Wurzburg, Vol. n. p. 153 (1869).

Gscheidlen, " Bemerkungen zu der Welcker 'sch en Methode der Blutbestimmung
und der Blutmenge einiger Saugethiere. " Pfliiger's Archiv, Vol. vn. (1873) p. 544.

Welcker, "Bestimmung der Menge des Korperblutes und der Blutfarbskraft, " &c.
Zeitschrift f. rat. Medicin. 3rd Series, Vol. iv. (1858) p. 147.

2 Malassez, "Recherches sur quelques variations que presente la masse totale du
sang." Archives de Phy&iologie normale et pathologique. 2nd series. Vol. n. (1875).
Consult especially pp. 277—280.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 217

finger were counted. By assuming that in the interval which had
elapsed between the venesection and the second enumeration the
volume of blood had become exactly the same as it had been at the
time of the first enumeration, and further that no formation of new
corpuscles had taken place in the same period, Malassez obtained data
for calculating the total mass of the blood.

It is obvious however that these assumptions are altogether
unwarrantable, and if in one case they led to a result not far removed
from the truth, such was a mere result of chance.

Medico-Legal Detection of Blood-Stains.

Detection It not unfrequently happens that the medical

of blood cells jurist is asked to decide whether a certain stain upon

clothes, weapons, floors, &c. is a stain of blood. When

recent, the identification of a blood- stain presents no

difficulties. By moistening it with diluted glycerine of specific

gravity 1025 and, after some time, expressing the liquid, we may

obtain microscopic evidence of the presence of blood corpuscles ;

when such is the case the observer may be able to state positively

that the blood was or was not the blood of a mammal, but cannot

venture upon any more definite expression of opinion.

Chemical Whether successful or not in the detection of blood

reaction of corpuscles, it is always desirable to obtain the chemical
chief blood proofs of the presence of blood ; and with proper treat-
constituents ment this is possible even with blood- stains of consider-
in stain. %k>\Q antiquity and of small size.

We shall suppose that the observer is examining a cloth stained
with blood; having selected the particular stain which he wishes
to examine he may, with pencil, draw a circle around it and mark
the circle with a letter or number, for purposes of identification and
description. He then will proceed to cut out the stain and to
pass a thread through it; the blood-stained piece of cloth is then
suspended in a very small test-tube containing a few drops of
distilled water ; the size of the tube must depend upon the estimate
which the experimenter forms of the amount of blood in the stain.
The piece of stained cloth is left to soak for one or two hours, at the
end of which time the water will usually have acquired a more or less
distinctly red colouration. By the aid of the thread which had been
attached to it the little piece of cloth is now withdrawn from the water
and pressed with a small glass rod against the upper part of the test-
tube so as to squeeze out the liquid which it had imbibed. A small
quantity of the liquid may be examined in a small cell with the aid
of the microspectroscope ; but only when the examination is carried
on by a person who has by considerable practice familiarized himself
with the use of the instrument and with the various absorption
spectra of colouring matters.

218 DETECTION OF BLOOD STAINS. [BOOK I.

In recognizing blood by means of the spectroscope the observer endeavours
to obtain a succession of characteristic spectra ; even when haemoglobin has
been decomposed and the stain contains methaemoglobin or haematin a
satisfactory series of spectrum observations may be made 1.

If the quantity of red solution be sufficient, a few drops may be
treated with solution of ammonia which will induce no change. The
greater part of the liquid, or, if its quantity be small, the whole of
it, is now heated to boiling; the red colour will disappear and a
turbidity or coagulation will be observed to form, the coagulum
having a dirty grey colour ; on now adding a drop of a solution of
caustic potash to the turbid liquid, this will be instantly cleared and
the solution will be observed to be green by transmitted and red by
reflected light ; on adding a small drop of nitric acid the precipitate
will be reproduced.

Guaiacum Another test which adds confirmatory evidence to

that afforded by other means, and which is of extreme
delicacy, rests upon the reaction developed by haemoglobin and its
derivatives when brought in contact with guaiacum and hydric
peroxide. To try this test it is best to moisten the stain with
distilled water, and then to press a piece of white filtering paper
firmly against it ; a little of the colouring matter will adhere to
the filtering paper. Having secured a slight stain on the filtering
paper, this is moistened with a drop of tincture of guaiacum, and
then with a few drops of an ethereal solution of peroxide of
hydrogen. A beautiful blue colour will be developed if the stain
is one of blood. It must be borne in memory, however, that
this test cannot be relied upon by itself, though the evidence
which it affords is valuable when taken in connection with other
facts.

TheHaemin In the case of very old blood-stains it may not only

test- be impossible to obtain blood corpuscles for microscopic

examination, but even to obtain a solution containing the colouring
matter and proteids of the blood. In such a case the haemin-test is
of special value. This test is based upon the fact that when
haemoglobin or haematin are heated with glacial acetic acid and
common salt, a hydrochlorate of haematin is formed, which, on
evaporation, is deposited in the form of reddish brown prisms, the
so-called haemin-crystals. The test is one of great delicacy and the
result is remarkably free from fallacy. The blood-stain, having been
cut out, is placed with a few drops of glacial acetic acid and a very
minute (indeed scarcely perceptible) crystal of common salt, in a
watch-glass, which is then heated to boiling over a spirit-lamp
flame. The liquid will soon assume a brownish red tint; the little
piece of cloth may then be squeezed with a rod against the side

1 Consult Sorby, " On some improvements in the spectrum method of detecting
blood." Monthly Microscopical Journal, Vol. vi. (1871) p. 9. Also MacMunn, The
Spectroscope in Medicine. London, Churchill, 1880.

CHAP. IV.] THE BLOOD. METHODS OF RESEARCH. 219

of the watch-glass, and the liquid is evaporated to dryness. The
watch-glass is then examined with a magnifying power of about
350 diameters. If no crystals are perceptible, more acetic acid
may be added and the process of boiling and evaporation repeated.
If present, the crystals present the appearance shewn in Fig. 24 (page
115).

Medico-Legal Detection of Carbonic Oxide in Blood.

As was mentioned at page 105, carbonic oxide expels the oxygen
from oxy-haemoglobin and forms a more stable compound, which is
not affected by the alkaline reducing solutions which readily reduce
oxy-haemoglobin. Blood of animals poisoned with carbonic oxide, if
nearly saturated with the gas, presents a remarkably persistent
vermilion colouration ; if not saturated, the colour may not be very
distinctly affected.

The action of a solution of caustic soda of specific gravity 1*3
establishes a very remarkable difference between CO-blood and
normal blood1. This reagent when added to normal blood converts it
into a black, slimy, mass, which when spread in thin layers over a
porcelain capsule appears of a greenish brown colour; blood which
has absorbed carbonic oxide presents, on the contrary, after treatment
with its own volume of the solution of caustic soda, the appearance of
a firmly coagulated mass, and, when spread on porcelain, appears of a
cinnabar-red colour.

It has been recommended that, instead of employing a simple
solution of caustic soda, a mixture of two parts of a solution of caustic
soda of sp. gr. T3, and 2J parts of a solution of chloride of calcium in
water (1 to 3), should be rubbed up with the blood in a porcelain
capsule, fifteen or twenty drops being sufficient for the reaction2.

A more conclusive proof of the presence of carbonic oxide is
obtained with the aid of the spectroscope. The suspected blood is
suitably diluted so as to exhibit with perfect distinctness the two
absorption bands of O2-Hb or CO-Hb. Then a small quantity of
Stokes's reagent (ammoniacal solution of ferrous tartrate or citrate) is
added. In the event of the blood containing carbonic oxide the two
bands will not wholly fade, but will persist more or less distinctly.
When the blood is saturated with carbonic oxide the spectrum
undergoes no perceptible change under the influence of the reducing
solution.

1 Hoppe, Virchow's Archiv, Vol. xi. Heft 3 (1857), p. 288.

2 Eulenberg, Die Lehre von den schadlichen und giftigen Gasen. Braunschweig,
1865, p. 48.

CHAPTER V.

THE LYMPH AND CHYLE. THE SO-CALLED
TRANSUDATIONS, NORMAL AND PATHOLOGICAL.

SEC. 1. THE LYMPH (INCLUDING THE CHYLE).
Preliminary Observations.

On the As the blood circulates through the capillaries of

nature of the the body there is a continual transudation, through
Lymph. their walls, of water holding in solution organic,

mineral, and gaseous constituents, which are destined for the nutrition
of the elements of the tissues. This nutritive fluid bathes the tissue
elements, and is the agent which supplies them directly with the
matters which they require for their maintenance and repair, whilst,
at the same time, it removes from them soluble effete matters which
would, if accumulating, impair the functional activity of the tissues
in which they have been formed. The fluid which has transuded
from the blood-vessels finds its way into the minutest radicles of the
lymphatic system, and is then carried, sooner or later, to lymphatic
glands, and through them into larger lymphatics which ultimately
empty their contents into the large venous trunks in proximity to the
heart. The term lymph, although usually applied to the liquid
contained in the lymphatic vessels, is also applicable to the fluid
which is found in those extra- vascular spaces from which the
lymphatics originate, or with which they communicate — to the fluid,
for instance, which bathes the lacunae of connective tissue, or which
moistens the interior of the great serous sacs. Inasmuch as divers
organs take from the fluid transuded by the blood different quantities
of organic, saline, and gaseous constituents, according to their wants,
and produce different kinds and different quantities of effete products,
it follows that the lymph must be a liquid which varies materially
in composition, according to the region from which it is derived, and
according to the greater or less functional activity of the organs
contributing to it.

CHAP. V.]

PHYSICAL CHARACTERS OF LYMPH.

221

Chyle is the
term applied
to the lymph
contained in
the lympha-
tics of the
small intes-
tine during
digestion.

quantity of
Lymph and
Chyle.

Whilst the lymphatics generally contain a liquid
which must be looked upon as a diluted liquor sanguinis,
deprived of a small fraction of certain of its constituents,
and augmented by certain other constituents, such as
urea or carbonic acid, which are the effete products of
tissue metabolism, certain of the lymphatics — those of
the small intestine — contain, during the period of diges-
tion, lymph which is laden with suspended fattymatter in
a fine state of division, and which gives to the fluid a milky appear-
ance. The fatty matter has passed from the interior of the alimentary
canal through, or between, the cylindrical epithelial cells of the villi
into the sub-epithelial connective tissue, whence it has made its
way into the commencements of the so-called lacteals, as the ab-
sorbents of the intestinal villi are called. Chyle is therefore the
lymph of the small intestine laden with fat whilst the absorption of
that substance is proceeding. The Chyle will be considered in detail
in relation to the functions of Digestion and Assimilation.

Circum- The amount of lymph which is discharged by the

stances which lymphatics of a part is much increased by muscular
influence the J ,. . » , , J . ,TT1

contractions and passive movements 01 the part. When

the arterial pressure is increased the amount of lymph
diminishes. When an obstruction to venous circulation
exists the amount of lymph increases. Poisoning with curare increases
the discharge of lymph. The amount of chyle is materially increased
by the digestion of food rich in fatty matters.

Mode of Small quantities of lymph for microscopic exami-

obtaining nation may be obtained by puncturing the subcutaneous

Lymph. dorsal lymph-sac of the frog, and aspirating with a

capillary pipette.

When large quantities of lymph are required they may be obtained
by tying a glass cannula into the thoracic duct of a deeply anaesthe-
tized animal, at the spot where that tube empties itself into the
junction of the large veins at the root of the neck1.

In large animals, such as the horse and ox, a cannula may be
tied into one of the large cervical lymphatics accompanying the
rotid artery.

For purposes of demonstration small quantities of lymph may
be obtained from the thoracic duct of a recently killed animal.

Physical characters of the Lymph.

When freshly drawn from the thoracic duct of fasting
animals the lymph is a transparent liquid, sometimes of
a slight yellow colour; when obtained from an animal
during the period of digestion, it presents a more or less milky colour
owing to the absorption of fatty matters from the alimentary canal.

1 This method was followed by Dogiel and by Hammarsten, in their researches on the
gases of lymph, conducted in the Leipzig Laboratory under the direction of Professor
Ludwig. (See ' Gases of Lymph, ' p. 225.)

Colour, and
microscopic
characters.

222 THE LYMPH AND CHYLE. TRANSUDATIONS. [BOOK I.

On microscopic examination, the transparent lymph of fasting
animals presents colourless corpuscles — lymph-corpuscles, identical with
the colourless corpuscles of the blood, floating in a clear liquid, the
lymph-plasma; mixed with these, a few coloured corpuscles are
often observed, even though great precautions have been taken to
prevent the admixture, with the lymph, of blood from wounded blood-
vessels.

It is certain that the lymph corpuscles are comparatively scanty
in the radicles of the lymphatic system, and that they are increased
in number as the lymph passes through the lymphatic glands. These
glands are the chief, though not the exclusive, formers of the lymph
cells, for wherever lymphoid or adenoid connective tissue exists, as
for instance in the mucous membrane of the stomach and intestines
(of which it forms almost the frame-work) or in the follicles of the
thymus, of the tonsils, of the spleen (Malpighian bodies), there
is doubtless a formation of lymph cells. It is, indeed, the wide
distribution of adenoid connective tissue, especially in the alimentary
canal, which accounts in great part for the fact, that the lymph of the
smallest lymphatics always contains some corpuscles, though some of
these are doubtless derived from the blood, and have wandered through
the capillary walls into the cell spaces of the connective tissue, and
so found their way into the lymphatics.

The lymph of animals in active digestion is milky from admixture
with the fatty chyle. It exhibits under the microscope, what has
been termed a molecular basis, i.e. innumerable very finely divided
particles, mainly fatty in nature, which manifest very characteristic
Brownian movements.

Reaction. The Lymph has an alkaline reaction, which is,

however, less marked than that of the blood.

Taste and Its taste is saltish, and it has a slight indefinite

Smeu. odour which varies somewhat in different animals.

Specific The statements of authors vary in regard to the

Gravity. specific gravity. According to Owen Eees and Marcet

the specific gravity varies between 1012 and 1022.

Coaguia- In a time which varies between 3 and 20 minutes

Won of Lymph. after it has left the vessels, the lymph undergoes
coagulation which is identical with that of liquor sanguinis. A soft
trembling jelly is at first formed, and after some time a contracted
colourless coagulum floats in a colourless or yellowish liquid, which
we may term the lymph-serum.

The fibrin which separates from coagulated lymph is identical with that
of blood. Very great differences exist in the rate of coagulation of lymph.
As a rule lymph which is flowing rapidly coagulates less rapidly than
lymph which is flowing slowly ; there is no rule to be laid down however.
Some lymph does not coagulate at all1.

1 Ludwig, quoted by Gorup-Besanez, Lehrbuch, &c., p. 378.

CHAP. V.] PROTEIDS AND FATS OF LYMPH. 223

The Proteids of the Lymph.

These consist of fibrinogen, of a globulin presumedly identical
with serum-globulin, and of serum-albumin.

The amount of fibrin which separates from the lymph varies
between 0'4 and 0*8 per 1000, being, therefore, much less in quantity
than that which separates from the blood. Accurate data are wanting
in reference to the amount of globulin, over and above the fibrinogen,
which the lymph contains.

The amount of serum-albumin, found in different specimens of
lymph, appears to have varied within wide limits, probably between
21 and 60 parts per 1000.

From certain observations of Wurtz it would appear that lymph
yields only about one-fourth of the amount of fibrin which is furnished
by the liquor sanguinis, and that it contains rather less than half the
amount of serum-alburnin contained in that fluid.

The Fats of the Lymph and Chyle.

The amount of fatty matters in the lymph of fasting animals is
small. Gubler and Quevenne on one occasion found the lymph
obtained from a lymphatic fistula in the leg of a woman, to contain
9'2 parts of fat per 1000, but this perhaps represents the highest
limit. In most analyses of lymph, the amount of fat found has
been smaller. In the chyle the amount of fat is immensely
greater. In his recent researches on the absorption of fat and its
passage through the thoracic duct, Zawilski1 has found that the fluid
obtained from the thoracic duct of animals fed upon a purely fatty
diet may contain the enormous proportion of 14'6 per cent, of fat, viz.
about three times as much fat as average milk. Under the heading
of fats are, however, included certain bodies which are not properly
fats, viz lecithin and cholesterin. Hoppe-Seyler analysed the ether
extract of chyle obtained from a fistula in the human subject and
found it to have the following composition : —

In 1000 parts of the ether-extract.

1st portion. 2nd portion.

Cholesterin . . . 113'2 1409

Lecithin .... 75'4 88'4

Olein 381-3

Palmitin and Stearin 4301 7707

The Extractive matters of Lymph.

Like the other constituents of the lymph, the so-called extractive
matters vary very greatly in proportion in different specimens. The
best known of these extractive matters are sugar and urea, though
others, such as lactic acid, leucine and tyrosine have been discovered.

1 Zawilski, "Dauer und TJmfang des Fettstromes durch den Brustgang nach Fett-
genuss." Ludwig's Arbeiten, Vol. xi. (1876) p. 147—167.

224

SALTS OF LYMPH.

[BOOK i.

chyle.

Sugar It has long been known1 that the lymph contains

suSar> and h has lately been shewu bJ v- Mehring2
that the amount of sugar in the lymph is approxi-
mately the same as in the blood. It had been stated
by Bernard that the lymph of the intestinal tract (chyle) does not
take up sugar when animals are fed upon a starchy or saccharine diet,
and the statement is confirmed by v. Mehring.

Urea Urea is a constant ingredient of the lymph and

present in the chyle, as was first pointed out by Wurtz 3. The amount
lymph. Qf ureaj }ike that of sugar, appears to be the same

in the lymph and blood.

The following are the results obtained by Wurtz; although,
owing to the method employed, the amount of urea found was much
below the actual amount, the observations are doubtless comparable
with each other.

QUANTITY OF UEEA FOUND IN 100 PAETS OF BLOOD, LYMPH AND

CHYLE.

Animal.

Blood.

Lymph.

Chyle.

Dog
Cow

0-009
0-019

0-016
0-019

0-019

Horse

0012

Bull

0-021

0-019

Other ex-
tractives

According to Lehmann the chyle of the horse
tractives contains alkaline lactates, and according to Frerichs

present in the and Staedeler, leucine and tyrosine are also present in

lymph and lymph; no definite information on these subjects is yet
chyle. J .V ->_-, J J

available.

The Salts of the Lymph.

Like the other constituents of the lymph, the sadts vary consider-
ably in proportion according as the fluid is more or less rich in water.

The salts are relatively much more abundant than the organic
solids, so that we may say that in transuding through the walls of
the blood-vessels, the liquor sanguinis furnishes to the lymph a small
quantity of its fibrinogen, about one-half of its serum-albumin, and a
much larger proportion of its salts.

The composition of the salts of the lymph and chyle appears to
be the same as that of the salts of the liquor sanguinis, in both cases
sodium chloride constituting the overwhelming constituent.

1 Gubler and Quevenne, Comptes Eendus, Vol. XLVI. p. 677.

2 v. Mehring, "Ueber die Abzugswege des Zuckers aus dcr Darmhohle." Ludwig's
Arbeiten, 1877.

3 Wurtz, Comptes Eendus, July, 1859.

CHAP. V.] THE LYMPH AND CHYLE. TPvANSUDATIONS. 225

The Gases of the Lymph.

The lymph contains carbonic acid, nitrogen, with, traces of oxygen,
all removable by the mercurial pump. The composition of the gases
of the lymph, especially the proportion and condition of the CO2
contained in that liquid, has formed the subject of elaborate investi-
gation in the laboratories of Leipzig and Bonn, because of the light
which the investigation promised to throw on the seat of the processes
of oxidation in the economy. In discussing that question, in another
section of this work, we shall again revert to the conclusions which
have been drawn from the study of the gases of the lymph, though
we think it right to give a systematic account of these in this place.

The first researches were made in the Leipzig laboratory, under
Professor Ludwig's direction, by Hammarsten1. They shewed that
pure lymph, unmixed with blood, contains either no oxygen or mere
traces of that gas ; that it contains C02 in quantity greater than is
contained in arterial, but smaller than is contained in venous blood ;
that it contains about the same quantity of N as is present in the
blood. The following are some of the actual results obtained by
Hammarsten.

VOLUMES OF GASES (MEASUEED AT 0°C. AND 760 MM. PEESSTJEE) YIELDED
BY 100 VOLUMES OF LYMPH, OBTAINED FEOM DIFFEEENT LYMPHATIC
VESSELS OF THE DOG. (HAMMAESTEN.)

0 C02 N
I. Lymph from the left foreleg, quite free

from blood . . . ' . . O'OO 41-89 112

II do 010 4713 1-58

IIL do. . .' . . . . 0-00 44-07 1'22

IV. Lymph from the thoracic duct 010 37'55 T63

V. The same lymph as IV. after being kept

for 24 hours in ice ... 0'05 37*50 T82

VI. Lymph from the thoracic duct, con-

taining a little haemoglobin . . 0'04 38'88 118

A second observer, Tschiriew2, pursuing the same subject, under
Ludwig's direction, obtained the following results, which shew the
simultaneous composition of the gases of lymph, of blood, and of
serum of blood, in dogs in an asphyxiated condition.

1 Hammarsten, " Ueber die Gase der Hundelymplie." Ludwig's Arbeiten,l87l.

2 Tschiriew, "Die Unterschiede der Blut- und Lyrnphgase des erstickten Ihieres.
Ludwig's Arbeiten, 1875.

G. 15

226 THE GASES OF THE LYMPH. !

BOOK I.

VOLUMES OF GASES (MEASURED AT 0°C. AND 760 MM.) YIELDED
BY 100 VOLUMES OF LYMPH, BLOOD AND SERUM (TSCHIBIEW).

I. Dog not under the influence of curare, bat

asphyxiated.

0 C02 N

Lymph 001 42-06 079

Blood .... 0-04 4278 170

Serum 0'09 48'3S 0'56

II. Same conditions as in I.

0 C02 N

Lymph O'Ol 5375 0'83

Blood 0-04 58-28 1-38

Serum . . . O'Oo 65'83 T92

III. Dog poisoned with curare, and

asphyxiated.

0 C02 N

Lymph O'Ol 41'25 T38

Blood Ill 45-18 1-84

Serum 013 5078 1*50

A third observer, Buchner1, continuing the observations of
Tschiriew, found that in asphyxia, as the quantity of carbonic acid
in the blood increased, that in the lymph diminished.

Tension of From these researches, which do not, it is true, teach

theco2of us the comparative tension of the gases of the lymph

Lymph. anc[ blood, it was reasonable to come to the conclusion

that probably the tension of the CO2 of the lymph was smaller
than that of the blood. Direct experiments made by Pfluger2 and
Strassburg3 indeed shewed that the carbonic acid of the lymph has
a tension, slightly but decidedly, less than that of the blood. Ac-
cording to the views which formerly at least were held by many very
eminent physiologists, this result seemed to localize the formation of

1 Buchner, "Die Kohlensaure in der Lymphe des athmenden und erstickten
Thieres." Ludwig's Arbeiten, 1876.

2 Pfluger, "Die Gase der Secrete." Archiv f. die aesammte Physiologie, Vol. n.
(1869) p. 156.

3 Strassburg, " Topographie der Gasspannungen im thierischen Organismus."
Pfliiger's Archiv, Vol. vi. pp. 65—96.

CHAP. V.] THE LYMPH AND CHYLE. TRANSLATIONS. 227

carbonic acid within the blood-vessels rather than in the tissues ; if, it
might be argued, C02 is formed in the tissues and passes into the
blood, it can only do so in virtue of the CO2 having a higher tension
in the extra-vascular liquids than in the blood. The answer which
has been given to this objection may be summarized as follows : —
It is conceivable, and indeed most likely, that the tension of the CO2
at the seats of its formation (in and near the anatomical elements
of the tissues) may be much higher than that of the lymph. If
.instead of analysing the lymph we analyse the normal secretions
of the body, such as the urine, bile, saliva, &c., which result
more directly from the action of the anatomical elements, we shall
be analysing liquids whose gaseous tension will, in all probability,
more nearly represent that of the tissues which are the seat of the
respiratory combustion. Now the tension of the C02 of these liquids
is much higher than that of the lymph, and higher even than that
of venous blood.

All difficulty in explaining the passage of carbonic acid into
the blood has, however, been removed by the last investigation on the
gases of the lymph made in the Leipzig laboratory. Gaule1 has
determined the comparative tension of the CO2, of blood, lymph and
serum, and has shewn that whilst the quantity of that gas in the
serum is greater than in the lymph, the tension of the C02 is
much greater in the lymph than in the serum. The same difference
will doubtless hold between the tension of the lymph and the tension
of the liquor sanguinis, and as we may consider the exchange of CO2
to occur in the first place between those two liquids, its passage into
the blood is easily accounted for.

The following are the results of one of Gaule's experiments :

PEECENTAGE OF C0.2, AND TENSION OF THE GAS, IN THE BLOOD-
SEEUM AND LYMPH OF AN ASPHYXIATED DOG.

C02 in 100 vols. Tension in mm. of Mercury.

(Temp. 40° C.)

Blood 24-6 567

Serum 34'5 33'4

Lymph 25%5 521

This subject will again be referred to at length in discussing the
Respiration of the Tissues.

1 Gaule, "Die Kohlensaurespannung im Blut, im Serum und in der Lymphe."
Ludwig's Arbeiten, 1878, and Archiv fur Physiologic of Du Bois-Keymond, 1878, p. 469.

15—2

228

ANALYSES OF THE LYMPH.

[BOOK i.

RESULTS OF THE QUANTITATIVE ANALYSES OF LYMPH
AND CHYLE MADE BY VARIOUS OBSERVERS.

I. ANALYSES OF THE LYMPH OF MA.N.

Constituents

Gubler i Marchand

Dahnliardt

Odenius

in

and

and

Scherer.

and

and

100 parts.

Quevenne.

Colberg.

Hensen.

Lang.

I.

H.

HI.

IV.

V.

VI.

Water

93-99

93-48

96-93

95-76

98-63

94-36

Solid Matters

6-01

6-52

3-07

4-24

1-37

5-64

Fibrin

0-05

0-06

0-52

0-04

0-11

0-16

Albumin

4-27

4-28

0-43

3-47

0-23

2-12

Fat

0-38

0-92

0-26

-1

0-1 5

2-48

Extractive Matters

0-57

0-44

0-31

f
~ )

0-16

Salts

0-73

0-82

1-54

0-73

0-88

0-72

II. ANALYSES OF THE LYMPH OBTAINED FEOM THE LYMPHATICS
OF THE HORSE (C. SCHMIDT).

Constituents in 1000 parts.

I.

II.

Water

963-93

955-36

Solid Matters

36-07

44-64

Fibrin

•\

Albumin
Fats and fatty acids

1

28-84

34-99

Other organic matters

J

Inorganic matters

7-22

7-47

NaCl

5-43

5-67

Na2O

1-50

1-27

K20

003

0-16

SO8

0-03

0-09

P2O6 combined with alkalies

0-02

0-02 '

Ca3(P04)2
Mg3(P04)2

}

0-22

0-26

In the serum from 1000

parts of Lymph

Schmidt found :

Albumin
Fats and fatty acids

)

23-32

30-59
1-17

Other organic matters

4-48

1-69

CHAP. V.] THE LYMPH AND CHYLE. TRANSUDATIONS.

229

HI. ANALYSES OF CHYLE OF THE HOKSE, DOG AND MAN1.

Constituents
in
1000 parts.

I.

Chyle of
Horse.

H.

Chyle of
Horse.

III.
Blood-
serum.

IV.

Chyle of
Dog.

V.

Blood- se-
rum of
Dog IV.

VI.

Chyle of
Man.

Water

960-97

956-19

930-75

906-77 '

936-01

904-80

Solids

39-03

43-81

69-25

96-23

63-99

95-20

Fibrin

2-57

1-27

Ml

1

Albumin

22-60

29-85

56-59

21-05

45-24 J

70-8

FatsCholes-]

terin and > 0'09

0-53

—

64-86

6-81

9-2

Lecithin '

Fatty acids)

in the formV 0'76

0-28

1-571

of soaps J

I

2-34

2-91

10-8

Other organ- 1
ic matters j

5-37

2*24

3-85 J

Haematin

0-05

0-06

,

Mineral salts

' 7-59

7-49

7-14

7-92

8-76

4-4

Loss

0-27

NaCl

5-76

5-84

5-74

Na0O ]

i -^i

1-17

0-87

K26 }

1 ol

0-13

0-14

S03

0-07

0-05

0-11

PA

0-01

0-05

0-01

Ca3(P04)2 I
Mg3(P04)2 /

0-44

0-25

0-26

C02

1-02

0-82

0-56

SEC. 2. THE LIQUIDS CONTAINED IN THE HEALTHY SEROUS
SACS. — SYNOVIA. — THE CEREBRO-SPINAL LIQUID.

The internal surface of the serous sacs of the body, such as
the pericardium, the peritoneum, the pleurae, &c., is, during life,
moistened by a small quantity of a liquid which must be looked
upon as lymph. These serous sacs are, indeed, in direct communi-
cation with lymphatic vessels, and offer the most highly differentiated
examples of the lacunar origin of those vessels.

After death it is usual to find in certain of the serous sacs,
especially in the pericardium, a small accumulation of the so-called
liquor pericardii ; its presence in them in quantity is, however, not
to be considered as affording any ground for the belief that such
accumulations exist during life, but is rather to be accounted for as
due to the changes in the circulation which immediately precede

1 This Table is extracted from Hoppe-Seyler's Physiologische Chemie, pp. 595 and
596. Analyses I., II. and III. are by C. Schmidt; IV. and V. are previously unpub-
lished analyses by Hoppe-Seyler. VI. is the analysis of the chyle of a beheaded person.

230

ANALYSES OF SYNOVIA.

[BOOK i.

death. Probably, in the most healthy condition, the serous sacs are,
as was said above, merely moistened with lymph, the excess finding
its way, as soon as it is formed, into the open mouths of the lymphatics.
Our knowledge of the physical characters and chemical composition
of the liquids of serous cavities is, therefore, almost entirely derived
from their examination when increased in quantity, and will be fully
referred to in the succeeding section of this chapter.

The secretion of the synovial sacs of joints requires
a special description, as it differs in some important
particulars from the contents of the other serous sacs.

Synovia is a transparent, faintly yellow, slimy liquid, of alkaline
reaction. It contains a larger proportion of solid matters than the
fluid of other serous sacs, and is specially distinguished from them
by containing mucin.

According to the observations of Frerichs l whose analyses of
synovia are given below, the joints of animals which have been kept
at rest furnished more synovia than those in active exercise ; in the
latter it is more concentrated.

ANALYSES OF SYNOVIA (FBERICHS).

I.

H.

III.

Constituents

Synovia

Synovia

Synovia

in

of a

of a

of an

1000 parts.

new-born

stall-fed

Ox at

Calf.

Ox.

grass.

Water

965-7

969-9

948-5

Solid matters

34-3

30-1

51-5

Mucin

3-2

2-4

5-6

Albumin and Extractives

19-0

15-7

35-1

Fats

0-6

06

0-7

Inorganic Salts

10-6

11-3

9-9

Cereforo-
spinal Liquid.

Although not contained in a serous sac, the so-called
cerebro-spinal liquid must be placed by the side of the
liquids of serous cavities, inasmuch as it also is essentially identical
with lymph. It is a liquid which is contained in the meshes
of the sub-arachnoid connective tissue (as that tissue is called
which lies between the arachnoid and dura mater) and in the
ventricles of the brain, the latter being connected with the sub-
arachnoid space by a narrow canal leading into the fourth ventricle,
and sometimes termed the foramen of Magendie. A certain quantity
of cerebro-spinal liquid, which probably never exceeds two ounces, is
contained in the sub-arachnoid space during life, and permits of an
equalization of intra-cranial pressure under different conditions of
fullness of the cerebral blood-vessels.

Cerebro-spinal liquid is alkaline, of low specific gravity (about
1005), and usually does not coagulate distinctly when heated, though

1 Frerichs, quoted by Gorup-Besanez, Leltrbnch derpJnjs. Cliemie. 4te Auflage, 1873.

CHAP. V.] THE LYMPH AND CHYLE. TRANSUDATIONS. 231

it contains appreciable quantities of globulins. It contains a body
which, like glucose, reduces cupric oxide, as was first pointed out
by Professor Turner 1. The cerebro-spinal liquid is occasionally much
increased in quantity and the analyses of the liquid made under these
circumstances will be considered in the next section.

SEC. 3. THE LIQUID IN DROPSIES.

Preliminary remarks on the mode of production of Dropsies.

It has been stated that the lymph consists of the liquid which
has transuded from the capillaries and which brings into intimate
contact with the anatomical elements of the tissues those elements,
of the blood which they need for their maintenance and repair.

Under normal circumstances, the composition of the blood, and the
differences between the pressure in arteries and veins are so adjusted,
that only as much liquid transudes from the blood-vessels as can
find its way back to the venous system through the lymphatics.
Two sets of circumstances may, however, arise to disturb the
normal relation. Firstly, the composition of the blood may be so
changed that the transudation from it into the tissues may increase
very greatly. This is the case when the relative proportions of the
water and proteids of the liquor sanguinis are disturbed, the former
increasing and the latter diminishing.

Secondly, the normal difference between the arterial and venous
pressure may be disturbed by an actual increase of the latter, as
for example by some mechanical obstacle pressing upon large veins
and diminishing their lumen, or by an obstacle to an easy passage of
blood through the heart ; or, locally, the normal difference in pressure
may be disturbed by vaso-motor changes (as in local inflammations).

Under any of these circumstances, dropsical accumulations may
result, i.e. accumulations of liquid which has transuded from the
capillaries into extra-vascular spaces, and which cannot be carried
back to the venous system by the lymphatics — of liquid which must
be looked upon as lymph, modified though it is, no doubt, by the
circumstances under which it has been formed. The dropsies which
are due to a change in the composition of the blood are most apt
to be general and to affect, at any rate in the first place, the loose
areolar tissue, especially in dependent parts of the body. The most
typical example is afforded by the dropsy which occurs in the course
of Bright's disease, in which the loss of albumin, by transudation
through the renal capillaries into the urine, may in a few days so
alter the blood that general anasarca comes on. Another example
is afforded by the general dropsy which comes on in some cases of
anaemia, which may be due to a derangement of the metabolic pro-
cesses of the body, and is not necessarily (though it frequently is) de-
pendent upon the draining away of same important blood constituent.

1 Turner, "Examination of the Cerebro-spinal fluid." Proceedings of the Royal
Society, vii., 1854—55, p. 89.

232

THE LIQUIDS IN DROPSIES.

[BOOK i.

Dropsy due to an altered relation between arterial and venous
pressure is aptly exemplified by the dropsy -in certain cases of heart
disease, or which is due to the pressure of an abdominal tumour, or a
cirrhosed liver, upon the inferior vena cava. In these cases the dropsy
is not general, but only affects the vascular area connected with the
obstructed veins.

It was long ago pointed out by C. Schmidt that where dropsical
accumulations occur simultaneously in various regions as, for example,
in the subcutaneous connective tissue and in several serous sacs,
the composition of the extravasated liquid varies in the different situ-
ations in consequence of local peculiarities, so that if the liquids were
withdrawn and were to accumulate again, (the condition of the blood
remaining constant in the interval), the second accumulations would ex-
hibit the same absolute composition and relative differences as the first.

The different dropsical fluids may be arranged in the following
order, according to their richness in proteids :

(1) Pleuritic fluid : (2) Peritoneal fluid : (3) Cerebro-spinal fluid :
(4) Fluid of subcutaneous oedema.

The quantitative differences in composition presented by fluids
removed at the same time from different serous cavities and from the
subcutaneous areolar tissue may be illustrated by quoting the two
following series of analyses.

I. Composition of various dropsical fluids removed simultaneously from
the body of a person who had died of albuminuria (C. Schmidt1).

Fluid from

Pleura. Peritoneum.

Water in 1000 parts
Solid matters „
Organic „ „
Inorganic „ „

963-95

36-05

28-50

7-55

978-91
21-09
11-32

9-77

Oedernatous

Sub-

connective

arachnoid.

tissue of

extremities.

983-54

988-70

16-46

11-30

7-98

3-60

8-48

7-70

II. Composition of the dropsical liquid removed simultaneously from a
patient affected with albuminuria (Hoppe-Seyler2).

Fluid from

Pleura. Per
Water in 1000 parts
Solids ^ „ „
Albumin „ ,,
Ethereal Extract]
Alcoholic „ in

Aqueous „ \- 1000
Inorganic salts parts.
Loss

1 Schmidt, Zur diarakteristi'k der epid. Cholera, p. 116 et seq., quoted by Hoppe-
Seyler, Phys. Chemie, p. 602.

2 Hoppe-Seyler, Yirchow's^rc/iiy, Vol. ix. (1856) p. 257.

Pleura.

Peritoneum.

Oedema of feet.

957-59

967-68

982-17

42-41

32-32

17-83

27-82

16-11

3-64

5-27

0-50

1

3-71

14-59 i

1-10

r

10-94

9-00

1

0-12

CHAP. V.] THE LYMPH AND CHYLE. TRANSUDATIONS. 233

The constancy of composition presented by successive dropsical
translations into the same sac is well exemplified by the two following
series of analyses1.

I. Analyses of fluid removed from the pleural and peritoneal cavities
on two separate occasions (Scherer).

Fluid from Pleura. Fluid from Peritoneum.

1st

2nd

1st

2nd

Paracentesis.

Paracentesis.

Paracentesis.

Paracentesis.

Water . . .

935-52

936-06

952-99

960-49

Solid matters .

64-48

63-94

47-01

39-51

Fibrin . . .

0-62

0-60

0-32

Albumin . .

49-77

52-78

34-58

29-73

Ethereal extract

2-14

1-35

1-26

1-63

Alcoholic extract
Aqueous extract

l-84\
1-62J

1-61

3-02

2-12

Inorganic salts

7-93

740

7-22

5-94

II. Analysis of the fluid removed from the peritoneal sac in a case of
Cirrhosis of the Liver (Hoppe-Seyler2.)

1st Paracentesis. 2nd Paracentesis.

Water in 1000 parts 984-50 982-53 983-33

Solid matters „ 15-50 17-47 16-67

Albumin „ 6-17 773 6'11

Ethereal extract „ 0'34 0'16 0'25

Alcoholic extract „ 0'24 0-56 2'16

Aqueous extract „ 0'67 1'12 0'84

Inorganic salts, soluble 8'30 7'99 8'05

„ insoluble 0-16 0-14 0'19

Errors of analysis 0'38 0'23 0'93

Pressure of liquid in ) =23 '5 mm. 25 '25 mm.

peritoneal cavity J of mercury. of mercury.

General Characters of Dropsical Fluids.

Resemblance Dropsical fluids always present more or less resem-

to diluted blance to diluted liquor sanguinis. In most cases where a

Liquor San- serous sac which contains the liquid is not inflamed,
this does not coagulate spontaneously, but does so on
the addition of fibrin-ferment. The transudations of an inflamed
serous membrane, on the other hand, which are rich in formed elements,
yield spontaneous coagula of fibrin.

TheProteids Whether coagulating spontaneously or not, the trans-

contained in udations which accumulate within serous sacs contain

dropsical ac- some flbrinogen, as evidenced by the formation of a

cumulations. coaguium on tne addition of fibrin-ferment, or on

heating to 56° — 59° C. Serum-globulin and serum-albumin are

1 These Analyses are transcribed from pages 602 and 603 of Professor Hoppe-
Seyler's Physiologische Chemie.

2 Hoppe-Seyler, Virchow's Archiv, Vol. ix. (1856) p. 250.

234

THE LIQUIDS IN DROPSIES.

[BOOK i.

Saline
constituents.

Extractive
matters.

present in addition. Old dropsical accumulations within the serous
sacs are richer in proteids than those recently formed.

The salts of dropsical transudations are similar in
character and usually in amount also to those present in
the liquor sanguinis. They are most abundant in recent accumu-
lations.

The extractive matters of the blood, such as urea,
uric acid, sugar, occur in the transudations, in much the
same proportions as in the liquor sanguinis. In old extravasations
cholesterin is occasionally present, and more rarely bilirubin.

Gases. All dropsical extravasations contain gases, C02, 0,

and N, removable by boiling in a Toricellian vacuum.

Of these gases the first is most abundant, the second sometimes
absent, and the third is present in about the same proportion as in
the blood. The tension of the C02 is, in some cases, considerably
higher than in the blood (Ewald).

TABLE EXHIBITING THE VOLUMES OF GASES, MEASUEED AT 0<> C. AND
760 MM. FOUND IN VARIOUS PATHOLOGICAL TRANSUDATIONS BY
PLANER, STRASSBURG AND EWALD (HOPPE-SEYLER1.)

Ci

^

Translation.

loosely

firmly

Total
C02

0

N

Authority.

com-

com-

bined

bined

Fluid of Peritoneum

9-404

4-866

14-270

0-139

2-107

Planer.

„ Hydrocele

32-49

33-45

64-94

0-16

2-05

Strassburg.

„ Oedema of extremities

22-25

9-15

31-36

traces

traces

Ewald.

„ do. chronic

nephritis

21-88

31-18

53-06

)?

5)

55

„ Pleurisy

39-34

15-59

54-93

0-68

1-33

11

,, do. in a case of

phthisis

18-54

25-99

44-53

0-54

1-87

)J

„ Hy drothorax, in a case

of Bright's disease

18-99

34-82

53-81

0-36

1-95

11

T)1 * J?J.

i

j

,, Jrleurisy alter recur-

rent fever

20-92

38-03

58-95

3-

16

11

„ Pleurisy with peri-

carditis

18-64

41-16

59-80

i

J

)

„ Tubercular pleurisy

25-47

46-82

72-29

0-17

1-04

J

„ Hydrothorax

25-34

48-67

74-01

0-29

i

0-87
__^

J

„ do. (of left pleura)

27-70

56-30

84-00

— IT

3-.

24

)

„ do. (double)

25-71

55-50

81-21

1-01

2-47

J

Iloppe-Seyler, Pliysiuloyisclie Chemie, p. 611.

CHAP. V.] THE LYMPH AND CHYLE. TRANSUDATIONS. 235

Characters of particular Transudations.

Having discussed the general characters presented by the transu-
dations which constitute the various forms of dropsy, it is necessary
to refer to special facts connected with certain of these liquids.

Pleura! The liquid which accumulates in the pleural cavities

transuda- in hydrothorax is clear, faintly yellowish, inodorous, and

free from viscosity; it is possessed of an alkaline re-
action ; its specific gravity is low, usually between 1010 and 1015.

In acute pleurisies, the liquid removed by paracentesis soon
coagulates, the fibrin which separates amounting to 0'4 or even 0'5
per thousand. Its specific gravity is above 1018. The amount of
solid matter exceeds 50 parts per 1000.

In chronic pleurisies fibrin does not usually occur, and the propor-
tion of albumin in the transudation increases.

Several analyses of pleural transudations have been given at
pages 232 and 233.

Peritoneal Possessed of a faint yellow colour, density varying

transuda- between 1005 and 1024. The liquid does not co-

tions, Ascitic adulate spontaneously unless there have existed some
liauid ° •, ',•

peritonitis. .

Some analyses are given at pages 232 and 233.

Liquid ef- Usually is colourless ; often spontaneously coagulable.

fused into the Contains a larger quantity of fibrinogen than other
Pericardium. transudations. Contains from 0'879 to 2'468 p. c. of
albumin (Kuhne).

Liquid of The density of hydrocele liquid oscillates between

Hydroceie. 1016 and 1022. Its colour is usually a very faint lemon-
yellow> but may be much darker ; it sometimes has a greenish tint ;
sometimes it is slightly viscous.

It contains a large quantity of globulins and serum-albumin, in
addition to the fibrinogen which has caused it to be the favourite
liquid for experiments on the formation of fibrin. In some cases it
contains a large quantity of cholesterin (1 — 5 p. c.). Succinic acid
has sometimes been found in it.

The following is the mean of 17 analyses of hydrocele liquid made
by Hammarsten.

Water in 1000 parts 938'85

Solid matters J „ 6M5

Fibrin (derived from fibrinogen) 0*59

Globulins 13'52

Serum-albumin 35'94

Ethereal extract 4'02

Soluble salts 8'60

Insoluble salts 0'66

NaCl 619

23G ANALYSES OF LYMPH, CHYLE, &C. [BOOK I.

In 12 analyses made by Hoppe-Seyler the solid matters of the
liquid of hydrocele varied between 41 '4 and 80'2 and the proteids
between 29 5 and 65 parts per 1000.

Cerebro-spi- In cases of spina bifida and chronic hydrocephalus

nai Liquid. large accumulations of liquid occur, which presents a
close resemblance to normal cerebro-spinal liquid. The liquid is
clear, has a low specific gravity, and contains usually from 10 — 13
parts of solid matters per 1000.

Sugar has been described as a normal constituent of cerebro-
spinal liquid, or at least a substance having a similar reducing action
as sugar upon cupric oxide (see p. 231). According to Hoppe-Seyler,
sugar is not a normal constituent of this fluid, and only occurs as
a result of irritation or inflammation of the brain or spinal cord.

Cerebro-spinal liquid, when boiled, becomes opalescent, without
yielding a flocculent precipitate, which only separates after the
addition of acetic acid (Hoppe-Seyler).

Cerebro-spinal liquid differs from other transudations in being
usually free from fibrinogen, and therefore not yielding a coagulum
of fibrin when treated with fibrin-ferment.

Carl Schmidt found that the cerebro-spinal liquid is remarkably
rich in salts of potassium — an observation which is well worthy
of being checked by fresh analyses of the liquid obtained by punc-
turing in cases of spina bifida.

The following are analyses by Hoppe-Seyler of the cerebro-spinal
liquid, obtained by puncture, in cases of spina bifida1.

ANALYSES OF THE CEEEBEO-SPINAL LIQUID, OBTAINED BY PTJNCTUKE,
IN TWO CASES OF SPINA BIFIDA (HOPPE-SEYLEE).

I. II.

1st 2nd 3rd 1st 2nd

Puncture. Puncture. Puncture. Puncture. Puncture.

Water 987*49 986'88 98672 989'33 989'80

Solid matters 12'51 1312 13'28 10'67 10'20

Albumin T62 2'64 2'46 0'25 0'55

Extractives \ 10'27 2'83 2'65 2'30 2'00

Inorganic salts, soluble/ 7'52 . 8'21 7'67 7'20

insoluble 0'25 115 0'2S 0'45 0'45

SEC. 4. METHODS OF ANALYSING LYMPH, CHYLE, AND OTHER
TRANSUDATIONS NORMAL AND PATHOLOGICAL.

The methods of investigation are precisely similar to those
pursued in the analyses of liquor sanguinis and serum (see p. 187
et seq.}, with the exception of the estimation of fibrinogen.

The amount of fibrin which separates spontaneously may be

1 Hoppe-Seyler, Physiologische Chemie, p. 001.

CHAP. V.] THE LYMPH AND CHYLE. TRANSLATIONS. 237

ascertained by washing the coagulum from a known weight of the
transudation and proceeding as stated at page 180.

The fibrinogen may then be determined by one of two methods :
firstly, by Frederique's method (see p. 188) ; secondly, by adding to a
weighed quantity of the liquid separated from any coagulum some
very active solution of fibrin-ferment (see p. 49), and then placing for
36 hours in an incubator heated to 40° C. ; then, collecting any
coagulum which has separated, washing, and proceeding as directed
in the case of blood-fibrin at p. 180. In giving the results of the
analyses the amount of fibrin, corresponding to fibrinogen, is then
stated.

After separating fibrin and fibrinogen, the globulins remaining in
solution are estimated by Hammarsten's method (precipitation with
magnesium sulphate, see p. 188).

In a fresh portion of the fluid the total proteids are estimated by
precipitation with alcohol (see p. 187).

By then subtracting from the result thus obtained the weight of
fibrinogen and of globulins, the amount of serum-albumin is ascer-
tained.

The extractive matters, salts and gases, are determined exactly
as in the case of blood or serum.

CHAPTER VI.

PUS.

SEC. 1. INTRODUCTORY REMARKS ON THE PHYSICAL PROPERTIES
OF PUS AND ON THE NATURE OF PUS.

CLOSELY connected with the liquids which have been considered in
the preceding chapter is one which, unlike these, forms no part of
the healthy body, but is invariably the result of a morbid process.

Pus is sometimes found in one of the natural cavities of the body,
as, for example, within the interior of a serous sac : sometimes
covering an epitheliated surface on the exterior, or opening on the
exterior, of the body : most commonly contained within an abscess —
a cavity whose walls are constituted by inflamed and usually indu-
rated tissues.

Physical Fresh, healthy, laudable pus presents the appear-

characters. ance of a somewhat creamy yellow liquid, which
unless it have been obtained from the vicinity of the intestines, is
destitute of fostid odour and possesses, at most, a mawkish smell.

Its reaction is usually said to be alkaline, but, according to Ewald,
it is often acid. Its specific gravity varies between 1020 and 1040,
being on an average 1032. The fluid does not coagulate spon-
taneously.

Microscopi- Under the microscope pus is seen to be composed

cai charac- of a clear liquid — the pus serum — in which closely
ters* float a large number of cells which, when first formed,

resemble, if they are not identical with, the colourless cells of the
blood.

These cells are usually more or less spherical : destitute of a cell
wall : somewhat granular, and contain one or more (often three, some-
times more) nuclei, which are rendered evident by the action of acetic
acid, which causes the protoplasm of the cell to become transparent
and indistinct. When very young, pus cells may exhibit amoeboid
movements, though the opportunity for observing this phenomenon
does not often present itself.

The diameter of pus corpuscles usually varies between 8/j, and
by the action of water they swell and become transparent,

CHAP. VL] PUS. 239

allowing their nuclei to be seen, and the latter may then be readily
stained with magenta or even with carmine.

As usually obtained, pus corpuscles resemble dead rather than
living colourless blood-cells, as evidenced by the absence of contrac-
tility.

Pus corpuscles are liable to undergo certain changes, of which the
most common is fatty degeneration ; the cells then contain a number
of highly refracting, obviously fatty, granulations ; at a more ad-
vanced stage, the cells break down and the fatty granulations thus set
free float in the pus-serum.

Nature of The liquid portion of pus — pus serum — resembles

PUS and the liquor sanguinis and the normal transudations very

origin of Pus closely, and, doubtless, is in great part, in the first
cells. instance, a transudation from the blood. With regard

to the pus corpuscles, they are, for the most part, either colourless cells
of the blood which have wandered through the capillary walls into
the extra- vascular spaces, or the offspring of such emigrated cells ; in
some cases, however, it is possible that the pus cells are derived from
the normal cells of the tissues amongst which they are found,
especially from epithelial and endothelial cells.

SEC. 2. THE Pus SERUM.

The liquid in which the pus corpuscles are suspended may be
separated in an unmixed condition by filtration ; the process is, how-
ever, a tedious one ; it may be obtained more readily, by mixing pus
with an equal volume of a solution of one part of sodium sulphate in 9
parts of water arid then filtering; the liquid which passes through the
filter is then a mixture of pus serum and solution of sodium sulphate.
Doubtless the separation would, in either case, be much facilitated by
the use of the centrifugal apparatus.

Pure pus serum is a turbid liquid which has a brownish tint
when examined by reflected light, whilst by transmitted light thin
layers appear of a yellow colour. Its reaction is usually alkaline.

Proteid Pus serum contains substantially the same proteid

matters of matters as blood serum, viz. serum-globulin and serum-
pus serum. albumin ; the former is partly precipitated by C02, but
may, as in the case of blood serum, be completely precipitated by
saturating with magnesium sulphate.

Extractive These consist of a mixture of neutral fats, cholesterin,

matters and of a derivative of glycerin-phosphoric acid. This

soluble in derivative is, according to Hoppe-Seyler, probably the

same as lecithin, the phosphorized proximate principle of

the yolk of egg; according to Fischer1 it is protagon. The matter is yet

1 Fischer, Centralblatt f. d. med. Wissenscliaften, 1865, p. 225.

240 EXTRACTIVE AND SALINE MATTERS IN PUS. [BOOK I.

altogether unsettled. In the analyses quoted below the phosphorus
in organic combination is supposed to be present in lecithin.

Extractive Hoppe Seyler has found leucine and tyrosine in

matters perfectly fresh pus ; urea and sugar may also be present,

soluble in It has been alleged that gelatin and chondrin occasion-

alcohol and a}}y occur in pus? but these bodies, though sought for
by Hoppe-Seyler, have never been found by him.

The saline Of these the chief is sodium chloride. The other

constituents saline constituents supposed to be present by Hoppe-

of pus serum, geyler will be learned by referring to the next para-
graph.

Results of Probably the only analyses of the serum of pus

Hoppe- which can be looked upon as really trustworthy are

Seyier's those made by Hoppe-Seyler, of which the results are

appended. The pus was in each case obtained from an

acute abscess.

ANALYSIS OF THE SERUM OF PUS (HOPPE -SEYLERi).

I. II.

Proteids in 1000 parts 63'23 77'21

Lecithin „ „ 1-50 0'56

Fats „ „ 0-26 0-29

Cholesterin „ „ 0'53 0'87

Alcohol-extractives „ 1'52 073

Water-extractives „ 11 53 6'92

Inorganic matters „ 773 777

Solid matters „ „ 86'30 94'35

Water „ „ 91370 905'65

1000-00 1000-00

ANALYSIS OF THE ASH OBTAINED BY INCINERATING THE PUS-SERUM
EMPLOYED IN THE ABOVE ANALYSES.

Quantity of various Saline Constituents in 1000 parts of Pus Serum.

I. H.

NaCl . . . 5-22 5-39

Na2SO4 . . 0-40 0-31

Na2HP04 . . 0-98 0'46

Na2CO8 . . 0-49 113

Ca8(P04)2 . . 0-49 >31

Mg3(P04)2 . . 019 012

PO 4 found in excess 0'05

"777 Wf

1 Hoppe-Seyler, " Ueber die quantitative Zusammensetzung des Eiters." Med.
chem. Untersuchung , p. 490.

CHAP. VI.] PUS. 241

SEC. 3. Pus CORPUSCLES.

As has been mentioned in a preceding section, these corpuscles
may be obtained by mixing fresh pus with solution of sodium sulphate
and filtering ; the corpuscles left on the filter may be freed from
adhering pus serum by washing with an additional quantity of the
solution of sodium sulphate.

Action of Solution of common salt cannot be employed in the

Naci on the place of solution of sodium sulphate in the separation of

proteids of pus cells, for, under the influence of sodium chloride, the

the cell ceiis are converted into a slimy, opaque, jelly, precipitable
protoplasm.

The Proteids present in the Cell- protoplasm.

It was formerly supposed that the pus corpuscles contained a
considerable quantity of a proteid identical with myosin. The most
careful investigation yet made (by Miescher1) of the constituents of
pus cells failed to detect myosin.

According to this author three proteids soluble in water can be
obtained from the protoplasm of the pus cells, viz. (1) alkaline
albuminate, partially precipitated by C02, and more completely pre-
cipitated by acetic acid, insoluble in solution of sodium chloride, and
soluble in very dilute hydrochloric acid (1 to 1000 of water): (2) a
proteid coagulable at 48° — 49° ; the flakes which separate are insoluble
in dilute HC1 and in solution of NaCl: (3) a proteid which
coagulates at the same temperature as serum-albumin. In addition
to these, two proteids insoluble in water are also present, in prepon-
derating quantity, viz. (1) a body insoluble in water, swelling up in
solution of NaCl, soluble in very dilute hydrochloric acid (1 to 1000)
giving rise to acid-albumin : this is the body, formerly considered to
be identical with myosin, which occasions the peculiar phenomenon
observed when pus is mixed with solution of common salt ; (2) a body
unacted upon by water and by solution of NaCl, and attacked with
difficulty by dilute hydrochloric acid (1 to 1000).

The matter of the Nuclei. Nude in (?).

When pus corpuscles are subjected to the repeated action of
weak- hydrochloric acid, it occasionally happens that a considerable
number of free nuclei are obtained; the greater number, however,
have some remains of the cell protoplasm yet adhering to them.

By digesting pus cells in artificial gastric juice (made by di-
gesting the mucous membrane of pig's stomach in water containing
10 c.c. of fuming HCi in 1 litre) the nuclei of the pus cells are

1 Miescher, "Ueber die cliemische Zusammensetzung der Eiterzellen." Hoppe-
Seyler, Med.-chem. Untersuchungen, p. 441 et seq.

G. 1C

242 NUCLEIN. [BOOK i.

isolated in large quantities. In order to obtain them uncontaminated
with organic phosphorus compounds soluble in alcohol and ether, it is
advisable to treat the pus cells with hot alcohol before digestion.

The nuclei, isolated by the above method, form a grey mass,
insoluble in very dilute HC1, but soluble in very weak solutions of
sodium hydrate. From this solution, acids added in excess precipitate
an insoluble body, which, according to Miescher1, consists of a definite
organic body containing phosphorus, to which he ascribes the name of
Nuclein. This body is, according to Miescher, found in the nuclei of
the segmentation spheres of the yolk; according to Plosz2 it is the
principal constituent of the nuclei of the coloured blood corpuscles
of Birds ; and according to Hoppe-Seyler3 it is found in yeast. It
is said also to be present in brain and liver. Indeed, wherever nuclei
are found, nuclein has been surmised to exist.

Elementary Miescher4 has investigated the nuclein obtained from

composition of salmon-melt more closely than that obtained from pus-
Nuciein. cells, and has come to the conclusion that nuclein is a

tetra-basic acid, having the formula C^H^N^O^.

The following is the composition of nuclein according to Miescher.

Calculated. Found.

COQ 35-95 36-11

Hq 5-01 5-15

N ' 1302 13-09

P3 9-61 9-59

O0(1 36-41 3606

Hoppe-Seyler prepared and analysed nuclein from pus and
obtained numbers which differed entirely from those of Miescher.

The1 following numbers shew the wide discrepancy between the
analyses of Hoppe-Seyler and Miescher.

ANALYSES OF NUCLEIN.

I. II.

(From pus.) (From spermatozoa of the Sahnon.)
(Hoppe-Seyler.) (Miescher.)

C-. ...49-58 36-11

H 710 5-15

N 15-02.... 13-09

P 2-28 9-59

1 Miescher, Op. cit.

2 Plosz, "Ueber das chemische Verhalten der Kerne der Vogfl- und Schlangen-
blutkorperchen." Hoppe-Seyler's Med.-chem. Untersuchungen, p. 461.

3 Hoppe-Seyler, "Ueber die chemische Zusammensetzung des Eiters." Med.-chem.
Untersuchungen, p. 500.

4 Miescher, "Die Spermatozoen einiger Wirbelthiere, " (Protamin, Nuclein). Se-
paratabrduck aus den Verhandlungen der naturforschendcn Gesellschaft in Basel, Vol. vi.,
1874. Abstracted in Maly's Jahresbericht, Vol. iv. p. 337 et seq.

CHAP, vi.] PUS. 243

Does a Whether the body obtained by Miescher from

definite body spermatozoa be a definite body or not, there can
Nuciein exist? be no question that as yet, all proof is wanting to
establish the proposition, that the substance composing cell nuclei
generally is a definite chemical individual, possessed of constant
composition. On the contrary, the evidence of most trustworthy
observers shews, that by following the processes which have been re-
commended for the preparation of nuclein, substances of widely
differing composition are obtained.

In different samples of the nuclein of yolk of egg, Worm Miiller1 found
2'2, 2-68, and 7'9p. c. of Phosphorus. In miclein from the same source
Miescher found 6-7 and 7'1 p c. In nuclein prepared from casein Lubavin2
found 4'6 p.c. of P. In nuclein from pus Hoppe-Seyler found 2 '28 p.c. of
P. ; in that prepared from an epithelial tumour he found 3'35 p.c. In the
xmclein from, pus Miescher found 2*6 p.c. of P.

The statements as to the state in which the P. exists in the so-called
nuclein are also discrepant.

We must therefore agree with the conclusions of Worm Miiller,
and deny the existence of a definite body, Nuclein. Probably, as this
author surmises, the different nucleins are mixtures of organic
phosphorus compounds with varying quantities of proteid bodies.

The Extractive Matters of Pus Cells soluble in water.

Alleged It was asserted by Boedecker3 that pus occasionally

presence of contains, besides proteid matters proper, gelatin and

Gelatin and chondrin. Miescher examined the aqueous extract of

pus cells for these bodies with entirely negative results.

Boedecker's By the name of chlorrhodinic acid Boedecker described a

chlorrhodinic crystalline acid which he obtained from pus by the following
aci<l. method : — The liquid is evaporated to dry ness, and the

powdered residue is successively treated with ether, alcohol, and then with
water. The aqueous extract is precipitated with lead acetate, and the
precipitate decomposed by means of sulphuretted hydrogen and boiled in
alcohol. On evaporation, the alcoholic fluid deposits groups of microscopic
needles, mixed with some crystals of sodium chloride. The former are
composed of the acid, which, according to Boedecker, contains nitrogen.
Iodine colours it yellow, and chlorine water of a rose colour, or of a dark
red tint, according to the amount of acid present. These facts suggest a
re-examination of the subj ?ct.

1 Worm Miiller, " Zur Kenntuiss der Nucleine." Pfliiger's Archiv, Vol. vin. (1873),
p. 190.

2 Lubavin, "Ueber die kiinstliche Pepsin-Verdauung des Caseins." Hoppe-Seyler's
Med.-chem. Untersuchungen , p. 477.

3 Boedecker, Zeitschriftf. rat. Med., N. F., Vol. vi.

16—2

244 CONSTITUENTS OF PUS CELLS. [BOOK I

Presence of It has been shewn by Salomon1 that pus generally

Glycogen in contains very appreciable quantities of glycogen, and
PUS- this fact agrees with certain histological observations of

Kanvier2. Thus a distinction which Hoppe-Seyler3 sought to esta-
blish between the colourless cells of the blood and their descendants,
the pus corpuscles, can no longer be maintained.

The Extractive Matters of Pus Cells soluble in alcohol and ether.

The chief of these are cholesterin, fats and derivatives of glycerin-
phosphoric acid ; free fatty acids may be likewise present in old pus
and form crystalline deposits.

The Mineral Matters of Pus Cells.

These consist of a small quantity of sodium chloride and of earthy
phosphates. Their amounts, as found by Hoppe-Seyler, will be
learned in the next paragraph.

Results of

Hoppe- The following are the results of the analyses made

Seyier's ana- by Hoppe-Seyler of two samples of pus cells isolated by
lysis of PUS sodium sulphate.
Corpuscles.

a. Organic constituents in 100 parts of dried pus contained

(1) (2)

Proteids 13762 1

Nuclein 34'257[ 68'585 67'369

Insoluble matters 20-566J

Lecithin) I/L.QQQ 7"564

Fats ) 7-500

Cholesterin 7'400 7'283

Cerebrin 5199J 10'284

. Extractive matters 4'433J

100-000 100-000

b. Mineral constituents
100 parts of dried corpuscles contain

NaCl 0-435 parts

Ca3(P04)2 0-205 „

Mg3(P04)2 0113 „

Fe2(P04)2 0-106 „

P04 0-916 „

Na 0-068 „

K traces.

1 Salomon, " Untersuchungen betreffend das Yorkommen von Glycogen im Eiter
und Blut." Deutsche med. Wochenschr. 1877, No. 35. Abstracted in Maly's Jahres-
bericht, Vol. vn., p. 130. Salomon, "Ueber das Vorkommen von Glycogen im Eiter."
Verhandlungen derphysiol. Gesellsch. zu Berlin. Jahrg. 1877-78, No. 19.

2 Eanvier, Progrts Med. 1877, p. 422.

3 Hoppe-Seyler, "Ueber die chemische Zusammensetzung des Eiters.'' Med. Chem-
Untersuch., p. 497.

CHAP. VI.] PUS. 245

SEC. 4. COLOURING MATTERS FOUND IN Pus.

Pus is sometimes coloured with bilirubin. At other times it
presents a brownish colouration very similar to that due to bilirubiu,
but without giving the reactions of this body. More rarely it has a
blue or green colour.

Pyocyanin.

It has long ago been known that the pus of old sores sometimes
presents a blue or green colouration, especially the former.

Fordos1 shewed that under these circumstances a blue colouring
matter is formed to which he gave the definite name of pyocyanin.

Mode of The material employed by Fordos consisted of

preparation bandages stained with blue pus. These are steeped in
of Pyocyanin. a weak solution of ammonia, which dissolves the colouring
matter and acquires a blueish or greenish tint. The ammoniacal
solution is shaken with chloroform, which dissolves pyocyanin, fat,
and a yellow colouring matter.

The chloroformic solution is shaken with water holding a little
sulphuric acid in solution, when the colouring matter assumes a red
instead of a blue or blueish-green colour, and, leaving the chloroform,
is dissolved by the acid solution. The supernatant red liquid is then
separated, mixed with chloroform, and a little solution of caustic
baryta added, which causes the red colouring matter to become blue
again and to be taken up by the chloroform. On now evaporating
this liquid, pyocyanin is obtained, in the form of blue needles or of
rectangular plates. These are soluble in water, alcohol, and chloroform.
Pyocyanin possesses, as will have been gathered by the reader from
the above description of the mode of preparation, some of the pro-
perties of a blue vegetable colouring matter.

Properties Pyocyanin is soluble in water, alcohol, chloroform,

of Pyocyanin. and ether. It is decolourized by chlorine, concentrated
nitric acid, and ozone : it is blue in the presence of alkalies, red
in that of acids. When blue pus is kept from contact with air the
colour disappears, to reappear again when the liquid is shaken with air.
Pyocyanin has not hitherto been analysed.

Pyocyanin a It has long been known that by being placed in

vegetable proximity to a wound of which the purulent discharges

are blue, suppurating surfaces which produced yellow pus

are apt to furnish a liquid which is also blue or green.

Liicke2 thought he had discovered that the blue colour is due to a blue

vibrio developing in the pus. The more recent investigations of Fitz3

1 Fordos, " Recherches sur la matiere colorante des suppurations bleues : Pyocyanin."
Paris, Comptes Rendus, LI., 1860, p. 215.

2 Liicke, "Die sogenannte blaue Eiterung u. ihre Ursachen." ArcMv f. KliniscTie
Chirurgie, in., 1862, p. 135.

3 Fitz, see an abstract entitled 'Recent Researches on Bacteria,' Quarterly Journal
of Microscopic Science, Jan. 1880, p. 106, from which the above account of Fitz's
researches is taken almost verbatim.

24G COLOURING MATTERS AND GASES OF PUS. [BOOK I.

have shewn that the generator of pyocyanin is a species of bacterium,
which has the form of a bacillus, and which possesses in addition the
property of decompo'sing glycerin in the presence of calcic carbonate,
with the formation of hydrogen, carbonic acid, butyl- alcohol and butyric
acid. Fitz cultivated the pus-bacillus in a solution of calcium lactate
and ammonium chloride, and obtained, in the solution, a colourless
reduction-product of the colouring matter, which was only blue on the
surface, but which when shaken with air assumed throughout a deep
blue colour similar to that of a solution of copper sulphate. These
properties of the blue colouring matter agree exactly with those
described by Fordos and Llicke. The bacteria which produce it,
and which multiply luxuriantly in the cultivating liquid, are small
elliptic bodies, from one to one and a half micromiJlimetres in length,
and generally occur hi couples.

It has been asserted by Herapath that indigo-blue occasionally occurs
in blue pus.

Pyoxanthose.

The above term was applied by Fordos1 to a greenish-yellow
colouring matter, already referred to in the description of the prepara-
tion of pyocyanin, accompanying that body, and like it soluble in
chloroform. It may be separated from pyocyanin by ether, in which
it is more soluble than the latter body. It crystallizes in yellow
prisms. It is soluble with difficulty in water, but readily soluble in
alcohol, ether, chloroform, bisulphide of carbon and benzol. It is
coloured red by acids, and violet by alkalies.

SEC. 5. THE GASES OF Pus.

When pus is subjected to the process employed for the extraction
of the gases of the blood — viz. heated in a Toricellian vacuum — it
gives off a mixture of carbon dioxide, oxygen and nitrogen, in which
the first-named is much the most abundant constituent.

Ewald2, to whom we owe the greater part of our
from Ewaid'a knowledge of the gases of pus, has arrived at con-
researches. elusions which may be stated briefly as follows :

1. Fresh pus yields to the Tori3ellian vacuum only carbon
dioxide, oxygen and nitrogen, the two latter gases being present
in very small quantities. It never evolves hydrogen (as had been
asserted by Mathieu and Urbain3), or sulphuretted or carburetted
hydrogen.

1 Fordos, " Recherches sur la mature colorante des suppurations bleues : Pyocyanin
et Pyoxanthose." Paris, Compies Rendus, LVI., 1863, p. 1128.

s< Ewald, " Untersuchungen zur Gasometrie der Transudate des Menschen." Archie
filr Anatomie und Physiologie, 1873, pp. 663—698. An exceedingly full and clear
abstract of this most valuable memoir is to be found in Maly's Jahresbericht, Vol. iv.,
pp. 421—431.

3 Mathieu et Urbain, Gazette Itebdomadmre, 1871, No. 24, and 1872, No. 21 (quoted
by Gautier).

CHAP. VI.]

PUS.

247

2. The amount of C02 contained in a purulent or sero-purulent
exudation increases with the age of the exudation.

3. The amount of C02 contained in a purulent exudation
is small in proportion as the exudation approaches pure pus in its
characters. This is not only true of the total C02, but particularly of

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248 QUANTITATIVE ANALYSIS OF PUS. [BOOK I. CH. VI.

the combined CO2 which is given off on the addition of an acid;
indeed pure pus contains only free CO./.

4. Pus corpuscles, — as doubtless also the colourless cells of
the blood, — possess the property of decomposing sodium carbonate
(Na2C03) and evolving from it COa.

5. Pus corpuscles and the colourless cells of the blood are either
altogether free from oxygen or contain mere traces of this gas.

SEC. (j. DIRECTIONS FOR THE QUANTITATIVE ANALYSIS OF Pus.

1. Determine the specific gravity by means of the bottle (see
p. 174).

2. Ascertain the reaction.

3. Determine the total solids, water and salts, as in the case of
blood (see p. 177).

4. Evaporate a known weight of the fluid, say 25 grin., to dryness.
Extract with ether and determine the amount of the ether extract.
If wished, determine in the latter the amount of cholesterin, lecithin
and fats, as in the case of blood (see p. 187).

5. Treat the residue after extraction with ether, with boiling
absolute alcohol, filter, evaporate the solution to dryness; weigh,
then ignite and weigh again. By subtracting the second from the
first weight the amount of the alcoholic extractive matters is found.

6. Mix a weighed quantity of pus with ten times its volume of
alcohol, set aside for 24 hours and proceed exactly as in Schmidt's
method for determining the total amount of proteids in the serum
(p. 188). In this way will be found the combined weight of the
proteids of the pus serum and of the corpuscles, together with
nnclein. The residue may then be boiled in water, and, after cooling,
subjected to artificial peptic digestion in the incubator for 24 hours.
The insoluble matters may be treated with a fresh portion of artificial

fastric juice and the process continued for a second period of 24
ours. The insoluble residue is then collected on a weighed filter,
washed successively with boiling water, alcohol, and ether, and then
dried; thus will be found the weight of the dry nuclei (nuclein?).

7. A portion of the pus may be filtered and the solids, salts,
extractives, &c. of the pus serum be determined, by following precisely
the methods recommended in the case of blood serum.

8. If it be required to determine the presence of urea, sugar, or
any other extractive matter, the methods recommended in the case of
blood may be followed.

9. For the separation and estimation of the gases of pus the same
proceedings are adopted as with blood.

1 The liquid contained in the pleural cavity, if not purulent and of old standing,
always yields a larger proportion of firmly combined than of loosely combined or
free C02.

CHAPTER VII.

THE CONNECTIVE TISSUES.
INTRODUCTION.

UNDER the term of 'the connective tissues,' histologists have grouped
together several tissues which at first would appear to have few
points in common — to wit: connective tissue proper, including the
white connective tissues and yellow or elastic tissue : cartilage : bone :
and dentine.

When we enquire into the grounds of this classification we
find that they are the following : — The tissues above named are
derived from the same embryonic layer1 (mesoblast) ; they all perform
similar, subordinate, functions of support or connection; they all
contain cells which develop a matrix or ground substance, which
has various characters in the various tissues; they shade off, as it
were, into one another, and represent each other in different species
of animals. "In one and the same organism typical development
brings with it a substitution of one member of the connective-
substance group for another. There, for instance, where in the
embryonic state gelatinous tissue existed, the latter is found trans-
formed into connective tissue or fat at a later epoch ; cartilage with
its derivatives takes on the form of bony substance. Finally we
encounter every kind of this substitution in the richest abundance,
brought about by the formative activity of a system modified by
disease. Almost every member of the group of connective tissues
may be replaced by very nearly any other, firstly by immediate
metamorphosis, then again more particularly by reconstruction from
the offspring of the original tissue2."

1 Tkis is not strictly true. The neuroglia, or connective tissue of the great nerve
centres and of the retina, is epiblastic in its origin; chemically, however, this tissue
differs from connective tissue, so that it is really true that true (collagenous) connective
tissues are derived from the mesoblast.

3 Frey: The Histology and Histochemistry of Man, translated from the fourth
German edition by Arthur E. J. Barker, London, 1874, p. 167.

250 VARIETIES OF CONNECTIVE TISSUE PROPER. [BOOK I.

SECT. 1. CONNECTIVE TISSUE PROPER.

structural By ^is term may be designated a tissue which pre-

Eiements of sents many very important modifications in different
connective situations, and whose function it is to connect together
Tissue. contiguous organs or parts of the body, or actually to

bind together the different anatomical elements which enter into the
composition of each organ.

Typical connective tissue presents for examination :

(1) Certain cells, which are especially abundant in the early
stages of development of the tissue, and which are termed connective
tissue cells or corpuscles.

(2) Bundles of fine fibres of a white colour, arranged in
parallel rows ; or crossing one another, so as to leave spaces between
them ; or so interwoven as to give rise to tough fibrous membranes.
These fibres swell up and become so transparent as almost to dis-
appear from view when the tissue is treated with acetic acid.

(3) Other fibres, usually much less numerous than the white,
presenting dark outlines, often intercommunicating by processes,
having when seen in large numbers a yellowish tint, and exhibiting
altogether distinct chemical reactions ; they are unacted upon by
acetic acid. These are the yellow, or elastic fibres of connective tissue.

(4) A ground substance or matrix in which the other elements
are imbedded and which serves to connect them together, so
that we apply to it indifferently the name of ground substance, or con-
necting substance, or cement.

By the preponderance of certain of these elements over others or
by the peculiar forms which certain of these elements may present,
the different varieties of connective tissue are distinguished. Thus,
for example, in 'white fibrous tissue,' of which tendons and ligaments
are formed, the white fibrillae preponderate over the other elements,
so that on superficial examination of the fully developed structures
neither cells nor yellow elastic elements are seen, and the structures
might be likened to cords formed of dense bundles of white fibrillae
firmly agglutinated together. Again in the yellow elastic ligaments,
such as the ligamentum nuchae of large herbivores, or the ligamenta
subftava of the human vertebral column, there is such a preponder-
ance of the yellow elastic over the white fibres, that the former confer
upon the structures their peculiar physical properties.

In the adenoid or retiform connective tissue, by a peculiar modifi-
cation of the connective tissue cells, which give off branching pro-
cesses which join together, a network of fine fibres is established,
radiating at many points from connective tissue cells — a network
admirably adapted to afford support to other structures.

In the gelatinous connective tissue, the matrix or ground sub-
stance in which cells and fibres are imbedded is abundant and has a
'gelatinous consistence.

CHAP. VI I.I THE CONNECTIVE TISSUES. 251

Connective Tissue Cells.

These are the anatomical elements which are alone present in
the earliest stages of the development of connective tissue ; and it is
probably by the differentiation of their protoplasm that the inter-
cellular structures are ultimately formed which give to 'the different
varieties their peculiar characters.

We must refer the reader to works on Histology for a full
description of the various forms of connective tissue cells. We shall
in this place confine ourselves to the following categorical statements.

(1) Connective tissue cells consist invariably of a more or less
finely granular and contractile protoplasm, in which lies imbedded a
nucleus (sometimes more than one), usually of a vesicular nature.
The connective tissue cell, whilst it is in its state of typical activity,
is destitute of a cell wall, though occasionally one may be developed
(as in the fat cell) by the differentiation of the peripheral portion of
the cell-protoplasm. In certain cases branching processes are given
off from the protoplasm of the connective tissue cells and may serve
to connect adjoining cells together.

(2) In certain regions connective tissue cells are found (e.g. in
the cornea) imbedded in cavities in the ground substance; these
cavities sometimes communicate by minute canals, so that there is
established a canal system through which liquids may permeate
(Saftcanalchensystem). Such spaces or cavities are in certain
situations doubtless continuous with the smallest lymphatic vessels.

(3) In other situations the cells are discontinuous and resemble
rows of cells laid against the bundles of white fibres, one row to
each small bundle : being connected to them, and supported, by the
ground substance or matrix.

(4) In some situations pigment is deposited in the protoplasm of
the connective tissue cells (e.g. in the outer layer of the choroid) ;
in others, fat is formed at the expense of the protoplasm.

(5) Lastly, there occur in the connective tissues certain cells, which
are in all respects similar to the colourless cells of the blood, and
which wander through the connective tissue spaces, in virtue of the
amoeboid movements with which they are endowed. These are,
doubtless, either colourless cells of the blood which have passed
through the capillary walls, or they are the offspring of cells which
have thus emigrated.

Micro-Che- ^ur mf°rmation in reference to the chemistry of

micaireac- the connective tissue cell is, necessarily, of the most
tions of the limited character and is almost confined to a knowledge
connective that the protoplasm is proteid in nature and that the
nucleus shares the characters of nuclei elsewhere and
has probably the same composition.

It may be convenient however to summarize the effects of certain
reagents upon these cells.

252 REACTIONS OF CONNECTIVE TISSUE CELLS. [BOOK I.

(1) The connective tissue cells are unaffected by iodized serum
which constitutes, therefore, the best neutral liquid for their examina-
tion.

Iodized serum is a reagent of very great value to histologists1. It
is best made by dissolving iodine in the amniotic liquid of the cow ; this
fluid is placed in a thin layer in a bottle containing fragments of
iodine, with which it is frequently shaken. The iodine gradually dissolves,
conferring upon the solution a yellowish tint ; in the course of time iodates
are formed which increase the solvent action of the serum on iodine,
so that after one or two months a dark brown iodized serum is obtained ;
it is when of this colour that it is most serviceable (Ranvier2).

(2) Solution of perosmic acid (1 to 100) fixes the cells in the
form which they present during life, and permits of their being subse-
quently stained with picrocarminate of ammonia.

(3) Silver nitrate (from 0'25 to 0'5 per cent.) is of great
use in examining fresh connective tissue. Solutions of this salt
acting on the tissue fix the cells in the form which they possess
whilst alive, but without colouring them. It is however absorbed by
the ground substance and on subsequent exposure to light reduction
takes place, so that the unstained cells stand out on a stained back-
ground. The treatment with silver does not prevent the subsequent
action of certain colouring matters (ammoniacal carmine solution,
solution of picrocarmine).

(4) Solution of gold chloride (1 to 100) is of great use in
demonstrating the arrangement of the connective tissue cells of the
cornea. It is absorbed by the cells, which it helps to preserve in
their natural condition ; the absorbed salt is afterwards reduced and
confers upon the cell a reddish violet colour.

(5) Acetic acid causes the protoplasm to become very transparent,
whilst it brings out the nucleus very distinctly.

The White Fibres of Connective Tissue. — Collagen and Gelatin.

The most abundantly distributed forms of adult connective tissue
contain as their principal anatomical element bundles of white fibres,
which, as was previously stated, are rendered so transparent by the
action of acetic acid, as to be almost invisible. The fibres of which
the bundles are made up are connected together by an agglutinating
substance, which is soluble in dilute solutions of caustic baryta, or
lime. The substance of which the fibrils are composed has received
the name of Collagen, from the fact that when subjected to the
action of boiling water it is converted into gelatin or glue (KO\\O).

Prepara- Tendons, being composed almost entirely of the

tion of Coiia- white fibrils of connective tissue, are best employed in
gen. Roiiett's the preparation of collagen. They are cut into thin
process. slices and are then soaked in water until all matters

1 Max Schultze : " Die Anwendung mit lod conservirter thierisclier Fliissigkeiteii
als macerirendes und conservirendes Mittel bei histologisclien Untersuchungen."
Virchow's Archiv, Vol. xxx. (1864), p. 263.

8 Ranvier: Traite technique d'Histologie, Vol. i., p. 76.

CHAP. VII.] THE CONNECTIVE TISSUES. 253

soluble in water are removed. The watery extract contains a little
alkaline albuminate, but very little coagulable albumin. The frag-
ments of tendon are then soaked for some days in very weak solution
of baryta or lime, by the action of which the connecting substance is
dissolved so that the individual fibrils fall asunder. The insoluble
matter is then washed, first in water and afterwards in weak acetic
acid, finally again in water. The residue consists almost entirely of
the substance of the white fibrils (collagen), mixed however with
small quantities of yellow elastic tissue and cell nuclei.

When placed in very diluted acids and alkalies, the fibrils swell
up and become transparent, their original appearance being restored
if the acid is exactly neutralized. When digested in very dilute
acetic acid at ordinary temperatures for some days, the fibres
gradually dissolve, yielding a solution which contains gelatin, and
also a little acid-albumin, produced by the action of the acid upon the
residual matter of connective tissue cells.

When the white fibrils are subjected to long-con-
Gelatin. ,. , , .,. . ,, ,. J &,, ,,

turned boiling in water at the ordinary pressure of the

atmosphere, or, still better, to the action of water heated under pres-
sure (as in Papin's digester), they dissolve, and the solution is found
to contain a substance termed Gelatin. The same body is produced at
the temperature of the animal body by the prolonged action of very
diluted acids on collagen.

Prepara- It is most convenient to employ commercial gelatin

Geiatto PUre for the PreParation of tne Pure substance. The finest
commercial gelatin is allowed to soak for some days in
large quantities of distilled water, which is frequently changed; in
this way the soluble salts of the gelatin diffuse out. The swollen
gelatin is now dissolved in distilled water by the heat of a water
bath; after allowing insoluble matters to subside the solution is filtered,
with the aid of a hot-water funnel, directly into 90° per cent, alcohol.
The gelatin separates in the form of white, thready masses, which are
collected, reduced to a fine state of division by cutting, and allowed
to dry, first of all in the air, and then in a water oven. Gelatin thus
prepared contains about 0'6 per cent, of ash1.

Gelatin is a body which is insoluble in cold water ; the sole
action of cold water is to cause it to swell up. On adding boiling
water to the swollen solid, it dissolves with readiness, and a clear
limpid solution is obtained, which when it is cooled sets as a more or
less firm jelly — gelatinizes. This property is possessed by solutions
which contain only 1 per cent, of the solid substance.

The power of gelatinizing is gradually lost when solutions of
gelatin are subjected to prolonged heating, and instantly lost when
they are heated to 140° in sealed tubes.

1 Hofmeister: "Ueber die chemische Structur des Collagens." Zeitschr. f. phys.
Chem. Vol. n. (1878), p. 315.

254 COLLAGEN A^TD GELATTX. [BOOK I.

Gelatin is insoluble in alcohol, ether and chloroform. It is dis-
solved with the aid of heat in glycerin, and on cooling a jelly (glycerin
jelly) is obtained.

Aqueous solutions of gelatin are powerfully laevo-rotatory, the
rotatory power being very much influenced by temperature and by
the reaction of the solution (Hoppe-Sevler). In aqueous solutions
at 30° (a) j = -130°.

Gelatin is not precipitated from its solutions by acetic acid and
ferrocyanide of potassium — a character which distinguishes it from
any proteid substance. It is not precipitated by acetic acid — a
character which distinguishes it from the closely allied body Chondrin.

Tannic acid precipitates gelatin even when its solutions are very
dilute. Solutions of mercuric chloride also precipitate it.

On the other hand gelatin is not precipitated by solutions of lead
acetate (which precipitates chondrin) nor by the majority of metallic
salts which do precipitate the proteids.

Composi- ^ke u^imate analyses made of collagen and of

tion of Geia- gelatin drew the attention of observers to the fact that
tin, and its the composition of these two bodies is very similar if
relation to no£ identical. They contain carbon, hydrogen, oxygen,
Collagen. nitrogen, and, according to some authors, sulphur.

The following analyses indicate the composition of these bodies.

Substance of Gelatin from
tendons. tendons.

Carbon 50"9 50'2

Hydrogen 7 '2 67

Nitrogen 18-3 17'9

O and S 23-5 25'0

According to Schiitzenberger and Bourgeois, sulphur is not an
essential constituent ; these authors ascribe to gelatin the formula

%HJ?«Q*;

The relations of gelatin to collagen have been made the subject

of a very interesting study by Hofmeister. This author has found
that by heating gelatin for Some time at 130°, it loses about
0755 per cent, of water and becomes converted into a body in all
respects identical with collagen. Collagen is, therefore, probably an
anhydride of gelatin.

The following are the mean results of the analyses of collagen
(Hofmeister).

Carbon 50-75

Hydrogen 6 -47

Nitrogen 17 '86

Oxygen 24 -92

To gelatin Hofmeister ascribes the formula C]02Hj51N31039, and
collagen is probably related to it as shewn in the following equation:
^N-O^ - H20 - C^AA,

Gelatin Collagen

.

CHAP. VII.] THE CONNECTIVE TISSUES. 255

Products of a. Action of boiling water. When solutions of

WonofPgeia- gelatin are subjected to the action of boiling water for
tin. a longer period than 25 hours they lose the property of

gelatinizing, and are found to contain a mixture of two peptone-like
bodies, to which Hofmeister has given the name of Semiglutin and
Hemicollin.

Semiglutin is very little soluble, whereas hemicollin is soluble, in
70 — 80 per cent, alcohol.

The former substance is precipitated by platinum tetrachloride,
the latter is not. Both bodies furnish glycine and leucine when
treated with boiling hydrochloric acid and stannous chloride. To semi-
glutin Hofmeister ascribes the formula C55H8gN17O22 and to hemicollin
C47H70N14019. In the process of decomposition into these bodies collagen
takes up water, increasing 2'22 per cent, in weight. The following
equation exhibits the relationship of these bodies, according to
Hofmeister,

C,JW>S + 3H20 = CJIssN.A, + C^HJS.A,

Collagen Semiglutin Hemicollin

b. Action of boiling sulphuric acid. When subjected to the
action of boiling sulphuric acid, collagen and gelatin yield, amongst
other products, ammonia, leucine, glycine, and, perhaps, aspartic
acid.

c. Action of caustic baryta in heated sealed tubes. When
heated with solution of caustic baryta in sealed tubes, ammonia,
carbon dioxide, acetic and oxalic acids, and a mixture of amido-acids
(containing glycine and alanine, amido-butyric acid, traces of glutamine,
&c.) are obtained.

d. Action of pancreatic ferments. When subjected to the com-
bined action of the pancreatic ferment and putrefaction gelatin yields
gelatin-peptones, leucine, glycine, volatile fatty acids, ammonia and
carbon dioxide. Amongst the fatty acids are acetic, butyric and
valerianic acids (Nencki1).

The Elastic Fibres of Connective Tissue. — Elastiri.

When ordinary connective tissue is treated with acetic acid, the
white fibres swell up and become transparent, whilst the elastic fibres
remain unaltered and are therefore very distinctly seen. When the
same connective tissue is subjected to prolonged boiling in water, the
collagen of the white fibres undergoes solution and there is left a
network of elastic fibres. These fibres are composed of an elastic
substance which offers remarkable resistance to the action of
chemical reagents, and to which the term Elastin has been applied.

1 Nencki : Ueber die Zersetzung der Gelatine und des Eiweisses bei der Fdulniss mit
Pancreas. Barn, 1876. Abstracted in Maly's Jahresbericht, Vol. vi., p. 31.

25G ELASTIN. — GROUND SUBSTANCE. [BOOK I.

Prepara- The lig amentum nuchae of the ox, horse, or still

tionofEias- better of the giraffe, is cut into thin slices, which are
tin- boiled in ether and in alcohol, and then for at least 36

hours in water. The two first of these reagents free the tissue from
fatty matters, whilst the prolonged action of boiling water converts
all the collagen into gelatin which passes into solution. The
insoluble matter is boiled in strong acetic acid for a long time :
then after removal of the acid by water it is boiled in concen-
trated solution of caustic soda until the fibres begin to swell.
The tissue is then successively heated with dilute acetic acid and
with water, and lastly placed for 24 hours in moderately concentrated
hydrochloric acid. The substance remaining is washed with water
until all acid is removed. It is then found to retain all the original
characters of fresh elastic tissue. 9

Composi- Miiller1 analysed elastin which he had prepared by

tion of Elas- following the previously described process and found it
tin- to have the following composition.

(1) (2) J3) (4)

Carbon 55*47 55*72 55 '55 55*09

Hydrogen ., 7"54 7*67 711 7'33

Nitrogen 16*09 1571 16*52 16*43
Oxygen 20'90 20*70 20'82. 21*15

Solubility. So far as is known elastin is not soluble in any liquid
which does not decompose it. It is soluble in boiling solution of
caustic potash, in cold concentrated suphuric acid, and in concentrated
nitric acid.

It is gradually dissolved when digested with pepsin and with
trypsin, the former ferment being more active than the latter 2.

Products of When- boiled with sulphuric acid elastin is said to

decomposition, yield leucine but no tyrosine.

Connecting or Ground substance of Connective Tissue.

Absorption When perfectly fresh connective tissue is immersed

of silver salts for a few minutes in a solution of silver nitrate

nectS^sub- (°'25 to 1<0 P*c*)' then freed from excess of silver by
stance and washing with distilled water, and afterwards exposed to

subsequent light, a deposition of metallic silver occurs which
reduction. appears to be deposited in the connecting substance

which is interposed between the various tissue elements. The proto-
plasmic elements of the tissue are left perfectly unstained, so that the
silver treatment furnishes the histologist with one of the best methods
of studying their arrangement. It has been held by many that
the absorption of silver, which is afterwards reduced, is a characteristic

1 Zeitschrift f. rat. Med. Third Series. Vol. x., part 2.

2 A. Ewald und W. Kuhne, Die Verdanung als histologische MetJiode.

CHAP. VII. ] THE CONNECTIVE TISSUES. 257

property of the substance, but this view is probably incorrect, the
absorption of the silver salt being merely due to the physical condi-
tions of the connecting substance rather than to any peculiar chemi-
cal property which it possesses1.

Solubility When any of the forms of connective tissue proper

if cttn Csub are macerate(l f°r some days in baryta- or lime-water,
s^ancefnsoiu- tne vari°us tissue elements fall asunder, in consequence
tions of tiie of the solution of the connecting substance. If
Alkalies. ^ the alkaline solution thus obtained be treated with
a dilute acid, a precipitate insoluble in excess of the reagent is
obtained, which, after careful washing with water, is found to be
insoluble in alcohol and ether, and when burned on platinum leaves
no perceptible amount of ash. This body is now looked upon
as identical with a^substance which is pretty widely distributed,
and which will be conveniently described in this place, viz. Mucin.

Mucin.

Distribu- This body, besides forming apparently a small

tion- proportion of all connective tissue proper, is present in

specially large quantities in embryonic connective tissue, and in that
form of the tissue occasionally met with in the adult animal and
which is termed Gelatinous connective tissue.

It is found in the epidermis, where it connects together the
epithelial cells. It is found in considerable proportions in synovia.

It is a frequent product of the activity of certain epithelial cells
and is the chief constituent in the viscid tenacious liquid called
Mucus, which often covers epitheliated surfaces.

Mucus is a colourless, viscid, semi-liquid substance varying very
greatly in consistence. It is sometimes transparent, but often turbid
from the presence of epithelial cells or foreign matters. It contains,
besides mucin, which is its chief organic constituent, small quantities
of proteid substances, and salts, amongst which common salt pre-
ponderates.

Mucin constitutes the chief ingredient of the tissues of certain in-
vertebrates, and indeed much of our knowledge of mucin is derived
from Eichwald's investigations of this body obtained from Helix
pomatia. It is not however certain that mucin from this source
is identical with that of the mucous membranes and tissues of man.

Modes of (&) From connective tissue (Rollett's method2). Ten-

preparation dons are finely divided and treated with large quantities
of distilled water, with the object of removing albumi-
nous and saline matters soluble in that liquid. They are then digested
for many days in large quantities of lime- (or baryta-) water. The
solution is precipitated by acetic acid, -which throws down a pre-

1 Consult Eobinsky, "Die Kittsubstanz auf Eeaction des Argentum nitricurn."
Archivf. Anat. u. Physiol., 1871, p. 184.

2 Eollett, Sitzuntjsberichte der Wiener Akadernie, Bel. 39, p. 308 : Strieker's Hand-
book, Vol. i. p. 72.

G. 17

258 MUCIN. [BOOK i.

cipitate which at first appears granular, but afterwards flocculent.
The precipitated body may be collected on a filter, and washed with
water or dilute alcohol.

(b) From bile. As bile often contains very large quantities of
mucin, it may serve as raw material for its preparation. The
bile is treated with its own volume of 80 p.c. alcohol, which throws
down a precipitate composed of mucin mixed with epithelium,
proteids, &c. The precipitate is separated by decantation and washed
with fresh alcohol, it is then suspended in a large quantity of
lime-water; after some days the solution is decanted and precipi-
tated with acetic acid, the precipitate is washed successively with
water, alcohol and ether1. This method may with slight modifica-
tions be employed to separate mucin from sputum, or other liquids
containing the body. In the case of sputum it would be well
to follow Gautier's advice, to commence by washing with water
acidulated with acetic acid.

Properties Mucin when freshly precipitated is a glutinous

of mucin. substance, which forms with water an opaque liquid

in which it is held in suspension without being dissolved. It is
soluble in weak solutions of the alkalies and alkaline earths,
from which it is precipitated by dilute acids, acetic acid being usually
employed for this purpose.

It is insoluble in dilute hydrochloric acid, containing from 01
to 1 p.c. of real acid ; but it is soluble in hydrochloric acid of 5 p.c.

Mucin which has been precipitated by acids is insoluble in
solutions of common salt.

Mucin is not digested by artificial gastric juice ; it is dissolved by
alkaline solutions of trypsin.

Mucin is precipitated by acetate of lead from neutral or weakly
alkaline solutions, but by no other metallic salts. It is not pre-
cipitated by acetic acid and potassium ferrocyanide : it is also
unaffected by tannic acid.

When treated with copper sulphate and caustic potash it prevents the
precipitation of cupric hydrate ; the solution is not reduced on boiling.

When boiled with Millon's reagent, mucin gives a rose colouration.
Elementary Mucin contains the elements C, H, N, 0, but no
composition sulphur. The following are the results of elementary
analyses by various investigators.

I. (Scherer). H. (Obolensky). III. (Eichwald).

Mucin from Mucin from Mucin from

mucous con- submaxillary Helix pomatia. .

tents of a cyst. gland.

Carbon 5217 52'31 48'94

Hydrogen 7'01 7'22 6'81

Nitrogen 1264 11'84 8'50

Oxygen 2818 2863 35'38

1 Gautier, Chimie appliquee a la M6decine, Vol. n. p. 126.

CHAP. VII.] THE CONNECTIVE TISSUES. 259

It is impossible to study these analyses without concluding that,
though agreeing in general chemical reactions, the mucin-like con-
stituent of the tissues of invertebrates is a different substance
from the rnucin obtained from mucous membranes.

Products of When boiled for twenty or twenty-five minutes

decomposi- with dilute sulphuric acid, mucin is decomposed with
tion of mucm. ^Q formation of acid albumin and a body possessing
the property of reducing salts of copper and bismuth similar to
those of glucose. That this body is not a sugar is proved by the facts
that it does not rotate polarized light, and that it is incapable of
alcoholic fermentation; it appears to be a nitrogenous body1.

When boiled with strong sulphuric acid for seven hours mucin
yields leucine and tyrosine (Obolensky).

When boiled with caustic soda, on neutralizing and shaking
with ether, the latter fluid dissolves a body which possesses the
reaction of pyrocatechin (06H602), i.e. is coloured of an emerald
green colour by solution of ferric chloride (Obolensky).

The products obtained when mucin is subjected to the action of
pancreatic ferment, prolonged until putrefaction sets in, have been
studied under Nencki's direction by Walchli2; amongst them were
found ammonia, indol, a large quantity of butyric acid, and a substance
possessed of a sweet taste and reducing copper salts.

Relations Mucin is unquestionably a product of the differentia-

of mucin. tion of the protoplasm of certain animal cells, and is

obviously derived from the proteids. It is conceivable that it may
result from a decomposition in which both collagen and mucin origin-
ate; what the nature of the decomposition may be is, however, quite
unknown.

SECT. 2. ADIPOSE TISSUE.

structural ^a* occurs in the animal body either in a state

Elements of of solution or minute suspension in its juices, or
Adipose deposited within the interior of cells. This is espe-

cially the case in cells which are developed in, and
supported by, the connective-tissue of certain regions ; these cells,
which originally are identical with connective-tissue cells, undergo
changes which ultimately result in the diminution of the cell proto-
plasm (at the expense of which, or through the agency of which*
oily matter is deposited within the cell) and in the development of a
well-marked cell- wall which serves to contain their oily contents. These
oily contents undergo remarkable fluctuations according to the state
of nutrition of the animal.

Fat cells are usually found in groups or clusters, supported by
the fibrous elements of connective tissue, and surrounded by a network

1 Obolensky, "Ueber Mucin aus der Submaxillardriise. " Pfliiger's Archiv, Vol. iv.
p. 336.

2 Walchli, Ber. d. deutschen chem. Gesellsch. xi. 1878, p. 509.

17—2

260 ADIPOSE TISSUE. [BOOK I.

of capillaries. They develope with special frequency in areolar tissue,
especially in the subcutaneous areolar tissue and in the connective
tissue which lies around and bet ween certain of the abdominal viscera.
In certain situations (as e.g. in the orbit) the areolar tissue is never
free from fat ; in other situations, as in the subcutaneous connective
tissue of the eyelids, of the penis, and scrotum, fat cells are never
present.

The fully formed fat cell presents 'the appearance of a little bag
distended with glistening oily contents, and with no appearance of a
nucleus or of typical protoplasm. In reality, however, it can be
shewn that even the fully formed fat cell contains a nucleus
with remains of the original cell protoplasm around it, though these
are so pressed upon and surrounded as to be invisible until some
cause comes into operation to remove a part at least of the oily
contents.

The contents of the fat cells are during life of flu id consistence and
only solidify when the tissue containing them is cooled below 25° C.
When solidifying, the oily matter often separates, at least in part,
in the form of groups of needles ; sometimes in small single needles.

When adipose tissue is treated with ether, this fluid extracts and
dissolves more or less completely the fatty contents of the cells, in
which the remains of the nucleus and protoplasm may be then
detected.

The wall of the fat cell is not acted upon by acetic acid nor by
dilute mineral acids; it is easily dissolved by natural or artificial
gastric juice. When treated with a solution of perosmic acid, the
fatty matter contained in fat cells is stained of an intense black
colour ; this reagent is admirably adapted for the microchemical detec-
tion of fatty bodies.

In this section a description will be given of the principal constituents
of the adipose tissue of man and the higher animals, the discussion of the
origin of fat in the economy being postponed to that division of this work
in which certain general chemical processes of the body are treated of
under the heading of Nutrition. A consideration of the fatty matters which
occur in particular organs or fluids (as for instance in the nervous organs and
in milk) is given in the chapters devoted to these subjects.

Mode of ex- The fatty matters which are contained in adipose

tracting the tissue are best extracted by drying the tissue so as to
fats of adipose expel the water which it contains, and then boiling the
finely divided or comminuted tissue in ether, which
dissolves all the fats. The etheral solution is then evaporated to
dryness. The residue contains the fats, mixed with small quantities
of other bodies soluble in ether, such as cholesterin and lecithin.

In some cases the fatty matters of adipose tissue can be separated
in large quantities by the combined action of heat (which causes the
fats to melt) and pressure ; or by boiling the tissue with water, when
the melted oil floats to the surface and can be skimmed off. These

CHAP. VII.] THE CONNECTIVE TISSUES. 261

two methods are employed in the commercial separation of fats from
certain animal products.

Chemical ^ke ^s wn^cn are contained within the fat cells of

constitution man and the higher animals are mixtures of the so-
of the neutral called neutral fats termed stearin, palmitin, and olein,
of which the two former are solid bodies at ordinary
temperatures, and are, at the temperature of the body, held in solution
by the third.

The neutral fats are the most abundant of the non-nitrogenous
organic proximate principles of the body and contain the elements
carbon, hydrogen and oxygen. These fats consist of ethers derived
from the triatomic alcohol glycerin C3H6(OH)3.

We may form a true conception of the relations of a neutral fat
to glycerin by examining the relations of an artificial neutral
fat, or glycerin ether, triacetin, to glycerin, and these wilLbe easily
illustrated by the aid of the two graphic formulae here appended. The
three carbon atoms of glycerin are seen to be linked to the 0 atoms of
three OH groups ; the H in any one or in all of these may be re-
placed by the oxidized radical of a fatty acid, e.g. by acetyl C2H3O ;
when all three of the hydrogens are thus replaced the neutral fat
called triacetin is formed, thus : —

H H H

H H H |

| | | H-C C C-H

H— C— C— C— H |

III 000

O O O I

I i i o-c o=c c=o

H H H ill

H— C~H H— C~H H-C-H

Glycerin. H H H

Triacetyl-glycerin ether, or triacetin.

The neutral fats of adipose tissue are constituted on the same
type as triacetin, except that, instead of acetyl, other acid radicals
take the place of the H in the OH groups.

In the two more solid fats, stearin and palmitin, the oxidized
radicals of stearic and palmitic acids occupy the position of the acetyl of
triacetin; in the more liquid constituent of the fats, viz. olein, the
oxygenized radicals of oleic acid occupy the same position.

Sometimes, instead of the terms stearin, palmitin, and olein, the
more precise designations of tristearin, tripalmitin and triolein are
employed.

The formulae of the three principal fats are appended and their
relationship shewn to glycerin and the acids.

Palmitin C3H5(OC16H310)3 Palmitic acid C16H310,OH

Glycerin C3H5(OH)3 Stearin C8H5(OC18Ha50)8 Stearic acid dgHwOjOH

Olein C3H5(OC18H330)3 Oleic acid C18H330,OH

General The neutral fats are all solid at a certain tempera-

properties of ture, above which they are fluid ; this temperature is
the neutral called their melting-point. They are all soluble in
boiling alcohol, in ether, benzol, carbon disulphide

262 PROPERTIES OF NEUTRAL FATS. [BOOK I.

and chloroform. When fluid they render paper which is coated
with them transparent (grease spots). When mixed with colloid
substances and water, they admit of being broken up into fine drops,
so that the fluid becomes white and opaque (an emulsion). Under
the influence of certain ferments (e.g. one of the ferments contained
'in the pancreatic secretion) they combine with the elements of water,
splitting up into glycerin and a fatty acid; thus : —

C3H5 (OCMH3,0)S + 3H20 = C3H5 (OH), + 3 (C H310, OH).

Tripalrnitin. Water. Glycerin. Palmitic acid.

The rancid smell of decomposing fats is due to volatile acids
which are set free.

When the neutral fats are boiled with solutions of the alkaline
hydrates or carbonates they undergo the process of saponification, i.e.
they combine with the elements of water, and decompose into
glycerin and fatty acids, the latter constituents combining at once
with the alkaline metal to form a soluble salt, termed a soap. Thus
when stearin, palmitin or olein is boiled with potash hydrate or with
sodium hydrate, the results of the operation are, in the first case,
potassium stearate, palmitate, or oleate, respectively and glycerin ; in
the second case sodium stearate, palmitate, or oleate, and glycerin.

When boiled with litharge the neutral fats yield an insoluble lead
soap and glycerin.

The term soap is applied to the metallic salts of the fatty acids,
and hence the term saponification is employed to designate the
process which results in the formation of these compounds.

Stearin or Tristearin. C^^O.C^H^O)^

Stearin is the chief constituent of the more solid fats. Its
melting-point is higher than that of any other neutral fat, but varies,
according to the treatment to which it has been subjected, between
53° and 66°. It is nearly insoluble in cold alcohol and ether, though
soluble in both these fluids when these are boiled. The insolubility
in cold ether is taken advantage of in the preparation of pure stearin.

Stearin may be obtained from the suet of the sheep by extracting
it repeatedly with cold ether, and dissolving the residue in boiling
ether, which, on cooling, deposits crystals of stearin, in the form of
little leaflets which shine like mother-of-pearl. From a boiling
alcoholic solution stearin is deposited in brilliant scales, which are
almost square rhombic crystals having angles of 90°'5.

Palmitin or Tripalmitin. C3H6 (0 . C16H310)8.

Is the more abundant of the two solid neutral fats in the adipose
tissue of man. It is more soluble in cold and in hot alcohol and ether
than stearin ; it is deposited from saturated solutions in the form of
fine needles, which radiate from a centre and appear as delicate
filaments.

CHAP. VII.] THE CONNECTIVE TISSUES. 263

Its melting-point varies, like that of stearin, according to its treat^
ment ; the temperature at which it solidifies, after being melted, is
said to be 45° C.

Margarin a From a mixture of stearin and palmitin, crystals

mixture of often separate which consist of a mixture or perhaps of
stearin and a combination of stearin and palmitin, but which were
formerly supposed to be a special fat to which the
name of Margarin was given and which was supposed to be a glycerin
ether of margaric acid (CJ7H3402).

The crystals which form not unfrequently in fat cells were
formerly supposed to consist of this compound.

FIG. 51. CRYSTALS OF THE SO-CALLED MARGARIN.

a. single needles. &. larger groups, c. crystals within fat cells, d. a fat cell con-
taining no crystals. (Funke.)

Olein or Triolein. CSH5(0 . C18H330)3.

This neutral fat is obtained in a state of comparative purity from
the more liquid fats by exposing them to a temperature of 0° C. and
then subjecting to pressure ; the liquid portion expressed consists of
olein.

When pure, olein is a colourless oil which is fluid at ordinary tem-
peratures and which solidifies when the temperature falls below 0° C.
When exposed to air olein absorbs oxygen, and in doing so it acquires
a faint yellow colour.

It dissolves all the solid fats, especially at a temperature of 30° 0.
Olein is easily soluble in cold absolute alcohol or in ether.

Glycerin. C3H6(OH)3.

Mode of As has been already stated, when the neutral fats

froTSe1011 are saponified, glycerin is set free. If the neutral
neutral fats. fats be boiled with litharge and water, the fatty acids are
all thrown down as insoluble lead soaps, and glycerin
dissolves in the water. By passing a stream of sulphuretted hydrogen,
it is freed from dissolved lead, and on filtering and evaporating the

264 PROPERTIES OF NEUTRAL FATS. [BOOK I.

solution a syrupy liquid — glycerin — is left. This instructive method
of preparing glycerin is not at present employed in the arts, the
substance being now obtained by decomposing and distilling the
neutral fats by means of superheated steam.

Properties Glycerin is a colourless, syrupy liquid of intensely

of Glycerin. sweet taste, having a specific gravity of T27, and
soluble, in all proportions, in water. It becomes solid at — 40° C. It
boils at 280°. When heated with the fatty acids it combines with
them, forming ethers which are constituted as the fats. Thus
by the action of acetic acid on glycerin at 100°C. a body termed
monacetin is obtained ; by the action of acetic acid at a higher tem-
perature diacetin is obtained, and again by reacting further with
acetic acid on the latter body, triacetin is formed. The H of the
three hydroxyls of glycerin is in this case successively replaced.

(OH (OH (OH (OCTLO

C3Hfi toH C8H6 JOH C8H6 toC2H30 C3H6 -toC2H80

(OH (OC2H30 (OC2H30 (OC2H30

Glycerin. Monacetin. Diacetin. Triacetin.

When glycerin is subjected to the prolonged action of yeast,

O TT Ol
it yields propionic acid 3 jj| 0.

When distilled with hydriodic acid glycerin yields isopropyl
iodide : —

CfLfl, + 5HI = CVHJ + SHp +2 I2.

Glycerin. Hydriodic Isopropyl Water,
acid. iodide.

When heated with phosphorus pentoxide, or acid potassium
sulphate, or subjected to destructive distillation, glycerin yields
acrolein (C8H40), which is the aldehyde of allyl-alcohol (C3H5OH) ; this
substance boils at 52'4°. Its vapour possesses an intensely irritating
and characteristic odour.

Fatty matters found in the adipose tissue of certain of the lower animals.

Spermaceti. In addition to the three neutral fats which have been men-
tioned other fats occur in certain members of the animal kingdom.
In spermaceti, which is a fatty substance contained in the cranial sinuses of
whales, there are no glycerides, but the fats appear to be derivatives of cetyl-

alcohol l6jg-34 0, a solid body melting at 50°, the chief compound being

cetyl-palmitate ; when saponified, spermaceti yields, in addition, stearic,
myristic and lauric acid. It is worthy of notice that cetyl-alcohol can
be artificially oxidized so as to yield palmitic acid.

Bees' wax In Chinese wax which is produced by the Coccus ceriferus,

and in bees' wax, the product of the common bee, the portion of

the substance which is soluble in boiling alcohol contains ceryl-cerotate

CHAP. VII.]

THE CONNECTIVE TISSUES.

265

OH )

naTTi" Af 0> which when saponified by boiling with caustic potash yields

^27^53 "J

OH")
ceryl-alcohol 27jr Y O, which is one of the series of primary alcohols, and

cerotic acid, C27H540, which is the normal fatty acid corresponding
to the above alcohol. In addition to ceryl-cerotate, free cerotic acid is
contained in bees' wax.

In the portion of bees' wax which is insoluble in alcohol there is
contained myricyl palmitate, an ether derived from myricyl alcohol

H

Analysis of the Fats.

A weighed quantity of the finely divided tissue in which
and^determi1- ^e ^a*s are *° ^e separated and determined is evaporated
nation of the to dryness in a water oven. The dry residue is then boiled
with ether for a long time. The process may be carried
on in a flask connected with an inverted condenser, the
flask being heated on the water-bath. The apparatus shewn
in Fig. 52, which was devised by Dr Drechsel2, is perhaps superior to
any other for the extraction of fats from animal matters. At A is a
flask containing ether, into which is fitted a tightly-fitting cork or stopper,

Extraction

total amount
of fat in a
tissue.

Fio. 52. PRECHSEL'S APPARATUS FOR THE EXTRACTION OP FATS.

perforated so as to allow the lower end of the bulb B to fit into it.
B is closed by a stopper. Into B can be passed a plaited filter such as
is shewn in the cut, and into this filter the solid is placed from which

1 Consult Schorlemm er's Organic Chemistry, p. 174.

2 Drechsel, Journ. f. prakt. 'Chemie, Vol. xv. (1877), p. 350 and Plate n.

26G METHOD OF SEPARATING FATTY ACIDS. [BOOK I.

the fats are to be extracted. PCbc is a glass connecter wlrich communi-
cates with an inverted Liebig's condenser, with a stopper which fits into
the upper part of £, and with the tube a which is joined to the side of A.
When the flask A containing ether is placed upon the water-bath, that
liquid boils and the vapour passes through a and b to bPC ; it ascends
into the Liebig's condenser where it is condensed, and it then flows
back into B over the matter placed upon the plaited filter, and thence
into A. A continual circulation of ether is thus kept up, and the
dissolved fats accumulate in A.

The ethereal solution is then evaporated to dryness in a weighed capsule,
and the weight ascertained.

Separation ^e n^ture of neutral fats is dissolved in boiling alcohol

of the fatty and the solution is poured into a silver basin, and then

acids contain- treated with an alcoholic solution of caustic potash. The

edintheneu- flu^ js boiled for some time over a water-bath and then

evaporated to dryness. Water is then added to the
residue so as to dissolve the soaps which have been formed, the solution
is now acidified by the addition of hydrochloric acid, then boiled
and allowed to cool. The fatty acids, which have been liberated from
the neutral fats, set in the form of an insoluble mass, which is collected
on a filter. This is then dissolved in hot alcohol and treated with an
alcoholic solution of lead acetate which precipitates all the fatty acids
in the form of insoluble lead salts. The precipitate is collected on a filter
and dried, and subjected to the action of boiling ether, which dissolves
only lead oleate, leaving lead stearate and palmitate. The ethereal
solution of lead oleate may be agitated with dilute hydrochloric acid,
which will decompose the salt, and the ether will then hold oleic acid
in solution which will remain on evaporation ; the treatment with HC1
should be carried out in an atmosphere of CO2. The mixture of lead
stearate and palmitate is heated with hydrochloric acid and thereafter
shaken with ether; the ethereal liquid is freed from acid by shaking
with water and the ether is then distilled off, when a mixture of palmitic
and stearic acids is obtained.

The melting-point of the mixture is then taken. In order to do
this a very thin glass tube is made and a small quantity of the
mixed acids is dropped in ; the tube is then drawn out slightly. The
glass tube is now attached by means of a little india-rubber band (which
may be made by cutting a thin circular slice from the end of a narrow
india-rubber tube) to a finely graduated thermometer, in such a manner
that the part of the tube in which the fat lies is on the same level as the
bulb of the thermometer. The latter is then plunged into a beaker
containing water, which is immersed into a larger beaker also containing
water. (See the arrangement employed for determining the temperature
at which solutions of the proteids coagulate at p. 15). The latter is then
heated so that the temperature of the water in the inner beaker rises
very gradually. The observer watches very carefully the temperature
at which the fat nielts; he then withdraws the heat from the outer
beaker and, as the temperature of the water surrounding the thermo-
meter bulb falls, he notices the temperature of solidification of the
previously melted fats.

CHAP. VII.]

THE CONNECTIVE TISSUES.

267

With the aid of the appended table based upon the observations
of Heintz * an approximate estimation of the composition of a mixture
of stearic and palmitic acids can be made.

A mixture of

Stearic acid Palmitic acid Melts at

Solidifies at

90 parts 10 parts 6 7° '2

62°-5

80 „ 20 , 65°-3

60°'3

70 30

62°-9

59°-3

60 40

60°-3

56°-5

50 50

56°-6

55°-0

40 60

56°-3

54°-5

30 70

55°-l

54°-0

20 80

57°-5

53°-8

10 90

60°-1

54°-,5

In order to demonstrate the separate presence of stearic and palmitic
acid it is, however, essential to proceed further. The mixture of the
acids is dissolved in boiling alcohol and treated with sodium carbonate,
then evaporated to dryness on the water- bath ; the residue is further
heated in the air-bath to 130°. The residue is pulverized and boiled
with absolute alcohol ; the solution is filtered hot.

Fractional This solution is now subjected to fractional precipita-

precipitation tion, by adding either solution of chloride or acetate of
of a mixture barium. "When one of these salts is added, little by little,
of fatty acids. ^o a solution containing both stearic and palmitic acids, the
precipitate which first falls is composed entirely of barium stearate ;
the further gradual addition of the barium salt leads to the precipi-
tation of a mixture of barium stearate and palmitate, and if the addition
of barium salt be continued after this pure barium palmitate falls.
Relying upon these facts the experimenter adds to the alcoholic solu-
tion of the mixed fatty acids one or two drops of a solution of barium
chloride or acetate, filters, heats the filtrate to boiling, then adds to it
one or two drops more of the barium solution, collects the new precipitate
on a separate filter, heats the filtrate, and repeats these operations until
the addition of barium salt occasions no new precipitate. Each precipitate
is collected on a separate small filter, is washed with warm alcohol
and dried at 120°C. The barium in each precipitate is then determined
by igniting in a porcelain capsule, and adding first hydrochloric and
then sulphuric acid to the ash, again igniting, and weighing the barium
sulphate. If stearic acid is present the first precipitates should contain
the amount of barium corresponding to barium stearate; if palmitic
acid, the last precipitates should agree in composition with barium pal-
mitate.

Comftosition of the barium salts of stearic and palmitic acids.

100 parts of barium stearate contain 19-49 parts of barium.
„ „ „ palmitate „ 21 '17 „ „

1 Poggendorff's Annalen, Vol. xcn. p. 588.

2G8 CARTILAGE. [BOOK I.

SECT. 3. CARTILAGE.

structural Cartilage is a tissue composed of certain cells,

termed cartilage cells, imbedded in a ground substance
or matrix. According to the predominating character
of this matrix the cartilage may be classified as (1) cellular cartilage,
(2) hyaline cartilage, (3) white fibre-cartilage, (4) elastic or spongy
cartilage. In the first of these varieties the matrix consists merely
of a transparent and very thin envelope termed the capsule, surround-
ing each cartilage cell ; these capsules possess the same chemical
properties as the matrix of hyaline cartilage ; in the second variety
the matrix is composed of a translucent homogeneous substance,
which occasionally presents an appearance resembling that of
ground glass, and sometimes exhibits fibrillation ; in the third,
the cartilage cells are surrounded by capsules which lie imbedded
in a preponderating mass of fibres identical with the white fibres of
connective tissue ; in the fourth the cartilage cells with their capsules
are imbedded in a meshwork of elastic fibres.

Cartilage The cartilage cell is a mass of protoplasm with one

cells. or two nuclei, contained in a cavity which it com-

pletely fills and which is bounded by the so-called capsule of the
cartilage cell, this being intercellular substance which is produced
by the differentiation of the cell protoplasm.

It is the capability of producing this intercellular substance which
is the very characteristic of the cartilage cell. In cellular cartilage we
find an aggregation of cartilage cells, each of which is surrounded by
its own capsule : in hyaline cartilage the homogeneous matrix has
been produced by the fusion of concentric, and successively developed,
cartilage capsules, as can be shewn by subjecting the fully formed
and homogeneous tissue to the action of certain reagents, such as a
mixture of hydrochloric acid and potassium chlorate, when the
appearance of concentric stratification of the matrix, around the
cartilage cells, is revealed.

In young cartilage cells the protoplasm often contains glycogen
(C6H10O5) ; in the cells of the fully developed tissue fat is often
seen.

General Composition of Cartilage.

Cartilage contains more than half its weight of water, though the
proportion varies remarkably. Its solid constituents consist mainly
of organic matter with a small proportion of salts, in which sul-
phates and phosphates preponderate.

' The following analyses exhibit the relative proportions of water,
organic matters and mineral matters in the cartilage of a young and
healthy man (Hoppe-Seyler1) :—

1 Hoppe-Seyler, quoted by Kiihne, Lehrbuch, p. 387.

CHAP. VII.] THE CONNECTIVE TISSUES. 209

Costal Articular Cartilages

Cartilages. from knee joint.

Water in 100 parts 67'67 73'59

Organic matters 3013 24'87

Mineral „ 2*20 1'54

Tlie following are the results of Hoppe's analysis of the ashes of
human costal cartilage.

Potassium sulphate in 100 parts 26*66

Sodium „ 44-81

„ chloride 6*11

„ phosphate 8'42

Calcium „ 7*83

Magnesium „ 4*55

Chondrigen.

The substance of which the matrix of hyaline cartilage and the
capsules of the cartilage cells in the other forms of cartilage is
composed, resembles in many particulars collagen, but differs from it
in the product which it yields by the prolonged action of boiling
water. As it is generally believed that by this action a body to
which the term Chondrin has been given is formed, the mother
substance has received the name of Chondrigen.

Chondrigen is unacted upon by cold water, and swells very
slightly in acetic acid. It is dissolved by concentrated mineral acids
and caustic alkalies. When heated (in sealed glass tubes or in a
Papin's digester) in water at a temperature of 120°C. for three or
four hours chondrigen dissolves ; the solution contains chondrin.

Chondrin.

Costal cartilage is boiled for a few minutes, arid
Preparation. • ,, ,, • i i •

is then scraped so as to remove the penchondrium.

It is then finely divided and boiled for twenty-four hours with water ;
or placed in a Papin's digester, and heated in water at 120°C., for
three or four hours. The solution thus obtained is filtered so as to
separate it from insoluble matters, such as elastic tissue, cellular
elements, &c., and it is then precipitated with acetic acid. The pre-
cipitate is then extracted with alcohol or ether. It may be again
dissolved in hot water and the solution poured into a large excess
of absolute alcohol, when the chondrin which precipitates is separated
and dried. When dry it presents the appearance of a hard, trans-
parent mass, devoid of smell and taste l.

General re- Chondrin is insoluble in cold water, in alcohol,

actions. ether, or chloroform. It is soluble in hot water, and

aqueous solutions of chondrin gelatinize exactly like

1 The description of the preparation of Chondrin is mainly borrowed from Gautier,
Chimie Appliquee, <&c., Vol. i., p. 346.

270

CHONDRIN.

[BOOK i.

solutions of gelatin. They are precipitated by the following reagents
which have no such action when added to solutions of gelatin : —
acetic acid, the precipitate being insoluble in excess of the pre-
cipitant, but soluble if some alkaline salt be added ; solutions of
alum, the precipitate being soluble in excess of the reagent ; solutions
of silver nitrate and copper sulphate, the precipitates being soluble in
excess of the reagents ; solution of lead acetate, the precipitate not
soluble in excess. Solutions of chondrin have been said to be
rendered only slightly turbid by mercuric chloride and by tannic
acid. The Author is inclined to rely on the very positive statement of
J. Miiller that these reagents exert the same action on chondrin
as on gelatin.

Elementary Very great discrepancies exist between the results

composition of various analyses of this assumed chemical individual,
as will be observed by a study of the following Table.

COMPOSITION OF CHONDRIN.

of Chondrin.

Mulder.

Fischer und
Boedeker.

Schiitzenberger
und
Bourgeois.

v. Meliring.

Carbon
Hydrogen
Nitrogen
Sulphur
Oxygen

49-3
6-6
14-4
0-4

20-3

50-0
6-6
14-4
0-4

28-6

5016
6-58
1418

29-08

4774
676
13-87
0-60
31-04

The following table exhibits the relative composition of gelatin,
chondrin and mucin, according to the analyses which appear to be
most trustworthy.

Gelatin
(Hofmeister).

Chondrin
(Schiitzenberger)

Mucin

(Obolensky).

Carbon
Hydrogen
Nitrogen
Oxygen

5075
6'47
17-86
24-92

50-16
6-58
14-18

29-08

52-31

7-22
. 11-84
28-63

Products of When finely divided cartilage is boiled with dilute

decom- hydrochloric or sulphuric acids there is formed a body

position of resembling acid albumin, and a substance which pos-
Chondrin. sesses a sweet taste and reducing properties analogous

to those of true sugars \ This body has been termed Chondri-glucose.

1 Fischer und Boedeker, "Kiinstliche Bildung von Zucker aus Knorpel (Chondro-
gen), &c." Ann ale. n der Chemie und Pharm., Vol. cxvu. (1861), p. 111.

CHAP. VII.] THE CONNECTIVE TISSUES. 271

According to Fischer and Boedeker, this body is laevo-gyrous and is
capable of undergoing the alcoholic fermentation. According to
Hoppe-Seyler1, the body which reduces cupric salts is a nitrogenous
body and is identical with the body obtained by boiling mucin with
dilute acids.

"When chondrin is subjected to prolonged boiling with dilute
sulphuric acid, it yields leucine, but no tyrosine or glycocine.

(Hoppe1).

When chondrin is heated with barium hydrate, Schiitzenberger and
Bourgeois 2 have found that the products of decomposition are some-
what different from those yielded by gelatin under the same circum-
stances. In both cases carbon dioxide, oxalic and acetic acid, and
ammonia are obtained, in addition to a mixture of amido-acids. The
quantity of acetic acid yielded by chondrin is, however, three times as
great as that yielded by gelatin. In the mixture of amido-acids no
glycocine is present.

Doubts as Amongst the tissues which are supposed to be

to the exist- composed mainly of chondrigen is the substance of the
ence of Chon- cornea. In an investigation on the chemical composition
of this structure, Morochowitz3 has arrived at the
conclusion that the primitive fibrillae of the ground substance of
the cornea consist of collagen, and that the supposed chondrin is a
mixture of gelatin and mucin. After extracting the tissue with lime-
or baryta-water or with lOp.c. solution of NaCl, it yields on being
treated with boiling water pure gelatin. From the alkaline solutions
mucin can be thrown down by the addition of an acid. Morochowitz
has further investigated cartilage from various sources, and has found
that after treatment with reagents which dissolve mucin, as lime- or
baryta- water, 10 p.c. solution of NaCl, or J p.c. solution of caustic
soda, whilst mucin is removed, the substance which is left undissolved
is on boiling readily converted into perfectly normal gelatin. Accord-
ing to this author chondrin is to be looked upon as no pure substance,
but as a mixture of gelatin, mucin and salts.

If these views be, as the Author believes, correct, all the tissues
belonging to the connective tissue group, possess common chemical
character in that their ground substance is in all cases a body
transformed into gelatin by the prolonged action of boiling water;
this being mixed in greater or less proportion with mucin which,
as we have shewn, undoubtedly plays the part in many forms of
connective tissue of a connecting or cementing substance.

1 Hoppe, "Ueber das Chondrin und einige seiner Zersetzungsproducte." Journ.f.
praJct. Chemie, Vol. LVI. (1852), p. 129.

a Schiitzenberger et Bourgeois, " Eecherches sur la constitution des matieres col-
lagenes." Comptes Rendus, LXXXII. 262.

3 Morochowitz, " Zur Histochemie des Bindegewebes." Verhandl. d. naturhht.
med. Vereins zu Heidelberg, Vol. i. part v.

272 BONE. [BOOK i.

SECT. 4. OSSEOUS TISSUE OR BONE.

structural TQe nard tissue which forms the scaffolding and.

Elements of support of the soft parts of our bodies, although on
Bone. superficial examination, appearing so different from the

other members of the group of connective tissues, possesses the closest
affinity to them, as is evident not merely from developmental
considerations but from a study of its chemical composition.

All the bones of the skeleton are invested by a fibro-vascular
membrane, the periosteum, which conveys to them the great majority of
the blood-vessels which supply them, and which contains on its inner
layer certain cells — osteoblasts — which are the active agents in the
growth of bone, and in virtue of which the periosteum possesses the
power of forming new bone.

Those bones which articulate with others have no periosteal
covering over their articular ends, which are tipped with cartilage.

The external part of all bones has a very dense structure ; the
interior is either hollowed out into a cavity termed the medullary
cavity, or is occupied by a trellice-work of bony plates which
constitute the cancellated tissue. Those bones which possess a
medullary cavity present near their articular ends much cancellated
tissue.

The medullary cavity lodges the medulla or yellow marrow, which
is composed of fat cells supported by a frame work of connective
tissue, and abundantly supplied with blood-vessels; the cancelli
or spaces of the cancellated tissue afford support to the so-called red
marrow, which is a tissue in which a large number of cells identical
with the colourless cells of the blood are found, besides certain cells
which resemble the nucleated coloured corpuscles of embryonic blood.

It also contains large giant cells with many nuclei to which
the name of myeloplaxes is applied, and which are identical in
appearance and probably in functions with those cells which under
the name of osteoclasts are supposed to be the active agents in
the formation of the medullary cavities of growing bone.

Though the chief blood-supply of bone is drawn from the perios-
teum, both arteries and veins of considerable size enter by so-called
' nutritious foramina ' and are distributed to the marrow and so-called
endosteum, as the connective tissue lining the medullary cavity is
called ; some of the branches of the nutrient vessels anastomose with
the blood-vessels of the hard tissue.

The blood-vessels contained in the hard substance of bone lie in
canals — the Haversian canals. Around these canals the bony sub-
stance is arranged in concentric lamellae. ' In these lamellae are
cavities, also arranged concentrically, called lacunae, and from these
proceed minute canals, the canaliculi, which establish a communica-
tion between adjacent lacunae, and between the lacunae which are in

CHAP. VII.] THE CONNECTIVE TISSUES. 273

the circle nearest the Haversian canal and the canal itself. The
lacunae lodge nucleated masses of protoplasm — the bone corpuscles or
bone cells — which do not send processes into the canaliculi.

The walls of the calcified lacunae and canaliculi, as well as of
the Haversian canals, appear to be composed of a tissue resembling
elastic tissue1 and are left surrounding the bone cells when softened
bone is boiled for many hours in water or when it is subjected to
digestion with trypsin2.

Besides the lamellae or sheets of bony substance which are
arranged in concentric layers around the Haversian canals, and which
may be termed the lamellae of the Haversian systems, other lamellae
are arranged concentrically around the medullary canal and imme-
diately beneath the periosteum ; these may be termed fundamental
lamellae.

The minute structure and arrangement of the soft parts of bone
can only be studied by making preparations of decalcified or softened
bones. In these preparations it may be shewn that the ultimate
lamellae of bone are transparent sheets which exhibit intercrossing
fibres, which possess the characters of the white fibres of connective
tissue. It may further be shewn that the fundamental lamellae are
perforated by fibres — the so-called perforating fibres of Sharpey —
which dip into the bone from the periosteum, and which appear to
have mainly the chemical characters of yellow elastic tissue.

The Water found in Bone.

All bones contain, when fresh, a considerable quantity of water.
The estimates of various observers differ remarkably in reference to
this matter. Volkmann estimates the mean percentage of water at
48 '6 p.c. of the fresh bone. According to Aeby's determinations
(which are certainly 'too low) bones just removed from the dead body
contain between 11 and 12 p.c. of water. According to this author
the water exists in a state of chemical combination, probably ana-
logous to that of water of crystallization. This view is based partly
on the constancy of the amount of water, but partly on the fact that
heat is evolved when dried bone is placed in water3.

The Animal or Organic basis of Bone.

Methods of When a bone is placed in a dilute mineral acid for

preparing de- some days, it gradually loses its rigidity, and although
calcified or retaining its form and general appearance, it becomes
bone- comparatively soft and pliable, so that a long and com-
paratively thin bone, such as the clavicle or the radius, may be tied
into a knot.

1 Hoppe, Virchow's Archiv, Vol. v. (1853) p. 170.

2 De Burgh Birch, "Erscheinungen bei Trypsinverdauung an Knochen." Central-
Matt f.d. med. Wissenschaft. 1879, p. 945.

3 Aeby, "Der Grund der Unveranderlichkeit der organischen Knochensubstacz."
Centralblatt f. d. med. Wissenschaft. 1871, No. 14.

G. 18

274 THE ORGANIC BASIS OF BONE. [BOOK I.

The following solutions may be employed for softening bones.

(1) A mixture of one part of hydrochloric acid and five of water.

(2) A mixture of nitric and chromic acids of the following com-
position : chromic acid 5 grms., nitric acid 10 cub. c., water 1000 c. c. l

(3) A solution containing from 2 to 5 parts of chromic acid in 1000
parts of water (Ranvier).

(4) A saturated aqueous solution of picric acid.

In the case of solutions 3 and 4 it is important that the fragments of
bone to be softened shall be very small.

Characters The organic basis of softened bone is insoluble in

of the organic cold water, but is for the most part soluble on pro-
basis of bone, longed boiling in water. The solution contains gelatin,
which is identical in its reactions with that body as it is obtained
from white fibrous tissue. The structures which are undissolved by
boiling water are the perforating fibres, and apparently the decalcified
walls of the lacunae, canaliculi, and of the Haversian canals, which
appear to be formed of a substance resembling elastin.

The organic basis of bone (which has by some writers been
termed ossein) then consists mainly of a body identical in chemical
reactions with collagen, mixed with a certain amount of elastin and
with the proteid matter of the bone cells. It is to be noticed that
the animal matter of cartilage before ossification does not consist of
normal collagen but of chondrogen (or, if we adopt Morochowitz's
theory, of a mixture of collagen and mucin). In the process of ossifi-
cation, which consists essentially in an intrusion of periostea! elements
into cartilage, which is pari passu removed by absorption, the animal
matter assumes all the characters of connective tissue proper.

There appears to be always some fat in bone, but its relations to
the organic basis are not known.

All organic matters are destroyed when bone is incinerated. The
following are the results of some analyses shewing the relative pro-
portion of organic and mineral matters in bone (Zalesky).

Organic matters. Mineral matters.

Bone of man (mean of 4 analyses) 34*56 65*44

„ „ ox (mean of 6 analyses) 32*02 67*98

„ „ guinea pig (mean of 2 analyses) 3470 65*30

The Mineral Matters of Bone,

The mineral matters of bone are deposited in the organic basis
in such a manner as to be invisible on microscopic examination.
They may for the most part be dissolved by employing the processes
already described as producing the decalcification of bone. They
may be obtained in a solid form by igniting or incinerating bone,
and they then retain the form of the original bones
bone).

1 Kutherford, Outlines of Practical Histology, pp. 3 and 82.

CHAP. VII.] THE CONNECTIVE TISSUES. 27-")

The following analyses l illustrate the composition of the mineral
matters of bone.

1. RESULTS OF THE ANALYSES BY HEINTZ3.

Ox. Sheep. Man.

(1) (2)

Ca 38-52 38-52 38'59 38'56

PO4 52-98 53-29 5375 53'87

C03 6-04 5-65 5-44 5'51

Fl 1-89 1-96 174 1*58

Mg 0-57 0-58 0-48 0'48

II. ANALYSES OF THE BONES OF CHILDREN. (RECKLINGHAtJSEN.)

Bones of skull Bones of child Bones of child of 6 years,

of child 3 14 days old. Femur,

days old. Skull. Femur. Cortical layers

(diaphysis). Epiphyses.

Ca 38-41 36-43 37'66 37'98 37*97

P04 56-20 56-96 54'81 54'86 5673

C08 4-85 6-02 7-06 6'88 4-97

Mg 0-54 0-5.9 0-47 0'28 0'33

HI. ANALYSES OF THE BONES OF MAN AND THE OX. (ZALESKY.3)

Man. Ox,

Ca 4013 40-69

P04 52-16 53-50

C03 7-81 8-45

Cl 018 0-20

Fl 0-23 0-30

Mg 0-29 0-28

From all the analyses which have been made we may legitimately
conclude that the proportion of mineral to organic matters in bone,
and even the relative proportion of the different elements, vary
remarkably little in animals of different species, and of different
ages.

The chief salts present in bone are five in number, of which four
are compounds of calcium, and one a compound of magnesium.
They are calcium phosphate, Ca32(P04); calcium carbonate, CaC03;
calcium chloride, CaCl2 ; calcium fluoride, CaFl2 ; magnesium phos-
phate, Mg32(P04). In addition to these, very small quantities of
sulphates and chlorides are always present.

1 Extracted verbatim from Hoppe Seyler, Physiologische Chemie, p. 105.

2 Heintz, "Ueber die chemische Zusammensetzung der Knochen." Poggendorff's
Annalen, Vol. LXXVII. (1849) p. 267.

3 Zalesky, " Zusammensetzung der Knochen von Menschen und Thieren." Med.
chem. Untersuchungen von Hoppe-Seyler. Part 1, p. 19 et seq.

18—2

276 THE MINERAL MATTERS OF BONE. [BOOK I.

The following exhibits the probable composition of the mineral
matters of bone calculated from the analyses of Zalesky.

Calcium phosphate (Ca32PO4) 83'889

Calcium carbonate (CaC03) 13'032

Calcium in combination with fluorine,

chlorine and organic acids 0'350

Fluorine 0'229

Chlorine 0183

98722

The occurrence of considerable quantities of a fluoride in bone
has, since it was first discovered, attracted the attention of many in-
vestigators (Chevenix, Morichini1, Gay Lussac, Berzelius 2, G. Wilson
and others). The presence of this element is readily proved by
heating powdered bone with strong sulphuric acid in a leaden
or platinum capsule, when hydrofluoric acid is given off, as can be
proved by its etching glass.

Constitu- ^ ^as been surmised that a combination of calcium

tion of the phosphate and calcium fluoride having the same consti-
minerai mat- tution as the mineral Apatite exists in bone ; the
ters of bone. composition of this mineral is shewn by the formula
Ca10M2( 6(P04).

In bone, however, the fluorine is present in very minute
quantities, the main compound having probably the composition
Ca10CO3, 6(P04). This matter is discussed again in connection
with dentine and enamel (see p. 291).

Zalesky has shewn that chlorides exist in bone in two conditions,
a portion being soluble in water, and another portion being only
dissolved by acids.

influence of The influence of food, rich or poor in earthy salts,

food on mine- upon the composition of bone has been studied by
rai matters of various writers with entirely different results. Thus
Forster 3 observed a diminution in the proportion of
calcium in the bones of dogs fed upon a diet in which calcium salts
were deficient. In similar experiments performed on dogs, Zalesky 4
obtained altogether negative results. Weiske 5 came to similar con-

1 An account of Morichini's discovery of fluorine in fossil teeth was given in a
letter addressed by Gay Lussac to Berthelot in the Annales de Chimie of 30 Fructidor,
an 13 (1805).

2 Berzelius, "Extrait d'une lettre a M. Vauquelin sur le fluate calcaire contenu
dans les os et dans 1'urine." Ann. de Chim.t Vol. LXI. (1807) p. 256.

3 J. Forster, "Ueber die Verarmung des Korpers speciell der Knochen an Kalk bei
ungeniigender Kalkzufuhr." Zeitschrift f. Biolog., Vol. xn. p. 464.

4 Zalesky, Op. cit., p. 44 et seq.

5 Weiske, "Einfluss verschiedener der Nahrung beigemengter Erdphosphate auf
die Zusamrnensetzung der Knochen." Zeitschr. f. Biolog., Vol. vm. p. 239. "Ueber
Knochenzusammensetzung bei verschiedenartiger Ernahrung." Zeitschr. f. Biol., Vol.
x. p. 410.

CHAP. VII.] THE CONNECTIVE TISSUES. '277

elusions. This matter will be referred to again (p. 282) in considering
the etiology of Rickets.

It was asserted by Papillon1 that when animals are supplied with
food specially rich in magnesium, aluminium, and strontium salts,
these elements enter into the composition of the mineral matter of
the bones. J. Konig2 contradicts the researches of Papillon in so far
as compounds of magnesium and aluminium are concerned, but
confirms them in respect to strontium. In the bones of rabbits
fed with strontium phosphate, he found as much as 5 '37 p. c. of
strontium. According to Weiske3 both Papillon and Konig have
fallen into error. In the bones of rabbits fed with strontium phos-
phate, Weiske found only minute traces of strontium.

The Composition of the Marrow of Bone.

As has been already said, it is customary to distinguish between
the yellow marrow, which is contained in the medullary cavity of the
long bones, and the red marrow which is lodged in the cancellated
tissue of spongy bone.

The former on microscopical examination has all the characters of
adipose tissue, being composed of fat cells supported by connective
tissue fibres and blood-vessels ; the latter contains cells which resem-
ble the white cells of the blood, and certain cells which resemble the
nucleated coloured corpuscles of the blood of the embryo.

The dried yellow marrow consists chiefly of fat which appears
to have the normal composition of the fatty matter of adipose
tissue. The red marrow is said to contain albumin and a free
organic acid, supposed by Berzelius4 to be lactic acid.

Heymann5 has detected hypoxanthin in marrow of healthy bones,
and Nasse 6 has found in the red marrow of the ribs of old horses,
microscopic agglomerations of granules, having a diameter of from
O'OOT — 0 015 mm., which contain oxide of iron (probably also ferric
phosphate) and organic matters and are coloured intensely blue by
ferrocyanide of potassium ; these are identical with similar bodies
found in the spleen of man and the horse.

These chemical facts, taken in connection with the observations
of cases of myelogenic leukaemia, give great countenance to the view

1 Papillon, " Recherches experimentales sur les modifications de la composition
immediate des os." Comptes Rendus, Vol. LXXVI. (Ib73) p. 352.

2 Konig, " Substitution des Kalkes in den Knochen und Einfluss kalkarmer Nahrung
auf die Zusammensetzung der Knochen." Ijeitschrift f. Biolog., Vol. x. p. 69.

» Weiske, Zeitschr. f. Biol, Vol. x. p, 410.

4 Berzelius, quoted by Gorup-Besanez, Phys. Chem., p. 631.

5 Heymann, " Ueber das Vorkommen von Hypoxanthin im normalen Knochen-
marke." Pfliiger's Archiv, Vol. vi. p. 184.

B Nasse, "Ueber das Vorkommen eisenhaltiger Korner im Knochenmarke." Ab-
stracted in Maly's JahresbericM, Vol. vn. (1878) p. 300. "Ueber den Eisengehalt der
Milz." Maly's Jahresbcricht, Vol. iv. (1874) p. 91.

278

RESULTS OF ANALYSES OF VARIOUS BONES. [BOOK I.

entertained by many histologists that the red marrow is an organ
concerned in the transformation of the coloured cells of the blood.

In the marrow of the bones of rabbits, Rustitzky1 found mucin ;
he was unable to discover this substance in the fat marrow of
ox bones.

RESULTS OF COMPARATIVE ANALYSES OF BONES BELONGING TO
DIFFERENT MEMBERS OF THE ANIMAL KINGDOM. (FREMY.2)

Name of Bone.

Ash
per
cent.

Calcium
Phos-
phate.

Mag-
nesium
Phos-
phate.

Calcium
Carbon-
ate.

Male foetus, 4 months ; femur . . .
,, ,, 6 months ,, ...
Female foetus ,, ...
„ „ 7 months; humerus . .
Girl, born at term ; femur ....
Boy, 18 months ,,

61-7
62-8
63-0
62-8
64-8
64-6

60-2
60-2

60-8
61-5

Woman, 22 years; scapula ....
,, „ cranium ....
femur ....
„ „ humerus. . . .
Man ; spongy part of femur ....
„ dense „ „ . . . .
Man, 40 years; femur.

63-3
64-1
64-6
64-1
61-0
65-0
64-2

60-0

57-8

56-9

1-3

10-2

Woman, 80 ,, „

64-6

60-9

1-2

7-5

64-5

58-1

1-2

10-0

00

64-3

57-4

1-3

9-3

„ 88 „ spongy part of femur
„ 97 „ femur

59-7
64-9

54-0
57-0

1-2
1-2

7-0
9-3

Egyptian mummy, female; femur . .
Saky ; femur

65-0
64-0

58-7

1-7

5-9

Kinkajou; femur

62-0

Genet
Bitch; femur

70-2
62-1

59-0

1-2

6-1

Youns[ lioness: femur .

64-7

60-0

1-5

6-3

Panther; femur . .

65-6

Walrus

63-1

53-9

1-5

9-3

Habbit; feuiur .

66-3

58-7

1-1

6-3

Guinea pig

71*8

Indian elephant

66-8

62-2

1-2

5-6

1 Rustitzky, Centralblatt f. d. med. Wissenschaft. 1872, p. 562.

2 E. Fr&my, "Recherches chimiques sur les os." Anhales de Chimie et de Physique,
ser. 3, Vol. XLIII. pp. 47—107.

CHAP. VII.]

THE CONNECTIVE TISSUES.

279

Name of Bone.

Ash
per
cent.

Calcium
Phos-
phate.

Mag-
nesium
Phos-
phate.

Calcium
Carbon-
ate.

Java rhinoceros
Horse ' femur . ...

65-3
70-4

60-0

2-3

5-2

Calf, still-born ; spongy part of femur
„ „ dense „ „
5 months j femur

61-5
644
69-1

60-5
59-4
61-2

1-2
1-7
1-2

5-2

8-4

Cow full grown

70-7

old

71-1

5) 5) ?>

Ox ' humerus

71-3

70-4

62-5
61-4

2-7
T7

7-9

8-6

70-2

62-4

1-7

7-9

Bull' femur

69-3

59-8

1-5

8-4

Lamb „
Sheep . .

67-7
70-0

60-7
62-9

1-5

1-3

8-1

7-7

Kid „

Cachalot

68-0
62-9

58-3
51-9

1 2
0-5

8-4
10-6

Whale ; spongy part of femur . .
Ea^le .

57-5
70-5

60-6

1-7

8-4

Vulture

66-2

Owl .

71-3

61-6

1-5

8-8

Ostrich ; dense part of femur
„ spongy part „ . . . .
Bustard

70-0
67-0
71-1

Chicken

68-2

64-4

M

5-6

Turkey

67-7

63-8

1-2

5-6

Partridge

70-7

65-4

Heron

70-6

62-5

1-5

10-2

Thrush
Humming bird ; bones of head . . .
„ „ „ limbs . . .
Teal

66-6
55-0
59-0
73-5

63-0
68-4

1-3

5-6

Turtle j carapace . . .

64-3

58-0

1-2

Land tortoise ; carapace
Crocodile ; cutaneous bone ....
Crocodile
Serpent

64-0
64-6
64-0
67-5

56-0
58-3
58-3

1-2
trace
0-5

10-7
9-7

7-7

Cod

61-3

55-1

1-3

7-0

Barbel

60-2

Sole

54-0

Shad

50-0

Carp

61-4

58-1

1-1

4-7

Pike

66-9

64-2

1-2

4-7

Eel

57-0

56-1

traces

2-2

Dogfish

62-6

Ray; cartilage

30-0

27-7

trace

4-3

280

FOSSIL BONES. OSTEOMALACIA.

BOOK I.

Fossil bones contain a smaller quantity of organic
matter than recent bones. This appears, however, to
yield normal gelatin on boiling. They contain the
same mineral matters as recent bones.

Composi-
tion of fossil
bones.

ANALYSES OF VAEIOUS FOSSIL BONES. (FEEMY.1)

Name of Bone.

Ash
per cent.

Calcium
Phos-
phate.

Mag-
nesium
Phos-
phate.

Calcium
Car-
bonate.

Calcium
Fluoride
and
Silica.

Organic
matter.

Ox, from the caves of Ores-

ton; rnetatarsal bone, exter-

nal portion having the aspect

of wood ....

80-74

71-1

1-5

11-8

10-3

— internal portion of same

(very friable).

80-6

71-5

1-7

11-3

11-0

— spongy portion of same .

84-2

63-3

1-2

5-2

17-2

8-0

Rhinoceros from Sansan

(Gers)

— vertebrae

83-4

59-0

41-3

2-6

trace

— ribs ....

83-1

66-8

27-5

1-4

trace

Hyena, from the caves of

Kirkdale; long bone .

75-5

72-0

1-3

4-7

20-0

Rhinoceros; dorsal verte-

brae ....

69-5

25-7

0-4

57-5

8-5

Rhinoceros ; humerus

73-0

32-4

0-4

64-0

6-2

Bear; dense part of bones

83-9

59-7

0-4

23-6

9-8

spongy part .

76-7

23-1

1-2

67-5

14-0

Anoplotherium; caudal

vertebra

84-0

53-1

04

20-4

19-4

Tortoise; vertebrae .

87-0

61-1

0-7

10-6

18-6

THE CHANGES WHICH BONE UNDERGOES IN DISEASE.

Changes of
the bone in
Osteomalacia
or Malacos-

teon.

Osteomalacia-.

By the name of Mollities Ossium, Osteomalacia
or Malacosteon, a disease is designated in which
the bones become deprived of a large part of their
mineral matter and liable to bend or to break.
Not only are the mineral matters removed, but the
organic basis undergoes marked structural alterations ; the medullary
cavity of long bones is enlarged and is often filled with hyperaemic
red marrow ; in some cases a yellow, in others a mucoid, marrow is
found.

In some cases of Osteomalacia the bones do not yield gelatin when
boiled ; in other cases they do.

1 Fr6my, Op. cit. page 88.

CHAP. VII.]

THE CONNECTIVE TISSUES.

281

The fatty matter of bone seems to be very greatly increased
in this disease.

Some observers have noticed that the bone possesses an acid
reaction and that this is due to the presence of lactic acid1. It has
indeed been surmised that the development of lactic acid is the
primary cause of the morbid change in the bones2. This view has
been supported by a narrative of experiments in which animals were
subjected to large and long-continued doses of lactic acid with the
result that they became affected with rickets which afterwards
passed into osteomalacia (Heitzmann2).

It has been justly remarked that the experience of physicians who
have experimented with Oantani's method of treating diabetes mellitus,
which consists in giving large doses of lactic acid, does not support
Heitzmann's statements, as no one has observed osteomalacia to result3.

ANALYSES OF THE BONES IN OSTEOMALACIA4.

I.

H.

III.

IV.

In 100 parts.

Femur of
man aet. 40

Eib from
same case as I.

Femur of
man aet. 60

Vertebrae
of child

(Lehmann).

(Lehmann).

(von Bibra).

(Marchand).

Organic basis

48-83

50-48

32-54

75-22

Fats

29-18

23-13

415

6-12

Soluble salts

0-37

0-63

1-35

1-98

Calcium phosphate

17-56

21-02

53-25

12-56

„ carbonate

3-04

3-27

7-49

3-20

Magnesium phosphate

0-23

0-44

1-22

0-92

Rachitis.

Changes Rickets is a general disorder of nutrition, accom-

in bone in panied by changes especially affecting the epiphyses

Rachitis or of bones. An abnormal proliferation of cartilage cells
occurs, leading to an enlargement of the epiphyses,
whilst the growing bones being deficient in earthy salts become
distorted. When calcification occurs the deformities which have
been produced are often rendered permanent. It is a disease which
affects bones in the process of development or rather cartilage which
is being converted into bone — and it therefore differs fundamentally
from osteomalacia, in which a morbid process causes the absorption

1 C. Schmidt, " Knochenerweichung durch Milchsaurebildung. " Annalen d.
Chemie und Pharm., Vol. LXI. (1847) p. 142.

2 Heitzmann, " Ueber die Wirkung der Milchsaurefiitterung auf Thiere." Anzeiger
der kais. Akad. d. Wissensch. Wien, 1873, No. 17. Abstracted in Maly's Jahresbericht,
Vol. in. (1874) p. 229.

3 The reader may consult a paper by Dr Ernst Heiss entitled " Kann man durch
Einfiihrung von Milchsaure in den Darm eines Thieres den Knochen anorganische
Bestandtheile entziehen?" Zeitschr. /. Biologic, Vol. xn. p. 151. Heiss found that
the results were negative, even though lactic acid was administered to annuals fed upon
a diet deficient in lime salts.

4 Gautier, Chimie appiique'e a la Physiologic, a la Pathologic, &c., Tome n. p. 541.

282

ETIOLOGY AND PATHOLOGY OF RICKETS. [BOOK I.

of the salts of fully formed bone and further serious changes in the
decalcified framework.

In rickets the bones become specifically lighter than in health ;
the unossified cartilage contains an increased proportion of water; the
long bones contain an increased quantity of fatty matter. The
amount of fat is, however, much less in the bones of rachitis than in
those affected with osteomalacia. It has been found by Lehmann
and Marchand that occasionally in rickets the bones do not yield a
normal gelatin when boiled.

COMPOSITION OF BONE IN RACHITIS1.

In 100 parts.

Femur
(Marchand).

Tibia
(Lehmann).

Humerus
(Bagsky).

Inorganic matters

20-60

33-64

18-88

Organic matters

79-40

66-36

81-12

Calcium phosphate
Magnesium phosphate
Calcium carbonate

1478
0-80
300

26-94 )
0-81 /

4-88

15-60
2-66

Soluble salts

1-02

1-08

0-62

Fats

7-20

6-22 )

Collagen

72-20

60-14 I

81-12

Calcium fluoride |
and loss j

i-oo

0-99 j

Etiology and Very different views have been advanced on the

Pathology of etiology and pathology of rickets. Petit 2 first sug-
Rickets. gested that the disease is caused by the too early

weaning of infants ; since his time others, with no less reason, have
maintained that too prolonged lactation often acts as a predisposing
cause, the impoverished milk being incapable of supplying the grow-
ing infant with all the materials which its organism requires.

Whilst some have considered that rickets is induced by an
improperly adjusted diet, in which the different groups of food con-
stituents are not in their proper proportions, a majority of writers
have advocated the view that the disease specially depends upon a
deficiency in the lime salts of the food, or in a deficient absorption
of lime salts. All these views have been supported by experimental
researches which have led to diametrically opposite conclusions ;
certain experimenters having, for instance, succeeded in inducing
rickets by feeding young growing animals upon meat instead of milk3,

1 Extracted from v. Gorup-Besanez, Lehrbuch d. phys. Cliemie, p. 635.

2 Petit, Traite des maladies des os, 1741.

3 T. Guerin, These de Paris, 1859, p. 24. Quoted, at secondhand, by Leon Tripier,
Dictionnaire ency elope dique des sciences me'dicales, Troisieme eerie (Paris, 1874).
Article " Bachitisme."

CHAP. VII.] THE CONNECTIVE TISSUES. 283

others by cutting off the salts of lime more or less completely
(Letellier, von Bibra, Chossat, Milrie Edwards1), whilst not a few have
found that although the animals subjected to these conditions suffered
in health and even died, they shewed no symptoms of rickets (Leon
Tripier2, Weiske3). We think, from a careful perusal of the experi-
mental evidence bearing on this question, that we may draw from
it the following conclusion. When young animals are subjected
to an insufficient diet or one in which certain of the alimentary con-
stituents are deficient, there is engendered a predisposition to rickets,
although there is no evidence to shew that such insufficient or improper
diet can, acting alone, induce the disease.

Amongst the views which have been promulgated and adopted by
eminent writers on this subject is that which ascribes the chief part
in the production of the disease to the formation of lactic acid in the
alimentary canal ; the acid thus formed is supposed to be absorbed
into the blood and to act ' as an irritant on the osteoplastic tissues'
and 'as a solvent on the calcareous salts deposited in the bones,
promoting their elimination4.'

This theory rests upon the most unsatisfactory evidence, as, that
the amount of lime excreted in rickets is increased (a fact which
has not been established by one single properly conducted observation) :
that rachitic bones have been found to contain lactic acid after death :
and that the urine of rachitic children contains lactates.

Even assuming that large quantities of lactic acid were generated
in the alimentary canal these would necessarily be converted into
lactates in the blood. No one has been bold enough to assume
that in rickets, or any other disease, the blood loses its alkaline reac-
tion, for no one could conceive of an acid reaction of the blood being
compatible with a prolonged continuance of its functions ; and yet
in order that lactic acid could exert any solvent action, it would
be necessary that it should exist in a free condition in the blood
or that, by an unknown chemical decomposition, alkaline lactates
should be decomposed in the bones. This theory like all crude

1 Amongst more recent researches which confirm the older writers on the possibility
of inducing rachitis by a diet poor in lime salts are those of T. Lehmann, " Ueber den
Einfluss der Nahrung auf die Knochenbildung." Abstracted in Maly's Jahresbericht,
Vol. vin. (1879) p. 272.

2 Leon Tripier. See the admirable article referred to in note 3, p. 282.

3 Weiske, "Einfluss Kalk- oder Phosphor saure armer Nahrung auf die Zusammen-
setzung der Knochen." Zeitschriftf. Biologie, Vol. vn. pp. 179 — 183 and pp. 333—337.

4 This view is adopted by Senator in his article on Kickets in Ziemssen's Cyclopaedia
of the Practice of Medicine, English edition. Vol. xvi. p. 178. We quote his very
words, " To sum up : the morbid process which underlies the development of rickets
may, in accordance with the results of experiments and the clinical observations we
possess, be explained in the following manner. Owing to digestive disturbance,, either
preexistent or brought on by improper feeding, lactic acid is generated in the system ;
this operates, on the one hand, as an irritant on the osteoplastic tissues ; on the other,
as a solvent on the calcareous salts deposited in the bones, promoting their elimination.
At the same time the supply of earthy matter is reduced, either directly (as in cases of
protracted lactation) or indirectly (as when diarrhoea carries off the lime-salts from the
intestines before they are absorbed)."

284

RICKETS. CARIES. NECROSIS.

BOOK I.

chemical theories of disease does not stand the test of even a super-
ficial criticism.

We shall probably form a nearly correct idea of the essential
nature of rachitis if we look upon it as a morbid process having its
seat in the ossifying epiphyses, and in newly-formed bone — a
morbid process which is the local expression of a general disorder
of nutrition. As a result of the latter, the cartilage cells undergo
an abnormal proliferation and the newly-formed bone cells are
more or less unfit to separate from the blood the lime salts which
are needed for the hardening of the newly-formed ground sub-
stance in which they lie. As a result of the excessive proliferation of
cartilage cells, the bones enlarge, especially at their epiphyses, and,
because as they grow they do not concomitantly harden, they yield
to external pressure and become deformed.

Caries.

The following is a tabular view of the composition
of the bone in caries, according to the analyses of
Becquerel and Rodier \

Changes in
the bone in
Caries.

Lumbar

/Meta-

Met. bone,

Phalanx

nffar>i-&A

vertebra

carpal

articular

of

of a

bone.

end.

finger.

caries.

woman

aet. 40.

Calcium phosphate)
„ fluoride /

4977

31-36

49-36

51-53

44-05

Calcium carbonate

7-24

4-07

8-08

5-44

3-45

Magnesium phosphate

1-11

0'83

0-98

3-43

1-02

Other salts

0-30

0-30

0-40

0-91

1-70

Collagen

37-97

59-36

37-47

35-69

41-42

| Fats

3-61

4-08

3-00

3-00

8-36

Changes in
the bone in
Necrosis.

von Bibra.

Collagen

Necrosis.

In necrosis the organic matter of bone is gradually
removed.

The following is an analysis2 of necrosed bone by

Fats

Calcium phosphate with)
a little calcium fluoride/
Calcium carbonate
Magnesium phosphate
Soluble salts

in 100 parts

19-58
1-22

72-63

4-03
1-93
061

1 Becquerel et Rodier, Traite de Chemie patholoyique, p.
3 Quoted by Gantier, Op. cit. Vol. n. p. 543.

546.

CHAP. VII.] THE CONNECTIVE TISSUES. 285

METHODS FOLLOWED IN THE QUANTITATIVE
ANALYSIS OF BONE.

i The bones to be analyzed are carefully denuded

ry prepara- °f thejr periosteum. They are then divided with a saw.

tion of bones The cancellated tissue is carefully removed, by means

to be subject- of a chisel, from the compact bone, and the latter

L to analy- js t]ien divided into somewhat small pieces. Each

of these is then wrapped in paper and being placed

on an anvil is struck with a hammer so as to crush it into minute

fragments. These are then powdered in a steel mortar and the

powder passed through a very fine sieve.

Some writers recommend that the crushed bones should, before
pulverization, be tied in a small muslin bag which is suspended
in distilled water, which is to be renewed several times, with the
object of separating from the bone soluble constituents which do
not properly belong to it, but which are of the nature of accidental
contaminations. If this process be followed, the fragments of bone
after extraction with cold water are dried in an oven and thereafter
pulverized.

Determination of the quantity of Fat in Bone.

A weighed quantity of powdered bone which has been dried
at 130° C. is extracted with ether as in the apparatus of Drechsel
(see p. 265). The ethereal solution is evaporated to dryness and
weighed.

Determination of the total quantity of Ash in Bone.

A quantity of the fat-free powder which has been dried at
1 30° C. is weighed in a platinum crucible and ignited until the ash
is perfectly white. The residue is moistened with solution of
ammonium carbonate, and then heated gently. The object of
this operation is to restore the carbon dioxide which may have
been expelled from the bases by the strong heat to which they have
been subjected.

Determination of the quantity of Chlorine in the Ash.

The ash resulting from the preceding operation is finely powdered
and dissolved, with the aid of heat, in dilute nitric acid. The
solution is concentrated and then treated with silver nitrate, which
precipitates all the chlorine as silver chloride. This is washed
by decantation, ignited and weighed according to the ordinary
rules of analysis. 1 part of AgCl corresponds to 0 '2 47 24 parts of
chlorine.

286 ANALYSIS OF MINERAL MATTERS OF BONE. [BOOK I.

Determination of the amount of Calcium in Bone.

The filtrate from the last operation is treated with solution of
NH4Cl, so as to precipitate completely, as AgCl, the silver which
it contains ; the nitrate is saturated with ammonia, and then acetic
acid is added so as to cause complete solution of the precipitate.
In the case of old bones which have been macerated or buried,
a certain quantity of phosphate of iron is present in the bone ash,
and it forms that part of the ammonia precipitate, just mentioned,
which is not dissolved by an excess of acetic acid. It may be
collected on a filter, washed, dried, ignited and weighed as Fe2(P04)2.
' From the acetic acid solution of the ammonia precipitate, the
calcium is thrown down by adding solution of ammonium oxalate.
The fluid, with the precipitate, is heated on a water-bath, set aside
in a warm place for 24 hours, and then thrown on a filter of which
the amount of ash is known ; the filtrate is collected and kept. The
precipitate is washed with water holding a little ammonia in solution.
It is then dried, and the precipitate and filter-paper with adhering
precipitate are separately ignited, as directed in works on quantitative
analysis. 100 parts of the resulting CaCO3 correspond to 40'00
of Ca.

Determination of the Magnesium.

The filtrate from which calcium has been precipitated by means
of ammonium oxalate is evaporated to a small bulk. It contains
all the magnesium of bone in the form of phosphate, which is preci-
pitated as ammoniaco-magnesian phosphate (Mg]SH4P04+ 6H20)
on saturating with ammonia. With this object an excess of ammonia
is added and the fluid is set aside for 24 hours in a warm place ;
it is then filtered through a small filter, the precipitate is washed
with ammoniacal water, dried and ignited.

100 parts of Mg2P207 correspond to 21'622 of Mg.

Determination of Phosphoric acid.

The filtrate from the precipitate of ammoniaco-magnesian phos-
phate in the last operation is now treated with magnesia mixture1
and set aside for 24 hours. Again a precipitate of ammoniaco-
magnesian phosphate forms, which corresponds to all the phosphoric
acid not combined with magnesium. The precipitate is treated as in
the last operation.

100 parts of Mg2P2O7 correspond to 78'37872 of 2(P04).

1 " Magnesia mixture is made by dissolving one part of recrystallized magnesium
sulphate and one part of pure ammonium chloride in eight parts of water, and adding
to the mixture four parts of moderately strong ammoniac solution. The liquid is
allowed to stand for a few days in a corked flask; it is then filtered and preserved
in a well-stoppered bottle." Thorpe's Quantitative Chemical Analysis, p. 111.

CHAP. VII.] THE CONNECTIVE TISSUES.

287

Dster mination of Carbonic acid.

About 5 grammes of the bone dried at 130° C. are employed
for this determination, which may be conveniently effected with the
aid of Geissler's apparatus as figured below. (Fig 53.)

The weighed quantity of bone is introduced through the tubu-
lature a into the bulb A, and then three or four cubic centimetres of
distilled water are added.

The stopper a is then inserted and the stopcock b leading from B
is turned so as to shut off the latter from A. The stopper which fits
into the upper part of B having been removed, pretty strong, but yet
non-fuming, hydrochloric acid is poured 'into B. The stopper is then
replaced. The small perforated stopper e at the upper part of C is

FIG. 53. G-EISSLER'S APPARATUS FOB THE ANALYSIS OF CARBONATES.

now removed, and with the aid of a small funnel, concentrated
sulphuric acid is poured into C to about the level shewn in the
drawing. The perforated stopper is then replaced. The whole
apparatus is then carefully dried with a clean cloth, placed in the
balance case for half an hour and then very carefully weighed.
After being taken from the balance the stopcock b is momentarily
opened, so as to allow a small portion of the contents of B to flow
into A. Carbonic acid is disengaged, and this passes through
the narrow tube c into the wider tube d, and thence, through two
small holes situated near its base, it bubbles through the 'sulphuric
acid contained in c. The effect of this passage of the moist carbon

288 ANALYSIS OF MINERAL MATTERS OF BONE. [BOOK I.

dioxide through the concentrated sulphuric acid is to dry the gas and
to retain the moisture in the apparatus. When the evolution of CO2
has ceased, stopcock b is again opened for an instant so as to allow a
fresh quantity of hydrochloric acid to act upon the bone. When the
evolution of C02 has ceased and does not recommence on the addition
of a few drops more of the acid, the whole apparatus is placed on
a water bath so as to heat it gently for a few minutes. The stopcock
b is then opened, the stopper at the upper part of B is temporarily
removed and a piece of narrow india-rubber tube is slipped over the
upper narrow portion of the perforated stopper e. The experimenter
then placing the free end of the india-rubber tube in his mouth,
draws air through the whole apparatus for about a minute, or rather
until the gas which is aspirated has lost the peculiar taste of CO2.
The india-rubber tube is then taken away, the stopper of B is
replaced, the whole apparatus once more wiped with a clean and dry
cloth, placed in the balance case for half an hour, and then again
weighed. On subtracting the weight after, from the weight before
decomposition, the weight of dry CO2 evolved is readily ascertained.

Determination of Fluorine.

Most chemists who have published analyses of bone have esti-
mated the amount of calcium fluoride indirectly, as follows. The
whole of the CO2 of the bone is supposed to exist as calcium carbonate,
(CaC03), then the whole of the phosphoric acid which does not exist
as magnesium phosphate is calculated in combination with calcium.
In a properly conducted analysis it will be found that on adding
together the calcium combined with carbonic and phosphoric acid
and subtracting the amount from the total weight of calcium found,
there is a small excess of lime left, which obviously must have existed
in some other form of combination. This is calculated as existing in
combination with fluorine.

Zalesky * determined the quantity of Fl directly, by a modification
of the method first suggested by Kobell. This consists in gently heat-
ing for a long period of time a weighed quantity of bone with strong
sulphuric acid and a weighed quantity of glass, the amount of silica
in which has been previously determined. In presence of the silica
and sulphuric acid all the fluorine contained in the bone unites to
form fluosilicic acid, SiFl4. The amount of fluorine present in the
bone is ascertained by determining the loss of weight which the glass
undergoes2.

Calculation of the results of the Analysis of the ash of bones.

The whole of the magnesium found is calculated as magnesium
phosphate (Mg8P208). The amount of phosphoric acid in this com-

1 Zalesky, Op. cit. p. 36.

2 Kobell's original paper was published in the Journ. f. prakt. Chemie, Vol. 162,
p. 385. The reader who desires to know the improvements introduced by Zalesky is
referred to the previously quoted memoir by this author.

CHAP. VII.J THE CONNECTIVE TISSUES. 289

pound is then deducted from the total weight of phosphoric acid,
the difference being calculated as calcium phosphate. The amount
of calcium in this compound is calculated and deducted from the
total amount of calcium found. Thus is found the calcium which
exists in other states of combination than as phosphate, viz. as
carbonate, chloride and fluoride. The whole of the carbon dioxide
found is assumed to have been derived from the decomposition
of calcium carbonate, so that the amount of the latter is easily
calculated. The chlorine found is calculated as present in calcium
chloride. By deducting then the calcium in combination with
phosphoric and carbonic acids and with chlorine from the total
quantity of calcium found, the amount of calcium present as CaFl2 is
obtained.

SECT. 5. TOOTH.

A tooth is a composite organ presenting for examination several
tissues; of these, three constitute the hard portion of the tooth, viz.
enamel, dentine, and crusta petrosa or cementum. In the interior
of the tooth is the so-called pulp cavity, which lodges the pulp,
which consists of a framework of connective tissue, to which are
distributed blood-vessels and nerves, and whence proceed processes
which are prolonged into the dentinal tubules.

Although two only of the hard tissues of tooth — viz. dentine and
crusta petrosa — belong to the group of connective tissues, the enamel,
which is a modified epithelial structure, will also, for reasons of
expediency, be considered in this place.

Dentine.

Dentine, or ivory, constitutes the chief part of the teeth. On
making a longitudinal section through a tooth it will be found
that the pulp cavity is bounded on all sides, except where the
cavities of the fangs open into it, by dentine ; the same tissue forms
the body of the crown (which is only covered by an external layer of
the harder enamel) and nearly the whole thickness of the fang or
fangs.

Dentine Dentine is distinctly mesoblastic in its origin, being

mesobiasticin formed through the agency of certain cells, termed
' odontoblasts,' which are modified connective-tissue
corpuscles arranged circumferentially over the surface of the papillary
protrusions which rise from the connective tissue of the buccal meso-
blast so as to meet and indent the downward dipping epiblastic
cells which give rise to the enamel organ.

G. 19

290 DENTINE. [BOOK i.

Microscopic Qn examining very thin sections of dentine it is

miCTotUchem£ found to consist of very fine tubes — the dentinal
cai reactions tubules, which are surrounded by a homogeneous ground
of dentine. substance; these tubules open internally in the pulp
cavity, from which they pass outwards, dividing and inter-communi-
cating. Sections made at right angles to their long axes exhibit the
tubes as minute round holes scattered through a translucent homo-
geneous matrix.

When teeth are placed in the acid solutions which have been
recommended for decalcifying bone (see p. 2740, the mineral matters
which give intense hardness to the hard tissues are dissolved, and it
then appears that around the lumen of the dentinal tubule there is a
structure which may be called the dentinal sheath, which, as it resists
the action of acids, obviously differs from the matrix more external to
it; the dentinal sheath possesses apparently the characters of yellow
elastic tissue. Under similar circumstances, the dentinal sheath may
occasionally be seen to contain a fine fibre, the dentinal fibre > which
is a process from the pulp, probably a process from the odontoblasts of
the pulp.

Relation of ^ we except the substance which constitutes the

dentine to dentinal sheaths and which is not affected by pro-

bone- longed boiling, nor by the action of acids or alkalies,

dentine has a composition which very closely resembles, or rather
which is almost identical with, that of bone; it consists, namely, of a
collagenous organic basis in which are deposited mineral matters
identical with those of bone.

The collagenous organic basis impregnated with salts is the result
of the activity of those connective-tissue cells which we term odonto-
blasts, just as the matrix of bone proper was originally formed through
the activity of those connective-tissue cells which we designated
osteoblasts. Though differing somewhat in arrangement and in texture,
the two tissues, dentine and bone, are, on developmental as well as on
chemical grounds, seen to be identical.

As Hoppe-Seyler has well shewn, the dentinal sheaths correspond to
the more internal portion of the ground substance of bone which may be
separated as a distinct investment bordering the lacunae, canaliculi and
Haversian canals (see p. 273).

water and Fresh dentine, when dried, loses about 10 per cent,

organic mat- of water ; the quantity of organic matter contained in
ter of dentine, ft varies between 26 and 28 per cent., on an average
being about 28 per cent.

constitu- Although a large number of analyses of tooth have

tion of the been made, we possess fewer absolutely reliable analyses
mineral mat- of dentine than of enamel. Dentine, like bone, cen-
ters of den- tains, as its chief mineral ingredients, calcium and
phosphoric acid ; the proportion of carbonic acid found
in its ash by most analysts is smaller than in bone. Hoppe-Seyler

CHAP. VII.] THE CONNECTIVE TISSUES. 291

is of opinion that in dentine, as in bone and enamel, the chief min-
eral ingredient is a definite compound (Ca10COs, 6(PO4)) of calcium
phosphate and carbonate, constituted like apatite (Ca10Fl2, 6(POJ).

Recalculating the results of an analysis of dentine of the ox made
by Aeby1, Hoppe-Seyler states its composition as follows :
Ca10C03, 6PO. in 100 parts 72'06

MgHP04 „ „ 075

Organic matter „ „ 2770

100-51

Numerous analyses of dentine by various chemists will be given
in the table exhibiting the general results of quantitative analyses of
tooth.

Enamel,

This tissue, the hardest in the body, as well as the richest in
mineral constituents, covers the crown or exposed surface of the tooth.

In its adult condition enamel is composed of polygonal (usually
hexagonal) prismatic columns which rest upon the dentine and
radiate out from it.

As has been already said, enamel is epiblastic in its origin, being
developed through the agency of the columnar epithelial cells of the
enamel organ, a structure produced by the proliferation and growing
downwards of the deeper epithelial cells of the oral mucous membrane.

When enamel is digested in acids, only a small quantity (2 — 6 per
cent.) of organic matter is left, which does not yield gelatin on boil-
ing. Enamel is thus seen, on chemical as well as on developmental
grounds, to differ from the connective tissues.

The mineral matters of enamel are essentially the same as those
of bone and dentine, and, according to Hoppe-Seyler, there are good
grounds to believe that they consist mainly of the same compound of
calcium phosphate and carbonate. Adult enamel contains a small
quantity of a fluoride, but Hoppe-Seyler failed to detect fluorine in
the growing enamel of the pig. It might be surmised that the
enamel consists of a mixture of apatite and bone earth, but there are
good reasons for believing that this is not the case.

The following formulae exhibit the relations between apatite and
the peculiar salt which Hoppe-Seyler believes to be the characteris-
tic mineral ingredient of bone, dentine and enamel.

Crystallized apatite .... Ca10Fl2, 6(PO4)
„ (another variety) Ga10Cl2, 6(P04)
The bone earth salt . . . Ca10CO3, 6(PO4)

In the annexed tables are given, firstly the results obtained by
Hoppe-Seyler from his analyses of enamel, and secondly the probable
amounts of the mineral compounds which he assumes to have been
present,

1 Aeby, Centrattlatt f. d. med. Wissenschaft. 1873. No. 7.

19—2

292

COMPOSITION OF TEETH.

[BOOK i.

Fossil
Elephant.

) 00 ,-•

b- co -^H cp <rq cq

co o b- 6 O O
to co

oo <n,O5 to i— i 1-1 co

^ T *9 T ^ 9* ^

•4jn o cb o o o cb
to co

i— I 00 *O 00 (M
O CO t-- r-i ,— (

CO O b- O O

»O CO

GO 00 — ' iO O

as c>i 10 ic cq

— o to o o
to co

10

(M CO O •*
00 -r^ to CO

co 6 «b o
to co

^ CD CO

00 O

to co

r— I O ""* tO CO tO tO

CO -^ GO to CO r-H O

^'00660 CT
to co

CO O ^O ^ ""^ ^
<^q co t- ^ cp ffi

<n o 4n o o o

to co

CO O

r^H CO

<N o

co

lO tO 00 t^
b- 17* O ^

tj- o cq o

•«T CO

IO O5 CO
60 I tp ^

o ' as o

O5

, jq

li

w

^
s <

GO 10 i>» <?q co

IQ O

Ci

co to to i— i

CO CO CM O

CO O

da

CO

CO O O
O5

CO ^ 10

Oi

O ^O GO
CO O rH

T-H 0

O <M CO iO

CO CO f r-l

4^ o <>i o

O (^ O

•-i . o o
op to ^

oo l~l S

O CO 1>- ^ Oi-

^ -q co co to

0^

CHAP. V[[.]

THE CONNECTIVE TISSUES.

293

Grusta Petrosa or Cement.

The cementum, or crusta petrosa, is found as a thin covering
over the dentine of the fangs and is developed from the periosteum

COMPARATIVE ANALYSIS OF TEETH AND PAETS OF TEETH OF VARIOUS
ANIMALS (VON BIBRA l).

Nature of body analysed.

Calcium
phosphate
withalittle
Calcium
fluoride.

Calcium
carbon-
ate.

Magne-
sium
phos-
phate.

Soluble
salts.

Total In-
organic
matter.

Organic
matter.

Enamel :

Woman, molar .

81-63

8-88

2'55

0-97

94-03

597

Man „ ...

8982

437

1-34

088

96-41

3-59

Wolf „ ...

87-82

1-21

1-10

0-83

90-96

9-04

Fox „ . . .

88-24

1-72

1-20

075

91-91

8-09

Lion, canine ....

8333

2-94

3-70

0-64

90-01

9-39

Bear „ ....

84-38

2-20

6-01

077

93-36

6;04

Seal „ ....

85-60

1-94

i-oo

0-63

89-17

10-83

Horse, molar ....

89-01

1-19

1-95

0-60

92-75

7-25

Ox, incisor ....

8377

7-00

1-32

0-61

9270

7-30

Dentine :

Woman, molar . . .

67:54

7-97

2-49

1-00

79-00

21-00

Man „ ...

66-72

3-30

1-08

0-83

71-99

28-00

Wolf „ ...

68-81

1-04

097

0-80

71-62

28-38

Fox „ ...

71-84

0-90

0-99

0-78

74-51

25-49

Lion „ ...

60-03

3-00

4-21

0-77

68-01

31-99

Bear „ ...

64-88

1-34

6-40

0-80

73-42

26-58

Seal „ ...

68-46

1-09

0-97

078

71-30

2870

Elephant (Indian), tusk

38-48

5-63

1201

070

56-82

43-18

» »

46-48

386

7'84

077

58-95

41-05

Dolphin

66-37

T84

1-36

099

70-56

20-44

Boar, tusk ....

60-00

251

6-43

0-43

69-37

30-63

Horse, molar . . .

61-28

6-08

175

074

69-85

30-15

Ox, incisor ....

5833

7-39

0-97

075

67-44

32-56

Goat, incisor ....

6304

283

1-70

0-93

68-50

31-50

Stag, molar ....

6351

399

372

058

71-80

28-20

Crocodile

53-47

6*33

1075

1-36

71-91

28-09

Crusta Petrosa :

Dolphin ....

69-42

1-79

1-47

093

73-61

26-39

Ox, incisor ....

58-00

7-22

099

0-73

66-94

33-06

Crocodile

5339

6-29

9'99

1-42

7109

28-91

Whole Tooth :

Saw-fish

61-99

3-64

1-70

1-81

6914

30-86

Pike

63-98

254

073

0-97

68-22

3178

Black-fish

59-94

9-01

2-00

177

7272

27'28

Plaice . . .

57-20

1-34

0-88

1-82

61-24

3876

1 Reprinted from Watts' Dictionary, Vol. v., p. 706.

294

COMPOSITION OF FOSSIL TEETH. [BOOK I. CH. VI [.

which covers them; histologically it is found to be composed of
true osseous tissue, presentiug lamellae, lacunae and canaliculi, per-
forating fibres, and occasionally Haversian canals ; chemical analysis
reveals no difference between it and bone proper.

COMPOSITION OF VARIOUS FOSSIL TEETH (VON BIBEA1).

Calcium
phos-
phate.

Calcium
fluoride.

Calcium
carbon-
ate.

Calcium
sul-
phate.

Magne-
sium
phos-

T)llclt6«

Silica,
iron, alu-
mine,
calcium

Organic
matter.

chloride.

Rhinoceros Tichorhinus

Upper molar, Enamel

83-11

414

7-66

0.95

073

0-24

3-17

„ Dentine

54-65

3-09

12-80

0-80

5-63

23-03

Elephas primigenius, molar

62-83

4-15

14-90

211

0-32

15-60

t) a »

68-43

3-72

15-40

1-34

1-91

9-14

Cave Bear, molar

64-03

2-51

1-46

8-25

0-30

23-45

Fish (Acrodus)

90-01

1-80

0-52

5-50

2-17

Fish from chalk

85-01

911

0-45

14-89

0-54

Analysis of Dental Tissues.

This is conducted according to the methods which have been
described for the analysis of bone.

1 Eeprinted from Watts' Dictionary, Vol. v.r p. 706.

CHAPTEE VIII.

EPITHELIAL TISSUES OR EPITHELIUM. KERATIN.
CHITIN. PIGMENTS DEPOSITED IN THE EPI-
THELIAL STRUCTURES. CERTAIN OTHER ANIMAL
PIGMENTS.

BY the term epithelium is designated a tissue, composed entirely
of cells, which covers the whole external surface of the body, and lines
the cavities which open externally. The term is generally held to
include also the tissue composed of a single layer of tesselated or
tile-like cells which lines the arteries, capillaries and veins, the serous
sacs and the lymphatics. This tissue to which the terms endothelium
or epithelioid tissue are more properly applied, as indicating that it
differs from epithelium in its development, in its characters, and in
the uses which it subserves, will be considered in this work in discussing
the chemistry of the so-called ductless glands and the lymphatics.

Confining our attention to epithelium proper we might classify
it in various ways : firstly, according to the form and arrangement of
the cells of which it is composed : secondly, according to the regions
in which it occurs : thirdly, according to the mode in which it is
developed : fourthly, according to the chemical characters which it
possesses ; we shall not, however, strictly follow any of these modes
of classification. There is no other tissue of which the individual
anatomical elements exhibit such marked differences in the chemical
operations of which they are the seat.

Speaking broadly we may, however, say that the epithelium
covering the external surface of the body is composed of cells which
are, even in their most active stages, the seat of but slow and
unimportant chemical changes, whilst a large number of them cease
to be the seat of any material exchanges whatever, or to manifest
any phenomena which characterize them as living, long before they
cease to form part of the Jiving body.

The function of such epithelium — and we are referring to that
which composes the cuticle and its appendages — is in the strictest
sense tegumentary. This epithelium possesses two characters which
may be taken together. Firstly, it is entirely derived from the
external layer of the blastoderm or epiblast. Secondly, however

296 KERATIN-PRODUCING EPITHELIAL TISSUES. [BOOK I.

different the arrangement of the cells and the physical characters of
the tissue which they compose, the main product which characterizes
them is an undefined horny substance to which the term Keratin
has been applied.

The epithelium, on the other hand, which covers the mucous
membranes and lines the interior of secreting glands, is composed of
cells, whose protoplasm is the seat of the most active and remarkable
chemical operations, tending to separate, from the blood, constituents
of which it has no longer need, or to build up, at the expense of
certain of those constituents, new bodies which are to serve important
functions in the organism.

This epithelium is mainly derived from the hypoblast, though
in some cases it takes its origin in the epiblast (epithelium of mouth
and salivary glands), in others from the mesoblast (certain portions of
the epithelium of genito-urinary tract). In short, the epithelium of
the mucous membranes is possessed of diverse chemical attributes
and is developed in several ways; it does not therefore possess any
common characters which permit of a general description.

We shall, therefore, in this chapter, confine ourselves, in the first
place, to a consideration of the chemistry of the keratin-forming
epiblastic tissues, postponing the exposition of the chemistry of other
epithelial tissues to future sections of this work, where they will be
treated of in relation to the organs in which they occur, and the
functions which they specially subserve.

SECT. 1. EPIBLASTIC KERATIN-PRODUCING EPITHELIAL TISSUES.
THE HORNY SUBSTANCE OF CUTICLE, NAILS, HORN, HAIR, AND
FEATHERS.

The cuticle or epidermis is composed of many layers
Structure of ~ .,, ,. , „ s . , T ,1 i . J i •

Epidermis. °* epithelial cells which overlie the derma or true skin.

The cells of the most superficial layers present the
appearance of distorted, shapeless, hardened scales; those of the
deeper layer are more or less spheroidal, soft, and present a well-
marked nucleus.

The most superficial cells, which are horny throughout, are
unacted upon by acetic acid ; this reagent renders the deeper cells
more transparent and their nucleus more evident.

The cells are connected together by a cementing substance which,
like the cementing substance of the connective tissue, is dissolved by
alkalies. In order to dissolve this connecting substance and effect
the dissociation of the epidermal cells, maceration in cold, or even
warm, solutions of caustic potash or soda should be had recourse to.
These reagents not only dissociate the cells but, in the case of the
more superficial cells, render their structure more evident.

The caustic alkali, at first, leaves the nucleus and the cell sub-
stance intact, merely causing the latter to swell and become more

CHAP. VIII.] EPITHELIAL TISSUES, &C. 297

transparent than previously ; subsequently, the nucleus may disap-
pear, leaving the cell body. In some cases, the separate anatomical
elements are best seen if after the action of alkali the tissue is placed
in water. Cold concentrated sulphuric acid also brings out the cells
of the epidermis, effecting to a certain extent their dissociation.
When heated, it dissolves the deepest cells (those of the rete mucosum)
but leaves undissolved the more superficial, in which the cell protoplasm,
has undergone conversion into horny substance.

structure of The Nails possess essentially the same structure as

Nails. the epidermis, and their cells may, like those of the

latter structure, be divided into an upper horny layer
and a lower softer stratum. The action of caustic potash or soda on
the cells of nail is similar to that exerted by these reagents on the
cells of the cuticle.

structure of Horn is constituted exactly as nail ; in the deeper

Horn. layers of cells pigment is sometimes present, as it is in

the rete Malpighii of the epidermis.

structure of Hoof is composed of compressed epithelial cells

Hoof. arranged concentrically around canals which run from

above downwards. The individual cells are made apparent by treat-
ment with solutions of caustic alkalies.

structure of Hairs have a more complex structure than the

Hair. epidermal tissues yet enumerated, and for a description

we must refer to treatises on Histology. It may be mentioned
however that the stem of the hair is seen to be covered by layers of
imbricated cells which are evidently modified epidermic epithelial
cells ; within these is the so-called fibrous substance which makes
up the greater part of the stem and which may be resolved into
elongated scales; and in the very centre of the hair is the medulla,
or pith in which sometimes air spaces are seen, sometimes cells which
are filled with fatty and pigmentary particles. The lower part of the
stem or shaft of the hair dips into the so-called hair-follicle, where
it is attached to, or rather grows upon, the papilla ; the imbricated
scales which cover the root of the hair are continuous with the inner-
most layer of cells of the epidermic lining of the hair-follicle.

Horny Substance or Keratin,

By the term Keratin is understood the organic substance, or
perhaps the mixture of organic substances, left as an insoluble residue
when cuticle, horn, nails, hairs, feathers, &a, are successively boiled in
ether, alcohol, water, and dilute acids. This insoluble residue retains
the form of the tissues from which it is prepared; it is little
affected by boiling with water at ordinary pressure, but is dissolved
when subjected to the prolonged action of water under pressure (as

298 HORNY MATTER. [BOOK I.

in sealed glass tubes heated to 150° — 200° C.), yielding a turbid
solution which furnishes on evaporation a dry mixture insoluble in
water. Keratin swells, and subsequently is dissolved by boiling in
alkalies, and on the addition of acids to the alkaline solutions
sulphuretted hydrogen is given off.

Horny substances swell when immersed in dilute acetic acid, and
are in great part dissolved by boiling glacial acetic acid.

When boiled with dilute sulphuric acid, Keratin yields aspartic
acid, volatile fatty acids, leucine, and tyrosine. Nitric acid dissolves
it, and oxalic acid is formed as an ultimate product.

When strongly heated, horny substances burn, evolving the charac-
teristic smell of burned feathers.

Results of Though we cannot obtain by any known process a

ultimate ana- definite substance Keratin, which can be considered as
lyses of Horny pure, yet the horny tissues present a very close resem-
Tissues. blance in the proportion in which their elements are

contained, as will be seen by perusing the analyses here appended1.

Hair, Nails, Cow's horn, Horse's hoof,

(v. Laer). (Mulder). (Tilanus). (Mulder).

0 50-60 51-00 51-03 51'41

H 6-36 6-94 6'80 6'96

N 1714 17-51 16-24 17'46

O 20-85 2175 22-51 19'49

S 5-00 2-80 3-42 4'23

The sulphur which is contained in these horny substances is very
loosely combined; it varies also remarkably in quantity in various
samples of the same tissue, as for example in human hair between
3 and 8" 23 per cent. When heated with barium hydrate and water
in sealed glass tubes, nearly the whole of the sulphur is obtained in
the form of Ba(SH)2 (Hoppe-Seyler2).

Inorganic Matters contained in the Horny Tissues.

All these tissues contain inorganic matters. In the nails the ash
is said to be specially rich in calcium phosphate. Hair contains from
0'5 to 7 per cent, of mineral constituents, and the latter contain
alkaline sulphates, iron and silica (40 per cent, of the ash). The pro-
portion of iron in the hair varies, and it has been stated that it is
larger in dark than fair hair; but this statement must be received
with some doubt.

The composition of the mineral matters of feathers varies, accord-
ing to von Bibra, with the nature of the food upon which birds feed;
thus, the silica may vary between 27 and 40 per cent, of the total
mineral matters.

1 Hoppe-Seyler, Physiologische Chsmie, 1 Theil, p. 90.

2 Hoppe-Seyler, Op. cit., p. 91.

CHAP. VIII.] EPITHELIAL TISSUES, &C. 299

SECT. 2. TISSUES WHICH YIELD CHITIN, SPONGIN, TUNICIN,
AND HYALIN.

The tissues of many groups of invertebrate animals contain
certain of the proximate principles which have been enumerated and
described as obtained from the tissues of man and the higher verte-
brates. Thus mucin is present in organisms low in the scale; as
we ascend, we find chondrin-yielding tissues, and in the Cephalopoda
tissues which yield gelatin1 when boiled.

In certain invertebrates we find, however, that the tissues contain
substances which do not occur in vertebrates. Amongst these are
the substances now to be considered, viz. Chitin, Conchiolin, Spongin,
Tunicin, and Hyalin.

Chitin.

Distribu- Chitin usually occurs throughout Invertebrates in the

in°tne Animal f°rm °f an investment to the outermost cellular layer
Kingdom. or ectoderm. The exceptions real and apparent to this

statement are noted in the following table of its distribution, which
however must be accepted as only approximately accurate, in the
absence of full chemical investigation, in any case except that of the
Arthropoda2.

Protozoa. Membrane of all "loricate" forms, cuticle of Infusoria,
&c. Oesophageal lining of toothed Ciliata (Nassula, Prorodon).
Central capsule of Radio! aria. Cyst wall of all encysted forms.

Coelenterata. Membrane of fertilized ovum. Mesodermal (?)
skeleton of Hydro-medusae (Velella).

Vermes. Membrane of ovum. Cuticle in all cases, including
the ectocyst of Polyzoa, and cuticular appendages, such as the
setae of Annelida. Oesophageal armature of Rotifera and some
Annelida. Mesodermal branchial skeleton of Balanoglossus.

Echinodermata. The presence of chitin is not indicated in this
group.

Mollusca. Membrane of ovum. Setae of larval Brachiopoda.
Byssus, shell-ligament and shell (in many cases, if not universally, the
organic base of the shell is composed not of chitin but conchiolin,
q. v.). Rings and hooks of suckers of Cephalopoda. Upper lip and
jaws of Cephalopoda and Gasteropoda. Radula of odontophore.
Mesodermal branchial skeleton of Lamellibranchiata.

1 Hoppe-Seyler, "Ueber Unterschiede im chemischen Ban und der Verdaunng
hoherer und niederer Thiere." Pfliiger's Archiv, Vol. xiv. p. 395—400. "Ueber das
Vorkommen von leimgebendem Gewebe bei Avertebraten." Med. Chem. Untersuchung .
p. 580.

2 It would seem that in many cases a chitinous composition has been ascribed to a
structure solely on the ground of its insolubility in caustic alkalies and dilute acids, or
even in only one of these two classes of reagents.

300 CHITIN. [BOOK r.

Arthropoda. Membrane of ovum. Cuticle with its appendages
external and internal (setae, apodemata, large tracts of alimentary
canal, gizzard when present, all excretory ducts, tracheae of Onycho-
phora, Arachnida, Myriapoda and Insecta).

Chitin is frequently found impregnated with calcareous matter, as
in Crustacea, or with silica, as in the radula of the higher Mollusca.

Pre aration wing~cases °f the cockchafer are boiled in

dilute solution of caustic soda until they have become
colourless; they are then washed with water, dilute acids, and
lastly with boiling alcohol and ether (Hoppe-Seyler).

From the shell of the crab or lobster it is obtained by the same
treatment, after previous digestion in hydrochloric acid, so as to
dissolve the earthy matters deposited in the chitinous tissue. The
chitin thus prepared may be dissolved in cold pure concentrated
hydrochloric acid, and the solution precipitated by the addition of
a large excess of water.

Pro erties Chitin is a colourless, amorphous body, which retains,

when prepared by the first of the above-mentioned
methods, the form of the parts composed of it ; when prepared by the
second method it appears as an amorphous gelatinous body. It is
insoluble in water, alcohol, ether, acetic acid, in dilute mineral acids,
and in solutions of the alkalies. It is dissolved by concentrated
mineral acids. Chitin resists in a very remarkable manner the action
'of^alkalies, and can be boiled in their concentrated solutions for long
periods of time without undergoing decomposition.

Elementary Chitin has been subjected to analysis by many

composition observers. The following is the mean of twelve analyses
and formula. ma(je by Ledderhose1, who has investigated the con-
stitution of chitin under the direction of Professor Hoppe-Seyler.

Carbon in 100 parts 45'69

Hydrogen „ „ 6'42

Nitrogen „ „ 7'00

Oxygen „ „ 40'89

Ledderhose * ascribes to Chitin the formula C15H26Na010.

Berthelot pointed out2 that when chitin is dissolved

Products of -in concentrated sulphuric acid it yields a fermentable

won"1' sugar; this statement has been disproved. The re-

searches of Ledderhose carried out under the direction

of Professors Hoppe-Seyler and Baumann have thrown great light

1 Ledderhose, "Ueber Chitin und seine Spaltungsprodukte." Zeitschrift fur
physiol. Chem. Vol. n. (1878), p. 213. Ledderhose: "Ueber Glykosamin. " Ibid.
Vol. iv. (1880), p. 139.

a Berthelot, Comptcs Rendus, XLVII. 227t

CHAP. VII I.] EPITHELIAL TISSUES, &C. 301

upon the decomposition, and have shewn that when heated with
acids chitin combines with the elements of water and splits up into a
nitrogenous body glycosamine and into acetic acid, thus : —

2 (CUIW)10) + 6H20 = 4(C6H13N05) + 3(C2H402).

'«'/.

Glycosamine, 06H13N05.

Prepara- Chitin is soluble in cold concentrated HC1, and the

tton. solution remains colourless when exposed to air, the dis-

solved body not undergoing decomposition and being thrown down
unchanged by the addition of water.

When the solution is boiled it becomes black, in consequence of
a decomposition which is completed in about an hour. On evaporation,
impure hydrochlorate of glycosamine is obtained, and is purified by re-
crystallizing repeatedly.

The amount of this compound formed amounts to 70 or 75 p.c. of the
weight of the chitin dissolved.

Properties. The hydrochlorate of glycosamine is easily soluble in
water, but soluble with difficulty in alcohol ; its solution has a sweet taste and
an acid reaction. It reduces alkaline solutions of cupric and silver salts,
and its solutions react like solutions of glucose when boiled with caustic
alkalies. It is dextro-rotatory (a) D = + 70°- 6. The pure base (prepared by
the action of barium hydrate on the sulphate of glycosamine) crystallizes
from alcohol in the form of needles. It is not fermentable.

Probable Ledderhose considers glycosamine to be an amido-deriva-

constitution. tive of grape sugar (dextrose), thus :

COH . (CHOH)4 . CH2OH COH . (CHOH)4CH2NH2

Dextrose. Glycosamine.

Conchiolin.

The organic matter of the shells of Mussels and Snails was
formerly supposed to be identical with chitin; this is not, however,
the case, and to the substance the name of Conchiolin has been
applied.

Prepara- The shells of mussels are macerated in dilute hydro-

tion- chloric acid ; then boiled in aqueous solutions of the

caustic alkalies.

Elementary The differences in composition between Conchiolin

composition. and chitin are shewn by the two analyses quoted below:

Conchiolin. Chitin.

Carbon .... 507 . . . 46'32

Hydrogen . . . . 6*5 . . . 6 '40

Nitrogen . . . . 167 . . . 614

Oxygen .... 261 . . . 4114

X

302 SPONGIN. HYALIN. TUNICIN. [BOOK I.

Reactions Conchiolin is insoluble in dilute acids and in alka-

of Conchiolin. line leys. It is soluble in hot concentrated hydrochloric
acid. When boiled with dilute sulphuric acid it furnishes leucine and
110 sugar-like body. By the two last characters it is as clearly dis-
tinguished from chitin, as by its much smaller amount of nitrogen.

Spongln.

When sponge is boiled with dilute hydrochloric acid, then with caustic
soda, water, ether and alcohol, there is left a body to which the name of
Spongin has been given.

This body (if a definite body it be), so far as it has been investigated,
appears to have the following composition :

Carbon 47*44

Hydrogen 6 -30

Nitrogen 1615

Oxygen 30-11

When boiled with water it yields no gelatin.

When boiled with dilute sulphuric acid it yields leucine and glycocine,
but no tyrosine.

Hyalin.

This term is applied to the principal constituent of the walls of hydatid
cysts.

Prepara- Hydatid cysts, emptied of their contents, are boiled in

tion. water, then in alcohol and ether. The residual matter is

soluble when heated in water (under pressure) at 150°C. The solution is
precipitated by alcohol, neutral and basic lead acetate, and by mercuric
nitrate.

Elementary The composition is said to vary according as the substance

composition. {s prepared from old or young cysts.

Composition of hyalin (Liicke1).

(1) (2)

From young cysts. From old cysts.

C ' 44-1 45-3

H 67 6-5

N 4-5 5-2

O 44-7 430

Products of When dissolved in strong sulphuric acid or boiled in

decomposi- dilute sulphuric acid, hyalin is said to yield 50 p.c. of its
tion- weight of a dextrogyrous sugar, susceptible of the alco-

holic fermentation.

Tunicin or Animal Cellulose, C6H1005.

This body, closely resembling, if not identical with, the cellulose so
widely distributed throughout the vegetable kingdom, occurs in the
mantle of the Tunicata.

1 Liicke, Virchow's Arcliiv, Vol. xix. p. 189.

CHAP. VIII.] EPITHELIAL TISSUES, &C. 303

Prepara- The cartilaginous investment of Ascidians, e.g. the

tion. "mantle" of Phallusia and Cynthia and the external coat of

Salpa consist mainly of Tunicin and may be employed in its preparation.

These structures are digested in hot water, then for a short time in
dilute acids and alkalies, lastly in alcohol and ether ; the residue, which
preserves the original form of the structures, consists of tuiiicin.

Tunicin is by the action of acids converted entirely into a reducing
sugar susceptible of the alcoholic fermentation (dextrose?). It is maintained
by Berthelot1 that tunicin presents certain differences from ordinary
cellulose, as, for instance, that it is coloured yellow by iodine and is less
affected by certain reagents.

SECT. 3. ON CERTAIN COLOURING MATTERS OF THE EPITHELIAL
TISSUES OF VERTEBRATES.

Brown and black Pigments. Melanin.

The cells of the rete Malpighii of the human skin often contain
granules of a black pigment; this is especially the case in the skin
of the negro, which owes its colour to these pigment-bearing cells.
A similar pigment is found in the hexagonal epithelial cells which
constitutes the most external layer of the retina, and which used
formerly to be considered as belonging to the choroid; also in the
connective tissue cells of the outer layer of the choroid. In the
bronchial lymphatic glands, of adults and aged persons, in the lung
tissue and in melanotic tumours, similar brown or black pigments are
discovered, which are all included under the name of Melanin, though
it is certain that the substance obtained from these various sources
does not present an uniform composition; in all probability, however,
all these colouring matters derive from the decomposition of haemo-
globin. The formation of such a black pigment has actually been
traced in the interior of the red blood-corpuscles, in cases of per-
nicious intermittent fevers (see p. 163).

Characters Melanin occurs in the form of minute amorphous

and reaction granules which when suspended in water exhibit
of Melanin. Brownian movements. It is soluble in ether, alcohol,
water and acids. When boiled with solution of caustic potash the
black colouring matter is slowly and imperfectly dissolved, a brown
liquid being formed, which is discolourized by chlorine.

In the lung tissue, particles of carbon sometimes occur ; these are some-
times in a finely granular condition, though occasionally they present the
appearance of minute fragments of coal. The latter are distinguished from
melanin by their complete insolubility in boiling caustic potash, in
boiling sulphuric acid, and when boiled in strong hydrochloric acid and
potassium chlorate.

1 Berthelot, Ann. de Chim. et de Phys., Vol. LVI. p. 149.

304 TURACIN. [BOOK i.

Percentage The analyses which have been made of the various

composition pigmentary matters included under the term Melanin
of Melanin. have led to widely discordant results. The carbon in
100 parts has varied between 51*7 and 58'3; the H between 4'02 and
5'09; the N between 71 and 13'8; the 0 between 22'03 and 35'44 l.

Pigments of the Feathers of Birds.

The brilliant colours of the plumage of birds is due in part to
the optical characters of the surface of the feathers (interference-
colours) : in part to the presence, within the feathers, of colouring
matters, which may usually be extracted from them by alcohol, ether,
or hot acetic acid, and which, as a rule, are very unstable, becoming
decolourized by exposure to air.

These colouring matters have hitherto not been subjected to a
thorough chemical investigation, with the exception of the one to be
described in the ensuing paragraph.

Turacin.

In various species of birds belonging to the family Musophagidae
and which, from the nature of their food, are designated Plaintain-
eaters, the primary and secondary pinion-feathers, are more or less of
a crimson colour. The colour is due to a pigment which has been
separated and analysed by Professor Church, who has applied to
it the name Turacin, from Tour aeon, the name by which the Plaintain-
eater is designated by the natives on the shores of the Gambia.

The barbs constituting the red part of the web are
parating° Tu- stripped from the shaft of the feathers, placed in a
racin. beaker, and washed with ether, then with alcohol.

They are then dried, by pressure between folds of
filtering paper, and placed in a very dilute cold solution of pure
caustic soda, a solution containing one part of soda in a thousand
of distilled water being quite strong enough. The crimson pigment
is soon dissolved ; its solution is then poured into dilute hydrochloric
acid (1 of acid to 4 of water), when the red colouring matter is
precipitated. It is then washed, first with water, until all acid
reaction is removed, and then in alcohol and ether, and dried.

Properties Occurs in scales which have a deep violet-purple

of Turacin. colour by reflected light, and a crimson tint when seen
in small fragments by transmitted light.

It has not yet been obtained in a crystalline form. It is very
slightly soluble in pure water, giving a pale rose-pink solution. It is
not soluble in alcohol or ether. It is insoluble in acid, but soluble in
alkaline liquids*

Spectrum Turacin and the feathers containing it possess a

of Turacin. spectrum which is almost identical with that of oxy-

1 See a paper by Hodgkinson and Sorby entitled " Pigmentum Nigrum, the black
colouring matter contained in hair and feathers." Jcurn. Chem. Soc. 1877, p. 427.

CHAP. VIII.] EPITHELIAL TISSUES, &c. 305

haemoglobin ; there is, namely, a shading of the blue end of the
spectrum and two absorption bands between D and E ; no change
is, however, produced by the addition of reducing solutions.

The author has carefully measured the positions of the bands of Turacin
(from Turacus persa) and he finds that the centre of the band corresponding
to that designated a in the spectrum of oxy-haemoglobin has a wave-length
of 578 ; the band in the green has a wave-length of 538 — 540.

composi- The remarkable feature of this red-colouring matter

"ura" is the constant presence of copper in it.

Church has made many analyses of several specimens of this
body, and these have yielded concordant results. From these analyses
Church has deduced the empirical formula CwH53CuN5Oi9, which
demands the following percentages: —

S;

Cu

100-00 100-00

The quantity of Turacin in a single bird does not exceed two or
three grains1.

Theory.

Experiment
(Mean).

54-87

54-63

5-12

5-22

5-81

5*90

6-39

6*38

27'81

27-87

o

SECT. 4. CERTAIN OTHER COLOURING MATTERS OCCURRING IN
THE ANIMAL KINGDOM.

The study of animal colouring matters apart from those found in
the blood has hitherto, with few exceptions, met with but little
attention. While u number have been examined spectroscopically
with the results given below, but few have been chemically analysed
with anything like thoroughness2. -

1 The above account is drawn from Professor Church's Memoir entitled "Researches
on Turacin, an animal pigment containing copper." Philosophical Transactions,
Vol. CLIX., Part ii. (1870), pp. 627—636.

2 The chief papers on this subject are the following: — E. Ray Lankester: "Report
on the Spectroscopic Examination of certain Animal Substances," British Association
Reports, 1869. "Abstract of a Report on the Spectroscopic Examination of certain
Animal Substances, presented to the Brit. Assoc. at Exeter, 1869," Journ. of Anat. and
Phys., Nov., 1869, p. 119. "On Blue Stentorin, the colouring matter of Stentor
coeruleus," Quart. Journ. of Micros. Sc., April, 1873. "Preliminary notice of some
observations with the spectroscope on Animal Substances," Joum. of Anat. and Phys.,
1868, p. 114. H. C. Sorby: "On the colouring matters derived from the decomposition
of some minute organisms," Month. Micro. Journ., Vol. vi. (1871), p. 124. "On the
colouring matter of some Aphides," Quart. Journ. of Micr. Sc., 1871, p. 352. "On the
colouring matter of Spongilla ftuviatilis," Quart. Journ. of Micros. Sc., 1871, p. 352.
"On the colouring matter of Bonellia viridis," ibid., p. 166. H. N. Moseley : "On
Actiniochrome," Quart. Journ. Micros. Sc., 1873, p. 143. "On colouring matters of
various animals," ibid., 1877, p. 1. This is a most important paper, giving the fullest
account of the Spectroscopic examination of a very large number of pigments.

G. 20

306

COLOURING MATTERS.

BOOK I.

The pigments, to be referred to in this section, occur either dif-
fused through the tissues, as in many marine animals, or in the form
of granules contained in certain cells or layer of cells, usually dermal
or subdermal, sometimes deeper in the mesoderm, very rarely in the
endoderm. Such granular mesodermal deposits are frequent in
cephalopoda, fishes, amphibia and even lizards. The Chlorophylls and
associated 'vegetable' pigments, when present in animals, are always
in granules, whether in the striae of the myophane of Infusoria, the
tissues generally of Spongilla, the sub-muscular mesoderm of Convo-
luta, or the endoderm of Hydra viridis.

The following is a list of such pigments as have hitherto been described,
arranged in the order of the animals yielding them1.

Sub-kingdom. Colouring Matters.

Protozoa. Chlorophyll. Blue Stentorin.

Porifera. Chlorophyll. Various other pigments shewing no bands.

Coelenterata. Chlorophyll, &c. in Hydra viridis and in Anthea Cereus,

var. smaragdina.

Actiniochrome in Bunodes crassicomis.
Polyporythrin in many simple Anthozoa and some

Hydroids.
Two distinct pigments with characteristic absorption

bands in Adamsia sp.

A red pigment with one band in Coenopsammia.
Other pigments yielding no bands.

Echinodermata. Purple Pentacrinin in many species of Pentacrinus.
Red Pentacrinin in a species from Meangis Is.
Antedonin from an Antedon and a deep-sea Holo-

thurian.

Hoplacanthinin from Hoplacanthus sp.
These four pigments all have absorption spectra with
definite bands, the other pigments obtained from
animals belonging to this class yield no bands.
Verines. Chlorophyll (?) in Convoluta2.

Bonellein in Bonellia viridis.
Other pigments yielding no bands, including a blue

one, reddened by acids, in a Rhyncodemus sp.
Crustacea. Chlorophyll in Telotea viridis.

Crustaceorubrin in many deep-sea Decapods ; in a Pan-
darus infesting Carcharius brachyurus ; in surface
Entomostraca.

Other pigments yielding no definite absorption spectra.
Insecta. Cochineal from Kermes cacti.

Aphidein from an Aphis on the apple.
Lac-dye from Coccus Laccae.

Other pigments not yet examined, or yielding no
definite spectra.

1 This list is compiled chiefly from Moseley's previously-quoted paper (Quart. Journ.
of Micr. Sc., 1877, p. 1).

2 Geddes, "Physiology and Histology of Convoluta Schultzii." Proceedings of
tlie Royal Society, Vol. xxvm. p. 449.

CHAP. VIII.] EPITHELIAL TISSUES, &C. 307

Mollusca. Aplysio-purpurin from Aplysia * and Doris.

Janthinin in Janthina.
Tyrian purple2 in several species of Murex and

Purpura.

Other pigments yielding no bands.

Vertebrata. In addition to colouring matters referred to in other

parts of this book — a bluish-green pigment with
a single band, extending from B to beyond 0,
destroyed by heat, acids and alkalies ; found in
Odax, 3 spp. and in Labrichthys Richardsonii 3.

A short account of the chief characters of certain of the above-
named colouring matters will now be given.

Blue Stentorin4.

This blue colouring matter obtained from Stentor coeruleus is
characterized by a spectrum with two absorption bands ; of these the
darker is on the red side of C; a second lighter band, between
D and E, occupies approximately the space intervening between the
middle of the a band of oxy-haemoglobin and the /3 band of the same
body. The colour is unaffected by acetic, hydrochloric, and sulphuric
acids; caustic potash causes the colour to become darker, the band
between D and E disappears, and that between B and C becomes
darker and is shifted somewhat nearer towards B.

Actiniochrome5.

This is a red colouring matter obtained by Moseley from
some specimens of Bunodes crassicornis. It possesses an absorption
band having approximately the position of the band a of oxy-
haemoglobin.

Bonellein*.

This is a colouring matter obtained by Sorby from Bonellia
viridis. According to Sorby it resembles blue chlorophyll in many
respects, but differs in only being temporarily altered by acids, the
original colour returning on neutralization. It occurs in fine granules
in the epidermal protoplasm, and is insoluble in water, soluble in
alcohol, ether and carbon disulphide. The following are the wave-
lengths of the centres of the absorption bands of Bonellein (expressed
in millionths of a millimetre) :

1. Alcohol solutions, alkaline or neutral
662, 636, 611, 587, 520, 490.

1 An Italian chemist has asserted that an aniline base is present in Aplysia,
(Moseley, op. cit., p. 13).

2 Lacaze-Duthiers, "Memoire sur la Pourpre," Annales des Sciences Naturelles, ZooL
Ser. iv., Vol. xn. pp. 5—84.

3 George Francis, Nature, Vol. xn. p. 167.

4 Lankester, Op. cit., (see foot-note to p. 305).
3 Moseley, Op. cit., (see foot-note to p. 305).

6 Sorby, Op. cit.

20—2

308 BONELLEIN. CARMINIC ACID. [BOOK I.

2. Alcohol solutions, slightly acidulated

636, 611, 588, 565, 543, 522, 492.
31 „ strongly acid

617, 590, 565, 552, 517.
Solutions of Bonellein are fluorescent.

Carminic acid.

The female Cochineal insect (Coccus cacti) contains from 26 —
50 p. c. of a splendid red colouring matter, to which the name of
Carminic acid is given, and from which commercial carmine is pre-
pared. This colouring matter is found in other species of Coccus,
and occurs in the vegetable kingdom, being found in the blossoms of
Monarda didyma.

Mode of One part of powdered cochineal is boiled in 40 parts

preparation. of water for half an hour; the solution is then decanted,
precipitated with lead acetate, care being taken to avoid an excess of
the precipitant ; the precipitate is washed with boiling water so long
as the washings give a precipitate with solutions of mercuric chloride ;
it is then decomposed by sulphuretted hydrogen; the filtrate from
the precipitate of lead sulphide is evaporated to dryness at a very low
temperature, and the residue is extracted with alcohol, which dissolves
the carminic acid.

Composition Carminic acid, C17H18O10, is an amorphous red powder

and proper- easily soluble in water, and alcohol, and in hydrochloric
ties. and sulphuric acids. It forms no salts of constant com-

position. The ammonium salt exhibits two absorption bands between
D and E nearer E than those of oxy-haemoglobin ; these bands are
more closely approximated than those of oxy-haemoglobin and have
less distinct edges. Aqueous and alcoholic solutions of cochineal
on the other hand absorb all but the red rays.

The author has determined the position of the centres of the two bands
of carminate of ammonia. The centre of the band which corresponds to
that designated as a in the spectrum of oxy-haemoglobin is approxi-
mately 530, the centre of the band corresponding to the band /? of
oxy-haemoglobin is approximately 570. An ammoniacal solution of carmine
ulso exhibits two absorption bands, of which the centres are respectively 570
and 528.

A solution of picro- carmine exhibits a spectrum which at first sight very
closely resembles that of oxy-haemoglobin. It will be observed, however,
that the band near the red (a) is less dark than the one in the green, the
centres being respectively approximately 565 and 520, and there is a third
band in the blue, very close to its junction with the green.

Carminic When boiled with dilute acids carminic acid

acid a Giuco- combines with the elements of water to form an
side- unfermentable sugar, which is optically inactive, and a

new pigment carmine-red (CnH12O7) :

C,,H180W + 2H,0 = C6H1005 + CUH1207°.

CHAP. VIII.] EPITHELIAL TISSUES, &C. 309

Tyrian Purple1.

This colouring matter, which was employed in remote antiquity
to dye the robes of royalty, and which even in the luxurious days of
Imperial Rome retained its position as the dye of greatest beauty
and value, is derived from the secretion of a glandular organ which is
situated at the lower part of the mantle, between the gill and the
rectum, of various species of Murex and Purpura. The secretion
when first poured out is colourless or yellowish, but when exposed to
the light, especially if it be first diluted with water, it assumes first a
bluish-green, then a red and lastly a purple-violet colour, at the
same time emitting a strong alliaceous smell. This change occurs
spontaneously in the case of Murex trunculus even though the juice
be kept in the dark, in sunlight it occurs in a few minutes. In
Murex brandaris the colour is produced only in the light and more
slowly. The dried juice, when powdered, appears red ; it is insoluble
in water, alcohol and ether, in dilute acids and cold alkaline leys.

Punicin tlie Schunck2, to whose investigations we owe so much of our

colouring knowledge of certain animal colouring matters, has examined

matter ob- tjie brig}jt purple colouring matter obtained by exposing to

action oHight light the secretion of the purpurogenous gland of Purpura
from the lapillus. This colouring matter is insoluble in water,

Chromogen of alcohol, and ether ; it is slightly soluble in boiling benzol,
'urpura an(j jn boiling glacial acetic acid. It dissolves entirely and

with comparative ease in boiling aniline. The solution is at
first green, but as it approaches saturation, it becomes purplish-blue ; on
cooling, it again becomes green, depositing at the same time small granular
masses of colouring matter, and retains at last only a faint greenish tinge.
The solution at its darkest stage, while still warm, shews a broad but
well-defined band, beginning near the line C of the spectrum, and extending
beyond D, but as the solution cools, depositing the substance contained in it,
the band becomes gradually narrower, until it occupies the space midway
between C and 2), and it then disappears. The masses of colouring matter
deposited from the solution in aniline, are seen under the microscope to
consist of star-shaped groups of irregular crystalline needles. Punicin,
when cautiously heated, furnishes a crystalline sublimate.

Chlorophylloid Colouring Matters.

The consideration of these will be postponed to Book ill. (Respira-
tion).

1 The reader interested in the subject of this paragraph is referred to the fine memoir
of Lacaze-Duthiers (see foot-note 2, p. 307) , and to an interesting article by Dr Schunck,
entitled " Note on the Purple of the Ancients." Journal of the Chemical Society, 1879,
p. 589.

2 Edward Schunck, " Note on the Purple of the Ancients." Journal of the Chemical
Society, No. 202 (1879), p. 589.

CHAPTER IX.

THE CONTRACTILE TISSUES.

SECT. 1. INTRODUCTORY.
THE STRUCTURE OF THE CONTRACTILE TISSUES.

The pro- WHILE Schwann and other observers, about the year
perties of 1835, were tracing the resemblances of vegetable "and

Protoplasm. animal tissues, Dujardin1, a French naturalist, was
investigating in some of the lower animals a remarkable substance
to which he gave the name of sarcode. This substance is amor-
phous, gelatinous of various degrees of consistence, and elastic ;
and it always contains granules of greater or less fineness. It
occurs in fragments whose shape is indefinite and indeed variable.
It is capable of developing within its mass vacuoles or cavities filled
with a pellucid fluid, which afterwards close so perfectly that no
crack or scar betrays their former presence. But it is chiefly remark-
able for its power of extending portions of its surface, at will, into
processes which may or may not inosculate, and which again, at will,
are withdrawn into the general mass. To this property the name
contractility is given. In what manner the protrusion is effected it is
impossible to decide ; but it is easy to imagine that the normal form of
the contractile mass of sarcode is spherical and that contraction may
be exerted in any chord: in which case the corresponding segment
would be pressed out as a process (Hermann). The projection of
columns or processes is not the only movement exhibited by sarcode.
The granules imbedded in its mass may undergo gliding and dancing
movements resembling the mechanical Brownian movements which
are seen when very minute particles are suspended in a liquid.
In each case the granules are passive. In the case of the gliding
movements, which are well seen along the extended processes of
foraminifera, the agent is the contractile sarcode; but in the case
of the dancing movements the cause may be the same as that of
the Browniao movements referred to. It is true that they may
be seen in contractile tissues which are unquestionably alive; but

1 Dujardin, "Recherches sur les organismes inferieurs." Ann. des Sciences naturelles^
2nd Ser. (1835), Vol. iv. p. 343.

BOOK I. CHAP. IX.] THE CONTRACTILE TISSUES. 311

it is also true that they are exhibited in dead tissue, and that
they often seem to depend upon a diluted state of the sarcode1.

It soon became apparent that the most remarkable properties
of sarcode, or, as it is now termed, protoplasm, were not peculiar to
it. Siebold2 discovered contractile powers in the yolk-spheres of
Planarian ova, and Wharton Jones3 in the white corpuscles of
vertebrates, while Kiihne 4 contrasted muscular tissue, Amoebae and
Vorticellae in respect of their excitability and death-changes. Thus
the way was prepared for the doctrine of the analogy of sarcode
to the body or contents of the animal cell, and the doctrine of
the cellular nature of infusoriarjs 5 ; from which we derive the unity
of the contractile power in such creatures as the Amoeba and in
the specialized muscular tissues of man.

Limited powers of contraction are enjoyed by very many cells
of the bodies of higher animals. The connective-tissue corpuscles
of the cornea 6, the cells of hyaline cartilage 7, and the walls of
capillaries 8, seem capable of contracting, at least when stimulated
by electrical currents. The gliding motion of granules in the pigment
cells of the frog's skin may be readily demonstrated. White blood
corpuscles and lymph cells exhibit movements in no respect different
from those of primitive sarcode; while ciliated epithelia and sper-
matozoa offer the simplest examples of movement as a specialized
function. But it is in muscles that contraction becomes prominently
the function of the tissue, and where its laws have been most fully
examined.

ciassifi- Of muscles there are, from the histological point of

cation of view, three sorts : (1) the smooth involuntary muscular

tissue of -intestines> uterus, arterial walls, &c. ; (2) the
striated muscles of the general voluntary system ; and
ture. (3) the striated involuntary muscle of the heart.

Structure of unstriped involuntary muscle.

This variety of muscular tissue consists of innumerable small
fibre-cells (0'045 to 0'230 x 0'004 to O'Ol of a mm.) extended in

1 Kecklinghausen, " Ueber Eiter- und Bindegewebs-Korperehen." Virchow's Archiv
f. path. Anat. u. Physiol., Vol. xxvin. p. 166, 1863.

2 Siebold, Froricp. Notizen, No. 360, p. 85. Quoted by Strieker, "Ueber die Zelle."
Handbuch der Lehre von den Gewcben, chap. i. p. 2.

3 Wharton Jones, "The blood corpuscle considered in its different phases of develop-
ment in the animal series." Phil. Trans. Roy. Soc. Lond. 1846, pp. 63—106.

4 Kiihne, " Untersuchungen ii. Bewegungen u. Veranderungen der contractilen
Substanzen." Archiv fur Anat. Physiol. u. wiss. Med. (Keichert u. du Bois-Eeymond),
1859, p. 816.

6 M. Schultze, "Ueber Muskelkorperchen und das, was man eine Zelle zu nennen
habe." Archiv f. Anat. Physiol. u. wiss. Med. (Eeichert u. du Bois-Eeymond) , 1861, p. 17.

6 Kiihne, Protoplasma, &c., p. 125. Eollett, Strieker's Handbuch, p. 1103.

7 Heidenhain, "Zur Kenntniss des hyalinen Knorpels." Studien des physiol. Inst.
zu Breslau, Part 2 (1863), p. 1.

8 Strieker, "Untersuchungen ii. die Contractilitat der Capillareu." Wiener Sitzungsber.
d. math.-naturwiss. Classe, LXXIV. p. 313, 1877.

312 INVOLUNTARY MUSCLE. [BOOK I.

the axis of contraction1, overlapping their neighbours, to which
they are united by means of an intervening substance well seen
in hardened transverse sections of the tissue. The importance of
this interposed substance has been called in question by Engelmann'2.
In perfectly fresh specimens, not only of the ureter but also of
other smooth muscular tissues, it is impossible to detect any de-
marcation of cell from cell ; the tissue forms, to all appearance,
a homogeneous mass, interrupted only by the nuclei ; it is an optical
continuum. -This homogeneity persists, under favourable circum-
stances, for a short time ; but frequently, after a few minutes
have elapsed, fine lines begin to appear, which speedily cut up
the field into elongated elliptical areas, enclosing the nuclei, and
clearly foreshadowing the cells. Thus the homogeneity claimed
by Engelmann for involuntary muscular tissue is the homogeneity of
an absolutely fresh cornea. It is merely optical and does not imply
a perfect structural continuity in the sense sometimes ascribed to
Engelmann's words3.

The cells are commonly spindleshaped, but sometimes forked and
flattened. They were formerly considered to possess no membrane;
but lately a sheath has been described, with annular swellings which
produce an appearance of transverse striae4. Their substance is
granular, and speckled with a varying number of refractile particles
soluble in alcohol; and they contain an elongated oval or rod-shaped
nucleus. Inside the nucleus one or more distinct nucleoli are found;
and beyond each pole of the nucleus, in the substance of the fibre-cell,
is a short row of larger granules, which diminish in size as they
approach the end of the fibre. The fibres frequently display a
longitudinal striation, especially when treated with reagents5; and,
although they are properly described as non-striated in a transverse
direction6, yet it is no uncommon thing, when they have been
macerated in certain hardening fluids, to find them snapped sharply
across so as to leave a truncated, praemorse surface. When examined
with polarized light, fibre-cells, like the transversely striated muscle

1 The power of contraction along tico axes at right angles to one another has been
suggested by Mr Gaskell in the case of the muscles of arterial walls. (Studies from
the Physiol. Lab. of the University of Cambridge, Part in. p. 164. Also Joitrn. Anat.
and Physiol., Vol. xi.)

2 Engelmann, "Zur Physiologie des Ureter." Pfluger's Archiv, Vol. n., 1869,
pp. 247, 274. "Beitrage zur allgemeinen Muskel- u. Nervenphysiologie." Vol. in.,
1870, p. 248.

3 See the discussion in the Archiv f. mikrosk. Anat. by Dogiel, Foster and Dew-
Smith, &c. Hermann (Physiology, 2nd ed. by A. Gamgee, p. 300) so understands Engel-
mann : but Engelmann always refers to a physiological continuity merely, although he
speaks of the ureter as a ' colossal fibre.'

4 E. Klein, "Observations on the Structure of Cells and Nuclei." Quarterly Journal
of Microscop. Science, New Series, July 1878, p. 331.

5 Flemtning, "Ueber die Beschaffenheit des Zellkernes." Arch. f. mik. Anat.,
Vol. xin. p. 693. Klein, Op. cit.

6 See however Meissner (" Ueber das Verhalten der muskulosen Faserzellen im con-
trahirten Zustande." Zeitschr. f. rat. Med., 2nd Ser., Vol. n., 1858, p. 316) who saw
transverse markings on contracted fibres ; also Klein, Op. cit.

CHAP. IX.] THE CONTRACTILE TISSUES. 313

about to be described, are found to contain doubly refracting, positive,
imiaxal particles scattered through their substance.

Structure of voluntary muscle.

The second kind of muscular tissue is commonly known as volun-
tary, and transversely striated. It consists of elements or fibres, which
are exceedingly large when compared with fibre-cells, being about -5^0 th
of an inch in diameter (10 to 80 /A1), and as much as from 1 to 1 J inches
long 2. Each fibre is enclosed in a structureless elastic sheath or sarco-
lemma, rounded at its extremities, which either become attached to
tendons or aponeuroses, or lie overlapped by neighbouring fibres.
The contents of the sarcolemma when examined in a perfectly fresh
condition, as they may be in the case of cold-blooded animals, are of
a pale grey translucent appearance. They exhibit a very regular
series of transverse markings, but hardly a trace of longitudinal
striation, if care have been exercised in the preparation. The
transverse striation is due to an alternation of dim and bright lines
which commonly run continuously across the long axis of the fibre,
but which are sometimes interrupted by 'faults,' (to use a geological
term,) as if one portion of the fibre had slipped to a lower level than
the rest. The striae in the frog's muscle are exceedingly fine
and somewhat confusing. If we examine in the normal condition
the muscles of animals lower in the scale, we find the corresponding
elements both larger and more complex. This examination may
be made with very little preparation in the case of the limb-muscles
of Hydrophilus, fragments of which may be snipped out and mounted
without any addition, after the chitinous covering of the thigh has
been split, while in the case of Cyclops no preparation whatever is
needed other than fixing the specimen beneath a covering glass3. A
muscle of small diameter and at rest should be selected for observa-
tion. In such a specimen the most striking feature will still be the
alternation of darker and lighter bands. But the dark, or, more
strictly speaking, the dim band will be found more or less marked by
longitudinal lines, and to be traversed by a zone or region less
cloudy than the rest, to which the name of Hensens disc is given.
The lighter stripe, in its turn, is still more clearly divided by a thin
dark line called Krauses membrane, which under a sufficiently
high power in hardened specimens appears as a series — often as
a double series — of dots4. If such a muscular fibre were seen in
cross section, and in a perfectly normal state, it would present
the appearance of a homogeneous clear substance, thickly and evenly

1 Eanvier, Trait'e technique d'Histologie, p. 468.

2 Quain's Anatomy, eighth ed., Vol. n. p. 115.

8 As was demonstrated to the author by Mr Marcus Hartog, in Cyclops the structure
of striated muscle, and the end-organs of the nerves in muscle, may be perfectly studied
in the Jiving, uninjured animal.

4 This line is said to have been first seen by Dobie (Ann. of Nat. Hist., 2nd Ser..
1849, Vol. m. p. 109).

STRUCTURE OF VOLUNTARY MUSCLE.

[BOOK i.

studded with fine dots ; but if the muscle were first frozen before the
section was made, it would be seen to be divided by fine lines into a
number of angular areas, known as Cohnheirris areas; as if the whole

FIG. 54. DIAGRAM ILLUSTRATING THE STRUCTURE OF A STRIPED MUSCULAR FIBRE.

(After Engelmann.)

The specimen was taken from the abdominal muscles of Telephorus melanurus, and
made rigid by being plunged into 50 p.c. alcohol: a represents the fibre in various stages
of contraction, when viewed in common light; 6 is a schema of the same fibre in polarized
light.

1, 2, represent the broad dim bands in the fully relaxed condition ; in, Hensen's
disc (Mittelscheibe) ; q, darker portion of broad dim band (Querscheibe) ; z, Krause's
membrane, appearing double (Zwischenscheibe) ; n, accessory band (Nebenscheibe) ;
i, i, intermediate substance, forming the broad bright stripe or band.

Of the various segments, 1 and 2 are fully relaxed ; 3, 4 and 5 are in the beginning
of contraction ; 6, 7 and 8 constitute the homogeneous stage of contraction ; and from 9
onwards the segments are in the stage of transposed bands, the original bright stripe
being now dimmer than the original dark stripe.

6 shews that there is no transposition of doubly refracting and singly refracting
substance on contraction.

For the sake of simplifying the diagram the double refraction of Krause's membrane
and the accessory bands is not indicated.

fibre consisted of a number of compressed columns or prisms sur-
rounded by the sheath of the sarcolemma. Were we to irrigate these
fresh specimens of muscular tissue with dilute acetic acid, the muscle
would swell up, and the transverse striation would become faint,
while here and there a third element of the tissue, viz. the nuclei,

CHAP. IX.] THE CONTRACTILE TISSUES. 315

would become prominent. These in the fresh state are oval, flat-
tened, structures containing nucleoli and usually surrounded by, or
associated with, a small fragment of unstriated granular protoplasm ;
but under the influence of the acid they frequently become shrivelled
and linear. In the frog and in the water-beetle they may be found
at any depth in the mass of the fibre; while in mammalian muscles
they are situated immediately beneath the sarcolemma.

If the leg of a water-beetle be torn from the body of a recently
killed specimen, and its chitinous covering split; and if it be then
plunged into absolute alcohol ; portions of fibres may be found in
all stages of contraction. Most frequently it happens that in the
contracted portion the sarcolemma is raised up from the contrac-
tile substance opposite the level of each dim band in the form of
a regular fold encircling the whole fibre. In consequence of this, the
sarcolemma at the edge of a longitudinal section (or optical longi-
tudinal section) of a fibre appears very regularly festooned, the
festoons being opposite the ends of the dim bands, and the fixed
points opposite the ends of the so-called membranes of Krause.
Upon this very remarkable appearance, taken in conjunction with
the appearance known as the areas of Cohnheim, Krause has
founded his theory that a muscular fibre is partitioned off into
superposed prismatic cavities or cells, by horizontal diaphragms
which are Krause's membranes, and by vertical walls which are the
boundaries of Cohnheim's areas.

Older views ^ne °lder views of the muscular fibre were chieflv

Bowman's based upon the effects of certain reagents on the tissue.
Sarcous ele- If a muscular fibre is steeped and hardened in a
solution of chromic acid or in alcohol — and this applies
to mammalian as to other fibres — the sarcolemma becomes brittle, and
the whole contents resolve, at the slightest touch, into innumerable
ftnQfibrillae, each of which exhibits an alternation of light and dark
parts corresponding with the light and dark bands of fresh fibres,
If, again, a fibre has been macerated in hydrochloric acid, the
tendency which it exhibits is, not to split longitudinally but rather
transversely, through the centre of the principal bright band, thus
breaking up into a number of superposed discs called the discs of
Bowman. It is clear that, if we imagine cleavage to occur at the same
time in both the longitudinal and the horizontal plane, the muscular
fibre will become broken up into a number of short prisms, rods,
or particles, which may be regarded as the structural units of
the dead fibres. To these Bowman1 gave the name of sarcous
elements, several of which are included in each area of Cohnheim.
The researches of Bowman remodelled the old representations of the
striated muscular fibre, and gave to them a form which they have
more or less preserved for forty years.

1 Bowman, "On the minute structure and movements of voluntary muscle."
Phil. Trans. Boy. Soc. Lond. 1840, p. 457.

316 VOLUNTARY MUSCLE IN POLARIZED LIGHT. [BOOK I.

•Pjjebe. The property of double refraction in muscular

Saviour of tissues has already been mentioned. Its discovery
muscle to po- Was made1 in striated muscle; where also its conditions
larized light. ^ave been more fully observed than in the smooth
variety. A convenient apparatus for demonstrating double refraction
in microscopic objects consists of two Nicol's prisms, one — the polar-
izer— fixed between the illuminating mirror of the microscope and
the object stage, and the other — called the analyser and capable of
rotating about the optical axis of the instrument — interposed between
the ocular and the observer's eye. When the planes of polarization
of the two Nicols are at right angles the prisms are said to be
crossed, and the field of view is darkened ; when they coincide the
field is brightest. If, when the Nicols are crossed, a doubly refracting
body is interposed between them; if for example a plate of doubly
refracting crystal, cut parallel to its axis, is laid upon the stage of
the microscope; the analyser no longer blocks the rays, and the
field again becomes bright. The degree of brightness varies according
to the direction of the axis of the doubly refracting plate: it is
greatest when this axis makes an inclination of 45° with each
Nicol's plane ; and it is nil when it coincides with either of
these. If muscle, or any part of muscle, behaved like such a plate
of crystal, we should ascribe to it similar double-refracting properties.
A more beautiful way of demonstrating the optical properties of
muscular tissue is to interpose a very thin plate of doubly refracting
selenite or mica between the crossed Nicols. In this case, as in the
above experiment, light is transmitted or not through the analyser
according to the inclination of the axis of the plate; but the light is
not white, it is coloured. The particular colour depends upon the
thickness of the plate; and the most useful thickness is that which
gives a purple tint to the field with the proper inclination of the
axis. Supposing this to be attained, and supposing also that we have
that relation of the plate to the prisms which secures the highest
intensity or fulness of colour, we shall find that, as we rotate the
analysing Nicol, the intensity of the tint will diminish to its
vanishing point, at 45°, beyond which the complementary tint will
appear, and increase to its maximum fulness at 90°; and so alter-
nately through every quadrant. If now we place upon the mica
plate a doubly refracting body, its colour will be found to differ
from that of the field according to its doubly refracting character, its
thickness and the inclination of its axis to the crossed Nicol planes.
The advantage of the arrangement is that we may discriminate
between isotropous and doubly refracting bodies, not merely by
different intensity of light, but by more easily detected differences of
colour.

By the aid of such appliances Brucke2 wals able to determine

1 C. Boeck (in 1839) ; reported in Arch. f. Anat. Physiol. u. wiss. Med. (J. Miiller),
1844, p. 1.

8 Brucke, "Muskelfasern im polarisirten Lichte." Strieker's Handbuch, Chap. vi.
p. 170.

CHAP. IX.] THE CONTRACTILE TISSUES. 317

that muscular fibres consist of isotropous and anisotropous or doubly
refracting substance; that the latter is found in the broad dim band
which is made up of a series of sarcous elements; and that these
optically resemble uniaxal crystals the axes of which coincide with the
length of the fibre. By comparing the optical phenomena of muscles
and rock-crystal he assigned to muscle a place among positive double-
refracting bodies; and on purely physical grounds he assumed the
double-refracting powers to be due to the presence in the sarcous
elements of innumerable doubly-refracting particles, to which hypo-
thetical particles he ascribed the name of disdiaclasts (8/9, twice, and
St,aK\aa), I break in twain).

The rest of the muscular fibre is isotropous in all meridians and
all positions; except Krause's membrane, which, like the sarcous
elements, is doubly refracting1.

Of late years the scheme of striated muscle in Arthropoda,
mamS views anc^ esPecially ^n Insccta, has grown to be still more compli-
cated than in this description. With suitable powers, that
which has been called Krause's membrane becomes resolved into three narrow
bands, an intermediate and two accessory2. The intermediate band is
continuous, when in the fresh state, and sometimes double ; but broken into
granules when hardened. The accessory bands are usually more or less
granular. The intermediate band is double-refracting, as may best be seen
in hardened specimens of broad-banded muscles ; whereas double refraction
in the accessory bands is faint and uncertain. The diagram on page 314
should be consulted; it will be noticed that the double refraction of
Krause's membrane is omitted for the sake of simplicity.

«,,Y^TT*oi«« While the above general description includes all

ouoaivision . . . • V • .

of voluntary conditions of voluntary striated muscular tissues,
muscles into varieties are distinguished in the muscles of some
:edt animals: these are the pale and the red, of which the
unlikeness of colour persists after bleeding. In the former, transverse
striation is extremely regular and longitudinal striation merely
indicated : the recti, the vasti and the adductor magnus of the rabbit's
hind limb are instances. In the latter, or red variety, of which the
adductor brevis and the soleus are types, the transverse bands are
broken up by a well-marked longitudinal striation3. Still more
interesting physiological differences will be spoken of hereafter.

1 See following note.

2 Engelmann, Proces verbaal d. k. AJcad. van wetenschappen. Afdeel. NatuurJc.
No. 6, Dec. 1871 ; and No. 7, Jan. 1872. Referred to in a paper by the same author
("Ueber die quergestreifte Muskelsubstanz ") in Pfliiger's Archiv f. d. ges. Physiol.,
Vol. vn. , 1873, pp. 36, 42, 50. Unhappily the term 'strie intermediate ' has been applied
by Eanvier (Traite Technique d'Histologie, p. 481) to Hensen's disc.

3 Eanvier, " De quelques faits relatifs a 1'Histologie et a la Physiologie des Muscles
strips." Arch. d. Physiol. norm, etpathol., 2nd Ser., Vol. i. p. 5, 1874. E. Meyer, "TJeber
rothe und blasse quergestreifte Muskeln." Arch. f. Anat. Physiol. u. wiss. Med.
(Eeichert und du Bois-Eeymond), 1875, p. 217. W. Krause seems to have been the
first to notice the distinction of colour in red and pale muscles (Anatomie des Kaninchens,

318 STRUCTURE OF MUSCULAR TISSUE. [BOOK I.

The blood-vessels of striated muscular tissue are
vessels of very abundant, and the capillaries are small. The

muscular latter are distributed upon the fibres, in elongated

tissue. meshes ; but they nowhere pierce the sarcolemma.

The capillaries of the red variety of voluntary muscles are wider
than those of the pale. They are disposed in shorter meshes, and are
marked by peculiar aneurismal dilatations1.

The structure of the Muscular Substance of the Heart.

The third kind of muscular tissue consists of quadrate cells with a
faint longitudinal striation and a rough transverse striation. In the
centre of each cell is an oval nucleus, usually associated with a
small amount of granular protoplasm ; and not unfrequently the cell-
substance contains a few scattered fat particles. The cells are joined
end to end, or side to side, and often by means of stout truncated
processes. They are apparently destitute of sarcolemma.

The heart-muscle of amphibia differs somewhat from the above
description. The cells are not quadrate, but spindle-like, overlapping
the neighbouring cells. They are transversely striated; and both
Krause's membrane and Hensen's disc are said to have been seen 2.
The dark striae are doubly refracting.

In mammals the size of the cells is from 50 — 70 //, long by
15— 23 Abroad3.

Terminations of Nerves in Muscle.

The mode of union of muscles and motor nerves cannot as yet
be said to have acquired more than anatomical significance. The cells
of smooth muscle are entangled in a net of nervous fibres, from which
fine offsets seem to end abruptly on the surface or in the substance
of the cells, or even to pierce the nuclei4.

A similar disposition of nerves has been claimed for the heart-
muscle 5.

In the case of striated voluntary muscle, the medullated nerve
fibres reach the sarcolemma, and pierce it. The sheath of Schwaim
or neurilemma becomes continuous with the sarcolemma. The

1 Ranvier, "Note sur lee vaisseaux sanguins et la circulation dans les muscles
rouges." Op. cit., p. 446. E. Meyer, loc. cit.

2 Langerhans, "Zur Histologie des Herzens," Virchow's Archiv, Vol. LVIII. p. 56
Gerlach, L., "Ueber die Nervenendigung in der Musculatur des Froschherzens."
Virchow's Archiv, Vol. LXVI. p. 187.

3 Schweigger-Seidel, "DasHerz," Strieker's Handbuch, chap. vii. p. 179.

4 Consult Arnold, "Gewebe der organischen Muskeln," Strieker's Handbuch, chap,
iv. p. 144; and, of the more recent authorities, Lb'wit, Wien. Acad. Sitzungsb. in.
Abth., Vol. LXXI. 1875, p. 355. Gscheidlen, R., Arch. /. mikr. Anut., Vol. xiv. p. 321.
Ranvier, Comptes Eendus, Vol. LXXXVI. p. 1142.

5 Consult Schweigger-Seidel, Strieker's Handbuch, chap. vii. ; and also P. Langerhans,
Virchow's Archiv, Vol. LVIII. p. 65. L. Gerlach, Virchow's Archiv, Vol. LXVI. p. 187.
E. Fischer, Arch. f. mikr. Anat., Vol. xm. p. 365.

CHAP. IX ] THE CONTRACTILE TISSUES. 319

axis-cylinder divides in Mammals, Birds and Reptiles, into a
short dendriform structure called an End-plate, which rests upon a
granular nucleated mass called a Protoplasmic foot. In Amphibia,
there is no protoplasmic foot, and the divisions of the axis-
cylinder are nucleated and long, extending mainly in the long axis
of the fibre, immediately beneath the sarcolemma. In all classes
the white substance of Schwann terminates somewhat abruptly at
the entrance to the sarcolemma or at a little distance within it1.
In muscular fibres destitute of sarcolemma the nerve ends in a
granular Eminence of Doy&re2, on the side of the fibres which usually
bears a nucleus.

CHEMICAL CONSTITUTION OF NORMAL LIVING MUSCLE, so FAR

AS IT CAN BE KNOWN OR INFERRED.

On the distribution of liquid and solid parts in a voluntary muscular

fibre.

In the section which has preceded, an account has been given of
the appearances presented by muscular fibres when subjected to a
high magnifying power; and we have shewn that, according to
all observers, there is contained within the elastic sarcolemma a
substance in which doubly refracting and isotropous structures
alternate. In the sequel it will be shewn that in the process of con-
traction the fibre becomes shorter and thicker, and that at the same
time the anisotropous elements become broader and shorter, the inter-
mediate isotropous substance also exhibiting some diminution in
height and perhaps (though this admits of doubt) diminishing in
amount. Whilst these changes in the form are proceeding, differences
in light- transmitting powers are perceived, though the behaviour to
polarized light remains as before.

We have referred to minor points in which different observers
disagree, but without laying very much stress upon them. We have
now, however, to discuss a question which is of paramount importance,
in reference to the physical constitution of voluntary muscle.

When we examine a dead muscular fibre, especially one which has
been acted upon by various hardening reagents, the contents of the
sarcolemma all unquestionably possess a solid consistence. Can
we, however, infer from such observations that they possess the same
characters during life ? Certainly not. It was shewn by Kiihne3 that

1 Kiihne, Strieker's Handbuch, chap. v.

2 Doyere, "MSmoire sur les Tardigrades. " Ann. des Sci. Nat., Ser. n. Vol. xrv.,
1840, p. 269.

3 Kiihne, ArcMv f. Anat. u. Physiol, 1859, p. 748. Untersuchungen uber das
Protoplasma und uber die Contractilitdt. Leipzig, 1864. (Consult section entitled
"Methoden zur Gewinnung des Muskelinhalts, " p. 2.) Lehrbuch der physiologischen
Chemie, Leipzig, 1866, p. 272.

320 STRUCTURES IN A MUSCULAR FIBRE. [BOOK I.

by subjecting yet living and rapidly frozen muscular fibres to pressure
we can express from the interior of the fibre a somewhat viscous
but yet perfectly liquid substance, to which, he gave the name of the
muscle-plasma, which shortly afterwards, if the temperature be
favourable, sets as a soft jelly: doubtless in consequence of the
coagulation of a proteid body of which the precursor or precursors
existed in solution. Kiihne had the good fortune to observe on one
occasion living muscular fibres, within the sarcolemma of which a
living nematode (subsequently again seen by Eberth1 and indentified
by him as tne Myoryctes Weismanni) freely moved about. This worm
was able to make its way from one end to the other of the muscular
fibre, displacing in its course (but only temporarily) the sarcous
elements, and in a way which left no room for doubt that the creature
was moving in a fluid medium in which were suspended the aii-
isotropous constituents of the fibre. Kiihne was thus led to the
conception that in the voluntary muscular fibre the contents consist
of anisotropous solid bodies — the sarcous elements — which are
suspended in a viscous liquid, contraction consisting essentially of a
change in the form of the suspended bodies.

Objections have been raised to this view of Klihne's, some of which
are based upon microscopic observations, others upon the difficulty
which their advocates have experienced in accounting satisfactorily for
the orderly arrangement of the anisotropous elements, on the hypothesis
that these are simply suspended in a viscous liquid.

Krause, as we have already pointed out, believes that the structure,
which since his description of it has gone by the misleading term of
'Krause's membrane' (viz. the anisotropous structure in the light band
of resting muscle), is attached to the sarcolemma, so that according to
him a muscular fibre would be divided into a series of transverse
compartments. But excellent observers who have followed him
(Engelmann), deny the existence of a membrane, the existence of
which is absolutely disproved by the fortunate observation of
Kiihne.

Such a view as Krause's might have been held before the time of
Kiihne's famous observation, but the latter, it appears to us, supplies
a certain criterion for rejecting the former. On physical grounds it has
been shewn by Briicke, and is maintained in his most recently pub-
lished writings by Hermann, that the existence of a system of trans-
verse partitions in the muscular fibre would oppose a great (and use-
less) resistance to the forces which bring about the changes in its form.

The difficulties wrhich some have experienced in explaining the
orderly arrangement of the anisotropous elements are, as Kiihne
points out, dispelled if we surmise that this orderly arrangement is
dependent (1) on the pressure exerted upon the contents of the fibre
by the sarcolemma, and (2) on the mutual attraction which leads solid
bodies, floating in a liquid, to adhere one to another.

1 Eberth, Zeitschriftf. wissensch. Zoologie, Vol. xn. (18.63), p. 530.

CHAP. IX.] THE CONTRACTILE TISSUES. 321

The facts then that (1) a large portion of the contents of the
sarcolemma can be expelled from it in the condition of a liquid,
and (2) that living bodies move in the interior of the living fibre, as
in a liquid holding solid bodies in suspension, appear to us to settle
definitely the great problem of the physical condition of the
doubly refracting and isotropous elements of muscular fibre. The
most weighty consequences follow the conclusion to which we are led.
We cannot, for instance, for one moment suppose that a liquid
can change its form in consequence of internal forces acting within it,
unless these lead to its becoming solid; we are therefore led on theo-
retical grounds to the conclusion that the sarcous elements must be
the structures which are directly concerned in the change of shape of
the fibre. Engelmann's observations, which in all respects are the
most consistent and satisfactory which have been advanced, since the
earlier classical investigations of Bowmann and Brticke were pub-
lished, seem to shew that in contraction the sarcous elements undergo
a change in form and in volume, increasing in bulk at the expense of
the isotropous substance, so that the combined volume of the contents
remains sensibly the same during contraction and during rest.

Were it possible, we should wish, in the first place, to study
the chemical history of the various structural elements which make
up the muscular fibre ; but this ideal aim can but most imperfectly
be realized ; so far as possible, we feel however bound to attempt the
task.

Chemical characters of the Sarcolemma.

The delicate transparent sheath which in voluntary muscle encloses,
as in a sac, the contractile matter which forms the chief substance of
the muscular fibre, was formerly supposed to be of the same nature as
elastic tissue; like the latter, it is unacted upon by acetic acid, and
resists long boiling with water, though it is ultimately dissolved.
It differs, also, from elastin in being slowly dissolved when heated in
dilute solutions of acids and alkalies. The fact that it is dissolved
gradually at the temperature of the body by the ferments of the
stomach and pancreas has also been adduced as proving that the
sarcolemma is not identical with elastic tissue ; in point of fact even
elastic tissue is slowly dissolved by these ferments, and particularly
by pepsin.

Chemical nature of the doubly-refracting elements of voluntary muscle.

The doubly refracting (anisotropous) matter of voluntary muscular
fibre is, during life, as after death, of solid consistence. It loses its
peculiar optical properties when the fibre containing it is subjected to
the action of either acids or alkalies, or when it is heated to boiling.
For these reasons it has been surmised that this matter is proteid
in nature. It has, however, been remarked that neither alcohol

G. 21

322 THE MUSCLE PLASMA. [BOOK I.

nor salicylic acid — reagents which coagulate the proteids — affect the
doubly refracting sarcous elements, so that one would be inclined
to believe that they consist rather of some derivative of the proteid
bodies than of proteid bodies pure and simple.

It is stated that ' Krause's membrane, ' though sharing the optical
properties of the sarcous elements, has a different deportment towards
dilute acids ; thus a three-per-cent. solution of acetic and a one-
per-cent. solution of hydrochloric acid are said to annul the
anisotropous character of the sarcous elements, but not of Krause's
membrane, which is, however, affected by caustic alkalies in the same
manner as the sarcous elements.

The Muscle Plasma1.

Kiihne's2 The ^9.u^ to which the name of muscle plasma is

method of given, and which constitutes, as has been shewn, the
obtaining isotropous material of the voluntary muscular fibre, can

Muscle Pias- orjy be obtained from muscle which has not passed into
ma" the state of rigor mortis, for when this change occurs, a

solidification of a proteid matter previously in solution occurs, and
muscle plasma, properly so called, ceases to exist. Cold delays the
coagulation of the plasma, as it does that of the liquor sanguinis, and
it is by its aid that the plasma can be obtained.

The muscles of cold-blooded animals alone preserve their vitality
sufficiently long to permit of the plasma being removed before rigor
has had time to occur ; practically those of the frog are always em-
ployed, and the process is the following :

The frog is bled, and salt solution (\ p.c.) is injected into the
aorta, so as to wash the whole of the blood out of the muscles. The
muscles are then cut up into small pieces, and washed in, or kneaded
with, more of the same salt solution cooled to 0° C., with the object of
getting rid of lymph. The fragments are then collected together,
enclosed in fine linen, and tied up so as to constitute a compact ball,
which is exposed to a temperature of about — 7° C. until it is in
such a condition that it can, by means of cooled knives, be conveni-
ently cut into very fine slices ; this operation can only be carried
out in very cold weather. The frozen slices are then pounded in
cooled mortars, the pounded muscle tied up in strong linen, and
expressed in a strong press at the temperature of the room. The
muscle thaws at 0°, so that the liquid which flows from the press has
this temperature ; it is then filtered through small paper filters
moistened with ice-cold salt solution ; as the filters speedily clog, the
fluid must frequently be transferred to fresh filters.

1 This account of the Muscle Plasma and Muscle Serum, is taken almost verbatim
from Kiihne's Lehrbuch der physiologischen Chemie, It would have been vain to
attempt to give a more succinct or a more satisfactory account than that of the eminent
physiologist to whom we owe almost every fact known in relation to the proteids of
muscle.

2 Kiihne, Untersuchungen iiber das Protoplasma^ p. 2. Lehrbuch, p. 272.

CHAP. IX.] THE CONTRACTILE TISSUES. 323

The filtrate obtained by the above operation is a faintly yellow
opalescent liquid. It is muscle plasma.

Properties Muscle plasma is of syrupy consistence ; it flows,

of the muscle however, forms drops, and possesses all the characters
plasma. Of a liquid. It has a faint alkaline reaction.

At ordinary temperatures muscle plasma coagulates exactly like
blood plasma. Coagulation is accelerated by contact with foreign
matter and commences at the points of contact ; it is also accelerated
by stirring with a glass rod.

Myosin.

The solid body which separates from muscle plasma when this
liquid coagulates has received the name of Myosin. This body differs
from fibrin in being a gelatinous mass when first formed, and though
it subsequently contracts, it never becomes fibrous, nor has the
opacity of blood fibrin.

Reactions The separation of myosin is hindered by cold. At

of muscle temperatures about 0°C. it occurs very slowly, whilst at

plasma de- 40° C. almost instantaneously.

^S UP°n When muscle plasma is diluted with cold water,

myosin is instantly precipitated, so that a drop of
muscle plasma allowed to fall into water sets instantly in the form
of a solid elastic ball. Dilute acids, and solutions of NaCl containing
from 10 to 20 per cent, of the salt, cause instantaneous coagulation.

Muscle plasma may be mixed with ice-cold salt solutions contain-
ing from 5 — 7 p.c. of NaCl without myosin separating.

When plasma is allowed to flow guttatim into dilute hydrochloric
acid (containing 01 per cent.), the little balls which are at first
formed dissolve as they sink through the column of liquid, and give
rise to an opalescent solution.

Pure myosin is obtained by dropping muscle plasma

Prepara- 'n^0 ji^Hed water, whereby a precipitate consisting of

myosin. little balls is obtained, which is easily washed with water.

Myosin which has been thoroughly washed with water

has a neutral reaction, is quite insoluble in pure water, but readily

soluble in solutions of common salt containing between 5 and 10 per

cent, of NaCl.

Another method of preparing myosin is based upon the solubility
of coagulated myosin in weak solutions of common salt. Muscle
is thoroughly washed with water, is finely divided, and rubbed up to
the consistence of a fine paste with powdered common salt, the
amount of salt which has been added being determined. Water is
then added in such quantities as to form with the salt of the muscle
a solution containing 10 per cent, of NaCl. The mixture of finely
divided muscle, salt, and water, which should have the consistence of
a thin magma, is set aside for 24 hours, then pressed in linen, and
filtered through paper. The yellowish, syrupy, solution when poured

21—2

324 MYOSIN. MUSCLE SERUM. [BOOK I.

into water furnishes at once pure myosin, and resembles muscle
plasma in all respects, except in not coagulating spontaneously.

Coagulation When gradually heated a solution of myosin begins

of myosin. to be turbid at 55° C., and deposits flakes of proteid

matter at 60° C., which consists of a coagulated product which
resembles other proteids coagulated by heat.

Powdered common salt, added in excess, precipitates myosin from
its solutions in common salt.

Myosin, like fibrin, decomposes peroxide of hydrogen.
Myosin- Liebig1 shewed that when muscle is placed in dilute

Syntonin. hydrochloric acid containing 1 part of acid in 1000, the

proteid natter is in great part dissolved, to be precipitated when the
solution is neutralized. Liebig believed that the body dissolved in
the acid was a special body, muscle-fibrin ; it is now known, however,
that the solution merely contains acid-albumin or syntonin. differing
in no respect from acid-albumin obtained from other proteids. It
has been suggested that the ease with which it is converted into
syntonin, under the influence of dilute hydrochloric acid, specially
distinguishes myosin ; it is probable that the rapid formation of acid-
albumin is due to the fact that muscle always contains a trace of pepsin.
The facility with which a solution of acid-albumin can
Characters ^ obtained from muscle, causes us to examine in this place
°f S B°toninn *^e reac^ons °f such a solution in greater detail than was
thought advisable in Chapter I.

To prepare acid-albumin from muscle this tissue is finely divided and
then placed in a large quantity of dilute hydrochloric acid (1 part of HC1,
1000 parts of water). The solution is after some hours filtered. On
neutralizing, gelatinous flakes are obtained, which are collected on a filter
and washed. These contain in 100 parts :— C, 54-06 • H, 7'2S ; N, 16-05 ;
S, Ml; O, 21-50.

Acid solutions of syntonin are not coagulated by heat; they are
precipitated by sodium chloride, ammonium chloride, calcium chloride,
sodium sulphate and magnesium sulphate.

Syntonin is soluble in a solution of sodium carbonate of 1 p. c., and the
solution is not coagulated by heat. It is soluble in cold solution of lime
\vater, and the solution does not coagulate when boiled ; it froths when shaken.

Muscle Serum.

Following the analogy of the blood, we may designate, by the
name of muscle serum, the liquid which remains after the separation
of the spontaneously coagulating substance from the muscle plasma.
The muscle serum from which myosin has separated at a low tem-
perature has a neutral or faintly alkaline reaction. Kept at the
ordinary temperature of our dwelling-rooms it acquires an acid re-
action, in consequence of the development within it of sarcolactic acid.

Proteids of Muscle serum contains three proteids in solution : —

the muscle 1. A proteid body which coagulates when the

Berum. muscle serum is cautiously heated (if needs be after

1 Liebig, Ann. d. Chem. u. Pharm. Vol. LXXIII. pp. 125—129.

CHAP. IX.] THE CONTRACTILE TISSUES. 325

careful neutralization) to 45* C. This body is not myosin, being dis-
tinguished from it by its insolubility in weak solutions of NaCl1.

2. An alkaline (potassium) albuminate, which is only precipi-
tated when the reaction is made strongly acid.

3. Albumin apparently identical with serum-albumin, and
coagulating, like it, at a temperature between 70° and 75° C. and not
coagulated by the addition of ether. This proteid is much more
abundant than either the first or second mentioned.

The great majority of the constituents to be discussed in the
sequel are contained in the muscle serum ; they will, however, for
convenience, be considered under separate headings.

The Haemoglobin of Muscles.

We have already stated that certain of the voluntary muscles are
distinguished by their red colour, due to the presence of haemoglobin
which colours the contents of the sarcolemma. In warm-blooded
animals, indeed, the majority of muscles are red, whilst in cold-
blooded animals frequently the heart is the only red muscle. In
certain gasteropod molluscs (Limnaeus and Paludina) Lankester
made the remarkable observation that whilst haemoglobin is not
present in the blood, it colours the muscular fibres which occur in the
walls of the pharynx, these muscles being among the most active in
the body.

Wherever haemoglobin occurs in the substance of muscle it colours
the plasma and not the anisotropous sarcous elements ; when the
plasma coagulates, a portion of the colouring matter adheres to the
myosin, whilst a portion remains in solution in the muscle serum.
To demonstrate the presence of haemoglobin in muscle, the blood-
vessels are washed out with salt solution, and thereafter the blood-
free muscle is held between a light and the slit of the spectroscope.
The muscular portion of the diaphragm of the rabbit lends itself
particularly well to this observation. Crystals of haemin may by
suitable treatment be obtained from the red muscles, or from the
plasma which they yield (Kiilme).

NITROGENOUS (NON-PROTEID) ORGANIC CONSTITUENTS OF MUSCLE.

Extract of When finely divided dead muscle is repeatedly

meat. treated with cold water, this liquid dissolves the whole

of the constituents of the muscle serum and, in addition,
perhaps, soluble matters derived from the insoluble anisotropous
sarcous elements. The solution thus obtained has a red colour due
to the haemoglobin extracted from the muscular fibres and (unless
the blood-vessels have been thoroughly washed out with salt solution)
derived from the blood contained in the vessels of the tissue.

1 The reader is referred for some recent observations on the proteids of Muscle to a
paper by Demant entitled " Beitrag zur Chemie der Muskeln." Zeitschrift f. physiol.
Chemle, 1879, p. 241.

326 EXTRACT OF MEAT. CREATINE. [BOOK I.

When the solution is boiled, the haemoglobin and the soluble
proteids which it contains are coagulated and, on filtering, a clear
liquid is obtained containing the salts of muscle, certain non-nitro-
genous organic bodies, such as glycogen, inosit, lactic acid, &c., and
a mixture of nitrogenous organic bodies, mostly basic in their cha-
racter; these are creatine, creatinine, carnine, xanthine, hypoxan-
thine, and perhaps urea. To the residue obtained by evaporating an
aqueous infusion of muscle, the name of extract of meat is given ;
several extracts of meat exist in commerce, which are substantially
obtained in the way we have mentioned and which may be employed
in the laboratory for the preparation of the various nitrogenous
organic bodies to be now described. Beef-tea is an aqueous extract
of meat, and contains the same substances as are present in the
solid extract of meat.

Creatine. C4H9N302 + H20.

This body occurs in only two of the elementary
tissues of the body, viz. in muscular and nervous tissue.
It has never been found in any glandular organ. In

small quantities it occurs in the blood, but it is present in muscle in

largest amount.

I. (Liebig's method1). Muscle is reduced to a fine state of division,
as for example by the use of a sausage machine, and then mixed with
one half its weight, or its own weight, of cold water, and set aside for
some hours. The insoluble matter is separated on a linen filter
from the liquid, and the former is subjected to strong pressure. It is
then treated with a quantity of water equal to that first used, and,
after some hours, the process of filtration and pressing repeated
as before ; the water used for the second extraction may be employed
afterwards to extract a fresh quantity of meat. The liquid thus
obtained is then boiled, by which means the albumin which it
contains is coagulated ; after removing the albumin by filtration,
baryta water is added to the filtrate, so as to precipitate the whole of
the phosphates present. The excess of baryta present in the solution
is removed by passing through it a current of C02, and, after filtering,
the filtrate is concentrated by evaporation on the water-bath, until
it has a syrupy consistence ; it is then set aside for some days.
Creatine separates out in the form of crystalline crusts adhering to the
bottom of the vessel; the mother liquor is poured off and the
crystals washed with cold alcohol; these are then dissolved in
boiling water, and the solution decolorized by means of animal
charcoal. On evaporation, crystals of creatine separate which are
purified by recrystallization.

II. (Neubauer's method2). The watery extract of muscle is preci-
pitated by solution of lead acetate, the solution is treated with
sulphuretted hydrogen to remove the excess of lead, and is cautiously

1 Liebig, Ann. d. Chem. u. Pharm. Vol. 62, p. 257.

2 Neubauer, Ann. d. Chem. u. Pharm. Vol. 119, p. 27.

CHAP. IX.] THE CONTRACTILE TISSUES. 327

evaporated until, on cooling, crystals commence to separate. It is then
set aside for some days to crystallize. The liquid from which the
crystals have separated is then treated with twice or three times its
own volume of 88 per cent, alcohol, and the crystals which readily
fall from this mixture of mother liquor and spirit are collected
on a filter and, if necessary, weighed. They are at first yellow,
but are obtained perfectly colourless by recrystallizing.

III. (Stadeler's process1). Finely divided meat is digested on the
water-bath with twice its volume of alcohol. The insoluble matter is
pressed and the filtrate is heated on the water-bath so as to drive off
a great part of the alcohol. Solution of basic lead acetate is then
added and the process continued as in II.

Pro erties Creatine crystallizes in the form of transparent,

colourless, shining, oblique, rhombic columns, which
when heated to 100° C. lose their water of crystallization (12'17 p.
cent.) and become opaque.

The crystals belong to the monoclinic system. Inclination of the clino-
diagonal to the principal axis— 70° 20'. Inclination of the faces oo P : co P
in the plane of the orthodiagonal and principal axis = 132° 2' (nearly). Spe-
cific gravity of the crystals 1'35 to 1/34.

Creatine is soluble in 74 parts of cold water at 18° C. : freely
soluble in hot water: slightly soluble in spirits of wine: but almost
insoluble in absolute alcohol and ether. One part of creatine requires
9400 parts of absolute alcohol at ordinary temperatures to dissolve it.

Compounds Though solutions of creatine have a neutral reaction,

the body is a weak base which, when dissolved in hydro-
chloric, sulphuric, and nitric acids, forms compounds which crystal-
lize well. The following are the formulae of these compounds :

Hydrochlorate of Creatine C.H9N3O2 . HC1
Sulphate „ „ C4H9N302.H2S04

Nitrate „ „ C4H9N3O2 . HN03.

A compound of mercury and creatine is formed in which two atoms
of hydrogen are replaced by a single atom of mercury (C4H7HgN3O2).

1 Stadeler, Journ. f. pract. Chem. Vol. 72, p. 25P.

328 CREATINE. [BOOK I.

I. When creatine is dissolved, in the cold, in the

ti strong mineral acids, or heated for some time in dilute

mineral acids, or heated for a much longer time (several days)

with water, a molecule of water is removed and Creatinine is formed, thus :

C4H9N3O2 = C4H7N30 + H20

Creatine. Creatinine. Water.

II. When boiled with baryta water, creatine yields urea, sarcosin,
methylhydantoin, methylparabanic acid and ammonia; these bodies are
products of several reactions which go on side by side, as shewn by the
following equations :

(a) C4H9N3O2 + H20 = CH4N20 + C3H7N02

Creatine. Water. Urea. Sarcosin.

(b) C4H9N3O2 = C4H6N2O2 + NH3

Creatine. Methylhydantoin. Ammonia.
(c) C4H9N3O2 + 02 = C4H4N208 + NH3

Creatine. Methylparabanic Ammonia.

acid.

III. When boiled with mercuric oxide, an aqueous solution of
creatine yields oxalate of inethylurarnine and water, thus :

C4H9N3O2 + 2HgO = C2H7N3 . C2H204 + 2Hg

Creatine. Mercuric oxide. Oxalate of Methyl- Mercury.

uramine.

Synthesis When the alcoholic solution of 2 parts of sarcosin

and constitu- (methylglycosine) is heated for some hours at 100°C. with 1
tion of crea- part of freshly prepared cyanamide, creatine is formed ; thus :
tine.

Methylglycosine, Cyanamide. Methyl-guanidine

or acetic acid,

Sarcosin. or Creatine.

Quantity of The quantity of creatine in the muscles of different

creatine pre- animals and in different muscles of the same animal
sent in mus- does not vary greatly. The following are the results of

Voit's1 determinations.

100 parts of Muscle of Frog yield 0' 21—0' 35 pts. of Creatine.
„ ., „ Fox „ 0-206—0-237 „ „ „

„ „ „ Ox „ 0-219—0-276 „ „ „

„ „ „ Dog „ 0223—0-248 „ „ „

„ „ „ Rabbit,, 0-269—0-336 „ „ „

„ „ „ Man „ 0-282—0-301 „ „ „

According to Voit tbe muscular substance of tbe heart contains
less creatine than the voluntary muscles, and not, as Liebig stated,
a larger quantity. According to Demant the amount of creatine*
increases remarkably in starvation.

1 Voit, Zeitschr.f. Biolog. Vol. iv. (1868) p. 77.

2 Demant, " Zur Kenntniss der Extractivstoffe der Muskehi." ZeitscTiriftf.phys.
Chemie, Vol. HI. (1879) p. 387.

CHAP. IX.] THE CONTRACTILE TISSUES. 329

Creatinine. C4H7N30.

This body, which is a strong base, and which, as has already been
stated, can be obtained from creatine by the prolonged action
of dilute acids or water, is according to Neubauer1 and Nawrocki2
not present in muscle. On the other hand, C. Voit believes it to be
occasionally present in that tissue.

Creatinine forms with zinc chloride a sparingly soluble compound
having the composition represented by the formula (C4H?N3O)2 ZnCl2,
and it is by conversion into this compound that it is always
estimated. Creatinine will be treated of more fully in the Chapter
on Urine.

Hypoxanthine or Sarcine. C5H4N4O.

This body was first discovered by Scherer in the splenic pulp, but
was shewn by Strecker to be constantly present in muscle. Unlike
creatine, hypoxanthine is pretty widely distributed, being found in
the blood, in many glands, in the marrow of bones, &c.

Prepara- The mother liquor from which creatine has sepa-

tion. rated is diluted considerably with water, and ammonia

is added until the reaction is alkaline; it is then treated with an
ammoniacal solution of silver nitrate, which throws down a flocculent,
gelatinous precipitate (C5H2Ag2N40), which is allowed to subside,
and first washed by decantation with weak solution of ammonia, and
then collected on a filter. The precipitate is then boiled in nitric
acid of specific gravity 1*1, which dissolves the hypoxanthine com-
pound, leaving undissolved any silver chloride which may be mixed
with it. The latter is separated from the solution by decan-
tation. On cooling, the nitric acid deposits a white crystalline com-
pound of hypoxanthine and silver nitrate, having the composition
C5H4N4O . AgNO3 ; these crystals often present the form of isolated
spindles or of rosettes formed of spindles radiating from a centre.

The silver compound is suspended in water, and treated with
sulphuretted hydrogen, then heated ; the clear fluid filtered from the
precipitate of silver sulphide deposits, on concentration, crystals of
nitrate of hypoxanthine ; when this compound is dissolved in hot
water and treated with ammonia, it deposits crystalline nodules (never
needles) of hypoxanthine8.

Pro erties " ^e foregoing paragraph, hypoxanthine

crystallizes in the form of nodules which never (Kiihne)
exhibit any needles. It is scantily soluble in alcohol; it is soluble in

1 Neubauer, Zeitschr. f. anal Chemie, Vol. n. (1863) p. 22.

2 Nawrocki, Centralbatt f. d. med. Wissensch. 1865, p. 417. Zeitschr. f. anal. Chemie,
1865, p. 336.

3 Very admirable woodcuts exhibiting the crystalline forms of the compound
of hypoxanthine with silver nitrate and of nitrate of hypoxanthine, are to be found in
Kiilme's Lehrbuoh, etc. p. 295 and 296.

330 HYPOXANTHINE. XANTHINE. [BOOK I.

78 parts of boiling and 300 parts of cold water ; it is soluble in dilute
acids and alkalies. It forms compounds with acids, bases, and metallic
salts. Certain of the latter have already been referred to; a com-
pound with platinum, having the composition of C5H4N40 . HC1 . PtCl4,
may be mentioned, as well as one with copper which is formed when
a solution of hypoxanthine is boiled with solution of cupric acetate ;
this compound is a brownish flocculent body insoluble in water, which
does not admit of purification ; it yields however impure hypoxanthine
when it is decomposed by HaS.

Relations Hypoxanthine is very closely related to xanthine

of Hypoxan- and to uric acid, as would appear probable from an exami-
thine to other nation of their formulae.

bodies< Uric acid . . C5H4N403.

Xanthine . . C5H4N4O,.
Hypoxanthine . . C6H4N4O.

From the two first of these bodies, hypoxanthine can be obtained
by the action of sodium amalgam; when oxidized with nitric acid
it yields xanthine.

Proportion According to Neubauer1 the flesh of the ox contains

of hypoxan- 0*022 per cent, and that of the rabbit 0'026 per cent,

thine found in of hypoxanthine.
muscle.

Xanthine. C5H4N4O2.

Xanthine a This constituen* °f muscle was first discovered by

rare consti- Marcet2, as a constituent of a urinary calculus, and by
tuent of uri- him called xanthic oxide. It was afterwards analyzed
nary calculi, by Liebig and Wohler3 and linger4. It has been
of guano and discovered in guano5, and in some cases in the urine
of man6 and the lower animals7.

Preparation ^n preparing hypoxanthine that body was directed

from muscle, to be precipitated with ammoniacal solution of nitrate
Neubauer's of silver, and the precipitate dissolved in nitric acid of

method. Sp gr j.-^ jt wag stated that when the acid cooled

the compound of hypoxanthine and silver nitrate separated. Now
the first precipitate (viz. that thrown down by ammoniacal silver
nitrate) contains, in addition to hypoxantlnne, a silver compound
of xanthine; the latter compound being more soluble in nitric acid of

1 Neubauer, Zeitschrift f. anal. Chem. vi. 33.

2 Marcet, Essay on 'the Chemical History and Chemical Treatment of Calcvlons
Disorders. London, 1819.

3 Liebig und Wohler, Poggendorff's Ann. Vol. XLI. p. 393.

4 Unger, Ann. d. Chem. u. Pharm. Vol. LVIII. p. 18.

e Unger and Pbipson, Chem. News, Vol. vi. 1862, p. 16.

6 Bence Jones, Quart. Journ. of Chem. Soc. Vol. xv. p. 78.

7 Weiske, "Xanthin und Harnsaure im Ham eines kranken Scbafbockes."
Zcitechr.f. Biol. xi. p. 254.

CHAP. IX.] THE CONTRACTILE TISSUES. 331

sp. gr. 1*1 than the former, remains in solution after the hypoxanthine
compound has crystallized out. On supersaturating with ammonia,
a gelatinous compound of xanthine and silver (C5H2Ag2N402 + H20)
separates. By dissolving in warm nitric acid, the compound
C5H4N4O2 . AgN03 is again formed, and from the latter xanthine
can be prepared by following a process analogous to that which
has been described in the case of hypoxanthine.

Properties Xanthine when freshly separated from its solutions

of Xanthine. presents the appearance of white amorphous granules.

Xanthine is almost completely insoluble in cold water, requiring
about 14000 parts of water at 16° G. to dissolve it, and 1400 parts of
boiling water. It is easily soluble in solution of ammonia, which
deposits it, on evaporation, in the form of indistinctly crystalline
plates (Kiihne) : from the ammoniacal solution it is completely pre-
cipitated by lead acetate.

Solutions of other alkalies likewise dissolve xanthine, and from
these it is precipitated by acids.

Reactions Ammoniacal solutions of xanthine when heated with

>f xanthine. Snver salts reduce the silver to the metallic state.

On heating xanthine in hot hydrochloric acid, and evaporating,
microscopic crystalline masses, composed of aggregations of hexagonal
plates, separate ; these consist of the hydrochlorate of xanthine,
C5H4N4O2 . HC1 + H2O. In a similar manner the nitrate is formed
(C5H4N402 . HN03), and this crystallizes in rhombic plates arranged
in clumps. The solution of the nitrate is precipitated by silver
nitrate in a flocculent form, and the precipitate may be dissolved
in hot nitric acid and allowed to crystallize on cooling ; it has the
composition (CSH4N402 . AgNO3) ; this body is much more soluble
in nitric acid than the corresponding hypoxanthine silver compound.
It separates irom its solution in nitric acid in the form of groups of
fine needles, which do not resemble the hypoxanthine compound.

Xanthine (like hypoxanthine) is soluble in pure (colourless) warm
nitric acid without the disengagement of gas; on cautious evapo-
ration a colourless nitrate is left ; the residue is not rendered purple
by ammonia. By these reactions xanthine is distinguished from uric acid.

Heated with fuming nitric acid containing nitrous acid, a citron-
coloured residue is left, which becomes orange or red on the addition
of caustic soda, and which when heated exhibits at its margin a
fine purple red colour.

of xanthine According to Scherer1 the fresh muscles of the

found in mus- horse contain 0'0026 p. c. of xanthine.

cle.

Relations This body, as has been said in treating of hypoxan-

of xanthine. thine, is closely related to that body and to uric acid.

1 Scherer, Ann. d. Chem. u. Pharm. Vol. cvn. (1858) p. 314.

332 CARNINE. [BOOK i.

Artificial Salomon1, and Krause2 who worked under his direc-

proiuction of tion, have shewn that both hypoxanthine and xanthine
xanthinefrom are formed in small quantities during the digestion of
proteids. fibrin with trypsin and pepsin. Both bodies are like-

wise formed when fibrin is digested at the temperature of the body
with weak hydrochloric acid.

Carnine. C7H8N403 + H20.

This base, discovered by Weidel3, has only hitherto been found in
Liebig's extract of meat, though doubtless it is a regular constituent
of muscle ; it constitutes about 1 p.c. of extract of meat.

Prepara- Liebig's extract of meat is dissolved in 6 — 7 times

tion* its volume of warm water, and cautiously precipitated

with a strong solution of baryta water, great care being taken to avoid
an excess of the precipitant. The filtrate is treated with solution of
basic lead acetate, and the precipitate collected and boiled with
water; the compound of carnine and lead being comparatively soluble
in boiling water is extracted by repeatedly boiling the precipitate
with water. The warm solution is treated with sulphuretted hydro-
gen, and the precipitate of lead sulphide having been separated by
filtration the filtrate is concentrated and treated with solution of
silver nitrate which precipitates a flocculent silver compound
(C7H?N4O3)2 . AgNO3 mixed with some AgCl. The latter is separated
by digesting the precipitate in ammonia. The precipitate is then
suspended in water, subjected to the action of H2S, and the filtrate
being concentrated yields crude carnine, which is purified by re-
crystallizing and by the action of animal charcoal.

Pro erties Carnine is very little soluble in cold, but easily and

completely soluble in boiling water. It is insoluble in

alcohol and ether. Its aqueous solution has a neutral reaction ; . it

has a scarcely perceptible taste at first, but leaves a slight bitter

after-taste.

Chemical When a hot solution of carnine is treated with a

relations. saturated aqueous solution of bromine, a slight evolution

of gas takes place, the solution is decolorized, and on concentration
deposits crystals of the hydrobromate of hypoxanthine ; at the same
time bromide of ethyl is formed ; thus :

C7H8N403 + 2Br = C6H4N4O.HBi- -f CH3Br + CO,.

Carnine. Bromine. Hypoxanthine- Bromide Carbonic

hydrobromate. of ethyl. acid.

1 Salomon, "Bildung von Xanthinkorpern aus Eiweiss durch Pankreasverdam
Ber. d. deutsch. chem. Ges. Vol. xi. p. 574.

2 Krause, " Ueber Darstellung von Xanthinkorpern aus Eiweiss." Inaug.
Berlin, 1878. (Abstracted in Maly's Jahresbericht, Vol. vm. p. 80.)

3 Weidel, "Carnin, eine neue .Basis aus dem Fleischextract." Ann. d. Chem.
Pharm. Vol. CLVIII. (1871) p. 353—368.

CHAP. IX.] THE CONTRACTILE TISSUES. 333

From this decomposition it would appear not improbable that car-
nine is one of the intermediate products between the proteid mole-
cule and bodies belonging to the uric acid group.

Uric acid. C5H4N403.

It is most questionable whether uric acid exists in muscle; by
Meissner it has been fouud in traces in the muscles of fowls1.

Urea. CH4N20.

It is yet doubtful whether urea should be reckoned amongst the
constituents of normal muscle. Liebig came to the conclusion that
it did not occur in that tissue, and this opinion has been generally
entertained. Of late Picard2 has stated that the muscular tissue of
rabbits contains as much as 3 per cent, of urea. This statement is
entirely without foundation, Picard having by the method of analysis
which he employed reckoned other bodies, and especially creatine, as
urea. It is probable that muscle does contain an exceedingly small
quantity of urea, though its separation from the other nitrogenous
constituents offers peculiar difficulties.

Inosinic acid. C10II14N4On.

By this name Liebig3 designated a syrupy acid which he believed
to be a distinct proximate principle of muscle, though present in
extremely small quantity. With bases this acid is said to form
crystallizable salts. A fresh investigation is needed even to establish
the existence of this body.

Taurine. C2H7NS03.

This body has been found in the muscles of the horse by Lim-
pricht4 and Jacobsen5, and in those of fishes by Limpricht. It is,
according to Valenciennes and Fremy, found in the muscles of
molluscs.

NON-NITROGENOUS ORGANIC CONSTITUENTS OF MUSCLE.

Living muscle contains in addition to the previously described
nitrogenous organic constituents, considerable quantities of non-nitro-
genous organic bodies ; these are fats, glycogen, dextrin (?), inosit,
and perhaps small quantities of a fermentable sugar.

1 Meissner, Zcitschr. f. rat. Med. Vol. xxxi. (1868) p. 144.

2 Picard, Comptes Eendus, Vol. LXXXVII. (1878) No. 15 and 25.

3 Liebig, Ann. d. Chem. u. PJiarm. Vol. LXXII. (1847) p. 317.

4 Limpricht, Ann. d. Chem. u. Pharm. Vol. cxxxm. (1865) p. 293.
6 Jacobsen, ibid. Vol. CLVII. (1871) p. 227.

334 FATS. GLYCOGEN. . [BOOK I.

Fats.

The connective tissue which separates the muscular bundles
always contains some fat-cells, and as it is impossible to separate the
muscular fibres absolutely from these, we cannot readily determine
the amount of fat which belongs to the isolated muscular fibres.
There is reason to believe, however, that quite independently of the
fat-cells of the connective tissue of muscle, the muscular fibres con-
tain fat which they give up to ether ; we know nothing, however,
either as to its amount or composition.

In phosphorus poisoning a fatty degeneration of muscle occurs,
similar to that observed to occur as an idiopathic affection, specially
affecting the muscular substance of the heart. In all probability,
in this as in other cases, the fatty degeneration is an evidence of
impeded nutrition (probably of imperfect oxygenation) of the tissue.

Glycogen. (C6H1005)u.

This body, which will be treated of fully in connection with the
liver, is a constant ingredient of the living muscular tissue.

It was at first supposed to be only present in the muscles of the
embryo1, but it was afterwards shewn to occur in muscles of adult
animals under certain conditions, and later still it was found to
be constantly present by Nasse2, Briicke, Abeles and others.

As the glycogen of muscle, on the cessation of the
separating vitality of the tissue, is very rapidly converted into
and deter- sugar, in order to separate muscle-glycogen the tissue
mining the must, whilst yet living, be placed in boiling water, with
amount of t^e Object of destroying the amylolytic ferment which
muscle611 would effect the change. It is then taken out of the

boiling water, reduced to a very fine state of division,
and boiled again in water. From this liquid, impure glycogen may
be precipitated by concentrating it and then adding an excess of
alcohol. The method of effecting the purification (by Briicke's
method) of the impure glycogen obtained by this method will be
described in connection with liver-glycogen.

Abeles' method ^ *s exceedingly difficult, indeed almost impossible,
to extract the whole of the glycogen from muscles by
boiling them with water. Briicke3 suggested that the muscle should
be boiled in a dilute solution of caustic potash. In this way the
whole of the glycogen is extracted, but a large quantity of proteid
matter passes into solution. Abeles4 gets rid of this by boiling with
zinc chloride. His method is the following : —

1 Claude Bernard, Comptes Rendus, Vol. XLVIII. (1859) p. 673.

2 Nasse, "Beitrage zur Physiologie der contractilen Substanz." Pfliiger's Archir,
Vol. ii. (1869) p. 97—121.

3 Briicke, Sitzungsber. d. Wien. Akad. Vol. LXIII. p. 214.

4 Abeles, "Beitrage zur Kenntniss des Glycogens." Mcd. Jahrbiicher, 1877, p. 551.

CHAP. IX.] THE CONTRACTILE TISSUES. 335

The muscle of which the glycogen is to be separated, is subjected
to long boiling in a solution of caustic potash ; the solution is then
almost neutralized with hydrochloric acid, care being taken, however,
that the reaction still continues distinctly alkaline; solution of chloride
of zinc is then added to it and it is boiled for a period varying
between 20 and. 40 minutes ; the proteid bodies are precipitated and
an easily filtered clear liquid is obtained. It is of importance that
just enough of the zinc salt should be added to effect the precipita-
tion. When this point has been attained and the clear liquid is no
longer rendered turbid by boiling with a fresh quantity of zinc
chloride, it is filtered, and the precipitate carefully washed ; the filtrate
and washings are concentrated on a water-bath, allowed to cool, and
then treated with much alcohol which has been faintly acidulated
with hydrochloric acid. Glycogen is thus precipitated ; it is collected
on a filter, washed with weak spirit containing about 60 p. c. of
alcohol, and acidulated with hydrochloric acid, until the washings
contain no zinc ; the acid alcohol is then displaced by pure alcohol,
and lastly the substance is dried, and weighed, or heated with dilute
mineral acids for 2 or 3 hours, and the sugar formed determined.

, Nasse scalds a known weight of muscle, then

pounds it up in .a mortar with a weighed quantity
of quartz sand, and digests it in a beaker with water and filtered
saliva for some hours. He then heats the mixture to 100° C. on a
water bath, to precipitate soluble proteids, then weighs the beaker
and its contents, and determines the quantity of sugar which a
weighed quantity of the clear liquid contains, employing for this pur-
pose a Fehling's solution of which 1 c.c. corresponds to 1 milligramme
of dextrose. Assuming that the sugar formed from the muscle-
glycogen has the same reducing power as dextrose, and that it is
equally distributed throughout the scalded muscle and water, the
amount of glycogen originally present can be easily calculated.

Proportion The amount of glycogen found in different muscles

of glycogen of the same animal and in different individuals of the

same species, varies so much that no general statement

can be made. In Nasse's experiments the glycogen of

resting muscles of frogs amounted, on an average, to 0'43 per cent.

In rabbits the amount varied between 0'47 and 0'95 per cent.

Abeles' results were decidedly higher. As yet, however, the total

number of reliable determinations of the amount of glycogen in

muscle is too small to allow of any statement being made as to the

average amount of this constituent present.

We shall examine in a future section the changes in the amount
of glycogen brought about by the passage of muscle from the state
of rest into that of activity or rigor.

1 Nasse, Op. cit. p. 101 and 102.

336 DEXTRIN. SUGAR. IXOSIT. [BOOK I.

Dextrin. (C6H1006)H.

Limpricht1 and Scherer asserted that horse-flesh contains dextrin ;
the former observer obtained large quantities from the muscles of
young horses.

It may be taken as certain2 that the body was however glycogen-
dextrin, produced after death from glycogen.

Fermentable Sugar.

It was formerly believed that muscle in a state of rest contained
a small quantity of sugar. From the observations of Nasse3 it would
appear that sugar is only developed during activity or rigor and that
none is actually found in muscle at rest.

Inosit. C6H1206 + 211,0.

This non -fermentable isomer of grape-sugar was discovered by
Scherer4 in the muscular substance of the heart, and has since
been found in the voluntary muscles, of which it is said, however, not
to be an invariable constituent ; it is said to occur especially in the
muscles of drunkards. It is likewise present in the tissues of the
nervous system (Miiller), and in the lungs, liver, kidneys and spleen of
oxen (Cloetta6); it has been found in the kidneys of man, in the
urine of certain cases of Bright's disease, in the urine of diabetes
mellitus; in the liquid contents of hepatic hydatid cysts. Inosit
is found in many plants6, as in green kidney beans, the unripe fruit of
Phaseolus vulgaris (by Vohl, who gave to it the name Phaseomannite) :
in the green pods and unripe seeds of the garden pea (Pisum sativum) :
in the unripe fruit of the lentil (Ervum lens), and of the common
acacia (Robinia pseudacacia) : in the heads of the common cabbage
(Brassica oleracea, var. capitata) : in the herb of foxglove (Digitalis
purpurea) ; in the leaves and stem of dandelion (Taraxacum dens
leonis), not from the flowers or roots; in the shoots of the potato; in
the green herb and unripe berries of asparagus ; and' in two crypto-
gamic plants, viz. Lactarius piperatus, L. and Clavaria crocea, Pezs7.

1 Limpricht, Ann. d. Chem. u. Pharm. Vol. cxxxm. (1865) p. 295.
3 Nasse, "Chemie u. Stoffwechsel d. Mu skein." Hermann's Handbuch der Phy-
siologic, Vol. i. part i. p. 280.

3 Nasse, "Beitrage zur Physiologic der contraction Substanz." Pniiger's Archiv,
Vol. ir. (1869) p. 103.

4 Scherer, Annal. d. Chem. u. Pharm., Vol. LXXIII. (1850) p. 322.
6 Cloetta, ibid. Vol. xcix. p. 289.

6 This list of plants in which inosit occurs is copied verbatim from Watt's
Dictionary, Vol. in. p. 274.

7 As to the identity of the inosit from vegetable and animal tissues consult Janret et
Villiers, Comptes Rendus, Vol. LXXXVI. p. 486.

CHAP. IX.] THE CONTRACTILE TISSUES. 337

Perparation An aqueous extract of muscle (preferably of the

from Muscle. muscular tissue of the heart) is prepared. This is freed
joedecter's from aikumm by "boiling &c., then treated with baryta
water to free it from phosphates; it is concentrated, set
aside, and the creatine is allowed to crystallize out; the mother liquor
is boiled with four times its volume of alcohol; a precipitate is formed
which, according as it adheres to the bottom or separates in a floccu-
lent form, is separated by decantation or nitration. The clear liquid
is set aside for 24 hours, when crystals of inosit often separate ; if not,
ether is added and the mixture of alcohol and ether shaken again
and again ; inosit then separates out gradually in the form of leaflets
having the lustre of mother-of-pearl. An excess of ether does not
interfere with the precipitation, but merely causes the separating
crystals to be smaller (Hoppe-Seyler). The impure inosit obtained
by the above methods is collected on a filter, washed with cold
alcohol, and recrystallized from water.

Pr parties Inosit crystallizes in the form, of large, colourless,

roonoclinic tables, sometimes arranged in groups like
cauliflower-heads.

The crystals of inosit have a specific gravity of 1*1154 at 5° ; they
effloresce in dry air, or in vacuo ; at 100° C. the whole of the water of

FIG. 56. INOSIT FROM THE MUSCULAR SUBSTANCE OF THE HEART OF MAN. (FREY.)

crystallization is given off, and the anhydrous inosit melts at 110°,
setting, on sudden cooling, in fine needles. Inosit dissolves in 6 parts
of water at 19° 0. ; it is insoluble in absolute alcohol and ether.
Solutions of inosit when boiled with basic lead acetate yield instantly
a transparent jelly (containing the compound C6H12062Pb20) ; this
reagent has been, indeed, employed by some observers in the separation
of inosit.

Inosit has a sweet, saccharine, taste ; it is not fermentable ; it
does not rotate the plane of polarization ; it does not yield a yellow or

1 Boedecker, Ann. d. Chem. u. Pharm., Vol. cxvu. p. 118.
G. 22

338 INOSIT. FERMENTS. INORGANIC SALTS. [BOOK I.

brown colour when boiled with solutions of the caustic alkalies ; it
does not reduce Fehling's solution, but changes its colour to green.

Scherer's When inosit is treated with nitric acid, the solution

reaction. evaporated nearly to dryness, then moistened with

ammonia and a small quantity of calcium chloride, and again evapo-
rated, a rose-red colouration is produced. Scherer's test only succeeds
with nearly pure inosit.

Gaiiois' When a solution containing inosit is evaporated, at

reaction1. a gentle heat, nearly to dryness, then treated with

a small drop of solution of mercuric nitrate (the solution used in
Liebig's method for the estimation of urea answers very well) and
evaporated carefully to dryness, a yellowish white residue is obtained;
on further cautiously heating, the yellow changes to a deep rose-
colour, which disappears on cooling but reappears on again heating.
This constitutes a delicate and characteristic reaction, helping to
confirm the presence of inosit, the identification of which should
however depend also upon a knowledge of the conditions under which
the body was obtained, and upon such properties as crystalline form,
solubility, sweetness, &c.

Inosit yields when treated with nitric acid nitro-substi-
of inosit tution compounds which are soluble in alcohol and have the

composition C6H6(NO2)6O6 (Hexanitroinosit) and C6H9(NO2)3OC
(Trinitroinosit) ; these bodies explode when struck.

In the presence of decomposing proteids, inosit is decomposed, with the
formation of propionic, butyric and ordinary lactic acids (Yohl).

Proportion As has previously been stated, inosit is not an invari-

of inosit in able constituent of muscle. According to Jacobsen,
Muscle. horse flesh contains 0'003 per cent, of inosit.

The Ferments present in Muscle.

Muscle contains a trace of pepsin, as was shewn by Briicke. It is
perhaps in consequence of its presence that muscle so readily dis-
solves in very dilute hydrochloric acid. Dead muscle also contains an
amylolytic ferment which readily converts the muscle-glycogen into
sugar. Other hypothetical ferments have been surmised to exist in
order to explain the processes going on in muscle during activity
and rigor,

THE INORGANIC CONSTITUENTS OF MUSCLE.

The proportion of water in muscle varies between

74 and 80 per cent., the average being about 75. It is

said to be larger in young than in adult animals. The muscles of

cold-blooded animals contain more water than those of warm-blooded

animals.

1 Gallop De VInosurie, Paris, 1864. The Author has been unable to consult
this Memoir.

CHAP. IX.]

THE CONTRACTILE TISSUES.

339

The variations in the proportion of water which occur during
muscular contraction will be referred to in a subsequent section.

Mineral Fresh muscle yields on ignition from 1 to 1;5 per

Salts. cent, of mineral matters, containing as their principal

constituent potassium and phosphoric acid. 100 parts of the mixed
mineral matters contain about 36 parts of phosphoric acid. There
are also small quantities of calcium and magnesium phosphates and
traces of chlorine and iron. The remarkable preponderance of
potassium over sodium in the ash is to be remarked.

SUMMARY OF THE QUANTITATIVE COMPOSITION OF MUSCLE.

la the following Table, which is quoted from Hofmann1, the average amounts of the
various constituents in 1000 of the muscles of vertebrates is given.

Mammals.

Birds.

Cold-blooded
animals.

Solid constituents in 1000

217—255

227—282

200

Water „ „ „

745—783

717—773

800

Organic matters ,, „

208—245

217—263

180—190

Inorganic „ „ „

9—10

10—19

10—20

Coagulated albumin. Sarco-

lemma, nuclei, vessels

145_167

150—177

(*)

Alkaline albuminate

28-5—30-1

Creatine

2-0

3-4

2-3

Xanthine and Hypoxanthine

0-2

Taurine

0-7 horse

0-0

1-1

Inosit

0-03

Glycogen

4-1—5-0

30—50

Lactic acid

0-4—0-7

Phosphoric acid

3-4—4-8

Potash

3-0—3-9

Soda

0-4—0-43

Lime

0-16—0-18

Magnesia

0-4—0-41

Sodium Chloride

0-04— 0-1

Oxide of Iron

0-03—0-1

SECT. 2. GENEKAL PHENOMENA OF LIVING MUSCLE.

Muscle in a state of Rest

We have in the previous section made ourselves acquainted with
the normal appearances and composition of living muscular tissue.

1 Hofmann, Lehrbuch der Zoochcmie, p. 104.

22—2

S40 MUSCLE IN A STATE OF REST. [BOOK I.

General Ths maintenance of this normal — or in other words,

Phenomena the preservation of the life of the tissue from moment
of Resting to moment — is a physiological act consisting in suc-

Muscie. cessive and simultaneous degenerations and regene-

rations of parts. The apparent changelessness of repose depends
upon the regularity and equilibrium of many hidden changes. The
general nature of these changes is roughly indicated by comparing
the constitution of the blood flowing to and from muscles. The
blood enters muscle comparatively rich in oxygen and poor in carbon
dioxide; it leaves the tissue relatively poor in oxygen and rich in
carbon dioxide. Therefore the changes of degeneration and regene-
ration proceeding within the muscle are, collectively, changes in which
at least carbon and oxygen are implicated ; and further, they are, at
least in part, of the nature of oxidations. Hence it appears that a
supply of new matter to the tissue is ultimately indispensable. The
tissue is the channel of a continuous circulation or migration of matter ;
it is the theatre of constant material exchanges. The blood conveys to
it the substances which are needed; these are elaborated, rearranged,
and converted into other forms within the tissue ; and are finally again
cast out into the blood-current. These operations, wrought within the
tissue of muscle during repose, are included in the term Nutrition ;
which may be denned, in a figure now well known to Physiology,
as the sum total of the processes which maintain the * stock ' of the
organism at the normal. In reality these internal processes of
muscle at rest are but little understood ; most of our knowledge
of them is derived by inference from the processes of muscular con-
traction. But we may assume that they occur in at least three well-
defined stages. In the first, what may be called raw material is
received into the tissue and stored up in some proximate modification ;
in the second, this store is elaborated into an intermediate form by a
process independent of the process of storing ; and in the third stage
this intermediate body suffers decomposition into certain ultimate
products, some of which, but probably not all, are discharged into the
blood.

The grounds for assuming this threefold process will be understood
when the contraction of muscle has been discussed. It may be
remarked that it finds an exact analogy in the processes of glandular
tissues.

The transformations of matter within the substance of resting
muscle, though the more obvious, are not the sole phenomena of
nutrition. Running parallel with them are certain transformations of
energy. The energy implied in the mutual affinities of the elemen
involved in the transformations of matter, undergoes conversion in
energy of other forms during the various nutritive changes.

What these forms are, is again largely a matter of inference.
is unquestioned that heat is the most important. A short time ago
electrical inequalities or tensions would probably have been set down as
a second form ; but this is no longer admissible. The various mov

ot

:

?°

as

•

CHAP. IX.] THE CONTRACTILE TISSUES. 341

ments of small masses, where they occur, such as in the fission of
nuclei and of fibres, constitute a third form. A fourth, if entirely
hypothetical, deserves to be mentioned. It is not indeed another form
of actual energy, like heat or mechanical motion, into which the energy
of chemical affinity is converted ; but rather a re-distribution of
the original potential energy. The elaboration of the intermediate
product in the above series of tissue-changes is entirely unknown to us
in its nature ; but it is at least conceivable that it is not altogether
a process in which stronger affinities are satisfied at every step. It
may, in part, be a dissociation ; in which case some of the energy set
free in the chemical changes of the final stage may again at once
become potential.

The intensity of the chemical and physical processes of resting
muscle depends upon temperature, the supply of the necessary sub-
stances, and what may be called the nutritive instinct, or inherent
capacity, of the tissue for the changes. Both the supply of matter
and the capacity of the tissue for the changes in question, are exalted
in the active state of muscle.

Muscle in Action.

General The phenomena of resting muscle thus consist of

Phenomena of two parallel and associated series of transformations,
Contracting one of matter, the other of energy. The same dualism
is seen in active muscle. The whole phenomenon of
contraction comprises (1) a sudden acceleration and extension of
chemical decompositions, and (2) a sudden and extensive conversion
of the potential energy of chemical affinity into actual energy of
various forms. The manifestations of actual energy in the case of
contracting muscle are pronounced and admit of a careful study;
they assume the form of heat, electrical inequality, and mechanical
motion.

Special Contraction may be started either by the normal

Phenomena of stimulus proceeding from a nerve ; or by electrical,
Contraction. chemical, thermal, or mechanical stimuli applied to
muscle even in the absence of nerves. In its mechanical aspect con-
traction is a shortening and thickening both of the whole muscle and
of its individual fibres, associated in the case of entire muscles with a
small reduction of bulk. Contracted muscle is less elastic and more
extensile than resting muscle.

Micro- Viewed under the microscope, the act of contraction

scopic ap- falls into well-marked stages. In the first, the bands

pearances. draw near together as the muscular fibre shortens ; and
the dark and light bands approximate in tint, until the whole fibre
is evenly dark with little or no striation: this is called the homo-
geneous stage. As contraction proceeds, striae again appear, but this

342 MUSCLE IN ACTION. [llOOK I.

time in a regular alternation of simple dark and simple bright bands.
There is however this difference : those parts of the fibre which
before were dark are now bright, and those parts which formerly
were bright are now dark. The fibre has emerged from the homo-
geneous stage with its bands interchanged as far as regards their tint ;
and this, therefore, may be called the stage of transposed lands. Their
transposition only affects the shade or tint of the stripes, the isotropous
and double-refracting elements of the fibre maintaining their original
relationship. This is well shewn in the figure on p. 314, where the
same contracting muscle is exhibited by ordinary and by polarized
light

It may be added that Eugelmann believes he has demonstrated that,
in contraction, the volume of the main double-refracting zone increases
at the expense of the isotropous layers.

If contraction is started at one point with a given
intensity it does not instantly extend over the whole fibre;
but travels along it as a wave with a velocity of 3 metres a second in
frogs. In the case of excised muscles, the wave suffers in its course a
diminution1 of intensity.

Latent When muscle is directly stimulated the contractile

period, force does not at once begin to develope. An interval

elapses between the application of the stimulus and the beginning of
contraction ; this is known as the period of latent stimulation, or latent
period. Its value was determined by Helmholtz to be about ^ sec.,
but it is found to vary in different circumstances, and under favour-
able conditions it is said to become as short as ^ or ^fa of a sec.2

Course, or When once started, the force does not spring

Curve, of suddenly to perfection, but developes in course of time.

Contraction. if a resistance be opposed to the shortening of the
muscle it is clear that no contraction can occur until the contractile
force has grown large enough to overbalance the resistance ; hence,
the greater the resistance, the longer the interval which must elapse
between the moment of stimulation and the beginning of actual
contraction. During contraction the contractile force does work and
becomes spent. Hence a smaller resistance serves to check contraction
near its end than near its beginning. The rapidity of contraction is
not equal throughout its course; it first increases and then diminishes
until the summit of contraction is reached, as it usually is within
^ or jlfo- of a second after stimulation. Beyond the maximum
the contractile force dies gradually away3; this is rendered probable
by the course of re-extension under the influence of a weight, which

. J See Hermann's Handbuch der Physiologie, Bd. i. Abth. i. p. 55.

3 See Hermann's Handbuch, Bd. i. Abth. i. p. 36.

3 Heidenhain, "Ueber Ad. Pick's experimentellen Beweis fiir die Giiltigkeit des
Gesetzes von der Erhaltung der Kraft bei der Muskelzusammenziehtmg." Pfluger's
Arch., Vol. ii. p. 426.

CHAP. IX.] THE CONTRACTILE TISSUES. 343

is not such as to admit of the assumption that gravity alone deter-
mines it; and it becomes more than probable when we learn that
the heat-developing processes1 and the chemical processes2 are also
carried over into the relaxing period. The duration of the period of
relaxation is about ^ of a second. Muscle appears to have no power
of active re-extension3.

Tetanus When stimuli are thrown into muscle with sufficient

rapidity, contractions overtake one another, sum their
effects, and maintain the muscle against extending forces, in a
position more or less of maximum contraction. Such continued con-
traction is called a tetanus, the laws of which belong to the physics
of muscle.

The course of contraction is not similar in all kinds of muscle;
nor is the course the same in any one muscle under all circumstances.
Thus the rate of contraction is much quicker in the muscles of
insects than in those of frogs ; and quicker in the latter again than
in those of the tortoise ; and in these than in the heart-muscle ; and
in this than in the smooth muscles of the intestines or ureter. Such
differences are of the greatest interest to the physiologist as indicating
either differences in the machinery for the conversion of energies,
or different capacities for the chemical changes upon which contraction
depends.

Red and In respect of such internal machinery, or capacity, the

Pale striated differences of red and pale striated muscles are remark-
Muscles, able. The contraction of the red variety is slow and
enduring, 10 stimuli a second being enough to cause almost unbroken
tetanus; while of the pale variety, the contractions are short and sharp,
20 — 30 stimuli a second being needed for a perfect fusion of them4. In
the former the latent period is so long as TXH sec. ; while in the latter
it has the value of ^ sec.3.

These physiological differences are all associated with varieties of
structure; but differences of a similar nature may be exhibited by the
same muscle when it contracts under varied conditions. Thus cold,
many poisons, and incipient exhaustion prolong contraction and
diminish its amplitude. Indeed the stimulus of a sharp blow to dying

idio-mus- muscle often produces a local contraction which may be
cuiar Con- likened to a wheat, and which may persist for a long
traction. time : such a contraction is described as 'idio-muscular'.

Absolute The force of muscular contraction is measured by

force. the weight which is just sufficient to prevent the

1 Steiner, "Ueber die Warmeentwicklung bei der Wiederausdehnung des Muskels. "
Pfliiger's Arch., Vol. xi. p. 196.

2 Heidenhain with Landau and Faculty, Loc. cit. Pfliiger's Arch., Vol. n. p. 429.

3 Kuhne, Loc. cit. Arch. f. Anat. Physiol. u. wiss. Med. (Reichert u. du Bois-
Reymond), 1859, p. 815.

4 Kronecker and Stirling, "The Genesis of Tetanus." Journal of Physiol. (Foster)^
Vol. i. p. 395.

5 Ranvier, Loc. cit. Arch, de physiol. norm, et path., 2 s6r. Vol. i. p. 5.

344 MUSCLE IN ACTION.- [BOOK I.

shortening of the muscle \ The force varies with the stimulus : as
this gradually increases, that enlarges, quickly at first but after-
wards more slowly, until a maximum is gained, which is known as
the ' absolute force '. The absolute force is usually stated to be 2800 —
3000 grms. per sq. centimetre of tetanized frog-muscle ; and between
6000 and 8000 grms. per sq. centimetre in the muscles of man
voluntarily contracted2.

These numbers cannot be taken as the direct or exact equivalent
of that portion of the chemical changes which is devoted to me-
chanical effect. For the result so obtained is less than the true
absolute force of the muscle experimented on by an amount which
depends on its extensibility. If muscle were more elastic than it is,
although the process of contraction with all its chemical changes re-
mained the same, the absolute force would seem to be less. This may
readily be demonstrated by interposing an elastic band between the
muscle and the weight about to be raised. The absolute force of such
a system is less than that of the muscle alone; whence we may conclude
that the proper extensibility of muscle has a like diminishing effect3.

If a loaded muscle be made to contract by the application of
a stimulus, the height through which the load is raised is called the
lift; and this multiplied into the load gives the value of the mechani-
cal work actually done. As the stimulus is increased, the lift grows
proportionally to the stimulus up to a maximum, beyond which
it remains constant4. Inasmuch as muscle is extensible, and
its extensibility is increased during the state of contraction, it is
clear that the lift is the expression of the actual shortening of
the muscle minus the difference between the extension of the
uncontracted and the extension of the contracted muscle. For the
full illustration of this the reader is referred to Ed. Weber's article
'Muskelbewegung' in Wagner's Handwarterbuch, and to the Text-
books of Physiology.

The lift varies also with the load, becoming smaller as the load
increases; and the variation is such that the product of lift into load
first of all increases and afterwards decreases as the load varies from
nothing onwards. In other words, within certain limits, the more a
muscle is weighted the more mechanical work will a given stimulus
produce. The increased tension to which the muscle is for the time
subjected converts it into a body capable of yielding a larger amount
of mechanical work than the same muscle less tense. Not only does
the state of tension in the very act of contraction influence the work
done, but the state of tension immediately prior to contraction has
the same effect : the greater the tension the greater within certain
limits6 the yield of mechanical energy.

1 E. Weber, Wagner's Handivorterbuch, in. 2, p. 84. Helmholtz, Arch. f. Anat.
Physiol. u. wiss. Med. (Miiller), 1850, p. 276 ; 1852, p. 199.

2 See Hermann's Handbuch, Bd. i. Abth. i. p. 64.

3 See Hermann's Handbuch, Bd. i. Abth. i. p. 65.

4 Fick. See Hermann's Handbuch, Bd. i. Abth. i. p. 108.

5 Heidenhain, Mechanische Leistung, Wamieentwicklung und Sto/umsatz lei
Muxkelthatigkeit. Leipzig, 1864, p. 84.

CHAP. IX.] THE CONTRACTILE TISSUES. 345

Maximum The maximum work done under most favourable

work. conditions is said to vary between about 3500 and 5500

gram-meters per gram of frog-muscle1.

Heat of Con- The mechanical motion of contraction is not the

tracting only exhibition of kinetic energy which accompanies

Muscle. t^ chemical changes of acting muscle. During tetanus2

and in single contractions3, the muscles become raised in temperature ;
and since this occurs in muscles removed from the circulation or
even in muscles entirely removed from the body, it must be due
to the heat-developing processes of the tissue itself.

Helmholtz, in a 2 — 3 minutes-tetanus through nerves, found the
thigh of a frog raised '14 to "18° C. ; and Heidenhain observed the
temperature of the gastrocnemius to be raised '001 to '005° C. in
a single contraction.

If the weight of the gastrocnemius be known, and also the specific
heat of muscular tissue, it is possible to estimate in heat-units the amount
of heat generated in one contraction. Fick4, taking the specific heat
of muscle to equal that of water, found that in one energetic contraction,
under most favourable conditions for activity, every gram of the contracting
muscle generates heat enough to raise 3'1 mgr. of water through 1° C.
The specific heat of muscle is however stated to be '7692 by Adamkiewicz5,
and -825 by Eosenthal6.

The evolution of heat in contraction is amenable to the same
influences as the evolution of mechanical work ; but though amen-
able in the same sense it is not so in the same degree. Thus, as the
stimulus gains in strength, not only does the lift become higher,
but the heat liberated is also increased, with this difference, that
the heat evolved is increased more rapidly than the lift7. So also the
greater the tension of a muscle, whether before or during contraction,
the greater, within bounds, will be the heat evolved as well as
the work done ; but here again, as the tension increases, the heat
evolved reaches a maximum and begins to decline sooner than
the mechanical effect8.

1 Hermann's Handbuch, Bd. i. Abth. i. p. 79.

2 Bunzen in Gilbert's Annalen, 1807, vol. xxv. p. 157 : quoted in Heidenhain, Mecha-
nische Leistung, etc., p. 33, where also will be found an account of the earlier researches
in which the heat of the body, and of muscles within the body, was found to be in-
fluenced by exercise. Helmholtz, " Ueber die Warmeentwicklung bei der Muskelaction. "
Arch. f. Anat. Physiol. u. tviss. Med. (Miiller), 1848, p. 144.

3 Heidenhain, Mechanische Leistung, Warmeentwicklung und Stoffumsatz bei Mus-
kelthdtigkeit. Leipzig, 1864, p. 73.

4 Fick, "Ueber die Warmeentwicklung bei der Muskelzuckung." Pfliiger's Archiv,
Vol. xvi. p. 84.

5 Adamkiewicz, "Die Warmeleitung des Muskels." Arch. f. Anat. Physiol. u. wiss.
Med. (Eeichert and du Bois-Eeymond), 1875, p. 254.

c Rosenthal, "Ueber die specifische Warme thierischer Gewebe." Monatsber. d.
Berliner Acad., 1878, p. 307.

7 Nawalichin, " Myothermische Untersuchungen." Pfliiger's Archiv, Vol. xiv.
p. 295.

8 Heidenhain, Mechanische Leistung, etc. p. 84 et seq.

346 HEAT DEVELOPED BY CONTRACTING MUSCLE. [BOOK I.

When muscle becomes exhausted, both the work done and the heat
generated decline; but the latter more quickly than the former1. The
cause of this dissimilarity may be one or other of the two following. It
may be that the heat-evolving and the work-evolving appliances in
muscle are totally distinct, and variously affected by the same condi-
tions. Or it may be that the heat and mechanical work of muscle, like
the heat and mechanical work of a steam-engine, arise in a common
fundamental combustion ; and that the relative proportions of the
two are to some extent determined by external conditions, just
as some steam-engines work more economically than others, i.e. with
a larger proportionate yield of mechanical work2.

It is impossible to say whether heat-developing processes are oc-
curring during the latent period3; but there is little doubt that they
continue beyond the period of maximum contraction. It is at least
certain that the heat developed in a muscle is influenced by the load
which it bears during relaxation as it is by that which it bears in
contraction ; and such influence cannot be explained as the result
of mere forcible extension4.

The proportion of heat and work evolved in contraction has been
determined by Fick5 in the case of excised frog-muscles to vary
according to the load ; the greater the load the larger the proportion
of the total actual energy taken up by mechanical work. Under the
most favourable circumstances for the performance of mechanical
work the relation of work to heat was 1 : 3'8, and in the least
favourable of Fick's experiments the relation was 1 : 23'(?. It is
extremely uncertain how far these fractions can be applied to muscles
within the body, or to the muscles of warm-blooded animals.

Fick's demonstration of this interesting relationship depends upon the
fact that, when the motion of a falling body is suddenly arrested, an
amount of heat appears, equivalent to the mechanical motion destroyed.
By direct experiment he proved that, if a weight suspended from a muscle
is raised by external means to a certain height and then let fall, the muscle
suffers a heating proportionate to the fall, i.e. which is the precise
equivalent of the work done in lifting the weight. He therefore caused
a loaded muscle to contract and afterwards allowed it to re-extend under
the weight which it had lifted; and then observed by how much the
temperature of the muscle had been raised. From the specific heat of
muscular tissue he was able to calculate the total quantity of heat gained
by the muscle in the process; and by subtracting from this total the heat-
equivalent of the work done in raising the weight, he was able to compare
the heating of a muscle under a certain load and the work done in raising
the load.

1 Heidenhain, Mechanische Leistung, etc. p. 74.

2 See Hermann's Handbuch der Physiol, Ed. i. Abth. i. p. 168.

3 Nawalichin, Loe. cit. Pfliiger's Archiv, Vol. xiv. p. 311.

4 Steiner, Loc. cit. Pfliiger's Archiv, Vol. xi. p. 204. See also Heidenhain with
Landau and Pacully, Loc. cit. Pfliiger's Archiv, n. p. 423.

5 Fick, Loc. cit. Pfliiger's Archiv, Vol. xvi. p. 79.

CHAP. IX.] THE CONTRACTILE TISSUES. 347

Electrical The third form under which the actual energy

tensions of of contracting muscle appears, is that of electrical
contracting disturbance. Muscles within the body, or absolutely
uninjured muscles, are electrically homogeneous: they
exhibit no current1. But whenever a stimulus is applied to a
muscle, the spot stimulated assumes a lower, or negative, potential
as compared with the rest of the muscle; so that if the two electrodes
of a galvanometer were applied to an excited and a non-excited spot
of muscle respectively, a current would be discovered. This func-
tional current increases to a maximum very rapidly and afterwards
disappears, but more slowly. It begins instantly on stimulation, i.e. it
has no latent period; and the whole phenomenon lasts about ^
sec. Hence it falls entirely within the latent period of contraction.
It travels down excised muscles from the point of stimulation with a
velocity which agrees with that of the contraction-wave, namely
about 3 metres a second in the frog. In the normal muscles of the
human fore-arm the velocity has been determined to lie between 10
and 13 metres per sec. Like the wave of contraction, the negative
wave diminishes in intensity during its course along excised muscles;
but no such diminution has been detected in the case of muscles in
which the processes of restitution are active2.

As is the case with the evolution of heat and mechanical
effect, the disturbance of electrical tension which follows stimulation
differs in degree according to the different conditions of stimulus,
irritability and tension. It increases up to a maximum as the
stimulus becomes more and more intense; it diminishes as exhaustion
approaches3; it increases with the lift4; and it varies directly as
the tension of the muscular fibres5.

Rigor Mortis.

Besides the conditions of rest and activity, there is a third
condition of muscular tissue with characteristic phenomena and a
singular bearing on the theories of muscular function, viz. the moribund
condition.

After the death of the body, or after the ligature of their tributary
arteries, or on subjection to a certain temperature, muscles become
rigid. That is to say, they become shorter and thicker, and of less

1 Hermann, "Ueber das Fehlen des Stromes in unversehrten ruhenden Muskeln."
Pfliiger's Archlv, Vol. in. p. 35.

2 Bernstein, "Ueber den zeitlichen Verlauf der negativen Schwankung des Muskel-
stromes," Monatsber. d. Berliner Acad., 1867, p. 444. Untersuchungenu. d. Erregungs-
vorgang im Nerven- u. Muskelsystem. Heidelberg, 1871. Hermann, "Ueber den
Actionsstrom der Muskeln im lebenden Menschen." Pfliiger's Archiv, Vol. xvi. p. 410.

3 Hermann, Handbuch der PhysioL, Bd. i. Abth. i. p. 220.

4 Harless, Gel. Anz. d. bayr. Acad., xxxvn. p. 267, 1853; quoted by Hermann,
Handbuch, Vol. i. Abth. i. p. 220.

5 S. Lamansky, "Ueber die negative Stromesscliwankung des arbeitenden Muskels."
Pfliiger's Arch., Vol. in. p. 202.

348 CHEMICAL CHANGES OF LIVING MUSCLE. [BOOK I.

bulk, as in the act of contraction. The lift of a muscle passing
into rigor is greater with a small load, but less with a heavy load,
than during a single contraction, and the absolute force is in the same
circumstances sometimes greater and sometimes less. No similar
comparisons have yet been made between rigid and tetanized muscles1.

Rigid muscle is less extensile, as well as less elastic, than normal
resting muscle, thus differing from contracted muscle, which is more
extensile. It is, farther, distinguished from contracted muscle by its
peculiar doughiness and opacity.

Rigor is associated with the evolution of heat — post mortem
elevation of temperature. This is doubtless in part a mere conse-
quence of the physical changes of density, and the transformation
from the fluid to the solid state. But a physical explanation will not
account entirely for the phenomenon ; for no rise of temperature can
be detected during the quasi-rigor — which is a simple coagulation —
induced by acids or alcohol, and true rigor is unquestionably attended
by chemical changes2.

The passage into rigor is further associated with a difference of
electric potential; dying muscle, like contracting muscle, is negative
to normal resting muscle3.

Thus the last event in the life-history of muscle resembles a
common contraction very closely in the nature of its physical
phenomena. We shall find that they are alike also in their chemical
changes.

SECT. 3. SPECIAL STUDY OF THE CHEMICAL CHANGES OF LIVING

MUSCLE.

It has been pointed out that the whole life of muscle consists
of two parallel series of transformations, of constitution and of
energy. The characters of one series, the transformations of energy,
have been rapidly sketched ; and it now remains to describe
in detail the changes of the other, or chemical series. It may at
once be stated that our knowledge of these two series is, and
must be, of very different extent. In the case of the physical trans-
formations wre are able to study their course in time, to fix their
maximum and trace their decline. In the case of the chemical
series the steps are entirely hidden ; we can merely compare the
constitution of a muscle before and after, but not during, an act of
contraction. We cannot say whether the chemical changes run pari

1 E. Walker, "Die absolute Kraft der Erstarrung." Pfliiger's Arch., Vol. iv. p. 186.

2 Hermann, Handbuch der Physiol, Vol. i. Abth. i. p. 171. Schiffer, " Ueber die
Warmebildung erstarrender Muskeln." Arch. f. Anat. Physiol. u. wiss. Med. (Keichert
and du Bois-Keymond), 1868, p. 442.

3 For a full account of the demarcation-current and its relation to the so-called
natural muscle-current of du Bois-Reymond, see Hermann's Handbuch, Vol. i. Abth. i.
p. 173 et seq.

CHAP. IX.] THE CONTRACTILE TISSUES. 349

passu with the physical phenomena of work, heat, and electrical
disturbance, which are in some fashion linked to them; or whether
the contraction of a muscle is not rather like the firing of a gun,
in which the progress of the bullet affords no clue whatever to the
course of the explosion.

Methods of Since then the chemical history of any event in the

the chemistry life of muscle rests on an analysis of chemical constitu-
of Muscle. tion before and after the event; and since certain of the

constituents of muscle may be exhaled into the surrounding medium ;
it is clear that the chemistry of living muscle comprises two lines of
enquiry :

1. Into the chemical composition of the muscle itself.

2. Into the chemical composition of the medium surrounding

the muscle.

These have for the most part been carried on independently; and
the latter has, beyond question, led to the more important results.
The enquiry into the chemical composition of the medium sur-
rounding muscle has been followed under two sets of conditions,
not however essentially different : the simpler, in which the muscles
are exposed to the air as a medium; the more complex, in which
the muscles remain in the body, or in which the blood is the surround-
ing medium. In the latter circumstances the enquiry is complicated
by the occurrence of institutional changes. Finally, when muscles
are examined while still within the body there are two ways of
obtaining a knowledge of the changes in their surrounding medium,
viz. by contrasting before, during, and after a muscular act,

1. The blood of muscle,

2. The general excreta.

While this summary includes all the methods of muscular chemistry,
it is necessary to state that they have not been equally applied to
living muscle in each of its three possible conditions, the resting, the
active and the moribund. The latter two conditions have been most
freely investigated, and it will be convenient to describe the results of
their investigation together, since they have much in common ; and
before the results of the examination of the normal state of rest are
stated.

THE CHEMICAL CHANGES OF CONTRACTION AND KIGOR.

A. Changes in the chemical composition of muscle itself.

Changes in the gaseous constituents1.

Apparatus. The air pump, which has proved so valuable a means of

research in the chemistry of the blood, has been also

The first rudimentary attempt at the gaseous analysis of muscle which the
author has met with, is described in a memoir sur V Irritability, by Girtanner, contained

350 GASEOUS ANALYSIS OF MUSCLE. [BOOK I.

employed in the analysis of muscle, but with much greater difficulty.
The difficulty is due in part to the nature of the method, for the
muscles cannot be transferred to the vacuum without preliminary
exposure to the contamination of air and indifferent fluids; and in
part to the nature of fresh muscle, whose tissue entangles bubbles
of gas, and whose gaseous contents, owing to the acidification of rigor
(p. 359), and to putrefaction, rapidly undergo change even at ordinary
temperatures.

For the analysis of muscle a special boiling-flask is necessary, such
for example as is figured in the following diagram.

A and B represent two views of the same apparatus, and the letters
are identical in their reference.

v is the froth-chamber, a globe provided with a short neck g, fitting on
to the drying-chamber t but shut off from it by the stop-cock b. It is
provided also with a longer neck, h, at right angles to the other, interrupted
by a stop-cook c, and fitting into the boiling flask f.

f is the boiling-flask, with a rounded bottom and a wide neck; it
is provided with three platinum wires melted through the sides and
reaching almost to the bottom of it. It is fitted on to the neck, h, of the
froth-chamber not quite at right-angles, as B shews, and in a plane at right-
angles to that of the neck g. f contains the muscle to be exhausted ; and at
the mouth of the neck h is a cork k grooved at the sides to permit the
passage of gases from f to v, while stopping any solid fragments which
might do damage to the stop-cock c.

v serves a double purpose besides that of a froth-chamber : Firstly, any
liquid which spirts over from f during ebullition is collected here, and may,
by turning v round the axis of its neck g, be made to trickle back into f.
Secondly, a reagent, such as an acid, may be kept in v during the
preliminary exhaustion of a muscle in/, and by a similar tilting of v, may
be brought to play on the muscle at any given moment.

t is a small drying chamber containing sulphuric acid, sufficiently large
to keep the vacuum of the pump dry so long as the stop-cocks c and b are
never open together ; by this means the access of watery vapour to the
absorption tube of the gas-analyser is prevented. The capacity of the
boiling-flask /, and the part of the neck h up to the stop-cock c, may
be about 200 c.cm.1

in Eozier's Observations sur la physique, Vol. xxxvii. 1790, p. 148. Muscle, cut into
small pieces, was enclosed in a glass retort connected with a pneumatic apparatus. A
very gentle heat was applied by means of a lamp for more than two hours, and the
gases which passed over into the pneumatic receiver were examined at different stages
of the experiment. At first atmospheric air passed over "mele* a une tres-petite
quantity d'air vital, dont le gaz nitreux indiquoit la presence ; " the second portions were
vital air, "mele" a du gaz acid carbonique." Girtanner very innocently remarks: "On
peut retirer la m£me quantit6 de ce gaz [vital air or oxygen] plusieurs fois de suite, en
exposant des substances animates alternativement a 1'air atmosphe'rique et a une
chaleur de 60 a 70 degr6s du thermometre de Kdaumur." He found the exact adjust-
ment of the temperature a matter of great difficulty: "Si Ton applique un degre" de
chaleur trop fort, on aura du gaz acide carbonique au lieu de gaz oxigene." The
fallacies of the method lie on the surface, but do not destroy the historical interest of
the experiment. Girtanner further found that he could extract almost all the oxygen
which animal substances contain "par le moyen de 1'eau chaude. "

1 Hermann, Untersuchungenu* d. Stojfwechsel der Muskeln ausgehend vom Gaswechscl
denelben. Hirschwald, Berlin, 1867, p. 4.

CHAP. IX.]

THE CONTRACTILE TISSUES.

351

FIG. 57. HERMANN'S APPARATUS FOR THE EXTRACTION OP THE GASES OF MUSCLE.

Gaseous If the muscles of a frog deprived of blood be quickly

analysis of removed from the body (and frog-muscles, for obvious
'scalded' reasons, are made use of in these experiments); and if

they be then plunged instantly into a large volume of
briskly boiling salt-solution of indifferent strength, they will be coagu-
lated at once throughout their mass, and die without preliminary rigor

352 GASEOUS ANALYSIS OF MUSCLE. [BOOK I.

and acidification. This method is technically known as scalding. If
an inverted beaker be placed in the large dish of boiling fluid and very
accurately filled with the salt-solution or its steam, to the exclusion
of all air, the muscle may be thrown at once beneath it, and any gas
which escapes from the muscle in the process may be collected.
Under these circumstances it appears that scalded muscle, during
the process of scalding, loses no appreciable quantity of gases. If
the muscle is now reduced to a low temperature, and minced
to prevent the mechanical entanglement of gas-bubbles ; and then
subjected to the influence of a vacuum ; it is found to yield a small
per-centage of gases. If, after a first evacuation, phosphoric acid is
added to the minced muscle by tilting the froth-chamber, a further
escape of gases follows. In both cases the gas is carbon dioxide.
Thus scalded muscle — muscle in which the process of rigor has been
circumvented, and which may therefore be regarded as presenting the
gases of fresh normal muscle — contains an extremely small quantity
of carbon dioxide, both free (i.e. capable of withdrawal by the air-
pump), and fixed (i.e. needing an acid to drive it out). The former
may amount to 2'74 per cent., and may be due, in part, to the im-
perfect scalding of the central portions of the muscle; for, if the
temperature is not high enough, rigor follows, and not instant death ;
and rigor is associated with acidification and the production of carbon
dioxide. The fixed carbon dioxide may amount to 1*95 per cent.1

Gaseous -^ ^e a^ove figures be taken as indicating the gases

analysis of of fresh normal muscle, we shall observe a marked
muscle difference in muscle which is passing into rigor. A con-

passing into venient method of producing the gases of rigor, and at
rigor' the same time facilitating their liberation from the mus-

cular substance, is the following. Frogs, whose blood-vessels have
been well washed out with *5 per cent, solution of NaCl, are taken
into a cold atmosphere, and their belly-muscles and muscles of the
hinder limbs (excluding the feet and tendons) are quickly cut away
and weighed on a watch-glass. They are then placed, still on the
watch-glass, over a freezing mixture until they are frozen to a firm
mass ; and afterwards they are minced with cold knives and rubbed
up in a cooled mortar. The freezing preserves the normal composition
of the muscle more or less perfectly during the mincing and tritura-
tion2 ; and these processes are devised to facilitate the disentangle-
ment of the gas during evacuation.

The frozen and triturated muscle is introduced into the boiling-
flask which is filled to the brim with normal NaCl solution at
0° C., the various portions being so quickly dropped in that a constant
overflow of salt-solution is kept up. The object of this manoeuvre
is to wash away the air-bubbles which are carried into the salt-

1 Hermann, Op. cit. pp. 115, 116. Expt. 9.

2 Kuhne, Untersuch.u. das ' Protoplasma, Leipzig, 1864, p. 3. See Hermann, Stoff-
wechsel der Muskeln, p. 5.

CHAP. IX.] THE CONTRACTILE TISSUES. 353

solution clinging to the frozen muscle, and which, becoming
disengaged, rise to the surface. This simple method is more
successful in preventing the intrusion of air-bubbles than that of
introducing the muscle into the boiling-flask under a surface of
mercury. When all the muscle has been introduced, the boiling-flask
is at once attached to the froth-chamber, the stopcock c being closed,
and the greatest care being taken not to include bubbles of air.
Salt-solution may be used either after shaking it up with the air of
the room, or after exhaustion of its dissolved gases by the air-pump;
in the former case it is necessary to ascertain the gaseous impurities
of the salt-solution and allow for them at the close of the experiment.

If it is desired at any time during the experiment to treat the
muscle with acids, the acid must be carefully placed in the froth-
chamber v, before its neck g is attached to the pump. Meanwhile
the boiling-flask / is surrounded by a freezing mixture in the position
figured in B.

It will be seen that the whole object of the experiment is to boil
a mixture of salt-solution and muscular tissue reduced to as fine a
state of division as possible; the preparation (which may occupy
about two hours) being made at a temperature least favourable to
spontaneous changes of the tissue to be analysed.

The muscle having been lodged in the apparatus for collecting the
gases, all that is necessary is to induce rigor ; and this may most readily
be done by raising the temperature of the boiling-flask. The froth-
chamber v is then made vacuous and the gases which boil over are
passed through the pump into the absorption-tube for analysis. At a
temperature of 0° C. little or no gas is given off, and then only after
several hours exposure. As the temperature rises to 15°C. there
is an indefinite, dribbling discharge. At temperatures beyond this,
up to 30° C., there is at first a large escape of gas which afterwards
subsides. But it is at still higher temperatures, of 40 — 50° C.,
that the greatest discharge occurs; here, also, it is voluminous
at first, becoming less and less as the exposure continues. At this
time the muscles have become acid and have ceased altogether to be
irritable. When the temperature has been raised beyond 70° no further
yield of free gases is obtained.

If the muscle is treated with phosphoric acid at 0°C., and sub-
sequently heated to assist the liberation of gas, a sharp evolution
occurs, of short duration and yielding but a small amount. If
the acid be added to a preparation subjected to a temperature of
20° — 50° C., at a time when the discharge has ceased to be volu-
minous, a brief acceleration of the discharge will result, followed by
complete and final stoppage.

secondary, If the muscle is kept in the apparatus beyond

or putrefac- the time at which the first discharge subsides, the

tive, dis- liberation of gases begins again, even without the

addition of an acid. This constitutes a secondary

discharge, and is due to putrefaction; it may begin

G. 23

354

GASES LIBERATED IN THE RIGOR OF MUSCLE. [BOOK I.

Nature of
gases libe-
rated by
muscle in
rigor.

within a few hours of the commencement of the experiment, and
must not be confused with the primary discharge. "We need not
here further discuss it1.

The gases of the primary discharges obtained from
muscle raised from 0° to temperatures varying from 5°
to 70°, added to the gases liberated by phosphoric acid,
vary from 1 to 15 vols. per cent, of the muscle used.
These gases contain no oxygen whatever ; on the
contrary, if salt-solution containing dissolved air have been used,
some of the oxygen will have disappeared from solution. Nitrogen is
constantly present in small amount in the portions of gas first set
free; but subsequently no nitrogen can be detected until the
secondary discharge begins. Carbonic anhydride is the chief, and
indeed the only constituent of the gas discharged during the middle
of an experiment2.

The following experimental numbers will serve to illustrate these
conclusions.

Experiment. 57*3 grms. ( = 54*16 c.cm.) of frozen and triturated frog-
muscle (the muscles of three or four frogs may be used) were exhausted
in boiled salt-solution at a temperature of 50° C. After the cessation of
the primary discharge, phosphoric acid was added, and the gases collected at
a temperature of 60°, as long as they continued to be evolved.

PORTION I. BEFORE ADDITION OF ACID.

Constitu-
ents in
c.cm.
(0°C.and
Imtr.)

Per-cen-
tage of
muscle.

Total gas

7-051

COa (free)
N + error
O

6-385
0-666

o-o

11-79
1-23

o-o

PORTION II. AFTER ADDITION OF ACID.

C0a (fixed)

1-105

2-04

Total CO2 free
and fixed

7-490

13-83

Experiment. 34-2 grms. (= 32-33 c.cm.) of frozen and triturated frog-
muscle, in boiled salt-solution, exhausted at 0°C. yielded no gas. Phos-

1 Hermann, Op. cit. p. 11.

2 Hermann, Op. cit. p. 10.

CHAP. IX.]

THE CONTRACTILE TISSUES.

355

phoric acid was then added without causing any discharge. The acidified
muscle was heated to 50°C., and thereupon liberated quickly a quantity of

Constitu-
ents in
c.cm.
(0°C. and
Imtr.)

Per-cen-
tage of
muscle.

Total gases

4-025

C02
N + error

2-222
1-803

Q-S7
i

Thus the change in gaseous composition which muscle undergoes
on passing into rigor may be summarized as a large increase of that
carbonic anhydride which is denned as free, or capable of simple
withdrawal by an air-pump.

The discharge of gases is a primary phenomenon of rigor and
not due to the decomposition of carbonates already existing in the
muscle, by the acid formed in the same process. For the addition of
phosphoric acid at a time when the discharge is free, tends rather to
diminish the total discharge than increase it, and never leads to an
evolution of gas proportionate to the yield of free carbon dioxide. In
other words, the. carbon dioxide is formed step by step with the pro-
cess of rigor ; but, although it is actually formed during rigor, it may
still first appear in some fixed and stable modification ; and this is a
possibility which we have as yet no means of testing 2.

Gaseous With very similar appliances to those just described

analysis of the gaseous alterations of muscle during activity may
contracting be determined ; but this research is beset with great
difficulties, if a complete analysis of the gases is
desired. In the first place it is indispensable to employ boiled salt-
solution and to use scrupulous care in excluding air-bubbles. In the
second, there is great danger of electrolytic action due to the strong
currents needed to stimulate muscles immersed in salt-solution. And
in the third, it is impossible to adopt wholly the method of tritu-
ration in order to facilitate the escape of gas in the vacuum ; all
that may be attempted for this purpose is to mince the muscle
coarsely, or to select muscles of small bulk like the sartorius. But
many of the difficulties of experiment may be avoided after it has once
appeared that, as in the case of rigor, carbon dioxide is the only
important constituent. No oxygen is ever detected ; and the nitrogen
evolved behaves like the nitrogen set free in the rigor of muscle.
Hence it is at once possible to dispense with the troublesome salt-
solution and the strong tetanizing currents ; and to examine muscles

1 Hermann, Op. cit. p. 114.

Hermann, Op. cit. p. 16.

23—2

356 GASES LIBERATED IN CONTRACTION OF MUSCLE. [BOOK I.

stimulated in a vessel of air. The method of the experiments is to
take a preparation of frog-muscles and expose it before, during, and
after tetanus under exactly identical conditions, collecting the yield of
gases separately, for comparison. For this purpose the pelves and
hinder extremities of three or four frogs divested of their skins may be
arranged to form a chain attached at each end to a platinum electrode
of the boiling-flask/. At the bottom of this flask is a little normal
salt-solution to keep the atmosphere moist ; salt-solution being pre-
ferred to water in order to defend the preparation from injury during
the accidental spurtings of the fluid. The temperature of the boiling-
flask is carefully maintained constant throughout the experiment,
at about 16 — 20° C. The muscles are first exposed to a vacuum for an
hour and the gases (A) collected. They are then tetanized at intervals
during another hour and the gases (B) again collected, care being
taken not to force tetanus into rigor. And lastly, they are again
allowed to rest for an hour while the escaping gases (C) are a third
time collected. On analysis it appears that

A contains the least amount of CO2.
B „ the greatest „ „
C „ somewhat less than B.

B and C may each contain more than three times as much carbon
dioxide as A ; and C may contain more than B, if, from any cause,
rigor happen during the third hour \

Thus in tetanus, as in rigor, the gaseous changes consist in an in-
crease of the carbonic anhydride capable of withdrawal by an air-pump.
The increase is due to a special production of the dioxide within the
muscle, and not to the decomposition of some pre-existent stable
form of it, by means of the acid which appears during tetanus. This
is demonstrated by comparing the gases of normal and tetanized
muscles from the same animal — an experiment which is practicable
from the circumstance that muscles when tetanized in the cold
lose a very small quantity of gases 2. Frogs are taken and buried
in snow until almost rigid. Their vessels are then washed out with
ice-cold salt-solution ; and one leg from each is amputated and
scalded in the manner already described : if the scalding has been
perfectly done the reaction of the muscle to litmus paper is neutral,
not acid. The scalded limbs are minced in a vessel kept cold over
a freezing mixture, and put into the boiling-flask with (unboiled)
salt-solution at 0°. Phosphoric acid is placed in the froth-chamber v
ready for use. The minced muscle is evacuated at 50° ; then acidi-
fied, and again evacuated. Meanwhile the rest of the cold carcases
are arranged in series on a cold plate and tetanized at intervals
during many hours. At the end of this time these muscles also
are scalded : they should have an acid reaction. They are minced

1 Hermann, Op. cit. pp. 116, 117. Expt. 11 and 12.

2 Hermann, Op. cit. p. 25.

CHAP. IX.]

THE CONTRACTILE TISSUES.

357

and exposed to the vacuum both before and after treatment with
phosphoric acid.

These experiments may be illustrated by the following notes :

Experiment. Three frogs prepared as above described. Tetanus was
induced at intervals during 3| hours1.

MUSCLES IN EEPOSE. WEIGHT 13-4 grms. ( = 12-67 c.cm.).

Constitu-
ents in
c.cm.
(0°C.and
1 mtr.)

Per-cen-
tage of
muscle
used.

C03 (free)
C03 (fixed)

0-381
0-620

3-01
4-90

MUSCLES TETANIZED. WEIGHT 20-2 grms. (=19-09 c.cm.).

CO, (free)'
CO2 (fixed)

1-462

0-843

7-66
4-42

Whence it appears that in tetanus the carbonic anhydride which
a vacuum can extract, added to that which is liberated by acids, may
rise as high as 12'08 p. c. by volume of the muscle used. Further,
thafc the carbonic anhydride set free by an acid is constant in resting
and tetanized muscle.

Experi-
ments of
Pfliiger and
Stintzing.

From the experiments of Hermann, which have
just been detailed, we may conclude that muscle
contains some constituent which in the course of
contraction or of rigor suffers a decomposition and
yields carbon dioxide in a condition to be removed by the air-pump.
Further, that after scalding (p. 352), or after acidification by phos-
phoric acid (p. 353), this constituent is no longer capable of decompo-
sition by the means which commonly bring rigor about. But although
it is then incapable of decomposition by a vacuum at a temperature
of 50° C., it appears to yield to tbe prolonged action of boiling water2,
splitting up with the liberation of carbon dioxide. In the experi-
ments in question the muscles of rabbits were used. They were
deprived of blood, finely minced, and then plunged into a large
volume of briskly boiling water, which was kept boiling for two or
three hours. The carbon dioxide which escaped was absorbed,
with every precaution to avoid losses, by means of caustic solutions,
and afterwards determined both by weighing the potash bulbs and
also by the gasometric analysis of the carbonate formed.

1 Hermann, Op. cit. p. 118. Expt. 14.

" E. Stintzing, " Untersuchungen u. die Mechanik der physiologischen Kohlen-
saurebildung." Pfluger'a Arch.f. d. ges. Physiol. Vol. xvm. 1878, p. 388.

358 GASES LIBERATED IN RIGOR AND CONTRACTION. [BOOK I.

Under these circumstances mammalian muscle yields on prolonged
boiling about 100 vols. p. c. (at 0° and 760 mm.) of carbon dioxide.
The source of this large volume of gas is not the decomposition of a
preexistent compound of it; since, if muscle is well acidified and
afterwards washed for many hours at an ice-cold temperature before
being boiled, the yield of carbon dioxide on boiling is but little less
than when acidification is omitted. There is, in short, little doubt
that the constituent of mammalian muscle which liberates carbon
dioxide on prolonged boiling, is the same as that which is decomposed
in tetanus and rigor; for if muscles are tetanized or made rigid, while
at the same time opportunity is offered for the escape of the carbon
dioxide which is known to be generated in those processes, the yield
of dioxide on subsequent boiling is reduced to a mean of 20 or 30
vols. p. c. instead of 100.

Relation- ^ a comParison be made of the carbon dioxide

ship between produced during rigor and during tetanus, a very
the gases of curious relationship will be found between them,
rigid and con- Such a comparison should be made with the limbs of
tracting ^Q same fr0g . one jimb being passed into rigor by a

temporary exposure to 45° C. while still in its skin ;
the other limb being tetanized frequently during a long interval.
After this preparation each limb should be scalded and otherwise
made ready for the extraction of its gases. It will be observed that
during the induction of rigor by a temperature of 45° there is an
opportunity for the escape of gases, which is however diminished as far
as possible by preserving the skin, and making the operation as short
as may be.

After this experiment it will appear that the rigid muscle
contains more carbon dioxide than the tetanized. Similar experi-
ments further shew that tetanized muscles produce less carbon dioxide
on passing into rigor than muscles which have not been tetanized
previously. Now, the total carbon dioxide set free by the rigor of
muscle which has been tetanized is made up of

a. the amount in the muscle at the moment of bleeding it ;

b. the amount produced during tetanus, minus v, the small
amount lost to the air in tetanizing ;

c. the amount produced in rigor,

while the total carbon dioxide set free by rigor in an untetanized
muscle is made up of

a. the amount in the muscle at the moment of bleeding it ;

d. the amount produced in rigor.

But experiment shews that the difference between

(a + b — v + c) and (a + d)

is about 2 per cent. Hence, if we assume that v = 2 per cent,
(and such is not an improbable assumption), then

(a + b + c) = (a + d), and b + c = d1.

1 Hermann, Op. tit. p. 26.

CHAP. IX.] THE CONTRACTILE TISSUES. 359

Changes in the non-gaseous constituents of Muscle in the states of
Activity and Rigor.

1. Change in Reaction and its causes.

During The flesh of dead animals, however fresh in the

rigor the ordinary sense of the word, has an acid reaction.

iTaTbeeT1011 Berzejiusl> who discovered this fact, concluded from his
neutral or experiments that it was due to the presence in muscle
alkaline be- of that acid which his countryman Scheele had separa-
ted from sour milk. The lactic acid of muscle was shewn,
by the subsequent researches of Engelhardt, Heintz and Strecker,
to differ from the common lactic acid produced by fermentation.
Liebig, who at first denied the presence of lactic acid in muscle, after-
wards based many ingenious hypotheses upon its supposed presence
in the muscular tissue during life. All these chemists, because
they had discovered lactic acid in the flesh of recently killed animals,
concluded that it must have been present during life ; for, at that
time, the conception had not yet been formed that when a tissue dies
processes set in which may give rise to new bodies — products of the
decomposition. This conception was due to Du Bois-Reymond. In
his papers on the reaction of the muscular tissue, and the changes
which it undergoes at death2, he established the immense importance
of distinguishing between a tissue which is yet living, though it
may be separated from the living body of which it once formed a
part, and one which has ceased to manifest the phenomena which it
possessed during life. With the cessation of these phenomena — and
in warm-blooded animals that cessation follows so soon upon somatic
death as to be almost coincident with it — there is a change in physical
properties and chemical structure. Thus whilst muscle is alive and in
a physiological condition it possesses a neutral reaction ; so soon as
it dies the reaction becomes acid. This change takes place so rapidly
in warm-blooded animals as to render it almost impossible to ascertain
the normal reaction; in cold-blooded animals, in which the vitality
of the tissues continues long, the acidification goes on so slowly as to
permit of its careful study.

It is impossible to over-estimate the importance of these, the first
researches which pointed to the subtle differences which may exist, even
from a chemical point of view, between living and dead tissues ; the
conception which guided them and which was securely based upon
them, immediately led one of Du Bois-Reymond's pupils, Kuhne, to
the discovery of the most important points in the chemistry of living
muscle ; and it has since then so influenced the progress of Physiology
that we can scarcely realize how much we owe to it. Our knowledge
of the changes which occur in secreting glands in various conditions
of functional activity ; of the variations in the objective characters of

1 Berzelius, Lehrbuch der Chimie, tibersetzt von "Wohler, 4th ed. Vol. ix. p. 569.
Ann. d. Chem. u. Pharm. Vol. i. p. 1.

2 Du Bois-Eeymond, " Ueber angeblich saure Keaction des Muskelfleiscb.es." Gesam-
melte Abhandlungen zur allgeineinen Muskcl- u. Nervenphysik. Leipzig, 1877. Vol. n. p. 3.

300 ACIDIFICATION OF MUSCLE DURING RIGOR. [BOOK I.

the retina, &c., has been gained by researches which prove the value
of the conception of Du Bois-Reymond.

Methods of Strips of red and blue litmus paper are pinned

determining alternately in rows to a varnished board, so that the
the reaction edges of adjoining pieces are in contact. A section of
the muscle of which the reaction is to be determined is
then pressed firmly over the boundary of two slips. In normal muscle
it is then observed that both the red paper assumes a bluish tinge
and the blue paper a reddish tint. This so-called amphichromatic
or amphoteric reaction, depends upon the muscle having in reality
often a neutral reaction ; when this is the case, though it affects both
blue and red litmus paper, it does not alter the tint of violet litmus.

When a muscle passes into the state of rigor mortis the reaction
becomes ipso facto decidedly acid, except in cases where the rigor is
brought on by plunging the muscle into hot water, when the
reaction is found to remain neutral or alkaline.

It would appear that the amount of acid which can be produced
in a muscle when it passes into rigor is a definite quantity, doubtless
depending upon the quantity of the body which, by decomposing, sets
acid free.

Acidifica- As was first shewn by Du Bois-Reymond, when a

tion of mus- separated muscle is tetanized and its reaction is deter-

cies, removed mined from time to time, it is observed to become more

from the in- , . -, i

fluence of the and m?re aci(? •

Wood, when Heidenhain shewed that the amount of lactic acid

they are formed during contraction increased with the resistance

tetanized. which the muscle had to overcome2.

It was shewn by Ranke that in this case as in tetanus there
is a maximum amount of acid which can be generated in the muscle
which is cut off from the blood-stream, and then tetanized. If two
muscles were taken for the determination of the amount of acid
formed during rigor mortis, but if one were subjected to prolonged
tetanus until rigor set in, whilst the other was allowed to remain
at rest, the quantity of acid formed in the first case would exceed
that formed in the second ; there is, therefore, a consumption of acid-
yielding substance during tetanus3.

Causeof the The acidity of muscle in the state of rigor or which

acid reaction has been tetanized is chiefly, if not entirely, due to the
wh^h^in liberation of lactic acid. In the very earliest stage of
the state of rigor it is probable that the acid reaction is really
rigor. due to an acid potassium phosphate, produced from

the alkaline phosphate by the action of lactic acid. Soon however
the reaction is acid because of the presence of lactic acid.

1 Du Bois-Beymond, Op. cit. p. 26.

2 Heidenhain, Mechanische Leistung, p. 143 et seq.

3 In a thesis presented to the University of Bonn on 4th June, 1880, and entitled
"Beitrage zur physiologischen Chemie des Muskels," Dr Joseph Warren communicated
preliminary observations tending to shew that the amount of lactic acid which can be
obtained from muscles which have been tetanized is smaller than is yielded by similar
muscles which have been maintained in a state of rest.

CHAP. IX.] THE CONTRACTILE TISSUES. 361

v/';'

The isomeric Lactic Acids.

At least three acids are known to chemists which have the
composition expressed by the formula C3H6O3. These acids all agree
in being syrupy, colourless, liquids of strongly acid reaction, soluble
in water, alcohol and ether, and yielding when heated first lactic
anhydride (C6H1005) and afterwards lactide (C8H402). Though pos-
sessed of many common characters, a careful examination of their
behaviour to polarized light, of the crystalline form and the amount
of water of crystallization of their salts, and of their products of
decomposition, has clearly established the existence of three perfectly
distinct lactic acids.

These three acids are (1) sarcolactic or paralactic acid, the chief
acid of dead muscle : (2) ordinary lactic acid : (3) ethylene-lactic acid.
The two first of these acids appear to possess the same chemical
constitution, and they may be spoken of as ethylidene lactic acids ;
still they exhibit certain well-marked differences, the first being,
for instance, dextrogyrous whilst the second is optically inactive, and
the salts of the two acids differing in the amount of their water of
crystallization, and in their solubility.

a. Sarcolactic Acid. (Optically active ethylidene lactic acid.)

1. Liebig's extract of meat is dissolved in four parts of

of^actf^acids Warm water and 8 Parts of 90 Per cent< alcoho1 are theu
from muscle. added to the liquid which is continually stirred. The
mixture is allowed to stand until the insoluble matter
has subsided and a clear supernatant liquid is obtained, and the latter is
then separated by decantation. In order to separate any lactic acid from
the insoluble residue, the latter is mixed with twice its weight of warm
water, and then precipitated with four or five times its volume of
alcohol. The alcoholic fluids obtained by these two operations are then
evaporated on the water-bath to the consistence of a thin syrup, and the
' latter is again precipitated by the addition of three or four times its volume of
alcohol ; the insoluble matter may be kept for the preparation of creatine,
hypoxanthine, &c. The alcoholic solution is now evaporated to dryness,
the residue is mixed with water, some dilute sulphuric acid added, and
then shaken up several times with ether. On evaporating this liquid, a
residue is obtained which consists of a mixture of sarcolactic and ethylene-
lactic acids.

2. Instead of employing the above method, the syrupy liquid from
which creatine has crystallized, in Liebig's method of preparing creatine, is
acidulated with sulphuric acid and then shaken with ether, and the ether
evaporated.

Separation Having obtained a mixture of the two acids, their

of sarcolactic separation is effected by converting them into zinc salts, and

from ethylene- the separation of the two salts is based upon their different

solubility in alcohol. With this object, the mixture of

362 SARCOLACTIC ACID — LACTIC ACID OF FERMENTATION. [BOOK I.

raw acids is dissolved in water and boiled with suspended zinc carbonate or
zinc oxide ; the clear liquid is separated by filtration from the insoluble
zinc compounds, and then evaporated until crystals commence to form.
The liquid is now treated with four or five times its volume of 90 per cent,
alcohol; after some time the liquid becomes turbid and deposits needle-
shaped crystals, consisting of zinc sarcolactate ; the ethylene-laetate, being
far more soluble in alcohol, remains in solution. The crystals of the former
body are then collected on a filter, washed with cold absolute alcohol; and
they may with advantage be re-crystallized. From the zinc compound
sarcolactie acid may be obtained by dissolving the salt in water, decompos-
ing by means of sulphuretted hydrogen, filtering the solution, concentrating,
shaking with ether, and then evaporating the ethereal solution, when the
pure acid is left.

Properties Sarcolactie acid is distinguished from the two other

of sarcolactie varieties of lactic acid by its property of deviating the plane
acid and its of polarization to the right. The specific rotatory power is
compounds. greatest immediately after the acid is dissolved; it then
sinks rapidly, and afterwards slowly rises again, without however again
reaching its initial value. It is worthy of note that whilst free sarcolactie
acid is dextrogyrous, its salts are laevogyrous.

Zinc sarcolactate has the composition Zn (C3H5O3)2 + 2H20. When
heated for half an hour at 100° C. it loses the whole of its water of crystal-
lization (12-9 per cent.). It is soluble in 17'5 parts of water at 14 — 15° C.
It is almost insoluble in absolute alcohol, requiring 1000 parts of boiling
absolute alcohol for solution.

x

The specific rotation of this salt is =- 7°'6.

Calcium sarcolactate, 2 [Ca (C3H503)2] + 9H2O, crystallizes in the form
of tuits of microscopic needles. The specific rotation of the salt is = - 3° '8.

b. Ordinary Lactic Acid. (Optically inactive ethylidene lactic acid.)

This acid is perhaps not present in acid muscle, though it has been stated
to be so by Heintz. Its quantity is at least inferior to that of the other
isomeric lactic acids.

Preparation. This acid is formed when saccharine liquids ferment in
the presence of certain decomposing matters of animal origin (Milk, Cheese),
which serve as vehicles for a peculiar organized ferment ; to the fermenta-
tion thus induced the term lactic acid fermentation is applied. For this
reason this variety of lactic acid is often designated 'lactic acid of fermenta-
tion.' For the details of the methods for preparing this variety of lactic
acid the reader is referred to any systematic work on organic chemistry.

The acid resembles sarcolactie acid except in its not
of inactive possessing the power of rotating the plane of polarization,

ethylidene Its salts differ in crystalline form and in the amount of water

lactic acid of crystallization which they contain, from those of sarcolactie

and its com- add>

Zinc lactate has the composition Zn (C3H503)2 + 3H2O.
When heated in the water-bath for half an hour it loses all its water

CHAP. IX.] THE CONTRACTILE TISSUES. 363

of crystallization (18-178 per cent.). It is insoluble in alcohol ; but soluble
in 6 parts of boiling water and 58 — 63 parts of water at 14° — 15° 0.

Derivatives Both sarcolactic acid and the acid product of fermenta-

of the ethyli- tion yield exactly the same derivatives or products of

dene lactic decomposition when subjected to identical processes, so

that we are justified in considering them to have the

same chemical constitution.

When heated for some hours in the water-bath, the lactic acids yield
the body termed lactic anhydride, C6H10O5; when heated at a higher
temperature, 140° or 150°, lactide, C3H4O2, is formed.

When oxidized with dilute chromic acid the ethylidene lactic acids
yield acetic and formic acids but no malonic acid.

Many synthetic processes are known which yield the inac-
Synthesis tive ethylidene lactic acid of fermentation ; the most instruc-
tive, as bearing upon the constitution of the acid, consists in

tic acid. heating ethyloxycyanide of ethylidene 4 with

\C2H5 JO/

aqueous solutions of the alkalies, when ammonia and common lactic acid
and a small quantity of ethylene-lactic acid are formed.

c. Ethylene-lactic Acid.

This acid undoubtedly accompanies optically active ethylidene lactic
acid in the juice of flesh.

Prepara- This has been described in part under 'sarcolactic acid.'

tion. The alcoholic fluid from which sarcolactate of zinc has

separated contains ethylene-lactate of zinc, which may be obtained from it
by evaporation. From the zinc compound, the free acid may be liberated,
by following precisely the same process as was recommended for the separa-
tion of sarcolactic acid.

Is optically inactive.

Its zinc salt has the same composition as that of sarco-
lactic acid, losing when heated in the water-bath 12 '9 per cent, of its
weight. The zinc salt, unlike that of the isomeric lactic acid, is exceedingly
soluble in water ; it is also much more soluble in alcohol.

Products of When oxidized by means of chromic acid, ethylene-

oxidation. lactic acid yields malonic acid (C3B[4O4).

By heating ethylene chlorhydrate with potassium cyanide
the nitrile of ethylene-lactic acid is formed j thus

CH2C1 OH,. ON

+ TTf — +

CH2OH CH2.OH

Ethylene Potas- Cyanhydrin. Potas-

Chlorhydratc. sium siuin

cyanide. chloride.

364 CHANGES IN MUSCLE DURING ACTIVITY. [BOOK I.

When cyanhydrin is boiled with caustic potash, the potassium salt of
ethylene-lactic acid is obtained ; thus :

CH2.CN CET.COOK

+NH

CH2.OH CH2.OH

Cyanhydrin. Potassium Potassium ethylene- Ammonia,

hydrate. lactate.

2. Changes in the proportion of Water.

According to Ranke1 the amount of solid matter in muscles
undergoes diminution when muscles are tetanized, so that there
appears to be a relative increase of water. Further, according to this
author, the proportion of water in muscle is inversely proportional to
its power of doiag work.

3. Changes in the water and alcohol extractives.

It was first shewn by Helmholtz2 that when muscles are tetanized
they yield a smaller quantity of matters soluble in water (water-
extractives}, but a larger quantity of alcohol-extractives than before.
This has been fully confirmed by Ranke3.

Heidenhain and his pupils Nigetiet and Hefner4 have more
recently shewn that as the resistance which an active muscle has to
overcome increases, the amount of the alcoholic extract increases and
that of the aqueous extract decreases.

4. Changes in the Proteids.

The total nitrogen of resting and tetanized muscle was found by
Ranke to be the same (about 14*4 p. c.). Ranke5 fancied that he had
made out that the preparation of proteids removable by water
diminishes in the tetanized muscle. We may fairly say that no
trustworthy experimental results exist to prove that the proteids
of muscle undergo changes during activity.

5. Changes in the amounts of Creatin.

According to Sarokin6 the amount of creatin in muscle is the same
whether it be in a state of rest or activity ; according to this author
a large production of creatinine occurs, however, during tetanus.

1 Eanke, Tetanus, Chap. n. p. 63 ("Der Wassergehalt des Muskels").

2 Helmholtz, "Ueber den Stoffverbrauch bei der Muskelaction." Arch. f. Anat. u.
Phys. 1845, p. 72.

'3 Eanke, Tetanus, p. 141.

4 Heidenhain, Nigetiet und Hefner. " Versuche iiber die Abhangigkeit des Stoffum-
satzes in den thatigen Muskeln von ihrer Spannung." Pfliiger's Archiv, Vol. in. (1870)
p. 574.

5 Eanke, Tetanus, p. 119. See also Nawrocki, Centralblatt, 1866, p. 385.

6 Sarokin, Virchow's Archiv, Vol. xxvui.

CHAP. IX.] THE CONTRACTILE TISSUES. 365

This statement has been contradicted by Nawrocki1. Voit has found
the creatme to be diminished by activity2.

6. Changes in the proportion of Glycogen and Sugar.

As was first pointed out by Ranke, during tetanus sugar is
produced in muscle. At the same time there is a diminution in the
amount of glycogen (Nasse3). It is quite unknown whether the sugar
is produced at the expense of the glycogen.

7. Changes in the amount of Fat and volatile fatty acids during

activity.

Ranke4 thought he had established that the quantity of fat
increases in muscle during activity, but the conclusion is probably
not warranted by the experimental data5. The same remark applies
to the statement of Sczelkow6 that during tetanus there is a
diminution of the volatile fatty acids contained in muscle5.

8. Oxidizing and reducing properties of Muscle during rest and

tetanus.

Griitzner shewed7 that whilst resting muscle is able to oxidize
pyrogallic acid, muscle which has been tetanized fails to do so. He
further shewed that solutions of sulphate of indigo undergo a change
of colour when circulating through tetanized muscle, which is of
such a kind as to point to the production of reducing substances7.
Gscheidlen8 has further shewn that during activity nitrates are con-
verted into nitrites.

B. Changes in the chemical composition of the medium surrounding

muscle.

a. When muscle is exposed to the air.

,_ . The study of 'muscular respiration,' or the study

1 Respira- rj, , . J, . , £ 11 ••

tion' of 0* the physiological processes of muscle by examining

muscle : the air to which muscle is exposed, seems to have

methods of first been systematically practised by George Liebig9
in 1850. His method was the simple one of en-
closing the muscles in a tube inverted over a surface

1 Nawrocki, Centralblatt, 1865, p. 417.

2 Voit, Zeitschr. f. Biol. iv. 1868 (p. 77).

3 Nasse, Pfliiger's Archiv, Vol. n. (1869) p. 97.

4 Eanke, Tetanus, p. 190.

6 See Hermann's criticism, Untersuchungen uber den Stojfwechsel, &c., p. 86 and 87.

6 Sczelkow, " Die fliichtigen Fettsauren des Muskels und ihre Veranderung wahrend
des Muskeltetanus." Archiv f. Anat. u. Phys. 1864, p. 672.

7 Griitzner, "Ueber einige chemische Eeactionen der thatigen und unthatigen
Muskeln." Pfliiger's Archiv, Vol. vii. (1873) p. 255.

8 Gscheidlen, "Ueber das Keductionsvermogen des thatigen Muskels." Pfliiger's
Archiv, Vol. vin. (1873) p. 506.

9 Georg Liebig, "Ueber die Eespiration der Muskeln." Archiv fur Anat. Phys.
u. wiss. Med. (J. Miiller), 1850, p. 393. The method was first suggested by Du Bois-
Beymond.

366 'RESPIRATION' OF EXCISED MUSCLES. [BOOK i.

of mercury. On the top of the mercury in the tube, floated a
caustic solution to absorb the carbonic anhydride. A rise of the
surface of mercury betokened absorption of oxygen ; for Liebig paid
no regard to the nitrogen, and assumed that the oxygen and the
carbonic anhydride were interchanged volume for volume. He
stated that excised frog-muscles, whether bloodless or unbled, on
exposure to an atmosphere of common air or of oxygen, absorb oxygen
and excrete carbonic anhydride. He made out also that the excretion
of carbon dioxide may occur into an atmosphere containing no oxygen1.

Valentin After Liebig, in 1855, Valentin2 took up the same

question of the influence of excised muscle on its
surrounding medium, with a view to discover differences of composition
between irritable and non-irritable muscle. The muscular hind limbs
of frogs were exposed to air in closed tubes for 1 — 6 days, and the air
examined at intervals. Irritability was abolished in various ways, as by
spontaneous death, by subjection to high temperatures, or by beating
to death ; and comparisons were established between the gaseous
exchanges of normal muscle, of non-irritable muscle, and of various
tissues, such as the skin and bones of the frog's body. He discovered that
other organs besides muscle abstract oxygen and excrete carbonic
anhydride; and, which was more important, that the gaseous exchanges
of muscle continue uninterruptedly after the death of the muscle.
In a word, not only living muscle, but skin and even dead muscle have
a 'respiration.' The gaseous exchanges of dead muscle are indeed
different from those of the yet living ; and the kind of exchange which
is characteristic of the dead state is established as soon as ever death
of the muscle intervenes, whether it be suddenly induced or whether
it be reached by a protracted decline. Active muscle does not
appreciably affect the nitrogen of the surrounding air; but after
irritability is lost, nitrogen escapes from the muscle. This difference
in the gaseous exchanges of the living and dead state was considered
by Valentin to betray some difference of constitution so subtle as
to escape chemical analysis or electrical tests.

Matteucci ^ne year ^ater' ^n 185^> Matteucci published a paper

in which the same subject of muscular respiration is
treated of3; but especially the respiration of muscles during contraction.
He divested frogs of their skin, took the hinder extremities and freed
them roughly from blood with filter-paper, and arranged them in a
closed air-space over mercury, for electrical stimulation. The air-space
measured about 70 — 80 c.cm. Stimulation was carried on at intervals
so as not to fatigue the muscles.

1 Op. dt. p. 408.

2 G. Valentin, "Ueber die Wechselwirkung der Muskeln und der sie umgebenden
Atmosphere." Arch. f. physiol. Heilkunde, 14th year, 1855, p. 431. His methods of
analysis are explained in Valentin's treatise on Physiology.

3 Ch. Matteucoi, "Recherches sur les phenomenes physiques et chimiques de la
contraction inusculaire." Ann. de cltimie et de physique, 3 s6rie, Vol. XLYII. 1856,
p. 129.

CHAP. IX.] THE CONTRACTILE TISSUES. 307

The air was analysed by absorbing the carbon dioxide with caustic
solutions and the oxygen with phosphorus. He found that muscle,
whether at rest or in contraction, caused a diminution of oxygen and
an increase of carbon dioxide, and usually of nitrogen also, in the sur-
rounding air — changes which were greater during contraction than
during repose of the muscle. The oxygen absorbed was greater than
the carbon dioxide exhaled. He exposed muscles to a vacuum, then
to pure hydrogen for two or three hours, then to an exhausted receiver,
which was subsequently filled with pure hydrogen. Notwithstanding
this careful removal of oxygen from about the muscles, carbon dioxide
was yielded up by them, especially on stimulation. Hence Matteucci
concluded that the oxygen which, in muscular respiration, helps to
form the carbon dioxide is not the oxygen of the air, but oxygen
which exists in muscle in a state of chemical combination1.

Valentin ^n 1857 Valentin published researches on the effect

of contracting frog-muscles upon the atmosphere2. His
apparatus consisted of a glass cylinder, abed, 2 decimetres high and be-
tween 2 and 3 centimetres bore. The bottom was closed by an iron
plate,/, with a hook, e, externally for the attachment of a battery wire.
The top was provided with a short iron flange, gh, which supported an
iron plate or lid, ik, capable of being hermetically fixed to the cylinder
by means of an interposed washer and screws. This lid was provided
with an exit-pipe, x, guarded by a stopcock, as well as with a ther-
mometer, I, and a gauge, nrt, open to the air, all being securely fixed
in an air-tight fashion.

In order to determine exactly the volume of air in the cylinder after
everything had been arranged for an experiment, the following preliminary
calculations were made.

Let w = cubic contents of the glass cylinder and the proximal limb
of the gauge when all is screwed up and the gauge stands at zero (or.
Fig. 58).

Let h be the height of mercury which must be poured into the distal
limb of the gauge to raise the mercury in the proximal limb up to o : i. e.
to diminish the volume v by the volume between q and o, or p.

Then, if b equals the barometric pressure, it follows that

v : (v - fj) = (b + h) : b,

b + h
fA" h '
b

Now suppose that a volume, p, of mercury be poured into the glass cylinder
to the level vw, before the lid is screwed up ; and let the gauge again stand

1 Matteucci, Op. cit. p, 138.

2 G. Valentin, "Die Wirkung der zusammengezogenen Muskeln auf die sie um-
gebenden Luftmassen." Arch. f. physiol. Heilkunde. New Series, Vol. i. 1857, p.
283.

368

'RESPIRATION' OF EXCISED MUSCLES.

[BOOK i.

at zero. The cubic contents of the cylinder and proximal limb of the gauge
now =(v— p}. And let hf be the height of mercury which must be

FIG. 58. APPARATUS FOR DEMONSTRATING THE RESPIRATORY EXCHANGES or LIVING
MUSCLE. (VALENTIN.)

introduced into the distal limb of the gauge to reduce the new volu
(v— p) by the same volume /n; i.e. to raise the proximal mercury again
from q to o.

CHAP. IX.] THE CONTRACTILE TISSUES. 369

Then, by (i),

and (v-p) = nfi + l .................. ...... (iii);

whence /w, = - -. ........................ (iv).

This gives the value of //,, which is the distance between two fixed points
q and o of the proximal limb of the manometer.

Having determined /u, carefully, once for all, it is clearly easy to
determine the volume of air under observation, diminished as it is by a
muscle preparation or any apparatus of unknown volume which may, in
an experiment, be introduced into the cylinder. It is only necessary to
ascertain the height of mercury which must be poured into the distal
limb to raise the proximal surface of mercury from q to o, and apply the
formula

V (the unknown volume)

By means of this apparatus Valentin was able to confirm Mat-
teucci's statements ; but he pointed out that comparable results could
only be obtained by employing small preparations and by restricting
the experiment to the first half-hour. He found that the relative
amount of absorbed oxygen, which Matteucci had discovered to be
greater than the exhaled carbonic anhydride, became less during con-
traction: that is to say, during contraction more carbonic anhydride
is exhaled than oxygen is absorbed. He noticed also that muscles
which have been fatigued by prolonged tetanus yield more carbon
dioxide and absorb more oxygen absolutely (though less relatively
to the carbon dioxide) than sound and vigorous muscles ; and this
he thought to be due to an enfeebled power of - resisting the
disintegrating action of the atmosphere, owing to some altered consti-
tution of the tissue which is characteristic of the state of exhaustion.

To Valentin, therefore, belongs the credit of pointing out that tbe
so-called 'respiration' of muscular tissue is in part a phenomenon of
putrefaction ; but it was Hermann who clearly enunciated this fact and
perfectly discriminated the living and the dead factors in the process.

Hermann metn°d adopted by Hermann was extremely

simple1: The muscle-preparation was suspended by a
platinum wire in a wide absorption tube, which was then inverted over
a mercury bath in such a manner that tbe open mouth dipped below
the surface, and the mercury stood at the same level outside and inside
the tube. The tube thus enclosed an unmeasured volume of air at

1 Hermann, Stojfwcclisel der Muskeln, p. 32.

G. 24

370 'RESPIRATION' OF EXCISED MUSCLES. [BOOK i.

the atmospheric pressure, together with the muscle, and usually a little
moisture floating on the surface of the mercury ; while the muscle
was kept within reach by means of the attached wire. If stimulation
of the muscle were desired, it is easy to see how it would he ac-
complished. At the close of the experiment the muscle was dragged
down through the mercury, an operation which was found to be
attended by no loss of gases whatever. The gases left in the
tube were then passed into several dry absorption tubes in succession,
in order to get rid of the moisture on the surface of the mercury;
and finally they were ready for analysis. The carbon dioxide was
directly absorbed by caustics; the oxygen was exploded, and the
nitrogen read off. The oxygen of the original air was estimated from
the nitrogen left behind. All deviations from this in the final
analysis were considered to be due to the absorption of oxygen by
the muscles during the experiment. The method is open to three
trivial objections: The small amount of carbon dioxide normally
present in the air (4 parts in 10,000) is neglected; the atmosphere
is not absolutely constant in its composition as regards oxygen and
nitrogen ; muscle itself yields up nitrogen on exposure, but only in
excessively small amounts, and during the earlier parts of an ex-
periment (Hermann).

Resting In this apparatus, by comparing during 15 or 20

muscles hours the gaseous exchanges of bloodless muscles, some

absorb 0. of which were rigid while others were still living, it

appeared that the absorption of oxygen is practically equal in the case
of both living and rigid muscles ; and therefore is probably not con-
nected with the functional mechanisms of the tissue. It is, in short,

Absorption dependent upon a process of putrefaction which is accele-
of o is in part rated according to the extent of muscular surface exposed
I utref active. to the action of the air, and which begins at a very early
period of exposure.

And in part But while the absorption of oxygen, as determined

physiological, by an analysis of the surrounding gaseous medium, can-
not be ascribed to any but putrefactive causes, we must hasten to
explain that other and more delicate tests would lead us to infer an
extremely slight but constant employment of oxygen which is truly
physiological.

Influence of ^-s earlj as 1795 Humboldt1 observed that muscles

medium upon preserved their irritability longer in oxygen than in air
irritability of or gases containing no oxygen — an observation which
muscle. has since been confirmed by Georg Liebig2 and others,

and with especial accuracy by Prof. Hermann. In Hermann's
experiments a muscle was suspended in an absorption-tube by means

1 Al. Humboldt, Versuche uber die gereizte Muskel- und Nerven-faser, 1797, Vol. n.
p. 282.

2 Georg Liebig, Op. cit.

CHAP. IX.] THE CONTRACTILE TISSUES. .371

of platinum wires melted through the sides of the tube, the wires
being adapted to an electrical apparatus for stimulation. The
absorption-tube was filled with salt solution after the muscle had been
fixed within it; and then inverted over a vessel containing a lower
stratum of mercury and an upper one of salt solution, in such a manner
that the open mouth of the tube passed through the upper layer
to the mercury. A bent tube proceeding from a reservoir of electro-
lytic hydrogen, or pure nitrogen, or air, or detonating gas, served to-
introduce the gas into the absorption-tube, and* at the same time
to drive out the salt solution. Muscles of different degrees of
thickness were kept under observation ; but the sartorius of the frog
was found to be peculiarly favourable for these experiments. The
results were not modified by previously curarizing the muscles.

Tested in this manner, it appeared that the thickness of the
muscles had a singular influence on the result. The sartorius, of
large surface and small bulk, lived longer in hydrogen than in gases
containing oxygen; while thicker muscles agreed with the muscles
observed by Humboldt, in retaining their irritability longer in
oxygen. Another form of this experiment led to the same con-
clusions. If muscles were enclosed in tubes which were then made
as vacuous as possible, until nothing remained in them but traces of
carbon dioxide, the sartorius was found to live longer in the vacuum,
in the absence of oxygen, than in air ; while thicker muscles lost their
irritability sooner in the vacuum. It should be noticed that all
muscles exhibit an exalted irritability when first the vacuum is
produced.

This influence of oxygen upon the irritability of thick and thin
muscles seems to admit of but one explanation. There are two
concurrent processes in muscles exposed to the air, in which oxygen
plays a part : one tends to destroy, the other to preserve irritability.
The former is, beyond doubt, the putrefactive process already demon-
strated in living and rigid muscle, which spreads the more rapidly
the greater the surface exposed. Hence in the thin sartorius the
process invades all parts of the tissue within a short time, and death
results : to defend the muscle from oxygen is to preserve it alive. On
the other hand, the second process implicating oxygen is a true
physiological process of revival. In the thicker kinds of muscles, whose
internal mass is long shielded from the putrefactive action of oxygen,
this process of revival is a marked benefit ; and hence the muscle
removed from the influence of oxygen by enclosure in a vacuum
more rapidly becomes enfeebled than one exposed to the air.

Of the nature of this functional absorption of oxygen and process
of revival we have as yet no exact conception. The process itself
is but of small value in prolonging the normal irritability of frog-
muscles exposed to the air and of no appreciable moment in the
function of contraction ; but, as will be hereafter explained, it is-
extremely potent in restoring irritability to mammalian muscles
exhausted by interruption of their blood current (p. 380). It is

24—2

372 'RESPIRATION' OF CONTRACTING MUSCLES. [BOOK i.

probably not effected by haemoglobin but by the tissue juices ; for
many invertebrates have no hemoglobin which yet have muscles
not essentially different from those of the frog. It may be simply
that the presence of oxygen assists the escape of the deleterious
carbon dioxide better than hydrogen or nitrogen, as was found
to be the case with the gases of the blood by Ludwig1.

It may, therefore, be said respecting the oxygen of the surrounding
medium, that while an appreciable amount is abstracted and absorbed
in the inevitable putrefaction of exposed muscle, a small portion,
altogether too slight to affect a gas-analysis, is taken up to preserve
irritability.

Resting The atmosphere surrounding exposed muscles,

muscles ex- besides losing oxygen, suffers an increase of its carbon
hale C02. dioxide. As was pointed out by Valentin, this is

not wholly an exchange of a functional character ; but is common to
living and dead muscular tissue. In other words, it is one of the
early phenomena of putrefaction, together with the absorption of
oxygen. But a comparison of the exhalations of living muscle
and of muscle made rigid, discloses that less carbon dioxide escapes
from the normal, on mere exposure, than from the rigid. Since
putrefaction is more, and not less, rapid in normal muscle than in
muscle made rigid by heat (the method of inducing rigor usually
adopted in these experiments), it is clear that putrefactive changes
cannot be called in to explain this difference. There can be little
doubt that it is due to the increased amount of carbon dioxide which
rigor is known to generate in muscle (vide supra). The amount
of carbon dioxide given off is very irregular and has no relation
to the oxygen at the same time absorbed.

-Hitherto we have considered the case of muscles
m rePose ; and we have come to the conclusion that,
sorb more o apart from an inappreciable quantity of oxygen, ab-
and exhale sorbed or otherwise employed, the exchanges of
more co2 than ^he 'respiration' of exposed muscle are not func-
muscies tional, but putrefactive, and are shared alike by muscle,

skin and other tissues. When we turn to the case
of muscles in tetanus we find that the gaseous exchanges have
a greater value, and especially as regards the carbon dioxide
excreted. The increase in the amount of oxygen absorbed is
indeed slight, and is due to the agitation of the tissue during
tetanus; for if the air about an unstimulated muscle be mechanically
kept in motion, a similar increase of the oxygen absorbed is found to
occur. It is not due to any increased activity of the putrefactive
processes brought about in the passage of the electrical currents, since
such currents have no influence over the putrefactive absorption of
oxygen by rigid muscles. The more remarkable increase of the carbon

1 Hermann, Op. cit. p. 52.

CHAP. IX.]

THE CONTRACTILE TISSUES.

373

dioxide exhaled is due, in the first place, to the increased production
of it within the muscle during, tetanus1; and in the second to the
increased facility for its escape offered in the agitation of the muscle.

The following experimental figures will serve to illustrate the extent of
the gaseous exchanges of muscle2.

Experiment. Comparison of gaseous exchanges of living muscle, and
muscle made rigid by exposure to a temperature of 45° C.

Duration of experiment 19h. 15 m. Temp. 14 — 17°C. Gases estimated
at 0° and 1 mtr.

1. Living muscle = 7-352 grms. = 6*948 c.cm.

2. Rigid muscle = 7 '6 31 grms. = 7 -2 13 c.cm.

Oxygen absorbed.

Carbon dioxide
exhaled.

c.cm.

p.c.

c.cm.

p.c.

Living muscle (6 -948 c.cm.)
Rigid muscle (7*213 c.cm.)

1*277
1*127

18-37
15*62

0*605
1*184

8*70
16-41

Experiment. Comparison of the gaseous exchanges of muscle in repose
and in tetanus.

Tetanus induced at intervals during the experiment.

Duration of the experiment 3 h. 10m. Temp. 15 — 16° C. Gases esti-
mated at 0° and 1 mtr.

1. Resting muscle = 9*468 grms. = 8*949 c.cm.

2. Tetanized muscle = 9*480 grms. = 8 -960 c.cm.

Oxygen absorbed.

Carbon dioxide
exhaled.

c.cm.

p.c.

c.cm.

p.c.

Resting muscle (8*949 c.cm.)
Tetanized muscle (8*960 c.cm.)

0-548
0*746

6*12
8*33

0*128
0*836

1*43
9-33

B. Changes in the chemical composition of the medium surrounding

muscle.

ft. When muscle is still in the body.

In the previous section, we have discussed the so-called
'respiration' of muscles removed from the body, or the gaseous
exchanges between excised muscles and their surrounding medium;

1 See the preceding section.

2 Hermann, Op. cit. pp. 123, 125 : Expts. 23 and 27.

374 GASEOUS EXCHANGES OF MUSCLES IN THE BODY. [BOOK I.

and we have tacitly assumed that the medium in question is
the air. This is not strictly true. The medium enclosing the
elementary parts of contractile substance consists of the tissue
juices, which come into contact with the air only at the surface
of the muscle : the tissue juices mediate between the muscular
substance and the air. Now the fluid which bears to muscle
within the body the same relation which air has to excised
muscle, is the blood ; and as in the former case, the tissue juices
are those which deal directly with the muscle-substance, mediating
between this and the blood. By the ramification of blood-capillaries
between the fibres of muscle, the opposed surfaces of the muscle and its
medium become enormously more extended than in the case of air ;
and by how much the more extensively the muscle is presented to the
medium, by so much the more readily will exchanges be effected.
This is a circumstance favourable to exchanges between muscle and
blood apart from any peculiar fitness or endowment of blood for the
work of exchange, which are matters for discussion in the Chapters on
Respiration.

In consequence of the organization of the body, and the necessity
which it is under of preserving a normal standard or equilibrium, there
are two methods of determining the influence which muscles exert upon
the blood. We may, in the first place, contrast the blood flowing to
and that flowing from the muscle, while the muscle is left in repose,
or is thrown into activity. This is the direct method, and is equivalent
to exposing muscles to an atmosphere of known constitution and after-
wards analysing the atmosphere. In the second place, we may observe
the changes of the general ingesta and excreta of the body which are
brought about when muscle is converted from one state to the other.
This is essentially an indirect method. Hitherto it has only been
employed to ascertain the chemical processes of muscle on passing from
the state of rest to that of activity, when the same animal is
compared, in respect of its ingesta and excreta, during repose and
during exertion. But a simultaneous comparison of the daily food and
excreta on the one hand, and the proportion of muscular and non-
muscular elements of the body on the other, in different kinds of
animals enjoying the same conditions of rest, might be employed to
ascertain the normal exchanges between muscle and its medium when
the former is at rest.

It is only necessary here to point out in general terms the uncer-
tainty of the indirect method of analysis. Everything which is given
up by muscle to blood is not of necessity given up by the blood to
the general excreta. The method of excretion is only one of the
means employed by the body to preserve its equilibrium. Some
part of the substances cast by muscle into the blood may be appro-
priated by other organs or tissues, and never appear at the surface of
the body ; and some part of the substances excreted, though brought
to the surface during muscular activity, may not have arisen within the
active muscle.

CHAP. IX.] THE CONTRACTILE TISSUES. 375

Changes of the medium surrounding muscle as shewn in an analysis
of the blood of muscle.

Analysis of ^ne general nature of the exchanges between muscle

the blood and blood has been long known or inferred from the

flowing to and physical character of the blood flowing out of the muscles,
from muscle. rf his blood is defined in general terms as venous ; and
ttwgases°b ^e distinction between arterial and venous blood is one
the method of °f ^ne most obvious and interesting problems of the
Ludwig and physiologist. But it is to Professor Ludwig that we
Sczeikow. owe the first accurate examination and account of the

exchanges. Assisted by Sczeikow1, and subsequently by A. Schmidt2,
he determined by means of the air-pump the composition of the
inflowing and outflowing blood of mammalian muscle. The blood
was collected from the muscles with as little disturbance to the general
circulation as .possible, by the following means. A cannula was inserted
into the femoral vein, below the opening of the vena profunda, with its
mouth towards the heart, and a loop of ligature was slipped beneath
the femoral vein above the opening of the profunda. When the ligature
was tightened, the normal current of blood from the profunda into the
femoral towards the heart was at once turned from its course and
flowed without obstruction through the cannula; when it was slackened
again, the current at once resumed its original channel, without the
tension having for a moment been raised. In this manner a supply of
venous blood from resting muscle was obtained. Arterial blood was at
the same time drawn through a cannula in the carotid artery. If it
were desired to stimulate the muscles of the leg, electrodes tipped
with moistened sponges were applied, one in the inguinal hollow and
the other behind the sacrum, opposite the origin of the sciatic
plexus. The extraction of the gases was at once undertaken in a
Lud wig's blood-pump. As a rule the bloods were examined in the
folio wing order: (1) the arterial blood; (2) the venous blood from
stimulated muscle ; (3) the venous blood from resting muscles. Very
frequently the examination of the last had to be postponed until the
following day; sometimes both the second and third kinds were
examined on the day after their withdrawal. In this case the blood
was kept, surrounded by ice, in the tube into which it had been
drawn.

The analysis of the gases was made by Bunsen's method.
On examination it was found that :

1. The colour of venous blood from active muscle is sometimes
brighter and sometimes darker than the colour of venous blood from

1 Sczeikow, "Zur Lehre von Gasumtausch in verschiedenen Organen :" presented
by Prof. Ludwig. Sitzungsber. d. k. Akad. Wien, Vol. XLV. Abth. i. 1862. Second
series of experiments.

2 A. Schmidt, "Das Verhalten der G-ase, welche mit dem Blut durch den reizbaren
Sliugethierinuskel stromen." Sitzungsber. der math.-phys. Classe der k. s. Gesellsch. der
Wissensch. Vol. xx. p. 12. See also Arbeiten aits der physiol. Anstalt zu Leipzig, 3rd
year, 1868 (Leipzig, 1869), p. 1.

376 INFLUENCE OF CONTRACTING MUSCLE ON THE BLOOD. [BOOK I,

muscle in repose. It may be brighter even when the blood contains
less oxygen.

2. Blood streams more rapidly out of contracting, than out of
resting, muscle.

3. Taking arterial blood as the standard, the following table
represents the condition of venous blood from resting and from
active muscle.

Venous blood : —

0, less than
arterial blood.

C02, more
than arterial
blood.

of resting muscle

9 p. c.

671 p. c.

of active muscle

12-26 p. c.

1079 p. c.

Since blood streams more rapidly from active muscle than from
muscle at rest, these differences of the blood in the two cases are
really much more considerable than the table shews; since in a given
interval of time more blood, with its reduced oxygen and increased
carbon dioxide, flows from active than from passive muscle.

4. If Q represent the numerical relation between the increase of
carbon dioxide and the decrease of oxygen as blood is converted
from the arterial into the venous state; that is to say,

ifQ

difference between C02 of arterial and venous blood
difference between O of arterial and venous blood

then this quotient Q is found in most instances to be greater during
contraction of muscle than during repose. This might be due to the
fact that, in contraction, more carbon dioxide is generated for every
volume of oxygen absorbed, than in repose ; but since the precise seat
of the production of carbon dioxide is as yet but a matter of hypo-
thesis, we cannot at once draw this conclusion from the above experi-
ments. It may be merely that the forces which determine the
diffusion of carbon dioxide and oxygen respectively, are differently
affected by the condition of contraction; whence the change in the
relationship Q would be brought about, not by an increased gene-
ration of carbon dioxide, but by an increased elimination.

The method of experiment which has just been de-
scribed is not free from objection. The uncontrollable
changes of the blood current in the course of an experi-
ment introduce a variable element which deprives the
results of all exact quantitative value. To meet this objection, and
to obtain results which should be strictly comparable, Ludwig and
Alex. Schmidt1 devised 'a method of investigating the changes which

Ludwig and A. Schmidt, loc. cit.

CHAP. IX.]

THE CONTRACTILE TISSUES.

377

defibrinated blood undergoes as it is artificially forced in a constant
stream, through separated though still living muscles. For this purpose
the biceps and semitendinosus muscles of the dog's hind-limb may be
employed. These muscles are supplied with blood by a branch of the
hypogastric artery, and by three or four branches indirectly from the

FIG. 59. APPAKATUS OF LUDWIG AND A. SCHMIDT.

TT is a glass vessel containing the muscles B, resting upon a support G ; the upper
edge is ground, and smeared with grease, to permit the hermetical closure of the vessel
by means of a glass plate.

Q is a glass vessel containing mercury, resting on blocks r, r: by raising Q the
blood in F may be driven through the vessels of the muscles in T.

F is a, vessel containing blood.

A connects the vessel F with the blood-vessels of the muscles under experiment.

V conducts the venous blood from the muscle.

R, R' are two graduated pipette-like vessels connected by means of a flexible tube
and containing mercury. Venous blood flows into R, displacing the mercury.

E, E, electrodes.

M, M, mercurial manometer.

femoral artery. Cannulae should be tied into the hypogastric vessel
and into the main branch from the femoral vessel, and all the arterial
twigs going to neighbouring parts should afterwards be carefully liga-
tured. A corresponding number of cannulae should be introduced into
the chief veins. The muscles may then be separated from the surround-
ing tissues, a portion of the tuber ischii being sawn off with their origin.
The two arterial cannulae are connected with the two limbs of a
T-tube of glass, the third limb leading to the reservoir which contains
the blood. A similar arrangement connects the two veins with a vessel

378 INFLUENCE OF CONTRACTING MUSCLE ON THE BLOOD. [BOOK I.

into which the venous blood may flow, and where it may be collected
for analysis.

It is convenient and advisable to take the blood which is needed in
the experiment, from the dog whose muscles are examined. The dog
should be .first bled nearly to death, and its blood then defibrinated
and made ready for the experiment.

Various conditions of blood must be used for comparison ;
arterialized blood, or blood perfectly reduced, or asphyxiated blood,
or asphyxiated blood restored by oxygen1.

After its blood has been prepared, the dog should be killed and its
muscles separated in the manner already described. They are then to
be transferred to a glass vessel, T, T, such as is figured in Fig. 59, and
covered over by a glass plate. It will be observed that the vessel is
perforated so as to allow the tubes conveying arterial and venous blood,
A, V, to pass into and out of it, and to permit the passage of wires, E, E,
connecting the muscles with an induction coil by which tetanus may be
induced. If it is thought necessary, the muscles may be attached to
a lever so arranged as to record its movements upon a revolving
cylinder. The blood may be forced into the arteries by means
of a column of mercury, M, the pressure of which admits of careful
regulation.

It will be found necessary to increase the pressure of mercury in
the course of an experiment in order to maintain a constant flow of
blood. The pressure of mercury (40 — 60 mm.) which, at the beginning
of an experiment, serves to drive 2-5 to 3 c.cm. of blood per minute
through a biceps muscle of 150 — 200 grms. weight, will have to be
more than doubled (100 — 150 mm.) after four hours in order to do the
same amount of work. It must not be supposed that the resistance
to flbw suffers a regular increase during this time ; on the contrary, the
gradual increase is interrupted by frequent variations to and fro,
for which there is no assignable cause. The observer must pay
unremitting attention to the rate of outflow, if he wishes to maintain
it constant even for a few minutes2.

The cause of these variations in the rate of outflow is left obscure by
Ludwig and Schmidt ; but it is extremely probable that part of the obstruc-
tion is due to the gradual death and contraction of the smaller arteries.
When the driving pressure is raised, the constricted vessels will again be
opened for the passage of blood, and the original rate of flow will be restored.
If this cause of obstruction is admitted, it follows that the more rapid out-
flow of blood from a muscle which is brought about by raising the driving
pressure, may be due not so much to accelerating the current of blood, as to
enlarging the number of channels for it. In other words, raising the pressure
of injection does not bring a larger volume of blood to play upon the same
amount of muscular tissue, but rather brings more muscle under the influence
of the blood 3.

1 Ludwig and Schmidt used small pieces of iron wire to effect the reduction.

2 Ludwig and A. Schmidt, Op. cit. p. 27.

3 Pfliiger, " Ueber die physiologische Verbrennung in den lebendigen Organisinen.'
rfliiger s Arch. Vol. x. 1875, p. 350.

CHAP. IX.] THE CONTRACTILE TISSUES. 379

It will not have escaped the attention of the reader that these
experiments are complicated by the exposure of the muscle to what
is practically an enclosed space of air. In other words, two methods of
experiment are being employed side by side — the method of exposure
to air as a medium and the method of exposure to blood as a
medium. And, as a matter of fact, Ludwig and Schmidt determined
that the air in the glass vessel, after an experiment of some hours'
duration, had lost some of its oxygen and gained in carbon dioxide.
The value of this exchange is, however, relatively slight. Another
defect in the method of experiment is also deserving of mention.
The blood as it flows into and out of the muscle is necessarily exposed
to the air of the 'glass chamber through the membranous walls of
the arteries and veins into which the cannulae are inserted. This
possible source of error was determined by Ludwig and Schmidt1 to
have no effect upon the analyses as regarded the oxygen which the
blood might take up from the air. While to counteract the error
as regarded the carbon dioxide which the blood might yield up to
the air in the same manner, only those experiments were compared
in which the facilities for the escape of it were approximately the
same in the rate of flow, and amount of carbon dioxide contained in
the blood.

Although in these experiments the authors above referred to
succeeded in imitating to a great extent the changes which go on in
the blood in its circulation through muscles, they found that in
separated muscles the gaseous exchanges were not so great as in
muscles connected with the body, the latter appearing to act more
energetically upon the oxygen of the blood. ]n fact, the conditions
of temperature adopted by Ludwig and Schmidt but little favour the
diffusion of oxygen amongst the tissues and the dissociation of oxy-
haemoglobin 2.

From the experiments which have been made in the manner
described it may be concluded that :

1. When a muscle through which an artificial stream of blood has
been circulating, is deprived of blood, the capacity for doing work is
not immediately lost. In the first stages of bloodlessness the
irritability increases ; but soon it sinks, at first with rapidity, then
more slowly.

The circulation of blood freed from oxygen, or of the blood
obtained from asphyxiated animals, exerts the same action on the
irritability of muscles as the absence of blood.

2. As regards the oxygen absorbed :

a. The quantity of oxygen taken up by muscle increases
directly with the increase in the rate of flow, apart from contraction.
Hence the greater proportion of oxygen absorbed in contraction is, in

1 Op. tit. P. 41.

2 Pfliiger, Op. cit., Pfliiger's Arch. Vol. x. p. 354.

380 PRESERVATION OF MUSCULAR IRRITABILITY. [BOOK I.

part at least, accounted for by the greater rapidity of blood current
which then occurs.

This is only true tinder the conditions of Ludwig and Schmidt's
experiments, in which an increased flow of blood through the muscle was
probably due to the blood being driven over a wider capillary area. More
O was taken up under the circumstances because more muscular substance
was brought to act upon the blood. It does not imply that the assumption
of O is dependent upon the rapidity of the blood stream, which is expressly
denied by Pfliiger1 and Tinkler2.

1). The more oxygen is contained in the blood flowing through
muscle, the greater is the ease with which the muscle takes up oxygen
from the blood.

c. The amount of oxygen consumed by muscle in activity, or
by muscle exhausted by doing work, is usually perceptibly greater than
that consumed during rest. But the oxygen consumed bears no
definite relation to the work done.

3. As regards the carbon dioxide excreted :

a. In most cases, but not in all, the venous blood flowing
from contracting or exhausted muscle contains an increased amount
of carbon dioxide. The exact cause of the less usual condition, in
which the carbon dioxide of the blood is diminished, is not clear.

6. The relationship between the carbon dioxide excreted and

, , , .j 009 excreted . xl

the oxygen absorbed, or the quotient ^ 2 , — , , , in these

experiments underwent no constant variation as the muscle passed from
the resting to the active condition.

Dependence The value of oxygen in preserving the irritability of

of muscular excised mammalian muscles may be readily demonstra-
irritabiiity ted. The circulation of a stream of oxygenated blood
upon a supply through muscle prolongs its life 17 or 20 hours beyond
the time when it would have died if left bloodless.
Hence Ludwig and Schmidt3 concluded, in opposition to the doctrine
then current, that a peculiar respiration goes on within muscle which
proceeds independently of the so-called vital properties of the con-
tractile matter. Furthermore, irritability may not only be preserved
in muscle by means of oxygenated blood, it may also be restored after
it has become lost by exhaustion of the tissue. For the purposes of
such restoration of muscle extremely minute quantities of oxygen are
sufficient. In one experiment, when a muscle had completely lost its
irritability owing to the interruption of its blood current for 128
minutes, and when for 38 minutes more a stream of reduced blood
had been let flow through the muscle without beneficial effect, the
passage of 13'5 c.c. of arterialized blood through it, occupying the

1 Pfliiger, " Ueber die Diffusion des Sauerstoffs, den Ort und die Gesetze der Oxida-
tionsprocesse im thierischen Organismus. " Pfliiger's Arch. Vol. vi. p. 48.

2 Tinkler, " Ueber den Einfluss der Stromungsgeschwindigkeit und Menge des Blutes
auf die thierische Verbrennung." Pfliiger's Arch. Vol. x. p. 368.

3 Op. cit. p. 46.

CHAP. IX.] THE CONTRACTILE TISSUES. 381

three succeeding minutes, restored the muscle almost perfectly. It
would seem that so little as 1*8 mgr. of oxygen is sufficient to restore
the irritability of a muscle weighing 209 grms.1

Analysis of As compared with the changes wrought in the

the non-gase- gaseous constituents of the blood by the exercise of

ents arte11" muscle> the changes in the non-gaseous constituents
wood of due to the same circumstance are small and less

muscle. certain.

It has been stated that during muscular activity, the amount of
the aqueous extractive matters removable from muscle diminishes, whilst
the alcoholic extractives increase : that whilst glycqgen diminishes, sugar
increases and lactic acid makes its appearance ; further that tetanized muscle •
possesses considerable reducing powers, which we may surmise to be
associated with the production of new substances within the muscle.

Were our knowledge of the chemical composition of the blood complete,
we should expect to find variations in the composition of that fluid, after it^
has passed through a muscle, which should be the correlatives of the changes
which occur in the muscle itself. In so far as the gases are concerned this
has been shewn to be the case. In reference to non-gaseous constituents,
our information is, however, of the scantiest character ; it indeed is limited
to the two following statements.

1. During tetanus, blood circulating through muscle becomes charged
with reducing substances.

Alexander Schmidt passed two different quantities of blood free from
oxygen through muscle at rest, and through muscle which was tetanized,
and then agitated the two specimens of blood with oxygen. He found that
the blood which had traversed teteunized muscle took up more oxygen than
that which had traversed resting muscle, and from this he concluded that
tetanized muscle gives up reducing substances to blood.

2. During tetanus blood acquires sarcolactic acid (Spiro2). We yet
possess very slight information on this point.

Changes in the medium surrounding muscle as shewn in the
analyses of the general excreta of the body.

The excretions which are modified by muscular exercise are those
of the lungs and kidneys. The description .of the methods of
collecting and examining these excretions properly belongs to the
Chapters on Respiration and the Urine. It will, therefore, merely be
necessary here to speak of the methods of experiment having a
peculiar bearing on the question of muscular work.

Effects of As regards the excretion of the lungs, it has long

muscular ex- been known that the volume of respired air is increased
ercise on the .. , . , , * , . «

pulmonary during muscular exertion, and that the proportion 01

exchanges. carbon dioxide and oxygen involved in the process of

1 Ludwig and A. Schmidt, Op. cit. pp. 58 and 61.

2 Spiro, "Beitrage zur Physiologie der Milchsaure." Zeitachrift f. phys. Chemie,
Vol. i. (1877—78) p. 111.

382 INFLUENCE OF MUSCULAR CONTRACTION ON RESPIRATION. [BOOK I.

respiration is enlarged both during and immediately after the period
of exercise1.

But the variations of the carbon dioxide exhaled and the oxygen
absorbed do not occur pari passu ; the relationship of rest is
different from that of activity. The exact determination of this
relationship, although theoretically very simple, is a matter of
considerable practical difficulty ; and the gradual improvement of the
apparatus employed may be traced in the series of papers already
referred to. No attempt will be made here to explain the practical
methods, inasmuch as the classical apparatus of Regnault and Reiset'2,
Sczelkow3 and Pettenkofer and Voit4, will be described in the Chapter
on Respiration. The object of all the improved appliances is to
exactly estimate the oxygen absorbed, and the carbon dioxide excreted,
while the air entering the animal's lungs is of fairly normal constitution
and pressure.

Methods of To effect this there are three chief methods. The

Experiment. animal may be enclosed in an air-space disconnected
from the external air, the carbon dioxide being removed, and the
oxygen being replaced, as they are formed and consumed respectively
(Regnault et Reiset). Or the animal may be made to breathe out of
one vessel and into another, the loss and gain respectively being
accurately measured, while the pressure in each vessel is maintained,
by suitable apparatus, the equal of that of the atmosphere (Sczelkow).
Or the animal may be kept in a space through which air is continually

1 Among those who discovered and investigated the influence of muscular exercise
upon the exchanges of respiration may be mentioned the following : Jurine, quoted in
the Encyclopedie methodique, Art. "Medecine," Vol. i. p. 494: ed. by Vicq. D'Azyr,
1787. Seguin et Lavoisier, "Premier Memoire sur la Respiration des animaux."
Mem. Acad. 1789, p. 575. Other researches of Lavoisier will be referred to under
' Respiration.' W. Prout, "Observations on the quantity of carbonic acid gas emitted
from the lungs during respiration." Thompson's Annals of Philosophy, Vol. n. 1813,
p. 328. Translated into Schweigger's Journal fur Chemieu. Physik, 1815, Vol. xv. p. 47.
E. A. Scharling, " Dritte Reihe der Versuche um die Menge der Kohlensaure zu
bestimmen welche vom Menschen in einen gewissen Zeit ausgeathmet wird." Journal
fur prakt. Chemie, Vol. XLVIII. 1849, p. 440. Vierordt, Physiologic des Athmens.
Andral et Gavarret, "Recherches sur la quantity d'acid carbonique exhale par le
poumon dans 1'espece humaine." Ann. de Chimie et de Physique, Ser. in. Vol. vui.
1843, p. 129. Regnault et Reiset, "Recherches chimiques BUT la Respiration des
animaux des diverses classes." Ann. de Chimie et de Physique, Ser. in. Vol. xxvi.
1849, p. 299. Ed. Smith, "Experimental Inquiries into the phenomena of Respiration."
Trans. Roy. Soc. Lond. 1859, Vol. CXLIX. pt. ii. p. 681. Pettenkofer und Voit,
"C02-Ausscheidung u. 0-Aufnahme wahrend des Wachens u. Schlafens." Sitzungsber. der
fc. hayer. Akad. d. Wissensch. zu Miinchen, 1866, Vol. u. p. 236. Ibid. 1867, Vol. i. p. 255.
Speck, Schriften d. Gesellsch. z. Be/order, d. ges. Naturwissensch. zu Marburg, Vol. x. p. 3,
1871. Rohrig und Zuntz, "Zur Theorie der Warrneregulation und Balneotherapie "
Pfluger's Archiv f. d. ges. Physiol. Vol. iv. p. 57. Zuntz, "Ueber den Einfluss der
Curarevergiftung auf den thierischen Stoffwechsel." Pfluger's Arch. Vol. xn. p. 522.

2 Regnault et Reiset, " Recherches chimiques sur la Respiration des animaux des
diverses classes." Ann. de Chimie et de Physique, 3rd series, xxvi. p. 299, 1849.

3 Sczelkow, Op. cit. 1862.

4 Pettenkofer, "Ueber einen neuen Respirations- Apparat. " Abhandlunr/en der matli.-
phys. Classe d. k. layer. Akad. d. Wiss. Vol. ix. Miinchen, 1863, p. 229. Voit, "Beschrei-
Ijung eines Apparates zur Untersuchung der gasformigen Ausscheidungen des Thier-
korpers." Ibid. Vol. xn. Miinchen, 1876, Abth. i. p. 219.

CHAP. IX.] THE CONTRACTILE TISSUES. 383

being drawn, and the air analysed as it emerges either in whole
or in sample (E. A. Scharling1, Pettenkofer and Voit). ',

In a fourth plan (Rohrig and Zimtz)2, oxygen is respired instead of
air. A rabbit whose lungs have been cleared of nitrogen by the free
respiration of pure oxygen for some time, is made to breathe into and out of
the same gasometer of oxygen, the bell of which is carefully counterpoised.
The oxygen passes from the gasometer to the rabbit, and back again, through
water-valves which contain a caustic solution instead of water. In this
manner the carbon dioxide formed in respiration is completely absorbed.
As the oxygen is used up and the gasometer sinks, the counterpoise is
adjusted from time to time in order to maintain the pressure within the
apparatus equal to that of the atmosphere. A pump for artificial respira-
tion may be readily adapted to this apparatus.

Effect of In whichever way the experiment is made, the fact

exercise on is clearly elicited that muscular exertion increases both
the gases of the oxygen absorbed and the carbon dioxide excreted,
but in 110 ratio of equivalency. To be precise we may
take the experiments of Scz.elk.ow, inasmuch as they were specially
devised to demonstrate this fact.

Rabbits were the animals employed. They were fed on a diet of
wheat and milk; and the gaseous exchanges of the whole body were
determined during rest, and during tetanus of the hind limbs brought
on in the manner already described. A study of the numerical results
shews that

1. Much more carbon dioxide is excreted during tetanus.

2. Usually, but not always, more oxygen is absorbed; but never
so much as corresponds with the carbon dioxide at the same time

exhaled. In other words, the quotient „-*= ^— is increased

O absorbed

during tetanus.

It should be observed that all the other conditions of the animal
besides those of movement specially contrasted, should be taken into
account in these comparisons ; and particularly the condition of food.
According to the food the relation of carbon dioxide exhaled and
oxygen absorbed is found to vary. This most probably explains the

CO

different values assigned to the relationship —~ during a period of

repose by different observers3.

The above conclusions are illustrated in the following table of three

, COn excreted „,, , . .-,

experiments. Q indicates the quotient g — r~5 — • *-"Q numbers in the

last column ("experimental errors + N in c.c.") are found by subtracting

1 E. A. Scharling, "Versuche ii. die Qnantitat der von einem Menschen in 24
Stunden ausgeathmeten Kohlensaure." Ann. der Chemie u. PJuirm. Vol. XLV. 1843.
Heft ii. p. 214.

2 Op. cit. Note, p. 382.

3 See Begnault and Reiset, and Sczelkow, Op. cit.

384 INFLUENCE OF MUSCULAR CONTRACTION ON RESPIRATION. [BOOK I.

the N" of the inspired air (estimated by the method of difference) from the
N of the expired air ; a + sign indicates that the N of the expired is
greater than that of the inspired. It is evident that all experimental
errors, i.e. errors in reading off measurements, etc., will sum themselves
algebraically to the 1ST so determined.

The duration of the experiments was recorded on a revolving cylinder ;
and the gases were analysed by Bunsen's method.

The amounts of CO2 and O, reduced to the standard temperature and
pressure, are averaged per minute of the experiment.
II = repose : T = tetanus.

Duration
of experi-
ment in
minutes.

Respira-
tions.

C.c. in one minute.
C02. 0.

Q.

Experi-
mental
errors + N :
total in c.c.

i. R

7-6

92

4-97

12-29

•404

+ 13-54

T.

6-5

82

13-69

12-11

1-13

+ 31-74

ii. R.

9-2

80

7-85

12-76

•615

-18-19

T.

5-1

106 ;

17-62

19-02

•927

+ 11-73

v. R

9-2

140

6-99

17-47

•400

- 5-3

T.

5-1

130

, 19-61

30-35

•646

+ 16-4

Cause of A simple consideration of tbe amount of carbon

the increase dioxide excreted, serves to shew that the increase during
of co2 excre- tetanus is not due simply to favoured elimination.
A very large rabbit rarely weighs more than 2 kilogs.,
of which TV, or 105*23 grms., may be considered to be the weight of the
blood. Taking 30 vols. per cent, as the proportion of carbon dioxide
in it, this weight of blood includes about 31'6 c.c. of carbon dioxide —
the total carbon dioxide in the blood of a very large rabbit at a given
moment. Now, even if we make the large assumption that the
carbon dioxide of the blood is, by rapid elimination, reduced to
one half in the course of an experiment, we shall still be quite unable
to account for the extraordinary excretion of carbon dioxide in
tetanus ; for if this enormous reduction were supposed to be effected

during a short experiment like Exp. ii., of the above Table, it would

(01 .(* \
or — — -i- 5*1 ) of carbon dioxide

per minute — a quantity far less than the observed. Hence the excess
of carbon dioxide excreted by the lungs during tetanus must be due
to an increased production of it within the body.

What part of this production has its seat in the tetanized limbs,
and what part in the rest of the body, these experiments fail to
discover. It is more than probable that the material exchanges of the
body at large do not preserve their equilibrium during the manifold

CHAP. IX.] THE CONTRACTILE TISSUES. 385

disturbances of tetanus ; but in what direction they are influenced,
whether they are checked or accelerated, is not clear. During
tetanus the blood becomes deprived of oxygen and charged with
carbon dioxide. Both these circumstances are unfavourable to the
gaseous exchanges of tissues generally ; nor are they compensated
by an increased respiratory and circulatory activity, for on prolonged
tetanus general asphyxia may arise1.

These considerations do not, however, entirely make clear the
origin of the carbon dioxide; and the uncertainty must always be
kept in mind when conclusions obtained in a study of the general
exchanges of the body are applied to muscles alone.

The absorp- A small correction is necessary in respect to the oxygen
tion of o dis- absorbed. After tetanus, the blood generally contains
cussed. a &maller proportionate quantity of oxygen. This defect

is to be ascribed to tetanus just as much as the defect of oxygen from
the air inhaled ; and hence it must be added to the latter before
the oxygen absorbed during tetanus can be compared with the carbon

CO

dioxide excreted. The effect of this addition is to make the quotient -~ -*

somewhat smaller than before ; but in no remarkable degree. For,
assuming the blood to contain 13 per cent, of oxygen, and half of
it to be found wanting at the close of a short experiment, it would
merely raise the oxygen absorbed by about 1 — 2 c.<j. per minute of
the experiment, in the case of a rabbit of 2 kilogs.

Effects of While the changes of respired air which are

muscular brought about by muscular exertion are so pronounced

exercise on as to have been remarked by the earliest observers, and
secretion ^ moreover give a decisive indication of the changes
which muscular exercise works in the blood, the changes
of the urinary excretion under the same circumstances have been
a matter of continual uncertainty. An examination of the methods
of the earlier investigators discloses the causes of this uncertainty.
The urine — the great drain of effete nitrogenous subtances — although
it is beyond doubt affected by the activity of the eminently nitro-
genous substance of muscle, is affected to an unexpectedly small
degree. In a word, the urine, in so far as it represents the tissue-
waste of muscle, is dependent upon the nutritional rather than
upon the functional processes of muscular tissue. Moreover nitro-
genous muscle, although an important source of the nitrogenous
excreta of urine, is by no means the only source. The character
of the food, its composition, the proportion of its elements, the
time of its ingestion, are all conditions which exert a large influence
over the constitution of the urine. It is to these two circumstances,
viz. to the smallness of the change which muscular exercise effects in
the urine, and above all to the omission from the calculation of the

1 Sczelkow, Op. cit.
G. 2$

386 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

various simultaneous modifying agents, that the discrepancies in the
statements of different observers are due.

In order that the reader may have some idea of the
Statements extent of these discrepancies, the results of some of the
observers. earlier observers may be here stated briefly. In most

cases, for the reasons already given, they cannot be con-
sidered either to support or to confute the results of later experiments.

C. G. Lehmann1 found that the excretion of urea in man was raised
from 32 grams per diem to 36 — 37 grams during exercise.

J. Fr. Simon2 also found an increase during exertion.

Mossier3 failed to discover any marked increase immediately after
exercise.

H. Beigel4 made experiments on six men under conditions of spare
diet and rich diet, water being taken at pleasure. With the former
diet the excretion of urea was raised from 31-86 grms. to 33 '32 grms. per
diem, on a day of labour; with the rich diet, from 46 '10 grms. to 52-26
grms. per diem.

W, Hammond5 found that the excretion of urea, which during 24
hours of rest was 31 -51 grms. (487 grains), rose to 42-4 grms. (682-09
grains) during a working day, and to 56 grms. (864 '97 grains), during a
day of hard labour.

Genth8 observed that the area excreted during prolonged labour was
increased beyond the normal.

Beneke7 found an increase on exertion.

Franque8 found an increase during exertion.

J. C. Draper9 found that a powerful man kept perfectly quiet in bed
for a long period, excreted on an average 2 6 '47 grams of urea daily, a
quantity which did not differ much from the normal excretion in
health.

L. Lehmann10 found a very small, or no increase in the excretion of urea,
on excessive exertion.

C. Speck11 found on exertion only a slight daily increase of urea, viz.
8 grms., with a rich nitrogenous diet, and 4 grms. with a poor nitrogenous
diet.

I C. G. Lehmann, Wagner's Handworterluch, Vol. n. p. 21. Physiological
Chemistry (Cavendish Soc. Trans.), 1851, Vol. i. p. 163.

u J. Fr. Simon, Animal Chemistry (Sydenham Soc. Trans.), 1846, Vol. n. pp. 144
and 168.

8 Mossier, " Beitrage zur Kenntniss der Urinabsonderung." Diss. inaug. Giessen,
1853. Quoted by Voit, op. cit. infra.

4 H. Beigel, " Untersuch. ii. den Harn- u. Harnstoff-mengen. " Verhandl. der
k. Leopold. Akad. d. Naturforsch., Vol. xxv. pt. i., 1855, p. 477. Quoted by Voit, op.
cit. infra.

5 W. Hammond, "Eelation existing between urea and uric acid." American
Journal of Med. Sci., new series, Vol. xxix., 1855, p. 119.

6 Genth, Untersuch. ii. den Einfluss des Wassertrinkens auf dem Stoffwechsel.
Wisbaden, 1856. Quoted by Voit, op. cit. infra.

7 Beneke, Nord. See Bad., 1855, p. 83. (Quoted by Playfair, Food and Useful
Work, p. 46.)

8 0. von Franque: Extract in Schmidt's Jahrbuch, 1856, Vol. xcn. p. 9.
tj J. C. Draper, New York Journal, March, 1856. See Voit, infra.

10 L. Lehmann, Arch. f. wissensch. Heilkunde, Vol. iv. pt. iv., 1860. See Voit, infra.

II C. Speck, Arch.f. wissensch. Heilkunde, Vol. iv. See Voit, infra.

CHAP. IX.] THE CONTRACTILE TISSUES. 387

Expert- Voit1 seems to have been the first to examine very

mentsof Volt, carefully this question of the influence of exertion upon
nitrogenous excreta. He selected the dog, as being better fitted for
the conditions of rigorous experiment than men, who had been
previously observed. The dog underwent severe exertion under two
sets of conditions : (1) when fasting from all food except water, and (2)
when on a diet just enough to cover all loss of weight when no work
was done. In each set of conditions the average excretions of resting
days and working days were compared. Sometimes resting days and
working days alternated ; but usually two or three days in succession
were devoted to rest, and two or three to labour. In this way there
was less danger of urine being retained in the bladder over the period
of a working day and expelled during the following resting day. In
all the experiments the dog was allowed to drink as much water as
he desired. Work was done in turning a tread-wheel, the number of
turns being registered, and the work carefully calculated. The dog
was taught to drive the wheel with great rapidity, encouraged by his
master's voice; and it was found that 10 minutes at a time was
sufficient to thoroughly fatigue the animal. A working day consisted
of about six periods, of 10 minutes each, spent in the wheel, with
about an hour's rest between each period. The greatest care was
taken to obviate loss of excretions ; thus the dog was taught to
micturate at a given spot and at regular intervals. The weight of the
dog, the water ingested, the urine, the urea, and the faeces were all
carefully determined from day to day. The dog weighed about 33
kilograms.

A studv of Volt's data conclusively shews us that
Conclusions. . . ( , J , ., . . J . ,,

during tne days ot exertion the excretion ot urea is

increased ; but it discovers also that,

1. The increase is absolutely very small; viz. an increase of
about 1 — 5 grams in the fasting experiments when the excretion
of repose was 10 — 15 grams a day ; and an increase of 5 — 10 grams
in the experiments with food, when the excretion of repose was about
110 grams a day.

2. The increase has no constant relationship to the work done.

3. The increase is evidently more influenced by the diet (viz. by
the amount of water ingested; by the fact of food being taken; &c.)
than by the circumstance of work.

Volt's dis- Impressed by the smallness of the increase of nitro-

cussion of Ms genous excreta during severe labour, Voit endeavoured
experiments. ^0 discredit the inference that some part of it, however
small, is directly owing to the activity of muscles. He dwelt upon
the circumstance that the free mgestion of water is, of itself, enough
to raise the excretion of urea by the urine. He pointed out that the

1 C. Voit, UntersuchungeniL denEinfluss des Kochsalzest des Kaffce's undder Muskel-
bewcgungen auf den Stqffwechsel. Mimchen, 1860.

25-2

388 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

circulation is much stimulated by muscular exercise, and that respira-
tion and the consequent ingestion of oxygen are enlarged by the same
means; all of which conditions, apart from the fact of muscular
contraction, favour the flow of parenchymatous juices and the manu-
facture of urea. He concluded, very justly, that the nitrogenous waste
was wholly incompetent to account for the mechanical work done.
He believed that the oxygen imported so largely into the body during
exercise served to burn up fat in extraordinary quantity and repair
the losses of heat which occurred by evaporation and radiation ; and he
formed the opinion that the small increase of urea excreted was due to
some of the nitrogenous tissues falling a prey to the oxygen during this
increased activity of combustion. In short, the increased oxidation of
nitrogenous matter was an accident of the general extension of
processes of combustion within the body, which would probably have
been avoided by a larger ingestion of fats.

These experiments have been since repeated by Voit1 with a view to
determine not merely the urea, but the total excretion of nitrogen in the
urine; and similar experiments have been made by Pettenkofer and
Voit8 on men ; with confirmatory results in each case.

It must ever be borne in mind, in following the early discussion of the
influence of muscular exercise upon the excretion of nitrogen, that the
question which divided observers was not whether exercise produces any
increase of nitrogenous excretion, but whether it produces an increase
corresponding with the mechanical effect of exercise. In respect of the
former question, a much greater unanimity would probably be found
than is commonly supposed to exist among those who have entered
into the discussion.

Experiments Although Voit selected the lower animals as best

of Pick and fitted for exact experiment, yet it was by a very simple
wisiicenus. observation conducted upon men that the question in one
of its phases was finally laid at rest. In the summer of 1865 Professors
Fick and Wisiicenus3, while abstaining from all nitrogenous food,
performed a definite amount of muscular work ; and, having estimated
the destruction of albuminous matters in the body from the nitrogen
excreted during the same time, they discovered that the combustion
of so much albuminous material was quite inadequate to account for
the mechanical effect. The work done was an ascent of the Faulhorn
to a height of 1956 metres. Therefore the mere mechanical lifting of
their bodies through this height was, in the case of

Fick (wt. 66 kilog.) 129096 kilogm.

Wisiicenus (wt. 76 kilog.)... 148656 kilogm.

1 Voit, Zeitsch. filr Biologie, Vol. n. p. 307. 1866.

2 Pettenkofer u. Voit, Zeitsch. fur Biologie, Vol. n. p. 459. 1866.

3 Fick und Wisiicenus, Vierteljahresschrift d. naturf. Geselhch. in Zurich, x., 1865.
Land., Edin. and Dnb. Phil. Mag,, Ser. 4, Vol. xxxi. p. 485. Suppl. number, 186G.
This important paper is also translated in the Ann. des Sci. Nat., S<5r. v. Vol. x., 18G8,
p. 257.

CHAP. IX.]

THE CONTRACTILE TISSUES.

380

This by no means represents the total work done during the
ascent ; for there is omitted the muscular work of circulation and
respiration ; the muscular work of maintaining the upright posture ;
the accidental, adventitious movements of the arms, &c. ; none of
which assisted directly in raising the experimenters to the top of the
Faulhorn, but all of which would swell the products of muscular tissue-
waste. For 17 hours before the work began no albuminous food was
taken ; during this period, during the period of the ascent, and for
some hours afterwards, the solid food consisted of rice, fat, and sugar,
taken in the form of cakes, with beer, tea and wine as fluid food.

The following diary serves to shew the relation of the urine as it was
collected, to the various parts of the experiment :

Aug. 29.

12 A.M.

Ceased to take

albuminous food.

Aug. 30.

6.15 P.M.

5.10 A.M.

Evacuated bladder.
Slept.
Started; made the

1 Urine of night before
J labour (A.)

ascent.

f Urine of labour (B.)

1.20 P.M.

Reached summit:

j

|

perfect rest.

Urine after labour (C.)
*

7 P.M.

Partook freely of

j

Aug. 31.

5 A.M.

meat, etc.
Slept.
Arose.

I Urine of night after
labour (D).

The urine was analysed at the summit for urea &c. ; and sealed samples
of each period were again examined for the total N, in the laboratory
on descending. Neubauer's method was used to determine urea; and
Liebig's method for chlorine.

The total N was determined by heating the urine with soda-lime,
collecting the NH8 as chloride, and determining as usual with JPtCl4.

From the analyses it appeared that the total elimination of nitro-
gen by the urine during the different periods was as follows : —

FlCK.

A. 6-9153

B. 3-3130)

C. 2-4293]

D. 4-8167

WlSLICENUS.

A. 6-6841

B. 31336)

C. 2-4165J

D. 5-3462

That is to say, reducing the numbers to a comparable form, the
excretion of nitrogen per hour was as follows :

FlCK.

A. 063

B. 0-41

C. 0-40

D. 0-45

WlSLICENUS.

A. 0-61

B. 0-39

C. 0-40

D. 0-51

390 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

It will be observed that, in both cases, the nitrogen actually excreted
during and immediately after exercise appears to diminish from the
standard of A. This fact will be afterwards discussed. It is not so
much the relative as the absolute elimination of nitrogen which we are now
considering.

Conciu- Having determined the excretion of nitrogen for

sions. some hours before, during, and after a period of muscular

exertion, Fick and Wislicenus had a key to the amount of albuminous
matter which had in the same interval of time been decomposed with-
in the body ; but one which required certain assumptions for its use.

In the first place it was assumed that all the nitrogen escapes by
the urine. This is not strictly true, since some is removed in the
faeces and some in the sweat; both these sources of loss were
neglected.

In the second place it was assumed that the nitrogen excreted
during labour (B), together with that excreted during the six hours
succeeding labour (C), fully represented all the albuminous matter
decomposed during the period of labour. This is the most vulnerable
point of the argument, and deserves some consideration.

It is clearly improbable that the nitrogen eliminated in the urine
(B) emitted during the occurrence of labour exactly represents the
albuminous decomposition during the same period. The act of
decomposition may not — and probably does not — occur at one step.
Intermediate stages between proteid and the ultimate form in which
the nitrogen escapes, are first formed and may possibly remain at
the seat of manufacture, or in some other organ, until long after
the period of exercise. Such intermediate stages would there-
fore have no representative in the urine excreted during labour.
But this lagging of elimination behind the time of formation, as the
observers themselves pointed out, is true also of the nitrogenous pro-
ducts of the period before labour. If the excretions of the period B
lack some of the decomposition-products proper to that period, it is
also true that they possess some which are proper to A. In order
amply to cover the effects of this retardation of excretion, Fick and
Wislicenus added to the nitrogen excreted during the eight hours
of the ascent the whole of that which was excreted during the six
hours following.

In the third place an assumption was necessary in order to con-
vert the nitrogen excreted into terms of albuminous substance. The
albuminous substances differ in constitution among themselves ; and
it is impossible to say which kind is chiefly taxed to supply the
nitrogen excreted during exercise. But all albuminous bodies except
permanent cartilage, contain more than 15p.c. of nitrogen. If,
therefore, we assume that every 15 parts of nitrogen excreted repre-
sent 100 parts of albuminous substance decomposed, we shall obtain
a quantity of albuminous matter greater than could possibly have
been destroyed in the body within the given time

CHAP. IX.] THE CONTRACTILE TISSUES. 391

If now we turn to the first table of the total nitrogen excreted,
and add the quantities B and C together, we find that the nitrogen
which Fick and Wislicenus agreed to take as representing the
albuminous decomposition of the period of exercise, is as follows : —

Fick 57423 grms.

Wislicenus 5*5501 grms.

And assuming that this represents an albuminous substance con-
taining 15 p.c. of nitrogen, we get as the quantities of nitrogenous
matter decomposed during the time of exercise, by

Fick 38-282 grms.

Wislicenus 37'000 grms.

Owing to a slight mistake fully explained in the original memoir, this
number in the case of Fick had to be reduced to 37*17 grams.

Thus were obtained numbers representing all the albuminous
matter which could possibly have been consumed during the period
of the ascent. Knowing this, we can easily calculate the kilogram-
metres of work which the burning of so much matter would represent.
According to Frankland1 the burning of 1 grm. of beef-lean dried

CMs 2161 kilogram-metres : whence, if the material consumed in the
y of each experimenter had been wholly burnt up, the oxidation
would have corresponded

In Fick to (37'17 x 2161) or 80324-37 kilog.-met.
In Wislicenus (to 37 x 2161) or 79957 kilog.-met.

As a fact the oxidation is not carried to its ultimate conclusion in
the body, since urea is the product. Hence the mechanical equiva-
lent of each combustion is really considerably less than that stated.

If we refer to the calculation of the work done in the mere
ascent of the Faulhorn, without taking into account for a moment
the large circulatory and respiratory work, we shall at once see what
a large proportion of the mechanical effect of muscular exercise is
wholly uncovered by the combustion of albuminous material.

Fick and Wislicenus were not acquainted with the heat of combustion
of dried albumin as directly obtained; they therefore made an approxi-
mation which now appears to have been rather in excess of the truth,
Neglecting the N altogether, they assumed that the heat of combustion of
albumin would not be greater than the sum of the heats of combustion of
the C and H which it contained.

1 grm. of albumin contains '535 grm. C.

•07 grm. H.
But the value of the heats of combustion are,

of C 8080 grm.-degrees*,

ofH 34462 grm.-degrees.

And

of *535 grms. C ... 4320 grm.-degrees.
of '07 grms. H ... 24 10 grm.-degrees.

1 Frankland, "Origin of Muscular Power." Lond., Edin. and Dub, Phil. Hag., 4th
Sor., Vol. xxxii., 18G6, p. 187.

392 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

Therefore that of the gram of albumin was assumed to be

6730 grm. -degrees.

Hence the heat of combustion of the decomposed albumin amounted,
In Fick to (37-17 x 6730) or about 250000 grm.-degrees.
In Wislicenus to (37 x 6730) or about 249000 grm.-degrees.
This converted into mechanical equivalents of that day, gave
For Fick, 106250 kilog. -metres.
For Wislicenus, 10"3825 kilog.-metres.

What Fick and Wislicenus' experiments shew beyond all doubt
is, that during and after muscular contraction no quantity of effete
nitrogenous material passes out of the body which is at all adequate
to the mechanical work done in contraction. What they do not shew
is whether or not any nitrogenous waste occurs in muscle during
activity.

It will have been observed that the experiments of Fick and
Wislicenus do not afford a comparison of the same organism during
repose and during activity, while all the other conditions are rigorously
the same. It is true that the food during the period immediately
before and immediately after exercise was non -nitrogenous, and so far
identical with that of the time of exercise itself. Nevertheless the
experimenters were not under precisely similar conditions in the three
periods named, because these periods were not equally remote from
the last ingestion of nitrogenous food. When the supply of nitro-
genous food is suddenly stopped, it is well known that the excretion
of nitrogen sinks to the starvation-standard with diminishing velocity.
Unless we make ourselves acquainted with the rate of this descent,
we cannot estimate the effect upon the excretion of nitrogen of other
circumstances or conditions.

Expert- In order to furnish such a comparison, and at the

merits of same time to control the observations of Fick and

E. A. Parkes. Wislicenus, Parkes1 undertook some experiments upon
soldiers at the Victoria Hospital, Netley. Two sets of experiments
were carried on. In one, work and repose were contrasted, in respect
of their influence upon nitrogenous excreta, on an unrestricted diet
containing no nitrogen ; and in the other, on a normal diet including
nitrogen, which was maintained practically constant during the whole
time of the experiment. We will confine ourselves in the first
instance to the experiments with a non-nitrogenous diet.

I. The two soldiers were kept for four days at light em-
ployment, with a normal temperate diet including meat, bread,
ale, etc.

> II. During two days they were put on a non-nitrogenous diet of
arrow-root, suga/and fat, and kept in perfect repose.

1 E. A. Parkes, " On the Elimination of Nitrogen during Kest and Exercise." Proceed.
Roy. Soc. Lond., Vol. xv. p. 339; Vol. xvi. p. 44, 1867.

CHAP. IX,]

THE CONTRACTILE TISSUES.

393

III. For four days they returned to the normal occupation >a*d
diet of period I.

IV. During two days they were again put on a non-nitrogenous
diet, and made to perform a long march each day, with intervals
of rest.

V. They again returned to their normal occupation and diet.

The diet was not limited ; the men took what they needed. The
nitrogenous excreta of urine and faeces were carefully determined ;
and periods II. and IV. were then compared as to the nitrogen
excreted.

We may illustrate this comparison by the numbers relating to one
of the soldiers experimented upon.

TOTAL DRY FOOD FOR THE TWO DAYS OF THE RESTING AND
WORKING PERIOD, IN KILOG.

K

W.

1-0044

1-3066

WATER, IN KLLOG.

4-592

5-1595

TOTAL EXCRETION OF NITROGEN, IN GRMS.

Urine,

Intestines.

Hours.

K.

W.

R.

W.

1—24
24—36

36—48

9-33
4-005
3-017 .

10-048
4-5331
3-361 J

•3875

•5318

Totals

16-352

17-942

•3875

•5318

Conclu-
sions.

From these tables it appears that there is a slight
total increase of nitrogen eliminated during muscular
exertion. It must be remembered that more non-nitrogenous food
was at the same time taken into the system, and more fluids were
drunk. Both these circumstances, apart from muscular exercise,
imply the use of digestive and assimilative apparatus which like
muscles are mainly nitrogenous in constitution.

In the second set of experiments the fluid and solid diet
(including 19'61 grms. of nitrogen) was constant during the whole
time.

394 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

The alternation of light labour, with repose or long marches, was
conducted as in the first experiment, viz. :

I. Four days of ordinary occupation.

II. Two days of perfect repose.

III. Four days of ordinary occupation.

IV. Two days of hard marching.

V. Four days of ordinary occupation.

Again we may take the case of one of the soldiers observed, — viz. the
one whose data served us in the first experiment.

COMPARISON OF TOTAL EXCRETION OF NITROGEN DURING REST AND
HARD LABOUR, IN GRMS.

Urine.

Intestines.

Hours.

R.

W.

IL

W,

1—24
24—36
36—48

20-094
9-855
8-315

18-478
7-3571
13-457 J

1-486

2-138

Totals

38-264

39-292

1-486

2-138

Means

19-132

19-646

The above table shews us that there is a slight increase in the total
amount of nitrogen excreted during labour as compared with a time of
perfect repose. This agrees with the facts elicited during the first set
of experiments. But it further appears that during the first 36 hours
of the period of labour the excretion of nitrogen is actually less than
during the corresponding period of rest. This fact is not indeed
supported by the data from the same man in the former experiment
when the diet was non-nitrogenous; but in the case of his fellow
in the previous experiment, the excretion of nitrogen passed through
the same phases ; that is to say, in the case of the other soldier on a
non-nitrogenous diet just as in the case of this one on a diet including
nitrogen, the immediate effect of hard muscular exercise was to diminish
the excretion of nitrogen : it is not until the night of the second 24
hours that we observe such an out-pouring of nitrogen as to raise the total
excretion of the two days of labour above that of the two days of rest.
This curious circumstance of the elimination of nitrogen will at once
remind us of the case of Fick and Wislicenus, in both of whom, on
a non-nitrogenous diet, muscular exercise seemed at first to diminish
the excretion of nitrogen.

If the above table be compared with the following one shewing
the nitrogenous excretions in the four days preceding rest, in the four
days following rest and preceding work, and in the four days following
work, a very interesting contrast will arise.

CHAP. IX.]

THE CONTRACTILE TISSUES.

395

TOTAL EXCRETION OF NITROGEN BY THE URINE IN THE PERIODS
INDICATED, IN GRMS.

Normals
before
Rest.

After
Rest.

After
Work.

1st day
2nd „
3rd „
4th „

17-886
16-810
19-212
17-520

15-920
17-608
19-382
17-54

21-25
19-942
23-488
19-530

Means

17-857

17-612

21-054

Conclu-
sions.

It will be seen that there is (1) a slight augmentation
of the nitrogen excreted during perfect repose as com-
pared with periods of light occupation (i. e. 19 '132 grms. per diem, as
against 17'857 and 17'612 grms.) ; and (2) that, after the period of
hard labour there is an enlarged excretion of nitrogen in the urine
which may continue for several days.

The observations of Edward Smith1 upon prisoners at
ments of ~Ed- hard labour, which were published shortly after the experi-
ward Smith. ments of Voit, and before those of Eick and Parkes,

tend to the same conclusions.

In an alternating series of days of hard and light labour, with a fixed
diet, the average excretion of urea on the days of labour was not indeed
markedly greater than on the days of comparative rest. Nevertheless, the
excretion of urea underwent from day to day a succession of oscillations
which beyond a doubt had reference to the character of the daily labour.
In most cases the total excretion of nitrogen during a day of hard labour
was somewhat greater than that of the days of light labour just before and
just after it; but in some cases it was apparent that the elimination of
nitrogen had been held over until the day after labour, making the excretion
of that day unusually large, and destroying the value of the averages.

The general fact of an increased excretion of nitrogen
mentso? Flint during periods of hard labour is also supported by the
and Pavy. observations of Austin Flint, Jun.2, and of Pavy3, made

upon the celebrated pedestrian Weston. This man was
kept under observation on two separate occasions during the performance
of extraordinary feats of walking, lasting for five or six days; and not
only was he observed during the days devoted to walking, but for five
or six days before and after. The ingested food in these periods was
accurately weighed, and the nitrogen estimated from the tables of Pay en4

1 Ed. Smith, " On the Elimination of Urea and Urinary Water." Phil. Trans. Roy.
Soc. Lond., vol. cli. pt. iii. p. 747. 1862.

2 Austin Flint, Juu., New York Medical Journal, June, 1871. " The source of
muscular power, as deduced from observations upon the Human Subject under conditions
of Kest and of Muscular Exercise." Journal of Anat. and Physiol, Vol. xn. p. 91,
1878 (contains the same facts as the former article).

3 F. W. Pavy, "The effect of prolonged muscular exercise on the system," Lancet,
London, 1876, Vol. i. pp. 319, 353, 392, 429, 466 ; Vol. n. pp. 741, 815, 848, 887.

4 Payen, Substances Alimentaires. Paris.

39G

INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

or from original determinations. The urine, and in Flint's experiment the
i'aeces also, were collected, and their nitrogen determined ; in the case
of the urine, only the nitrogen of the urea and uric acid was taken.
In Flint's experiments the weight of Weston fell from an average of
119 or 120 Ibs. to 116 '5 Ibs. on the first day, and gradually to about
115*75 Ibs. on the fifth day: it quickly regained the normal during
the rest succeeding the march. In Pavy's experiment there are no
data for exactly comparing Weston's weight before, during and after
the walk; but during the six days of exercise the weight fell from 134i£
Ibs. on the first day to 130T5g- Ibs. on the sixth.

The following tables will sufficiently bear out the general conclusion
of these experiments. The numbers in the case of Flint's observations
are the revised ones published by him in the Journal of Anatomy and
Physiology, after Pavy's observations had appeared.

OBSERVATIONS OF PROF. FLINT.

Nitrogen

Relation of

Nitrogen
ingested,
in grains.

elimi-
nated as
urea and
uric acid,

Nitrogen
ingested
and Nitro-
gen elimi-

in grains.

nated.

BEFORE THE WALK.

First 24 hours

361-22

304-55

Second „

288-35

276-84

Third

272-27

305-08

Fourth „

335-01

283-87

Fifth „

440-43

299-31

Average

339-46

293-93

1 : -8658

DURING THE WALK.

First 24 hours (80 m.)

151-55

331-44

Second „ (48 m.)

265-92

328-05

Third „ (92 m.)

228-61

399-16

Fourth „ (57 m.)

144-70

324-59

Fifth „ (40 J m.)

38304

306-08

Average

234-76

338-01

1 : 1-439

AFTER THE WALK.

First 24 hours

385-68

277-00

Second „

499-10

334-44

Third

394-83

358-78

Fourth „

641-71

348-19

Fifth

283-35

379-79

Average

440-93

339-64

1 : -7702
1

CHAP. IX.]

THE CONTRACTILE TISSUES.

397

OBSERVATIONS OF DR. PAVY.

Nitrogen
ingested,

Nitrogen
elimi-
nated as
urea and

Relation of
Nitrogen
ingested
and Nitro-

in grains.

uric acid,

gen elimi-

m grams.

nated.

BEFORE THE WALK.

First 24 hours

378-29

292-70

Second ,,

451-14

301-75

Third „

472-29

231-54

Fourth „

539-95

363-71

Fifth „

441-15

342-59

Sixth „

582-80

387-17

Average

477-60

319-92

1 : -6698

DURING THE WALK.

First 24 hours (96 m.)

491-80

524-59

Second „ (77 m.)

826-43

582-42

Third „ (70J m.)

759-15

600-29

Fourth „ (76J m.)

547-57

503-21

Fifth „ (67 m.)

790-78

450-06

Sixth „ (63 m.)

614-61

468-70

Average

671-72

521-54

1 : -7764

AFTER THE WALK.

First 24 hours

384-40

Second „

(not de-

238-39

Third

termin-

381-22

Fourth „

ed).

278-46

Fifth

299-19

Sixth „

268-17

Average

306-64

Before these figures can be compared it is necessary to remark that
the tables do not include the nitrogen eliminated in the faeces, or that
eliminated in the urine otherwise than as urea and uric acid. Compared
with the nitrogen of the urea and uric acid, the nitrogen of the other
egesta named is indeed small and unimportant; still inasmuch as the
nitrogen eliminated as urea and uric acid does not bear a constant relation-
ship to that otherwise excreted, no comparison is perfect which does
not include the latter. Unfortunately the experiments of Flint and
Pavy do not, together, furnish us with the necessary data for a perfect
comparison.

398 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK 1.

If now we contrast the experiments of Flint and Pavy

IU1U' ^rn

sions.

we observe in the first place that both shew an increase

. of nitrogen eliminated during exercise : the proportion of
nitrogen ingested to nitrogen excreted is, during the days before the walk,

in Flint's case 1 : '8658

in Pavy's case 1 : '6698
while during the walk it is

in Flint's case 1 : 1439

in Pavy's case 1 : '7764

But in the second place, we are struck by the remarkable difference
in the degree of the increase. This discrepancy, which has led Prof. Flint
into a long discussion of the question of muscular power, it is not necessary
to examine fully here. Assuming for the moment that the calculations
of both observers are well-founded, it is sufficiently clear that some
circumstance must have existed in one or the other experiment to destroy
their precise analogy. For example, in Flint's case the food of Weston
fell off considerably during the period of exercise ; while in Pavy's it
increased. Moreover, in Flint's experiment Weston urged himself to
the very extreme of endurance. "The most notable event in the course
of the five days' walk was what appeared to be a total collapse of muscu-
lar and nervous power. * * * * At 10.30 P.M. on this (the fourth) day,
Mr Weston broke down completely. He could not see the track, and
was taken staggering to his room, having reached apparently the limit
of his endurance. * * * The calculations as well as the general condition
of the system, shew that the period had probably arrived when repair of
the muscular system had become absolutely necessary1." If we may
suppose that over-exertion brings about a condition of muscular tissue in
which disintegration proceeds with unusual ease2, the very marked increase
of urea and uric acid in Prof. Flint's case admits of a simple explanation ;
and especially if we may further suppose that over-exertion, in certain
extreme cases, leads to the absolute rigor of individual fibres, as it does in
the case of muscles out of the body. In this condition the rigid albuminous
fibres would rapidly degenerate and serve to increase the common nitro-
genous excretions of the body3.

In addition to demonstrating a slight increase of nitrogenous excreta
during exercise, Dr Pavy endeavours to shew that the nitrogenous waste
during the walking period is wholly incompetent to account for the
mechanical work done. His argument is similar to that of Fick and
Wislicenus.

Still more recently W. North4 in experiments upon him-

mentsTof " se^' ^as arr^ve(i a* confirmatory results. He determined the

W North. urea excreted and, by Payen's Tables, the nitrogen ingested

during eight days, beginning on Monday, Sept. 3. On

Monday, Tuesday and Wednesday, nitrogenous food was taken ad libitum:

1 Flint, Journ. of Anat. and PhysioL, Vol. xn. p. 134.

2 T. R. Noyes, "Experimental Researches on the excretion of urea." American
Journal of the Med. Sci., New Series, Vol. LIV., 1867, p. 354.

3 Hermann, Stnffwechsel der Muskeln, p. 100.

4 W. North, " An Account of two Experiments illustrating the effects of Starvation
with and without severe Labour, on the Elimination of Urea from the Body." Journal
of Physiology (ed. by M. Foster), Vol. I. p. 171.

CHAP. IX.] THE CONTKACTILE TISSUES. 399

on Thursday the nitrogenous ingesta were reduced from an average of
about 15-5 grams per diem to 4*228 grams; on Friday to 1'365 grams;
and on Saturday to '399 grams. On Saturday, Sept. 8, Mr North
underwent severe exertion on the tread- wheel, the previous days having been
days of comparative rest. On Sunday nitrogenous food was again taken
ad libitum. Mr North first makes the assumption that the nitrogen excreted
as urea is influenced by the nitrogen ingested on the previous day, rather
than by that ingested on the same day ; and then finds that the total nitrogen
excreted as urea from Tuesday to Sunday is indeed greater than the total
nitrogen ingested from Monday to Saturday (the work-day), but only by 1/17 6
grams, which is wholly insufficient to account for the loss of weight (3J
Ibs.) sustained during the experiment, or for the work done.

Later experiments have led to a like result, in a majority of cases.
But whilst as a rule the nitrogen appears to be practically unaffected by
exercise, occasionally the excretion of nitrogen is decidedly increased. This
phenomenon, in Mr North's as in other cases, may, we think, depend upon a
temporary condition resembling fever engendered by the exercise, when it is
not due to causes already referred to.

General Ef- ^ may> therefore, be regarded as established that

feet of Mus- muscular exercise somewhat enlarges the total excretion

cuiar Con- of nitrogen. There is no reason to doubt that this

traction upon enlarged excretion is due, in the last instance, to the

^tif11^?611 degradation of the nitrogenous tissues of muscle ; but
of the Urine. ,, ° , , .. ,, ° „ . , ,

the degradation is far too small to account, by me-
chanical equivalence, for the work done in contraction. Moreover it
appears that the actual elimination of waste nitrogenous matters
does not coincide with, or very closely follow, the period of muscular
contraction. Sometimes, perhaps most frequently, the immediate
effect of exercise is rather to dimmish the elimination of nitrogen,
and to postpone the enlargement of excretion for some hours, or even
days. This fact is well illustrated in the experiments of Parkes ; by
whom it was thought to be of such essential importance as to warrant
the hypothesis that muscle, in activity, gains rather than loses nitro-
gen. "When a voluntary muscle is brought into action by the

influence of the will, it appropriates nitrogen and grows A

state of rest ensues, during which time the effete products are
removed, the muscle loses nitrogen, and can again be called into
action by its stimulus1." And this also, according to Parkes, is the
explanation why the elimination of urea is greater during absolute
rest than during light and regular labour.

Such a hypothesis is not, however, necessary. The formation of
effete nitrogenous matters in muscle, and their elimination at the
kidneys, are separate operations conducted by different protoplasmic
structures. The conditions favourable to one are not necessarily
favourable to the other ; blood, for example, is received into muscles
in large quantity during contraction, and at the same moment is

1 Parkes, "On the Elimination of Nitrogen." Proc. Roy. Soc. Lond., Vol. xvi.,
1867-68, p. 58.

400 INFLUENCE OF MUSCULAR EXERCISE ON THE URINE. [BOOK I.

diverted from the kidneys. It is therefore as probable that the
kidneys act ill during excessive muscular exertion, as that digestion
is imperfectly performed in the same circumstances. Further,
the formation of urea, the end*product of nitrogenous waste, takes
place in all probability in several stages, of which the earlier only
have their seat in muscle itself. This much at least is certain, that
muscle contains little or no urea, either at rest or after contraction ;
whence it must probably be concluded that if the proteids of muscle
contribute to the urea excreted normally, their contribution takes the
immediate form, not of urea,- but of some antecedent of urea. It is
not necessary to suppose that this antecedent form is creatine or any
body like creatine : indeed, as will be urged elsewhere, the tendency
of the experimental evidence is to render it very improbable that any
of the urea excreted passes through a preliminary creatine-stage ; for
when creatine is introduced artificially into the blood it is invariably
excreted not as urea, but as creatine. The form in which muscle-
proteid leaves the muscles, after having become effete as contractile
matter, may still be proteid ; in which case the whole oxidation of
muscle-proteid to the urea-form would occur altogether outside the
muscular tissues. But, if urea is not at once formed in muscle — if
the nitrogenous waste of muscle escapes into the blood in a proteid
(or other) form — the elaboration of the waste material into the form
in which it is actually excreted must go on elsewhere ; and in what-
ever organ this elaboration has its seat, it is very probable that the action
of the organ is hampered during prolonged or excessive muscular
exercise.

The manufacture of urea in two stages also fully explains the
other fact elicited in Parkes' experiments, viz. that during absolute
rest the elimination of nitrogen is slightly increased, if the diet
remains the same. Of the nitrogen ingested as food, part is de-
composed more or less directly and appears in the urine at once as
urea, and part serves to repair the nitrogenous waste of muscles,
reaching the urine by a circuitous path through muscular tissue;
the latter portion appearing in the urine at a later date than the
former. If anything occurs to diminish the wasting of muscle, less
nitrogen is yielded up by muscle to the urine, but at the same time
less is called upon to repair waste, and more, therefore, passes
directly into the urine from the food. Thus the same quantity of
nitrogen should appear in the urine whether muscles be exerted or
not, so long as the food remains constant. But this is only true if the
nitrogenous waste products of muscle pass at once from muscle into the
urine ; which ex hypothesi is not the case. On the contrary, they are
intercepted by some other organ and delayed. This organ, therefore,
at any given moment, contains waste products derived from muscle,
in course of preparation for excretion as urea. If, now, muscles are
suddenly thrown out of employment, less nitrogen of the food is called
upon to repair waste of tissue and more passes directly into the urine;
but at the same tinie the waste nitrogenous matters which happen

CHAP. IX.] THE CONTRACTILE TISSUES. 401

to be in the intermediate preparatory organ as the consequence of
the preceding day's exertions, are perfected into urea and excreted.
Thus the urine receives not only the nitrogen corresponding to its
proper day ; but also some which should have formed part of muscle,
and have been excreted on the morrow.

The nitrogenous constituents of the urine are not
muscular ^ie onty constituents which suffer change during mus-

contraction cular exercise; but the non-nitrogenous elements have
on tne non- excited less attention and still remain an object for exact
nitrogenous research. Kliipfel found that the acidity of the urine
as estimated by titration with soda solution was some-
times increased and sometimes diminished during mus-
cular exercise, even when the food remained constant. The experi-
ments of Sawicki gave the same results, and disclosed also that the
acidity was influenced by the quality and quantity of food taken far
more markedly than by the circumstance of muscular exercise. On
the other hand, Pavy noticed that the acidity of the urine was
increased during severe exercise; and Janowski seems to have come
to a similar conclusion. Kliipfel has surmised that the diminution
of acidity which is sometimes observed after muscular exertion may
coincide with an abnormally large excretion of acid sweat; but no one
has yet established this1. In the experiments of both Flint2 and
Pavy3 the proportion of sulphuric acid and phosphoric acid excreted
during labour was greater than during rest; while that of sodium
chloride was less under the same circumstances. In Sawicki's experi-
ments, referred to above, the phosphoric acid suffered no constant
variation from rest to labour.

THE CHEMICAL CHANGES OF LIVING MUSCLE WHEN AT KEST.

Many of the chemical changes of normal resting muscle have been
already described or implied in the Section on Muscle in action.
The "respiration" of excised muscles and the preservation of irrita-
bility by means of oxygen, were among the earliest discoveries of
muscular chemistry, and have been stated at length in the account of
the researches of Humboldt, Georg Liebig, Valentin, Matteucci and
Hermann. The general nature of the material exchanges of muscles
which are still in the circulation, is indicated in the conversion of
arterial into venous blood by muscles at rest ; but a point of special
interest is this, that the blood flowing from muscles paralysed by

1 Kliipfel, "Ueber die Aciditat des Harnes bei Kuhe u. bei Arbeit." Hoppe-
Seyler's Med. chem. Untersuch., iv. p. 412, Berlin, 1871. Pavy, Lancet, London, 1876,
Vol. ii. p. 888. Sawicki, " Sauregehalt der Harnmenge in Arbeit und Kube." Pfliiger's
Archiv, 1872, Vol. v. p. 285. Janowski, "Sauremenge des Harnes in Verbaltniss zur
Muskelarbeit." Dis. inaug., Moscow, 1876. (The original is in Kussian, it is ex-
tracted into Hofmann and Schwalbe's Jahresberichte, Vol. v. pt. ii. p. 274.)

2 Flint, New York Medical Journal, June, 1871.

3 Pavy, Lancet, London, 1876, Vol. n. p. 881.

G. 26

402 CHEMISTRY OF RESTING MUSCLE. [BOOK I.

section of their nerves is less venous than that flowing from quite
normal muscles1. We are naturally led to compare this with the
fact which we already know, that the blood from contracting muscles
is far more venous than that from the same muscles in repose, and to
ask whether the cause is not the same in both cases. In short, do
the nervous centres exert a tonic influence, automatic or reflex, over
the muscles, keeping them constantly in a state of partial contraction
at the expense of certain chemical decompositions ? Such a tonus
has, for physiological reasons, been ascribed to voluntary muscles ; but
not, as yet, upon grounds which are absolutely beyond question
(see the various Text Books of Physiology). While, then, we may
bear in mind this tonic contraction of voluntary muscle, as a possible
or, it may be, a partial explanation, the fact itself must be taken
as established, that separation of a muscle from its nervous centres
is followed by a diminution of the normal chemical changes of repose.
Whether or not the nervous centres induce a constant contraction
of voluntary muscles, they certainly bring about a constant chemical
tonus (as it has been called) in the same tissues2.

The method of investigating this chemical tonus has hitherto been
that of comparing the general excreta of the body before and after the
separation of large tracts of voluntary muscles from the central nervous
system. The excretion of the lungs was collected in an apparatus
for the respiration of a definite amount of gases at an unvarying
pressure. This apparatus, which differs from that of Ludwig and
Sczelkow devised for a similar purpose, will be found described in
the original memoirs. By means of it the oxygen consumed and
the carbon dioxide excreted by an animal could be measured with
considerable accuracy, while the arrangements were such as not
sensibly to impede the normal respiratory movements of the animal.
The apparatus was fitted with appliances for artificial respiration.
The separation of the muscles from the central nervous system was
brought about in one of two ways: by curare-poisoning3, or by division
of the spinal cord between the cervical and dorsal regions4.

A rabbit was attached to the apparatus and its respiratory exchanges
determined, respiration being carried on artificially to such an extent as
to leave the rabbit apnoeic for 3 to 4 seconds on stopping the injection of
air. This degree of apuoea is known not to affect the material exchanges
of the body5. The rabbit was then curarized with 2 — 3 mgr. of curare
subcutaneously injected, and its respiratory exchanges again determined.

1 Claude Bernard, Lemons sur les proprie"tes des tissus vivants, p. 221. Paris, 1857.

2 Bb'hrig und Zuntz, "ZurTheorie der Warmeregulation und der Balneotherapie."
Pfluger's Arch.f. d. ges. PhysioL, Vol. xv., 1871, p. 57.

3 Rohrig und Zuntz, Op. cit. Zuntz, "Ueber den Einfluss der Curare vergiftung auf
den thierischen Stoffwechsel. " Pfl tiger's Archiv f. d. ges. PhysioL, Vol. xn., 1876,
p. 522. Pfliiger, "Ueber Warme und Oxydation der lebendigen Materie." Arch. f. d.
(jes. PhysioL, Vol. xvm., 1878, p. 247.

4 Pfliiger, Op. cit., p. 305.

B Finkler und Oertmann, "Ueber den Einfluss der Athemmechanik auf den Stoff-
wechsel." Pfluger's Arch. f. d. ges. PhysioL, Vol. xrv., 1877, p. 38.

CHAP. IX.]

THE CONTRACTILE TISSUES.

403

During this time the animal was kept wrapped up in wadding to prevent
such an excessive cooling as might of itself depress the tissue-changes.
Artificial respiration was carried on at the same rate as before. Under
these circumstances both the consumption of O and the excretion of C03
were diminished by more than one-half in the course of an hour or two.
Thus l in one case the consumption of O fell

from 1740 cm. to 750 cm. per hour;
the excretion of CO2 fell

from 1560 cm. to 591 cm. per hour.

The diminution observed by PMger and others2 working in his
laboratory, as the mean of many experiments, was somewhat less than
this, viz.

Normal rabbit
Curarized rabbit

O consumed per kilog.
per hour in c.cm. (at
08C., and 760 mm.)

C02 excreted per kilog.
per hour in c.cm. (at
0°C., and 760 mm.)

673-21

436-20

570-41
356-9

that is to say,

in the amount of O 35 '2 p.c.

in the amount of CO2 37 "4 p.c.

This diminution of the gases interchanged in respiration is not
due to a deficient circulation; for both the blood-pressure and the
heart were observed in similar experiments to be unhampered and
normal. Nor can it be set down to the cessation of the ordinary
muscular contractions of repose, viz. those of respiration and those
which serve to maintain the upright position ; unless, indeed, they
are assumed to have unexpected proportions. It might be caused by
some direct action of the curare upon the tissues which give rise to
carbon dioxide in the body ; and experiments were undertaken by
Colasanti3 to test this supposition. Curarized and non-curarized blood
was made to traverse, under precisely similar conditions, the right
and left hind limbs respectively of a recently killed muscular dog.
On comparing the outflowing blood from the two limbs it appeared
that there was no difference in the relative proportions of oxygen
and carbon dioxide contained in them. Hence the diminution must
be due to some influence of the central nervous system which is cut
off when the animals are paralysed by curare.

The experiments in which the spinal cord was divided shewed that,
even when the respiratory muscles were left freely acting, the separation

1 Zuntz, Op. cit., p. 527.

2 Pa tiger, Op. cit. Archiv f. d. ges. Physiol., Vol. xvin., 1878, p. 302. Finkler und
Oertmann, Archiv f. d. ges. Physiol., Vol. xiv. p. 62.

3 Giuseppe Colasanti, "Zur Kenntmss der physiologischen Wirkungen des Curare-
giftes." Pfliiger's Arch. f. d. ges. Physiol., Vol. xvi., 1878, p. 157.

26—2

404 FATIGUE, EXHAUSTION AND REVIVAL. [BOOK I.

of the rest of the muscles from the nervous centres was followed by a
diminution

of 0 consumed 37*1 p.c.

of C02 produced 29 -92 p.c.

but here the heart was weak and the circulation disturbed l.

Whatever may be the nature of the influence exerted by the
nervous system, it is probably reflex in its origin, and excited by the
difference of temperature between the skin and the external medium.
Liebermeister and Gildermeister2 shewed in the case of men that the
production of heat and the formation of carbon dioxide increase on
the application of cold to the surface of the body ; and Rohrig and
Zuntz confirmed this in the case of rabbits, by immersing them in
cold baths. If, however, the animals be first curarized, immersion in a
cold bath no longer stimulates the interchange of oxygen and carbon
dioxide, but rather tends to diminish the amount of both3. In other
words, the reflex mechanism is in abeyance, and the bath, by directly
cooling the tissues, renders their various processes more sluggish.

Curare-poisoning seems to have no diminishing influence over the
nitrogenous excretions of the urine4.

With regard to the constitution of muscle itself after separation from its
cerebro-spinal centres, it is said that it contains less creatine5 but more
glycogen".

SECT. 4. FATIGUE, EXHAUSTION AND REVIVAL.

Signs of Muscles are incapable of contracting continuously

Fatigue. for an indefinite time. They become fatigued more

or less quickly, and are finally exhausted, when the most powerful
stimulus fails to cause a contraction. The evidence of fatigue is a
slow contraction of small amplitude. The muscle contracts slowly
to its maximum, which is abnormally small; but especially does
it elongate more slowly and less perfectly than usual, approximating
the condition of the 'idiomuscular contraction' (p. 843) 7. The
rate of transmission of the wave of excitation is also probably
diminished during fatigue8.

1 Pfliiger, pp. eft., p. 320.

2 Quoted by Eohrig und Zuntz, pp. cit. Pfliiger's Arch. f. d. ges. PhysioL, Vol. iv.,
1871. a Pfliiger, Op. eft., p. 303.

4 Voit, Zeitsch. fiir Biol., xiv. p. 57, 1878. See Hofmann and Schwalbe's Jahres-
berichte, Vol. vn. pt. iii. p. 272.

6 Sczelkow, Centralblt. /. d. med. Wiss., 1866, p. 481 (Original not seen).

6 Macdonnel, "On the formation of Sugar and Amyloid substance in the Animal
Economy," Proceed. Roy. Irish Acad., Vol. vn. p. 276, 1860. Also Observations on the
Functions of the Liver, Dublin, 1865, p. 23. Ogle, "A Hypothesis as to the ultimate
destination of Glycogen, " St George's Hospital Reports, in. p. 149, 1868. Chandelon,
" Ueber die Einwirkung der Arterienunterbindung u. der Nervendurchschrieidung auf den
Glycogengehalt der Muskeln." Pfliiger's A rch. /. d. ges. PhysioL, Vol. xin. p. 626, 1878.

7 See also the condition known as "Contractur, or remnant of contraction," which
follows powerful direct stimuli: Tiegel, Pfliiger's Arch., Vol. xm. p. 71, 1876, and
Hermann, Pfliiger's Arch., Vol. xni. p. 370.

8 See Hermann's Handbnch der Physiologic, Vol. i. Abth. i. p. 58.

CHAP. IX.] THE CONTRACTILE TISSUES. 405

Measure of The progress of fatigue may be gauged by the effects

Fatigue. Ex- produced in a muscle on applying at intervals a constant
periments of stimulus. When a muscle is tetanized the course of
H. Kronecker. fatigue is indicated by the gradual extension of the
tetanized muscle, which takes place at first with accelerating, but
afterwards with diminishing velocity, until the original length is almost
attained. When the muscle (of a frog), which is moderately weighted,
is stimulated at regular intervals of 2 — 12 seconds with a constant
(maximal) stimulus from an induction-machine, the heights through
which the weight is lifted diminish regularly in an arithmetical
series; i. e. the curve of fatigue forms a straight descending line1.

Such a series of lifts may be called a fatigue-series, the members
of which exhibit a constant difference D. The value of D diminishes
as the intervals of stimulation increase ; but with constant intervals
it is independent of the load which the muscle is made to lift. That
is to say, the curves of fatigue with different loads are a parallel series
of descending lines so long as the intervals of stimulation remain the
same for all the loads.

Causes of The causes of fatigue and exhaustion are very

Fatigue and obscure ; but since the fatigue of muscles in which the
circulation has ceased may be readily removed by
renewing the current of blood or even by washing out the blood-
vessels with indifferent salt solutions, especially such as contain per-
manganate of potash (*05 per cent.)2, we may suppose them to be due
either to the accumulation of the products of contraction, or to the
defect of constituents, such as oxygen, which the blood can supply, or
to both these causes combined. It is at least certain that carbon
dioxide has an injurious influence upon muscles 3, which is shared by
the lactic acid arising during contraction4; while it has been observed
that the addition of '05 to *1 p. c. of sodium carbonate to a '6 p. c. salt-
solution enhances its power of maintaining the activity of a frog's
heart fed with such a solution5 : the beneficial effect of the addition
is gradually lost as the salt-solution continues to be used ; but it may
be regained by adding a fresh supply of the carbonate, or by shaking,
up the old solution in the air. Ranke found that all acids haTJre
a diminishing influence on irritability. It is said that lactic acid
diminishes also the electromotive force of muscle6. The accumu-
lation of these products, therefore, could not fail to promote ex-
haustion. It is also certain that the renewal of the blood current
through muscle is swiftly followed by the revival of fatigued muscles;

1 H. Kronecker, "Ueber die Ermiidung u. Erholung der Muskeln." Ber. der math-
phys. Classe der k. sacks. Gesell. der Wissensch., 1871, p. 690.

2 H. Kronecker, Loc. cit., p. 694.

3 Georg Liebig, Hermann, Op. cit.

4 J. Eanke, Tetanus. Leipzig, 1865, p. 350.

5 Sti^non, "Die Betheiligung der einzelnen Stoffe des Serums an der Erzeugung des
Herzschlages." Arch. f. (Anat. u.) PhysioL, 1878, p. 263.

6 Eanke, Op. cit. Eoeber, Arch. f. Anat. u. Physiol., 1870, p. 615. (Original not
seen.)

406 THEORIES OF MUSCULAR ACTIVITY. [BOOK I.

and that this is largely due to the presence of oxygen is shewn in the
experiments of Ludwig and A. Schmidt already described (p. 380)
(Kronecker). But how far the accumulation of the products of con-
traction, or the defect of oxygen, and probably of other constituents of
normal muscle, can be trusted to explain the fatigue of muscles which
are still within the body, it is impossible to say.

stenson's It is, well known1 that ligature of the blood-vessels

Experiment. supplying muscles is followed by paralysis and rigor
of the muscles: and the same consequence follows any stoppage
whatever of the blood-current. Renewal of the current, on the other
hand, is followed by a restoration of irritability provided that rigor
is not complete. There can be little doubt in this case, that the
paralysis is in part due to an interruption of the normal exchanges
between muscles and the blood, i. e. to the accumulation of carbon
dioxide and the acid of rigor, and to the defect of oxygen, etc.

Among the mechanisms which contribute to the revival of fatigued
muscles or the prevention of exhaustion, must be mentioned the vaso-
motor nerves. Ludwig and Sczelkow 2 discovered that venous blood
flows more rapidly from contracting than from resting muscles, — a
phenomenon which has been investigated by the pupils of Ludwig and
traced to the vaso-motor system. The dilatation of the blood-vessels of
muscle, upon which the accelerated outflow seems to depend, also
accompanies the reflex stimulation of muscles, and is said to be visible
under the microscope in the case of frog-muscles, even when the
circulation is stopped and the blood-pressure abolished 3.

It will be remembered that the first effect of subjecting a muscle
to a vacuum was to increase its irritability (p. 371) : and that the
same result followed the withdrawal of blood from muscles in the ex-
periments of Ludwig and A. Schmidt (p. 379), No explanation of
this has yet been given.

SECT. 5. THE THEORY OF MUSCULAR ACTIVITY.

The archives of Physiology contain many curious speculations as to
tha nature of muscular motion ; nor is this surprising when we reflect
that the power of self-movement must ever have appeared the chief
and most characteristic attribute of animals. Many of the hypotheses
are extremely fanciful, and most of them are incomplete, offering to
explain certain elements only of the complicated act of contraction. In-
deed few theories are burdened with a heavier task than that which
comprehends all the phenomena of a muscular act. It must explain
how nervous stimuli affect muscle, and how contraction is transmitted

1 Steno, quoted by Schwammerdam (de Bespiratione, Leydeu, 1679, p. 61), and by
Haller (Elementa PhysioL, iv. p. 544. 1762).

2 Ludwig and Sczelkow, Op. cit. See p. 375.

3 See inter alia the researches of Gaskell, Studies from the PhysioL Lab. of Cambridge,
in. 1877 (Journal of Anat. and PhysioL Vol. XL); Journal of PhysioL, ed. by M. Foster,
Vol. i., 1878.

CHAP. IX.] THE CONTBACTILE TISSUES. 407

along the fibres ; it must satisfy us as to the actual method of shorten-
ing— as to the momentary disturbance of the structural elements of
the fibre; and, finally, it must account for the origin of the heat,
electrical disturbance and mechanical motion which are characteristic
of muscular contraction, and suggest some relationship among them.
Most theories have been content to explain one or other of these
points without attempting the whole.

The point which most nearly concerns the physiological chemist
is the last mentioned, viz. the origin and interdependence of the heat,
electrical tension and mechanical motion of contraction, and this
alone will be considered in the present chapter. The probable nature
of nervous action will receive consideration elsewhere in this book ;
and the views of observers as to the appearances of contracting muscle
beneath the microscope will be found in Manuals of Histology.

John One of the most remarkable anticipations of modern

Mayow. discovery and opinion occurs in the works of John

Mayow, published in 1668 — 16741. Exactly one hundred years before
the discovery of oxygen, Mayow demonstrated by a series of con-
clusive experiments, that some portion or constituent of the air is
necessary to combustion, and that the same substance is equally
indispensable to living animals. To this portion of the air he gave
the name of particulae igneo-aereae. The same substance enters into
the composition of nitre ; since, when nitre is present, combustible
bodies can inflame and burn even in vacuo or beneath water. Hence
Mayow called the igneo-aerial particles, nitro-aerial particles and nitro-
aerial spirit. This substance, though indispensable to combustion,
does not itself burn ; but when antimony is calcined in the focus of a
burning-glass, or by exposure to the flame of nitre, the antimony
becomes not a little increased in weight — a circumstance which Mayow
could only explain by supposing the fixation of igneo-aerial particles in
the calcined antimony2. Respiration introduces igneo-aerial particles
into the blood, where they meet with saline-sulphureous (i.e. combusti-
ble) particles and produce the animal heat3. From the blood the igneo-
aerial particles are conveyed to the muscles4, where they meet with

1 John Mayow, Tractatus quinque Medico-physici. Oxford, 1674.

2 J. Mayow, de Sal-nitro et spiritu nitro-aereo, Chap. iii. p. 28. "Hue etiam facit,
quod Antimonium non tantum a Spiritu Nitri, radiisque solaribus, sed etiam a flamma
nitri, in qua particulae nitro-aereae densius agglomerantur, virtutem Diaphoreticam
acquirit. Neque illud praetereundum est, quod Antimonium, radiis solaribus calcinatum,
haud parum in pondere augetur; uti experientia conipertum est: quippe vix concipi
potest, unde augmentum illud Antimonii, nisi a particulis nitro-aereis, igneisque ei inter
calcinanduni infixis, procedat."

3 Ibid., Chap. viii. p. 151.

4 J. Mayow, de Motu Musculari et spiritibus animalibus, Op. cit., part ii. p. 3.
" Spiritum nitro-aereum respirationis ope in Cruoris massam transmitti, Sanguinisque
Fermentationem, et Incalescentiam ab eodem provenire, alibi a nobis ostensum est. lam
vero circa usum Spiritus istius inspirati addo insuper, quod idem in Motibus Animalibus
instituendis partes primarias sortiatur : quam quidem opinionem a me jam olim in
medium prolatam, etiamnum firmiter retineo, non quod praeconceptae Hypothesi man-
cipatus, earn, uti moris est, mordicus defendere constitui, sed quod eandem rationi
maxime consentaneam arbitror."

408 THEORIES OF J. MAYOW. [BOOK I.

other salino-sulphureous particles secreted from the mass of the blood,
and by union with these cause an effervescing which produces muscular
motion. How the nitro-aerial or igneo-aerial particles reached the
muscles, Mayow did not feel quite certain. He was sometimes inclined
to think that they proceeded directly to the muscles from the blood ; but
it appeared more probable on further reflection, that the salino-sulphur-
eous or motive particles alone were supplied directly by the blood; while
the nitro-aerial particles approached the muscles through the brain and
nerves, being the same, in short, as the animal spirits. Why, asked
Mayow, should not the animal spirits be derived from the air rather
than from the food ? Indeed, it seemed to him impossible that the
immense waste of animal spirits could be supplied from any other
source. The salino-sulphureous or combustible nature of the motive
particles was thought to be shewn by this, that in violent exercise no
small loss of fat occurs, and if exercise is long continued fat almost
disappears : on the contrary, animals leading an easy inactive life grow
fat, and fat appears in large quantity in the muscles. A supply both
of igneo-aerial and of salino-sulphureous particles is indispensable to
continued animal motion. As movement is increased and more of
each sort of particles is wasted, more of them must be added to the
body. Not only must respiration be enlarged, but more food contain-
ing salino-sulphureous parts must be taken. Hence those substances
which contain much volatile salts and sulphur (i.e. combustible matter)
are best fitted to recruit the frame worn out by protracted labour.
Finally, Mayow clearly recognized that the animal heat arises, not solely
in the union of nitro-aerial and combustible particles in the blood
generally, but in that special union which is accomplished in muscles
during muscular contraction : part of the heat of an animal in violent
exertion arises in the union of nitro-aerial and salino-sulphureous par-
ticles in muscle1.

This was in 1674. When we remember that it was nearly two
hundred years before physiological science fully overtook the specula-
tions of Mayow, — that, although oxygen was discovered in 1774, it
was not until 1861 that Moritz Traube definitely announced that
muscular contraction depends upon the combustion of non-nitrogenous
matters in muscles themselves — we shall feel no surprise that Mayow's
work was so speedily forgotten. Scientific judgment must have
been strangely uneducated to have allowed the experimenters of
that day to read and lose sight of observations which seem to
us now so exact and suggestive. It has been the fate of Mayow,
which his genius little merited, instead of leading science, to be twice
revived by antiquarian zeal, at the very moment when his dis-
coveries had been made over again by independent observers. After
the researches of Priestley, Scheele and Lavoisier had made brilliant

1 Op. cit., part i., de sal-nitro etspiritu nitro-aereo, p. 152. "Quanquam calor iste in
animalibus, per exercitia violenta excitatus, etiam ab effervescentia particularum nitro-
aerearum et salino sulphurearum in partibus motricibus ort&, partim provenit, ut alibi
ostendetur.

CHAP. IX.] THE CONTRACTILE TISSUES. 400

the close of the last century, the importance of Mayow's work was
proclaimed by the enthusiasm of Dr Beddoes1 and Dr Yeats2; and in
1864, at a time when the theory of muscular activity had already
received its present bent, the acute speculations of Mayow in refer-
ence to it were again most honourably made known by Professor
Heidenhain 3.

It would be pushing literary justice to the extreme verge of pedantry to
pretend to find in authors earlier than Mayow the germs of a theory which
they were not in a position even to comprehend ; still it is interesting to
observe that the general idea of a combustion of matters within the body,
upon which the powers of life depend, is to be found in a book with which
Mayow was probably familiar. Francis Bacon, in his Historic*, Vitae et
Mortis, taught that all living beings contained two kinds of spirits, spiritus
mortuales which fill inanimate objects, and spiritus vitalis which confers life.
The doctrine of a vital principle stirring and regulating the members of
living creatures had existed, in one form or another, from the earliest times ;
but more than this Bacon taught that the spiritus vitalis exhibited a certain
incensio, or combustion, which gave rise to peculiar motions and powers.
"In omnibus aniniatis duo sunt genera spirituum : spiritus mortuales, quales

insunt inanimatis ; et superadditus spiritus vitalis Sunt autem duo

discrimina praecipua inter spiritus mortuales et spiritus vitales Alterum

discrimen inter spiritus est ; quod spiritus vitalis nonnullam habeat incensi-
onem, atque sit tanquam aura composita ex flamma et ae're ; quemadmodum
succi animalium habeant et oleum et aquam. At ilia incensio peculiares
praebet motus et facilitates; etenim et fumus inflammabilis, etiain ante
flammam conceptam, calidus est, tenuis, mobilis; et tamen alia res est,
postquam facta sit flamma ; at incensio spirituum vitalium multis partibus
lenior est quam mollissima flamma, ex spiritu vini, aut, alias; atque
insuper mixta est, ex magna parte, cum substantia ae'rea ; ut sit et flammae

et aereae naturae mysterium ...Neque tamen ulla ex ipsis actioni-

bus unquam actuata foret (i. e. of the stomach, liver, heart, brain etc.),
nisi ex vigore et praesentia spiritus vitalis et caloris ejus4."

The obvious and extreme importance of air for the support of life,
and the muscular weakness which follows excessive bleeding, did not
escape the earliest observers, and were the foundation of hypotheses which
have been thought to foreshadow the modern view5.

After the time of Mayow, the doctrine of muscle was

Glisson. mainly given over to the Stahlists. Armed with his

conception of an immaterial and rational anima endowed

with unlimited spontaneous powers over matter, Stahl explained

nearly all things with equal facility, and among them muscular

1 Thomas Beddoes, Chemical Experiments and Opinions extracted from a work
published in the last century. Oxford, 1790.

2 G. D. Yeats, Observations on the claims of the Moderns to some discoveries in
Chemistry and Physiology. London, 1793.

3 Heidenhain, Mechanische Leistung, etc. Leipzig, 1864.

4 Francis Bacon, Historia Vitae et Mortis, 1(323, Can. iv. and v. "Works," by
Spedding, Ellis and Heath, 1857, Vol. n. p. 214.

5 Al. von Humboldt, Versuche u. die gereizte MusJcel- und Nervenfaser, Yol. n. pp.
91, 93. 1797.

410 GLISSON. HALLER. WHYTT. [BOOK I.

motion1. His opinion generally prevailed until the time of Haller,
whose doctrine of the independent irritability of muscle marks the next
advance in the theory of muscular contraction. The term Irritability
was not indeed new to Physiology. The name, and in a certain sense
the notion also, was introduced by Glisson in the latter half of the
seventeenth century. He taught that irritability was a property of
the elements of our bodies, even of the bones and juices, which was to
be attributed to a natural perception unaccompanied by any sensation
whatever. It was supposed to depend upon 'Archaeus who is the
framer of his own body' ; and it could be demonstrated after death by
the application of acid and pungent liquors2. Bat it was Haller who
first gave the idea a firm foundation in experiment. Resting on the
experiments of Haller and his pupils, this important doctrine was
definitely formulated in a Treatise on the Sensible and Irritable
parts of Animals3. Irritability was defined as the property, pos-
sessed by muscular fibres alone in the body, of shortening when
they are touched ; while those parts were called sensible which,
when handled, transmit the impression of the touch to the soul, or,
in animals, lead to evident signs of pain and disquiet. Irritability is
distinct from sensibility, since nerves, the most sensitive of structures,
-are absolutely^! e void of irritability. Stimuli applied to nerves, how-
ever, lead to convulsions and palpitations of neighbouring muscles,
but only in such as are directly supplied by the nerve stimulated.
Muscles contract after separation from the brain, after their nerves
are all cut away, and even after removal from the body. Hence
irritability is a property quit&^apart from the soul and the nerves.
Haller thought it probable that, ^me time or other, the use of the
nerves with regard to the muscles would be reduced to conveying to
them the commands of the soul, and to increasing and exciting that
natural tendency which the fibres have of themselves to contract4.
The property of producing motion is different from all other proper-
ties of bodies, and it probably resides in the glutinous mucus rather
than in the earthy parts of muscles. It is a property of muscles as
gravity is a property of matter generally, and it is doubtless owing
to a physical cause depending on the arrangement of ultimate parti-
cles. It is destroyed by drying the fibre, as well as by opium.

^ The most active opponent of Haller in this country

was Robert Whytt5, Professor of Medicine in the Univer-

1 Georg. Ern. Stahlius, Theoria Medica Vera, 1708. Ed. Lud. Choulant. Lips., 1831.
Tom. i. sec. vi. p. 466. Georg Ernst StahVs Theorie der Heilkunde. Dargestellt von
Wendelin Ruf. Halle, 1802, p. 206.

2 Francis Glisson, de Ventriculo et Intestinis, c. vii. Quoted by Haller, Op. cit.,
infra.

3 Haller, " de Partibus corporis humani sensilibus et irritabilibus." Commentarii
Soc. reg. Scientiarum Gotting. Tom. n. 1752, p. 114. A Dissertation on the Sensible
and Irritable parts of Animals. Translated from the Latin. London, 1755.

4 Haller, Loc. cit., p. 139.

6 Robert Whytt, Physiological Essays, Edinburgh, 1766. Third Edition. On the
Vital and other Involuntary Motions of Animals, Edinburgh, 1763, Second Edition.

CHAP. IX.] THE CONTRACTILE TISSUES. 411

sity of Edinburgh, whose criticism of Haller displays the greatest
ingenuity and address. This acute observer was not a disciple of Stahl;
indeed his doctrine was disclaimed by the purer Stahlists ; but it forms
a link between the teaching of Stahl and the doctrine of vital force
of the next generation. According to him all parts of the body are
pervaded by a Sentient Principle which is affected more or less acutely
by stimuli or irritants; and the motions which invariably follow
irritation and are always in proportion to the strength of it, are the
endeavour of the pervading principle to remove the part from the
source of irritation ; it acts upon the muscles through their nerves,
but in a manner altogether obscure. Whytt's sentient principle is
the soul of the Stahlists shorn of its rationality and spontaneity, and
bound by an original decree to the task of responding by movements
to every stimulus impressed upon the body. This principle remains
for a time in parts amputated from the body ; hence such parts are
capable of contractions when touched. It is the merit of Whytt to
have insisted upon tha importance of the stimulus in all involuntary
actions, and the invariableness of the motions excited by it.

The experiments of Haller dealt a fatal blow to
Jolm Stahlism and the like. Already in the writings of

Whytt we see this doctrine subsiding into the simpler
one of vital force, as it is implicitly adopted, for instance, in the pages
of John Hunter. Hunter1 was content to classify muscular motion as
one of the forms of the movement of matter; of which the attraction
of masses owing to gravitation was another form and the elective
attraction of chemical substances was a third. He thought that it
most probably arose from construction : but it was a principle in action
very different from the attractions in common matter, and equally
unintelligible with gravity and chemical attraction. In short, the
current view of the cause of muscular motion was that it was original,
a vis insita, a vital power peculiar to living tissue during its life.

Fothergiii, It was not long, however, before the doctrine of

and Girtan- vital force began to be expanded. The re-discovery of
oxygen had quickened philosophical speculation, and
seemed to have placed in the hands of physicians a remedy of the
greatest promise. One of the first methods of treatment to be bene-
fited by the new chemical discoveries was the art of restoring sus-
pended animation. Inflation of the lungs was empirically known to
be extremely useful in such cases ; but it was Dr. A. Fothergill2 who
first suggested an explanation of its value which, if not wholly true,
was true in the greater part. "In all cases of suspended animation
the grand intention ought to be, to excite the latent principle of
irritability on which the motion of the vital organs immediately
depends." And how, asks he, can this be better done than by

1 Croonian Lectiire, 1781. Works, Ed. by Palmer, Vol. iv. p. 255.
' 2 A. Fothergill, Hints for improving the art of restoring Suspended Animation, 1782,
pp. 15, 17, 18.

412 GIRTANNER. [BOOK I.

inflating the lungs, not with common air used again and again, but
with fresh supplies of dephlogisticated air? If dephlogisticated air
supports the name of a taper with a splendour hardly credible by
those who have not witnessed the experiment, is it not clearly
indicated as peculiarly fitted for restoring the vital spark when
nearly extinguished ?

The theory thus suggested soon received a remarkable amplifica-
tion in the hands of Girtanner1. In an essay, exhibiting great re-
search (but at the same time shewing that a restless desire to have an
answer of some kind to his questions had led its author to content
himself with mere specious explanations), the principle of irritability
is identified with oxygen. All organized parts, whether of the animal
or vegetable kingdom, are considered to be capable of irritability,
and the irritable fibre is one and the same in nature wherever
found. Even the fluids of the body exhibit an irritable contraction
(i.e. coagulation) and obey the general laws of irritability. The prin-
ciple of irritability, or oxygen, is received by the blood in the lungs,
and conveyed to all parts of the body, where it is stored up in the
irritable fibres. There is a normal quantity for each fibre, upon which
its tone depends ; and the normal is preserved by the ceaseless action
of habitual stimuli, such as heat, light, nourishment, circulation of the
juices, etc. which withdraw the surplus. Thus the health of the fibre
depends upon an even balance of gains and losses of oxygen. If the
gains are excessive, irritability becomes increased. If the losses are
excessive through the extraordinary action of the habitual stimuli,
irritability sinks or disappears altogether. All reagents which are
thought to be capable of acting upon irritable fibres are divided into
three classes according as their affinity for oxygen is greater than, equal
to, or less than, that of the fibres. The former abstract oxygen from
the fibres and depress their irritability : such are opium, alcohol, fat.
The latter impart oxygen to the fibres, producing a super-irritability
which is often extremely fatal : white oxide of arsenic, Vacide muria-
tique oxigene, are eminent examples of this class. The intermediate
class behave as neutral bodies, until change of temperature, or some
other condition, removes them into the first class or the third. Thus
irritability is always in proportion to the oxygen which an irritable
organ or fibre contains; and whatever augments or diminishes the
oxygen of the body likewise augments or diminishes its irritability.

Whether Girtanner had ever read Fothergiil's Hints does not
appear, nor is it of importance to enquire ; but it is certain that
Fothergill, four years after the publication of Girtanner's Memoirs,
expanded his original conception into a theory of irritability which is
practically identical with Girtanner's2. Vitality, in his view, consists

1 Girtanner, "Me'moires sur 1'Irritabilite, considered comme principe de vie dans la
Nature organisee." Observations sur la Physique, ed. by Rozier and de la Metlierie.
1790, Vol. xxxvi. p. 422; Vol. xxxvn. p. 139.

2 A. Fothergill, On the Suspension of Vital Action, Bath, 1795, p. 67 and else-
where. This Essay gained the Gold Medal of the Eoyal Humane Society in 1794.

CHAP. IX.] THE CONTRACTILE TISSUES. 413

in action and reaction between the vital organs and their respective
(habitual) stimuli, exactly in the sense of Girtanner. Irritability
co-exists with animal heat and keeps pace with it through life :
hence it probably has a similar origin. But, inasmuch as animal
heat can be shewn to be dependent upon vital air (oxygen), whose
latent heat, in short, is the source of the animal heat, may not vital
air be also the proximate cause of irritability I

Beddoes a^ove v^ew was received with much favour, and

therapeutical use was freely made of it. Dr Beddoes
discussed it in his Remarks on Girtanner s Essay, and is stated to have
specially pointed its reference to the case of muscle by the question,
Does muscular action or intumescence really depend upon the com-
bination of oxygen with hydrogen or azote (separately and combined in
various proportions) in consequence of a sort of explosion produced by
the nervous electricity1 ? The advances which scientific theory makes
are often so insidious that we are apt to underrate their importance.
It is hardly too much to say that this question of Beddoes marks the
real point of departure of the modern views of muscular irritability.
Previously, the prevailing tone of thought had been semi-metaphysical.
Oxygen had been regarded as a principle, the presence of which
conferred irritability upon tissues, and the withdrawal of which from
the organs and fibres happened to be effected by its union with other
elements. Now attention was concentrated upon the union itself
rather than on the uniting bodies, and irritability was regarded as the
result of a process, and not the attribute of a substance. A taper, a
lamp, a fire, became the fashionable metaphors wherewith to illustrate
various physiological acts. Food was not useful food if it had no
affinity to oxygen. Life itself was but the burning of a lamp of which
the body is the wick and food the oil.

Brandis Illustrations like these did not go uncriticised. "Such

metaphors," remarked Brandis2, the German translator of
Darwin's Zoonomia, "are apt to cast shadows where we would fain have
light." It is indeed certain, argues this author, that phlogistic pro-
cesses occur in the body as in the combustion of other substances :
carbon is united to oxygen and expired as carbonic acid gas ; phos-
phorus appears to become acidified in the body and is excreted in the
urine in combination with lime ; and such probably is the case with
other constituents of the fibres. But there is as yet nothing to shew
that these bodies can excite themselves to union. Some external force
is needed to start the combustion, and to determine the intensity of it
in any particular act. This is the unknown and subtle vital force,
which is as indispensable to the vital processes as the spark is to the
kindling of a fire.

1 Yeats, Op. ct«., p. 171. Al. von Humboldt, Op- cit., Vol. n. p. 105. The original
Remarks seems to be somewhat rare, as neither Prof. Eudolf Heidenhain nor the author
has succeeded in obtaining a copy.

2 J. D. Brandis, Versuch. ii. die Lebenskraft. Hannover, 1795.

414 EEIL. HUMBOLDT. J. LIEBIG. [BOOK I.

Reil and On the other hand, Reil and his brilliant pupil

von Madai. von Madai1 avoided even the semblance of a vital or
irritable principle by referring to the structure and chemi co-physical
changes of living matter all the peculiar phenomena of life. As to
the nature of the changes Reil was not explicit ; and, although von
Madai adopted provisionally the term phlogistic processes, because
carbon and oxygen play so important a part in them, it was with a
wise reservation as to their exact nature and method (loc. cit. p. 101).

It was, however, Humboldt2 who first denied with
Humboldt. . , , . ,, ' , . . ,,

special emphasis the exclusive importance of oxygen in

the vital processes. Many observations had disclosed to him vital
processes in which oxygen takes none but a subordinate part. To
speak of life as an oxidation is to take a one-sided and distorted view
of vital phenomena. Oxygen in his opinion is a most important
stimulus, but not the common basis, of irritability. It is true that
many phlogistic processes occur in the performance of vital functions :
but how many other chemical decompositions go forward which do
not so much express the affinity of oxygen for phosphorus, azote,
hydrogen and carbon, as the affinities of these for one another ?
Thus physiology became accustomed to the absence of a particular
contractile or irritable principle. First its office and dignity were
conferred upon oxygen, and then oxygen was reduced to the rank
and privileges of a common chemical element.

. The doctrine of vital force in muscular action by no

means at once gave place to the views of the physico-
chemical school. It remained indeed the prevalent doctrine until
the time of Baron Liebig8, who attempted to bring it into rela-
tion with the most recent discoveries of Physiological Chemistry.
The vital force, resident in animals and plants, finds its scope of
action in the presence of a certain structure and organization of parts.
It is governed by laws in harmony with the universal laws of resistance
and motion ; nevertheless it is independent of the matter in which
vitality is manifested. It is this vital force which keeps living matter
from decomposition even in the presence of oxygen, which determines
its growth, and which also causes the movements of animals. Besides
a special organization of substance, a certain temperature and a
constant supply of food are indispensable conditions of its activity.
A muscle therefore is an organ endowed, by virtue of its vital proper-
ties, with certain powers of self-preservation, growth and motion.
While it is at rest, and exposed to the influence of oxygenated blood,
the vital force is absorbed in restraining the natural tendencies to

1 J. C. Eeil, "Ueber die Lebenskraft," Arch, fiir die Physiologic (Eeil), Vol. i. Heft i.
p. 8. 1796. D. von Madai, "Ueber die Wirkungsart der Eeize und der thierischen
Organe." Ibid., Vol. i. Heft iii. p. 68. 1796.

2 Al. von Humboldt, Versuche it. die gereizte Muskel- u. Nervenfaser. Vol. n.
p. 106 et seq.

3 J. Liebig, Animal Chemistry, or Organic Chemistry in its applications to
Physiology and Pathology. Translated by W. Gregory. London, 1842.

CHAP. IX.] THE CONTRACTILE TISSUES. 415

disintegration and union with oxygen. But if a portion of the vital
energy is diverted for the purposes of contraction, the natural incli-
nation of the muscle to change is, in part, unchecked, and a certain
portion of the tissue becomes oxidized and dead. For every motion
of contraction there is a material exchange, with an absorption of oxy-
gen, and a certain amount of tissue cast off. The relationship among
these is invariable : for every portion of force expended in motion
there is a definite proportion of tissue wasted and oxygen absorbed.

Besides attacking living tissues which are for the moment left
unprotected by the vital force, the oxygen which is absorbed into the
body attacks those also which are already lifeless ; whence arises the
heat and the proper temperature of the body which is so important a
condition of vitality. The production of heat and force in the body
are indeed closely related; but heat can be produced without any
change in the living elements of the body, while mechanical effect
is always proportional to the amount of living matter which loses the
condition of life. However closely the conditions of this twofold
production seem to be connected in regard to mechanical effects,
yet the disengagement of heat can in no way be considered as in
itself the cause of these effects. All experience proves that there is in
the organism but one source of mechanical power, the conversion
of living into lifeless compounds.

Thus for every portion of oxygen taken into the body there is
a corresponding proportion of heat and mechanical force produced.
Further, the amount of tissue metamorphosed in a given time is
measured by the nitrogen in the urine; and the sum of the mechanical
effects produced in each of two individuals at the same temperature,
is proportional to the nitrogen excreted by each.

The views of Liebig were not left unchallenged.
J. R Mayer1, the early apostle of the conservation of
energy, in his treatise on organic movement in relation to material
exchange, exposed the inconsistencies of the doctrine of vital force as
expounded by Liebig, and stated that not only the heat, as Liebig
admitted, but also the muscular motions, of animals had but one
source in the oxidation of combustible matters. He calculated from
the combustion- heat of carbon, that the extraordinary consumption of
combustibles by a labouring animal, bearing in mind the enlarged
production of heat during labour, is fully competent to account in
a natural way for the work done. But the combustions in which the
animal movements take their origin are not combustions of the
substance of muscle itself. To account for the production of motion
in this way, we should have to assume a rapid destruction and
restoration of muscular tissue, of which there are no sufficient histo-
logical and physiological signs. The oxidations take place rather
in the blood, which is the true oil for the flame of life.

1 J. E. Mayer, Die organische Bewegung in ihrem Zusammenhang mit dem Stqft'~
wechsel. Ein Beitrag zur Naturkunde. Heilbronn, 1845.

416 VOIT. TRAUBE. [BOOK 1.

Thus the oxidations of the body result in the generation of heat
and of motion, which therefore, within limits, are complementary to
each other. With a certain chemical combustion, the greater the
mechanical effects produced, the less the amount of heat which
appears ; and conversely. During the performance of mechanical
work a proportion of the heat which would otherwise have been
sensible becomes 'latent,' — this proportion being equivalent to the
work done.

In this manner J. R. Mayer emancipated muscle from the doctrine
of vital force, and taught the true source of muscular power in the
chemical union of substances. Muscles, according to him, are an
apparatus for the conversion of chemical difference into mechanical
effect, just as plants are an apparatus for converting light into
chemical difference ; and this power of living muscle is what con-
stitutes irritability. At the same time experiment was surely estab-
lishing the other opinion which Mayer had, on theoretical grounds,
opposed to Liebig's teaching, viz. that muscular exercise is not
associated with an extraordinary destruction of the nitrogenous
substance of muscle.

Voit The experiments of Voit * on dogs, which have already

been described, may be said to have effected the final
overthrow of the older views. Voit himself seems to have mistaken
the meaning of these experiments. He was compelled to admit that
no more nitrogenous waste occurred in muscular exercise than in
muscular rest ; but he appears to have taken no account of the well-
marked increase of respiratory products in the same circumstances.
He drew the conclusion that no more energy is expended during
exercise than during rest, but the same energy takes another form ;
and as he found no evidence that this transformation was one of heat
into mechanical motion, he supposed that it was a conversion of
electrical energy. This view was never much encouraged.

Moritz Traube, on the other hand, who was inves-
tigating the subject when Voit published his researches,
recognized at once the great importance of the experiments, and
explicitly formulated the view that no albuminous body is used up in
muscular contraction. On the contrary, muscles contribute rather to
the non-nitrogenous respiratory excretions. They are a chief seat
of the oxidations of the body, and by means of their nerves the
oxidations which occur in them are made to yield mechanical energy.

_, «.. . A still more interesting advance in the theory of

Matteucci. ,. i • i

muscular contraction concerns the oxygen which serves

the oxidations of muscle. Already in 1856 Matteucci2 had remarked

1 C. Voit, Untersuch. u. den Einftuss des Kochsalzes, etc., auf den Sto/iuechscl.
Munchen, 1860.

2 Ch. Matteucci, ' ' Kecherches sur les phe'nomenes physiques et cbimiques do
la contraction musculaire. " Ann. de Chimie et de Physique, 3e Se'rie, Vol. XLVII., 1856,
p, 129.

CHAP. IX.] THE CONTRACTILE TISSUES. 417

that muscles which for some hours had been rigorously excluded
from contact with oxygen gas, were yet capable of yielding carbon
dioxide, especially when in the act of contraction ; and he had con-
cluded that the oxygen which in 'muscular respiration' forms the
carbon dioxide is not the oxygen of the air but oxygen which exists in
muscle in a state of chemical combination. The same circumstance
attracted the attention of Traube. In his view, oxygen enters muscle
from the blood and unites there in some loose chemical combination,
from which it is readily abstracted by the oxidizable bodies of the muscle
juices. Muscular substance reacts to oxygen and reducing substances
like indigo, cupric hydrate or the vinegar ferment, and its action is more
perfect and rapid at the higher temperatures. Complete deoxidation
of a muscular fibre brings with it death-rigor, while complete saturation
with oxygen implies a perfect irritability.

Heidenn i "^n ^s manner the doctrine of muscle was begin-

ning to assume its present outlines when Heidenhain1
demonstrated that the heat and mechanical work produced in con-
traction are not complementary — that, in short, they vary in a similar
although not quite identical manner, when subjected to the same con-
ditions of tension, etc. The hypothesis of J. R. Mayer that mechanical
work arises at the expense of heat in muscle, which many observers
had endeavoured to sustain, became finally untenable; and it was
now necessary to assume that the heat-evolving and work- evolving
processes of muscle were in some degree independent of one another^

„ At this point Hermann2 began his well-known

Hermann. • , • r /• , • , • > p ^

examination ot muscular respiration, most 01 the

results of which have already been presented to the reader. Although
it was granted that the oxygen made use of in the formation of
carbon dioxide was not taken from the blood at the moment of!
formation, but was rather stored up in muscle at some time beforehand,
yet it seems to have been assumed that the act of formation of carbon
dioxide was a true oxidation ; and for this reason it had been found
necessary to suppose the existence of some body with affinities for
oxygen intermediate between those of haemoglobin and the oxidizable
matters of muscle. The great and peculiar stability of this hypothetical
oxygen-carrier, which could, while easily parting with its oxygen to
the oxidizable portions of muscle in contraction, yet steadily resist the
action of a vacuum even at high temperatures, was however always
a point of great difficulty ; and, to avoid it, Hermann surmised that
the chemical operation in contracting muscle is not a true oxidation,
but rather the splitting up of some complex body with the formation
of simpler, more stable, substances. Such decompositions were already
known to be capable of yielding energy, and especially heat ; as, for
instance, when the complex molecule of sugar, in the process of
fermentation, splits up into alcohol, carbon dioxide, etc., without
the help of oxygen from the air.

1 Heidenhain, Mechanische Lcistungen, etc.

2 Hermann, Stoff'wechsel der Huskeln.

a 27

418 HERMANN. [BOOK I.

As to the supposititious substance itself, all that direct observation
suggested was that it must be of such a nature as to yield carbon
dioxide and some free acid, probably lactic ; but by reflecting upon
the analogies of contraction and rigor, Hermann was led to assign to it
a very complicated structure. The resemblances of contraction and
rigor are manifold. In each there occurs a shortening, thickening, and
small reduction of bulk of the muscle ; and a mechanical force is
developed to each, although at very different rates. Both processes
are associated with an evolution of heat ; and we may now add that
contracting muscle and muscle becoming rigid assume the same electric
potential in reference to living and resting muscle. With regard to
their chemical changes, both processes are independent of the oxygen of
the surrounding medium, and both are followed by the appearance of
a free acid and the formation of carbon dioxide. Further, there is this
relationship between the two in the case of excised muscles, that
the more free acid and carbon dioxide are produced by the previous
tetanus of the muscle the less are generated on subsequently passing
into rigor. Moreover, phenomena are known which seem to be most
naturally regarded as intermediate states between contraction and
rigor. If a fatigued muscle receives a sharp stimulus, as from a
sudden blow, a local (idio-muscular) contraction is produced which
lasts for a long time ; and if such an exhausted muscle be repeatedly
stimulated it may pass at once into true rigor.

If, then, contraction so closely resembles rigor, may we not consider
it as a transitory form of rigor, and assume that we have in contraction
what undoubtedly occurs in rigor, viz. the separation of a coagulum of
myosin ? In rigor the coagulum at once passes to a condition of con-
* tracted clot : here, therefore, the analogy must end ; for in normal
irritable muscle the clot never goes beyond the gelatinous stage.

Hermann's hypothesis may thus be summed up : The chemical
substratum of muscular activity is the falling to pieces of a complex
nitrogenous body, which has been called Inogene substance. The pro-
ducts of the decomposition include carbon dioxide, a fixed acid, and a
gelatinous albuminous body, of which the first is cast into the blood-
current, while the last, and possibly the second also, help to build
up again the original compound. The decomposition is constantly
occurring, even during the repose of muscle ; but in such circumstances
restoration keeps pace with destruction. In contraction, on the
contrary, destruction largely exceeds restoration. The blood supplies
to muscle the non-nitrogenous matter and the oxygen needed for
the reproduction of Inogene substance.

Along with the chemical changes of Inogene substance, other
changes occur in the regeneration of muscle itself. These affect the
nitrogenous as well as the non-nitrogenous elements of the tissue,
and help to swell the nitrogenous excreta. In cases of severe exertion
it is not improbable that these changes may be unusually large ; and
this especially would be the case were individual muscular fibres to
become rigid and stand in need of absorption and removal.

CHAP. IX.] THE CONTKACTILE TISSUES. 419

In this manner the theory of Hermann brings together all the
chemical facts of muscular contraction : but it has the further merit
of attempting to solve the electrical facts also. It is entirely beyond
the scope of this work to explain in detail how this is accomplished.
It will be sufficient to state that the key to the most complicated
electrical phenomena of resting and acting muscle is to be found in
the contact of heterogeneous substances; and that the heterogeneity
required by the theory is supplied by the chemical difference which
undoubtedly exists between resting muscle on the one hand, and dying
or contracting muscle on the other. Bat, while the theory is so far
satisfactory, we must not blind ourselves to the capital imperfection of
it, that it does not represent to us how contraction itself takes place.
To suppose that in the hypothetical formation of gelatinous myosin
the physical particles of muscle are drawn together or suffer a
rearrangement, is but to support one hypothesis by advancing another ;
the explanation of muscular contraction on the view that it is
due to the shortening of gelatinous myosin has always appeared to
us improbable, and it is certainly not countenanced by any known
facts.

27—2

CHAPTER X.

THE NERVOUS TISSUES.

SECT. 1. INTRODUCTORY.

ciassifica- THE organs which compose the nervous system of the
tion of nerve- higher animals may be classed as 1st central organs,
organs. ^^ ^ ^Q brain and spinal cord and the various

peripheral or sporadic ganglia ; 2nd conducting structures or nerves
which are engaged in bringing into communication the central organs
with, 3rd, end-organs wherein the fibres of certain of the nerves
(afferent) commence, and those of certain others (efferent) terminate.

Grey and ^he large nerve-centres are composed of grey and

•white matter white matter, properly so called. In the grey matter
of the nervous reside the nerve-cells which are the characteristic ele-
ments of the central organs, and which for the most
part certainly have a connection direct or indirect with nerve-
fibres.

The white matter is composed of nerve-fibres, making their way
to and from the grey matter, and only very exceptionally contains
nerve-cells. Both grey and white matter are supported by a connec-
tive-tissue framework termed the neuroglia ; both are supplied with
blood-vessels which penetrate from the surrounding membranes,
though the grey matter is much more vascular than the white ;
in both we can trace the commencement of lymphatic vessels.

Nerve-cells.

Nerve-cells are irregular masses of protoplasm, possessed of a well-
marked nucleus and nucleolus, and sending out one or more processes.
The protoplasm of the cell is often somewhat pigmented (greyish) ; in.
the nerve-cells of the ganglia of Aphrodite aculeata it has been shewn
to be tinged of a red colour, due to the presence of haemoglobin. That
part of the protoplasm which immediately surrounds the nucleus is
granular, while, in most cases, that which is disposed at the periphery
of the cell exhibits a striated appearance which seems to be similar
to, and indeed continuous with, that often exhibited by the axis
cylinders of the nerves. It is beyond the province of this book to

CHAP. X.] THE NERVOUS TISSUES. 421

describe all the various forms which nerve-cells present ; and mention
will merely be made of certain observations which refer to the most
marked and readily investigated of these structures.

The processes which are given off by many nerve-cells, as by the
nerve-cells of the grey matter of the spinal cord, are numerous, and
such cells are often spoken of as multipolar. These processes are
extremely fragile, but under favourable circumstances they may be
observed to give off a number of fine branches. In addition, it has
been maintained (Deiters) that one process which is usually distin-
guished from the rest by its much greater thickness and length
becomes continuous with the so-called axis cylinder of a medullated
nerve-fibre ; such a process would, on this view, place the nerve-cell
in direct communication with a nerve-fibre.

The other finely ramifying processes anastomose with similar
processes from other nerve-cells, giving rise to a reticulum from which
probably arise the axis cylinders of other nerve-fibres. Such a fine
reticulum can readily be seen in the grey matter, though it is some-
times difficult to establish which parts of it are purely nervous and
which belong to the connective tissue. Nerve-cells may o-r may not
have a sheath or investment.

We are acquainted with very few facts relating to the micro-
chemistry of the nerve-cells ; they are doubtless in the main proto-
plasmic in composition, and are therefore specially rich in proteid
substances. From the analysis of the grey matter as compared with
the white, we conclude that the nerve-cells are comparatively poor in
the complex phosphorized constituents, and in other bodies, such for
instance as cholesterin, which are found in large quantities in nervous
organs as a whole. From the abundant supply of blood to the grey
matter as compared with the white we may assume that respiratory
exchanges go on much more actively in nerve-cells than in nerve-
fibres, a conclusion strongly borne out by the previously mentioned
discovery of haemoglobin in the nerve- cells of the ganglia of Aphro-
dite aculeata*

Nerve-fibres,

We may conveniently divide nerve-fibres into the two classes of
(1) medullated, (2) non-medullated nerve-fibres ; the former are much
the more abundant.

1. Medullated nerve-fibres. When examined in its yet living
condition the medullated nerve-iibre presents the appearance of a
perfectly pellucid homogeneous structure which might at first be
thought to be a tube with transparent walls, containing a transparent
liquid ; a careful examination of all facts causes one, however, to
reject this view without hesitation.

At death the nerve-fibre undergoes changes in its physical consti-
tution, and it then can be shewn to present (1) a highly transparent
membranous envelope, termed the neurilemma, in which, or beneath
which, are oval flattened nuclei, (2) a central structure, the axis cylin-

422 NERVE-FIBRES. [BOOK I.

der, and (3) between the neurilemma and the axis cylinder a white,
highly refracting substance, known as the medullary sheath or white
siibstance of Schwann. At intervals, the white substance is inter-
rupted (nodes of Ranvier).

2. Non-medullated nerve-fibres. These differ from the medul-
lated variety in the absence of the white substance of Schwann.
They consist of an axis cylinder sheathed in a nucleated neurilemma.

The neurilemma may be absent from both medullated and non-
medullated nerve-fibres.

The neuri- The very transparent and thin membrane which

forms the wall of the nerve-fibre appears to possess
characters which closely resemble if they are not identical with those
of the sarcolemma. When a medullated nerve-fibre enters a muscular
fibre, the neurilemma loses itself upon, and becomes continuous with,
the sarcolemma (Kiihne). By prolonged boiling both neurilemma and
sarcolemma yield gelatin.

The axis This structure, which under a high power of the

cylinder. microscope presents the appearance of a cylindrical

band, exhibiting marks of fibrillation, is certainly of solid consistence
during life, and is composed of a mixture of proteid with complex
fat-like bodies. It is partly soluble in a weak aqueous solution
of hydrochloric acid (1 to 1000), and in a 10 per cent, solution
of common salt. It is not collagenous. It reduces gold solutions
very readily in the presence of light ; it is stained by ammoniacal
solutions of carmine, which leave the white substance of Schwann
unstained ; this action of carmine is probably dependent upon changes
which occur in the axis cylinder at death.

Chromic acid, potassium bichromate, ammonium monochromate
and certain other reagents, harden the axis cylinder and render it
more easily seen. Perosmic acid, though hardening it, does not stain
it black.

The white That the m-edullated nerve-fibre is not homogeneous

substance of while it is in a physiological condition, i.e. that a distinc-
tion between the axis cylinder and medullary sheath
exists, may be proved by various considerations, for which the reader
is referred to works on histology. The white substance of Schwann
appears during life to have a semi-liquid consistence ; from optical
considerations it would seem to contain suspended solid bodies. At
death it undergoes a kind of coagulation. The white substance of
Schwann instantly reduces solutions of perosmic acid and becomes
black from the presence of metallic osmium. When fresh, the white
substance can be squeezed out of the nerve-fibres, .and is found to be
insoluble in water in which it swells; it is partially soluble in alcohol.
The white substance of Schwann is doubtless specially rich in the
complex phosphorized fats, in the cerebrin group of bodies, and in the
cholesterin, which will be described as the chief constituents of the
nervous matter.

CHAP. X.] THE NERVOUS TISSUES. 423

SECT. 2. THE PROTEIDS FOUND IN THE NERVOUS TISSUES.

More than one-half of the solids contained in the grey matter, and
about one-fourth of the solids of the white matter of the nerve-centres,
consist of proteid substances, and yet our knowledge of these is but
scanty.

Amongst these proteid bodies are to be mentioned, (1) a proteid
substance which is soluble in water and is coagulated at 75° C. ; this
probably is derived from the grey matter ; (2) a globulin-like body
which is dissolved by a 10 per cent, solution of sodium chloride and is
precipitated from it when the same salt is added to saturation ; and
(3) an alkaline albuminate, which remains in solution when a 10 per
cent, salt solution of brain is boiled; in the solution filtered from
coagulated proteids, acetic acid produces an abundant precipitate1.

SECT. 3. NEUROKERATIN AND NUCLEIN.

NeuroJceratin.

If medullated nerve -fibres are treated with boiling alcohol and
ether, so as to extract the fatty matters of the medullary sheath, there
is left in its place an irregular framework which is highly refractile,
and which is scarcely affected by digestion with trypsin or pepsin.
This refractory substance swells when placed in concentrated sulphuric
acid and in solution of caustic potash, but only dissolves in these
liquids when boiled.

The substance resembles, indeed, the horny matter of epidermis in
its power of resisting powerful chemical agents, and has been called
by Kiihne and Ewald2, who have studied its properties, Neuro-
keratin.

The substance is found not only in medullated nerve-fibres, but in
the grey matter of the nerve-centres and in the retina ; it appears not
to be present in non-medullated nerve-fibres.

Ox's brain is washed in water, finely divided, digested for a

Mode of se- jong time in cold alcohol, again pounded, pressed, treated

rokeritf/6 with alcoho1- Then full7 extracted with ether ; dried in the air,

and powdered. The dry powder is shaken through hair

sieves and boiled with alcohol, until this liquid dissolves no more cerebrin.

The residue is boiled with water, pressed and digested with pepsin and the

insoluble residue washed ; it is then digested for 24 hours in a weak

solution of trypsin containing salicylic acid, and afterwards it is digested at

1 The most recent observations on the proteids of the brain are contained in a paper
by Petrowsky entitled " Zusammensetzung der grauen und der weissen Substanz des
Gehirnes." Pfl tiger's Archiv, Vol. vn. p. 367.

2 A. Ewald und W. Kiihne, " Ueber einen neuen Bestandtheil des Nervensystems"
(Neurokeratin). Verhand. d. naturh. med. Vereins zu Heidelberg, Vol. i. Heft 5.

424 NEUROKERATIN. NUCLEIN. [BOOK I.

40° C., for six hours, in a similar trypsin solution which has been rendered
alkaline. The residue is washed with cold, and afterwards with hot,
solution of sodium carbonate, and then extracted with a J p. c. solution of
caustic soda. The extracted matter is treated with a little acetic acid,
with the object of removing adhering alkali, and is then washed with
alcohol and ether. The residue presents the appearance of a yellowish
powder which amounts to from 15 to 20 p. c. of the dried residue left
after the brain has been fully extracted with alcohol and ether.

Pro erties Neurokeratin resembles in its general behaviour the

keratin of the horny tissues ; it differs from that
substance, however, in being less easily soluble in boiling solutions of
caustic potash.

When boiled with, dilute sulphuric acid for some hours neuro-
keratin does not, like horn, entirely dissolve. In the solution, both
tyrosin and leucin are found ; the former being in larger and the latter
in smaller quantities than when proteids are similarly treated.

Neurokeratin emits, when ignited, the odour of burning horn ; it
melts and then burns with a luminous flame. The body contains
nitrogen and 2'93 per cent, of sulphur, and leaves T6 per cent.
of ash.

Nuclein.

. When describing the constituents of pus, the propriety of admitting
the existence of a definite chemical individual termed nuclein was
discussed, and the conclusion arrived at, that under that term bodies
of the most varied composition had been classed, the common proper-
ties of which consisted in resisting the action of the digestive ferments
whilst they were soluble in weak solutions of caustic soda. By
following processes essentially similar to those by which the alleged
nuclein has been separated from pus, v. Jaksch1 thinks he has
discovered nuclein in human brain.

His analyses do not agree with any of the analyses of nuclein obtained
from other sources (see p. 242). As v. Jaksch alleges that his body
possessed the properties of Miescher's nuclein obtained from the milt
of salmon, we quote the ultimate analyses of Miescher's and v. Jaksch's
substances.

ANALYSES OF NUCLEIN,

From spermatozoa of salmon. From human brain.

Miescher. v. Jaksch.

(1) (2)

C. 3611 50-6 50-5

H. 5-15 7'4 7-8

N. 13-09 13-21 13-15

P. 9-59 171 208

1 v. Jakscb, "Ueber das Yorkommen von Nuclein im Menschengeiiim." Pfliiger's
Arehiv, Vol. xin. p. 469.

CHAP. X.] THE NERVOUS TISSUES. 425

Mr Geoghegan1 has estimated the amount of the hypothetical
nuclein in brain at 1*4 grms. per 1000 grms. of brain-substance.

SECT. 4. THE PHOSPHORIZED CONSTITUENTS FOUND IN NERVOUS

TISSUES.

There is no subject in Physiological Chemistry concerning which
it is more difficult to give a statement, which would be accepted as
correct by those who have devoted their attention to it, than the
chemistry of the complex phosphorized fats which exist in the nervous
tissue. In the following pages an attempt will be made to give as
impartial an account as possible of the present co-ndition of a subject
which is eminently in a transition stage.

PKOTAGON.

Discovery *n ^e year 18^5, ^r Oscar Liebreich published an

by Liebreich. important paper2 upon a new proximate principle which
he had separated from the brain. Unlike the numerous
bodies, possessed of ill-defined properties, which had,, by different
writers, received the names of cerebrin, cerebric acid, lecithine, or
phosphorized fats, this new body could be extracted by an easy process
in a state of purity, and to it, probably as indicating it as the first
definitively ascertained specific constituent of brain, Liebreich ascribed
the name of Protagon (Trpurayos, leading in advance).

Mode of ^"he substance was obtained by the following process.

preparation. An animal was bled to death from the carotid, and a
stream of water was passed through the vessels of the
head, so as to wash the blood out. The brain, freed from its mem-
branes, was then pounded in a mortar and shaken in a flask with
ether and water at 0° C. It was allowed to stand at a temperature of
0°, until the ether had separated, and the treatment with ether again
and again repeated.

The brain-matter having been separated by filtration from ether
and water, was digested with 85 per cent, alcohol, at a temperature of
45° C. The fluid was filtered hot and allowed to cool at 0°C. A
flocculent precipitate then separated, which was collected on a filter
and treated with cold ether, until it ceased to dissolve cholesterin.
The insoluble mass was dried in vacuo, and dissolved in spirit at 45° C.
The alcoholic solution was filtered, and allowed to cool very gradually,
when protagon separated in the form of microscopical needles, differing
a little in arrangement and form according to the concentration of the

1 Edward G. Geoghegan (aus Dublin). " Ueber die anorganischen Gehirnsalze nebst
einer Bestimmung des Nucleins im Gehirn." Zeitschrift f. phys.Chem, Vol. i. p. 330.

2 Oscar Liebreich, "Ueber die chemische Beschaffenheit der Gehirnsubstanz."
Annalen der Chemie und Pharmacie, Bd. cxxsrv. S. 29 — 44.

426 PROTAGOX. [BOOK i.

solution. The substance thus obtained could be recrystallized re-
peatedly. As a result of his analyses Liebreich ascribed to protagon
the formula C^H^N.OJP.

Properties Protagon is soluble with difficulty in cold, but more

of Protagon, easily in warm alcohol and ether. At higher tempera-
according to tures than 55° C., alcohol appears to decompose protagon.
In water protagon swells and presents the appearance of
an opaque jelly, ultimately dissolving so as to form an opaque
solution. Liebreich found that protagon was soluble in glacial acetic
acid, which deposited it again in a crystalline form, when subjected to
the action of cold.

When boiled with a solution of barium hydrate protagon is
decomposed, yielding glycerin-phosphoric acid, fatty acids of which
he isolated stearic acid in a state of purity, and a base to which he
gave the name of neurine, and to the platinum compound of which he
ascribed the formula C5H14NCl3Pt. This base was afterwards shewn
to be identical with the base which Strecker had separated from bile
and termed choline.

Although the absolute accuracy of a large number of Liebreich's
facts has been placed beyond question, the cardinal fact itself — that
protagon is a definite phosphorized principle contained in nervous
matter — had, until lately, come to be universally denied.

Hypothesis -By Diaconow, Hoppe-Seyler, and Thudichum it is

that protagon denied that any such definite substance exists, and
is a mixture Liebreich's protagon is held to be a mechanical admix-
ture of a ph°sPnorized bodJ termed lecithin, CJHJNPO,,
with a nitrogenous, non-phosphorized, body termed
cerebrin. The presence of phosphorus in protagon is said to be due
to contamination with lecithin, and in support of this view it is
alleged that by extracting protagon with ether, the substance loses more
and more phosphorus. According to Diaconow and Hoppe-Seyler's
admission, the phosphorus does adhere most obstinately and cannot be
entirely got rid of, though Dr Thudichum thinks he has, by mere
extraction with ether, obtained cerebrin (or cerebrines) quite free from
phosphorus.

According to Diaconow1 by repeated extractions with ether, the P
contained in protagon may be made to sink to 1 per cent., whereas,
according to Liebreich's formula, it should contain 1*5 per cent.
Actually, in the three determinations which he made, Liebreich
obtained 11, I'l, and 1*5, as the amount of phosphorus in 100 parts,
but, unfortunately, he seems to have concluded that the highest number
was correct and made it the basis of his calculation.

In the year 1877, the Author, assisted by Mr Leopold Larmuth,
Platt Physiological Scholar, commenced in the Physiological Labora-

1 Diaconow, "Das Lecithin im Gehirn." Centralblatt filr die medicin. Wissen-
schaften (8 Februar 1868). Nr. 7, Pag. 97.

CHAP. X.] THE NERVOUS TISSUES. 427

tory of Owens College, a series of experiments intended to determine
whether Liebreich's protagon existed or not. This preliminary inves-
tigation shewed that by Liebreich's process there is always obtained
a body having the physical properties of protagon, and containing
phosphorus in a proportion sufficiently near to that indicated by him;
it was found that the amount of phosphorus in specimens of protagon
which had been crystallized from alcohol four or five times, was not
smaller than that present in protagon which had only once been
crystallized, though a thorough treatment with ether preceded each
recrystallization.

These first experiments, so far as they went, were perfectly satis-
factory. It appeared, however, quite essential, before forming a
definite opinion, to extend them very considerably, and especially to
prove the definite nature of 'protagon by a large number of analyses,
indicating not merely the amount of phosphorus, but also that of the
other elements present in it.

This investigation was subsequently carried on by the Author in
conjunction with Dr E. Blank enhorn, with the result of proving to
their entire satisfaction that protagon is a definite chemical body \

Perfectly fresh ox's brains are freed from blood and
of BiaiSen-88 ^rom adneiing membranes as completely as possible,
horn and the and are then digested for many hours (18 to 24) in
Author for 85 per cent, alcohol in a large incubator kept constantly
preparing at 450 Q The guid ig filtered wnilst hotj and the

insoluble matter is again treated with fresh quantities
of spirit, the proceeding being repeated four or five times, as long,
indeed, as the filtrate when cooled to 0° deposits a fair quantity of
white flocculent precipitate. This precipitate is collected on a filter,
and being then transferred to a stoppered bottle is thoroughly and
repeatedly agitated with ether, in order to dissolve cholesterin and
other bodies soluble in ether2. The ether having been removed,
first by decantation and then by filtration, the substance left undis-
solved by it is first of all dried between sheets of filtering paper
in air, and afterwards over sulphuric acid or phosphorus pentoxide.

1 Gamgee und Blankenhorn, "Ueber Protagon," Zeitschriftf. pliysiol. Chemie, Vol.
in. (1879) p. 260. "On Protagon," Journal Physiology, Vol. n. (1879) p. 113. _

2 At first we commenced by repeating exactly the process of Liebreich in all its
details ; one of the steps of that process we had found fraught with peculiar difficulty,
and we soon ascertained that it could be dispensed with without prejudicially affecting
the success of the operations. The step to which we refer consists in agitating the
freshly pounded brain repeatedly with water and ether at 0° C., so long as the ether
dissolves considerable quantities of substance, then filtering and placing the insoluble
matter in 85 per cent, alcohol at 45° C. When pounded brain is so treated with water
and ether it swells up and the separation of the ether is most incomplete. The process
of filtration is one which is attended with great difficulty, even when carried out
in the only way in which we found it possible, viz., in the woven bags sold for household
purposes, for straining jellies, &c. It was, however, apparent that however prolonged
the ether washing, it never succeeded in freeing the brain from cholesterin and other
matters soluble in ether, and that the removal of these bodies from protagon was most
readily effected at a later stage of the operations.

428

PROTAGON.

[BOOK i.

The resulting mass, which has a snow-white colour, is reduced to
powder, moistened with a little water, and digested for many hours
with alcohol heated to 45° C. From the filtered liquid, if this be
allowed to cool very gradually, the protagon separates in the form
of microscopic crystals, mostly arranged in rosettes, the appearance
and arrangement of which differ somewhat, as Liebreich very exactly
pointed out, according to the degree of concentration of the solution.
The once crystallized protagon thus obtained is collected on a filter,
washed with ether, and dried first of all in air and ultimately over
P2O5. It is then recrystallized as many times as required, the process
always commencing by pulverizing and thoroughly shaking with cold
ether.

Results of With the object of proving the definite nature of

ultimate protagon, the Author and Dr Blankenhorn subjected

analyses of the body to repeated recrystallization, subjecting the

product of the successive operations to ultimate ana-

lysis. The following are the results of these analyses : —

C
H
N
P
O

Protagon once
recrystallized
(dog).

Protagon twice
recrystallized
(ox).

No. 1. No. 2.
66-46 66-58
10-96 10-72
2-3 2-6

Protagon twice Thrice re- I
recrystallized crystallized
(ox). (ox).

'our times
recrys-
stallized
(horse).
No. 7.

66-26
10-48

No. 5.

66-3
10-52

No. 6.

66-6
11-06

No. 3.

66-34
10-56
2-40

No. 4.

66-35

1078

No. 8.

66-30
10-467
2-29

1-094 1-107 1-032 1-081 1*027

From the above numbers we have deduced for protagon the

empirical formula C160H308N5P035.

Calculated.

1920

308
70
31

560

2889

10-66
2-42
1-07

19-40

Mean found.

66-39

10-69

2-39

1*068

100-00

From the results of these analyses it appears to the Author
that the existence of protagon as a definite chemical individual is
well-nigh proved.

stability of It has been alleged by Diaconow and Hoppe-Seyler

protagon. ^hat by prolonged treatment with ether and alcohol

the whole of the phosphorus of protagon may be removed, and these
authors regard protagon as a mixture of a nitrogenous body (cerebrin)
with a phosphorized substance (lecithin). The most careful investi-
gation of the matter by the Author has led to entirely opposite con-

CHAP. X.] THE NERVOUS TISSUES. 429

elusions. Pure protagon is remarkably rebellious to the action of even
boiling alcohol, though that action be continued for hours: and the most
persistent attempts to separate lecithin from it have failed. At the
same time there can be no doubt that protagon when treated with
certain reagents which decompose it, especially when digested with
alkalies, yields, amongst other bodies, certain of the most characteristic
of the products of decomposition of lecithin. But to conclude from
the presence of these, the presence of lecithin, is obviously unphiloso-
phical.

There can be no doubt that protagon is accompanied in the brain
by large quantities of a body or bodies which may provisionally be
conveniently classed under the term of cerebrin, and likewise by
smaller quantities of other phosphorized bodies, containing a per-
centage of phosphorus very close to that found in lecithin, yielding
the same products of decomposition, and the separation of which
presents extraordinary difficulties. Yet, the Author is convinced that
unquestionably the only well characterized phosphorized proximate
principle, which can with our present methods be separated with
certainty and whose existence will be confirmed by future researches,
is Protagon.

1. Action of Alkalies. Liebreich discovered that

re^earchlfon wlien Protagon is boi!ed for 24 hours with. a satu~
tfce products rated solution of barium hydrate, the solution con-
ofdecompo- tains the barium salt of glycerin-phosphoric acid,
sition of pro- (C3H5) (OH)2 . 0 . PO (OH)2 ; a base called N eurine, which
was afterwards shewn to be identical with a base pre-
viously obtained by Strecker from ox bile and termed by him Choline
(CSH15N02) ; and barium salts of several fatty acids, especially of
stearic acid.

There can be no doubt that there is formed, in addition, a certain
quantity of a cerebrin-like body or a mixture of such bodies, i.e. a
substance is obtained which is not soluble in ether, which dissolves
in boiling alcohol and is deposited from it in a nodular form on
cooling : which contains nitrogen but no phosphorus. The experiments
of the Author have shewn him that the quantity of this body which
is formed is very much smaller than would be the case on the
hypothesis that protagon is a mixture or even a conjugated compound
of lecithin and cerebrin.

2. Action of acids. When protagon is boiled with hydrochloric
acid in the absence of light, a yellowish liquid is obtained, which
deposits flocculi of a body free from phosphorus ; and which becomes
coloured when exposed to light. In this process there is set free a
laevogyrous, non-fermentable sugar.

The chief products of decomposition of protagon will be de-
scribed at length under the head of lecithin and cerebrin; under the
latter the cerebrin body which accompanies protagon will be dis-
cussed.

430 LECITHIN. [BOOK

LECITHIN.

Besides the nervous tissues, there are others in which organic
phosphorized ingredients occur in considerable proportions ; these are
found in large quantities in the ovum, in spermatozoa, etc.

The first satisfactory study of the phosphorized organic bodies
was made by Gobley *, who described under the name of Lecithine
a viscous proximate principle which he had separated from the eggs
of the carp. This body was entirely soluble in ether, soluble with
difficulty in cold, but readily in hot alcohol, from which it was
deposited on cooling. Gobley2 found that this body, when ignited,
yielded an ash possessed of a strong acid reaction, owing to the
presence of phosphoric acid. He further shewed that when decomposed
with acids or alkalies, his lecithin yielded glycerin-phosphoric acid
and fatty acids, amongst which he cited oleic and margaric. Besides
lecithin, Gobley separated, by a process wjiich would certainly cause
decomposition of any complex proximate principle, a body which
he termed Cerebrin, which contained Q'43 p.c. of phosphorus.

In researches on the brain which were anterior to his most mature
investigations on lecithin from eggs, Gobley arrived at the conclusion
that the phosphorized matter of the brain resembles, if it be not iden-
tical with, that obtained from eggs, and this view of Gobley's is the
one which has commended itself almost universally to physiological
chemists.

After the publication of Liebreich's memoir on Protagon attention
was again directed to the phosphorized proximate principles of the
body, it being doubtless surmised that the well-defined protagon
would be discovered where earlier observers had found less sharply
characterized bodies. This surmise was, however, soon disproved.

In a paper published in Hoppe-Seyler's Untersuchungen, by one
of his own pupils, Parke 3, " On the Chemical Composition of the
Yolk of Egg," in which the amount of protagon present was calculated
on Liebreich's data from the amount of phosphorus found in the
alcoholic extract of the yolk of egg, the observation was made that,
by calculation, more protagon was found than corresponded to the
whole weight of the alcohol extract.

In a paper immediately succeeding that of Parke's, Hoppe-
Seyler 4 clearly expressed his conviction that the yolk of egg contains
no protagon but lecithin, this being the name which Gobley had
given to the chief phosphorized constituent of the yolk. He further
stated, that experiments made in his laboratory by Herr Jiidell had

1 Gobley, Journal de Chimie etPharmacie, Vol. xvii. (1850) p. 401 : Vol. xvm. (1850)
107.

2 Gobley, Journ. de Pharm. et Chimie, Vol. xi. (1847) p. 409, and xii. (1847) 1.

3 Parke, "Ueber die chemisette Constitution des Eidotters," Med.-chem. Unter-
suchungen, Heft 2, p. 213.

4 Hoppe-Seyler, " Ueber das Vitellin, Icbthin und ihre Beziehung zu den Eiweiss-
Btoffen." Hed.-chem. Untersuchungen, Heft 2, p. 215.

CHAP. X.] THE NERVOUS TISSUES. 481

shewn that the ether extract of red blood-corpuscles, besides choles-
terin, contained a body, the amount of phosphorus in which corresponded
to S'25 p.c. of P2O5 (that is to say, containing 3'6 p.c. of phosphorus),
and which therefore could not be protagon. Hoppe-Seyler had by
this time obviously commenced to entertain doubts as to the existence
of protagon, though he did not commit himself to a denial of its
presence in the brain ; indeed, by implication he rather admitted his
belief in its existence1.

Another of Professor Hoppe-Seyler's pupils, Dr Diaconow 2, now
continued the investigation.

In a paper on the bodies containing phosphorus which are present
in the hen's egg and in the ova of the sturgeon, he came to the
following conclusions :

1. Gobley's lecithin and the phosphorized bodies which are
obtained from Ichthin and Vitellin yield on boiling the same products
of decomposition as protagon.

2. They contain about twice as much phosphorus as protagon,
so that they are either altogether distinct from protagon, or they
consist of a mixture of protagon with a second phosphorized body.

3. In any case protagon is not the only phosphorized proximate
principle of the body.

4. The discovery of phosphoric acid in alcoholic or ethereal extracts
of different animal organs, does not entitle us to conclude that protagon
is present.

5. The quantity of phosphoric acid found in an ethereal extract,
freed from cholesterin and fats, affords no estimate of the quantity of
protagon.

A short time after the appearance of the preceding, Diaconow
published a second paper3 in which he described the properties of
the phosphorized constituent of yolk of eggs, to which he correctly
ascribed the name which Gobley had given to it, giving the results of
analyses, and naming the chief products of its decomposition.

According to Diaconow' s description of lecithin from
Diaconow's eggs this body has the following characters :

°f Pure lecittlin presents the appearance of a yellowish-

white, waxy, very hygroscopic solid, which when in
thin layers shines with a silky lustre ; it is soluble in ether and
alcohol, it swells in water, and on shaking it in, or stirring it with,
water it forms a starch-like solution which filters with difficulty.
When ignited it burns away, leaving as only residue phosphoric
anhydride4. The chemical formula of the body calculated from its

1 "Ob aber neben Protagon auch Lecithin sich in der Hirnmasse findet habe ich
nich untersucht." Hoppe-Seyler, Ibid. p. 220.

2 Diaconow, " Ueber die phospborhaltigen Korper der Hiihner- nnd Htoreier" (Vor-
laufige Mittheilung). Hoppe-Seyler, Med.-chem. Untersuchungen, Heft 2, p. 221.

3 Diaconow, "Ueber die chemische Constitution des Lecithin," Centralblatt fur die
med. Wissenschaften, 1868, No. 1, p. 2.

4 This is a curious mistake which has been repeated by all physiological chemists ;
the residue is one of metaphosphoric acid.

432 LECITHIN. [BOOK i.

ultimate analysis (C = 64*27 p.c. ; II = 1 1/4 ; N = 1/8 ; P = 3'8) is
C4fE90NP09 + H2O. When decomposed it yields glycerin-phosphoric
acid, stearic acid and neurine — the very same products which Liebreich
had obtained from protagon.

Compounds Lecithin forms compounds both with bases and

of lecithin. acids, as for example with potassium and hydrochloric

acid, and the latter forms a platinum compound

(Strecker) ; in the case of distearyl-lecithin (see ' constitution of

lecithin ') this compound would have the formula

A similar compound with cadmium chloride exists. The platinum
compound is soluble in ether, but it may be precipitated from the
ethereal solution by an excess of alcohol.

Diaconow's Diaconow had in the meantime directed his investi-

assertion of gation to the brain, and one month after the publication
the presence of his first paper there appeared a second1 which has
of lecithin in exerted a most weighty influence, causing physiological
chemists to come to the conclusion that Liebreich's
protagon does not exist as a definite proximate principle, but that
it consists of a mixture of lecithin with a body free from phosphorus,
cerebrin.

Diaconow's Brain freed from its membranes and from blood, is

method of finely divided and repeatedly extracted with ether ;

separating the residual mass is digested with absolute alcohol
lecithin from at 40° C., and the alcoholic solution thus obtained is

ain' cooled to 0° ; the precipitate which separates is filtered

off, and washed with a little cold absolute alcohol and afterwards
once again with ether. A portion of the substance dissolves in ether,
whilst another, protagon, remains as a residue. The latter is
repeatedly extracted with ether at ordinary temperatures, and the
collected ethereal extracts are subjected to distillation ; the residue
is dried at 40°, dissolved in a little absolute alcohol, and the alcoholic
solution is cooled. There separates a white substance having the
composition and properties of lecithin. The substance is amorphous,
non-pulverizable, hygroscopic, swells in water and when shaken with
it forms an emulsion. When burned it leaves as a residue phosphoric
anhydride (?). When decomposed with lime- or baryta-water it
yields in addition to neurine, barium stearate and glycerin-phos-
phate.

Analyses of the body separated in this way by Diaconow furnished
the following results.

1 Diaconow, "Das Lecithin im Gehirn.'' Centralblatt filr die medicinischen Wissen-
schaften, No. 7, 8th Feb. 1868, p. 97.

CHAP. X.] THE NERVOUS TISSUES. 433

1. 0-0678 substance gave 0*0083 Mg2P207 = 7'83 # P205.

2. 0-0985 „ „ 0-0123 „ =7'98J „

3. 01833 „ „ 0024 Pt = TS5gN.

The formula of lecithin, C^H^NPO^ demands 8'378 g of P205 and
117 1 of N.

Upon these facts, and these alone, so far as the author is aware,
all subsequent writers have based their belief in the presence of
lecithin in the brain, adopting the views of Diaconow and Hoppe-
JSeyler that protagon is a mixture of lecithin with cerebrin *.

The Author's ^ *s unquestionably true that the precipitate

criticism of which separates from an alcoholic solution of brain,
the observa- contains, besides protagon, cholesterin, and a body for
which we may retain the name of cerebrin, small
quantities of bodies soluble in ether which have a
much higher percentage of phosphorus than protagon, and which
possess the general smeary characters of lecithin. But these bodies
are present in very small quantities, and are readily removed by ether
washing. Protagon which has been several times recrystallized yields
no such body as lecithin to ether, in which liquid it is at ordinary
temperatures practically insoluble.

From his own observations then he would conclude that whilst
it is true that the brain yields to alcohol other phosphorized bodies
than protagon, the latter is much the most abundant of the phos-
phorized products, and by no action of ether can it be split up into
lecithin and a non-phosphorized cerebrin, it is possible and indeed
probable that amongst the phosphorized principles, lecithin is to be
reckoned. No sufficient proof of its identity has however yet been
furnished. It is indeed apparent to the author from his own work,
no less than from a careful study of the researches of Thudichum,
that the phosphorized ingredients are numerous.

Description of some of the products of decomposition of lecithin
and protagon.

Whichever the view adopted, it appears that certain of the pro-
ducts of decomposition of lecithin and protagon are the same. The
chief of these are glycerin-phosphoric acid, neurine or choline, and
fatty acids; the two former of these bodies will now be described.

Glycerin-phosphoric acid (CSH9P06).

When distearyl-lecithin is decomposed by boiling with alkaline
solutions, it combines with the elements of water, with the formation

1 The author has attempted to separate lecithin from brains by Diaconow's method
and has failed entirely.

a. 28

4.34 GLYCERIN-PHOSPHORIC ACID. [BOOK I;

of stearic acid, glycerin-phosphoric acid and choline, as shewn by the
following equation : —

CJI^NPO, + 3H20 = 2 (CJ^OJ + C3H9P06 + C^NO,.

Lecithin. Water. Stearic acid. Glycerin- Neuriiie.

phosphoric
acid.

This acid may be prepared by the decomposition of
tion"? giy- lecithin by means of caustic baryta, or synthetically in the
cerin-phos ~ following manner :—

phoric acid. Phosphoric anhydride is added in equivalent propor-

tions to glycerin which is kept cool by a freezing mixture.
Much heat is evolved and the new acid is formed. According to the
author's experiments the yield of acid is extremely small. The solution is
diluted with water, neutralized with barium carbonate, filtered from the
large quantity of barium phosphate which is formed, and the filtrate is
exactly neutralized with dilute sulphuric acid. The filtrate from deposited
barium sulphate is concentrated in vacua at a low temperature; in this
manner a watery solution of the acid is obtained. This solution cannot
be concentrated beyond a certain point, as it decomposes.

Pro erties Glycerin-phosphoric acid is a syrupy liquid possessed of

and com- both an acid and sweet taste. It forms salts which are for

pounds. the most part soluble in water, but insoluble in alcohol ; the

lead compound is an exception to the former statement.

The barium compound has the composition CaHyBaPOg. Tim die-hum
and Kingzett l describe a hydrate having the composition C3H7BaP06H2O.

The normal calcium salt has the composition C3H7CaP06H20 ; it
is less soluble in hot than in cold water, so that a solution appears to
coagulate when boiled, like a solution of albumin. An acid salt has
been described having the formula C3H7CaPO , C3H9P06.

A soluble zinc salt (C3H7ZnO6) and an insoluble lead salt (C3H7Pb06)
also exist ; the latter is prepared by adding a solution of acetate of lead
to the barium compound.

Constitu- ^. stu(tying the appended graphic formulae of

tion> ' glycerin, phosphoric acid, and glycerin-phosphoric acid

the reader will apprehend the view which is held of the
constitution of the last-named body.

(OH (OH

(1) C3H5^OH (2) PCKOH

(OH (OH

C3H5(OH)3 PO(OH)3

Glycerin. Orthophosphoric acid.

1 Thudichum and Kingzett, " On glycero-phosphoric oxide and its salts, as obtained
from the phosphorized constituents of the brain." Journ. Cliem. Soc., July, 1876.

CHAP. X.] THE NEKVOUS TISSUES. 43j>

ron

/ON n TT <OH

(*} ^3^5 } (OH

(°-po{oS

C3H6(OH)2.O.PO(OH),

Glycerin-phosphoric acid.
Neurine1 (Choline). C5H15N02.

Pre oration k°dy, which was first obtained by Strecker2

from bile (and termed choline), and afterwards inde-
pendently discovered as a product of decomposition of protagon by
Liebreich, and of lecithin by Diaconow, may be obtained by decomposing
either of the latter bodies by boiling them for at least an hour with
baryta water. The liquid is then filtered, treated with C02, then boiled,
filtered and concentrated at a gentle heat. A syrupy residue is
obtained which is extracted with absolute alcohol; the solution is
filtered, hydrochloric acid is added, so as to induce a slight acid re-
action, and then a solution of platinum tetrachloride is added ; a yellow
precipitate (composed of a double salt of neurine and platinum tetra-
chloride) falls, which is washed in alcohol, dissolved in water and
decomposed by H2S. The filtrate from the precipitate of platinum
•sulphide is concentrated in the water bath. In this way hydrochlorate
of neurine is obtained and from this the base is set free by treatment
with silver oxide.

Pro erties Neurine is a syrupy liquid, soluble in alcohol and

ether and possessing a marked alkaline reaction. It
does not coagulate albumin. Its solutions dissolve fibrin.

Besides the compounds with hydrochloric acid.

Compounds. . - ,. J i •,/

previously reterred to, neurine lorms compounds with

carbonic and sulphuric acids ; its hydrochlorate forms double salts with
platinum tetrachloride and with gold chloride. The following are
the rational formulae of the hydrochlorate and of the platinum and
gold compounds :

(1) N(CH3)3(aH4-OH)Cl.

(2) [N (CH3)3 (C2H4 • OH) Cl]2 PtCl,

(3) N (CH3)3 (C2H4 • OH) 01 + AuCl3.

Products of Amongst numerous and highly interesting decom-

decomposi- positions it may be mentioned that, when heated,
tion- neurine splits up into glycol and trimethylamine :—

TT (OH XQTT

°"H- OH = C2H4 JJjg + N (CH3)S

*•— .--y- - -^ ^-- — ~"v* — " —

Glycol. Trimethylamine.

1 In writing this description of Neurine the author has availed himself very freely of
the excellent account in Hoffmann's Lehrbuch der Zoochemie, page 114 et seq.

2 Strecker, " Ueber das Lecithin." Ann. d. Chem. u. Pharm., Vol. LXXII. p. 77.

28—2

436 NEURINE. LECITHIN. [BOOK I.

Synthesis. Neurine has been produced synthetically in two ways:
1st. By bringing together a concentrated solution of trimethyl-
amine with ethene oxide, thus : —

CH. CH..OH

>0 + N(CH3)3 + HOH =

i:

Ethene oxide. Neurine.

2nd. By heating in a sealed tube a mixture of ethene-chlor-
hydrin and trimethylamine, thus : —

CH2 . OH + N (CH3)3 = CH2 . OH
3H2C1 T^^m*e- (1H2- N (CH3)3C1

Ethene- Chlorhydrin. Neurine Hydrochlorate.

Constitution of Lecithin.

Having now described the properties of lecithin as observed
by Diaconow and Strecker, we have to approach the question of the
constitution of that body.

The lecithin which Diaconow believed to exist in the brain was
supposed by him to yield as a product of decomposition, and as
the only fatty acid, stearic acid, and it may be therefore termed,
for reasons which will be apparent immediately, distearyl-lecithin.
This body and indeed all lecithins, for there are probably many
lecithins, are derived from glycerin-phosphoric acid.

If in glycerin-phosphoric acid two of the atoms of hydroxyl-
hydrogen in the glycerin be replaced by two molecules of stearyl, we
shall obtain an acid to which the name of distearyl-glycerin- phos-
phoric acid was given by Diaconow. This acid would have the
constitution CIH.(C18H88OS)80 . PO(OH)2: it was actually obtained by
Diaconow on shaking an ethereal solution of lecithin with sulphuric
acid, the products of the reaction being this body and neurine-
sulphate.

Now distearyl-glycerin-phosphoric acid may unite itself with
neurine to form lecithin.

The mode of attachment of neurine to distearyl-glycerin-phosphoric
acid cannot be represented with certainty by any formula. The three
following formulae for distearyl-lecithin have been suggested and
represent the different views which have been held : —

(A) Diaconow : x

-N(CH8)3-CaH,OH

1 Centralblatt f. d. med. Wiss., 1868, Nr. 1, s. 3.

CHAP. X.] THE NERVOUS TISSUES. 437

(B) Strecker : l

3 5o

0-C2H4-N(CH3)3OH

(C) Hoppe-Seyler : 2

'n

0-C2H4-OH

Distearyl-lecithin may be looked upon as the type of the lecithins ;
but we can easily conceive of lecithins which only differ from this one
by the radicals of oleic acid or of palmitic acid having taken the
place of stearyl. We should thus have dipalmityl-lecithin, dioleyl-
lecithin or oleyl-palmityl-lecithin formed ; such bodies probably
exist.

Phosphorized Principles other than Protagon and Lecithin.

In a very elaborate research on the phosphorized constituents of
the brain, Thudichum3 has come to the conclusion that he has
separated at least three well characterized bodies or groups of bodies.

The bodies belonging to these groups are distinguished as (1) the
Kephalines, (2) the Myelines, (3) the Lecithins.

Ke haiine Belonging to the first group is a body Kephaline,

which is said to be exceedingly soluble in ether ; this

body is soluble in hot alcohol, but less so than either lecithin or

myeline. To kephaline is ascribed the formula C^H^NPC^. It does

not form definite compounds with platinum or cadmium.

Myelines ^e myelines are far less soluble in ether than

kephaline or the kephalines, and less soluble in alcohol
than the lecithins.

Various myelines have been described by Thudichum including
bodies having the following formulae : — CJ&g-NPOg ; C^H^NPO^;
CUWO,.; Q.BUN.PO,; CMH81NPOS; C^N,PO,

Thudi- The following are the main characters of the phosphorized

chum's sum- principles of the brain as summarized by Dr Thudichum. In

mary of his quoting them the author in no respect expresses his agree-

researches on ment with Dr Thudichum's conclusions.

"The SrouP °f the PhoaPhorize(1 bodies contains the
ciples phosphorus in the form of phosphoric acid, combined proxi-

mately with glycerine, so that by chemolysis they all yield
glycero-phosphoric acid, but they differ in the manner in which they
contain the nitrogen, and the acid radicles which constitute the great bulk

1 Ann. Chem. Pharm., 18G8, Bd. CXLVIII. s. 77.

2 Physiologische Chem., 1877, 1 Theil. s. 80.

3 Thudichum, " Besearches on the Chemical Constitution of the Brain." Reports of
Medical Officer of the Privy Council and Local Government Board, 1874, p. 113 et seq.

438 THUDICHUM'S RESEARCHES. . [BOOK i.

of their substance, and according to these differences must be divided into
sub-groups. "We thus obtain the sub-groups of the kephalines, myelines, and
lecithines.

" Of these the kephaline sub-group, itself hitherto unknown, includes
members which contain the nitrogen in either one or two forms, one being
either choline or neurine, another hitherto unknown; and they contain
the fatty acid radicles also in forms with which chemistry is at present
unacquainted, and the members of this sub-group further vary in the
amount of oxygen which they contain in a manner so as to be sharply
characterised thereby. This variability of the constituent oxygen may
be transitional, but must not be confounded with that remarkable
reaction of the bodies of this group which I describe as their oxy dis-
ability.

" The myeline sub-group, also new, contains the nitrogen in two forms,
of which one is choline, the other amide in a fatty acid radicle. The
fatty acid radicles vary, and are mostly new forms, some known forms.
The members of this group consequently vary in carbon, hydrogen and
oxygen; little in nitrogen, never in phosphorus. They are not oxy-
disable after the manner of kephaline, though there is an oxy-myeline
after the mariner of oxy-kephaline. They are the least soluble of the
entire group, the least decomposable, and stand the highest temperatures,
being unchanged by fusion at a heat above the boiling point of water.

" The lecithine sub-group, well known from the chemistry of eggs, is
only with difficulty evolved from the brain, on account not only of the
many stages of the processes necessary for their isolation, but also on
account of a prominent feature of its members, namely, their readiness
to decompose when in the anhydrous state. This tendency to apparently
spontaneous lysis into proximate nuclei prevents the inquirer fixing
properties and varieties with the same precision as in the previous
groups; but it furnishes a valuable key to the explanation of many
changes in the sick body, which may arise, or have been proved to
arise, from their decomposition.

" The chemical characteristics of these sub-groups may be summarized
thus : the kephalines possess the tendency to be oxydised, oxydisability ;
the myelines are not easily changed by any agent or influence, and possess
therefore stability; the lecithines easily fall to pieces, they are afflicted
with lability.

"In language more technically chemical: the kephalines have on the
outside of their molecules free affinities for oxygen; this gas they bind
in several ways; when the oxygen combined with a molecule has
attained a certain quantity, the avidity (intensity of affinity) of the
molecule increases to this extent, that it monopolises all available
oxygen to itself until the limits of its oxydisability (at present unknown)
are attained ; until its free affinities are satisfied. Until then the rest of
the molecules, if the supply of oxygen be insufficient to oxydise all to
the same point, are not oxydised. The kephalines, however, are not in
a state of atomic tension, and therefore do not fall to pieces so easily as
the lecithines, but require for lysis the influence continued for some
time, of powerful extraneous affinities in the presence of water and heat.

"The myelines have no apparent free affinities for oxygen; they are
not affected by heat to and above boiling water except to the extent of

CHAP. X.] THE NERVOUS TISSUES. 439

fusion ; their atoms are not in a state of chemical tension, but require for
vibration into permanent decomposing distances the influence of strong
external affinities, water and heat.

" The lecithines, however, are in a state of great atomic tension, and
therefore slight external affinities or dissociating impulses suffice to effect
their decomposition. Such a slight impulse is the attraction of absolute
alcohol for their fatty acid radicles in the absence of external water. The
water given out by the alcohol in becoming ethylic ether serves to enable
the radicle of glycero-phosphoryle to become glycero-phosphoric acid, and
to remain in combination with the choline evolved1."

SECT. 5. NON-PHOSPHORIZED NITROGENOUS BODIES OF UNKNOWN

CONSTITUTION. .

Cerebrin (?) or Cerebrins (?).

Miiller's ^n ^e Jear 1858 Miiller published an account of

cerebrin. a non-phosphorized body which he had obtained from

brain by the following process. The brain was pounded
up with baryta water to the consistence of a thin milk and then
boiled ; the precipitate which formed under these circumstances was
extracted with boiling alcohol: on cooling, the alcoholic solution
deposited an abundant precipitate. This was treated with ether
to separate cholesterin and fats, and then recrystallized from boiling
alcohol. The purified body thus obtained, which was termed cerebrin
by Miiller, possessed the following characters : — it was a loose,
white, very light, powder, destitute of smell and taste, soluble in
boiling alcohol and ether, but insoluble in water, cold alcohol and
ether. Under the microscope the body presented the appearance of
small round balls2. Miiller published analyses of this body (only
two carbon determinations being given) and to it he ascribed the
formula C34H33N06.

The following are the results of the analyses of Miiller's cerebrin :

Calculated. Found.

(1) (2)

C 68-23 68-35 68*56

H 11-04 11-30 11-25

N 4-68 4-69 4'53

0 16-05 15-66 15-66

That a body produced by the prolonged action of a solution of
boiling barium hydrate on so complex an organic mixture as brain
should be a definite proximate principle of the unaltered organ would
appear in the highest degree unlikely ; even more unlikely than
that it should be a definite principle at all. Yet, without any sufficient
proofs, the existence of Miiller's cerebrin has found favour with all
those who have doubted the existence of protagon, and, since the time

1 Thudichum, Op. cit. p. 198.

2 Miiller, "Ueber die chemischen Bestandtheile des Gehirns." Ann. d. Chem.
u. Pharm., Vol. cv. p. 361, 2te Abth.

440 ' CEREBRIXS.' [BOOK i.

when Diaconow and Hoppe-Seyler first promulgated this idea, the
latter body has been generally considered to be a mixture of cerebrin
with lecithin.

The author, in his criticism of the position which had been taken
up by Diaconow and Hoppe-Seyler, tried to shew that those who
had manifested great scepticism in Liebreich's protagon had taken for
granted the existence of a body whose investigation had been infinitely
more incomplete1. The justness of the criticism has been thoroughly
confirmed by the subsequently published researches of Geoghegan2.

Geoghegan's Instead of boiling pounded brain with caustic baryta,

modeofpre- as Muller had done, Geoghegan extracted pounded
paring cere- ^rain with .cold alcohol and ether, then boiled it in
alcohol. The white body which separated on cooling,
and which according to Geoghegan is a mixture of cerebrin, cholesterin
and lecithin, was treated with ether so as to separate cholesterin,
and then boiled with baryta water. The insoluble residue was
dissolved in alcohol and crystallized.

It was analysed with the result of finding that it contained only
one-third of the amount of nitrogen which had been found by Miiller ;
to it the empirical formula Gj3J$f)m is ascribed.

f

Mean of Geoghegan's . Mean of Miiller's
analyses of cerebrin. analyses.

C ................ 6874 68-45

H ................ 1091 11-20

N ................ 1-44 4-50

The author's The author's researches on cerebrin, though far from

researches on complete, were made immediately prior to the publi-
cerebrin. cation of Geoghegan's paper, though they have been

hitherto unpublished. They have led him to the following conclusions :

(1) By the action of ether, however prolonged, or of alcohol, a
phosphorus-free cerebrin cannot be obtained from protagon ; though
by boiling with alcohol for many hours protagon appears to be decom-
posed, so that by separating the substance which falls first on cooling
and subjecting it again to prolonged treatment with boiling alcohol, a
body is obtained which differs somewhat in physical characters from
protagon ; if this body be many times subjected to the""" action of
boiling alcohol and to the above referred-to process of separation,
a substance is obtained containing less phosphorus than protagon
and having a different composition. This body, which was certainly
not absolutely pure, was analysed witli the following results : —

(1) (2)

C. in 100 parts 64'44 6*23
H. „ „ 10-46 10-54
N. „ „ 3-12

(2) By the action of caustic baryta on protagon there is obtained

1 Gamgee and Blankenhorn, Op. cit. Journ. of Phys., p. 121.

2 Geoghegan, " Ueber die Constitution des Cerebrins." Zeitschrift /. pliys. Chemit,
Vol. in. (1879), p. 332.

CHAP. X.] THE NERVOUS TISSUES. 441

a cerebrin-like body, which agrees fairly in so far as the C and II
with Geoghegan's body. The nitrogen has not yet been determined.

C. in 100 parts 68'95

H. „ „ 1132

(3) In addition to protagon, and other phosphorized matters,
there is always extracted from brain by alcohol at 45°, a very con-
siderable quantity of a body, which, in order to distinguish it, the author
provisionally termed pseudo-cerebrin. This body is less soluble in
80 p. c. alcohol at 45° than protagon, so that on subjecting impure
protagon to repeated crystallization from 80 p.c. alcohol there accumu-
lated residues consisting of the cerebrin-like body. The latter is a
white, pulverulent body, very unlike protagon to the naked eye and
separating under the microscope in the form of very large nodular
masses. After repeated recry>stallization from alcohol it was found to
be practically free from phosphorus (containing only 0*08 p. c.).

On analysis this body has given results which are not widely differ-
ent from those of Geoghegan, though they are sufficiently discrepant
to render it certain that the substances analysed by that observer
and himself were not identical. Whilst the author would refrain from
speaking with confidence of the absolute purity of ' pseudo-cerebrin!
he has, however, no hesitation in asserting that Geoghegan's substance,
from the method of preparation, is necessarily a mixture of the so-
called pseudo-cerebrin just referred to with a 'cerebrin' obtained by
the action of barium hydrate on protagon — and therefore much more
impure than the body now provisionally described by the term of
pseudo-cerebrin.

Analyses of ' pseudo-cerebrin' found by the Author to accomnany Protagon.

(1) (2) (3) (4) Mean.

C 6897 68-95 69*01 68'67 68-89

H 11-7 11-17 11-60 12-10 11-87

N 176 1-95 1-64 2-01 1'83

O. 17-41

100-00

From the above analyses the author deduces for ' pseudo-cerebrin'
the empirical formula C^H^NOg.

It would therefore appear to the author that whilst protagon
cannot be separated by the action of solvents into a non-phosphorized
cerebrin and a phosphorized body, yet such non-phosphorized bodies
exist by its side in the brain, and can be obtained from protagon by
the action of caustic baryta.

By the action of concentrated sulphuric acid on
gan's^e-6" cerebrin, this author has obtained a body to which he
searches in ascribes the formula C22H42O5 and believes to be a
the decompo- derivative of cetyl-alcohol, and which he terms Cetylid.
sition of cere- Qn fusion with caustic potash this body evolves a
an. Cetylid. mixture of QJJ^ H and Nj whilat palmitic acid is

formed ; a portion of the N is left in the form of an ammoniacal salt.

442 CHOLESTERIN. [BOOK I.

Thudi- Under the name of cerebrins, Thudichum describes a

chum's re- class of nitrogenous bodies free from phosphorus, which

searches on^ -^Q believes to exist in the brain. Certain of these bodies he
obtained by following substantially Miiller's process ; others
by extracting brains with alcohol at 45° C., and purifying the substance ob-
tained by various solvents. He believes Miiller's cerebrin to be the lowest
representative of a group of nitrogenous prinicples of the brain which
are free from phosphorus, contain nitrogen, and vary in the number
of carbon atoms which they contain, for each nitrogen atom, between
17 and 48. "Whatever may be the ultimate explanation of these differ-
ences of composition must be left for future inquiry. Meanwhile it is
certain that these differences do but slightly affect the external appearance
and bearing towards solvents of these bodies, so that by describing
the general properties of one we describe the general properties of all
members of the group, while differentiating characters and means are most
difficult of discovery and application."

"The cerebrins are all soluble in hot alcohol, particularly in absolute
alcohol, and deposited on cooling; they are very little soluble in cold
absolute alcohol, much less soluble indeed than myeline, which can thus be
separated from the cerebrins. The mixture is dissolved in hot alcohol and
allowed to cool j nearly all cerebrin falls down, much myeline remains in
solution. The deposit is separated from the liquid, and subjected to this
treatment until it is free from phosphorus."

The following are the three chief bodies which Thudichum classes amongst
the cerebrins.

Cerebrin C^H^O, (Mailer).

Phrenosine C^H^NOg.

Kerasine C^ELNOo.

SECT. 6. CHOLESTERIN (C26H440 + H20).

Amongst the most abundant of the constituents of the nervous
tissues, and especially of the white matter, is the beautiful, crystalline,
non-nitrogenous, body cholesterin. This body, which is very freely
soluble in ether, cold or hot, is also freely dissolved by warm alcohol,
which in great part deposits it on cooling; in consequence of its solu-
bility in these two fluids, cholesterin finds its way into both the
ethereal and alcoholic extracts of the nervous tissues2.

Prepara- The tissue from which cholesterin is to be extracted

tion of cho- may be placed in cold alcohol for some days, so as
lesterin from to deprive it of the greater part of its water. The
SP1" hardened substance is then finely divided and digested
in boiling alcohol. The alcoholic solution is filtered

1 Thudichum, " Eesearches on the Chemical Constitution of the Brain." Reports
of the Medical Officer of the Privy Council and Local Government Board, London, 187-1,
pp. 113—247.

2 The author some years ago performed a number of experiments with the object of
determining whether cholesterin preexists in the nervous tissues or is merely one of the
products of the decomposition of more complex bodies. These experiments led him to
the conclusion that cholesterin exists preformed in the brain.

CHAP. X.] THE NERVOUS TISSUES. 443

through a heated funnel and the filtrate is cooled. The deposit,
which consists of cerebrin, protagon, other complex phosphorized
bodies, and cholesterin, is collected on a filter, washed with cold alcohol,
and then, after being pressed between filter paper, is shaken in a
stoppered bottle with ether; the ethereal solution is filtered, the ether
is distilled off, and the residue, consisting of impure cholesterin mixed
with some lecithin, is heated in a water bath for an hour with an
alcoholic solution of caustic potash. The contents of the flask are
then evaporated to dryness on the water bath, and the dry residue
is washed with water and dissolved in a mixture of ether and
alcohol, from which it is allowed to crystallize by the spontaneous
evaporations of the solvents.

Properties Pure cholesterin separates from its solutions in

anhydrous ether or chloroform in the form of needles
containing no water of crystallization; but from alcohol it separates in
the form of rhombic tables.

FIG. 60. CRYSTALS OF CHOLESTEBIN AS IT SEPARATES FROM ALCOHOL OR ETHER
CONTAINING WATER. (Frey.)

Cholesterin is insoluble in water, alkalies and dilute acids; it is
very slightly soluble in cold, but soluble in 9 parts of boiling, alcohol.
It is highly soluble in ether cold and boiling, in chloroform, benzol,
and in solutions of salts of the bile acids.

Dry cholesterin melts at 145°, and distils in vacuo at 360°. Its
solutions exert a left-handed rotation on the plane of polarization. The
specific rotatory power of solutions of cholesterin (a)jD=— 31°'6. Amongst
the most useful reactions for detecting cholesterin are the following : —

1. When treated with concentrated sulphuric acid, and after-
wards a little iodine, a play of colours, of which blue, green, and
red are the most prominent, is produced. This reaction may be
employed as a more stringent proof than that offered by the
microscopic characters of the crystals, and it may be well observed
under the microscope.

2. When cholesterin is gently heated with five volumes of
sulphuric acid and one volume of water, the edges of the crystals
are seen to become of a carmine colour; this reaction admits of

444 CHOLESTERIN. EXTRACTIVES. SALTS. [BOOK T.

being performed on a microscopic slide, and the results may be
watched under the microscope.

3. When cholesterin is dissolved in chloroform, and the chloro-
formic solution is shaken with an equal volume of strong sulphuric
acid, the chloroform becomes successively blue, red, cherry-red, and
ultimately purple, whilst the subjacent sulphuric acid acquires a
marked green fluorescence.

4. When heated gently with a mixture of one volume of solution
of ferric chloride and two volumes of hydrochloric acid, cholesterin

assumes a violet or blue colour.

« *

Cholesterin is a monad alcohol, and it readily forms

compounds with certain acids as with the volatile fatty acids.

tives of cho- ^^ ^e acti°n of bromine upon cho! ester in, both bodies

lesterin being dissolved in carbon disulphide, Cholesterin dibromide

(CaH^OBrj) is formed.

By the action of phosphorus pentachloride on dry cholesterin, cholesteryl
chloride C^H^Cl is obtained. By the action of an alcoholic solution of
ammonia upon the chloride, cholesterylamine CggH^NHo is obtained. By
treating a boiling alcoholic solution of cholesteryl chloride with sodium
amalgam, a crystalline carbo-hydrate having the composition C H and
a melting point of 90°, is obtained.

By the action of boiling nitric acid on cholesterin, cholesteric acid
is obtained, C8H10O5. This body is one of the substances obtained when
cholic acid is oxidized in a similar manner.

When oxidized by means of chromic acid, cholesterin yields oxycholic
acid, C^H^Oe. The two last compounds establish a close relationship
between cholesterin and the bile acids.

SECT. 7. EXTRACTIVE MATTERS OCCURRING IN THE NERVOUS
TISSUES WHICH ARE COMMON TO THESE AND TO OTHER TISSUES,
ESPECIALLY THE CONTRACTILE.

It is a fact worthy of notice that the brain contains considerable
quantities of the same bodies which are found in muscle, viz. crea-
tine, xanthine, hypoxanthine, inosit, and lactic acids ; in addition
it contains leucine, uric acid and probably urea.

According to W. Mttller1 the quantity of inosit in ox brain
amounts to 0'8 parts per 1000. The same author separated 0'6 grms.pf
uric acid from 50 pounds of ox brains. Mtiller found creatine in
the brain of man, but not in that of the ox.

The quantity of lactic acid separated from ox brain is said to
amount to 0'5 per 1000, and, strangely, to be identical with the lactic
acid of fermentation. As Kuhne has remarked, this lactic acid may
take its origin from the inosit of the brain.

1 Muller, Annal der Chemie u. Pharm.t Vol. cm. (1857), p. 131.

CHAP. X.] THE NERVOUS TISSUES. 445

SECT. 8. THE INORGANIC CONSTITUENTS OF THE NERVOUS TISSUES.

Brain is extremely poor in inorganic matters, though it is difficult
from the discrepant results of various writers to give reliable facts as
to the exact amount of these ; the estimates vary between 01 and
1 per cent, of the fresh brain.

Not only are statements discrepant as to the total quantity
of brain ash, but also as to the relative amounts found in the white
and grey matters. It appears to be true that the ash of the grey
matter has an alkaline, whilst that of the white has an acid reaction.

The following table exhibits the results of the analyses of the
mineral matters of brain made in Hoppe-Seyler's laboratory by
Geoghegan1.

INORGANIC MATTERS CONTAINED IN 1000 PARTS OF BRAIN.

(1) (2) (3) (4)

Cl ..................... 1-20 0-430 1-320 T064

P04 .................. 1-40 0956 2-016 1'392

CO3 .................. 0796 0-244 0'548 0'330

S04 .................. 0-220 0-102 0-136 0132

Fe PO ............ 0-010 0-096 0'098 0'032

Ca ..................... 0-005 0-020 0'014 0'022

Mg .................. 0-016 0-068 0-080 0'072

K ..................... 1-630 0-580 1778 1-520

Na .................. 1-000 0-450 1114 0780

Total Ash.., 6-277 2'946 7'084 5'344

SECT. 9. GENERAL SUMMARY SHEWING THE RESULTS OF QUANTITA-
TIVE ANALYSES OF BRAIN, SPINAL CORD AND NEKVES.

1. Proportion of Water.

The amount of water is much larger in grey than in white
matter, in early than adult life. The following are observations made
by Weisbach on the brains of men.

PROPORTION OF WATER IN 100 PARTS.

Age Age Age

20 to 30 30 to 50 70 to 94

White substance of brain 69'56 68-31 72-61

Grey „ „ 83-36 83-60 8478

Cerebellum 78'83 77'87 80'34

PonsVarolii 73'46 72'55 7274

Medulla oblongata 74'43 73'25 73'62

In the foetus the brain contains between 87'9 and 92'6 per cent, of
water.

1 Geoghegan, "Ueber die anorganischen Gehirnsalze." Zeitschr. f. phys. Chern.,
Vol. i. p. 330.

446 CHEMICAL PROCESSES IN THE NERVOUS TISSUES. [BOOK I.

The proportion of water in the spinal cord is less than in the
brain. Thus Bernhardt obtained the following results : —

PROPORTION OF WATER IN THE SPINAL CORD AND MEDULLA

OBLONGATA.

Cervical portion of cord 73 05 p c.

Lumbar „ „ 76 '04

Medulla oblongata 73'90

Cortex of brain 85*86

White matter of brain 70'08

Sympathetic cord 64'30

2. Proportion of the chief organic constituents of ox brain
(Petrowski1).

Grey matter. White matter.

Solids 18-40 p. c. 31-65 p. c.

Water 81-60 „ 68'35 „

Albumin and Gelatin . . 5 5 '37 24*72

Cerebrin 0'53 9*55

Lecithin 17'24 9'90

Cholesterin .... 18'68 51-91

Substances insoluble in anhydrous

ether 671 3'34

Salts 1-45 0-57

The above analyses, though interesting as. shewing the varying
proportions of certain of the brain constituents, such as water,
proteids, andcholesterin,mustnot be considered as throwing any light
upon the nature or distribution of the phosphorized constituents. The
phosphorus present in the mixed alcoholic and ethereal extracts having
been determined, a calculation was made upon the unwarrantable
assumption that all the phosphorus was derived from lecithin. The
reader who has perused the preceding pages will understand the
unfounded nature of this surmise.

SECT. 10. THE CHEMICAL PROCESSES CONNECTED WITH THE ACTIVITY
AND DEATH OF THE NERVOUS TISSUES.

We are acquainted with singularly few facts which throw a light
upon the chemical processes which have their seat in the organs of
the nervous system.

The great vascularity of the central organs as compared with the
nerves, and especially of the grey matter of the central organs,
establishes a presumption that processes have their seat in the nerve
cells of the grey matter which demand an abundance of oxygen.

1 Petrowsky, "Zusammensetzung der grauen und der weissen Substanz des Gehirns."
Pfliiger's Archiv, Vol. vn. p. 367.

CHAP. X.] THE NERVOUS TISSUES. 447

Observation of the living organism also teaches us that the proportion
of oxygen which is supplied to certain of the central organs influences
their activity in a remarkable manner; thus the activity of the
respiratory centre in the medulla is affected chiefly by the amount
of oxygen of the blood which traverses it. Again, an adequate
supply of oxygen to the brain appears to be a condition essential
to the proper exercise of the mental functions, and it is probably
in consequence of deprivation of oxygen that the moment blood
is cut off from the brain, as by ligaturing or compressing some of
the large arteries supplying it, all mental acts cease. When, however,
we direct our inquiries to the nature of the processes which have
their seat in the nerve cells we are obliged to conclude that we are
yet altogether in the dark.

The nerve fibre is much less directly influenced by a supply or
absence of oxygen than the central organs, and it is probably for
this reason, amongst others, that it survives, even in warm-blooded
animals, after the brain and spinal cord have ceased to manifest any
signs of vitality.

The only change of a chemical nature which has been proved
to occur in nerves as a result of long continued activity, or at death,
is a change in the reaction of the axis cylinder, which from an alkaline
changes to an acid reaction. The grey matter of the brain having an
acid reaction even during life *, no change can be observed to occur at
death.

When nerve fibres are cut off from their connection with certain
nerve cells, whilst the life of the animal is preserved, they gradually
undergo a fatty degeneration which affects the axis cylinders and
ultimately leads to an abolition of their power to act as conducting
organs.

1 Gscheidlen, "Ueber die Reaction der nervosen Centralorgane. " Pfluger's Archiv,
Vol. vni. p. 171.

CHAPTER XI.

CHEMICAL HISTORY OF CERTAIN OF THE PERIPHERAL
TERMINATIONS OF THE NERVOUS SYSTEM AND OF
THE ACCESSORY STRUCTURES CONNECTED WITH
THEM,— THE TISSUES AND MEDIA OF THE EAR,
THE TISSUES AND MEDIA OF THE EYE.

Introduc- DIRECTLY or indirectly all the nerve fibres of the

tory- organism are connected centrally with nerve centres, of

which we have examined the chemical history, so far as it is at
present knovva to us. Peripherally nerve fibres either commence in
certain special end-organs capable of being influenced by movements
in the external medium and of transmitting the influence through
the nerves to the nerve centres (afferent nerve fibres), or they ter-
minate in structures of which the immense majority are concerned in
bringing about changes in the position of different organs of the body,
and changes in the relation of the organism to the medium which it
inhabits (efferent nerve fibres}. Fibres of the latter class terminate by
peculiar end-organs in the contractile tissues which have formed the
subject matter of Chapter IX.

In the present chapter there remains to be discussed the chemi-
cal history, so far as it is known, of the peripheral nervous end-organs
which are connected with afferent nerves, though unfortunately it is
only in connection with the eye that any detailed information is
available. For reasons of expediency we shall consider not merely the
chemical facts relating to the actual nervous structures, but also those
relating to the accessory apparatus with which they are connected.

SECT. 1. THE TISSUES AND MEDIA OF THE EAR.

The organ of hearing of vertebrates, reduced to its simplest form,
consists of a membranous sac of greater or less complexity, termed the
membranous labyrinth, on the inner surface of which are situated

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS. 449

epithelial structures which are directly continuous with fibres of the
auditory nerve. The sac contains a liquid termed endolymph and
is usually separated from the bony or cartilaginous structures wherein
it lies, by a liquid termed perilymph, through which sound-waves
have to be transmitted before they can affect the structures contained
in the membranous labyrinth. On the inner surface of the mem-
branous labyrinth, in certain situations, are crystalline bodies usually
termed otoliths or otocoma.

Perilymph and Endolymph.

Peril b According to Dahnhardt1 the perilymph of the had-

dock is a somewhat tough gelatinous liquid, rich in
mucin, and containing a proteid matter precipitable by acids but not
coagulated by heat. It contains from 2'1 to 2'2 per cent, of solid
constituents. Its chief saline constituent is common salt.

Endo- According to the same observer the endolymph of

lymph. ^ne haddock is a clear liquid, containing 1*5 per cent, of

solid matter ; the quantity of mucin is small, and albumin is absent.

Otoliths, Lapilli, or Otoconia.

In the vestibular sacs of the membranous labyrinth of most
(though not of all) fishes, lying free upon the surface of the epi-
thelium and bathed by endolymph, lie small concretions, termed
otoliths, lapilli, or otoconia, which are either pulverulent, as in the
plagiostomatous fishes, or hard and stony, as in the osseous fishes. In
these cases the otoliths rest freely on the surface of the long pro-
jections of the hair-cells which line the otolith sacs.

Although by no means universally distributed, similar concretions
are met with in the vestibular sacs and in the ampullar commencements
of the semicircular canals throughout the various groups of vertebrate
animals, though as a general rule they do not present themselves as
individual lapilli, lying free, but rather as pulverulent crystalline
concretions lying imbedded in the epithelial lining of the sacs. In
the latter case the individual crystals are surrounded and held
together by a slimy organic matter. Otoliths also occur in many in-
vertebrate groups.

According to Johannes Muller, the otcliths of the osseous fishes
have a structure similar to that of the enamel of the teeth, though
the statement is one which cannot, on morphological grounds, be
comprehended and invites further examination2.

1 Dahnhardt, "Endolymphe u. Perilymphe." Arbeit, d. Kieler physiol. Instituts,
p. 103.

3 For much interesting information on otoliths consult Miiller's Elements of Phy-
siology, translated by Baly. Vol. n. p. 1129 et seq.

29

450 OTOLITHS. COENEA. [BOOK I.

Crystalline fine pulverulent otoliths which occur in most

appearance animals present the appearance of microscopic crystals,

of puiveru- presenting remarkable variations in size. Their form

lent otoliths. ^g s]iewn in the accompanying illustration.

FIG. 61. OTOLITHS, COMPOSED OP CALCIUM CAKBONATE. (From Frey, after Funke.)

According to Dahnhardt1 otoliths contain from about 74'5 to 77'5
per cent, of mineral matter, composed chiefly of calcium carbonate in
the form of crystals. The organic matter resembles mucus.

The membranous Labyrinth.

As yet no information whatever is possessed in reference to the
composition of the walls of the labyrinth. Mainly these are com-
posed of connective tissue, which is said to resemble the cornea in
structure2. Of the chemical characters of the epithelium of the
labyrinth nothing whatever is known.

SECT. 2. THE TISSUES AND MEDIA OF THE EYE.

The Cornea.

The ground-substance of the cornea presents, as has already been
pointed out (p. 271), very great similarity in chemical composition to
that of hyaline cartilage, and until lately it was asserted without
contradiction that both tissues, when subjected to prolonged boiling,
yield chondrin, although that substance, as obtained from the cornea,
was said to possess somewhat special reactions. As we have already
pointed out, Morochowitz3 denies the existence of chondrin and looks

1 Dahnhardt, "Endolymphe u. Perilymphe." Arbeit, d. Kieler physioL Institiits, p.
106.

2 Hensen, Op. cit. p. 68.

3 Morochowitz, "Zur Histochemie des Bindegewebes." Separat-Abdruck aus den
Verhandlungen des naturhist.-med. Vereins zu Heidelberg. Vol. i. Part 5.

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS. 451

upon the ground-substance of the cornea, like that of hyaline cartilage,
as composed of collagenous and mucin-yielding bodies.

When digested in sulphuric acid the cornea may be split up into
lamellae, whilst potassium permanganate separates these into fibrils
which are broader than those of the fibrillar connective tissue (Kiihne).

Acetic acid first renders the cornea transparent and afterwards
causes it to swell, though the ground-substance does not dissolve.
After digestion in dilute mineral acids, the ground-substance of the
cornea becomes readily soluble in boiling water.

When the cornea is heated to 55° C. it becomes opaque, in con-
sequence partly of changes in the corpuscles: in part, however, because
of the coagulation of proteids previously existing in solution in the
parenchymatous fluid bathing the tissue (Kiihne)1. By treating the
cornea with water, this liquid dissolves alkaline albuminates and a
globulin which, according to Schmidt, possesses fibrinoplastic activity.

Presence of ^n tne C01irse °f h*8 beautiful studies on the

Myosin in the histological structure of the yet living cornea corpuscles

protoplasm of Kiihne x was led, from the eminently contractile character

of their bodies, from their behaviour to stimuli, and from

the changes which they undergo at death, to surmise

the close relationship of their protoplasm to the substance of muscle.

This relationship, according to Bruns2, is further evidenced by the

fact that the cornea contains myosin, doubtless derived from its

corpuscles.

To obtain myosin, Bruns separated tlie cornea from the sclerotic, and
placed the finely divided structure in saturated solution of ISTaCl for 24
hours. The solution on being treated with large quantities of distilled
water deposited a precipitate, soluble in weak solutions of NaCl (containing
less than 10 per cent.) and in water containing 1 part in 1000 of hydro-
chloric acid.

Results of The following is an analysis of the cornea by

analyses of His.
cornea.

Water in 1000 parts .... 758'3
Collagen ... . 203'8

Organic matters insoluble in water . 28'4

Soluble salts ... . 8'4

Insoluble „ . .

lOOO'O

Sclerotic.

No special information is possessed in reference to the sclerotic,
which, however, consists of collagenous connective tissue.

1 Ktthne, Untersuchungen uber das Protoplasma. See section entitled "Das Proto-
plasma der Zellen in der Cornea " (p. 123—131).

2 Bruns, " Chemische Untersuchungen uber die Hornhaut des Auges." Hoppe-
Seyler's Untersuchungen, p. 260.

29—2

452 AQUEOUS HUMOR. CRYSTALLINE LENS. [BOOK I.

Aqueous Humor.

This liquid which fills the anterior chamber of the eye is free from
all formed elements. Although the anterior chamber must be looked
upon as essentially a lymph space, and the secretion of the aqueous
humor like that of lymph is essentially dependent upon the arterial
pressure1, yet it possesses a very different chemical composition.

Aqueous humor is a perfect transparent liquid of
roSe? specific gravity 1003—1009, possessed of alkaline
reaction.

Aqueous humor contains a trace of a proteid matter
wtic1} is stated by Ktihne to be fibrinoplastic, it also
contains about 4 parts per 1000 of extractive matters,
amongst which is urea, and from 7 to 8 parts per 1000 of mineral
matters.

The following are the results of an analysis by Lohmeyer2 of the
aqueous humor of a calf.

Water per 1000 .... 986*87

Proteids 1'22

Extractive matters . . . . 4'21

Sodium chloride . . . . 6 '89

Other mineral matters . . . 0'81

1000-00

Crystalline Lens.

The crystalline lens is composed of concentric layers of fibres,
which are essentially elongated cells, and which usually present more
or less marked serrated edges. The structure is bounded externally
by a capsule composed of a structureless membrane which appears to

Eossess physical and chemical characters similar to those of the sarco-
jmma of muscle.

The crystalline lens is not homogeneous, as its refractive index
increases as we pass from the more external to the more internal layers
— an optical property which probably bears a relation to the fact
that the specific gravity of the central portion of the lens is, according
to Chevenix3, greater than that of the superficial layers, in the propor-
tion of 1194 to 1076.

Chemical The lens contains about two-thirds of its weight of

constituents water; its solid matters consist chiefly of a globulin
of the Lens. (about 24'6 p.c.) besides some serum albumin; in addi-
tion they contain small quantities of fat, traces of cholesterin and salts.

1 Chawas, " Secretion des Humor aqueus im Bezug auf die Frage nach den Ursachen
der Lymphbildung." Pfliiger's Archiv, Yol. xvi. p. 143.

2 See Gorup-Besanez, Lelirbuch d. phys. Chemie, 4to. ed. (1878), p. 401.

3 Chevenix, quoted by Kuhne, Lehrbuch, p. 404.

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS. 453

The giobu- Under the name of Crystallin, Berzelius described

lin contained the proteid belonging to the group of globulins which
forms the chief solid constituent of the crystalline lens.
This substance is soluble in water holding oxygen in solution, forming
an opalescent liquid, which is precipitated by C02. According to
Laptschinsky1, acetic acid does not precipitate this body, which, how-
ever, separates in a flocculent form when its solution is heated to
70 ° C. According to this author the lens behaves fibrinoplastically :
according to Kiihne it does not possess that property.

The cornea becomes opaque after death ; it is believed (Kiihne)
that this is not due to any coagulation of a soluble proteid, but to
diffusion phenomena, leading to the formation of vacuoles in the
lens-fibres, which necessarily impair the passage of light.

Results of The following are the results of four analyses of the

analyses^! ° crystalline lens of the ox, made by Laptschinsky.
the lens. Proteids in 100 parts . 34'93

Lecithin . . . . 0'23

Cholesterin . . . 0'22

Fats .... 0-29

Soluble salts . . . 0'53

Insoluble salts . . 0'29

The following are the results of other analyses of the lens of the
ox made by Hoppe-Seyler and Laptschinsky.

Hoppe-Seyler Laptschinsky

Proteids in 100 parts . 33'03 . . 3472

Aqueous extract . . 0'94 . . 0'95

Alcohol extract . . 0'52 . . 0'37

Insoluble salts . . 012 . . 0'17

Soluble salts . . . 0'61 . . 0'50

Ash obtained on incinerating) . QQ

the aqueous extract j

Ash obtained on incinerating) A . _ - .,

the alcoholic extract j U'U

Ethereal extract . . 0'45

Changes of By introducing solutions of salts or of sugar under

the lens in the skin of frogs a form of cataract is induced in which
cataract. ^e structure presents vacuoles ; these have been pro-

duced apparently by the abstraction of water from the lens ; the
cataract which occurs in the course of some cases of diabetes is proba-
bly induced in this way.

In genuine cataract the more common change consists, however,
in a fatty degeneration of the lens in which cholesterin is abundantly
deposited ; occasionally it is said that the lens is the seat of a depo-
sition of calcareous salts.

i Laptschinsky, Ein Beitrag zur Chemie des Linsengewebes.

454 THE VITREOUS BODY. THE RETINA. [BOOK I.

The Vitreous Body.

The vitreous body or humor consists essentially of mucous
connective tissue. In the very loose and large meshes of the tissue is
contained a large quantity of watery fluid, containing a small
quantity of proteids and said to be specially rich in urea (according to
Picard containing 0*5 per cent, of that body).

The following is an analysis of the vitreous body by Lohmeyer1.
Water in 1000 parts .... 986*400
Membranes . . . . . 0*210

Proteids and mucin (chiefly the latter) 1*360

Fats 0-016

Extractive matters (urea, &c.) . . 3*206

Sodium chloride .... 7757

Other mineral matters . . . 1*051

1000000

The Choroid.

The middle coat of the eye or Choroid is eminently vascular and
contains, imbedded within its substance, branched pigment-cells very
similar to those of the frog's skin. Until the researches of Max
Schultze had shewn that they properly belonged to the retina, the
layer of hexagonal pigment-cells (retinal epithelium) which we shall
describe in the sequel were described as an integral part of the
choroid.

All that need now be said in reference to the chemistry of the
choroid is that its branched pigment-cells contain a pigment which
appears to be similar to, if not absolutely identical with, that which
under the term of Fuscin we shall describe as the pigment of the
retinal epithelium.

THE RETINA.

Introductory.

The retina is the most internal of the tunics of the eye, and
contains the complex terminations of the optic nerve.

This membrane which by its internal surface lies in contact with,
or applied to, the external surface of the vitreous body and which
is covered externally by the vascular choroid, possesses during life
an exquisite transparency and doubtless throughout its greater part
absolute optical homogeneity, so that undulations of light which have
traversed the transparent media of the eye and impinge upon the
inner surface of the retina, may penetrate to the very peripheral struc-
tures which they are destined to throw into action.

The retina possesses a connective-tissue framework, wherein lie
imbedded the greater part of its nervous elements, but which does

1 Quoted by Gorup-Besanez.

CHAP. XL]

PERIPHERAL NERVOUS END-ORGANS.

455

not extend so far as to afford support to those structures (the rods
and cones) which are eminently the end-organs of the optic nerve or
to those pigrnented epithelium cells which afford a close investment
to the outer limbs of the rods.

Description ^ri ^e accomPanymg engraving the structure of

of the ten the retina is semi-diagrammatically represented, so as

layers of the to shew with clearness the position, and the mutual

relations, of the ten layers which, since Max Schultze's

description, histologists have agreed to distinguish.

The first layer (1, Fig. 62) is composed of the so-called membrana
limitans internet, which is a fibrillated membrane belonging to the con-
nective-tissue framework.

The second layer (2, Fig. 62) is the nerve-fibre layer and is composed
of naked axis-cylinders continuous with the optic nerve fibres which
having pierced the sclerotic and cornea enter the retina at the ' col-
liculus nervi optici!

The third layer (3, Fig. 62) the nerve-cell or ganglionic layer, is com-

FlO. 62. DIAGRAMMATIC SECTION OF THE RETINA. (Max

456 STRUCTURE OF THE RETINA. [BOOK I.

posed of multipolar nerve-cells which unquestionably communicate
by certain of their processes with the fibres of the second layer,
and by other more delicate processes with the delicate reticulum
which constitutes a great part of the fourth layer.

The fourth layer (4, Fig. 62), termed the inner molecular layer, is
composed in part of fibres belonging to the connective-tissue frame-
work, which afford support to a delicate reticulum which doubtless
is the medium of communication between the layers which lie internal
and external to it.

The fifth layer (5, Fig. 62), or internal granular layer (also inner
nuclear layer), is conspicuous for the presence of the so-called granules,
viz. small transparent nucleated spherical bodies with two poles, of
which one points towards the inner the other towards the outer
molecular layer, and which are doubtless connected with the networks
of those layers. These granules are considered by all to belong to
the nervous elements of the retina.

In addition we observe, however, in the fifth layer certain granules
which are probably connective-tissue cells, and radiating fibres (fibres
of Muller) which belong to the connective-tissue framework, which is
specially well developed in the granular layers.

The sixth layer (6, Fig. 62), or outer molecular layer, possesses a
structure similar to that of the fourth or inner molecular layer, consisting
of fibres of which some doubtless belong to the connective-tissue frame-
work and merely afford support for a truly nervous reticulum ; this
outer molecular layer is much less deep than the inner molecular layer.

The seventh layer (7, Fig. 62), or external granular layer (also outer
nuclear layer), presents many strata of bodies resembling in the main
those characteristic of the inner granular layer, and like them
presenting nuclei and two poles of which the inner pass to the
reticulum of the outer molecular layer ; the granules offer however
peculiarities : — Istly their external poles very clearly are connected
with either the rods or cones of the ninth layer : 2ndly the granules
which are connected with the cones (cone- granules) are larger, are
situated in the more external strata of the layer, and are directly
joined to the cones without the intermediation of fibres, whilst the
rod-granules are smaller, are joined to the rods by fibres and present
two transverse stripes.

The eighth layer (83 Fig. 62) is composed of the external limiting
membrane and, like the first layer, is composed of a fibrillated
membrane which forms the external boundary of the connective-
tissue framework of the retina; within this boundary the retina
is vascular, outside it is absolutely free from blood-vessels. The
external limiting membrane is perforated by the communications
between the rods and cones and the outer granular layer.

The ninth layer (9, Fig. 62), bacillary layer or layer of rods and
cones, is composed of the bodies which are by common consent and for
undeniable reasons considered to be the end-organs which are directly
excited by luminous undulations, which initiate the state of activity,

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS.

457

of the Visual
Epithelium.

which travels through the more internal layers of the retina and
ultimately stimulates the optic nerve fibres.

Particular The roc^s an(^ cones are included by Klihne in the

description term of visual cells (Sehzellen) or visual epithelium cells,
and must be distinguished from the retinal epithelium
cells, viz. the pigmented epithelium cells of the 10th
layer. Both rods and cones are distinctly nervous elements in so
far that they are doubtless in unbroken connection with the layer
from which nerve fibres ultimately spring. The rods and cones
possess some points in common and some which are distinguishing.
"Each consists of two distinct segments — an inner and an outer;
the division between the two occurring, in the case of the rods, about
the middle of their length (in man); in the cones at the junction
of the finer tapering end-piece with the basal part ; consequently, the
outer and inner segments of the rods are nearly similar in size and
shape, the inner being, however, slightly bulged as a rule, whereas
the inner segment of the cone far exceeds the outer
one in size, the latter appearing merely as an appen-
dage of the inner segment (fig. 63).

"The two segments both of the rods and cones ex-
hibit well-marked differences, both in their chemical
and optical characters, as well as in the structural ap-
pearances which may be observed in them. Thus while
in both the outer segment is doubly refracting in its
action upon light, the inner is, on the contrary,
singly refracting : the inner is stained by carmine,
iodine, and other colouring fluids, whilst the outer
remains colourless. The outer segment in both shews
a tendency to break up into a number of minute super-
imposed disks, whereas the inner segment is itself again
distinguishable into two parts — an outer part, appa-
rently composed of fine fibrils, and an inner part,
homogeneous, or finely granular1," by which they com-
municate directly, in the case of the rods with a rod-
fibre, in the case of the cones with a cone-granule.
The inner limbs of the rods are longer than those of
the cones ; on the other hand the outer limbs of the
cones are much shorter than those of the rods so that
the latter project above the former.

Retinal Epi- The tenth layer (10, Fig. 62), or pig-

theiium. mentary layer, is composed of a single

layer of hexagonal pigmented epithelium cells, form-
ing a mosaic which covers the outer limbs of the
rods ; these cells are characterized by possessing long
processes which extend from their anterior faces in a
beard-like fashion, and lie in the crevices between the rods ^and cones.

FIG. 63. A BOD

AND CONE FROM
THE HUMAN RETI-
NA. (Schultze.)

Quain, Elements of Anatomy, 8th ed. (edited by Schafer), Vol. n. p. 613.

458

STKUCTUKE OF THE RETINA.

[BOOK i.

Kiihne has made the discovery that the protoplasm of these cells is
the seat of remarkable movements, as proved by the different distri-
bution of the pigment in them, dependent upon the degree of
illumination to which the eye has been subjected. In the pigment
cells of a frog which has been kept for several hours in the dark, the
pigment is found to be confined to the cell bodies and the roots of the
processes coming from these. But if microscopical sections be made
of the eyes of frogs which have remained for some time in the sun-
light, the pigment will be found to have extended itself much further
forward in the cell processes towards the membrana limitans externa,
a proportionally smaller quantity remaining in the cell bodies. In the
eyes of frogs which have been exposed to light, the retina, when removed,

Tapetum.

FIG. 64. KETINAL EPITHELIUM CELLS. (Max Schultze.)

(a) Cells seen from external surface; (b) and (c) Cells seen in profile, with
processes projecting inwards.

has much epithelium attached to it. Conversely, in the eyes of frogs
which have not been exposed to light, the retina can be removed without
its epithelial covering. These facts will be again referred to in discussing
the functions of the retinal epithelium in regenerating the visual purple.
In animals possessing a tapetum, the epithelial layer
of the retina is unpigmented in the tapetal area, and the
choroid is composed anteriorly of a dense, strongly light-reflecting tissue.
In some animals, as the sheep or ox, the tapetum is composed of fine
fibrous tissue. In others, as the dog and cat, it is made up of several
layers of unpigmented cells which are filled with exceedingly fine
crystals (Max Schultze). Some fish, as the bream (Abramis Brama)
possess a so-called psuudo-tapetum (Briicke, Kiihne and Sewall1) ; in
the bream the retinal epithelium contains, in certain areas, both dark
pigment and amorphous strongly light-reflecting Guanin, so that the
epithelium seen from before presents a bright or a dark surface ac-
cording as the pigment, under the influence of darkness or light, is
found in tLe bases or processes of the cells.

y The preceding description of the structure of the

in the struc- retina does not apply to every part of its surface, though
ture of the we must refer the reader who requires detailed infor-
mation on this subject to treatises on histology. Suffice
it to say that at the entrance of the optic nerve ( Colli-
culus nervi optici) the nerve fibre layer is immensely

1 Kiihne und Sewall, "Zur Physiologie des Sehepithels. " Verhandl. d. naturhist.
Vereins zu Heidelberg, N.S. Vol. n. Heft v. (June, 1880).

retina in
different
regions.

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS. 459

developed, whilst the other nervous layers are absent. At the so-
called macula lutea, and especially at its central depression, tho
fovea centralis^ cones are found to the exclusion of rods ; at the
periphery of the macula lutea cones are found, each surrounded by a
circle of rods, whilst over the rest of the retina the cones are found
sparsely distributed amongst the rods.

Variation ^e ^acillary lajer °f the retina does not always

in the distr- possess both rods and cones ; in some animals we find
button of rods rods and in others cones, or where both are present their
and cones in relative number varies. Thus the following animals
the retinae of nave no cones : — the ray, the shark, the sturgeon, the
cifsseTof kat, ^e nec%en°g> tne mole. The following animals

animals. nave no rods: — lizards, serpents, tortoises, and perhaps

all reptiles. All mammals have more rods than cones;
nearly all birds have more cones than rods, though in the owl, which
is a nocturnal bird, the cones are very few.

Chemical composition of the Retina as a whole.

In consequence of the scanty material at the chemist's disposal
little is known as to the general composition of the retina. The
reaction of the retina is said to be acid. According to C. Schmidt1
the retina, besides containing albumin, yields, on boiling, gelatin and
mucin. Its alcoholic extract yields a body which gives a crystal-
lizable compound with platinum chloride and which smells of tri-
inethylamine, doubtless due to the decomposition of neurine. As
Klihne remarks, we may on general grounds surmise that the retina
contains the same bodies as the central nervous systems.

Whilst the living retina is perfectly transparent, at death it
becomes opaque, doubtless in consequence of the coagulation of some
proteid constituent.

General Chemical facts relating to Rods and Cones.
Chemical -^e inner segments of both rods and cones are corn-

structure of posed of protoplasm which during life is possessed of
the inner marvellous transparency ; after death this becomes

limbs of the opaque and presents granular deposits, nuclei, and in
some cases spherical, lenticular, or paraboloid, highly
refractile bodies.
rho^i^i The outer limbs of the rods are composed of an

dnemicai . . . -1 .

structure of external envelope, which agrees, m physical characters
the outer and in its power of resisting various agents, with neuro-

limbs of the keratin. This external envelope encloses contents which
morphologically appear as little disks which are sepa-
rated by an intermediate substance ; it is impossible to distinguish
between the chemical characters of these two kinds of substances.
Ktilme has pointed out that the contents of the envelopes consist of a
mixture of proteid bodies and of substances soluble in alcohol and

1 See Kiihne, « Chemie der Netzhaut. " Hermann's Handbuch, Vol. in. Part 1, p. 239.

460 CHROMOPHANES. [BOOK I.

ether and doubtless similar to, if not identical with, those extracted by
these solvents from the nervous tissues. Kiihne has found, indeed, that
the contents of the outer limbs behave to osmic acid in almost the
same manner as the medullary sheath of nerves ; to the substance in
the contents which exhibits this reaction Kiihne ascribes the name
of rod-myeloid (Stabchen-myeloid), though he does not wish thereby
to indicate that it is a definite proximate principle.

Solubility -^ax Schultze first pointed out, and his observations

of the outer are confirmed by Kiihne, that the contents of the
limbs of both outer limbs of the rods and cones are dissolved with
rods and extraordinary rapidity and ease by bile, the envelope

cones imbue. alone remaining

Colouring matters associated with cones. (Chromophanes?)

The outer limbs of the cones differ from those of the rods in being
invariably free from colouring matters. In birds, reptiles and fishes,
however, the inner segment of each cone presents a minute globular
body, apparently of a fatty nature, and possessed of brilliant and
varied colours, violet, blue, green, yellow, and red, though red and
yellow are most frequently met with.

The fact that the pigments are held in solution by fats is proved
by the intensely brown colour which the coloured globules acquire
when treated with perosmic acid and by the fact that they are
dissolved by such solvents of fatty bodies as a mixture of alcohol and
ether, carbon disulphide, and benzol.

Whilst the colouring matters of the cones are grouped together
under the name of Chromophanes, Kiihne1 has succeeded in separa-
ting, and examining the physical properties of, three distinct colouring
matters, a green, a yellow, and a red, which he distinguishes by the
names of chlorophane, xanthopJiane and rhodophane respectively.

Method of A large number of eyes (50 to 300) of doves or hens

separating are bisected so as to cut off the anterior segments ; the
the Chromo- vitreous humor being removed, the posterior segments
of the eyes are placed at once in absolute alcohol ; as
soon as possible the alcohol is poured away and the eyes are tho-
roughly exhausted with ether. On evaporating the ether, a fiery-red"
fat is obtained which is dissolved in hot alcohol and saponified by the
action of caustic soda, water being used to replace the alcohol as it
evaporates. The hard soap which separates from the mother liquor
is well dried and then treated successively with petroleum ether, then
with ether, lastly with benzol, which dissolve in order Chlorophane,
Xanihophane, and Khodophane ; for the methods of purification the
reader is referred to the original paper.

General All the chromophanes when treated with solution of

characters of iodine assume, as Schwalbe pointed out, a blue colour

the chromo- which differs in intensity and shade according to the

phanes. shade of the particular chromophane. The chromo-

1 Kiihne und Ayres, Ueber lichtcbestdndige Farben der Netzhaut.

CHAP. XI.] PERIPHERAL NERVOUS END -ORGANS. 461

phanes slowly become decolourized even in the dark. They are
much more rapidly bleached in the light, though very much less
rapidly than the visual purple to be afterwards described. Under
the most favourable circumstances a solution of chlorophane exposed
to the direct rays of the sun will be bleached in a few hours ; a
solution of xanthophane under similar circumstances will resist for a
period three times longer, and a solution of rhodophane for a period
twenty times longer. The process of decolourization is stated by
Kuhne to be dependent upon the presence of oxygen and to be there-
fore probably due to oxidation changes.

Special Chlorophane is of a greenish yellow colour; its

characters of alcoholic and ethereal solutions possess this tint. They
Chlorophane. present two absorption bands; these (in the case of a
petroleum ether solution) are situated between F and G.

Special Xanthophane, ' unlike Chlorophane, is but slightly

characters of soluble in petroleum ether, but readily soluble in al-
Xanthophane. cohol, ether and carbon disulphide. The solutions ex-
hibit a strong absorption of the violet end of the spectrum and a
single absorption band, which in the case of the ethereal solution is
situated near F, and on its violet side. In the case of solutions in
bisulphide of carbon the absorption band is situated between b and F.

Special This colouring matter is not at all soluble in petro-

characters of leum ether or carbon disulphide. It is most readily
Rhodophane. soluble in oil of turpentine and in alcohol which has
been acidified with acetic acid ; these solutions become decolourized,
after some hours, even in the dark. Solutions in benzol may be kept
indefinitely. These solutions exhibit marked absorption of the violet
end of the spectrum and a single absorption band between b and F.

Colouring matters associated with the rods,
(Visual Purple or Ehodopsin)

Historical In the year 1851 Heinrich M tiller1 pointed out that

Notes. the rods of the retina of the frog when seen en masse

often present a reddish colouration. In 1857 Leidig2 referred to the
satiny-red colour possessed by the retina of the frog. Later Max
Schultze drew attention to the satiny-red colour of the rods of the
retina of the rat and owl.

These observations did not however attract marked attention and
were lost sight of until the publication of a remarkable paper by
Boll, presented to the Berlin Academy towards the close of the year
18763, in which that observer announced the startling fact that the
bacillary layer of the retina of all animals is during life not colourless,
but of a purple red colour.

1 H. Mueller, Zeitschr. f. wiss. Zoologie, Vol. in. p. 234.

2 Leidig, Lehrbuch d. Histologie, p. 238.

3 Boll, "Zur Anatomie u. Pbysiologie der Retina." Monatsber. d. Berl. Acad.,
12 Nov. 1876.

4G2 VISUAL PURPLE OR RHODOPSIN. [BOOK I.

During life, according to Boll, the peculiar colour of the retina is
perpetually being destroyed by the light which penetrates the eye;
darkness, however, restores the colour, which vanishes for ever almost
immediately after death.

The wonderfully suggestive nature of Boll's discovery led Kiihne
to repeat his observations1. Whilst generally confirming the funda-
mental statement of Boll, Kiihne at once was able to correct and
amplify Boll's account. In the first place, relying implicitly upon the
statements of Boll, he examined, as soon as possible after death, the
retinae of animals (frogs and rabbits) which had been kept for some
time in darkness. He soon found that the beautiful purple colour
persists after death if the retina be not exposed to light; that the
bleaching takes place so slowly in gas-light that by its aid the retina
can be prepared and the changes in its tint deliberately watched;
that when illuminated with monochromatic sodium light the purple
colour does not disappear in from twenty-four to twenty-eight hours
even though decomposition has set in. These first observations of
Kiihne on the vision-purple (Sehpurpur), as he termed it, whilst they
shewed that the disappearance of the colour is not, as Boll had assert-
ed, a necessary concomitant of death, removed many of the difficul-
ties which stood in the way of a careful investigation. Carrying out
his preparations in a dark chamber illuminated by a sodium flame,
Kiihne was able almost at once to discover the conditions necessary
to the destruction of the vision-purple, as well as the most important
facts relating to its restoration or removal. Since then the investi-
gation of the retinal pigments and of photo-chemical processes in the
eye ha,ve formed the subject of continuous and successful studies on
the part of Kiihne and his pupils, and it is to them that we owe all
the important facts relating to this fascinating subject2.

1 Kiihne, "Zur Photochemie der Netzhaut." Gelesen in der Sitzung des Natur-
historisch-medicinischen Yereins zu Heidelberg, den 5 Jan. 1877.

2 The following is a list (in the order of publication) of the researches of Kiihne and
his pupils on the retinal pigments and photochemical processes hi the retina which have
appeared in the Untersuchungen aus dem physiologischen Institute der Universitdt
Heidelberg.

(1) Kiihne, " Zur Photochemie der Netzhaut." (2 Abdruck.) UntersucJiung. Vol. i.
Part i.

(2) Kiihne, "Ueber den Sehpurpur." Ibid.

(3) Kiihne, "Ueber die Verbreitung des Sehpurpurs im menschlichen Auge."
UntersucJiung. Vol. i. Part ii. p. 105.

(4) Kiihne, "Weitere Beobachtungen iiber den Sehpurpur des Menschen." Ibid.
p. 109.

(5) Kiihne, ' 'Das Sehen ohne Sehpurpur." Ibid. p. 119.

(6) Ewald u. Kiihne, "Untersuchungen iiber den Sehpurpur." Ibid. p. 139.

(7) Kiihne, "Ueber die Darstellung von Optogrammen im Froschauge." Ibid.
Vol. i. Part iii. p. 225.

(8) Kiihne, "Eine Beobachtung iiber das Leuchten der Insectenaugen. " Ibid.
p. 242.

(9) Ewald u. Kiihne, "Untersuchungen iiber den Sehpurpur." (Fortsetzung. )
Ibid. p. 248.

(10) Kiihne, "Ueber lichtbestandige Farben der Netzhaut." Ibid. Vol. i. Part
iv. p. 341.

CHAP. XL] PERIPHERAL NERVOUS END-ORGANS. 463

Having given this brief account of the progress of discoveries on
the visual purple, a short abstract of all the more important facts
which have been brought to light may be given.

Distribution If the retina of a rabbit or a frog — preferably of

of the Visual one which has been kept for some time in the dark —
Purple in the ke quickly removed from the perfectly recent eye, in a
fcetina. room lighted with the help of a monochromatic yellow

light, and be taken into the daylight, it will be observed to be of a
purple-red colour, which quickly bleaches on exposure. On a closer
inspection it will be found that in a horizontal plane cutting the
retina the purple colour is more intense, forming a distinct purple
band, whilst the macula lutea and a rim 3 — 4 millimetres broad, at
the ora serrata, are devoid of colour. If the retina be examined
under the microscope the purple colour will be found to be limited to
the rods, and to the outer segments of these, all other parts of the retina
looking greenish by contrast. Thus the purple colour varies in fulness
directly with the richness of the retina in rods. The more cones, the
less visual purple : and vice versa. Hence the absence of purple from
the fovea centralis which contains cones only, and its entire deficiency
in the rod-less retinae of reptiles. But, although the colour is
confined to the outer limbs of the rods, it must not be supposed that
every rod is purple. The rods in the neighbourhood of the fovea
centralis (viz. in the macula lutea} lack colour, as also do the rods in
the colourless margin near the ora serrata. The cause of the greater
depth of purple in the horizontal zone previously referred to has not
been discovered, as, for instance, whether it is due to a more intense
colouration of each rod segment, or to a greater length of the rod
segments.

(11) Ewald u. Kiihne, " Untersuchungen iiber den Sehpurpur. " (Scliluss.) Ibid.
Vol. i. part iv. p. 370.

(12) Kiihne, " Beobachtungen iiber Druckblindheit." Ibid. Vol. n. Part i. p. 46.

(13) C. Fr. W. Krukenberg, "Ueber die Stabchenfarbe der Cephalopoden. "
Ibid. p. 58.

(14) Kiihne, "Beobachtungen an der frischen Netzhaut des Menschen. " Ibid.
p. 59.

(15) Kiihne, " Fortgesetzte Untersuchungen iiber die Retina und die Pigmente des
Auges." Ibid. p. 89.

(16) Ayres u. Ktihne, "Ueber Regeneration des Sehpurpurs beim Saugethiere."
Ibid. p. 215.

(17) Ewald, "Ueber die entoptische Wahrnehmung der Macula Lutea und des
Sehpurpurs." Ibid. p. 241.

(18) Kiihne, "Zur Abwehr einiger Irrthu'mer iiber das Verhalten des Sehpurpurs."
Ibid, p. 254.

(19) Kiihne, "Notiz iiber die Netzhaut der Eule." Ibid. p. 257.

(20) Mays, "Ueber das braune Pigment des Auges." Ibid. Heft in. p. 324.

(21) Kiihne, "Notizen zur Anatomic und Physiologie der Netzhaut." Ibid. p.
378.

The first two papers in the above list were translated from the German by Mrs
Foster, edited with notes by Dr Michael Foster and published under the title "On
the Photochemistry of the Retina and on the Visual Purple." London, Macmillan and
Co.* 1878.

Kiihne has recently given a systematic- account of his researches under the title of
"Chemische Vorgange in der Netzhaut" in Hermann's Handbuch der Physiologie, Vol. i.
Part i. (1879) p. 235—337.

464 VISUAL PURPLE OR RHODOPSIN. [BOOK I.

With regard to the distribution of the visual purple in the animal
kingdom, it is to be remarked that whilst the rod-bearing retinae
of vertebrates generally possess it, in a few isolated animals it is
inexplicably absent. Thus a species of bat (Rhinolophus hipposideros)
has no purple, and hens and pigeons want it, though bats have none
but rods in their retinae, while the birds mentioned, with a prepon-
derance of cones, yet possess rods also. With these exceptions, all
vertebrates with rod-bearing retinae possess the visual purple, and
all invertebrates hitherto examined lack it. It is found in day-
loving and night-loving animals, — in the sunward-flying eagle and
the nocturnal owl, in fishes which inhabit the sombre depths of the
ocean, and in the embryo into whose eye light has never fallen.

Method of Kiihne's study of the visual purple and of the

separation of changes which it undergoes by the action of light were
Visual Purple much aided by the discovery of the fact that the
or Rhodop- colouring matter is soluble in aqueous solutions contain-
ing from 2 — 5 p. c. of crystallized bile.

Colourless crystallized bile is obtained by evaporating ox bile to dryness
on the water-bath after mixing it thoroughly with much animal charcoal.
The perfectly dry residue is "heated with absolute alcohol and a large excess
of ether is added to the filtered solution ; by this means the salts of the bile
acids are precipitated and ultimately acquire a crystalline structure. The
precipitate which consists of sodium glycocholate and taurocholate is termed
' crystallized bile.'

The perfect isolation of rhodopsin by this solvent is beset with
difficulties, the greatest of which is to avoid contamination with blood-
colouring matter. The retinae of certain animals disappoint all
attempts to free them from haemoglobin and are therefore unfit for
the extraction of visual purple. Fortunately the frog is not among
these. Twenty to thirty frog retinae separated in the chamber by
the aid of sodium light, are moistened with about 1 c.c. of a 2 p. c.
solution of bile salts and shaken, but without violence, for an hour.
The mixture is allowed to stand so as to allow of the subsidence of the
grosser particles, and the supernatant fluid afterwards poured on to a
filter. The solution thus obtained is of a red-purple colour, bleach-
ing to a water-like fluid on exposure to light. The solution is
perfectly clear and transparent and does not fluoresce or seem
opalescent, if absolutely free from fuscin. It may be concentrated
rapidly in vacuo, yielding solutions of progressively deeper tints of
purple and finally a dark residue resembling ammoniacal carmine,
containing dark violet or black amorphous particles. This mass
reacts to light after the manner of solutions. It is hygroscopic and
its amorphous particles redissolve. If the bile solution of rhodopsin
is thrown upon a dialyser the bile escapes, leaving a violet magma
capable of being bleached in the sunlight.

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS. 465

Optical When solutions of rliodopsin are exposed to light,

characters the colour changes from a purple tint, through red and

of Rhodop- orange, to yellow Jbefore becoming colourless. According
to the rapidity of our observation, therefore, will be our
notion of the pristine tint, when we bring the solution into the light
to examine it. If our eye fixes it in the red stage first, and then we
begin to note the fading, we shall be led to conclude that the original
tint was a deep red rather than a violet ; and in fact, many observers,
as Boll who proposed for the colouring matter the name 'Seh-Roth',
have fallen into this error of description. To obviate this self-decep-
tion we must prepare, in the dark, solutions of the visual purple, of
strengths becoming progressively weaker, and bring them (one by
one in the order of their concentration) into the light. It will be ob-
served under these conditions that the tints of the different strengths
run from purple-violet (in the strongest solution) through purple-red,
carmine-red and rose-colour, to lilac in the weakest. In other words,
the fading of the colour on exposure to light is different from the
fading of the colour on progressive dilution. In the former case
appears a yellow admixture which is absent from the original colour.
Indeed it is to some extent a misnomer to speak mostly of a 'fading' of
the visual purple, for besides itself fading, the visual purple is converted
into a visual yellow, which in its turn fades. The hypothesis that
visual purple becomes visual yellow in the sunlight, while visual
yellow fades in the same circumstances, suffices to explain all the
diversities of tint presented by the retina. According to the rapidity
of the conversion of purple to yellow, and according to the rate at
which the yellow is dissipated altogether, will be the particular tint
of an exposed retina. It will be shewn that different regions of the
spectrum have different powers of converting and bleaching rhodopsin.

spectrum When light is passed through visual purple and after-

of Visual wards through a prism, there is obtained a spectrum

Purple and offering no defined absorption bands, but presenting a
Visual Yellow. generaj absorption of rays in the centre of the spectrum,
from a little to the red side of D to the violet side of G. Visual yellow
blocks the rays from the red side of F to the blue end of the spectrum.
The most complete absorption by the visual purple is in the region of
E : that by the yellow is at G.

The characteristic transformation of visual purple in

tne presence of sunlight opens up a number of in-
different teresting questions. Is it to the highly refrangible
wave-lengths invisible rays or to the coloured rays that the change is
Purple Vl dae ? Are rays of a11 degrees of refrangibility in the

visible spectrum equally concerned in the action?
What is the nature of the conversion of purple into yellow; is it
physical or chemical, a synthesis or a splitting asunder of complex
into simpler groups ? Several of these questions have already re-
ceived a satisfactory solution, as we shall now attempt to shew. The

G. 30

466 VISUAL PURPLE OR RHODOPSIN. [BOOK I:

entire beam of white light is by far the best transformer of the visual
purple — superior to light of any particular wave-length. The less
refrangible dark rays at the red end of the spectrum do not bleach
the visual purple. Whether the actinic rays at the violet end are
capable of slowly bleaching is not yet ascertained, though it is certain
that if possessed of activity it is almost immeasurably weaker than
that exerted by the coloured rays. Of the visible rays of the spectrum
those bleach the visual purple most freely and quickly which the
visual purple in solution most effectually quenches. Thus the order
of activity in the bleaching of the purple is as follows : — yellowish-
green, green, blue, green-yellow, yellow, violet, orange and red.
Between yellow-green and yellow the time of bleaching is con-
siderable ; it is less between yellow-green and green up to blue.

But intensity of light, or the quantity on the unit of surface, has
an influence which renders the above classification very general and
bespeaks for it some latitude. We may in connection with this
subject draw attention to a practical point, which has already been
referred to incidentally. The inconvenience of the preparation of the
visual purple in the dark or in a dim twilight may be met by using
a monochromatic light of slight decomposing powers. A glance at
the previously mentioned orders of decomposing-activity of light of
different colour will convince us that red would be the best light for
the preparation of visual purple, were it not that in red light it is
impossible to detect and avoid blood stains. But yellow light from a
sodium flame, which takes about two hours to bleach a frog's retina,
is a useful substitute.

Although rays from different regions of the spectrum differ in
their powers of transforming visual purple, yet no visible ray fails to
bleach it if the exposure be prolonged enough. Further, the rays
differ among themselves in the rate at which they convert and bleach
the retinal colours. White light, to which we refer as a standard,
brings about the following transitions from purple : — red-purple, pure
red, orange, yellow, chamois-yellow to no colour whatever.

On the other hand, the red rays of the spectrum produce a change
through pure red and orange to the palest yellow, whilst taking an
extraordinarily long time to do so. And the rays from the opposite
end of the spectrum cause the purple to merge into a final stage which
is not yellow but bright red or lilac. In other words, as the wave-
lengths diminish less and less visual yellow is produced ; or, perhaps,
the yellow which is produced is bleached as quickly, or more quickly,
than the still unaltered purple.

Influence of Light is not the only agent which affects the visual

temperature purple. When retinae are exposed to temperatures
upon the varying from about 50° C. to 76° C. the colour fades with

?le> a rapidity which increases with the temperature. At
52 — 53° in the absence of light it takes some hours to disappear, at 76°
it disappears instantly. That a low temperature does not interfere
with the action of light upon the purple-stained rods is shewn by the

CHAP. XI.] PERIPHERAL NERVOUS END-ORGANS. 467

fact that a temperature of - 13° C. does not materially impede the
bleaching by light.

Action of Caustic alkalies, acids, alcohols, ether, and chloro-

various che- form decolourize the retinae of recently killed frogs. On
micai agents the other hand, many agents whose activity niight
PurpieUal kave keen presumed upon, such as ammonia, alum,

the process of putrefaction, trypsin, are ineffectual in
changing the visual purple.

When de- ^n describing the effect of various agents upon the

prived of visual purple that body has usually been under con-

water the ditions which presupposed the presence of water. If,

Visual Purple however, water be withdrawn from the structure or

sukstance coloured with visual purple, though that
substance continues to be affected by sunlight, the time
during which the light must act is enormously increased.

optograms ^G ^act ^a* ^ie ^V'IUS retina possesses a colouring

matter which is decomposed by light led Kiihne very
early to enquire whether it was possible, under certain circumstances,
to obtain actual images on the retina, corresponding to objects which
have been looked at. After his first experiments Ktihne endeavoured
to observe, on the retinae of rabbits, bleached spots corresponding to
the images of external objects, but his endeavours failed. In the
course of his researches Kiihne discovered the remarkable fact which
will be described in the succeeding section, viz. that there exist
within the retina agents which are concerned in the restoration
of the visual purple. Taking for granted that such agent or agents
exist, it will follow that, in order to obtain on the retina a picture
of external objects, the effect of the light would have to be so
prolonged or so intense as to destroy the balance between the
destruction of the visual purple and the power possessed by certain
retinal elements to restore it.

Kiihne took a coloured rabbit and fixed its head and one of its
eye-balls at a distance of one metre and a half from an opening
thirty centimetres square in a window shutter. The head was
covered for five minutes with a black cloth, and then exposed for
three minutes to a somewhat cloudy mid-day sky. The animal was
then instantly decapitated; the eye-ball which had been exposed was
rapidly extirpated by the aid of yellow light, then opened, and
instantly plunged in a 5 per cent, solution of alum. Two minutes
after death, the second eye-ball, without removal from the head, was
subjected to exactly the same processes as the first, viz. to a similar
exposure to the same object, then extirpated. On the following
morning the milk-white and now toughened retinae of both eyes were
carefully isolated, separated from . the optic nerve, and turned ;
they then exhibited on a beautiful rose-red ground a nearly square
sharp image with sharply-defined edges; the image in the first eye

30—2

468 RETINAL EPITHELIUM. FUSCIN. [BOOK I.

was somewhat roseate in hue and less sharply defined than that in
the second, which was perfectly white. The size of the images was
somewhat greater than one square millimeter.

To the images obtained by following such a method as that
described Kiihne gives the name of Optograms. The process may be
modified by taking the retinae from the alum solution and then
drying them in vacuo in the dark. They are in this way rendered
•very resistant to the action of light.

Chemical facts relating to the Retinal Epithelium.

The retinal epithelium cells (formerly termed hexagonal pigment
cells of the choroid) present most externally a covering of neuro-
keratin; more internally they present a protoplasm wherein are
found imbedded one or more nuclei, and still more internally that
protoplasm presents large numbers of pigment granules. From this
part of the cell proceed processes which make their way between
the outer segments of the rods.

The protoplasm of these epithelium cells presents deposits which
are described by Kiihne as consisting of Myeloidin, besides a fat tinged
with yellow colouring matter termed Lipochrin ; in the more internal
part of the cell and in the processes is found the dark pigment now
termed Fuscin.

idin " term Kiihne indicates the fact that the

retinal epithelium cells contain deposits of a substance
closely resembling, if not identical with, that forming the medullary
sheaths of nerve fibres.

Fat is not a constant ingredient of retinal epithelium,

being absent in that of man, the ox and the pig. In the

frog it always is tinged of a deep golden or citron colour. It remains

fluid at low temperatures, and is readily extracted by ether, benzol and

carbon disulphide.

_. . . This is a yellow colouring matter extracted by ether

Lipochrin. r J P ,. ^ • •> ±\ J i

from the eyes ol irogs, irom which the retinae have

previously been removed.

Lipochrin presents two absorption bands, the position of which
differ according to the nature of the solvent. "When dissolved in
ether these bands a'e situated between F and G. This colouring-
matter seems to resemble somewhat a yellow colouring matter
which has been' named lutein, in consequence of its being readily
extracted from the corpora lutea of cows. Lipochrin is slowly bleached
in sunlight.

Fuscin.

The brown pigment of the fetinal epithelium has usually, like
other pigments of the same colour and appearance, been termed
Melanin; of late Ktihne has proposed to term it Fuscin. This

CHAP. XL] PERIPHERAL NERVOUS END-ORGANS. 4G9

pigment occurs in the form of elongated, sometimes spindle-shaped,
rods in the epithelium cells; these rods are protruded into the
processes of the cell protoplasm.

Method of More than 500 hens' eyes are bisected and the

separating posterior halves are placed whilst yet fresh in alcohol ;
Fuscin — Mays' they are then boiled in alcohol and afterwards extracted
with boiling ether and water. They are then subjected
to energetic trypsin-digestion for 24 hours. The pigment is then left
in little masses which are collected on gauze and triturated with alkali
with the object of separating nucleins. It is mechanically separated
from adherin neurokeratin.

Pro erties ^"° c^em'ca^ reagent dissolves fuscin, except con-

centrated acids and alkalies, and these only do so very
gradually, or by the aid of heat. On long boiling in concentrated
sulphuric acid fuscin dissolves, colouring the acid of a dark brown
colour. By long digestion in caustic alkalies and their carbonates,
fuscin dissolves.

In the presence of oxygen, fuscin is slowly bleached, apparently in
consequence of an oxidation process ; the sensibility of the pigment
obtained from different animals appears to differ.

Fuscin is a nitrogenous body and, on ignition, leaves a small
quantity of ash containing iron.

Action of Light upon the Visual Purple of the Living Eye.
Regeneration of Visual Purple.

Though when the eye is exposed to diffuse daylight the visual
purple is not destroyed, by exposing frogs for considerable periods to
direct sunlight the retinae are found to have been bleached. Byallo wing
such frogs to remain in comparative darkness the colour is however
soon restored. Amongst the earliest of Kiihne's experiments were
those which threw light upon the structures which retard the bleaching
of visual purple or are concerned in its restoration.

If an equatorial section be made through a recently extirpated
eye, and a flap of retina be lifted up from the underlying choroid to
which the retinal epithelium cells are adhering, and if the whole be
exposed to light, it will be found that the purple colour of the flap
will be destroyed, whilst the colour of the rest of the retina will
persist. If, however, the bleached portion of flap be carefully
replaced, so that it is again in contact with the retinal epithelium
cells, complete restoration of the visual purple occurs. The restora-
tion is a function of the living cells, and it appears to be independent
of the fuscin which they contain. As it is absolutely dependent on
the life of the structures which overlie the rods, it is natural that, it
should persist for a longer time after somatic death in the frog than in
the rabbit.

470 VISION WITHOUT VISUAL PURPLE. [BOOK I.

While the epithelium at the back of the retina is the agent in the
restoration of the visual purple, it is ascertained that it may impart
something to the rods themselves, leading to " auto-regeneration" as
Kiihne terms it. Frequently when an isolated retina is bleached it
will on being removed from the light regain somewhat of its purple
colour ; and similarly bile solutions of the visual purple, if they contain
no ether, may also exhibit this " auto-regeneracy," especially if both
retina and epithelium have been employed in making the solution.

Do the rods then contain a something out of which the visual
purple may be regenerated, and are the epithelial cells the agents of
this elaboration, withdrawing the supposed substance from the rods
and working it up into visual purple ?

In concluding this account of the visual purple it is
Vision expedient to point out what bearing, if any, the facts

Without i j "L j i_ i IT

Visual Purple which nave been described nave upon our knowledge
of the physiology of vision.

The most sensitive region of our eye — that which we turn upon
any object which we wish to see with the utmost distinctness — con-
tains cones only, and cones are just those elements of the bacillary
layer which are destitute of visual purple. Again, many animals which
are keen-sighted may be seen to have retinae which are quite free
from this colouring matter.

Under these circumstances we cannot assert that these beautiful
discoveries relating to the visual purple have succeeded in solving the
tempting problem as to the mode in which light affects the retina.
They, however, open up a promising field for speculation and hold out
inducements to those who would pursue similar lines of enquiry. The
changes in the visual purple are perhaps little more than accidental
accompaniments of more important, though by our senses unseen,
chemical processes — processes which may in reality be initiated by
the undulatory movements of the ether and of which the results may
be the real stimuli which normally throw the optic nerves into
activity.

INDEX.

ABELES, sugar in blood, 65 ; glycogen in

muscle, 334
Abramis Brarna, presence of guanin in

retinal epithelium of, 458
Abscess, 238

Absolute force of contracting muscle, 345
Absorption spectrum of blood, 91; map^

ping of, 94 ; see Spectrum
Acetonaemia, 168
Acetone in blood, 168 ; effect of on blood,

169

Acetylene, action on blood, 107
Acid-albumins, 17
Actiniochrome, 306, 307
ADAMKIEWICZ on specific heat of muscle,

345

ADDISON, pernicious anaemia, 154
Adenoid tissue, 250

Adipose tissue; occurrence of, 259 ; charac-
ters of, 259 ; extraction of fats of, 260 ;
chemical constitution of, 261; fats of,
general properties of, 261 ; saponifica-
tion of fats of, 262 ; of lower animals,
fats of, 264
AEBY, on water in bone, 273 ; analysis of

dentine, 291

Albumins, 16; derived, 17
Albuminoids, 22
Alkali-albumins, 17
ALVERGNIAT'S mercurial pump, 204
Amphichromatic reaction of living muscle,

360

Amphoteric reaction of living muscle, 360

Amyloid substance, 18

Anaemia, 137, 145; BECQTJEREL and Bo-

DIER'S classification of, 145; blood

changes in, 147 ; causes of, 146 ; HAYEM'S

classification of, 150; effect of iron on

blood of, 152 ; progressive pernicious,

154 ; nature of, 154 ; blood changes in,

155 ; of heart disease, 165 ; blood changes

in, 166

Analysis, of ash of serum, 69; salts of
plasma, 69; phosphoric acid and lime
of serum, 68, 70; soluble salts in se-
rum, 70; magnesium in serum, 70;
red corpuscles, 80; ultimate, of oxy-

haemoglobin, 88; ultimate, ofhaematin.
114; mineral constituents of red cor-
puscles, 122; "blood cells and plasma,
122; quantitative of blood, 127, 128;
quantitative of blood of Cephalopoda,
134 ; blood in chlorosis, 151 ; blood in
leucocythaemia, 154; blood in scurvy,
156; blood in diabetes, 172; lymph and
chyle, 228; synovia, 230; dropsical
fluids, 232 ; hydrocele fluid, 235 ; cere-
brospinal liquid, 236; pus serum, 240;
nuclein, 242, 424 ; pus corpuscles, 244 ;
gases of pus, 247; gelatin, 254, 270;
collagen, 254; elastin, 256; mucin,
258, 270; fats, 265; cartilage, 269;
chondrin, 270; bone (mineral matters
of), 275; comparative, bones in different
animals, 278; fossil bones, 280; bone in.
osteomalacia, 281; in rachitis, 282;
in caries, 284; in necrosis, 284; of bone,
methods of, 285; dentine, 291; enamel,
292; comparative, tooth, 293; fossil
teeth, 294; dentinal tissues, 294; ulti-
mate, of horny tissue, 298 ; hair, nails,
cow's horn and hoof, 298; of chitin, 300;
conchiolin, 301; spongin, 302; hyalin,
302; melanin, 304; turacin, 305; quan-
titative of muscle, 339 ; gaseous, scalded
muscle, 351: gaseous, muscle passing
into rigor, 352 ; gas in muscle, 354 and
355; gaseous, contracted muscle, 355 —
358 ; blood flowing to and from muscle,
375; non-gaseous constituents of blood
of muscle, 381; ultimate, protagon, 428;
cerebrin, 439, 440; pseudo-cerebrin,
441; inorganic matters in brain, 445;
general summary of brain, spinal cord
and nerves, 445, 446; water in nerve
substance, 445 ; cornea, 451 ; aqueous
humor, 452 ; crystalline lens, 453 ; vitre-
ous body, 454
ANDRAL, scurvy, 156

ANDRAL and GAVARRET, blood in disease,
138; blood in chlorosis, 151, 152; ty-
phoid fever, 159; intermittent fevers, 162
Annelids, blood of, 131
Antedonin, 306

472

INDEX.

A]'liicTcin, 306
Aplysio-purpurin, 307

Appaiatus, for changes in gaseous consti-
tuents of muscle, 319, 350

Aqueous Humor, 452

ARN STEIN, blood pigment in intermittent
fever, 162

ARONSTEIN, pure serum-albumin, 63

Articular Rheumatism, blood in, 158

Ascitic Fluid, 235

Ashes of Serum, composition of, 69

• Blood, determination of, 177 ; by

Rose's method, 179

Bone, determination of, 285

Axis-cylinder of nerve fibres, 421

AYRES und KUHNE, retina, 463

BABBINGTON, blood in diabetes, 170

Bacillus anthracis, in splenic fever, 161

Bacillus malariae, 162

BACON, FRANCIS, views as to a vital spirit,
409

BAUMANN on chitin, 300

BECQUEREL and RODIER, specific gravity
of blood, 26; blood in disease, 138 ; fibrin
in disease, 142 ; fats of blood in disease,
143; classification of anaemia, 145 —
147 ; blood in chlorosis, 151 ; scurvy,
156; purpura hemorrhagica, and hae-.
mophilia, 157 ; blood in acute rheuma-
tism, 157 ; febricula or ephemeral fever,
159; typhoid fever, 159; classification
of anaemia in heart disease, 166 ; fibrin
in pulmonary affections, 167; bone in
caries, 284

BEDDOES, calls attention to the older re-
searches of Mayow, 409 ; his discussion
of Girtanner's essay, and views of
muscular irritability, 413

Beef-tea, 326

Bees' wax, 264

BEIGEL, excretion of urea, 386

BENEKE, excretion of urea, 386

BERNARD, CLAUDE, on CO-haemoglobin,
104, 105 ; sugar in lymph, 224 ; on glyco-
gen, 334

BERNHARDT, water in spinal cord and me-
dulla oblongata, 446

BERNSTEIN on functional current, 347

BERT, gases of the blood, 204

BERTHELOT on chitin, 300; tunicin, 303

BERZELIUS, reaction of dead muscle, 359;
marrow, 277; blood in disease, 138

BIBRA (VON), blood of Helix pomatia and
Cephalopods, 132; osteomalacia, 281;
rachitis, 283; comparative analysis of
teeth, 293; analysis of fossil teeth, 294;
analysis of feathers, 298

BIERMER, pernicious anaemia, 154

Bile, preparation of rnucm from, 257

BILHARZ. parasites in blood, 161
Pnlharzia haematoliu, 161
Bilirubin in pus, 245
BIRCH, DE BURGH, bone, 273

BISTEOW, C2H2-haemoglobin, 107

BLAINVILLE (DE), on blood cells

BI.ANKENHORN, protagon, 427

Blood, physical characters of, 23 ; method
of collection of pure arterial or venous
blood for analysis &c., 23; organic con-
stitution of, 25 ; specific gravity of, 26 ;
reaction of, methods of its determination,
26; composition of, 126; gases of, 126;
effect of reducing agents on, 99 ; guaia-
cum test for, 102 ; amount of haemo-
globin in, 1 03 ; amount of iron in, 103 ;
amount of water in, 139. Action of car-
bonic oxide on, 104; action of nitric
oxide on, 106; action of acetylene or
ethine on, 107; action of hydrocyanic
acid on, 107

• Coagulation of, 27 ; determination of

its commencement, 27; duration of
various stages, 27; rapidity of in dif-
ferent animals, 28; formation of fibrin
in, 28 ; circumstances affecting coagu-
lation, 29; theories of, 42; influence
of salts on, 53; non-coagulation of
the blood in living vessels, 55

Methods of investigating: — determi-
nation of sp.gr., 174; of reaction, 176; of
water, 177; of solids, 177; of ashes, 177
(by Rose's method, 179); of fibrin, 180 ; of
haemoglobin, 182; of iron, 186; of cho-
lesterin, 187 ; of lecithin, 187 ; of fats,
187 ; of urea, 190 ; of uric acid, 193 ;
of sugar, 194 ; of w eight of moist cor-
puscles, 195; of gases, 196. Analysis of
gases of, 206. Total quantity in body,
determination of, 215. Carbonic oxide
in, its detection, 219

of muscle, see Muscle

of Invertebrates, 129: — functions

of, 129; characters of, 129; chemical
composition of, 130; colouring matter
of, in Sipunculus mulus, 131; haemo-
globin of, 130 ; green colour of, in An-
nelids, 131; blue, of Mollusca and
Molluscoida, 132 ; blue, of Octopus,
133; composition of in Cephalopoda,
134

changes in, produced by iron, 152

changes in, in disease, 136: — Intro-
duction to, 136 ; proportional variations
of principal constituents, in diseases in
general, 139; e.g. water, 139; coloured
corpuscles, 139; haemoglobin, 139; fibrin,
142; serum-albumin, 142; fats, 143;
cholesterin, 143 ; lecithin, 143 ; sugar,
143; urea, 143; uric acid, 143; ex-
tractives, 143 ; hypoxanthine, 143; salts,
143; gases, 143; respiratory capacity
of, 144

changes in, in particular diseases,

145: In disorders of nutrition, 145: — •
e.g. anaemia, 145; chlorosis, 150; leu-
cocythaemia, 152 ; progressive perni-
cious anaemia, 154; scurvy, 156; haemo-

INDEX.

phflla, 157; puipura haemorrhagiea, of, 273; organic basis of, 274; deeal-

- 7: anico

rification of, 273; oasein of, 274; incine

158 ; rheumatoid arthritis, 158; rickets, : . :. : i7i: ;: i : ...:-. :

158 ; osteomalacia. 158. In fevers, organic and ™»>g»-«l Blatters of, 274 ;

159:— febneula, 159; typhus fewer, 159; ^ :.::.::: :. L7-1: : :

typho*d&rrer,153;reUp^ngfeT«-, Ia9; mineral matters of, 275; oonstitotion of

splenic fever, 161; intermittent fevers, mineral matters of, 276; mneral mat-

162; scarlet fever, 163; measles, 163; ten of, influence of food on, 276. Mar-

-:_-~ .-. -.'.._: vrv-;: .!.>. \- :_:;.:.. r.~ :. .•..:.:..-.::. r. .:. i"7 :: -~_1 " :.•>.

163. In diseases of the heart, 164:— in- analysis aad composition of , 280. Chan-

.:' l.r^n 1;^.-,^ mtaatMfimtiam gesofiu<

:• -.. : .-_- -:::._ : ::. _, _:: -, .> . 280; in caries, 284; in necrosis, 284;

1 1^ ;-,::-:_ -_: Methods of analysis of, 285; p

167; bronchitis, 167; pleurisy, 167; of, for anatjais, 285. Fat»of,<istermnv
onalis, 167. In diseases of ation of, 285. Determinaton of ashes
Tn dJabfrtrn «M»lHfaig ifift - of, 285; of calcium, 286; of magnesium

'in the blood in diabetes, 168; in, 286; of phosphoric acid, 236; of car-
a diabetes, 170. In diseases V :. : ! — : .- 1 .

liaM k Was, 00
in, in disease, 162 Bonelkin, 306; source and properties of,

307

fever," 160; BaciU** mmtkncu, 161; Boracie Acid, action on coloured blood.
BUhmrzia kmrmmtaUm, 161; Distama corpuscles, 74

161; bacteria and micro- BOUELU, coagulation of blood, 42

161; septicaemia, BdncHKs on red corpuscle, 72

161; diphtheria, 161; scarlatina, 161; BOCBOWHB, gelatin, 254; cbondrin, 270

*Uri***ms*iMi* htnuMi^lte- baciUms BOWXAX'S sareous elements, 315

'malarias, 162 BKASBIS, objections to the chemical Tiers
BJood-Corposdes: in coagnlation, 34; ana- of muscular contraction. 413

lysis of, 123 Brain, wafer in, 446 ; chief organic con-
— — eoionred, general characters of, 71; Ktifry^ity in, 446 (tee Xerve tissues);
•hwtan :1 72: WMofal aatan ;:. 7i : reaction of grey matter of, 446

Etromaof.74, 80; effects of «fa and BsAinrKU, pernicious anaemia, 155

salines on, 72; action of bnaoe acid BKHHTJC, diseases of kidney, 173

on, 74; ^^^^ti^ methods of, 74; Bronchitis, blood in, 167

number in blood, 78; Density and BBOCABDEL, on gases of blood in disease,
of, 79; composition of, 79; sepa- 144

of stroma from pr^eids, 80; BRCCXE on muscle in polarized L-ght, 31 6;

of CO. on, 81; action)of O on, am fn**h awaml ane, : J J: .Ijc _ :i

81; nuclei of, 82; haemngjohin, in in muscle, 334 ; paraglobulin, 39; blood,
r,ll:;:-l : _ : 1 ,; : --•- 55; blood-cells, 73, 74

of, 121; gaaea^of, 123; BKCSTOS, nudei of red corpuscles, 82

BCCHAXAS'S discoveries relating to coagn-
of, 123; lation, 43; washed clot, 45; serum, 59

BrcHXKB, gases in lymph, 226

Bony-coat of blood, 23

of, 124; behaviouT m"eoagnlation, 124 BCXGK, on Xa and K in blood corpuscles,
Blood-stains, detection of by microscope, 122

- by spectroscope, 217; by chemical Buszn, on heat of contraction, 345

reactions, 217; guaiaeum test for, 218; BUSK, scurvy, 156

Blood^vesaeta of muscular tissue, 313 Calcium, determination of in bone, 286

Blue Stentorin, 307 Calcium sareolaetate, 362

Bi-rMEn^CH on blood-cells, 73 Carbamie Acid, in the blood, 65 ;

BOBCK on nutscle in polarized light, 316 Carbonates, Geissler's apparatus for
Bone***, method fix m<raf i- ^ anarysis of, 287

337; choodrin, 270; ehondri-gmeose, Carbonic Acid (CO,), tension of in lympb,
270; extractive matters of pus, 243; 226; in dropsical effusions, 234

cblorrhoiinie acid, 243 Carbonic Oxide, action of on blood, 104;
Bois-&raosi> (»c) on eftect of death in medico-legal detection of in blood, 287

muscle, 359; on afidtficstion in tetanus, Caries, bone-changes in, analyses of, ^234

360 ymwmmmS* Acid, 308; source, mode of pre-
Boii, retina, 461 paration, I^operties andcompositionaf,

Bone, structural elements of, 272; water 308; spectrum, 303; aglucoade, 308

474

INDEX.

Carnine, 332 ; preparation, properties and
chemical relations of, 332

Cartilage, structural elements of, 268;
varieties of, 268 ; cells of, 268 ; compo-
sition of, 268 ; analyses of, 269

Casein, 17

Cellulose (animal), see Tunicin

Centrifugal machine, 58

Cerebrin, 83, 426, 439 ; Geoghegan's pre-
paration of, 440; analyses of, 435, 439,
440; pseudo-cerebrin, 441; decomposi-
tion of, 441

Cerebro-spinal fluid, position of, 230;
characters of, 230; accumulations of,
236; analyses of, 236

Cetylid, 441

CHAKCOT, crystals in leucocythaemia, 153

CHAVVAS, aqueous humor, 452

CHEVENIX, lens, 452

Chitin, 299, 300; distribution, 299; pre-
paration, 300; properties of, 300; ele-
mentary composition and formula of,
300 ; products of decomposition of, 300

Chlorocruorin, 131 ; spectrum of, 131

Chlorophane, 460

Chlorophyll, 306

Chlorosis, definition of, 150; blood in,
150; composition of blood in, 151;
effect of iron on blood in, 152

Chlorrhodinic Acid, 243; preparation of,
from pus, 243

Cholera, blood in, 163

Cholesterin, 442; preparation, 442; pro-
perties, 443 ; crystals of, 443 ; reactions
of, 443 ; composition and derivatives of,
444; in the blood, 65; in corpuscles of
blood, 80, 84; proportional variations
of, in blood of diseases in general, 143 ;
determination of, in blood, 187; in
lymph, 223; in dropsical effusions,
234; in hydrocele fluid, 235

Choline, see Ne urine

Chondrigen, 269

Chondri-glucose, 270

Chondrin, 269 ; preparation of, 269 ; reac-
tions of, 269; composition of, 270;
analyses of, 270; decomposition products
of, 270; existence of, 271; in pus, 243

Choroid, the, 454

CHOSSAT, rickets, 283

CHBISTISON, blood in disease, 138 ; urea in
blood in disease, 143; blood in kidney
diseases, 172, 173

Chromophanes, 460; method of separating,
4CO ; general characters of, 460

CHUBCH, on turacin, 304, 305

Chyle, 221; see Lymph

Chyluria, 162; filaria sanguinis hominis
in, 162

CLOETTA on inosit, 336

Coagulated proteids, 18

Coagulation of proteids by heat, 14 ; how
to determine temperature at which it
occurs, 15; table exhibiting tempera-

ture at which it occurs, 16; of blood,
see Blood ; of lymph, 222 ; of dropsical
fluids, 233

COBBOLD, parasites in blood, 161

Cochineal, 306, 308

COHNHEIM'S areas, 314

Collagen, 252 ; preparation of, 252 ; analy-
sis of, 254 ; relation to gelatin, 254

Conchiolin, 301, 302; preparation and
composition of, 301 ; reactions of, 302

Cones of the Eetina, 457 ; general chemical
facts relating to, 459 ; colouring matters
associated with, 460 ; distribution of
in various classes of animals, 459

Connective tissues, the, 249

Connective Tissue proper, structural ele-
ments of, 250; varieties of, 250; cells
of, 251 ; effect of reagents upon cells of,
251 ; white fibres of, 252 ; collagen of,
252; gelatin of, 253; elastic fibres of,
255; ground substance of, 256; cement
of, 256; effect of silver salts on, 256;
action of alkalies on cement of, 256;
mucin of, 256; preparation of mucin
from, 257

Contraction of muscle, 342 (see Muscle);
rate, latent period, and course or curve
of, 342; idio-muscular, 343, 404; absolute
force of, 343, 344; maximum work of,
345; heat of, 345; electrical tensions
of, 347 ; chemical changes of, 349 ; more
O absorbed and more C02 exhaled dur-
ing, than during repose, 372 ; heat and
work in, 417

Cornea, 450; myosinin,451; analysis of, 451

Corpuscles of blood, 25, see Blood-cor-
puscles

Corpuscles of perivisceral fluid in sea-
urchins and Holothurians, 134

COSSAB-EWABT, bacteria in splenic disease,
161; see GEDDES

Crassamentum, 28

Creatine, 326—328 ; preparation of, 326 ;
Liebig's method for preparing, 326;
Neubauer's method, 326; Stadeler's
method, 327; properties of, 327; crystals
of, 327 ; compounds of, 327 ; derivatives
of, 328 ; synthesis and constitution of,
328 j quantity present in muscle, 328 ;
changes of amount in muscle, 364 ; in
brain, 444 ; in the blood, 65

Creatinine, 329 ; in the blood, 65

Cruor, 85

Crustaceorubrin, 306

Crusta petrosa or cement, 293 ; histology
of, 294

Crusta phlogistica, 28

Cyan-haematin, 115

DAHNHABDT, perilymph and endolymph,

449
DAHNHABDT and HENSEN, analysis of

lymph, 228
DAVAINE, bacteria in splenic disease, 163

INDEX.

475

DAVY, coagulation of blood, 29, 30

DELIA TOBEE, on red corpuscles, 72

DEMANT, on creatine in muscle, 328

DENIS, on preventing coagulation of blood,
33 ; fibrin, 36 ; plasmine, 46

Dentinal sheath, 290

Dentine, 289—291 ; origin, 289 ; micro-
scopical structure and micro-chemical
reactions of, 290; relation of bone to,
290 ; water, organic and mineral matter
of, 290 ; analysis of, 291

Derived Albumins, 17

Dextrogyrous, definition of term, 8

Dextrin in muscle, 336

Diabetes mellitus, blood in, 168 ; com-
parative state of blood and urine in,
168; coma of, 169; lipaemia of, 170;
fat emboli in, 170; analyses of blood in,
172

Diacetin, 264

DIACONOW, protagon, 426, 428, 431; phos-
phorized principles of yolk of egg, 431 ;
description of lecithin, 431; lecithin
in brain, 432 ; separation of lecithin
from brain, 432; neurine, 435; consti-
tution of lecithin, 436; distearyl-lecithin,
436 ; distearyl- glycerin-phosphoric acid,
436

Dialysers, 6, 7

Dialysis, 6

Dioleyl-lecithin, 437

Dipalmityl-lecithin, 437

DIPPEL'S oil, 19

Disdiaclasts, 317

Disease, blood changes in, 136

Disorders of Nutrition, definition of, 137;
blood changes in, 145

Distearyl-glycerin-phosphoric acid, 436

Distearyl-lecithin, 436, 437

Distoma haematobium, 161

DOBIE'S stripe, 313

DOGIEL, mode of obtaining lymph, 221

DONDEBS, on CO-haemoglobin, 106

DONNE on blood-cells, 73

DOYEEE, eminence of, 319

DBAPEB, excretion of urea, 386

DBECHSEL on extraction of fat, 265

DBESCHFELD, emboli in diabetes, 171

Dropsical fluid, varieties of, 232 ; analyses
of, 232 ; characters of, 233 ; coagulation
of, 233; proteids of, 233; salts of, 234;
extractives of, 234; gases of, 234;
pleural, 235 ; peritoneal, 235 ; peri-
cardial, 235; of hydrocele, 235; cerebro-
spinal, 236; methods of analysing,
236

Dropsy, production of, 231 ; fluids of, 232 j
composition of fluids of, 232

DBOSDOFF, absorption of peptones, 64

DUJAEDIN, on sarcode, 310

DUMAS, on blood-cells, 73 ; blood in
disease, 138

DUNCAN, JOHANN, blood corpuscles in dis-
ease, 147

DUPEE'S apparatus for determination of
urea, 192

EBEBTH on muscle plasma, 320

Egg- albumin, 16, 17

EICHHOBST, leucocythaemia, 152; per-
nicious anaemia, 155

Elastin, 255; preparation of, 256; com-
position of, 256; analyses of, 256;
solubility of, 256 ; products of decompo-
sition of, 256

Eminence of Doyere, 319

Enamel, 291, 292; structure and origin,
291; composition of, 291; analyses of, 292

Enamel organ, 289

Eudolymph, 449

Endothelium, 295

End-plates, 319

ENGELHABDT, lactic acid of muscle, 359

ENGELMANN, on involuntary muscle, 312;
voluntary muscle, 314, 317; on Krause's
membrane, 320; on contraction of
muscle. 342

Ephemeral fever, blood in, 159

Epidermis, 296, 297; action of reagents on,
297

Epithelium, 295, 296; tegumentary, 295

Erysipelas, blood in, 163

Ethidene-lactic acid, preparation, proper-
ties, products of oxidation and synthesis
of, 363

Ethine, action of, on blood, 107

Ethylidene-lacticacid, optically active, 361;
optically inactive, 362; derivatives of, 363

EULENBEBG, CO-blood, 219

EWALD, gases in dropsical fluids, 234 ; gases
in pus, 246, 247 ; solubility of elastin,
256; neurokeratin,423; retina, 462,463

Exercise, effects of, on gases of respira-
tion, 383

Exsiccators, 178

Extractives, 64

Extractive matters, of plasma and serum,
64 ; of blood in diseases in general, 143 ;
of lymph, 223 ; of dropsical effusions,
234; of pus serum, 239; of pus cells,
243, 244

Extract of meat, 325, 326 ; preparation of,
325; constituents of, 326

Fat emboli in diabetes, 170

Fatigue of muscle, 404; signs of, 404;
measure of, 405 ; causes of, 405

Fats, of blood, 65; of red corpuscles, 83;
proportional variations of, in disease,
143 ; determination of in blood, 187 ; of
lymph, 223 ; of adipose tissue, proper-
ties of, 261; of adipose tissue in lower
animals, 264: analysis of, in tissues,
265; Drechsel's apparatus for extrac-
tion of, 265 ; extraction of, 265 ; deter-
mination of, 265; separation of fatty
acids from, 266 ; of bone, determination
of, 285 ; in muscle, 334

476

INDEX.

Fatty acids, separation from neutral fats,
266; fractional precipitation of, 267

Feathers of Birds, pigments of, 301

Febricula, blood in, 159

Fermentable sugar in muscle, 336

Fever, blood in, see Blood

Fibrin, general characters of, 18; action
of pepsin on, 18; formation of, 49;
relation to coagulation of blood, 28;
arrangement in blood-clot, Eanvier's
demonstration of, 34; mode of separa-
tion for chemical examination, 35;
properties of, 36; composition of, 36;
amount in blood, 37 ; precursors of, in
liquor sanguinis, 37; amount of, in dis-
ease (augmentation and diminution of),
142; quantitative determination of, 180 ;
Hoppe-Seyler's method, 180; in coagu-
lated blood, 181; amount of in lymph,
223

'Fibrine ordinaire' of Denis, 46

'Fibrine soluble' of Denis, 46

Fibrin ferment, 39, 48; preparation of
solution of, 48; origin of, 49

Fibrinogen, 17 ; of blood, gener 1 charac-
ters of, 17; methods of obtaining,
40 ; coagulating influence of paraglobu-
lin on, 47 ; determination of, in liquor
sanguinis, 189; of lymph, dropsical
effusions, 233; of hydrocele fluid, 235

Fibrinoplastic substance, 37, 39

FICK, on lift, 344 ; on heat of contraction,
345 ; on heat and work of muscle, 346

FICK and WISLICENUS, experiments on the
excretion of nitrogen during muscular
work, 388 et seq.

Filaria sanguinis hominis, 162

FINKLER, blood in muscle. 380

FISCHER, chondrin, 270; chondri-glucose,
271; extractive matters of pus serum,
239

FITZ, pyocyanin, 245, 246

FLINT, experiments on the excretion of
nitrogen, in the case of the pedestrian
Weston, 395 et seq.

Fluorine, determination of in bone, 286

FOLEY and LEONARD, intermittent fevers,
162

FORDOS, pyocvanin, 245; pyoxanthose,
246

FORSTER, influence of food on composition
of bone, 276

Fossil bones, analyses and composition of,
280

FOSTER, B., blood in diabetes, 170

FOTHERGILL, A., views concerning the in-
fluence of oxygen on irritability, 411

FRANQUE, excretion of urea, 386

FRANKLAND, analysis of gases, 207 — 213 ;
determinations of the heat evolved in
oxidation &c., 3'J1

FREDERIQUE, temperature of coagulation
of proteids, 16; serum -globulin, 39;
coagulation of fibrinogen, 41; on co-

agulation blood, 55; haemocyanin, 133,
134 ; determination of fibriuogen in
liquor sanguinis, 183

FREMY, comparative analyses of bone, 278,
279 ; composition of fossil bones, 280

FRERICHS, pigment in blood in intermit-
tent fever, 163 ; uraemic phenomena,
173; synovia, 230

FREY, haemin-crystals, 115; haernatoi-
din, 120 ; connective tissue, 249

Functional current of muscle, 347

FUNKE, oxy -haemoglobin, 84; haematoi-
din, 120

GALLOIS, reaction for inosit, 338

GAMGEE, A., fibrin-ferment, 49; dissocia-
tion tension of 0 in 0-haemoglobin,
102; on C 0-haemoglobin, 106; action of
nitrites on haemoglobin, 109; blood in
diabetes, 171; estimation of urea in
blood, 191; protagon, 426, 427; on
Diaconow's observations, 433; on cere-
brin, 440 ; pseudo-cerebriu, 441 ; cho-
lesterin in brain, 442

GARROD, blood in disease, 138; uric acid in
blood in disease, 143 ; gout, 157 ; deter-
mination of uric acid in blood, 193, 194

Gases, of the blood in health, 126; of
liquor sanguinis, 70; of coloured blood
corpuscles, 123; of the blood in diseases in
general, 143; separation of, from blood,
196 ; determination of, in blood, 196 ;
methods of analyses of, 206 ; absorptio-
metric methods, 207; eudiometric
methods, 211; of lymph and chyle, 225;
of dropsical fluids, 234; of pus, 246; of
muscle, 349 ; nature of gases liberated iu
rigor, 354; relation between gases of
rigid and contracting muscle, 358

GASKELL, speculation as to muscles of
arterial walls being able to contract
along two axes, 312; observations on
flow of blood through muscle, 406

GAUTIER, preparation of mucin, 258; v
preparation of chrondrin, 269 ; action
of water on proteids, 19 ; sp. gr. of plasma,
34; serum, 60; fats of blood in disease, 143

GAULE, C02 in lymph, 227

GAVARRET, see ANDRAL

GAY-LUSSAC, fluorine in bone, 276

GEDDES, PATRICK, on perivisceral fluid of
sea-urchins, 134, 135

GEDDES, PATRICK, and J. COSSAR-EWAET, on
the life history of Spirillum, 161

GEISSLER'S specific gravity bottle, 177;
apparatus for analysis of carbonates, 287

Gelatin, in pus, 243; obtained from con-
nective tissues, 253; preparation of, 253 ;
nature of, 253 ; composition of, 254 ;
relation to chondriu, 254; analyses of,
2")4; products of decomposition of, 255;
effects of reagents on, 255

GENTH, blood of Limulus cyclops, 132 ; ex-
cretion of urea, 386

INDEX.

477

GEOGIIEGAN, imcleinin brain, 425; method
of preparing cerebrin, 440; analysis of
cerebrin, 440; decomposition of cerebriu,
cetylid, 441; mineral matter in brain,
445

GEKLACH, on salts in serum, 66; ash in
serum, 67 ; magnesium in serum, 70

GIRTANNER, gaseous analysis of muscle,
349, 350 ; his views of irritability, 413

GLISSON, ' Irritability,' 410

Globin, 112

Globulins, their general characters, 17;
members belonging to the group of, 17

Glucose, in the blood, in health, 65; in
blood of diabetes mellitus, 168

Glycerin, 263 ; preparation of, 263 ; pro-
perties of, 264; effects of reagents on,
264

Glycerin-jelly, 254

Glycerin-phosphoric Acid, 433; prepara-
tion, 434; properties, composition, and
constitution, 434

Glycogen, in white blood corpuscles, 125;
in pus, 244; in cartilage cells, 268; in
muscle, 334; separation and determina-
tion of in muscle, 334 ; Abeles' method,
334 ; proportion in resting muscle, 335

Glycogen-dextrin, 336

Glycosamine, 301; preparation, properties
and probable constitution of, 301

GMELIN'S reaction, 120

GOBLEY on phosphorized principle of
coloured corpuscles, 83; lecithine, 430;
cerebrin, 430

GORUP-BESANEZ (VON), on gelatin in blood,
153

GOSDEW on haematin, 117

Gout, blood in, 157

GOWERS, enumeration of blood corpuscles,
77 ; clinical estimation of haemoglobin,
184

GRAHAM, egg-albumin, 63

Granular red blood corpuscles, 124

GREHANT, determination of urea in blood,
191 ; gases of the blood, 204

GRIESMEYER, parasites in blood, 161

GRUTZNER oa oxidizing and reducing
properties of muscle, 365

GSCHKIDLEN, sp. gr. of blood, 26; oxy-
haemoglobin, 88; determination of urea
in blood, 190; volume of blood in
bodies of animals, 216; on oxidizing
and reducing properties of muscle, 365 ;
reaction of grey matter of brain, 447

Guaiacum test for blood, 102; test for
blood stains, 218

GUBLER AND QUEVENNE on lymph, 223

GUERIN, on rickets, 282

GULLIVER, coagulation of blood, 29, 43;
red corpuscles, 71

HAAS, specific rotation of proteids, 12;

serum- albumin, 64
Hair, 297

HARLESS, on functional current, 347;
blood of Helix pomatia and Cephalo-
poda, 132

Haematin, 84, 108; production of, 112;
spectrum of, 113; preparation of, 114;
properties of, 114; percentage composi-
tion of, 114; action of HC1 on, 115; of
potassium cyanide on, 115; hydrochlo-
rate of, 115 ; reduced, 108, 118

Haematinometer, 92

Haematoblasts, 124

Haematoidin, 120; identity with bilirubin,
120; spectrum of, 121; action of nitric
acid on, 120

Haematolin, 118

Haematoporphyrin, 115; preparation of,
117; properties of, 117; spectrum of,
117

Haematoscope, 92

Haemin, 115; preparation of crystals of,
115; properties of, 116; preparation in
large quantities, 116; test for blool
stains, 218

Haemochromogen, 108; spectrum of, 111,
118; preparation of, 118

Haemocyanin, 133

Haemocytometer, 77

Haemoglobin, 71; in corpuscles of various
animals, 80; respiratory oxygen of, 91 ;
spectrum of, 99; reduced, 99; effect of
nitrogen or hydrogen on, 97; effect of
oxygen on, 100; amount in blood, 103;
relation of to number of corpuscles,
104; compounds with oxygen, 84;
carbonic oxide, 104; nitric oxide, 106;
acetylene, 107; hydrocyanic acid, 107;
products of decomposition of, 108; in
blood of Invertebrata, 130; amount of,
in general diseases, 139; quantitative
determination of, 182; Hoppe-Seyler's
method, 182; Preyer's method, 184;
Gower's method, 184; by amount of
iron contained, 186; clinical estimation
of, 184; of muscle, 325. See Oxy-hae-
moglobin.

Haemoglobinometer, 186

Haemophilia, .blood in, 157

Haemorrhagic diathesis, see Haemophilia

Haemoscope, 92

HALLER, views on irritability, 410

HAMMARSTEN, temperature of coagulation
of proteids, 16; preventing coagulation
of blood, 33; method of separating se-
rum-globulin, 37, 38; serum-globulin,
39, 61; fibrinogen, 40; coagulation of
blood, 51 — 53; determination of serum-
globulin in serum, 189; mode of ob-
taining lymph, 221; gases in lymph,
225 ; hydrocele fluid, 235

HAMMOND, excretion of urea, 386

HARTOO, MARCUS, on muscle in Cyclops,
313

HAYCRAFT, determination of urea in blood,
192, 193

478

INDEX.

HAYEM, enumeration of bloocUcorpnscles,
77; haematoblasts, 124; blood corpus-
cles in disease, 148, 149 ; classification
of cases of anaemia, 150
Heart disease, blood in, 164; influence of,
on composition of blood, 164: anaemia
of, 165

Heart-muscle, structure of, 318

Heat of contracting muscle, 345; influ-

.. ence of load upon, 346 ; relation of heat
and work, 346

Heat, specific, of muscle, 345

HEFNER, changes in water and alcohol
extractives of muscle, 364

HEIDENHAIN, volume of blood in animals,
216; on contraction of cartilage cells,
311 ; curve of muscle contraction, 342,
343; heat of contraction, 345, 346; lactic
acid in contraction, 360 ; water and al-
cohol extractives of muscle, 364; heat
and work in contraction, 417

HEINTZ, lactic acid in muscle, 359; fatty
acids, 267; analysis of calcined bone,
275

HEITZMANN, osteomalacia, 281

HELMHOLTZ, on latent period, 352; abso-
lute force of contraction, 344; heat of
contraction, 345 ; water and alcohol ex-
tractives in muscle, 364

Hemicollin, 255

HENSEN'S disc, 313

HEBAPATH, indigo-blue in pus, 246

HERMANN, phosphorized principle of
coloured corpuscles, 83; haematoscope,
92; NO-haemoglobin, 107; on contrac-
tility of protoplasm, 310 ; Krause's mem-
brane, 320 ; maximum work of muscle,
345; muscle-current, 347; functional cur-
rent, 347 ; rigor mortis, 348 ; apparatus
for extracting gas from muscle, 351;
gaseous analysis of scalded muscle, 352;
secondary discharge of muscle gases,
354; and gases in rigor, 354, 355;
method for analysis of gas of contracted
muscle, 356; gas in rigor, 358; fat in
muscle, 365 ; respiration of muscle, 369,
370; on influence of medium on muscle,
370, 371; muscular respiration, 417 ;
'Inogene' theory, 417

HEUCK, leucocythaemia, 156

HEWSGN on coagulation of blood, 29, 30,
33, 43, 55 ; red corpuscles, 72

Hexanitroinosit, 338

HEYDENREICH, relapsing fever, 160

HEYMANN, marrow, 277

HEYNSIUS, on nature of paraglobulin, 40 ;
serum-albumin, 63

Hippuric Acid in the blood, 65

HOFMEISTER, gelatin, 253, 270; relation
of gelatin to collagen, 254 ; semiglutin
and hemicollin, 255

HOLM, haematoidin, 120

Holothurians, peri visceral fluid of, 134

HOME, Sre EVERARD, on haematoidin, 120

Hoof, 297
Hoplacanthinin, 306

HOPPE-SEYLEE, chondrin, 271'; bone, 273;
CO-blood, 219 ; on gelatin, 299 ; inosit,
337; cartilage, 268; chondri-glucose,
271; relation of dentine to bone, 290;
dentine, 290, 291 ; enamel, 291, 292 ; anal-
ysis of horny tissue, 298 ; percentage com-
position of proteids, 5 ; specific rotation
of proteids, 12 ; temperature of coagula-
tion of proteids, 16; preparation of serum-
albumin, 62; lecithin in serum, 65;
composition of red corpuscles, 80 ; sepa-
ration of proteids of stroma, 81 ; phos-
phorized principle of coloured corpuscles,
83 ; neutral fats in red corpuscles, 84 ;
oxy-haemoglobin, 84, 87, 88, 90, 91; spec-
tra of haemoglobin, 100 ; methods for re-
ducing 0 -haemoglobin, 101; CO-haemo-
globin, 105, 106 ; compound of HCN and
haemoglobin, 107 ; methaemoglobin,
111, 112; haematin, 113, 114, 115; hae-
min, 116; haematoporphyrin, 117; hae-
matolin,118;haemochromogen, 118, 119,
120; fat in diabetic blood, 172; method
for separation of fibrin, 180 ; determina-
tion of haemoglobin in blood, 182, 183;
determination of cholesterin, lecithin
and fats in blood, 187 ; determination
of proteids in serum, 188; by polari-
meter, 189 ; analysis of chyle, 229 ;
dropsical liquid, 232, 233; hydrocele
fluid, 236; cerebro-spinal liquid, 236;
serum of pus, 239 — 240 ; nuclein, 242 ;
glycogen in pus, 244 ; analysis of pus
corpuscles, 244 ; gelatin, 254 ; protagon,
426, 428, 431 ; lecithin in yolk of egg,
430 ; distearyl-lecithin, 437

Horn, 297

Hot-air oven, 178

Hot -water oven, 179

HUFNER, amount of respiratory 0 in 0-hae-
moglobin, 102

HUIZINGA, serum-albumin, 64

HUMBOLDT, AL. VON, on action of medium
on muscle, 370, 371; denies that oxygen
is the common principle of irritability,
414

HUNTER, JOHN, coagulation of blood, 29,
30, 42; views on muscular motion, 411

Hyalin, preparation and composition of,
302 ; products of decomposition of, 302

Hydrocele, liquid of, 235 ; analyses of,
235

Hydrogen, action on blood, 97

Hydrolytic decomposition, 19

Hypoxanthine or Sarcine, 329 ; prepara-
tion and properties of, 329 ; relations to
other bodies, 330 ; proportion of, in
muscle, 330 ; in the blood in health, 65 ;
in the blood in diseases, 143 ; in leu-
cocythaemic blood, 153

Idio-muscular contraction, 343, 404

INDEX.

479

Inogene substance, 418

Inorganic matters in brain, 445

Inosinic acid, 333

Inosit, 336— 338 ; distribution, 336; prepar-
ation of, 337 ; Boedecker's method, 337 ;
properties and crystals of, 337 ; deriva-
tives of, 338 ; proportion of, in muscle,
338 ; in brain, 444

Intermediate blood corpuscles, 124

Intermittent fevers, blood in, 162 ; Bacil-
lus malariae in, 162

Invertebrate animals, blood of, 129

Involuntary muscle, 311, 312 ; see Mus-
cle

Iodized serum, preparation of, 252 ; action
on connective-tissue cells, 252

Iron, in blood, amount of, 103 ; effect of
on blood, 152; determination of in blood,
186

JACOBSEN, on taurine in muscle, 333

JADERHOLM, spectrum of CO-haemoglobin,
105 ; compound of CO with haematin,
106 ; on Gamgee's nitrite-haemoglobin
spectrum, 109 ; haemochromogen, 120

JAKSCH (VON), nuclein in brain, 424

Janthinin, 307

JONES, BENCE, on Xanthine, 330

JONES, WHARTON, on white corpuscles, 123,
311

JUDELL on composition of red corpuscles,
80 ; phosphorized principle of coloured
corpuscles, 83 ; cholesterin in red cor-
puscles, 84

Kephaline, 437, 438

Kerasine, 442

Keratin, 297, 298; reactions of, 298; ulti-
mate analyses of, 298 ; inorganic matter
in, 298

KEYE'S solution, 78

Kidney, diseases of, blood in, 172

KINGZETT, glycerin-phosphoric acid, 434

KLEBS and TOMMASI-CRUDELI, on Bacillus
malariae in intermittent fevers, 162;
pigment in blood in intermittent fevers,
163

KLEIN, on sheath of involuntary muscle-
cells, 312

KLUPFEL, speculations on the causes of
change of reaction in urine, 401

KOBELL, determination of fluorine in bone,
288

KOCH, bacteria in splenic disease, 161

KOLLIKER, on oxy-haemoglobin, 84

KONIG, 277; influence of food on com-
position of bone

KRAUSE'S membrane, 313, 320; reaction
of, 322

KRAUSE, blood-cells, 73; theory of struc-
ture of muscular fibre, 315; first ob-
served distinction between red and pale
voluntary muscles, 317 (foot note)

KRONECKER, on tetanus of pale muscles,
343 ; fatigue of muscle, 405

KUHNE, reaction of blood, 26; stroma of
red blood corpuscles, 80, 81; globin,
113; liquid of pericardial effusion, 235;
solubility of elastin, 256; on contractility
of protoplasm, 311 ; nerve-endings, 319 ;
on muscle-plasma, 319, 320; method of
obtaining muscle-plasma, 322; xanthine,
331 ; re-extension of muscle, 343 ;
gaseous analysis in rigor, 352; neuri-
lemma, 422; neurokeratin, 423; lactic
acid in brain, 444; researches by himself
and his pupils on the chemistry of, and
photo-chemical processes in, the retina,
458—470

KUNDE, oxy-haemoglobin, 84

LACAZE-DUTHIERS, Tyrian purple, 307, 309

Lac-dye, 306

Lactic acids, the isomeric, 361 — 364; in
brain, 444; in leucocythaemic blood,
153

Lactic anhydride, 361

Lactide, 361

LAER (VON), analysis of hair, 298

Laevogyrous, definition of term, 8

LAMANSKY, on functional currents, 347

LANDOIS, haemoglobin-crystals in blood
of insects, 131

LANG, crystals of oxy-haemoglobin, 90

LANGERHANS, histology of heart muscle,
318

LANKESTER, on blue stentorin, 307; on
haemoglobin in muscle, 325; on Gam-
gee's nitrite -haemoglobin spectrum, 109;
haemoglobin, 129 ; haemoglobin in in-
vertebrates, 130; chlorocruorin, 131, 132

Lapilli, 449

LAPTSCHINSKY, crystalline lens, 453

Lardacein, 18

LARMUTH, LEOPOLD, protagon, 426

LAURENT'S Polarimetre a Penombres, de-
Ascription of, 8 ; theory of, 10

LEBERT, relapsing fever, 160

Lecithin, 426, 430—432; description of
(Diaconow), 431; compounds of, 432;
presence in brain, 432 ; separation from
brain (Diaconow), 432; products of de-
composition of, 433 ; , constitution of,
436; in the blood, 65; in blood cor-
puscles, 80 ; of blood in diseases in
general, 143 ; determination of in blood,
187
LEDDERHOSE, on chitin, 300; on glycos-

amine, 301
LEGEROT, respiratory capacity of blood in

disease, 145

LEHMANN, C.G., analysis of bone in osteo-
malacia, 281; salts in serum, 66 ; density
of blood corpuscles, 79 ; oxy-haemo-
globin, 84, 85 ; blood cells and plasma,
122 ; analysis of blood, 128 ; extractives
in lymph, 224 ; excretion of urea, 386

480

INDEX.

LEHMANN, L., excretion of urea, 336

LEIDIG, oxy -haemoglobin, 84 ; retina, 461

Lens, crystalline, 452 ; chemical con-
stituents of, 452; results of quantitative
analysis of, 453 ; changes of, in cataract,
453

LEONARD and FOLET, intermittent fevers,
^162

LEPINE, cyanosis, 144

LETELLIEB, rickets, 283

Leucocythaemia, 152; varieties of, 152 ;
blood in, 153 ; crystals in, 153; analysis
of blood of, 154 ; Eichhorst's corpuscles
in, 156; myelogenic, 152, 277

Leukaemia, 152 ; see Leucocythaemia

LlEBERMEISTER and GlLDERMEISTER, TC-

searches on the influence of cold on the
production of heat and the formation of
carbon dioxide, 404

LIEBIG, J. VON, method for preparing
creatine, 326; on xanthine, 330; on
urea in muscle, 333 ; iiiosinic acid, 333 ;
lactic acid in muscle, 359 ; salts in
serum, 67; views on muscular force,
415

LIEBIG, Gr., respiration of muscle, 365 ; on
action of medium on muscle, 370

LIEBREICH, reaction of blood, 26; CgHj-
haemoglobin, 107 ; protagon, 425 ; pro-
ducts of decomposition of protagon, 429;
neurine, 435

Lift of muscle, 344

LIMPRICHT, on taurine in muscle, 333 ; on
dextrin in muscle, 336

Lipaemia in diabetes, 170

Liquid in dropsies, see Dropsy

Liquor Pericardii, 229 ; see Pericardium

Liquor Sanguinis, 25, 31 ; methods of
obtaining, 31 ; properties of, 33 ; speci-
fic gravity of, 34 ; reaction of, 34 ;
coagulation of, 34 ; action of C02 on,
37, 40 ; extractive matters of, 64 ; salts
of, 66 ; composition of salts of, 69 ;
gases of, 70 ; mineral matters of, 122 ;
analyses of, 128; determination of fibrin-
ogen of, 189

LISTER, coagulation of blood, 30, 55 — 57

Liver, blood in diseases of, 167

LOWER, coagulation of Wood, 42

LTJBAVIN, nuclein, 243

LUCKE, pyocyanin, 245; composition of
hyalin, 302

LTJDWIG, mercurial pump, 199 ; lymph, 222 ;
on gases of blood, 372 ; methods for
investigating gases of blood of muscle,
375, 376, 377, 378

Lungs, blood in diseases of, 167

Lymph, nature of, 220 ; resemblance of to
dilute liquor sanguinis, 221; circum-
stances affecting quantity of, 221; modes
of obtaining, 221 ; physical characters
of, 221 ; corpuscles of, 222 ; plasma of,
222 ; molecular basis of, 222 ; reaction
of, 222 ; specific gravity of, 222 ; coagu-

lation of, 222; serum of, 222; pro-
teids of, 223; fats of, 223; extractives
of, 223; salts of, 224; gases of, 225;
analysis of, 228; methods of analysing,
236
Lymphatic leukaemia, 152

MACDONNEL, on the amount of glycogen in
muscles separated from the nerve-cen-
tres, 404

MAC MUNN, spectroscopy, 94

MAGENDIE, foramen of, 230

Magnesia mixture, 286

Magnesium, determination of, in bone,
286

Malacosteon, see Osteomalacia, 280

MALASSEZ, enumeration of blood corpus-
cles, 75, 78; relation of haemoglobin
to number of corpuscles, 104, 147;
blood-corpuscles in disease, 148 ; deter-
mination of haemoglobin in blood, 183 ;
method for estimating volume of blood,
216

MARCET, on xanthine in urinary calculi,
330

MARCHAND, osteomalacia, 281

MARCHAND and COLBERG, analysis of lymph,
228

MARCHIAFAVA, intermittent fever, 162 ; pig-
ment in blood in intermittent fever,
163

Margarin, 263 ; nature of, 263 ; crystals of,
263

Marrow of bone, 277

MATHIEU AND URBAIN, gas in pus, 246

MATTEUCCI, muscular respiration, 366, 367,
416

MAYER, J. E., views on irritability and
source of muscular power, 415

MAYOW, theory of respiration and muscular
activity, 407

Measles, blood in, 163

Meat, Extract of, 325

Medullary sheath, 422

Medullated nerve-fibres, 421 ; see Nerve
fibres

MEHRING (VON), sugar in blood, 65, 195 ;
sugar in lymph, 224 ; chondrin, 270

MEISSNER, determination of urea in blood,
190 ; uric acid in blood, 193 ; on
uric acid in muscle, 333

Melanin, 303 ; occurrence, characters and
reactions of, 303 ; percentage compo-
sition of, 304

Membrane, Krause's, 313; see Krause's
membrane

Mercurial pumps, 198; Ludwig's, 199;
Pfluger's, 200; Alvergniat's, 204

Metabolic processes, 5

Metabolism, 5

Methaemoglobin, 90; spectrum of, 109;
action of reducing agents on, 109 ;
production of under influence of nitrites,
109; nature of, 109, 112

INDEX.

481

Micro-spectroscopes, 96

MIESCHER, nuclein, 83, 242, 243 ; pus
cells, 241

MILLON'S reaction, 13

MILNE-EDWARDS, 011 rickets, 283; green
blood, 131

Mollities osseum, see Osteomalacia, 280

Mollusea, blue blood of, 132

Molluscnida, blue blood of, 132

Monacetin, 264

MORICHINI, 011 fluorides in bone, 276

MOROCHOWITZ on constitution of chondrin,
271

MOSELEY on certain animal pigments, 305,
306

MOSSLEE, excretion of urea, 386

Mucin, distribution of, 257; preparation
of, from connective tissue (Rollet),
from bile and sputum (Gautier), 257,
258; properties of, 258; composition
of, 258 ; analyses of, 258 ; products of
decomposition of, 259; relations of, 259

Mucus, nature of, 257

MULDER, on chondrin, 270; analyses of
hoof and nails, 298

MULLER, JOHANNES, on separation of blood
corpuscles, 33 ; chondrin, 270

MULLER, HEINRICH, retina, 461

MULLER, W., elastin, 256; inosit,336, cere-
brin, 439; inosit in brain, 440; uric acid
and creatine in brain, 444

MUSSY, GUENNAU OB, typhus fever, 159

Muscle, classification of, 311; structure of
unstriped involuntary, 311,312; struc-
ture of voluntary, 313 — 315 ; in'polarized
light, 316 ; blood-vessels of, 318 ; struc-
ture of heart muscle, 318; termination
of nerves in, 318 ; chemical constitution
of normal living, 319; distribution of
liquid and solid parts in, 319 ; plasma
of, 320; chemical nature of double-
refracting elements of voluntary muscle,
321; serum of, 324; haemoglobin of,
325; nitrogenous (non-proteid) organic
constituents of, 325 — 333; quantity of
creatine present in, 328; proportion of
hypoxanthine in, 330; proportion of
xanthine in, 331; uric acid in, 333;
urea in, 333; inosinic acid in, 333;
taurine in, 333; non-nitrogenous or-
ganic constituents of, 333—338; fats
in, 334 ; glycogen in, 334, 335 ; dextrin
in, 336; fermentable sugar in, 336;
inosit in, 336, 338; proportion of inosit
in, 338; ferments in, 338; inorganic
constituents of, 338, 339; water in,
338; mineral salts in, 339; summary
of quantitative composition of, 339.
General phenomena of living, 339 — 348 ;
at rest, 339, 340; phenomena of con-
tracting, special and general, 341; mi-
croscopic appearances, 341 ; rate of con-
traction of, 342; tetanus of, 343; red
and pale striated, 343 ; absolute force of

G.

contracting, 343, 344; maximum work
of, 345 ; heat of contracting, 345 ; specific
heat of, 345; proportion of heat and work
yielded by active, 346 ; electrical tensions
of contracting, 347; functional current
of, 347; rigor mortis of, 347. Special
study of chemical changes of living, 348
— 349; methods of studying chemistry
of, 349 ; chemical changes of contraction
and rigor, 349 ; changes in the gaseous
constituents of, 349; gaseous analysis
of scalded, 351; gaseous analysis of, in
rigor, 352; secondary or putrefactive
discharge of gases of, 353. Gaseous
analysis of contracted muscle, 353 — 358 ;
relation between gases of rigid and
contracting, 358 ; changes in non-gase-
ous constituents of muscle in activity
and rigor, 359; changes in reaction
and its causes, 359 ; methods of de-
termining reaction of, 360; acidification
of tetanized muscle removed from in-
fluence of blood, 360; cause of acid
reaction in rigor, 360; changes in pro-
portion of water in, 364; changes in
water and alcohol extractives of, 364;
changes in proteids of, 364; changes of
creatin of, 364; changes of proportion
of glycogen and sugar in, 365; changes
in fat and fatty acids in, 365 ; oxidising
and reducing properties of, in rest and
tetanus, 365; changes in chemical
composition of medium surrounding,
365. Eespiration of, 365 ; influence of
medium upon irritability of, 370; in-
fluence of oxygen on thick and thin,
371; resting muscles exhale CO2, 372;
contracting muscles absorb more 0 and
exhale more C02 than resting, 372;
changes in chemical composition of
medium surrounding muscle, 373;
influence of O on irritability of, 380;
analysis of non-gaseous constituents of
blood of, 381 ; changes in medium sur-
rounding muscle determined by general
excreta of body, 381 ; effects of muscu-
lar exercise on pulmonary exchanges,
381; chemical changes in living, when
at rest, 401; fatigue of, 404; measure
of (Kronecker's experiments), 405 ; causes
of, 405

Muscle-plasma, 320; Kuhne's method of
obtaining, 322; properties of, 323

Muscle-serum, 324; proteids of, 324

Muscular activity, theories of, 406; John
Mayow, 407 ; Glisson and Haller, 409 ;
Whytt, 410; John Hunter, 411 ; Fother-
gill and Girtanner, 411 ; Beddoes, 413 ;
Brandis, 413 ; Eeil and von Madai, 414 ;
Humboldt, 414; Leibig, 414; J. E.
Mayer, 415; Voit, 416; M. Traube,
416 ; Matteucci, 416 ; Hermann, 427

Myelines, 437, 438

Myelogenic leukaemia, 277

31

482

INDEX.

Myeloplaxes, 272

Myosin, 17, 323, 324; reactions of muscle-
plasma depending on, 323 ; preparation
of, 323 ; coagulation of, 324

Myosin-syntonin, 324

NACHET, enumeration of blood corpuscles,
77

Nail, 2D7

NASSE, nitrogen in proteids, 19 ; specific
gravity of blood, 26; coagulation of
blood, 28; glycogen in muscle, 334, 365;
method of determining it, 335; glycogen-
dextrin, 336; on fermentable sugar in
muscle, 336 ; on marrow, 277

NAWALICHIN on heat of contraction, 345,
346

NAWROCKI on creatinine, 329 ; on creatin,
365

Necrosis, bone changes in, analysis of,
284

NEXCKI, action of pancreatic ferments on
gelatin, 255

Nerves, termination of, in muscle, 318

Nerve-cells, 420, 421 ; histology of, 420,
421 ; micro-chemistry of, 421

Nerve-fibres, 421, 422; medullated, 421;
histology of, 421

Nerve-organs, classification of, 420

Nervous tissues, 420; grey and white
matter of, 420 ; phosphorized constitu-
ents of, 425; phosphorized principles
of, other than protagon and lecithin, 437;
non-phosphorized nitrogenous bodies of,
439; extractives in, 444; inorganic
constituents of, 445; water in, 445;
chemical processes connected with ac-
tivity and death of, 446

NEU BAUER, method for preparing creatine,
326; on creatinine, 329; onhypoxanthine,
330 ; preparation of xanthine, 330

NEUMANN, leucocythaemia, 152

Neurilemma, 421

Neurine or choline, 426, 435, 436; pre-
paration, properties and products of
decomposition of, 435 ; synthesis of, 436

Neuroglia, 420

Neurokeratin, 423, 424; reactions and
mode of preparation of, 423 ; properties
of, 424

Neutral fats in the blood, 65

NIGETIET, changes in water and alcohol
extractives of muscle, 364

Nitric Oxide, action of on blood, 106

Nitrites, action of on the blood, 109

Nitro-aerial particles of Mayow, 407

Nitrogen, action on blood, 97

Nitrogen, excretion of by the urine : sum-
mary of general effects of muscular
contraction upon, 399

Non-coagulation of the blood within living
vessels, 54

NORTH, experiments on the excretion of
urea during rest and work, 398

Nuclei of red corpuscles, chemical compo-
sition of, 82

Nuclein, 83, 424 ; nature of, 241 ; compo-
sition of, 242; analysis of, 242; exist-
ence of, 243

OBERMEIER, blood in relapsing fever, 159,
160

OBOLENSKY, mucin, 259, 270

Octopus, blood of, 133

ODENIUS and LANG, analysis of lymph,
238

Odontoblasts, 289

Oekoid, 74

Olein, 263; preparation of, 263; nature
of, 263

Optograms, 467

Ossein, 274

Osseous tissue, see Bone

Osteoblasts, 272

Osteomalacia, blood changes in, 158;
bone changes in, 280 ; analyses of bones
in, 281

Otoconia, 449

Otoliths, 449

Oxalic Acid in gout, 157

Oxy-chlorocruorin, 132; spectrum of, 131

Oxy-haemocyanin, 133

Oxy-haemoglobin, 84 ; methods of prepara-
tion of, 85; elementary composition of,
88; percentage composition in various
animals, 88; crystalline form of, 89;
chemical reactions of, 90 ; general cha-
racters of, 90; decomposition of, 90;
absorption spectrum of, 91; spectra
which may be derived from, 97; effect
of nitrogen or hydrogen on, 97 ; reduc-
tion of, 100 ; amount of oxygen in, 102 ;
reaction with guaiacum, 102; activity
of oxygen of, 103; action of carbonic
oxide on, 104 ; action of nitric oxide on,
106 ; action of acetylene on, 107

Palmitin, 263 ; how obtained, 263
Pancreatic ferment, action on reduced

haemoglobin, 101
PANUM, serum-casein, 61; volume of

blood in bodies of animals, 216
PAPILLON, blood of octopus, 133;
Paraglobulin, 17; general characters of,
17 ; methods for obtaining, 37 ; pro-
perties of, 38; coagulating influence on
fibrinogen, 47; derivation of, 39, 50,
52 ; importance of, in formation of
fibrin, 47, 51; in serum, 60
PAEKE, protagon in yolk of egg, 430
PARKES, observations of, on the elimination
of nitrogen during muscular work, 392
et seq.

'Particulae igneo-aereae ' of Mayow, 407
PAVY, sugar in blood, 65 ; blood in diabetes
mellitus, 168; determination of sugar

INDEX.

483

in blood, 194, 195 ; observations on the
excretion of nitrogen in the case of the
pedestrian Weston, 395 et seq.

Pentacrinin, purple and red, 306

Peptones, general characters of, 17; rela-
tion to proteids, 5

Pericardium, liquor of, 229; characters of
effusions into, 235

Perilymph, 449

Periosteum, 272

Peritoneal transudations, characters of,
235; analyses of, 232

Pernicious anaemia, 154; see Anaemia

PETIT, rickets, 282

PETROWSKY, proteids of brain, 423 ; chief
organic constituents of brain, 446

PETTENKOFER, method for determining
O absorbed and C02 excreted, 383

PETTENKOFER and VOIT, experiments on
the total excretion of nitrogen in the
urine, 388

PETTERS, acetonaemia, 169

PFLUGER, on constitution of proteids, 21,
22; sp. gr. of blood, 26; on activity of
O in 0-haemoglobin, 103; gases of
blood, 126; C02 in lymph, 226; mer-
curial pump, 200; gaseous analysis of
muscle, 357 ; on the changes of blood in
muscle, 378, 379, 380

Phaseomannite, 336

PHIPSON on xanthine, 330

Phosphoric Acid, estimation of, in blood-
serum, 69 ; in bone, 286

Phrenosine, 442

Phthisis pulmonalis, blood in, 167

PICARD, urea in blood in disease, 143;
determination of urea in blood, 190 ; in
muscle, 333

Picro-carminate of ammonia, preparation
of solution of, 183

Pigments, brown and black of epithelial
tissues of vertebrates, 303; of feathers
of birds, 304; turacin, 304; in animal
kingdom generally, 305—309

Pigmentum Nigrum, 304

PLANER, gases in dropsical liquid, 234

Plasma Sanguinis, 25, 31 ; see Liquor
Sanguinis

Plasma of muscle, 320

Plasmine, 46

Pleural transudations, characters of, 235;
analysis of, 232, 233

PLOSZ, nuclei of red corpuscles, 82; nuclein,
242

Pneumonia, blood in, 167

Polarimeters, 8

Polaristrobometers, 8

Polyporythrin, 306

POTAIN'S solution, 78

POINTING, J. H., on Laurent's Polarimetre,
10—12

Pseudo-cerebrin, 441; analysis of, 441

Punicin, 309

Purple, Tyrian, 309 ; see Tyrian purple

Purpura haemorrhagica, blood in, 157

Pus, 238; physical characters of, 238;
microscopical character of, 239; seruni
of, see Pus Serum; corpuscles of, see
Pus Corpuscles; nature of, 239; colour-
ing matters of, 245 ; gases of, 246 ; analy-
sis of, directions for, 248

Pus Corpuscles, origin of, 239 ; action of
NaCl upon, 241 ; proteids of, 241 ; nu-
cleus of, 241; extractive matters of,
243—244; gelatin of, 243; chondrin of,
243; preparation of chlorrhodiuic acid
from, 243; glycogen of, 244; mineral
matters of, 244

Pus Serum, 239; proteids of, 239; ex-
tractives of, 239 ; salts of, 240 ; analyses
of, 240

Pyocyanin, 245; preparation of, 245; pro-
perties of, 245 ; bacterium of, 246

Pyoxanthose, 246

Pyrocatechin, 259

PBEVOST, blood-cells, 73 ; blood in disease,
138

PREYER, oxy- haemoglobin, 85, 88; spectra
of 0-haemoglobin, haemoglobin and
CO-haemoglobin, 98, 99; amount of
respiratory 0 of 0-haemoglobin, 102;
proportion of Fe and haemoglobin in
blood, 103, 104; globin, 112; haematoi-
din, 121 ; determination of haemoglobin
in blood, 184

PRIBRAM, salts in serum, 66; ash in serum,
67

Protagon, preparation of, 83, 425, 427;
nature of, 83; discovery by Liebreich,
425; formula and properties of, 426;
constitution of, 426 ; Gamgee and Blan-
kenhorn's method of preparation, 427;
ultimate analysis of, 428; stability of,
423; products of decomposition of, 429,
433; action of alkalies and aeids on,
429

Proteids, occurrence of, 3 ; general cha-
racters of, 4; proportion of, in various
liquids and solids, 4; origin of, 4; per-
centage composition of, 5 ; solubility
of, 5, 13; diffusibility of, 6; rotatory
power of, 7 ; chemical reactions of, 13 ;
detection of, in solution, 13; methods of
completely separating, 14; determina-
tion of temperature of coagulation of,
14 ; synopsis of the chief, 16; coagulated,
18; products of decomposition of, 18;
theoretical views as to the constitution
of, 20; bodies related to, 22; of serum,
60; determination of, in serum, 188;
relative proportion of, in serum of blood
of various animals, 61 ; of blood-corpus-
cles, 80; of lymph, 223; of dropsical
fluids, 233; of pus serum, 239; of mus-
cle seruni, 324, 325 ; changes of proteids
of muscle, 364; in nervous tissues,
423

Protoplasm, properties of, 310

484

INDEX,

Protoplasmic foot, 319

PYE-SMITH, pernicious anaemia, 154

QUINCKE, on amount of haemoglobin in

disease, 140, 141 ; diabetic coma, 170
QUINQUATJD, gases of blood in disease, 144

BABUTEAU, blood of octopus, 133

Rachitis, blood changes in, 158; bone
changes in, 281; composition of bone in,
282; etiology of, 282; pathology of, 282

EAJEWSKY, determination of haemoglobin
in blood, 183

EANKE, acidity of muscle, 360; changes
hi proportion of water in muscle during
activity, 364 ; changes in the water and
alcohol extractives of muscle in activity,
364; changes in proteids of muscle in
activity, 364; changes in the sugar
of muscle during activity, 365 ; changes
in fat of muscle during activity, 365

EANVIEE, on red variety of voluntary
muscle, 317, 343; on capillaries of
muscle, 318; fibrin, 34, 35; glycogen in
pus, 244; iodized serum, 252

EECKLINGHAUSEN (VON), on protoplasm,
311; analysis of calcined bone, 275

Eeduced haemoglobin, 99

EEGNABD, gases in blood in disease,
144

EEGNAULT, method for determining 0
absorbed and CO2 excreted, 382

EEICHEET, oxy-haemoglobin, 84

EEIL and VON MADAI, views of muscular
contraction, 414

EEISET, method for determining O ab-
sorbed and CO2 excreted, 382

Eelapsing fever, blood in, 159; organisms
in, 159; spirillum of, 160

Eespiration, effect of exercise on gases of,
383

Eespiration of muscle, 365; G. Liebig's
method, 365—366; Valentin, 366, and
367; Matteucci, 366—367; Hermann, 369

Eespiratory capacity of blood, definition
of, 144 ; in disease, 144 ; effect of putre-
faction on, 144

Eespiratory Oxygen of haemoglobin, 91 ;
dissociation-tension of, 102

Eetiform tissue, 250

Eetina, 454 ; description of the ten layers
of, 455; variations in the structure of,
in different regions, 458 ; variations in
the distribution of rods and cones in the
retinae of different classes of animals,
459; chemical composition of, as a
•whole, 459; general chemical facts re-
lating to rods and cones of, 459;
colouring matters associated with the
rods of, 461 ; distribution of the visual
purple in, 463; retinal epithelium, 457;
chemical facts relating thereto, 468

Eheumatism articular, blood in, 158

Eheumatoid Arthritis, blood in, 158

Bhoclophane, 460

Ehodopsin, see Visual Purple

Bickets, see Eachitis

Eigor Mortis, 347, 348; chemical changes
of, 349; nature of gases liberated in,
354; cause of acid reaction in, 360

EODIEK, typhus fever, 159 ; see BECQUEEEL.

Bods of the retina, 457 ; colouring matters
associated with, 461; general chemical
facts relating to, 459; distribution of
in various classes of animals, 459

BOHEIG, method for determining 0 ab-
sorbed and C02 excreted, 383 ; soaps in
blood, 65

EOHEIG and ZUNTZ, researches on the
influence of curare on the material
exchanges of muscle, 402 ; on the
influence of division of the spinal cord
on the same, 403

EOLLETT, preparation of oxy-haemoglobin,
86; preparation of collagen, 252; pre-
paration of mucin, 257

EOSE, determination of ashes of blood,
179

EOSENTHAL, specific heat of muscle, 345

Eotation of plane of polarization, determi-
nation of, 8

'Eotatory power, specific', determination
of, 8 ; of some proteids, 12

EUPSTEIN, effect of acetone on blood, 169

EUSTITZKY, on marrow, 278

Saccharimeter, 8

SALKOWSKI, ash of plasma, 67 ; haematoi-
din, 121; Charcot's crystals, 153

SALOMON, on xanthine from proteids, 332;
leucocythaemia, 153 ; glycogen in pus,
244

Salts of Blood, see Liquor Sanguinis,
Serum and Blood-corpuscles; of lymph,
see Lymph; of dropsical effusions, 234;
pus serum, 240 ; of blood in diseases ill
general, 143 ; of muscle, 339

SANDEBS and HAMILTON, action of acetone
on blood, 169 ; lipaemia and fatty em-
bolisms in diabetic coma, 170

SANDEBSON, BUEDON, apparatus for col-
lecting plasma, 32 ; Frankland's method
for the analysis of gases, 207 — 213

Saponification, 262

Sarcine, 329 ; see Hypoxanthine

Sarcode, 310

Sarcolactic acid, 361 ; preparation, 361 ;
separation from ethidene-lactic acid,
361; properties and compounds of, 362

Sarcolemma, chemical characters of, 321

Sarcous elements, 315

SAEOKIN, changes in amount of creatin
in muscle, 364

Scalded muscle, gaseous analysis of, 351

Scarlet fever, blood in, 163

SCHAFEB, white corpuscles, 125

SCHAELING, method for determining 0 ab-
sorbed and C02 excreted, 383

INDEX.

485

SCHERER, on xanthine in muscle, on
dextrin in muscle, 336 ; inosit, 336 ;
urea in blood in disease, 143; leucocyt-
haemia, 153 ; analysis of lymph, 229 ;
analysis of dropsical liquid, 233
SCHMIDT, CAKL, sp. gr. of blood corpuscles,
79 ; composition of red corpuscles, 80 ;
mineral constituents of red corpuscles,
121, 122; analysis of blood, 127, 128;
blood of pond-mussel, 132; blood in
disease, 138 ; serum-albumin in disease,
142; corpuscles in disease, 147; blood
in cholera, 163; blood in diabetes,
172 ; analysis of lymph, 228 ; liquid in
dropsy, 232 ; cerebro- spinal liquid, 236;
osteomalacia, 281

SCHMIDT, A., on blood, 33; fibrinoplastic
substance, 37 ; source of paraglobulin,
39 ; fibrinogen, 40 ; hypotheses on coagu-
lation of blood, 46 — 48 ; fibrin ferment,
48; pure dialysed serum-albumin, 63;
salts in serum, 66; action of O-haemo-
globin on guaiacum, 102; determination
of proteids in serum, 188 ; determina-
tion of serum-globulin in serum, 189 ;
Ludwig's pump, 200 ; blood of muscle,
375, 376 — 378 ; on non-gaseous constitu-
ents of muscle blood, 381

SCHREINER, Charcot's crystals, 153

SCHULTZ, C. H., blood-cells, 72

SCHULTZE, MAX, on nature of Infusoria,
311; iodized serum, 252; structure of
retina, 455 et seq.

SCHUNK, Tyrian purple, 309

SCHUTZENBERGER, chondrin, 270; decompo-
sition of proteids, 19, 20; views as to con-
stitution of proteids, 21 ; gelatin, 254

SCHWANN, blood-cells, 73 ; white substance
of, 422

SCHWEIGGER-SEIDEL, on muscle-cells, 318

Sclerotic, 451

SCUDAMORE, coagulation of blood, 29

Scurvy, blood in, 156

SCZELKOW, on fat in muscle, 365 ; method
for analysis of gases in blood of muscle,
375 ; method for determining 0 absorbed
and CO2 excreted, 382 ; effect of exercise
on gases of respiration, 383 ; on the
amount of creatine in muscles sepa-
rated from the nerve-centres, 404

Sea-urchins, perivisceral fluid of, 134

' Sehpurpur ' or Visual Purple, 462

Semiglutin, 255

SEMMER, on preventing coagulation of
blood, 33; red granular corpuscles, 124

SENATOR, on rickets, 283

SERTOLI, phosphoric acid in serum, 68 ;
soluble salts in serum, 70

Serum of blood, 27, 57 ; modes of obtain-
ing, 57 ; physical characters of, 59 ;
effect of diet on, 59 ; specific gravity
of, 60 ; composition of, 60 ; proteids of,
60 ; presence of peptones in, 64 ; ex-
tractive matters of, 64 ; pigment in, 65 ;

salts of, 66; composition of soluble
salts in serum of ox's blood, 70 ; gases
of, 70 ; determination of water, total
solids and salts, 188 ; of proteids, 188

Serum of pus, 39 ; see Pus Serum

Serum-albumin, 16 ; preparation of, 62 ;
relative proportion of in serum of blood
of various animals, 61 ; quantitative
determination of serum-albumin, 188 ;
proportional changes of in disease, 142

Serum-casein, 61

Serum-globulin, 37; methods of obtain-
ing : viz. Schmidt's, 37 ; Hammarsten's,
37 ; properties of, 38 ; relative propor-
tion of, in serum of blood of various
animals, 61 ; determination of in serum,
189

Serum of muscle, 324 ; see Muscle-serum

SETSCHENOW, Ludwig's pump, 198

SIEBOLD, on planarian ova, 311

SIMON, blood in disease, 138

Small-pox, blood in, 163

SMITH, EDWARD, observations of, on the
elimination of nitrogen during rest and
work, 395

Soap, 262

Solar maps, 94

SOLEIL, saccharimeter, 8

Solids of blood, determination of, 177

SORBY, micro-spectroscope, 97 ; spectrum
of CO -haemoglobin, 105 ; on Gamgee's
nitrite- haemoglobin spectrum, 109; hae-
moglobin, 130, 131; bonellein, 307; pig-
men turn nigrum, 304

Specific gravity of blood, determination
of, 174 ; bottles for determination of, 175

Specific rotation, 8

SPECK, observations of, on excretion of
urea, 386

Spectra, of oxy -haemoglobin, 97 ; of hae-
moglobin, 99 ; of CO -haemoglobin, 99,
105 ; of NO-haemoglobin, 107 ; of hae-
moglobin with hydrocyanic acid, 108 ;
of methaemoglobin, 109; of haematin,
113; of haemato-porphyrin, 117; of
haemochromogen, 111, 118; of haema-
toidin, 121; of blood of Planorbis, 130;
of chlorocruorin, 131; of oxy-chlorocru-

. orin, 131; of turacin, 305; of chlorophane,
xanthophane and rhodophane, 461 ; of
rhodopsin or visual purple and of
visual yellow, 465 ; of lipochrin, 468

Spectroscope, 93 ; examination of blood
by, 91 ; with scale indicating wave-
lengths, 94

Spectrum maps> 94

Spermaceti, 264

SPIEGELBERG, volume of blood in bodies
of animals, 216

SPIRO, on sarcolactic acid in blood of
muscle, 381

Splenic fever (of cattle), blood in, 161 ;
Bacillus anthracis in, 161

Splenic leukaemia, 152

48G

INDKX.

Spongin, 302 ; preparation, analysis and

reactions of, 302
STADELER, method for preparing creatine,

327
STAHL, views on muscular motion, 409 ;

reference to his views on muscular

motion, 410
Stearin, 262; nature of, 262; preparation

of, 262

STEELL, GRAHAME, diabetic coma, 170
STEINEB, on curve of muscular contraction,

342; on heat of contraction, 346
STENO or STENSON, observations on the

influence of stoppage of the circulation

through muscle, 406
STEWART, GRAINGER, pernicious anaemia,

156
STINTZING, gaseous analysis of muscle,

357—358

STIRLING on tetanus of pale muscles, 343
STOKES, haemoglobin, 100
STRASSBURG, C02 in lymph, 226; gas in

dropsical liquids, 234
STRECKER, choline, 426, 435; compounds

of lecithin, 432 ; distearyl-lecithin, 437
STRICKER, on contractility of capillary

walls, 311; lactic acid in muscle, 359
Stroma of coloured blood corpuscles, 74,

80 ; method of obtaining for microscopic

examination, 80 ; separation of pro-

teids of stroma, 81 ; peculiarities of, 81
Succinic Acid in hydrocele fluid, 235 ; in

uraemia,
Sugar, in the blood, 65 ; variations of, in

blood in disease, 143 ; determination of

in blood, 194; in lymph, 224; in drop-
sical effusions, 234; in cerebro-spinal

fluid, 236; in pus, 240; fermentable, in

muscle, 336
Synovia, 229; characters of, composition

of, and analysis of, 230
Syntonin, 324; characters of solutions of,

324; preparation of, 324

Tapetum, 458

Taurine in muscle, 333

TEICHMANN, haemin crystals, 116

Tetanus of muscle, 343; acidification of
muscles in, 360; sarcolactic acid in, 381

THACKRAH, coagulation of blood, 29

THE'NARD, blood in disease, 138

Thrombosis, 54

THUDICHUM, protagon, 426 ; glycerin-phos-
phoric acid, 434; phosphorized prin-
ciples in brain, 437—439; kephalines,
myelines, and lecithines, 437; cerebrins,
442 ; phrenosine arid kerasine, 442

TIEGEL, researches on muscular contraction
following powerful direct stimuli, 404

TILANUS, analysis of cow's horn, 298

TOMMASI-CRUDELI, see Klebs

Tooth, 289 — 294; obvious structure of,
289 ; comparative analyses of, 293 ; com-
position of fossil teeth, 294

Transudations, 220

TRAUBE, views on nature of muscular
contraction, 416

Triacetin, 264

Trinitroinosit, 338

Triolein, 263

Tripalmitin, 262

TRIPIEB, LEON, rickets, 283

Tristearin, 262

TSCHIRIEW, gases in lymph, 225

Tunicin (Animal Cellulose), 302, 303; oc-
currence, 302 ; preparation and reactions
of, 303

Turacin, 304 ; occurrence and mode of
separation of, 304; properties and spec-
trum of, 304 ; composition of, 305 ;
analysis of, 305

TURNER, cerebro-spinal fluid, 231

Typhoid fever, blood in, 159

Typhus fever, blood in, 159

Tyrian purple, 307 ; source and properties
of, 309

UNGER on xanthine, 330

Urea, in the blood, 65 ; in blood of diseases
in general, 143 ; in gout, 157; in cholera,
blood of, 164; in Blight's disease, blood
of, 173; determination of, in blood,
190; Picard's method, 190; Grehant's,
191; by sodium hypobromite, 191; in
lymph and chyle, 224 ; in dropsical ef-
fusions, 234; in pus, 240; in muscles,
333

Uric Acid, in the blood, 65; in blood of
disease, 143; in gout, 157; determina-
tion of in blood, 193 ; in dropsical effu-
sions, 234; in muscle, 333; in brain, 444

Urinary secretion as influenced by mus-
cular exercise, 385 ; statements of early
observers concerning, 386 ; experiments
of Voit, 387 ; of Fick and Wislicenus,
388 ; of Parkes, 392 ; of Edward Smith,
395 ; of Flint and Pavy, 395 ; of North,
398

VALENTIN, respiration of muscle, 366—367;
apparatus, 368, 369

VIERORDT AND WELCKER, method for enu-
meration of blood corpuscles, 74

VIRCHOW, haematoidin, 120; pigment in
blood in intermittent fever, 163

Visual Purple or Khodopsin, 461; dis-
tribution of in retina, 463 ; method of
separation of from retina, 464; spec-
trum of, 465 ; effects of light of different
wave-lengths upon, 465; influence of
temperature upon, 466; action of various
chemical agents upon, 466; action of
light upon, 469; regeneration of, 469;
vision without, 470

Vitellin, 17

Vitreous body, 454

VOHL, on inosit or phaseomannite, 336,
338

INDEX.

487

VOIT, on quantity of creatine in muscle,
328 ; on creatinine, 329 ; on creatin,
365 ; method for determining 0 absorbed
and CO 2 excreted, 383; researches on
excretion of urea during work, 387 ;
views on the transformation of energy
in muscle, 416

VOLKMANN, water in bone, 273

Voluntary muscle, 313 — 315 ; subdivision
into pale and red, 317; red and pale,
343 ; see Muscle

VULPIAN, Charcot's crystals, 153

WALCHLI on mucin, 259

WALKER, E., on absolute force in rigor
mortis, 348

WARREN, J., lactic acid in muscle, 360

'Washed blood-clot,' 45

Water, amount of, in the blood, 139 ;
variations in disease, 139; in blood of
cholera, 163; density of, at various
temperatures, 176; determination of
amount in blood, 177

Water, in muscle, 338; changes of pro-
portion of in muscle, 364; in nerve
substance, 445, 446

Wave-lengths, spectral bands referred to,
95 ; scale of, 95

WEBER on absolute force of contraction,
344

WEIDEL on carnine, 332

WEIGERT, relapsing fever, 160

WEISBACH, water in brain, 445

WEISKE, mineral matters of bone, 276,
277; rickets, 283; xanthine, 330

WELCKER, red corpuscles, 71; enumera-
tion of blood corpuscles, 74, 78; total
quantity of blood in body, 215; volume
of blood in bodies of animals, 216

WELLS, red corpuscles, 72

WESTON, account of experiments made

upon, by Flint and Pavy, 395 et seq.
WEYL, temperature of coagulation of

proteids, 16; serum -globulin, 39
White substance of Schwann, 422
WHYTT, doctrine of a 'Sentient Principle,'

411

WILES, pernicious anaemia, 154
WILSON, fluorine in bone, 276
WINOGRADOFF on serum-albumin, 63
WOHLER on xanthine, 330
Work, maximum, of muscle, 345
WORM-MILLER, dissociation tension of 0

in O-haemoglobin, 102; corpuscles in

healthy blood, 147 ; nuclein, 243
WURTZ, sugar in lymph, 224

Xanthine, 330, 331; rare constituent of
iirinary calculi, 330; source of, 330;
preparation by Neubauer's method, 330 ;
properties of, 331; reactions and rela-
tions of, 331 ; proportion in muscle, 331 ;
artificial production of xanthine from
proteids, 332 ; in the blood, 65

Xanthophane, 460

Xanthoproteic reaction, 14

ZAHN, thrombosis, 54, 55

ZAWILSKI, on lymph, 223

ZEISS, C., spectroscope, 94 — 96; micro-
spectroscope, 96

ZENKER, Charcot's crystals, 153

Zinc sarcolactate, 362 ; lactate, 362

Zooid, 74

ZUNTZ, reaction of blood, 26; CO-haemo-
globin, 106 ; method for determining O
absorbed and C02 excreted, 383; see
also KORHIG and ZUNTZ

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