MANUAL OF VITAL FUNCTION
TESTING METHODS AND
THEIR INTERPRETATION
WILFRED M. BARTON
!TY OF CALIFORNIA
Manual of Vital Function
Testing Methods and
Their Interpretation
BY
WILFRED M. BARTON, M.D.
A ssociate Professor of Medicine, Medical Department, Georgetown
University, Attending Physician to Georgetown
University Hospital andW asking-
ton A sylum Hospital
BOSTON: RICHARD G. BADGER
TORONTO: THE COPP CLARK CO., LIMITED
Copyright, 1916, by Richard G. Badger
All Rights Reserved
THE GORHAM PRESS, BOSTON, U. S. A.
TO DE. GEORGE M. KOBER, DEAN OF THE
GEORGETOWN MEDICAL SCHOOL, THIS LITTLE
BOOK IS DEDICATED, AS A MODEST TOKEN OF
APPRECIATION OF HIS UNSELFISH DEVOTION TO
OUR SCHOOL AND HOSPITAL
PREFACE
THERE should always be some valid reason for writing
a new book, especially a medical book. My reason for
writing, or perhaps to express it more accurately, for
compiling this work, is, that the information that it
contains is scattered quite broadly thro a wide and
extensive medical literature, that may not be readily
accessible to all, and which has never, so far as I know,
been collected together in book form.
Inasmuch as every progressive physician and sur-
geon at the present day is making more or less frequent
use of the different functional tests, to determine the
efficiency of vital organs, it occurred to me that to
collect them all in one volume, together with the neces-
sary data whereby they might be intelligibly inter-
preted, might prove to be useful, particularly to the
busy practitioner.
The following work, which is an effort to carry out
that idea, includes only the tests for so called vital
functions, namely those applied to the liver, kidneys,
heart, pancreas, and ductless glands. To go beyond
this and take up the functional tests of all the organs,
such as the eye, ear, nervous system, etc., would be to
exceed the legitimate field of true function testing and
to encroach upon the well-trodden fields of general
semiology and diagnosis.
I hope the book may prove to be useful and con-
venient to all who are interested in this fascinating and
ever-developing field of clinical pathology.
5
6 Preface
It is a matter of which American physicians may
well be proud that the most substantial and brilliant
progress in the development of tests of organic func-
tion, particularly in reference to the kidneys and liver,
has been brought about by the assiduous efforts of some
of their own fellow countrymen.
In the chapter on heart I have received valued assist-
ance from my friend and colleague, Dr. Thos. S. Lee.
CONTENTS
TESTS OF LIVER FUNCTION
PAGE
GENERAL CONSIDERATIONS 13
FUNCTIONAL TESTS TO DISCOVER DISTURBANCES OF THE GLYCO-
GENIC FUNCTION OF THE LIVER 14
The Cane Sugar Test 15
The Glucose Test 16
The Levulose Test 16
The Galactose Test 18
Conclusions Concerning the Carbohydrate or Sugar
Tests 18
FUNCTIONAL TESTS TO DETERMINE DISTURBANCE OF THE UREA-
GENETIC FUNCTION OF THE LIVER 20
Urea Elimination and Nitrogen Coefficient as Criteria of
Liver Function 22
Quantitative Urea Estimation in Urine (Marshall's
Method} 24
Total Nitrogen Estimation in Urine (Kjeldahl's
Method) 28
Augmentation of Urinary Ammonia as an Index of Urea-
genetic Liver Function 30
Estimation of Ammonia Nitrogen in Urine (The
Formalin Method) 31
Experimental Provocative Ammoniuria 32
Aminoaciduria as a Criterion of Liver Function .... 33
Experimental Provocative Aminoaciduria .... 33
Estimation of Residual Nitrogen in Blood Serum as an
Index of Hepatic Function 34
Summary of the Value of Ureagenetic Tests of Liver
Function 35
FUNCTIONAL TESTS TO DETERMINE DISTURBANCE OF THE ANTI-
TOXIC FUNCTION OF THE LIVER 36
Methylene Blue Test of Toxopexic Function of the Liver
(Chauffard-Castaigne Test) 36
Roche's modification of Methylene Blue Test ... 37
Indicanuria, Spontaneous and Provocative as Means of
Testing Integrity of Hepatic Fixation 38
Tests for Urinary Indican 38
7
8 Contents
PAGE
FUNCTIONAL TESTS TO DETERMINE DISTURBANCE OF THE SAN-
GUINOPOIETIC FUNCTION OF THE LIVER 40
Estimation of Blood Coagulation Time as an Index of Liver
Function. Wright's Method of Fixing Coagulation Time 41
The Fibrinogen Test of Whipple and Horwitz .... 42
Estimation of Fibrinolysis Time as an Index of Liver Func-
tion (Goodpasture' s Test) 43
Estimation of Lipase in the Blood as an Index of Liver
Function (Whipple's Test) 44
Lowenhart's Methods of Lipase Estimation ... 45
Ghedini's Test .... 46
Application of Abderhalden's Method to Estimation of
Sanguinopoietic Liver Function 46
FUNCTIONAL TESTS TO DETERMINE DISTURBANCE OF THE EXOCRIN-
ous OR BILIARY FUNCTION OF THE LIVER 47
Tests for Urobilinogen, Urobilin and Bilirubin in the Urine.
Interpretation of Results with Reference to Hepatic
Function. The Urobilinogen Test (Ehrlich's Test) . . 51
Tests for Urobilin 54
Tests to Determine the Global capacity of the Liver to
Eliminate Foreign Substances 55
Phenoltetrachlorphthalein Test of Liver Function (Rown-
tree, Horwitz and Bloomfield Test) 55
TESTS OF KIDNEY FUNCTION
GENERAL CONSIDERATIONS 64
URINALYSIS AS A CRITERION OF RENAL FUNCTION .... 70
Estimation of Urinary Water. Experimental Polyuria
(Albarran's Method) 72
The Water Tests (Straus-Grunwald Method) 75
The Diuretic Tests (Pharmacological) 75
Estimation of Sodium Chloride as an Index of Renal Func-
tion 76
The Sodium Chloride Test. Test of Alimentary
Chloruria 79
Sodium Chloride Estimation 79
Estimation of Urinary Nitrogen as an Index of Renal Func-
tion 80
Diminished and Delayed Excretion of Urea .... 82
Forced Urea Elimination. Provocative Urea Test of
McKaskey 83
Estimation of Urinary Coloring Matter as an Index of
Renal Function 84
Estimation of Urinary Diastase as an Index of Renal Func-
tion . 85
Contents 9
PAGE
STUDY OF THE PHYSICAL AND BIOLOGICAL CHARACTERISTICS OF
THE URINE AS CRITERIA OF KIDNEY FUNCTION .... 86
Estimation of Freezing Point of Urine (Cryoscopy) as
Index of Renal Function (v. Koranyi's Test} .... 86
Electrical conductivity of the Urine as Index of Renal
Function 89
Estimation of Urinary Toxicity as Index of Renal Function 89
STUDIES OF THE BLOOD AS CRITERIA OF RENAL FUNCTION . . 90
Urea and Incoagulable or Rest Nitrogen in the Blood as
Indexes of Renal Function 90
Marshall's Method for Determination of Urea in Blood . 94
Estimation of Incoagulable Nitrogen. Morris' Modifica-
tion of Hohlveg-Meyer Method; Folin and Denis Method 99
Estimation of Blood Coagulation Time as Index of Renal
Function 104
Cryoscopy of Blood as Index of Renal Function .... 104
STUDIES OF THE ELIMINATION OF FOREIGN SUBSTANCES, BY THE
KIDNEY AS CRITERIA OF RENAL FUNCTION 105
1. Miscellaneous Substances:
The Potassium Iodide Test 105
The Phloridzin Test 107
The Hippuric Acid Test 108
The Lactose Test 108
2. Elimination of Dyes — Urinary Chromoscopy. . . . 110
The Methylene Blue Test Ill
The Indigo Carmine Test 114
The Phenolsulphonephthalein Test of Rowntree and
Geraghty 115
GENERAL SUMMARY OF RENAL FUNCTION TESTS 128
SELECTION AND PRACTICABILITY OF RENAL FUNCTION TESTS . . 138
TESTS OF PANCREATIC FUNCTION
GENERAL CONSIDERATIONS 142
TESTS OF PANCREATIC FUNCTION WHICH CONCERN THE EXTERNAL
OR DIGESTIVE ACTIVITY OF THE ORGAN 144
Proteid Digestion Tests: Estimation of Undigested Protein
in Stool as Means of Determining Pancreatic Hypofunc-
tion 146
Schmidt's Cell Nuclei Test 148
Sahli's Glutoid Capsule Test 148
FAT DIGESTION TESTS. DETERMINATION OF EXCESS OF FAT IN
THE STOOLS 150
10 Contents
PAGE
STARCH DIGESTION TESTS 153
IDENTIFICATION OF FERMENTS IN EXCRETA AS EVIDENCE OF PAN-
CREATIC FUNCTION 153
Demonstration of Trypsin in Stools 154
Demonstration of Trypsin in Stomach Contents . . . 156
Demonstration of Diastase in Feces 158
Demonstration of Lipase in Stools 160
TESTS FOR PANCREATIC FUNCTION WHICH CONCERN THE INTERNAL
OR METABOLIC FUNCTION OF THE ORGAN 161
The Cammidge Reaction 162
Loewi's Pupillary Test 165
Spontaneous and Provocative Glycosuria 166
GENERAL CONCLUSIONS CONCERNING PANCREATIC INSUFFICIENCY
TESTS 167
TESTS OF HEART FUNCTION
GENERAL CONSIDERATIONS 168
Reaction and Muscular Exertion as Basis for Estimating
Cardiac Function 171
The Staircase Case (Selig's Test) 173
The Ergometer Test (Graupner's Test) 175
Mendelsohn's Test 179
Katzenstein's Test 181
Herz's Self-Checking Test 183
Gymnastic Resistance Test 184
The Russian Test 185
The Venous Pressure Test (Schott's Test) .... 185
Cardiac Reflex Estimations in Determining Heart Function 187
Sodium Chloride Elimination and Cardiac Function . . 188
Modern Clinical and Instrumental Methods in Cardio-
Pathology; Their Applicability to Estimation of Heart
Function 188
Sphygmomanometry. Work- Velocity Ratio . . . 188
Cardiac Efficiency Factor of Tigerstedt 193
Cardiac Strength, Cardiac Weakness Ratio . . . .194
Cardiac Overload Factor of Stone 197
Reontgenoscopy and Cardiac Function 199
Sphygmography and Cardiac Function 199
GENERAL CONCLUSIONS AS TO TESTS OF CARDIAC FUNCTION . . 202
TESTS OF DUCTLESS GLAND FUNCTION
GENERAL CONSIDERATIONS 240
The Thyroid Gland 270
Contents
HYPERFUNCTION OF THYROID GLAND
Hypophysis-Extract Test of Claude, Baudouin and Porak
Adrenalin-Mydriasis Test of Loewi
Experimental Hyperthyroidism Test
Aceto-nitril Test of Reid Hunt
Metabolic Tests of Hyperthyroidism
Complement-Deviation Test of Hyperthyroidism
Abderhalden Test in Hyperthyroidism
HYPOFUNCTION OF THYROID GLAND
Therapeutic Test of Hypof unction
The Parathyroid Glands
The Thyrnus Gland
The Suprarenal Glands
HYPOFTTNCTION OF SUPRARENAL GLANDS
Sugar Tolerance and Hypoadrenal Function ....
HYPERFUNCTION OF SUPRARENAL GLANDS
Adrenalinemia and Hyperadrenalism
Adrenalin Glycosuria and Hyperfunction ....
Complement-fixation in Suprarenal Disease ....
The Hypophysis
STATES OF HYPERPITUITARISM 246
Increased Gas Exchange and Hyperpituitarism .... 247
Glycosuria and Hyperpituitarism 248
STATES OF HYPOPITUITARISM 249
INDEX . 251
MANUAL OF VITAL FUNCTION
TESTING METHODS
CHAPTER I
TESTS OF LIVER FUNCTION
GENERAL CONSIDERATIONS
THE liver cell represents the whole organ in minia-
ture. If we possessed an adequate knowledge of all
the functions of this cell, we would understand com-
pletely the functions of the organ.
The functions of the liver cell are numerous and each
individual cell performs its quantitative quota of all
these functions. The liver is provided with two evacu-
ating channels, one internal, by way of the blood (he-
patic vein), the other external, by way of the biliary
passages. It receives blood by way of the portal vein
from the digestive tract and its related organs, and dis-
charges into the duodenum its more or less complex
excretion.
Regarded as a center of elaboration, the liver cell
has several important functions, chief of which are:
1. ureagenetic, 2. glycogenetic, 3. lipasic, 4. antitoxic or
cytopexic, 5. sanguinopoietic, 6. thermic, 7. ferric.
The above mentioned functions are spoken of as en-
docrinous. The liver has, however, an exocrinous func-
tion which by some is regarded as the most character-
istic, namely, the excretion of bile by way of the biliary
passages.
Tests of hepatic function are directed towards the de-
termination of the integrity of one or the other of
these phases of activity.
13
14 Manual of Vital Function Testing Methods
There is no organ whose functional and clinical ex-
amination is fraught with more difficulties than the liver.
Slight functional disturbances of the organ are attended
by very uncertain symptomatology. When severe or-
ganic lesions exist, such as acute yellow atrophy, cirrho-
sis, abscess, cancer, etc., modifications in the size of
the liver, as well as its consistence, together with
symptoms of portal hypertension (ascites) and of bili-
ary obstruction (icterus), offer a combination of objec-
tive evidence which makes the diagnosis usually clear.
But there is a longer or shorter period in all these
diseases, usually in the earlier stages during which a
condition of hepatic insufficiency (hypohepatism) and
more rarely, hepatic hyper function (hyperhepatism)
exists, which might be recognized by appropriate tests.
The trend of modern investigation is in the direction
of the development of practical functional tests of
sufficient simplicity to enable the clinician to judge the
functional capacity of the liver, before gross organic
lesions or indubitable symptoms have appeared.
Tests for liver functional capacity may be profitably
considered under five headings.1 The first four are
endocrinous, the fifth exocrinous. They are as follows :
I. Disturbance of the glycogenic function. II. Dis-
turbance of the urcagenetic function. III. Disturbance
of the antitoxic function. IV. Disturbance of the
hemapoietic function. V. Disturbance of the biliary
function.
I. FUNCTIONAL TESTS TO DETERMINE DISTURBANCES OF
THE GLYCOGENIC FUNCTION OF THE LIVER
Normally the liver cells retain in the form of glycogcn
almost all the glucose brought to them from the ali-
1 See Les Precedes actuel d'etude de 1'insuffisance hepatique.
Gaz. d. hop. Par. 1914, no. 25, p. 408 (Brule, Garban). To this
article is appended an extensive bibliography.
Tests of Liver Function 15
mentary tract. Under certain conditions of liver in-
sufficiency, glucose is not fixed by the cells, and passes
immediately into the blood, producing a hyperglycemia,
from whence it is excreted in the urine. This fact has
been utilized as a basis for testing the glycogenetic
integrity of the liver cell.
The name of Claude Bernard is closely linked with
the history of the physiology of hepatic glycogenesis.
Soon after the discovery by Claude Bernard of the role
of the liver in carbohydrate metabolism, the use of
sugars as tests for hepatic function began.
In applying the sugar test, different varieties of
sugars have been employed, particularly saccharose
(cane sugar), glucose, levulose, and galactose. The
first sugar employed for this purpose was saccharose
(cane sugar).
The sugar tests of hepatic function are four in
number, as follows: 1. The Cane Sugar test. 2. The
Glucose test. 3. The Levulose test. 4. The Galactose
test.
1. The Cane Sugar Test. Colrat, Lepme Test
150 to 200 grams of cane sugar syrup are adminis-
tered to the subject in the morning while fasting. The
urine is collected every hour or two and examined for
sugar with Fehling's solution or other means. The
presence of glycosuria renders the test positive.
The cane sugar test was considered for a long time
the best criterion of liver insufficiency. The clinical
results of the test have, however, been contradictory.
A rather weighty theoretical objection is the fact that
cane sugar must be converted into glucose in the ali-
mentary tract before it can be utilized by the organism,
and the power of the intestinal juices to produce this
16 Manual of Vital Function Testing Methods
conversion is in each case an unknown quantity and
therefore a source of error.
#. The Glucose Test
The patient takes in the morning on an empty stom-
ach, 150 grams of pure dextrin-free glucose dissolved
in 300 c. c. of water. The ingestion of this amount
should not take over a quarter of an hour.
The urine is then collected every hour or two for ten
hours in separate vessels and tested for sugar, the
patient remaining on a milk diet during the time re-
quired by the test.
Castaigne has advised the following details with a
view of perfecting the glucose test. For several days
prior to the performance of the test, the subject should
be kept on a certain known quantity of carbohydrate.
The renal permeability should be investigated and the
possibility of spontaneous glycosuria especially after
meals eliminated.
The results of the glucose test have been rather con-
flicting and some investigators have appeared to find
glycosuria following the test in apparently healthy sub-
jects and its absence in certain cases of hepatic cirrhosis
where the liver parenchyma would have been acknowl-
edged on general clinical grounds to have been damaged.
3. The Levulose Test. Strauss Test
This test was introduced by Strauss 2 in 1901 as a
substitute for the saccharose and glucose tests.
' Berl. klin. Wchnschr., 1898, XXXV, p. 398, and 1899, XXXVI,
p. 1.59; Deutsch. med. Wchnsch., 1901, XXVII, p. 756, also 1903,
XXXIX, p. 1T80.
Tests of Liver Function 17
To apply the test 100 grams of levulose are given in
the morning on an empty stomach and the urine evacu-
ated every four hours thereafter for a day and examined
for sugar by the fermentation test or polariscope.
Owing to the high price of levulose, honey, which
contains a large percentage of it, has been advised as a
substitute.
A normal person should tolerate 100 grams of levu-
lose without levulosuria.
The rationale of this test was founded on the experi-
mental work of Sachs,3 who found that frogs whose
livers had been removed had a lower tolerance for levu-
lose than intact controls. With dextrose and galactose
this was not the case. It was contended therefore that
there is no mechanism besides the liver capable of
handling levulose, while there is such an extra hepatic
mechanism in the case of glucose.
Immediately after its introduction, this test came
Into pretty general use and was commended by Ferra-
nini 4 v. Halasz,5 Bruining,6 and others, and was con-
demned by Landsberg,7 Churchman,8 and others. Much
was expected from the levulose test because as above
stated it was believed that the liver alone is concerned
in levulose metabolism. However this may be, experi-
ence has apparently failed to substantiate the hopes
which the test originally inspired and it is not now be-
lieved that the levulose test is essentially superior to
other sugar tests of hepatic function.
"Zeitschr. f. klin. Med., 1899, XXXVIII., p. 87.
*Zeitschr. f. inn. Med., 1902, XXIII, p. 921.
8Wien. klin. Wchnschr., 1908, XXI, p. 44.
"Berl. klin. Wchnschr., 1902, XXXIX, p. 587.
'Deutsch. med. Wchnschr., 1903, XXIX, p. 563.
8 Johns Hopk. Hosp. Bull., 1912, XXIII, p. 10.
18 Manual of Vital Function Testing Methods
4- The Galactose Test. Bauer's Test
Forty grams of milk sugar dissolved in 400-500 c.c.
of tea are taken in the morning on an empty stomach.
The urine is passed every four or five hours thereafter
and examined for sugar.
In icterus gravis and catarrhal jaundice this test has
been reported as giving fairly constant results.
Bauer 9 considered the galactose test especially
adapted to determining the condition of liver function in
catarrhal jaundice. The amount of sugar recovered
in the urine after the galactose test was found to be
greater in catarrhal jaundice than in obstructive jaun-
dice, consequently it was supposed that the test would
be of importance in differential diagnosis between the
two conditions. This opinion was upheld and confirmed
by Bondi and Kb'nig,10 Riess and Jehn,11 and Hi-
rose.12
Outside of catarrhal jaundice the results were pro-
nounced inconstant by Falk and Saxl,13 v. Frey,14 and
others.
Conclusions concerning the Sugar Tests. — It may be
said that the investigation of hepatic insufficiency by
any or all of the sugar tests, is to be regarded merely as
supplementary or complementary to other means of
investigation, since the results of these tests alone are
not conclusive. Nevertheless the results which may be
obtained by their help when associated or corelated with
"Wien. med. Wchnschr., 1906, LVI, p. 2557.
10Wien. med. Wchnschr., 1910, LX, p. 2617.
"Deutsch. med. Wchnschr., 1912, XXXVIII.
12Deutsch. Arch. f. klin. Med., 1912, CVIII, p. 187.
13Ztsch. f. klin. Med., 1911, LXXIII, p. 131, 325.
"Ztschr. f. klin. Med., 1911, LXXII, p. 383.
Tests of Liver Function 19
those obtained by other methods are of sufficient value
to justify their retention in clinical medicine.
It is now understood that the mechanism whereby
alimentary glycosuria is produced is more complex than
was formerly supposed, and that the liver is not the
only organ involved in the process. Other tissues are
now known to be concerned in glycofixation and mobil-
ization. Furthermore, the individual coefficient of sugar
utilization has been found to vary within quite wide
limits, 50-350 grams of levulose for example. The co-
efficient varies also in the same individual for the dif-
ferent sugars so that in reporting results of sugar tests
it is deemed expedient to specify the particular kind of
sugar used.
The unknown factors of intestinal absorption and
renal permeability complicate all sugar tests.
The sugar tests have been found positive, especially
in severe bivenous cirrhosis, in icterus gravis and in
cholelithiasis. They are, however, of no prognostic
value.
A recent comprehensive study of the applicability of
carbohydrates as tests for hepatic functional activity
has been made by Bloomfield and Horwitz.15 These
authors call attention very properly to the factors
which tend to render the sugar tests for hepatic func-
tion unreliable. The great theoretical stumbling block
in the way of accepting the finding of the carbohydrate
tests is the fact that extra-hepatic factors of consider-
able importance are concerned in the sugar regulating
metabolism. Some of the glands of internal secretion
take part in this. Certain lesions of the hypophysis
may cause glycogenolysis and glycosuria, while with
other hypophyseal lesions the sugar tolerance is in-
15 Johns Hopk. Hosp. Bull., 1913, XXIV, p. 375:— a good bibli-
ography is appended to this article.
20 Manual of Vital Function Testing Methods
creased.
The internal secretion of the pancreas is generally
considered to exert an inhibitory effect on the mobiliza-
tion of glycogen by the liver. The suprarenals have an
accelerating effect on the same mechanism. The thy-
roid gland also takes part in sugar metabolism, the
exact nature of which is unknown. Both the autonomic
and sympathetic nerves likewise affect the mobilization
of glycogen. These last facts, of course, tend to render
the sugar tests less definite and satisfactory.
From a practical standpoint it cannot be denied that
the sugar tests have certain disadvantages. It is diffi-
cult for patients to ingest the large quantities of sugars
required without the occurrence in certain cases of
nausea, vomiting and diarrhoea. Faulty absorption,
intestinal fermentation, portal obstruction with col-
lateral circulation, sugar retention due to nephritis,
inconstancies in the diet also combine to render the
results inaccurate.
It will remain for the future, however, to determine
whether the sugar tests of hepatic insufficiency can be
developed or modified in such a way that the rather
numerous objections with which impartial observers are
agreed the tests are encumbered, may be eliminated.
In such an event these tests, which from an historical
standpoint are so interesting, may take a definite place,
even though subsidiary, in that important group of
tests by which one seeks to obtain an insight into the
functional integrity of the liver.
II. FUNCTIONAL TESTS TO DETERMINE DISTURBANCES OF
THE UREAGENETIC FUNCTION OF THE LIVER
All the tests for hepatic function which deal with
the ureagenctic activity of the liver are concerned with
Tests of Liver Function 21
the question of nitrogen metabolism. The liver plays a
large and important part in this metabolism; perhaps
not so exclusive a role, however, as was formerly sup-
posed.
The ureagenetic function tests are, for the most part,
merely studies of nitrogen metabolism. They consist
mostly of estimations of the amount of nitrogen elimi-
nated in the urine with accurate partition of this nitro-
gen into different groups, particularly urea and am-
monia. The relation between the amount of nitrogen
eliminated in these two forms when compared with the
total nitrogen eliminated will afford valuable criteria
for estimating the ureagenetic functional capacity of
the liver.
The practical estimation of ureagenetic functional
capacity of the liver involves much more complex series
of chemical processes than were found to be involved in
the investigation of the carbohydrate function by means
of the sugar tests. Inasmuch as the first criterion of
normal liver function from the standpoint of nitrogen
metabolism involves the relation between the amount of
urea excreted in the urine and the amount of total
nitrogen excreted therein, the investigator must make
two separate chemical analyses. First the exact amount
of urea excreted must be estimated, and second, the
exact amount of total nitrogen eliminated. In discuss-
ing these analyses, no attempt will be made to give any
data beyond the description of such methods which, on
account of their comparative simplicity and established
accuracy, have become firmly established in the clinic.
The estimation of total nitrogen in the urine by the
standard method — that of Kjeldahl — requires a good
deal of time and some technical skill. But it is a method
which can be easily performed in any well-equipped
laboratory attached to a hospital, and should be car-
22 Manual of Vital Function Testing Methods
ried out by a competent person. A rapid and accurate
method of quantitative estimation of urea has been
lately devised — that of Marshall — and this method has
already become the standard one in the clinic.
The estimation of urinary ammonia, or ammonia
nitrogen, is another step required in the study of the
efficiency of the liver from the standpoint of nitrogen
metabolism. Here also it is best for the clinician to
familiarize himself with one method, preferably that of
Folin, the details of which are further on discussed.
The actual administration of ammonium-bearing or
amino-acid-containing substances to the patient, with
the subsequent examination of the urine to determine
the capacity of the liver to convert these substances into
urea, is another method of clinical investigation of the
liver function. Finally it is conceded that the estima-
tion of rest or residual nitrogen in the blood serum may
be utilized to determine whether or not the liver cell is
capable of producing an adequate urea synthesis of
nitrogen products in the body.
To recapitulate, we may tabulate the different meth-
ods of testing the ureagenetic functional power of the
liver under the following heads and in this order they
will be discussed: 1. Urea Elimination and Nitrogen
Coefficient as Criteria of Ureagenetic Liver Function ;
2. Augmentation of Urinary Ammonia as an Index of
Ureagenetic Liver Function ; 3. Amino-aciduria as a
Criterion of Ureagenetic Liver Function ; 4. Estima-
tion of Residual Nitrogen in the Blood Serum as an
Index of Ureagenetic Liver Function.
1. Urea Elimination and Nitrogen Coefficient as
Criteria of Liver Function
Urea Elimination,- — The liver has long been regarded
as the chief source of urea, which substance is supposed
Tests of Liver Function 23
to be formed from ammonium salts, amino acids and
products of nitrogenous catabolism.
The urea synthesis is supposed to be the work of the
liver cell governed by ferments which it secretes.
It has long been recognized that in certain diseases of
the liver the percentage of urea eliminated in the urine
is lowered. Therefore diminution of urea excretion
may sometimes indicate hepatic insufficiency and an
estimation of the quantity of urea eliminated is there-
fore an available factor in testing the ureagenetic func-
tion of the liver cell.
The quantity of urea eliminated in the urine by a
healthy adult in 24 hours is 25 to 30 grams. In cases
of hepatic insufficiency this quantity may fall to 10, 5,
3 or .5 grams or even 0 in icterus gravis.
But before one can attribute a diminution of urea
elimination to functional disorder of the liver, certain
other factors of great importance must be taken into
consideration. One of these is the functional capacity
of the kidney. It is well understood that urea retention
in the blood may be due to defect of kidney permeability.
Another important factor is the amount of proteid
intake. Before attempting to draw any conclusions
with respect to the relation of urea elimination to the
ureagenetic functional capacity of the liver it will al-
ways be necessary that the individual to be tested shall
be placed for a sufficient length of time upon a fixed
ration in which the amount of proteid is known and
invariable.
The Nitrogen Coefficient. — The relation of urea ni-
trogen to the total nitrogen excreted in the urine is
known as the coefficient of nitrogen elimination. This
coefficient may be important since under certain circum-
stances its diminution may constitute a valuable sign
of ureagenetic hepatic insufficiency.
24 Manual of Vital Function Testing Methods
The nitrogen coefficient is usually expressed by the
following fraction:
N. urea
N. total
the arithmetical relation in other words between the
amount of nitrogen eliminated in the urine as urea and
the total quantity of nitrogen eliminated. This co-
efficient will diminish in proportion to the diminution
of urea nitrogen.
According to some authors a diminution of the ni-
trogen coefficient is absolutely constant in hepatic in-
sufficiency.
The normal figures of the nitrogen coefficient vary
from 85% to 95%. In icterus gravis it has been found
reduced to 40 ; likewise in phosphorus poisoning.
But just as in estimating the percentage of urea in
the urine and considering the same as an index of he-
patic function, so also in determining the coefficient of
nitrogen elimination for the same purpose the patient
must be placed on a fixed and invariable proteid regi-
men, though the absolute quantity of proteid taken is
negligible.
It must also be previously known that there is no
deficiency in renal permeability. To determine the co-
efficient of nitrogen elimination, two operations must be
performed. First, a quantitative estimation of urea
must be made, from which the calculation of urea nitro-
gen may readily be accomplished. Secondly, the total
nitrogen eliminated in the urine must be estimated and
this part of the operation is, unfortunately, somewhat
difficult and time-consuming. The simplest known
means for performing these operations will be given.
Quantitative Estimation of Urea in Urine. — Several
Tests of Liver Function 25
methods are in use for determining quantitatively the
amount of urea in urine. One of the most frequently
used is the hypobromite method, using the ureometer of
Doremus. In this method nitrogen is set free by sodium
hypobromite and measured in the apparatus. The re-
sults obtained by this method are extremely inaccurate
and it is not used, therefore, where absolute results are
required.
Three other methods which are much more depend-
able have been in use for some years in the laboratories.
These are: 1. The Morner-Sjoqvist;16 2. Folin's
method ;17 and 3. Schondorff's method.18 The details of
these methods may be found in any modern text book
on laboratory methods. We shall not give a descrip-
tion of them here.
Marshall's Method of Urea Estimation. — Quite re-
cently a rapid chemical method for the estimation of
urea in urine has been introduced by Marshall of Johns
Hopkins.19 It depends upon the conversion of urea into
ammonium carbonate by means of an enzyme prepared
from soy bean. This enzyme is called urease because of
its facility in effecting this conversion. Urease is found
in some bacteria and fungi. The following formula
represents the chemical decomposition produced:
NH2 ONH4
CO +2H2O = CO
NH2 ONH4
The presence of urease in soy bean (glycine hispida)
was first observed by Takeuchi in Japan. Its applica-
16Skand. Arch. f. Phys., 1891, II, p. 438; Zeit. f. phys. Chem.,
XVII, p. 140.
"Zeit. f. phys. Chem., 1901, XXXII, p. 504.
"Arch. f. d. Ges. Phys., 1896, LXII, p. 1.
19 Jour, of Biol. Chem., 1913, XIV, no. 3; 1913, XV, no. 3.
26 Manual of Vital Function Testing Methods
tion to the quantitative estimation of urea is due as
above stated to Marshall.
A convenient form of the enzyme urease is now to
be found upon the market under the name of urease
(Dunning). It is supplied in convenient 25 milligram
tablets put up 40 tablets per package by Hynson West-
cott & Co., pharmaceutical chemists of Baltimore,
Maryland.
Urease (Dunning) is a fine, almost white, powder,
with little taste or odor, soluble in slightly alkaline
water.
The apparatus and material required for the estima-
tion are as follows : Four 200 c.c. Erlenmeyer flasks
with cork stoppers ; one 50 c.c. glass-stoppered burette ;
one 5 c.c. bulb pipette; one small glass mortar; 100 c.c.
of solution of methyl orange; 1000 c.c. of decinormal
solution of HC1; 50 c.c. toluol and a package of urease
(Dunning) tablets.
Put 1 or 2 c.c. of toluol into each of two Erlenmeyer
flasks of 200 c.c. capacity. Into one of the flasks intro-
duce exactly 5 c.c. of a specimen of urine and 100 c.c.
of distilled water; stopper flask with cork. Crush a
urease tablet in a small glass mortar and dissolve in
about 5 c.c. of water. Transfer this solution without
loss into the other flask containing toluol and rinse
mortar with several portions of distilled water until
about 100 c.c. have been added to the contents of the
second flask. Add 5 c.c. of the urine and stopper with
a cork.
Each flask is now thoroughly shaken and allowed to
stand at room temperature over night or at least 8
hours. If it is necessary to get more rapid estimations
two tablets are used instead of one and the mixture
digested at 40 °C for an hour.
The test may indeed be completed in 15 minutes by
Tests of Liver Function 27
using only 1 c.c. of urine, two tablets and digesting at
40° Centigrade for 15 minutes. The factor would in
this case be 3 instead of .6, as will be seen later by
carrying out the longer time limit.
After the lapse of the time set the two solutions are
titrated to a distinct pink color with decinormal HC1,
using methyl orange as indicator.
The urease has converted the urea present in the urine
into ammonium carbonate. The amount of ammonium
carbonate formed by the urease is indicated by the
quantity of standard HC1 solution required to exactly
neutralize the contents of the flask containing urease
minus the quantity required for control specimen.
According to the chemical formula representing the
conversion of urea into ammonium carbonate (z?. s.) it
may be seen that 60 grams of urea are converted (by
urease) into 96 grams of ammonium carbonate. This
amount (96 grams) of ammonium carbonate would
require 72 grams of standard HC1 solution to neu-
tralize it.
As this quantity of HC1 solution (72 grams) is con-
tained in 20,000 c.c. of n/10 HC1 solution and is
equivalent to 60 grams of urea represented by 96 grams
of ammonium carbonate, then 1/20000 of the quantity
of 1 c.c. of n/10 HC1 solution will be equivalent of
1/20000 of 60 grams, equal .003 (60-^20000 =.003).
Therefore each c.c. of decinormal HC1 solution re-
quired to neutralize the enzyme treated specimen minus
the number of c.c. required to neutralize the control
specimen represents .003 of urea, and as the 5 c.c. speci-
men is the 1/ooo Part of a liter, multiply the number of
c.c. of n/io HC1 solution in excess of the control re-
quirements by the factor .6 (.003X200=.6) to find the
urea per liter when estimating the daily output.
One part of nitrogen is equivalent to 2.143 parts of
28 Manual of Vital Function Testing Methods
urea.
Estimation of Total Nitrogen in the Urine. Kjel-
dahl's Method.20 — Ten c.c. of urine are carefully meas-
ured into a Jena glass, round-bottom flask. Add a few
drops of concentrated solution of sulphate of copper,
15 c.c. of H2SO4 and 10 grams of potassium sulphate.
The flask is supported in an inclined position to pre-
vent loss by spurting. The mouth of the flask is loosely
closed by a glass bulb blown on end of a piece of glass
tubing.
The flask may be conveniently supported by a thick
piece of asbestos board with a hole in the center of a
size to permit the flame to come in contact only with the
portion of flask covered by fluid. Wire gauze protects
the flask from direct contact with the flame.
The flask should now be gently heated over a Bunsen
flame for half an hour. When foaming ceases the flame
is raised until the acid begins to boil gently. The whole
heating process is carried out under a hood because of
sulphurous acid fumes which are given off. When the
fluid in the flask is colorless or pale green, oxidation is
complete. This requires about two hours. The acid
and the oxidizer (CuS04) convert all nitrogenous mat-
ter into ammonium sulphate. The flask is then cooled.
The next step is distillation. The same or a larger
Jena glass flask may be used, but preferably a copper
apparatus which will not break. Two hundred c.c. of
water are added and enough 30% NaOH solution to
make the mixture strongly alkaline. Ammonia is set
free by the action of the alkali. This is distilled over
in 80 c.c. of decinormal H2SO4, which has been accu-
rately measured into a flask. The flask containing the
original solution is heated until about 2/3 of it have
passed over and there is considerable bumping from
80 Zeit. f. Chem., 1883, XXII, p. 3T8.
Tests of Liver Function 29
the separation of sodium sulphate. This usually re-
quires about thirty minutes. Bumping may be dimin-
ished by adding fragments of pumice, granulated zinc
or talcum powder at the beginning of distillation. A
simple form of apparatus can be extemporized in the
laboratory. [See Illustration.]
The tube D contains a few glass beads and some of
the H2SO4 is poured over these to prevent the escape of
any ammonia. A few drops of methyl orange are
added to the pearls and the flask C to indicate alkalin-
ity, in which event more acid is to be added promptly.
D is not necessary if a Liebig condenser is used.
The tube B prevents the alkaline fluid in A from
spurting over into E. The tubes E and B are made of
broken pipettes of 50 or 100 c.c. capacity.
The decinormal acid into which the ammonia has
30 Manual of Vital Function Testing Methods
condensed is titrated with n/10 NaOH. Methyl orange
is used as an indicator.
Subtract the number of c.c. of decinormal NaOH
used from the number of c.c. of acid taken and the
remainder will give the amount of ammonia distilled
over, for every c.c. of acid neutralized by the ammonia
is equivalent to so much decinormal ammonia.
Decinormal ammonia contains 1.4 grams of nitrogen
to the liter or .0014 gram per one c.c. The amount
of nitrogen can therefore be determined by multiplying
the number of c.c. of acid neutralized by .0014. This
gives the nitrogen in grams for 10 c.c. of urine used.
The description of the method given above is taken
from Wood's Chemical Diagnosis, N. Y., 1909, p. 408.
2. Augmentation of Urinary Ammonia as an Index of
Ureagenetic Liver Function
Ammonia, amino acids and carbonates constitute the
last intermediaries in the metabolic processes by which
the body proteids are catabolized into urea. As before
stated, the liver has been regarded as the chief factor in
this conversion.
If the functional capacity of the liver is deficient,
the amount of nitrogen eliminated in the form of am-
monia will be increased. Normally, .7 gram of am-
monia are excreted by the urine in 24 hours. In hepatic
insufficiency the amount may be doubled or trebled.
The relation of ammonia nitrogen to total nitrogen
in the urine is normally 3 or 4 parts per hundred.
Under pathological conditions, however, it may rise to
30 parts per 100 and such an increase may, under cer-
tain circumstances, indicate hepatic insufficiency. In
states of acidosis the ammonia nitrogen is also increased.
Tests of Liver Function 31
It is often useful to make a comparison between the
amount of nitrogen excreted as ammonia and the nitro-
gen eliminated as urea. This is done as follows : The
molecular weight of ammonia NH3 is 17, the nitrogen
fraction of ammonia is, therefore, 14/i7-
NH2
The molecular weight of urea CO is 60.
NH2
The nitrogen fraction is 28/6o or 7/i5.
The amount of ammonia in grams in a given sample
is estimated by the formalin method (v. «.), and 14/i7
of this represents the ammonia nitrogen. The urea is
calculated in grams in the same sample of urine, by
Marshall's method, and T/15 of the weight is nitrogen.
Under normal conditions the ammonia nitrogen is about
1/20 of the urea nitrogen.
Estimation of Ammonia Nitrogen in the Urine. The
Formalin Method. — The estimation of acidity is the
first stage in the estimation of ammonia nitrogen. Pro-
ceed as follows :
Measure out 25 c.c. of urine into a beaker and dilute
with about double the volume of distilled water. Add
2 or 3 drops of phenolphthalein. Run in n/10 NaOH
from a burette until a faint permanent pink color is
produced. Note the number of c.c. of NaOH used.
Measure about 10 c.c. of commercial (40%) formalin
into a second beaker. Add phenolphthalein. Neutral-
ize exactly with n/10 NaOH. Add the neutral formalin
to the neutral urine. The pink color disappears. Run
in n/10 NaOH until the pink color returns. Note the
number of c.c. of NaOH used. The result is calculated
in terms of n/10 NaOH for the acidity. That is to say
the acidity of the urine is given as the number of c.c. of
n/io NaOH required to neutralize 100 c.c. of urine to
phenolphthalein. Thus if 10 c.c. of soda were used in
32 Manual of Vital Function Testing Methods
the first titration to neutralize 25 c.c. of urine, the acid-
ity of the urine is 10/25X1QO:=40. The ammonia re-
sult should be expressed in grams of ammonia per 24
hours. The number of c.c. of soda used in the second
titration of the urine is the equivalent of the number of
c.c. of ammonia present in the 25 c.c. of urine. Sup-
posing the number of c.c. of soda used in the second
titration to have been 10, then 10 c.c. n/10 NaOH=
10 c.c. n/10 NH3 = 10 X -0017 gm. NH3. Therefore the
ammonia passed in the 24 hours = 10 X .0017 X
24 hours urine in c.c.
25
The reaction depends upon the combination of the am-
monium salts with formaldehyde to form urotropinc
and the consequent liberation of the acids previously
combined with ammonia.
Experimental Provocative Ammoniuria. — This test
is based upon the fact that normally, the liver trans-
forms all ammonia transported to it into urea. If there
is pathological alteration of the liver cell, this am-
monia will not be transformed and the quantity of
ammonia in the urine increases.
Before applying the test of provocative ammoniuria
in a given case the total ammonia excretion in 24 hours
should be estimated over two days. Meanwhile the
patient is put upon a fixed regimen.
In the morning, after having urinated, the subject is
given 6 grams of ammonium acetate. The urine for the
next 24 hours is collected and the ammonium content
estimated (v. s.) which can be compared with that
found prior to the test.
The value of this test is variously estimated. Some
have concluded that any considerable increase of am-
monia in the urine after the ingcstion will always in-
Tests of Liver Function 33
dicate an impairment of the functional integrity of the
liver cell. It is only claimed to be of value when posi-
tive. Others, on the contrary, have insisted that in spite
of very advanced disease of the liver the ammonium
salts ingested continue to be transformed into urea.
3. Aminoaciduria as a Criterion of Ureagenetic
Function
The existence of considerable quantities of amino
acids, leucine and tyrosine, in the urine in liver diseases
was noted in 1866 by Frerichs. In 1907 Glaessner 21
showed that in disease of the liver there is usually an
increase in the relation of amino nitrogen to total nitro-
gen in the urine. Normally this relation, or so called
coefficient, of aminoaciduria has been shown to vary
from .5 to 3.5 per 100. In diseases of the liver (chole-
lithiasis, cirrhosis), the ratio may rise to 11 or even 13
per 100.
Labbe and Bith have estimated the amount of amino
acids in the blood serum and have found it increased in
certain liver diseases.
Hyperaminoaciduria, according to Brille and Gar-
ban,22 is not necessarily an indication of liver disease.
It may occur during states of rapid emaciation, cach-
exia, pneumonia, typhoid fever and diabetes compli-
cated with acidosis.
These authors agree, however, that in all these states
it may not be improbable that the appearance of am-
inoaciduria is dependent upon a disturbance of the
function of the liver cell.
Provocative Aminoaciduria. — The principal sub-
stances which have been used are glycocol, alanin, as-
aZeit. f. Exper. Path. Therap., 1907, IV, p. 336.
22 Gaz. d. Hop., 1914, LXXXVII, p. 405.
34 Manual of Vital Function Testing Methods
paraginic acid, and commercial peptone. These amino
acids, also peptone, have been employed by different
experimenters.23 The patient is given certain quanti-
ties of these substances and the amount of amino-acid
excreted in the urine carefully measured.
If the proper care has been taken in the days which
precede the test to establish the normal amino nitrogen
coefficient for the individual and to keep the subject
under observation upon a fixed and invariable diet, then
provocative aminoaciduria tests may be of value. They
will frequently show a marked increase of the coefficient
after the ingestion of the amino-acids (or peptones)
as compared with the coefficient prior to the test, and
the marked increase according to some authors will
only occur when the liver parenchyma is diseased.
The insuperable difficulty in these methods of esti-
mating liver function arises from the fact that the esti-
mation of amino nitrogen in urine and blood requires
very complicated chemical manipulations which effectu-
ally prevent their introduction into clinical medicine.
4- Estimation of Residual Nitrogen in Blood Serum
as an Index of Hepatic Function. Chauffard Brodin
Test 24
The quantity of residual nitrogen in blood serum is
obtained by subtracting the urea nitrogen from the
total nitrogen estimated in dealbuminized serum. Re-
sidual nitrogen is made up of ammonia, amino-acid,
uric acid, etc. It has been contended that the elabora-
tion of residual nitrogen products is exclusively de-
""Ztschr. f. klin. Med., 1910, LXXI, p. 261; 1911, LXXIII, p.
325.
24 Jour. Biol. Chem., 1912, XII, p. 301; Jour. Amer. Chem. Soc.,
1913, XXV, p. 1567,
Tests of Liver Function 35
pendent upon the liver. In normal persons the re-
sidual nitrogen is always below 10 grams per liter
of serum. In many acute and chronic diseases of
the liver, the residual nitrogen has been found above
10 grams, the amount being in proportion to the grav-
ity of the hepatic lesion. The diet and condition of
the kidney are negligible.
The estimation of the amount of residual or rest
nitrogen in the blood serum has come to be regarded
as of more importance as a test of kidney than of liver
function. For this reason a more complete account of
the method and its interpretation will be found later
under that head.
General Summary of the Value of Ureagenetic Func-
tion Tests. — In the first place it must be admitted that
some of these ureagenetic tests are complicated to such
an extent that they cannot be carried out without the
assistance of an expert chemist, thus diminishing their
applicability to clinical work.
Further than this it must likewise be admitted that
many physiological causes of error exist which militate
against a too strict interpretation of results. Other
factors besides the liver collaborate in nitrogen metab-
olism.
Notwithstanding these valid objections it must be
admitted that the study of nitrogen has given some
useful results and it is to be hoped that future investi-
gations may bring a greater degree of order from the
now more or less confused and contradictory material
which comprises our stock of knowledge today concern-
ing the true relation of the liver to nitrogenous metab-
olism.
When this day arrives the clinician will be better
able to interpret the results obtained from ureagenetic
tests than he is at the present time.
36 Manual of Vital Function Testing Methods
in. FUNCTIONAL TESTS TO DETERMINE DISTURBANCE
OF THE ANTITOXIC FUNCTION OF THE LIVER
Poisons which obtain access to the organism thro
the portal circulation are fixed and destroyed under
normal circumstances by the liver cells. It is natural,
therefore, that this important function should be em-
ployed as a basis for testing the integrity of the organ.
The first efforts made in this direction consisted
in estimating the toxicity of urine and blood serum.
An increase of toxicity of these bodies, it was held, in-
dicates diminished toxicopexic power of the liver paren-
chyma. Tests of this character are carried out upon
lower animals. The urine to be tested being injected
intravenously into rabbits according to Bouchard's 25
method, and blood serum or urine into the cerebrum
of rabbits according to Widal's method. The numerous
sources of error and the technical complications sur-
rounding these two methods have prevented their intro-
duction into clinical medicine and they need not be
described.
The methods which are in use to determine the anti-
toxic functional power of the liver are two in number.
They are as follows: 1. The Methylene Blue Test of
Chauffard and Castaigne. 2. Estimation of Indica-
nuria, Spontaneous and Provocative.
1. Methylene Blue Test of Toxopexic Function of the
Liver. Chauffard, Castaigne Test 26
When the liver cell is unable to properly arrest and
fix poisons, it reacts to the passage of methylene blue
28 See also Estimation of Urinary Toxicity as a Test of Renal
Function.
""Presse M6d., 1898, April 23; Jour, de Physiol. Path., 1899,
May. See also Babaliantz, These de Geneve, 1912.
Tests of Liver Function 37
through its parenchyma by an intermittent elimina-
tion.
Test. — Inject 1 c.c. of 5% solution of methylene
blue subcutaneously. Collect and examine the urine
in half an hour, then every hour. Normally at the
end of half an hour after injection the urine becomes
colored, the color rising to maximum in 3 or 4 hours,
disappearing in about 50 hours. If the elimination
when it commences instead of being continuous occurs
in cycles, that is intermittently, the test is positive and
may indicate hepatic insufficiency.
It must be acknowledged that the value of this test
is greatly diminished by the fact that the state of
renal permeability must be considered, since this will
have a predominant influence upon the quantity and
rapidity of elimination. Where this factor is known
the test may be of value.
Roche's Modification of Chauffard's Methylene Blue
Test. — In this test the methylene blue is taken inter-
nally instead of being given hypodermically. .002 gm.
of methylene blue is swallowed at eight o'clock in the
morning on an empty stomach. The substance is given
in capsule. The urine is collected every four hours in
separate vessels. If the urine of the second recipient
is clearly colored it will denote inability of the liver
to retain the pigment and consequent hepatic insuf-
ficiency. Normally this amount of methylene blue
should be completely arrested and fixed by the liver
so that no coloring matter appears in the urine after
its administration. If the test is positive, the urine,
especially that passed four to eight hours after ad-
ministration, will be colored green.
Unfortunately the condition of the kidney must also
be reckoned Avith in applying this test, as failure of
38 Manual of Vital Function Testing Methods
elimination of the dye may be due to renal impermea-
bility.
The same remarks with respect to renal permea-
bility apply in interpreting the Roche modification as
when the dye is given by hypodermic injection. It
can never be known a priori whether a failure of elimi-
nation is due to renal insufficiency or to normal fixa-
tion of the dye by the liver cell. In other words, the
state of the kidney function must be previously known
or ascertained.
2. Indicanuria Spontaneous and Provocative as a
Means of Testing the Integrity of Hepatic Fixa-
tion
Spontaneous Indicanuria. — It has been urged that
the normal liver is always capable of arresting and
destroying indican which is formed in the intestine as
a result of putrefaction of albuminoids. Therefore
the spontaneous presence of indican in the urine has
been held to be evidence of hepatic insufficiency.
Provocative Indicanuria — .001 gm. of indol is in-
gested in the morning on an empty stomach. The urine
is collected every four hours and examined for indican.
The individual to be tested should be put on a milk
diet for a few days before the substance is ingested.
TESTS FOR INDICAN IN THE URINE
Qualitative.-— The principle involved is to decom-
pose the sodium or potassium compound of indoxyl
sulphuric acid present in the urine by strong HC1 and
oxidizing this compound.
Obermayer's reagent is usually employed. This con-
sists of strong HC1, sp. gr. 1.19, to which is added two
parts per thousand of ferric chloride. The liquid is
Tests of Liver Function 39
fuming yellow and keeps indefinitely.
Precipitate the specimen of urine to be tested with
a small amount of lead acetate or subacetate, avoiding
excess, and filter. This removes pigments. 15 c.c. of
filtered urine are mixed with equal quantity of Ober-
mayer's reagent and 2 c.c. of chloroform added. Cork
or cap the tube with rubber, and slowly invert. The
chloroform takes on a blue color whose depth will
roughly show the amount of indican present.
Quantitative. — Strauss' 2T method is a good one.
Twenty c.c. of urine are mixed with 5 c.c. of 20% lead
acetate solution and filtered. Ten c.c. of filtrate (cor-
responding to 8 c.c. urine) are placed in a small gradu-
ated separatory funnel and mixed with 10 c.c. of Ober-
mayer's reagent (v. s.).
Five c.c. of chloroform are added, the tube corked
and gently shaken. This is repeated in two minutes.
Pour the chloroform from the tube. Add 5 c.c. of
chloroform and repeat the extraction. Continue until
added chloroform remains colorless.
Two c.c. of united chloroform extracts are put in
a small test tube of same diameter as tube containing
standard solution. Chloroform is added drop by drop
until colors are matched, against white background.
The standard solution is made by dissolving .001 gm.
C.P. indigotin (Kahlbaum) in 1000 c.c. of chloroform.
Portion is sealed in test tube and kept in the dark.
If the total amount of chloroform used for extrac-
tion is equal to a and the amount of chloroform used
to dilute the 2 c.c. to the color of the standard tube
equals x, the total amount of chloroform necessary to
dilute all the chloroform used in extraction equals
27Deutsch. med. Wchnsch., 1902, p. 299.
40 Manual of Vital Function Testing Methods
The total number of c.c. used in the extraction and
in the dilution of the extraction mixture represents,
therefore, a bulk containing .001 gm. of indigo.
In normal urines 5-10 c.c. of chloroform are usually
all that is required to extract the whole amount.
To obtain the results in milligrams it must be con-
sidered that the amount of indigo extracted was from
8 c.c. of urine.
IV. FUNCTIONAL TESTS TO DETERMINE DISTURBANCES
OF THE SANGUINOPOIETIC FUNCTION OF THE LIVER
CELL
Some results of practical value have already come
from tests of hepatic function based upon the haemic
activities of the liver. The whole question of the rela-
tion of the liver to the biology of the blood is very com-
plex and not completely understood at the present
time ; but it is generally agreed that the liver is inti-
mately concerned in the elaboration of some of the
constituents of the blood, particularly fibrinogen and
certain of the ferments.
The haemic tests so far proposed have dealt with
two physiological aspects of the relation between the
liver and blood : first coagulability and second the
presence of miscellaneous ferments. If the liver is in
reality the principal source for the elaboration of
fibrinogen any notable diminution of this substance
in the blood will indicate a diminution of the func-
tional integrity of the liver cells. With diminished
fibrinogen and perhaps also fibrin ferment, a delay or
deficiency in the coagulability of the blood will ensue.
One test of liver function will therefore consist in
estimating the coagulability of the blood.
With respect to the tests for ferments in the blood
Tests of Liver Function 41
it may be said that the estimation of lipase is so far
the most important. The liver under normal circum-
stances inhibits the formation of this ferment so that
hepatic insufficiency results in an actual increase in
the amount of lypolytic enzyme in the blood.
The other ferment tests of hepatic function are ap-
parently too subtle and uncertain to be of any con-
siderable practical value.
The functional tests which are in use at the present
day to determine the sanguinopoietic capacity of the
liver are as follows: 1. Estimation of blood coagula-
tion time, and estimation of fibrinogen. 2. Estimation
of fibrinolysis time. 3. Estimation of the amount of
lipase in the blood.
1. Estimation of Blood Coagulation Time as an Index
of Liver Function
Prolongation of coagulation time of the blood is
said to be present when the liver is physiologically de-
fective and consequently the estimation of coagulation
time has been proposed as a simple and effective means
of determining hepatic insufficiency.
Unfortunately while the relation of normal hepatic
function to normal coagulation of the blood is un-
doubtedly intimate and important, it is also true that
so many other factors besides the liver enter into the
physiology of blood coagulation as to materially lessen
its importance as a basis for functional estimation.
Coagulation time of the blood has also been sug-
gested by Tettinger and Bachrach as a means of test-
ing renal function.
Wright's Method of Estimating Coagulation Time.
—The necessary apparatus consists of a series of cap-
illary tubes, elastic bands, a beaker, a jug of hot and
42 Manual of Vital Function Testing Methods
a jug of cold water, a watch with a second hand and
a thermometer. The capillary tubes are of the same
caliber and are provided with a 5 c. mm. mark. The
procedure is as follows: Clean the patient's thumb
with ether. Wrap a piece of elastic tubing round the
thumb from the base nearly to the tip. Puncture the
tip of the thumb with a sterile surgical needle. Draw
up blood to the mark on the pipette. It is not essential
to obtain the exact quantity of blood, slight variations
in the amount being of less importance than rapid
manipulation. Note the exact time by the watch.
Stretch a flat elastic band over the ends of the tube to
prevent entrance of water. Stand the tube in the
beaker filled with water at 37° C. Stir the water oc-
casionally with the thermometer and keep the tem-
perature constant by adding hot or cold water.
Prepare three or four more capillary tubes in the
same way, numbering each tube and taking the time of
each. At the end of three minutes take out the first
tube and blow out the blood. Give the second tube
3}/o minutes and if the blood is still fluid give the third
tube four minutes and so on. The tube from which
the blood fails to be expelled by blowing gives the co-
agulation time. The normal time for blood coagulation
is 31/2 minutes.
The Fibrinogen Test. Whipple Hormtz.28 — 20 c.c.
of oxalated blood plasma are heated to 59° C. for 20
minutes. Fibrinogen is precipitated. The precipitate
is isolated by centrifugation, washed with water, alcohol
and ether, dried at 120° C. and weighed. A rough esti-
mate of the amount of fibrinogcn in the blood is made
by clotting a little plasma with calcium, testing the
28 Jour. Exp. Med., 1911, XIII, p. 136; also Johns Hopk. Hosp.
Bull., 1913, XXIV, p. 207, 343.
Tests of Liver Function 43
toughness of the clot with a glass rod. The quantita-
tive or weighing method is, however, better.
Normally fibrinogen exists in the plasma in the pro-
portion of .30 to .40 gm. per 100 c.c. In case of liver
deterioration, alteration or injury the amount will
be found diminished. In some cases of cirrhosis the fibri-
nogen content has been found very low (.05 gm. or less.)
#. Estimation of Fibrinolysis Time as an Index of
Hepatic Function. Goodpasture Test 29
Very recently, Goodpasture has called attention to
the interesting and important fact that in chronic,
atrophic, hepatic cirrhosis the blood possesses the power
of completely digesting the clot in a few hours at body
temperature.
The clot in normal blood remains undigested for
days and sometimes even for weeks. Goodpasture be-
lieves that the dissolution of the clot is due to an
enzyme. The activity of this enzyme is destroyed by
heat and inhibited by normal serum. The fibrinogen
content of the blood in his reported cases (four in
number) was below normal. He suggests premature
digestion of clot durante vivo may account for the
frequency of spontaneous hemorrhage in atrophic
cirrhosis. The Goodpasture test will, in all likelihood,
be found of great value in estimating hepatic insuf-
ficiency in chronic cirrhosis. It will be interesting to
observe what the test for fibrinolysis will show in early
or latent cases of cirrhosis as well as in hepatic con-
ditions generally.
Technic of Goodpasture Test. — Blood is drawn from
an arm vein by means of the usual technic. The coagu-
29 Johns Hopk. Hosp. Bull., 1914, XXV, p. 330.
44 Manual of Vital Function Testing Methods
lation time is estimated.* A portion of blood is drawn
into 1 Jo solution of sodium oxalate to prevent clotting
of the specimen. This last is centrifuged and 20 c.c.
of supernatant plasma is used to determine fibrinogen
content if this is desired. Otherwise the entire oxalated
plasma is used for tests of coagulation time and fibrin-
olysis.
In testing the oxalated plasma for coagulation time,
use 1 c-c. of plasma -)- 1 gtt. CaCl2 (1%).
The original clot from the drawn blood and speci-
mens of clotted oxalate plasma are placed in the ther-
mostat at 37°. They are examined every hour. If the
test is positive, the blood clot liquefies and is dissolved
in 31/2 to 5 hours.
In negative (normal) cases there is no digestion of
the clot for several days, even where no aseptic pre-
cautions are taken.
3. Estimation of Blood Lipase as an Index of Liver
Function. Whipple's Test 30
There exists normally in the blood a lipolytic fer-
ment lipase. The percentage of lipase in normal blood
is remarkably uniform. In certain diseases of the liver
this ferment has been found to increase in amount indi-
cating that under normal circumstances the liver in-
hibits its formation. Any considerable increase, there-
fore, of lipase in the blood has been held to indicate
hepatic insufficiency. Whipple, in collaboration with
Mason and Peightal, found that after acute injury of
the liver from chloroform there was always found an in-
* For other methods than Wright's consult modern laboratory
manuals.
30 Johns Hopk. Hosp. Bull., 1913, XXIV, p. 207; ibid., 343;
ibid., 357.
Tests of Liver Function 45
crease in the lipase of serum or plasma. Sometimes this
rise amounted to 1—2 c.c. of 1/10 normal acid. Inas-
much as the content of lipase may increase five to eight
times the normal under certain conditions of hepatic
disease it was naturally suggested as a test of hepatic
insufficiency.
The value of the test is chiefly qualitative rather than
quantitative. It has been found of especially positive
value in suspected eclampsia, chloroform poisoning, yel-
low atrophy and cholangitis. In cirrhosis of the liver
there may be a subnormal lipase.
Tests for Lipase in the Blood. Lowerihart's 31
Method of Estimating Lipase Is Usually Employed. —
Blood serum is collected in four tubes and a little
toluene, .3 c.c., is added to prevent microbic contami-
nation. Each tube contains 1 c.c. plasma or serum
diluted with 4 c.c. of distilled water.
Two of these tubes are used as controls. To the
others a little ethyl butyrate (butyric ether), .26 c.c.,
is added. The four tubes are shaken, corked and put in
the thermostat at 38° for 18 to 24 hours.
At the end of this time the two control tubes are ex-
amined for their normal alkalinity by means of deci-
normal acid solution. The other two tubes are titrated
for free butyric acid by decinormal alkali solution.
The total lipolytic activity is measured by adding
the two figures, since the butyric acid formed had to
first neutralize the normal alkalinity of the serum.
The exact method is as follows : After incubation
the tubes are cooled in ice water, 3 drops of azolitmin
added and then titrated in pairs to a neutral reaction,
using Vio normal acid and alkali. The two control
tubes usually show the blood alkalinity to be
.1 c.c. Vio normal acid and the butyrate tubes show
81 Amer. Jour. Physiol., 1902, VI, p. 331.
46 Manual of Vital Function Testing Methods
the acid production to be .1 to .2 c.c. above the neutral
point. This means that the total lipolytic activity is
.2 to .3 c.c. Vio normal solution, that the plasma
lipase has split up the ethyl butyrate to this amount.
Normal plasma lipase is then in terms of Vio normal
acid equal to .20 to .30 c.c.
Ghedini's Test.^2 — This consists in an estimation of
the power of the blood serum to convert glycogen into
glucose. A reducing ferment supposed to be formed
by the liver cells effects the conversion.
The Test. — Blood serum to be examined is added to
a solution of glycogen. Scrum from a known normal
person is similarly treated and used as a control.
To both tubes is added a little sodium hydroxide,
then potassium sulphocyanide, and the solutions fil-
tered and examined with the polariscope.
If there is hepatic insufficiency the rotatory power
of the serum will be less than the normal control.
The factors upon which this test are founded are
rather subtle and imperfectly understood ; indeed it is
by no means accepted that the liver is the unique or
even the most important source of the ferment which
affects the conversion of glycogen into glucose.
4- Application of Abderhalden's 33 Method to Estima-
tion of Sanguinopoietic Functions of the Liver
Recently it has been held that destruction of liver
parenchyma by disease (autolysis) gives rise to the
presence of an excess of protcolytic ferments in the
blood serum and this fact has formed a basis for test-
32 Gazz. degli ospcdali, Milan, Jan. 12.
33 See Breitmann, Zentrbl. f. innere Med., XXIV, 1913, p. 857.
Tests of Liver Function 47
ing the functional capacity of the liver.
The technic of the method is too complicated to
admit of its use in clinical practice and the results ob-
tained are as yet too meagre to justify any conclusions
as to its value.
V. FUNCTIONAL, TESTS TO DETERMINE DISTURBANCE OF
THE EXOCRINOUS OR BILIARY FUNCTION OF THE
LIVER
Bile is partly a secretion and party an excretion, for
it not only plays a role in certain digestive processes,
notably the splitting or absorption of fats, but it also
contains certain waste products which escape in the
feces. It is important to remember, however, that cer-
tain constituents secreted and excreted in the bile are
reabsorbed by the intestine and carried back by the
blood stream to the liver.
Perfectly fresh hepatic bile, in contradistinction to
bile in the gall bladder, contains one pigment only,
namely, bilirubin, and from this pigment others are
gradually formed by processes of oxidation. Fresh
human bile from the hepatic duct is golden yellow in
color but becomes olive brown to grass green after re-
maining in the gall bladder and cystic duct, because of
the presence of biliverdin, which is an oxidation product
of bilirubin.
It is now universally accepted that the bile pigment
bilirubin originates from the disintegrated hemoglobin
of the red blood corpuscles. When these break up
their contained pigment is carried to and fixed by the
liver, where it is converted into an iron-free pigment,
bilirubin.
It appears that the power of transforming blood
pigment into bilirubin is not exclusively the property
48 Manual of Vital Function Testing Methods
of the liver cell, since in old blood extravasions any-
where in the body a substance, haematoidin, is found
which is chemically identical with bilirubin. Under
ordinary physiological conditions, however, the liver
seems to be the only place in the body where bilirubin
is formed. It is not known whether the actual dis-
integration of red corpuscles is confined to the liver,
but certainly this organ has the unique power of fixing
the hemoglobin set free by haemolysis, retaining its
iron and converting it into bilirubin.
Under normal conditions bilirubin does not appear
in the feces, but in its place is found a reduction prod-
uct which is known as urobilin (stercobilin). The reduc-
tion of bilirubin to urobilin takes place in the intestine
from nascent hydrogen liberated by bacterial action.
Part of this urobilin formed in the intestine escapes in
the stools. Another portion is reabsorbed by the in-
testinal mucosa as a protochrome, — urobilinogen. Part
of this substance is eliminated in the feccs. The re-
mainder passes back thro the portal circulation to
the liver, where it is changed back into bilirubin to be
again reexcreted by the bile when the cycle is re-
peated.
A very minute amount of the urobilinogen absorbed
by the bowel escapes in the urine. Under normal con-
ditions this quantity is very small.
Soon after urobilinogen is eliminated by the urine,
it changes to urobilin so that the urobilin is not present
in freshly passed urine but only appears from the break-
ing down of urobilinogen by light and air.
Urobilin was recognized as a constitutent of patho-
logical urines long before urobilinogcn, from which it
is formed by oxidation, was discovered.
Urobilinuria and urobilinogenuria may therefore be
regarded as practically identical and due to the same
Tests of Liver Function 49
substance in different forms. The pathological causes
of both are the same.
It has been known for a long time, however, that
urobilin appears in the urine in a comparatively large
number of pathological conditions, among which may
be mentioned the infectious diseases, particularly ma-
laria and pneumonia, cirrhosis of the liver, lead poison-
ing, decompensated heart disease, pernicious anaemia,
pulmonary infarction and visceral hemorrhages. Like-
wise certain drugs and poisons are known to produce
urobilinuria. In obstructive jaundice, it often appears
in the urine before bilirubin and may, in fact, alternate
with this substance. Urines which contain much uro-
bilin present a dark yellow color which may be imparted
to the foam on shaking. They are thus quite like or-
dinary icteric urines.
The hepatic conditions in which urobilin appears in
the urine fall into two groups: 1. Mechanical inter-
ference with biliary flow in the ducts. 2. Insufficiency
of the liver cell. In the first group, if the obstruction
is absolute as in some cases of stone, no bile reaches
the intestine and no urobilin is formed, hence no uro-
bilinogen appears in the blood or urine. If the ob-
struction remains complete the bile pigment bilirubin
begins to be absorbed by the blood and excreted in
the urine. If the occlusion of the ducts is only partial,
some bile reaches the intestine, urobilin is formed, ab-
sorbed and may appear as urobilinogen in the urine.
On removal of the obstruction the full flow of bile in
the intestine is resumed. The functional activity of
the liver is sufficient to eliminate the urobilinogen
brought to it by the blood from the intestine and hence
there is no call for its elimination by the kidney and
it becomes reduced to a mere trace in the urine or com-
pletely disappears therefrom.
50 Manual of Vital Function Testing Methods
As a sign of liver insufficiency urobilinogenuria may
be absolute or relative. Absolute when the liver paren-
chyma is totally unable to eliminate urobilinogen and
allows it to get into the blood; relative when an ex-
cessive breaking down of red corpuscles anywhere in
the body overwhelms the liver with pigment which the
cells are unable completely to convert into bilirubin.
In other words, if the liver is insufficient it will be un-
able to excrete increased amounts of urobilinogen
whether derived from intestinal urobilin or from blood
pigments. In either case there will be an increase or
accumulation of urobilinogen in the blood and it will
appear in measurable amounts in the urine.
Icterus with the appearance in the urine of normal
bile pigment (bilirubin) may be regarded as a further
step in such a pathological process. Here the defect
of excretory function of the liver is greater because
of actual inflammatory obstruction of the biliary tract.
Jaundice, as is well known, is the result of the absorp-
tion into the blood of the bile pigment bilirubin. Jaun-
dice, whether obstructive or toxemic, is always due to
some lesion of the biliary excretory passages by which
the flow of bile into the intestine is diminished or pre-
vented and is always accompanied by some inflam-
matory state of the biliary tubules. Icterus, when
well marked, becomes clinically quite evident from the
discoloration of tissue which results. The demonstra-
tion of bilirubin in the urine is then superfluous. In
such an event, icterus becomes an element in the semi-
ology of hepatic disease and has to be evaluated with
other symptoms in making a diagnosis.
The presence of small quantities of bile pigment
(bilirubin) in the urine is, however, an earlier sign of
exocrinous hepatic insufficiency than icterus. For this
reason the tests for bile pigment in the urine become
Tests of Liver Function 51
tests to a certain extent at least of hepatic disease and
perhaps insufficiency.
To summarize the above facts in their relation to
the estimation of the external secretory function of
the liver it may be said that disturbances of this func-
tion are identified by tests which disclose the presence
in the urine of the three substances above mentioned,
namely, urobilinogen, urobilin and bilirubin ; the tests
for these substances will therefore be given together
with special data bearing upon their individual sig-
nificance.
There is, however, another way by which the total
power of the liver to carry on its external secretory
function may be judged. This consists theoretically,
in finding a substance which if injected into the circula-
tion will be eliminated exclusively by the liver. It
was only in recent years that such a substance was dis-
covered, phenoltetrachlorphthalein.
Tests to determine the status of the excretory func-
tional power of the liver may, therefore, be properly
divided into two categories :
1. Tests for urobilinogen, urobilin and bilirubin in
the urine with the interpretation of results in reference
to hepatic insufficiency. 2. Tests to determine the
global capacity of the liver to eliminate foreign sub-
stances. In this category there is but one test so far
devised, namely that of phenoltetrachlorphthalein.
1. Tests for Urobilinogen, Urobilin and Bilirubin
in the Urine. Interpretation of Results with Refer-
ence to Hepatic Insufficiency
The Urobilinogen Test. Ehrlich's Test.34— The Ben-
zaldehyde Reaction. — While investigating the aniline
34Wien. med. Wchnschr., 1901, XV; Munch, med. Wchnschr.,
1901, XV.
52 Manual of Vital Function Testing Methods
dyes for their effects upon trypanosomes, Ehrlich found
that the addition of paradimethylamidobenzaldehyde to
certain fresh urines produced a bright red coloration.
This was in 1901. In 1903 Pappenheim 35 called atten-
tion to the fact that the reaction occurred only in those
urines which, on standing, gave the reaction of urobilin.
In the same year Neubauer 36 demonstrated that the
reaction was due to urobilinogen, a colorless chromogen
which gradually becomes converted into urobilin.
Technic. — The test is very simple. Add to fresh
urine in a test tube several drops of Ehrlich's reagent.
The reagent is as follows :
Paradimethylaminobenzaldehyde ... 8 gms.
Concentrated hydrochloric acid .... 80 gms.
Distilled water 200 gms.
If urobilinogen is present a red color appears. The
color reaction in the cold urine is of pathological sig-
nificance only when a distinct scarlet color is obtained.
There are a few sources of possible error. The in-
gestion of hexamethylamine or antipyrine may cause
the same reaction. The presence of acetone in the
urine must also be excluded as it produces a similar
coloration.
If the reaction persists after free purgation it is
more significant. The reaction is not constant in all
conditions in which it has been found because the liver
may be able to excrete enough urobilinogen to prevent
its appearance in the blood and urine.
Urobilinogen has been found in the urine in many
pathological conditions, chief of which are cirrhosis of
the liver, cholangitis, infectious diseases, heart diseases
80 Berl. klin. Wchnschr., 1903, II, p. 42.
36 Sitz. d. Gesell. f. Morph. u. Physiol., Munich, 1903, July, H. ii.
Tests of Liver Function 53
in the stage of decompensation, pernicious anaemia, pul-
monary infarction and visceral hemorrhage.
Ehrlich's test has been highly recommended as a clin-
ical method for determining hepatic insufficiency by
Miiller, Bauer, Neubauer, Hilderbrand and others. It
has been regarded by some observers as an adequate and
infallible criterion of hepatic function. Some observers
have claimed for it a prognostic significance. All of
these contentions have been found to overstate the facts.
The presence of urobilinogen in considerable quan-
tity in the urine may indicate that there is a partial
interruption in the biliary excretion and that some of
the intestinal urobilin absorbed into the blood is not
being thrown off by the liver. The primary cause may
be either a disorder of the liver cells or a congestion or
toxemic obstruction of the biliary channels.
The discovery of a persistent urobilinogenuria should
therefore point to a careful study of the hepatic func-
tions by all known means. If hepatic insufficiency can
be ruled out the only remaining explanation is that
excessive destruction of the blood corpuscles is taking
place somewhere in the body.
Recently rather strong criticisms of Ehrlich's test
have appeared. Wilbur and Addis 37 in 1913 stated
that they do not believe it constitutes a reliable criterion
of hepatic function. This opinion has been reiterated
by Chesney, Marshall, and Rowntree,38 last year. All
these authors maintain that observations on a single
or 24-hour specimen of urine have no significance. That
great variations appear from day to day and that
only when repeated tests are made covering a period of
two weeks, controlled by studies of urobilin content in
the feces, can the results be accepted.
87 Arch, of Int. Med., 1913, Feb., p. 235.
38 Jour. Amer. Med. Assn., 1914, LXXIII, p. 1533.
54 Manual of Vital Function Testing Methods
The simplicity of Ehrlich's test will insure for it a
permanent, if not paramount, place. It must always
be remembered that only a persistent and well-marked
urobilinogenuria is to be regarded clinically as sig-
nificant.
A well-marked and lasting reaction appears to in-
dicate one of two things, an hepatic insufficiency or
excessive hemolysis. There is nothing in the test itself
to enable a differentiation between the two to be made.
The test is therefore of some importance when regarded
only as a corroborative sign of insufficiency of the
excretory function of the liver.
Tests for Urobilin. — First add to the urine a few
drops of 10% solution of zinc chloride and enough am-
monia to dissolve the precipitate. Filter into a test
tube and hold same against a dark background ; a green
fluorescence denotes urobilin. Equal quantities of 1%
solution of zinc acetate and urine may be used in the
first part of the test. The fluorescence may be made
more visible by concentrating light on the tube with a
lens.
Another and perhaps more delicate method of de-
tecting urobilin is to extract 50 c.c. of urine with 50 c.c.
of pure ether. Pour off the ether into a tube and evap-
orate. Dissolve the brown residue in a little strong
alcohol. The solution will be pale yellow with a green
fluorescence if urobilin is present.
A third method is to acidify 20 c.c. of urine with
several drops of HC1. Shake gently with 5 c.c. of amyl
alcohol. This extracts the pigment and shows a bright
green fluorescence when treated with an alcoholic solu-
tion of zinc chloride and a little ammonia. By trans-
mitted light the amyl alcohol is a faint pink shade.
The most rapid method of testing for urobilin is by
Tests of Liver Function 55
the use of the pocket spectroscope. If the urine is dark
it should be diluted. The characteristic spectrum band
is seen between the green and blue, between the lines
C and F.
Tests for Bilirubin. — The tests usually employed are
those of Gmelin, Rosenbach, Huppert and Smith.
Gmelin's Test.— The urine is treated with sufficient
concentrated HNO3 containing a trace of nitrous acid,
sufficient to form a layer beneath. If bilirubin is pres-
ent there will be a play of colors at the zone of contact
from yellow through green, blue, violet, red and orange.
The green will lie nearest the urine and the orange in
the upper acid.
Rosenbach' 's Test.— Filter the urine through Swedish
filter paper ; apply a drop of HNO3 containing a trace
of nitrous acid upon the paper. The play of colors
above mentioned under Gmelin's test will appear.
Huppert' s Test. — Precipitate the urine with barium
chloride and ammonia. The precipitate is washed with
water. Wash the precipitate with alcohol into alcohol
acidulated with sulphuric acid. Boil for a time. If
bilirubin is present an emerald green color will appear.
Smith's Test. — A small amount of urine is placed in
a test tube and overlaid with a few c.c. of tincture of
iodine diluted with alcohol 1 :10. If bilirubin is present
a distinct emerald green ring will develop at the zone of
contact.
2. Tests to Determine the Global Capacity of the
Liver to Eliminate Foreign Substances
The Phenoltetrachlorphthalein Test of Liver Func-
tion. Rowntree, Horwitz, and Bloomfield Test. — Phe-
56 Manual of Vital Function Testing Methods
noltetrachlorphthalein was originally studied pharma-
cologically by Abel and Rowntree 39 in 1909. It was
proposed in 1913 as a test for functional capacity of
the liver by Rowntree, Horwitz, and Bloomfield. (Johns
Hopk. Hosp. Bull., 1913, XXIV, p. 327.) Important
experimental work was done on the drug in 1913 to
determine its behaviour in different liver injuries, by
Whipple, Mason and Peightal,40 and by Whipple,
Peightal and Clark.41
Great interest is attached to the phenoltetrachlor-
phthalein test for liver function ; first, because it bids
fair to become the most satisfactory test for the pur-
pose yet devised, and, secondly, because all the work
upon it has been done in America.
Professors Orndorff and Black 42 of Cornell Univer-
sity were the first to make the substance in 1909. The
pharmacological investigations of Abel and Rowntree
included a study of phenolphthalein together with the
new synthetic phthaleins, with special reference to their
behaviour as purgatives. They found that phenolphtha-
lein and its halogen substitution products, of which
phenoltetrachlorphthalein is one, do not differ greatly
in their physiological action. They are nonirritant to
mucous membranes and subcutaneously when injected in
oil. They are of low toxicity and possess no bacteri-
cidal action.
Both phthaleins are laxative when given by mouth,
subcutaneously or intravenously. When an oily solu-
tion of phenoltetrachlorphthalein (.4 gm.) is injected
under the skin of dogs or human beings a laxative action
is induced which continues from 4 to 6 days. When the
80 Jour. Pharmacol. and Expcr. Therap., 1909, I, p. 231.
"Johns Hopk. Bull., 1913, XXIV, p. 207.
"Johns Hopk. Bull., 1913, XXIV, p. 343.
0 Amer. Chem. Jour., XLI, 1909, p. 349.
Tests of Liver Function 57
tetrachlorphthalein is given subcutaneously it escapes
from the body exclusively in the bile. When the same
substance is given by mouth it is not absorbed. After
subcutaneous administration of phenoltetrachlorphtha-
lein the drug escapes dissolved in the bile and becomes
later absorbed by the mucous membrane of the large
intestine.
These facts form the physiological basis for the phe-
noltetrachlorphthalein test. The drug is eliminated ex-
clusively, or practically so, in the bile, and since this
excretion can be hurried through by purgatives, no time
will be given for its absorption, and thus the actual
amount of the drug eliminated may be found.
The experimental work done by Whipple, Mason and
Peightal, and by Whipple, Peightal and Clark has
established the fact that there is a striking parallelism
in animal observations between the amount of experi-
mental liver injury produced and the amount of phtha-
lein eliminated.
Their method of determining this fact was as follows :
Female dogs were used because of ease of catheteriza-
tion. The dogs were given intravenously .1 gm. of the
phthalein when weighing between 10-20 pounds ; .2 gm.
when weighing over 20 pounds. The injection was given
in the forenoon and 200-300 c.c. of water administered
by stomach tube. After 5 or 6 hours the urine was col-
lected and magnesium sulphate and croton oil given to
produce several semi-fluid stools. The feces were col-
lected next morning. The total feces were then diluted
to 1 to 2 liters. The mixture was made alkaline with
5-10 c.c. of 40% solution of sodium hydroxide and
shaken until uniform. One tenth of this quantity was
taken and diluted to 500 c.c. with water, 3 to 4 c.c. of
40% solution sodium hydroxide added and the mixture
thoroughly shaken. Of this second solution 100 c.c.
58 Manual of Vital Function Testing Methods
were precipitated with 5 c.c. of saturated solution of
basic lead acetate. After a few seconds a curdy pre-
cipitate fell. The solution was made up to 200 c.c.
with water containing 4 c.c. of 40 c/o solution sodium
hydroxide. On standing, the supernatant fluid showed
a clear phthalein color. This was filtered and the clear
solution read off in a colorimeter against a standard
solution .01 gm. phthalein to the liter.
In the hands of its authors, this method gave pretty
uniform results; on normal dogs the amount excreted
being 35 to 50% with .1 gm. injection and 40 to 50%
with .2 gm. injection. The drug did not appear in the
urine. About 10-15% of the phthalein injected was
lost from the time of the injection to that of its being
poured out by the bile into the intestine. This shows
that the liver is quite specifically concerned in the elim-
ination of the substance, phenoltetrachlorphthalein.
Whipple and his collaborators found that when the
liver parenchyma is artificially injured, as by chloro-
form, phosphorus or hydrazinc, there is a very notable
drop in the output of the substance in the feces down
to 20 or 10% or even a mere trace. The phthalein then
begins to be excreted by the urine. When the hepatic
lesion improves, the phthalein output in the bile in-
creases toward normal. The normal output of phe-
noltetrachlorphthalein in dogs is 45% of the amount
injected. The drop in phthalein output is always pro-
portional to the extent of liver injury. If the injury is
grave enough to produce death, the phthalein output
falls to zero. In some instances as the effects of the
hepatic injury are spontaneously repaired, there may
be an actual increase of phthalein output even above
normal, a hypersecretion of phthalein as it were.
Besides injury to the liver parenchyma by poisons,
the above investigators found that severe circulatory
Tests of Liver Function 59
disturbances artificially produced are followed by a
drop in the phthalein output. Actual destruction of
liver parenchyma produces similar results.
In experimental obstructive icterus there is, of course,
no phthalein output, since none of the drug reaches the
intestine. In experimental hematogenous icterus there
is no modification of phthalein output.
The experimental studies so far performed indicate
that the phenoltetrachlorphthalein will be valuable from
a quantitative as well as a qualitative standpoint in
the estimation of insufficiencies of liver function.
The clinical application of phenoltetrachlorphthalein
as a test for hepatic function was first worked out by
Rowntree, Horwitz and Bloomfield.
We owe to Rowntree the suggestion that the specific-
ity of the liver in excreting phenoltetrachlorphthalein
analogous to that of the kidney towards phenolsulphon-
phthalein would indicate that estimations in man of
the quantity of dye excreted by the liver after an in-
travenous injection ought to afford a practical clinical
method of determining the functional capacity of the
liver.
It will thus be seen that the phenoltetrachlorphthalein
test of Rowntree, Horwitz and Bloomfield is founded
upon rational theoretical considerations and that it was
subjected to a very rigid experimental investigation.
Technic of Phenoltetrachlorphthalein Test. — An
aqueous solution of the disodium salt is used. It is pre-
pared by placing 2.5 gms. of phenoltetrachlorphtha-
lein in a 200 c.c. Erlenmeyer flask with 5 c.c. of 2/n
NaOH solution and 45 c.c. of freshly distilled water.
This is boiled under a reflux condenser for 20 minutes.
The solution is filtered into a 100 c.c. flask. This solu-
tion is of 5% strength and is approximately isotonic
60 Manual of Vital Function Testing Methods
with blood. It is intensely purplish red in color. Since
the phthalein is precipitated by CO2 in the atmosphere
it will not keep more than a few days, hence requires
to be freshly prepared for use. The patient is given
two compound cathartic pills the night before the test
is applied.
In making the test 8 c.c. of the solution which will
contain about 400 milligrams of tetrachlorphthalein are
measured. It has been found that this amount is never
followed in normal persons with the excretion of any
dye in the urine and is sufficient to produce an intense
color in the final preparation of the feces. The 8 c.c.
is given intravenously as follows, of course under strict
aseptic precautions :
A funnel with properly connected intravenous system
is filled with freshly distilled water or salt solution and
the flow into the vein is started. When this is well
established, the phthalein solution is added. Fifty to
100 c.c. of water are used and the phthalein solution is
washed in with freshly distilled water until the fluid
entering the vein is clear. About a quarter of an hour
is required for the injection.
After the injection, the patient is given another pur-
gative, usually two compound cathartic pills, and this
dose is repeated the following morning if the bowels are
not running freely. The stools are collected for 48
hours in a covered vessel. The urine is collected for
24 hours.
The quantitative determination of the amount of
phthalein passed is made as follows: The total feces
collected are put in a wide mouth 2 liter bottle diluted
with water to 1 or 1.5 liters, according to quantity,
and the whole put in a shaking machine and well agi-
tated for from 5 to 20 minutes.
One-tenth of the total amount is immediately poured
Tests of Liver Function 61
off in a 1-liter flask. To this is added 5 c.c. of 40%
NaOH, which makes the mixture a dirty red color. The
flask is stoppered and thoroughly shaken. One hun-
dred (100) c.c. of the contents of this flask are placed
in a 200 c.c. flask, to which is added 5 c.c. of saturated
basic lead acetate. This decolorizes the mixture and
throws down a heavy precipitate which leaves a color-
less supernatant liquid. Five (5) c.c. of 40% solution
of NaOH are added, which produces the phthalein
color. More hydroxide solution may perhaps be needed
to bring out the full color, but excess should not be
used. The contents of the flask are then made up to
200 c.c., shaken, and the solution allowed to stand five
minutes. The supernatant fluid is clear and some can
be poured off for colorimetric examination. The read-
ing is made in a colorimeter similar to the one used in
testing kidney permeability with phenolsulphonphtha-
lein. The comparison solution is made by taking .4
c.c. of the original solution used for injection and
diluting it up to 1 liter plus sufficient NaOH to make
the deepest color. The per cent, of dye eliminated can
be read off on the instrument.
If the reading is low be sure that the maximum color
has been developed by adding a little NaOH again to
the 200 c.c. dilution above mentioned. NaOH must be
carefully added because excess will tend to render solu-
tions yellowish red instead of pure purple red.
In case the quality of color is unsatisfactory the
authors of the test recommend the following procedure :
After the addition of about 10 c.c. of 40% NaOH dilute
the feces mixture up to 1 liter. Take one-tenth of this
and add 5 c.c. of sodium hydroxide and water up to a
liter. One hundred c.c. of this is put in a 200 c.c. flask
and to it is added 5-10 c.c. or more of the following
solution :
62 Manual of Vital Function Testing Methods
CaCl2 90 gms.
Cone. NH4OH 10 c.c.
Water 50 c.c.
This brings out a good quality of color. Dilute up to
200 c.c. and allow to stand covered for some hours, per-
haps even 24. The supernatant liquid is then tested
in the colorimeter with the standard solution as above.
The lower limit of normal output is 20%.
A year's experience with the phenoltetrachlorphtha-
lein test of liver function has prompted a recent com-
munication from Chesney, Marshall, and Rowntree
(Jour. Amer. Med. Assn., 1914, LXIII, p. 1533) in
which these authors conclude that outspoken changes in
the liver can, in most cases, be demonstrated by the test.
It was found positive in advanced cirrhosis, in passive
congestion (cardiac liver) and in cancer and syphilis
involving the liver.
These authors recommend the association with the
phthalein test, of estimation of nitrogen partition in
the blood and urine, and fibrinogen estimation in the
blood serum.
They make the important general observation that
the information to be derived from tests of the liver
function does not compare in reliability with that ap-
plied to the kidney. Similar views have likewise been
expressed by other writers. The reason would appear
to be plain. We do not as yet understand the functions
of the liver regarded as a unit or dissociatively as we
do those of the kidney. The symptomatology of hepatic
insufficiency is not understood to an equal extent with
that of the kidney. However this may be, sufficient
progress has been made to afford ample congratulations
for the work of the past and an optimistic outlook for
Tests of Liver Fwnction 63
future developments.
Krumbhar 43 has recently stated as a result of his
researches and investigations that the phenoltetrachlor-
phthalein test of Rowntree, Horwitz, and Bloomfield
promises a greater value than all other tests so far
devised for estimating the functional capacity of the
liver.
43 N. Y. Med. Jour., 1914, c. 719.
CHAPTER II
TESTS OF KIDNEY FUNCTION
GENERAL CONSIDERATIONS
Historical. — In 1830 Hahn noticed after ingestion
of turpentine in gouty persons that the substance failed
to render the urine odorous as it was known to do in
healthy persons. In 1837 Rayer noticed the same
thing with regard to asparagus.
Clinicians for a long time have known that many
persons with nephritis can not take mercury, salicylates,
iodides, bromides and various other drugs without rap-
idly showing signs of intolerance. Todd wrote upon
this subject in 1857 and Roberts in 1865. Duckworth
and Bouchard in 1873 showed experimentally that
many drugs which normally pass quite readily through
the kidney, fail to do so in nephritis.
The first practical application of these facts was
made by Achard and Castaigne, who introduced methyl-
ene blue in 1897 as a direct test of the functional
capacity of the kidneys.
Physiological. — As well expressed by Blum the fun-
damental function of the kidney is its osmoregula-
tory power; its power to constantly maintain at an
unvarying point the molecular concentration of the
blood. This it does by removing a series of substances
whose accumulation in the organism would eventually
64
Tests of Kidney Function 65
produce serious and even fatal consequences. These
substances are removed in the urine.
Two general theories of urinary secretion have
existed, side by side, for several decades. They are
known eponymically by their originators and although
they have been added to or subtracted from in details
by a host of subsequent workers they yet stand as op-
posing schools of physiological interpretation. These
schools are known as that of Ludwig on the one hand
and that of Bowman-Heidenhain on the other.
According to Ludwig, the elimination of urine is a
simple process of physical filtration and diffusion. The
anatomical structure of the glomcrulus and the physio-
logical conditions existing therein, appearing to favor
the idea of filtration, Ludwig believed that water passes
through the epithelium of the capillary wall and the
glomerular epithelium as through a filter, carrying with
it sodium chloride and other inorganic salts and urea,
and that the diluted urine in its passage through the
uriniferous tubules becomes concentrated through loss
of water by diffusion into the more concentrated blood
and lymph.
According to the other theory, that of Bowman-
Heidenhain, the elimination of urine is fundamentally
a secretory act and not fundamentally a physical act.
It assumes that the glomerulus secretes water and inor-
ganic salts while the epithelial cells of the uriniferous
tubules secrete urea and the other specific constituents
of the urine.
We shall not attempt here either a historical or phys-
iological review of these theories. It may be said that
the majority of physiologists adhere to the more con-
servative and vitulistic hypothesis of Bowman-Heiden-
hain. The grounds for this belief, and, indeed, all of
the facts bearing upon both sides of this now classical
66 Manual of Vital Function Testing Methods
controversy, are properly to be found in any modern
text book of physiology.1
It is pretty generally agreed that whether by filtra-
tion or secretion, water leaves the kidney through the
glomerulus. Beyond this generally accepted fact there
is so little unanimity of opinion as to the exact place
where sodium chloride urea and the other solid constitu-
ents of the urine are eliminated, that it is impossible to
make any categorical statement with reference thereto.
Many attempts have been made to divide up the total
kidney function into categories or topical functions and
to locate these functions anatomically in parts of the
glomerulo-tubal structure. But it cannot be pretended
at the present time that any such differentiation has
been proven, certainly not to such an extent as to
justify deductions of great practical importance. This
question will, however, be more fully elaborated later.
The composition of the urine is far from simple. Its
chief constituents, apparently, are water, sodium chlo-
ride and urea. But besides these substances, urine con-
tains purine bodies (uric acid, xanthin, hypoxanthin),
creatinin, oxalates, glycuronates, phosphates, sul-
phates, various oxy-nitrogenous and fatty acids, and
the pigments urochrome and urobilin.
Nearly all of the nitrogen excreted from the body is
supposed to pass through the kidney. The total amount
of nitrogen eliminated in the urine in 24 hours is conse-
quently regarded as the most important index of proteid
metabolism. The actual estimate of total urinary nitro-
gen is usually done by the method of Kjeldahl (described
under Liver Tests, q. r.).
The total weight of nitrogen in the urine multiplied
by 6.25 gives the amount of protein broken down in
the body, since nitrogen forms 16% of the weight of
1 See Howell, Halliburton, etc.
Tests of Kidney Function 67
the protein molecule. The total amount of nitrogen
eliminated in 24 hours by a normal adult is between
14 and 18 grams, which corresponds to 88-117 grams
of protein.
The total nitrogen eliminated in the urine is divided
as follows: 1. Urea nitrogen; this averages 87.5% of
the total. 2. Ammonia nitrogen; this averages 4.3%
of the total. 3. Creatinin nitrogen, 3.6%. 4. Purin
nitrogen, variable.
Urea occurs in the urine as its chief nitrogenous
constituent (about 2% ). Since a normal adult secretes
1500 to 1700 c.c. of urine in 24 hours the amount of
urea eliminated will vary from 30 to 34 grams. Urea,
of course, is not manufactured by the kidneys, but is
merely eliminated by them. Urea is a normal con-
stituent of blood existing in that fluid in quantities
varying from .035 to .153%. If the kidneys are re-
moved or become impassible to urea this substance
accumulates in the blood.
Sodium chloride is the chief inorganic constituent of
urine, amounting to about 15 grams per day in a nor-
mal adult.
Under pathological conditions a variety of sub-
stances organic and inorganic may appear in the urine,
whose search and identification is a part of the routine
analysis conducted for clinical purposes.
A consideration of these substances belongs, of
course, to the domain of general clinical pathology.
The question of how far an ordinary urinary examina-
tion can serve to reveal the functional capacity of the
kidney will be taken up presently.
The kidney functionates normally at a point below
its maximum capacity, retaining unused a certain
amount of functional energy which constitutes its re-
serve. When this reserve becomes exhausted the func-
68 Manual of Vital Function Testing Methods
tional capacity of the organ will be irremediably dam-
aged unless it can recuperate. When uremia or edema
have appeared the amount of functional incapacity of
the kidney has become considerable. No special test of
function is required to discover it, perhaps, but even
under such conditions, it may be extremely useful to
determine just how far, in any given case, the deprecia-
tion of functional integrity has gone.
Classification. — There are various general plans by
which the functional state of the kidney may be in-
vestigated.
In the first place we may determine how far the kid-
ney is able to eliminate increased amounts of its normal
constituents, such as water, salt and urea.
In the second place we may select substances foreign
to the organism, but which are eliminated by the kid-
ney, and determine the rate and quantity of their excre-
tion. Iodide of potassium, lactose, phenolsulphon-
phthalein, phloridzin are examples of such substances.
Thirdly, the study of the blood will constitute an-
other avenue of approach to the problem of estimating
the function of the kidney, because one important result
of renal insufficiency will be the accumulation of sub-
stances in the blood which should be eliminated in the
urine. Among methods of this type may be mentioned
partitive estimations of nitrogen in the blood, particu-
larly incoagulable nitrogen. Such examinations will
often disclose an abnormal degree of accumulation or
retention of such products in the blood if a condition
of renal impermeability or insufficiency exists.
Thus it will be seen that all tests for kidney function
are based upon the broad principle that any deprecia-
tion of renal activity will be reflected in the urine on the
one hand and the blood upon the other. The urine will
contain less of its normal constituents than normally
Tests of Kidney Function 69
and less of any substance artificially eliminated by the
kidney, while the blood will show the effects of renal
inadequacy by disclosing an accumulation in the plasma
of substances which are normally excreted continuously
in adequate amounts.
All tests for renal function so far devised may be
satisfactorily divided into the three following cate-
gories :
1. The urine as an index of renal function — (a)
Urinalysis, (b) Physical and Biological characteristics.
2. The Blood as an index of renal function. 3. Elimi-
nation of foreign substances by the kidney as an index
of renal function.
In the following synopsis, we may see how the various
tests which have come to be used can be distributed
among the three classes :
I. Urinalysis as an index of renal function.
A. Urinalysis.
1. Estimation of water : experimental polyuria.
2. Estimation of sodium chloride : experimental
chloruria.
3. Estimation of urinary nitrogen ; urea, etc.
4. Estimation of urinary coloring matter.
5. Estimation of urinary diastase.
B. Physical and biological characteristics.
1. Cryoscopy of the urine.
2. Electrical conductivity of the urine.
3. Estimation of urinary toxicity.
IT. Studies of the blood as indices of renal function.
1. Estimation of blood urea and of incoagu-
lable (residual) nitrogen in blood.
2. Estimation of coagulation time.
3. Cryoscopy of the blood.
70 Manual of Vital Function Testing Methods
III. Studies of elimination of foreign substances by the
kidney as criteria of function.
A. Miscellaneous.
1. Potassium Iodide.
2. Phloridzin.
3. Hippuric Acid.
4. Lactose.
B. Dyes or colors: experimental urinary chromos-
copy.
1. Methylene blue.
2. Indico carmine.
3. Phenolsulphonephthalein.
I. THE STUDY OF UUINALYSIS AS AN INDEX OP RENAL
FUNCTION
A . Urinalysis
The urine represents, as it were, the concrete results,
almost the total results of renal activity. Inasmuch
as the entire urinary output of the kidney for any given
period of time may be readily collected it would seem
natural to assume that a chemical analysis would throw
all the light that is necessary upon the problem of renal
function ; but as a matter of fact, while it is true that
chemical analysis of the urine provides an adequate
insight into the amount of salts, of water and of urea
secreted by the kidney, it is not true that urinalyses
are sufficient to determine the functional capacity of
the organ. Gross anatomical and physiological disturb-
ances are often thus discovered and extremely important
information is thus derived concerning the diagnosis of
diseases of the kidney and urinary organs.
But this is a very different proposition from deter-
Tests of Kidney Function 71
mining thereby the functional capacity or incapacity of
the kidney, in the absence of evidence of gross organic
lesion. Even under apparently normal circumstances
the actual amount of the different urinary constituents
excreted may vary considerably. For example the
chlorides may be eliminated in excess and nitrogen re-
tained or vice versa. But such variations are not neces-
sarily dependent on anatomical lesions of the kidney
or even upon any disturbance of renal permeability.
If, in a given 24-hour specimen of urine, the figures
representing the elimination of the different important
constituents depart from the usual normal, we cannot
draw any absolute conclusions from this fact alone as
to whether the functional capacity of the kidney is
below or above normal. One reason for this is that the
chemical constitution of the urine is not dependent
alone upon kidney functional power but it is influenced
by a large number of extremely important extrarenal
factors, among which may be mentioned the intake of
food and fluids, the condition of the nervous system and
other organs, etc.
It is a well-known fact that before any important
conclusions as to nitrogen metabolism can be drawn
from the chemical constitution of the urine, these factors
must be taken into consideration, and if they are ade-
quately considered the task becomes complicated by
many necessary experimental refinements. The whole
question of body metabolism must be taken up. The
intake and output in every direction must be measured.
But even after this is done and it is demonstrated from
urinalysis that there is a deficit in nitrogen excretion,
the kidney function may be perfect and the nitrogen
simply retained in the tissues.
Variations in nitrogen elimination occur under so
many different conditions that interpretation is often
72 Manual of Vital Function Testing Methods
difficult or impossible. The same may be said with
regard to the output of other constituents of the urine.
Concerning the application of urinalysis to the inter-
pretation of kidney function it may be said that if the
figures are all consistently and invariably normal, the
kidney function is apt to be good; and if there is a
persistent and considerable departure from normal the
kidney function may be deficient.
But in order to be of real value, the tests, particularly
those regarding the nitrogen excretion, must often as-
sume the proportion of metabolism experiments, which
makes them impractical for clinical use.
Fortunately for the purposes of clinical medicine the
physician will not be called upon to consider the in-
tricate problems of nitrogen metabolism in his investi-
gations of renal function. His desire will be to know
the capacity of the kidney to do its work and fortu-
nately this object may be accomplished without recourse
to extremely elaborate and technical processes.
We are justified in expecting that under an average
regimen the kidneys will eliminate somewhere near the
average amounts of the urinary constituents. But
knowing how many and how variable the extrarcnal fac-
tors are which influence the absolute quantities of uri-
nary constituents eliminated, it will become apparent
that the results of a urinalysis, no matter how complete,
will require to be supplemented by other and better
means of determining the functional capacity of the
kidney.
1 . Estimation of Urinary Water as an Index of Renal
Function. Experimental Polyurla. Albarran's Method
The healthy kidney possesses the power in a high
degree to adapt itself to those tendencies, such as addi-
tion or subtraction of water to or from the circulating
Tests of Kidney Function 73
blood, which would tend to alter the molecular concen-
tration of the fluid. It quickly re-establishes both mo-
lecular and water equilibrium, thus maintaining an
equable osmotic tension in the blood and lymph. Super-
fluous water is, normally, quickly eliminated through
the glomerulus and reabsorption in the canaliculus is in-
hibited. When the supply of water to the organism is
deficient, the resorptive function of the canaliculus is
raised and a more highly concentrated urine is elimi-
nated. This function of the kidney may be properly
termed its diluting-concentrating power.
Since water secretion is a function of the glomerulus,
the diluting power of the kidney is a glomerular func-
tion. The functionally weak kidney is not only unable
to produce a highly concentrated urine but also unable
to elaborate a much diluted one. The concentrating
power of the kidney is a function of the epithelium of
the uriniferous tubes (canaliculus).
The diluting-concentrating power of the kidney suf-
fers in diffuse kidney disease in proportion to the
amount of parenchyma involved. In parenchymatous
nephritis the water secreting power of the kidney is
lowered ; in contracted kidney it is more or less retained.
The increased urine following an experimental pro-
vocative polyuria test differs from the increased secre-
tion following a heavy meal, since in the former case the
freezing point (A), molecular concentration, chloride,
phosphate and urea content (specific gravity) are all
diminished, while in the latter they are all increased.
With respect to continuity of function the diseased
organ possesses a greater constancy and invariability
in proportion to the amount of disease. The normal
kidney function tends to vary, that is, to adapt itself to
the constantly changing conditions in the organism.
The diseased organ has no such power.
74 Manual of Vital Function Testing Methods
The healthy kidney always functions below its maxi-
mum strength; always possesses, in other words, a cer-
tain reserve power which can be used under extraor-
dinary circumstances, such as great increase in water
and solid molecular intake. The insufficient kidney is
unable to meet these requisitions for added energy, and
responds but little if at all to extra stimulation. It
has lost its reserve.
As a corollary to the above it may be added that if
one kidney is diseased and the excretion from the two
organs be compared, the facts as above stated will
apply. The affected kidney is less able to respond to
adaptation requirements than the normal and the degree
of its failure to do so may properly be taken as the
measure of its incapacity.
For these reasons polyuria tests may be employed
with the view of conducting examinations upon the total
excretion or whenever necessary upon the excretion ob-
tained by ureteral catheterization from each organ
separately.
The Water Tests. — The provocative polyuria tests
are usually carried out with water. The tests should
be applied in the morning on an empty stomach. The
morning urine prior to the test should be measured and
examined for quantity and specific gravity, total sodi-
um chloride and urea elimination and perhaps cryo-
scopically. The patient is then given 500-700 c.c. of
mineral water or ordinary water. The urine should be
collected every half hour by voiding or catheter if the
total amount is to be examined (general renal function)
or by ureteral catheterization if the separate kidney
functions arc to be compared.
Under normal circumstances the polyuria appears
within the first half hour, reaching its maximum at this
time, and quickly sinking. The content of solids sinks.
Tests of Kidney Function 75
The freezing point (A) diminishes.
If the functional power of the organ is below normal,
the polyuria is delayed or does not occur and the
amount of variation from the normal may be taken as
a fair measure of the incapacity.
Straus-Grunwald Method. — The patient takes noth-
ing after 7 P. M. into the stomach. At 6:30 A. M. the
following morning a pint of water is ingested. The
night urine is collected; also that voided at 7, 8, 9, 10,
and 11 A. M. The amount and specific gravity of each
portion are recorded. The patient remains in a reclin-
ing position during the time of the test.
In normal cases an amount of urine is passed in the
first 3 hours equal to that which was drank. That is
by 10 A. M. at least a pint is voided. At 8 A. M. the
sp. gr. is lowest. Variations in the amount voided, time
required, and specific gravity will indicate abnormal
renal function.
The Diuretic Tests {pharmacological}. — Caffein,
diuretin, theophyllin (theocin), euphyllin and other
diuretic substances have been employed, but none of
these drugs has been found to possess much advantage
over the simple water test. The diuretic drugs appear
to increase the solid constituents of the urine as well as
the fluids. Blum, who introduced euphyllin, does not
recommend it because he has found a fall of blood pres-
sure follow its hypodermic or intravenous administra-
tion.
None of these tests have acquired a sufficient promi-
nence to justify their description.
While the so-called water or polyuria tests have
proven of some value in estimating relative function in
the separate kidney secretions, it is generally agreed
76 Manual of Vital Function Testing Methods
that they arc of much less importance in estimating
total kidney function.
The absolute quantity of urine voided varies very
greatly under normal and abnormal circumstances.
According to Rowntree and Fitz there is no constant
relation between the existence of polyuria or oliguria
and the condition of kidney function as shown by other
well-recognized tests. This applies to nephritic and
cardiac cases and to combinations of these.
The specific gravity of the urine in advanced nephri-
tis according to these authors is usually low.
2. Estimation of Sodium Chloride as an Index of
Renal Function
Ten to fifteen grams of sodium chloride are excreted
by a normal adult in 24 hours. The rapidity with
which the kidneys can excrete a considerable amount of
sodium chloride has been suggested and employed as a
test for renal function.
With regard to the method of so-called forced elim-
ination of sodium chloride, it must be freely granted
that the problem is a very difficult and complicated one.
If diminished secretion of sodium chloride always
indicated renal impermeability the problem would be
solved, but this is by no means the case. The tissue
fluids themselves, everywhere in the body perhaps, have
varying affinities for sodium chloride and a diminution
of chloride elimination may not signify a diminished
permeability of the kidney for salt but only an increased
retention of salt in the body.
Nevertheless there is, under normal circumstances, a
very close relation between the intake of sodium chloride
in the food and its elimination by the kidneys. So that
if an individual is placed for a period of time upon a
Tests of Kidney Function 77
regimen containing a low percentage of salt, the excre-
tion of that substance will become reduced to a lower
equilibrium. If now the quantity of sodium chloride
ingested be suddenly increased there will be an immediate
and proportionate increase in the amount secreted.
If there is no increase, it will be difficult to determine
whether there is chloride retention or defective excre-
tion so that the chloride test is seldom used alone as a
measure of renal function.
The exact situation in the glomerulo-tubular mechan-
ism where sodium chloride is excreted, is not known
with certainty. . According to the rather recent investi-
gations of Schlaycr and Takayasu and Von Monakow,
sodium chloride is excreted by the tubular epithelium,
or more exactly the excretion of salt following its ad-
ministration in amounts in excess of the usual daily
intake is accomplished by the tubules.
When large amounts of salt are ingested the excre-
tion, according to Schlayer, takes place in one of two
ways, depending upon the amount of water simultane-
ously absorbed. If the salt is given without extra water
it is almost entirely secreted within 24? hours without
diuresis, by an increased concentration of the urine. If,
however, it is given with an excess of water it is secreted
partially through increased concentration and partially
through diuresis.
If the vascular structure of the kidney is injured,
the ingestion of salt may be followed by marked diure-
sis, the salt all escaping in the urine in 21 hours with-
out the percentage content being increased. The spe-
cific gravity may be low and tends to remain at a fixed
point. To tin's combination of phenomena, Schlayer
gave the name vascular hyposthenuria, and, according
to his idea, the inability of the kidney to eliminate a
urine concentrated in salt is not due to tubular defect
78 Manual of Vital Function Testing Methods
but to a hypersensitive condition of the blood vessels
which allows the secretion of the salt in relatively large
amounts of water; in other words, the sensitive vessels
respond to the salt administration by the diuresis.
When the vascular injury is more marked the vessels do
not so react but respond with oliguria.
In severe tubular epithelial disease, however, the
quantity of salt eliminated is not raised by the adminis-
tration of salt. Here a urine of fixed low specific grav-
ity of moderate quantity is obtained. The salt is re-
tained. There is, according to Schlayer, a tubular
hyposthenuria.
These interesting findings reported by Schlayer and
his school have not been completely corroborated and
it does not appear to be agreed at the present time that
any absolute distinction between vascular and tubular
hyposthenuria can be founded upon the response of the
kidney to tests with sodium chloride.
In fact the number of extrarenal factors concerned
in the salt output are so many and so illy understood
that the salt test alone is considered of no value. But
when considered in conjunction with other functional
tests and with clinical findings in cardiac and renal
cases it may be of some diagnostic and prognostic
value.
In advanced nephritis there seems no doubt that
salt elimination is lessened to a certain extent but
if the patient has been on a salt-poor diet for a time
previous to the test, the tissues will retain salt when
administered regardless of the cardiorenal condition.
(Edema is the symptom in chronic nephritis which
is usually associated with the idea of chloride reten-
tion. It is not positively known just what factors are
concerned in this, or whether they are chiefly renal or
extrarenal.
Tests of Kidney Function 79
It is on the basis of the supposed connection be-
tween oedema and chloride retention that the now well-
known method of salt reduction in the treatment of
oedema in Bright's disease was introduced.
Technic of Sodium Chloride Test. Test of Alimen-
tary Chloruria.— The patient is placed on a diet con-
taining about 5 gms. of salt per day. After several
days or when the salt output is approximately con-
stant, 5-10 gms. of sodium chloride are given, dis-
solved in 125 c.c. of water. The quantity is taken in
three portions during the day. This is kept up for
four consecutive days. The daily output of chloride
is determined by the method described below. If the
patient is kept on a salt-poor diet, say 2.5 gms. daily
for some time previous to the test, it will be found that
excretion will always be lessened from a normal ten-
dency of the tissues to retain chlorides. For this
reason the best method consists in merely establishing
chloride equilibrium before the test is started.
Estimation of Sodium Chloride in the Urine. — The
principle of the test is that chlorides are precipitated
by solutions of nitrate of silver. Volhardt's method
with its various modifications is regarded as the most
accurate quantitative method of chloride estimation,
and is used when the exact amount must be known as
in metabolic experiments.
For all practical purposes Mohr's method will suf-
fice.2 The strength of silver solution used in the test
is such that 1 c.c. corresponds to .01 gm. of sodium
chloride. Such a solution contains 29.06 gms. of pure
fused silver nitrate to the liter. The technic of the
estimation is as follows :
2 The Lutke-Martius method is often recommended, see Sahli's
Diagnostic Methods, 1911, p. 455.
80 Manual of Vital Function Testing Methods
10 c.c. of urine previously freed from albumen are
put in an Erlenmeyer flask or porcelain capsule and
100 c.c. of water added and several drops of potassium
chromate solution, enough to produce a distinct yel-
low color.
The standard silver solution is added from a burette,
stirring until the reddish orange color which appears
first where the drop falls is distributed. The first
permanent orange color trace is the end of the reac-
tion. The operation should be repeated if necessary,
to make certain of results. The number of c.c. used
multiplied by .01 gives the amount of sodium chloride.
The results are a little high but near enough for prac-
tical purposes. A rough estimate of the amount of
chloride in the urine may be made as follows : To a
test tube of clear urine non-albuminous, add 10 drops
of pure HNO3 and one drop of AgNO3 (1 to 8). If
chlorides are normal or increased, the precipitate is
a compact ball which sinks to the bottom. If dimin-
ished, the ball is less compact. If much diminished,
only a cloud is produced without solid flakes. This
last represents a chloride content of \% or less.
3. Estimation of Urinary Nitrogen as an Index of
Hcnal Function
When functional tests of the liver were being dis-
cussed (z'.s.) considerable emphasis was laid upon in-
terpreting ureagenetic disturbance of that organ by
taking into consideration the different phases of nitro-
gen metabolism which arc so closely connected with
liver activity. It was then shown that a knowledge
of total nitrogen elimination in the urine and particu-
larly of the different forms in which the nitrogen is
eliminated, will shed light upon the condition of liver
Tests of Kidney Function '81
functional power.
The reasons for these assumptions were given in
their proper place, but it may be serviceable to refer
again to the fact that the liver is regarded as a very
important locus for the synthesis of urea in the body.
For this reason it is quite reasonable to suppose that
functional depreciation of the liver cells would be re-
flected to an appreciable extent by the quantitative
relative variations of the nitrogen constituents of the
urine.
When now we come to the significance of nitrogen
elimination in the urine to disorders of kidney activity,
it will be necessary to remember that the kidney has
nothing to do whatever in a specific way with the nitro-
gen metabolism. Its only function is to excrete the
nitrogenous waste products which are brought to it
by the blood. That urea is not manufactured by the
kidney is proved by the fact that if blood is perfused
thro an isolated kidney, no urea is formed even tho
substances such as ammonium carbonate, from which
urea is readily produced, are added to the blood. It
is well known that if the kidneys are removed in ani-
mals or their function paralyzed, urea will continue to
accumulate in the blood as long as the animal sur-
vives.
No physiological fact is bettor known than that
the relative amount of urea nitrogen in the urine varies
directly with the amount of protein food ingested.
Other nitrogenous constituents of the urine, the purin
bodies and creatinin, are unaffected by the intake. This
suggested to Folin that most of the urea in the urine
may come directly from food proteins which, having
been hydrolyzed in the intestine into amino acids, are
absorbed and further hydrolyzed and oxidized and
the nitrogen constituent immediately eliminated as
82 Manual of Vital Function Testing Methods
urea. The liver has most to do with this process though
the urea forming function of this organ is known to
be shared by some other tissues since even after the
removal of the liver some urea is formed.
These physiological principles being agreed upon,
it is easy to appreciate that the question of the estima-
tion of urinary nitrogen as an index of renal function
will be practically confined to the estimation of the
amount of urea eliminated under normal circumstances,
the patient being on a fixed diet, with an estimation
of the power of the kidney to eliminate more urea when
the proteid intake is increased, or when urea itself is
ingested. The question of nitrogen accumulation in
the blood as an index of renal insufficiency will be dis-
cussed below, when the study of the blood as an index
of renal function is considered. It may be admitted
here that the study of the partition of nitrogen and
particularly quantitative estimations of urea and in-
coagulable nitrogen in the blood serum are of much
greater significance in the estimation of kidney function
than the same or related studies applied to the urine.
Diminished and Delayed Excretion of Urea. — This
is an old criterion of functional renal power. The
physiological principles upon which it is based have
just been given. In order that this test of renal func-
tion shall be really conclusive, the patient must be put
for some days upon a fixed regimen in which the amount
of protein is definitely known.
The feces and urine must be examined to determine
what part of the nitrogen has escaped in both. In
carrying out such procedure, the experiment rises to
the dignity of a metabolic investigation and requires
great care and patience besides considerable technical
skill. For this reason such a procedure cannot be re-
garded as adapted to clinical use.
Tests of Kidney Function 83
Without the above precautions the value of ordinary
routine urea estimation in the urine as a criterion of
renal function is extremely doubtful.
If the dietary conditions can be reasonably con-
trolled and a perfectly and persistently normal output
of urea results, the renal function is at least equal to
the ordinary demand of that person. If the proteid
intake is increased and the urea excretion undergoes
a corresponding and immediate rise, it may be con-
cluded that the reserve force of the kidney is not
materially damaged.
But negative results need not necessarily be taken
to indicate a defect of renal function because of the
fact that digestive disturbance and diminished liver
function will cause the same thing to occur. If these
extrarenal factors of error can be eliminated, then a
diminished output of urea may become a valuable and
reliable index of renal inadequacy.
Under a later chapter, concerning study of the blood
as an index of renal function, it will be shown that the
quantitative estimation of nitrogen partition in that
fluid is a much more reliable test of renal functional
power than urea estimations in the urine.
Repeating what has been given under general con-
siderations it may be stated that a normal adult se-
cretes from 30 to 34 grams of urea in the urine every 24
hours.
The simplest quantitative test for urea in urine is
that of Marshall, which has been described (v. s.}.
This method is so simple and so accurate that every
clinician should familiarize himself with it.
Forced Urea Elimination. Provocative Urea Test of
McKasky. — Technic- Thirty grams of urea dissolved
in four to six ounces of water are given with a small
84 Manual of Vital Function Testing Methods
breakfast, such as a cup of gruel. Follow this with
four or five ounces of water to assist diuresis. The
urine is collected every two hours for twenty-four hours,
beginning two hours before the urea is given, so that a
standard for comparison may be had, to determine the
amount of increase. Quantitative determinations of
urea are made in the different specimens at the end of
the time.
The maximum excretion, its time, incidence, and the
curve for the 24 hours, is thus determined.
Under normal conditions there is a sharp rise in
urea excretion in the second two-hour period. When
the kidney function is deficient the sharp rise is absent
or is much delayed.
The only factor liable to disturb the interpretation
of this test is gastric stasis.
Theoretically, the provocative urea test should be
useful. There does not appear to be any reason why
the urea could not be injected in smaller amounts, di-
rectly into the circulation, in which event the liability
of misinterpretation through retention in the stomach
would be avoided. The test has not been subjected to
any clinical examination, so far as I know.
4- Estimation of Urinary Coloring Matter as Test
of Renal Function
Thudichum's Test. — This test is now of only histor-
ical interest. It was proposed by its author on the
clinical ground that in many chronic kidney diseases
the urine becomes distinctly paler in color. Therefore
careful quantitative estimations of color excretion
might be of value as an early sign of renal imper-
meability. But unfortunately for the value of the test,
the quantity of coloring matter excreted in the urine
Tests of Kidney Function 85
depends upon a great variety of factors (liver, intes-
tine, food, etc.), of which the most insignificant of all
is perhaps renal permeability.
5. Estimation of Urinary Diastase as an Index of
Renal Function
Wohlgemuth's Test? — Te clinic. After neutraliza-
tion urine is placed by means of an accurately grad-
uated pipette in a series of twelve test tubes, the amount
decreasing from .6 c.c., .5 c.c., A c.c., to .1 to .09 c.c.,
.08 c.c., to .04 c.c. A sufficient quantity of 1% sodium
chloride solution is then added, to bring the amount of
fluid in each tube up to 1 c.c. In order to more readily
obtain the fractional quantity of urine required, 1 c.c.
of the urine may be diluted up to 10 c.c. and from this
diluted urine the required measures may be taken.
To each tube is added 2 c.c. of a 1-1000 solution of
freshly prepared soluble starch. The tubes are im-
mersed in a water bath at 38° C. for 30 minutes, after
which they are placed in cold water for 3 minutes.
To each tube is then added sufficient 1/50 normal
iodin solution to elicit a permanent color, violet or blue
occurring where digestion is not complete.
The tube in the series immediately preceding incom-
plete digestion of the starch indicates the diastase
content of that particular urine. From this is calcula-
ted the diastasic activity represented by 8. 8 is expressed
as the number of c.c. of 1/10% starch solution which
can be digested by 1 c.c. of urine. This test can be
applied to the whole urine or to the samples obtained
unilaterally by ureteral catheterization. The diastase
test is particularly adapted to unilateral estimations
of kidney function applied to urine obtained by ureteral
3 Lancet Clinic, 1913, CX, p. 164.
86 Manual of Vital Function Testing Methods
catheterization. But altho it is capable of indicating
in the majority of cases which is the diseased or more
diseased kidney, in the opinion of most genitourinary
surgeons it is not necessary and adds nothing to the
information obtained from the phthalein test or urea
estimations, which latter are operations somewhat more
easily performed.
The diastase test has never been used to any extent
in estimating total functional capacity of the kidneys.
B. The Study of Physical and Biological Character-
istics of the Urine as Criteria of Renal Function.
There are three tests which come under this category :
1. Cryoscopy or determination of the freezing point
of the urine.
2. Electrical conductivity of the urine.
3. Determination of urinary toxicity.
1. Cryoscopy of the Urine. Significance for Estimat-
mg Renal Function. Von KoranyVs Test
The theoretical bases upon which this test is founded
are of extreme interest both from physical and physio-
logical points of view but naturally cannot be com-
pletely considered here.
The freezing point of distilled water is zero. The
freezing point of any solution is below zero, and the
depression of freezing point below zero is proportionate
to the molecular concentration of the solution. Conse-
quently the freezing point of a solution is a measure of
its molecular concentration. It is also a measure of its
osmotic pressure.
The freezing point of a solution is independent of
the nature, size, and molecular weight of the dissolved
Tests of Kidney Function 87
molecules and is only dependent on their number. The
specific gravity of a solution is on the contrary de-
pendent upon the nature and molecular weight of the
dissolved molecules. Solutions of similar molecular con-
centration have the same freezing point and the same
osmotic pressure but not necessarily the same specific
gravity.
By cryoscopy the molecular concentration of urine
and blood can be estimated and in this way a certain
insight into the functional power of the kidney can be
obtained, since the global function of this organ is to
regulate the osmotic pressure, or, what is the same,
the molecular concentration of the blood. One can
obtain from an estimation of the lowering of the freez-
ing point of the urine below the zero of distilled water
a somewhat more adequate idea of the functional ca-
pacity of the kidney than from the specific gravity.4
Of two urines of equal specific gravity the one with
the lower freezing point comes from the better func-
tioning kidney.
Investigations have shown that the freezing point
of the urine ( A ) in health varies between rather wide
limits (A= —.90° to — 2.30°) and that it is to
some extent affected by miscellaneous factors of extra-
renal nature.
Altho much was expected originally from the de-
termination of the freezing point (cryoscopy) of urine
in estimating the integrity of renal function, it is not
depended upon to any great extent at present.
The molecular concentration of the blood hence its
freezing point ( 8 ) is much more constant than that
of the urine. It is supposed to remain somewhere
4 Some authors do not agree that cryoscopy is superior to sp.
gr. estimation in determining renal permeability, v. Sahli, Diag-
nostic Methods, 1905, p. 551.
88 Manual of Vital Function Testing Methods
near — .56°. It was thought that insufficiency of renal
function by allowing the accumulation of molecules
in the blood which should be excreted would raise the
molecular concentration therein or, in other words,
lower the freezing point. Unfortunately, experience
has not confirmed the hopes of those who thought that
cryoscopic examinations of the urine and blood would
solve the great problem of estimating the renal func-
tions and the method is not extensively used.
The relation between the lowering of freezing point
of blood (& ) and urine (A) was proposed by Dreser as
a measure of the work done by the kidney and by
Bernard as the basis of a sort of mathematical con-
ception of the eliminatory power of the kidney. The
g
formula — X V = R will represent the molecular
A
elimination of the kidney according to this conception.
A represents the freezing point of urine, 8 that of
the blood, V the quantity of urine in 24 hours. In
normal cases R varies from 3000 to 5000, whereas in
renal insufficiency these numbers are considerably re-
duced. Unfortunately all attempts to reduce our con-
ceptions of organic function to mathematical terms
have not been eminently successful.
Technic of Cryoscopy.- — The technic of cr}roscopy
is not especially complex but requires a certain ap-
paratus for its performance. It is usually carried
out in the laboratory and has never become a routine
clinical procedure. Only the necessary outlines of
the method need here be given since those who desire
to master it can easily refer to numerous texts in
which the technique is minutely described.5
The technic of cryoscopy is carried out with Beck-
B Consult in this connection Wood's Chemical and Microscopical
Diagnosis, 1909, p. 61; Sahli's Diagnostic Methods, p. 546.
Tests of Kidney Function 89
mann's freezing apparatus carrying a special ther-
mometer. The freezing mixture is made of ice, water
and salt. In the freezing mixture is plunged an ordi-
nary thermometer and a mixer, thus enabling the tem-
perature to be kept at a fixed point ( — 3° to — 5°).
Thro an opening a tube containing the special ther-
mometer immersed in the liquid to be frozen can be
immersed in the freezing mixture. The estimation
of the exact points of freezing is not difficult and
usually the operation can be performed in its entirety
in 15 or 20 minutes.
2. Electrical Conductivity of the Urine
The electrical conductivity of the urine in health
and disease was first studied by Turner. The electrical
conductivity is estimated in ohms of resistance and
depends upon the number of ions of salts dissolved
in the urine. The method measures, in other words,
the amount of salts or mineral content of the fluid.
The Kohlrausch method of performing the test, which
is the one usually employed, requires a whetstone
bridge, a resistance box, telephone and cells, besides
other paraphernalia, and partly on account of this
complexity and also the fact that the practical results
obtained are meagre, the test has never come into gen-
eral use.
3. Estimation of Urinary Toxicity as a Test of Renal
Function
Bouchard's Test.- — It was for a long time believed
that the toxicity of the urine was proportional to the
functional power of the kidney. Urine of human be-
ings produces symptoms of poisoning and death when
90 Manual of Vital Function Testing Methods
injected intravenously into lower animals. Bouchard
developed from this fact a method of testing renal
function 6 by determining the quantity of a 24-hour
specimen of urine, required to kill a kilogram of lower
animal. Bouchard established a so-called urotoxic co-
efficient which was that quantity of poison elaborated
by every kilogram of body weight of the person whose
urine was tested.
The same objections exist with respect to the theo-
retical basis of this test, as in the case of chemical
urinalysis previously discussed, namely that so many
factors besides renal sufficiency or insufficiency enter
into the production of results that the test becomes
devoid of scientific value. It simply shows the toxicity
of a given urine injected intravenously into a given
animal and by no means reflects the actual functional
power of the kidney thro which it was derived. The
test is quite complex and has been abandoned.
II. STUDIES OF THE BLOOD AS CRITERIA OF RENAL
FUNCTION. ESTIMATION OF BLOOD UREA AND OF IN-
COAGULABLE (RESIDUAL) NITROGEN IN BLOOD
When it is considered that a major portion of the
nitrogenous waste of the body makes its escape thro
the kidney by way of the urine, it becomes evident that
a diminution of the functional capacity of these organs
must often result in an accumulation of nitrogenous
products of metabolism in the blood.
Such an idea is very old and as long ago as 1821
Prcvost and Dumas " reported an increase of urea in
the blood after extirpation of the kidneys in animals.
8 Also toxopexic liver function (y. .?.).
1 Quoted by Schondorff in Pfliiger's Archiv f. de ges. Physiol.,
1899, LXXIV, p. 307.
Tests of Kidney Function 91
The clinical importance of their experiments was rec-
ognized by Bright in his observations upon nephritis
in 1836.8
After B right's time it became well recognized that
in chronic nephritis there may be a tendency toward
accumulation of nitrogenous matters in the blood, but
the quantitative study of nitrogen retention was forced
to await the development of accurate chemical methods
of investigation. In this place we can speak only of
modern technic.
The development of accurate technic in nitrogen
estimation of the blood and its application to clinical
medicine is a subject which has been perfected only
since the beginning of the century. Ascoli,9 Strauss 10
and others showed that in many cases of chronic
Bright's disease nitrogenous matter accumulates in the
blood and the increase is more marked as death ap-
proaches. Muller 1: showed that in outspoken uremia
the accumulation becomes proportionately more
marked.
Obermayer and Popper 12 first laid stress upon the
increase of incoagulable nitrogen in the blood serum
in uremic states and found that in this incoagulable
nitrogen increase, urea plays the most important part.
Hohlweg13 substantiated these facts and altho he
showed that an increase of urea in the blood is not
necessarily pathognomonic of uremia, nevertheless its
accumulation therein may be regarded as an evidence
and to some extent at least as an index of the function
s Guy's Hosp. Rep., 1836, I, p. 358.
u Pfluger's Arch. f. de ges. Physiol., 1901, LXXXVII, p. 103.
10 Chronisch Nierenenzundung; ihrer Eimvirkung auf die Blut-
flussigkeit, etc., Berlin, 1902.
"Verb. d. Deutsch. path. Gesell., 1904-5, VII to IX, Erg. 80.
"Zeit. f. klin. Med., 1909, LXVII, p. 332.
MDeut. Archiv f. klin. Med., 1912, CIV, p. 216.
92 Manijuil of Vital Function Testing Methods
of the kidneys.
Widal,14 however, believes that the amount of urea
retention is an actual quantitative index of renal func-
tion and that the severity and prognosis of a given
case may be predicated upon the basis of such findings.
Widal's clinical method of estimating urea in the blood
is not commonly used in this country as it gives, ac-
cording to Rowntree and Fitz, an error of 10 to 60%
and is therefore useless as a quantitative method.
The estimation of urea in blood serum has remained
until recently a rather difficult chemical operation, but
of late one or two practical clinical methods have been
devised which render the test much more practical.
Marshall's method appears to be the simplest and
most practical of these and will he described in full
(».*.).
The normal figures for the elimination of urea are
between .300 to .500 gm. per liter of serum.
The simplest methods by which the total incoagulable
nitrogen in the blood serum may be determined still
remain even more difficult and complicated than those
of urea estimation, especially since they involve in the
end a nitrogen determination by Kjeldahl's method. A
fair laboratory equipment is therefore necessary.
The most practical method of determining the in-
coagulable nitrogen seems to be that of Hohlweg
and Meyer which has been modified by Morris ]r> in
this country. This method will be described below.
The normal figures for total incoagulable nitrogen in
the blood serum are .500 to .600 gm. per liter.
The recent work of Folin and Denis indicates that
a urea concentration in the blood of .5 gm. and total in-
14 Bull, et Mem. Soc. Med. d. hop. de Paris, 3, 1011, XXXTI, p.
627.
15Archiv. of Int. Med., 1911, VIII, p. 457.
Tests of Kidney Function 93
coagulable nitrogen content of .6 gm. per liter, which
was formerly considered normal, is too high an esti-
mate. They found the normal urea concentration .13
gm. and incoagulable nitrogen .26 gm. per liter. In
their experience no great prognostic significance is to
be attached to urea concentration of less than .55 gm.
per liter and incoagulable nitrogen less than .50 gm.
per liter. Greater concentration than this, especially
if the freezing point of the serum drops lower than
— 60 are of considerable prognostic significance.
In pure passive congestion Rowntree states that he
has never seen the rest nitrogen in the blood serum
higher than .63 gm. per liter.
Marked accumulation of incoagulable nitrogen or
of urea in the blood is now regarded as a valuable
evidence of renal insufficiency and in cases of nephritis
it is an unfavorable prognostic sign.
The relation between non-protein (incoagulable) ni-
trogen retention in the blood and the excretion of phe-
nolsulphonphthalein has been studied in experimental
uranium nephritis by Frothingham, Fitz, Folin and
Denis.16 These investigators found that the results of
the two tests paralleled very closely. For this reason
and because in clinical studies the same parallelism has
been found to obtain, these two tests have come to be
considered as among the best for conjoint use.
It will now be necessary to give the simplest, most
practical and accurate methods by which the clinical
investigator may determine the two important phases
of the blood which have been discussed, namely, the
amount of urea in the blood and the amount of in-
coagulable or rest nitrogen contained in the same fluid.
The chemical operations which are used at present for
these purposes are simple enough to bring about their
"Arch, of Int. Med., 1913, XIII, p. 2L5.
94 Manual of Vital Function Testing Methods
frequent use in the clinic. The best method of quickly
and accurately determining the amount of urea in the
blood is that of Marshall. The incoagulable or rest
nitrogen in blood serum is usually determined by either
of two general methods, that of Hohlweg-Meyer and
that of Folin and Denis. The details of these three
important methods will now be given.
Marshall's Method for the Determination of Urea
in the Blood.17 — The blood is drawn in the usual man-
ner and allowed to stand on ice until clotting is com-
plete. As shown below, the urea content of the serum
does not change after standing even for three or four
days ; the blood can, therefore, be kept on ice over
night, if desired.
Two equal portions of the serum are measured into
ordinary test tubes, 1 c.c. of the soy bean extract lf
added to one tube, and about 0.5-1.0 c.c. of toluene to
each. If sufficient serum is available, 10 c.c. portions
should be used; however, perfectly satisfactory results
can be obtained by using 5 c.c. or even 3 c.c. portions
of the serum. The tubes are tightly stoppered and
allowed to remain at room temperature until the con-
version of the urea into ammonium carbonate is com-
plete. Generally, they are allowed to stand over night,
altho four to five hours is usually amply sufficient for
the completion of the reaction. The contents of the
tube containing the serum and extract arc transferred
to cylinder A (see illustration), and washed in with a
"Jour, of Biol. Chem., 1913, XV, No. 3.
18 The preparation of the soy bean extract is as follows: Ten
grams of finely ground soy beans are treated with 100 c.c. of water
and allowed to stand with occasional agitation for one hour. 10 c.c.
of ^ hydrochloric acid are added and the mixture allowed to
stand about fifteen minutes longer. It is now filtered and pre-
served with toluene. Such a solution is perfectly satisfactory for
use at least five or six days after its preparation.
Tests of Kidney Function
95
very small amount of water (not more than 5 c.c.).
Two grams of sodium chloride, an equal volume of al-
cohol and a layer of kerosene oil are added to the cylin-
der. The contents of the other tube are transferred to
cylinder B, and treated in exactly the same manner. 25
c.c. of -— hydrochloric acid and about 25 c.c. of water
are placed in each of the 200 c.c. Erlenmeyer flasks
used for the absorption of the ammonia. The different
parts of the apparatus are now connected and about 0.5
gram of sodium carbonate added to each cylinder. A
rapid air current is passed through the apparatus until
all the ammonia has been removed from the cylinders.
With a good suction pump, one hour suffices. The
excess of acid in the absorption flasks is titrated with —
OU
sodium hydroxide and alizarin sodium sulphonate. The
amount of acid neutralized in the flask attached to
96 Manual of Vital Function Testing Methods
cylinder B corresponds, of course, to the ammonia 10
present in the serum, while the amount used in the other
two flasks represents the urea plus the ammonia. The
difference corresponds to the urea in terms of r-r
OU
hydrochloric acid, and multiplied by 0.0006 gives the
urea in grams present in the amount of serum taken
for the determination.
Details in Connection With the Apparatus and De-
termination.— 1. On account of the large quantity of
protein in serum, it is advisable to use both alcohol and
kerosene to prevent foaming.20
2. The tubes C and C' are ordinary calcium chlor-
ide drying tubes packed loosely with cotton. These
in conjunction with the bulbs prevent any splashing
or mechanical transmission of the alkali into the ab-
sorption flasks. While the bulbs are probably not
absolutely necessary, they are convenient in keeping
the cotton filters dry.
3. For the better absorption of the ammonia, the
tubes in the Erlenmeyer flasks are closed at one end,
and pierced with six or seven small holes, as suggested
by Folin.'1 Even with this device one absorption flask
is not always sufficient to completely absorb the am-
monia. Two flasks are always used for safety in con-
nection with the urea determination; however, since
from the serum alone only a very small amount of am-
monia (corresponding to 0.10-0.70 c.c. of -^ HC1)
oU
19 We can, however, place no value on this as a determination of
the true ammonia content of the blood, for on standing even a
few hours tin1 blood develops much more ammonia than the
original amount (Folin).
!0 This has been pointed out by Folin, in connection with the use
of the air current method for determining ammonia in blood.
(Zeitsch. f. physiol. Chem., XXXVII, p. !(>.>, l!)OJ-0.'i.)
-'Jour. Biol. Chem., XI, p. 493, 19U.
Tests of Kidney Function 97
is ordinarily obtained, one absorption flask is here
sufficient.
4. A layer of toluene is placed on the liquid in the
absorption flasks, for. due probably to the alcohol car-
ried over by the air current, considerable foaming some-
times occurs. If not prevented, this results in a loss
of a portion of the contents of the flask.
5. The bottle contains dilute sulphuric acid to free
the air from any traces of ammonia before passing it
through the apparatus.
6. No correction is necessary for the ammonia de-
rived from the 1 c.c. of soy bean extract used, as the
amount obtained from this source is unappreciable.
Another method also suggested by Dr. Marshall is
to draw 5 c.c. of blood from a vein with a hypodermic
needle, into a 5 c.c. pipette, and immediately transfer
the specimen to a test tube, containing 1 to 2 c.c. of
1% sodium oxalate solution. To this is added 25
milligrams (one tablet) of Urease-Dunning, the tablet
having been previously crushed and dissolved in 5 c.c.
of water. This mixture is allowed to stand until the
urea of the blood is decomposed; at ordinary room
temperature, one-half an hour is usually sufficient; it
is better, however, to place the test tube in a beaker
of water at 30° to 40° C. for one-half hour. After the
urea has been changed, the contents and sufficient wash-
ings of the tube are transferred to a cylinder. The
ammonia is then removed by a current of air collected
in the - hydrochloric acid and titrated with
50 50
sodium hydroxide.
Calculating Urea Content.— As the purpose in using
Urease-Dunning is to convert the urea present in a
specimen into an easily estimated substance — am-
monium carbonate — and as the amount of this salt
98 Manual of Vital Function Testing Methods
produced from this source, by the enzyme, is indicated
by the increased alkalinity of the specimen to methyl
orange, it is obvious that the quantity of standard
hydrochloric acid required to exactly neutralize the
contents of the flask containing urease, less the quan-
tity required for the control specimen, corresponds to
the ammonium carbonate formed by the conversion of
the urea originally present in the specimen.
By the following equation:
NH2 ONH4
CO +2H20=CO
NH2 ONH4
it may be calculated that 60 grams of urea would be
converted, by urease, into 96 grams of ammonium car-
bonate, which amount would require 72 grams of hydro-
chloric acid to neutralize it.
As this quantity (72 grams) of hydrochloric acid
is contained in 20,000 c.c. of decinormal (N/10) hydro-
chloric acid solution and is equivalent to 60 grams of
urea, as represented by 96 grams of ammonium car-
bonate, it follows that one twenty-thousandth of this
quantity or 1 c.c. of decinormal hydrochloric acid
would be equivalent to one twenty-thousandth of 60
grams = 3 milligrams (60 -f- 20,000 = .003), there-
fore each c.c. of decinormal hydrochloric acid required
to neutralize an enzyme-treated specimen, that is in
excess of the number of cubic centimeters required to
neutralize the control specimen, represents three milli-
grams of urea, and, as the 5 c.c. specimen is the one
two-hundredth part of a liter, it will be only necessary
to multiply the number of c.c. of the decinormal hydro-
chloric acid solution, in excess of the control's require-
ments, by the factor .6 (.003 X 200 = .6) to ascer-
tain the urea per liter, when estimating the daily output.
Tests of Kidney Function 99
1. Technic of Estimating Total Incoagulable or So-
catted Rest or Residual Nitrogen in the Blood Serum
Two methods are in common use. These are Morris'
modification of the Hohlweg-Meyer method, and the
method of Folin and Denis.
Morris' Modification of Hohlweg-Meyer Method. —
To 10 c.c. of blood serum obtained by venipuncture or
otherwise, in a 300 c.c. Erlenmeyer flask is added a
reagent consisting of equal parts of 1% acetic acid
and a 5% solution of monocalcium phosphate, until the
reaction is acid to litmus but neutral to Congo red.
The volume is brought up with distilled water to 80
c.c. and 80 c.c. of saturated solution of sodium chloride
are poured into the flask.
The mixture is boiled to precipitate the coagulable
proteins, and the filtrate, from which the proteins have
been shown to be completely removed, subjected to a
nitrogen determination by Kjeldahl's method.
For a description of the technic of Kjeldahl's method
see page 28.
Folin and Denis Method. — This method is considered
at the present time as the most practical way of quan-
titatively determining the amount of rest nitrogen in
the blood. It will be given in the words of its authors
from their communication published in 1912. (Jour.
Biol. Chem. 1912, XI, 527, Ibid. 1913, XIV, 29.)
Method for Drawing Blood. — "Before going into de-
tails of the chemical work it would seem worth while
to describe our method of drawing blood because sc
far as we have been able to learn it is somewhat different
from the procedures employed by physiologists and
because we believe it to be expeditious, neat and exact
and therefore particularly suitable for quantitative
work.
100 Manual of Vital Function Testing Methods
"We use neither cannulae nor syringes but simply
hypodermic needles and pipettes. The needles are about
1 mm. in diameter, and about 25 mm. long. They are
immersed in a dilute solution of vaseline in ether and
then allowed to drain and dry on a clean paper for
a few minutes before being used. (This does not apply
of course to the drawing of human blood when the
needles must be thoroughly sterilized.) An adequate
supply of these needles is kept on hand so that we do
not need to use any needle more than once in any one
experiment. The needle is attached to the tip of a
2 or 5 c.c. pipette by means of a short piece of narrow
pure gum tubing. A small pinch of powdered potas-
sium oxalate is introduced into the upper end of the
pipette (which must be clean and perfectly dry) and
is allowed to run down into the tip and the needle. The
other end of the pipette is connected with a rubber
tube which in turn connects with a mouthpiece con-
sisting of a short tapering glass tube. Close to the
pipette the rubber tube carries a pinchcock.
"To draw the blood insert the needle in the vein or
artery and regulate the flow of the blood by means of
the pinchcock and by suction. The exact quantity of
blood desired is thus obtained without any waste and
without clotting."
Isolation of Non-Protein Nitrogenous Constituents.
— "To separate the non-protein nitrogenous constitu-
ents from the protein materials we make use of pure
(acetone-free) methyl alcohol and an alcoholic solu-
tion of zinc chloride. Ordinary methyl alcohol cannot
be used because the impurities in it, particularly the
acetone, combine Avith more or less of the urea so that
it escapes decomposition in the subsequent treatment
and is not quantitatively recovered. We have satisfied
ourselves by means of determinations on pure urea
Tests of Kidney Function 101
solutions that the presence of acetone results in a loss
of urea.
"As soon as the blood is drawn it is transferred into
measuring flasks half filled with methyl alcohol and the
flasks are then filled up to the mark with methyl alcohol
and vigorously shaken. Two cubic centimeters of blood
are diluted to 25, while for 5 c.c. of blood use 50 c.c.
flasks. At the end of two hours, or as soon after that as
is convenient, the contents of the flasks are filtered
through dry filters. To the filtrate are then added two
or three drops of a saturated alcoholic solution of zinc
chloride and after standing for a few minutes the mix-
ture is again filtered thro a dry paper. The zinc chlor-
ide brings down an appreciable precipitate and the last
traces of coloring matters so that when the second
filtration is made, a perfectly colorless filtrate is ob-
tained. 5 c.c. of these filtrates, corresponding to 0.4
or to 0.5 c.c. of blood, depending on whether 2 or 5
c.c. of blood were drawn, are taken for each determina-
tion.
"The precipitation procedure described above is the
one which we ordinarily use. There are objections to
it. We are not certain that protein-like materials may
not escape precipitation by this as by every other
method and we do know that the filtrate does not con-
tain all of the non-protein materials. When relatively
large quantities (equivalent to 100 mgm. of nitrogen
per 100 c.c. of blood) of creatine or asparagine are
added to blood and treated as described above there is
invariably an appreciable loss of material. To over-
come this loss we have tried to triturate and wash the
first alcoholic precipitate with methyl alcohol, and with
some substance as, for example, with glycocoll, urea
and acetamide, we are thus able to get practically
quantitative results, while with others, such as creatine,
Manual of Vital Function Testing Methods
asparagine and tyrosine, we still do not get quite all.
Moreover, such trituration and washing docs leach out
a small amount of the coloring matters of the blood
so that except for special experiments with less soluble
substances we consider the simpler procedure rather
more satisfactory.
Determination of the Total Non-Protein Nitrogen. —
"To determine the non-protein nitrogen of the blood
5 c.c. of the alcoholic filtrate is transferred to a large
Jena test tube. One drop of sulphuric acid, one of
kerosene and a pebble are added and the methyl alcohol
is driven off by immersing the test tube in a beaker
of boiling water for five to ten minutes. When the
alcohol is removed 1 c.c. of concentrated sulphuric
acid, a gram of potassium sulphate, and a drop of
copper sulphate solution are added and the mixture is
boiled, cooled and diluted.
"From this digestion mixture the ammonia is removed
in the usual manner. It is, however, not collected di-
rectly in a measuring flask (as in urine analysis) but
in a second large test tube previously charged with 1
c.c. of — acid added to 3 c.c. of water. The reason
for this variation is that 0.4 to 0.5 c.c. of blood con-
tains only 0.1 to 0.2 mgm. of non-protein nitrogen.
The final Nesslerizcd solution cannot be diluted to 100
c.c. and smaller volumetric flasks cannot be used as re-
ceivers during the air current treatment because of
spattering. Large test tubes are therefore used as
receivers and the ammonia is Nesslcrized in these before
the liquids are transferred to measuring flasks.
"Ordinarily the colored solutions when obtained from
cat's blood are transferred to 25 c.c. flasks and are then
found to have a depth of color which permits of a
sure and accurate reading in the colorimeter. In some
Tests of Kidney Function 103
of our absorption experiments the total non-protein
nitrogen runs up to very high figures and then the solu-
tions are diluted to 50, sometimes even to 100 c.c., be-
fore being read in the colorimeter.
"Human blood contains scarcely more than one-half
as much non-protein nitrogen as cat's blood. In the
case of human blood we therefore never draw less than
5 c.c. and we take 10 c.c. of the filtrate for each de-
termination. In all other respects we use the same
procedure for human blood as for cat's blood.
"In all ordinary cases 7 to 8 c.c. of diluted Nessler's
reagent (dilution 1:5) are added for the production
of the color. If much ammonia is present so that the
resulting colored solution must be diluted to 50 or 100
c.c. correspondingly larger amounts of Nessler's reagent
are added.
"The calculation of the analytical results to milli-
grams of nitrogen per 100 c.c. of blood is not difficult,
but the formulae given below may prove useful. In
these formulae, the standard solution contains 1 mgm.
of nitrogen (as ammonium sulphate) Nesslerized in a
100 c.c. flask and the colorimeter prism of the standard
50
is set at 20 millimeters. ^- X D, in which R stands
XV
for the reading of the unknown and D represents the
volume to which its ammonia has been diluted, gives
the desired figure. The reason for the figures is that
we are working with 4 c.c. of blood.
"When 5 c.c. of blood is taken and it is diluted to 50,
40
the formula becomes -=? X D-
xv
"When working with human blood and taking 10 c.c.
of the filtrate obtained from 5 c.c. of blood diluted to
20
50 the formula is — X D-
xv
104 Manual of Vital Function Testing Methods
"It may be thought that we are using unnecessarily
small amounts of blood in these analyses. We are,
however, by no means sure that working with larger
amounts would yield more accurate results and we have
satisfied ourselves by scores of duplicate analyses that
the method as outlined gives trustworthy figures.
Further, the smaller the quantity of blood which can
be made to give reliable results the greater becomes
the usefulness of the method. The work which we
have already done on cats could not have been done
on such a small animal except by means of these micro-
chemical methods. Finally, small amounts of blood
must be used for the urea determinations because of
the disturbing effects of the sugar present."
2. Estimation of Blood Coagulation Time as Test of
Renal Function
Bachrach-Tittinger Test. — In cases of renal insuf-
ficiency, such cases as those in which it is claimed that
the freezing point of the blood is lowered (high fig-
ures for 8 ) the coagulation time has been found de-
layed. This delay is supposed to be connected with salt
retention in the plasma. A rather large amount of
blood is required according to the original technic of the
originators (20 c.c.). The test is not credited with
much value.
For method of estimating blood coagulation time
see page 41.
3. Cryoscopy of Blood as Test of Renal Function
Cryoscopy of the Blood. — This method has been
employed by some investigators on the principle that
under conditions of renal impermeability the waste
Tests of Kidney Function 105
products which fail to be eliminated in the urine will
accumulate in the blood, thereby increasing the molecu-
lar concentration. This means, of course, a lowering
of its freezing point.
The technic of estimating cryoscopy of the blood is
not different from that applied to the urine, which
has been already described (v. page 86). This method
has not come into general use, however, and for this
reason it will not be further considered here.
III. STUDY OF THE ELIMINATION OF FOREIGN SUB-
STANCES BY THE KIDNEY, AS CRITERION OF KIDNEY
FUNCTION
This category of tests may be divided into two parts :
A. Miscellaneous Chemical Substances :
1. Potassium iodide.
2. Phloridzin.
3. Hippuric acid.
4. Lactose.
B. Elimination of Dyes by the Kidney.
1. Methylcne blue.
2. Indigo carmin.
3. Phenolsulphonephthalein.
CHEMICAL TESTS
A. Tests with miscellaneous chemical substances.
1. The Potassium Iodide Test
This was one of the first chemical substances applied
to the estimation of renal function since it was sug-
gested by Duckworth as long ago as 1867. 22
22 St. Barthol. Hosp. Rep., 1867, III, 216.
106 Manual of Vital Function Testing Methods
Potassium iodide is rapidly absorbed from all the
mucous membranes.23 It is absorbed unchanged and
appears quickly in the excretions. Only a few min-
utes normally elapse before it can be demonstrated in
the urine. According to the investigations of
Quetsch,24 Roux 25 and Studeni 26 it appears at any
time from 9 to 18 minutes after doses of 1 to 3 grams
have been swallowed. The greater part of the iodide
ingested is excreted in the urine. Some, however, es-
capes in the saliva and other secretions.
Iodide is rapidly excreted, since 65-80% of the
amount ingested is eliminated in 24 hours. Several
investigators have reported the exact time required
for complete elimination to take place. According to
Antem,27 .5 gram requires 40 hours to be excreted, and
Schlayer and Takayasu,28 and Monakow 29 state that
they found the same amount required 48 hours to
eliminate. Schlayer concluded from his studies that
the demonstration of iodide in the urine beyond 60
hours after its administration may be considered de-
layed, therefore a pathological excretion.
Schlayer and his followers endeavored to fix as they
did with sodium chloride the exact locus of elimination
for iodide in the kidney. They believed that iodides are
excreted by the tubular epithelium. They also con-
tended that the elimination of iodide is not delayed
in passive congestion (cardiac) while it is delayed
in chronic tubular nephritis. These suppositions have
neither of them been substantiated by subsequent in-
23 Gushing Pharmacology, 5 Ed., 1910, 510.
M Berl. klin. Wchnschr.,' 1884, XXI, 353.
28 Thfcse de Paris, 1890, no. 248.
20 Inaugural Dissertat., Zurich, 1897.
27 Arch. f. Pathol. u. Pharmacol., 1902, XL VIII.
'"Deutsch. Arch. f. klin. Med., 1911, CI, 354.
""Deutsch. Arch. f. klin. Med., CII, 248.
Tests of Kidney Function 107
vestigations.
Rowntree and Fitz in their experience with the iodide
test have found it to vary markedly and they believe
that the observation of excretion time of potassium
iodide as a test of renal function is unreliable.
Technic of Iodide Test. — .5 gm. (7^ grains) of
potassium iodide is given in solution by mouth in
the morning on arising. The urine is collected at the
end of 48 hours and every four hours thereafter and
tested for iodide until a negative result is obtained.
One of the simplest and best qualitative tests for
iodide in the urine is that of Sandow. The test is
made by taking 30 c.c. of urine, 2 c.c. of 2% solution
of sodium nitrate and 2 c.c. of dilute sulphuric acid,
adding chloroform and shaking. A purplish or violet
color appears in the chloroform if iodide is present.
2. The Phloridzin Test
Von Mering 30 discovered the fact that the injection
of the glucoside phloridzin into animals, produces a
glycosuria without a hyperglycemia, thus proving that
the conversion is a vital act of the renal parenchyma.
Achard and Delamare 31 built upon this fact a method
of testing the functional capacity of the kidney.
Technic of Phloridzin Test. — .005 gm. of phloridzin
in fresh aqueous solution is injected hypodermically.
At 15-minute intervals the urine collected by catheter or
voided spontaneously is examined for sugar. The
maximum excretion takes place normally in an hour
and disappears in 2 or 3 hours.
00 Centralbl. f. med. Wissensch., 1885, 531.
31 Bull, et Mem. Soc. Med. d. Hop. de Paris, 1899, 379.
108 Manual of Vital Function Testing Methods
3. The Hippuric Acid Test
It has been long known that bcnzoic acid or benzoates
are eliminated by the kidney as hippuric acid which
is synthesized in the kidney from benzoic acid and
glycocoll. Altho the fact has been used as a basis for
testing kidney function the results have been disap-
pointing and the method lias been abandoned.
4- The Lactose Test
Voit 32 was the first to demonstrate that lactose is
eliminated by the healthy kidney following its sub-
cutaneous or intravenous injection. De Bonis 33
claimed that the elimination takes place exclusively in
the glomerulus.
Lactose was suggested as a means of estimating renal
functions in 1911 by Schlayer and Takayasu,34 who
with their co-workers studied the question of renal func-
tion in experimental and clinical nephritis. These
workers studied the elimination of lactose, potassium
iodide, salt and water, dividing the nephritides into
vascular and tubular varieties with subdivisions.
Schlayer believed that the elimination of lactose be-
ing exclusively, as he thought, a glomerular function
could be taken as an index of the vascular functioning
power of the kidney. Lactose being a foreign sub-
stance that is not found in the organism would not
be influenced in its elimination by extrarenal factors,
and should therefore be an ideal criterion of glomerular
function, any delay in its passage through the kidney
indicating glomerular insufficiency.
MDcut. Arch. f. klin. Med., 1897, LVII, 545.
"Giorn. intern, cl. scien. Med., 1907, XXIX, 4iG.
:" Deutsch. Arch. f. klin. Med., 1911.
Tests of Kidney Function 109
Following Nussbaum's 35 technic of obtaining in the
frog an exclusively tubular secretion from the kidney
by artificially excluding the glomerular secretion,
Rowntree and Fitz 36 concluded that the tubular epithe-
lium can secrete a certain amount of lactose, hence its
elimination is not exclusively a function of the glomeru-
lus. But they concluded also from their clinical experi-
ments with lactose in the study of renal function in
practice and also from some studies they have made
in experimental passive congestion, that the mechanism
of lactose excretion differs essentially from that of
phthalein, salt, indigo carmin, etc., and that estima-
tions of lactose excretion may be looked upon as a
satisfactory index of the vascular, if not exclusively
the glomerular function of the kidney.
Technic of Lactose Test. — Two and five-tenths (2.5)
gms. of chemically pure lactose are dissolved in 25 c.c.
of freshly distilled water, placed in small cotton stop-
pered Erlenmeyer flasks and pasteurized for four
hours for four successive days at 75 to 80° C. By this
method the dose injected amounts to a little over 2
gms. lactose in 20 c.c. of water. A fresh solution is
used for each injection and a careful technic for in-
travenous injection carried out.
Following the injection there are usually no consti-
tutional disturbances altho occasionally there may be
some headache malaise or even chill followed by fever.
The urine is collected four hours after the injec-
tion and every hour or two hours after for twelve
hours. Each specimen is tested for sugar by Nylander's
reagent, using the same amount of urine, solution and
length of time for boiling. Polarimetric readings may
be made.
^Pfliiger's Arch. f. d. ges. Physiol., 1878, XVI, 179; XVII, 580.
36 Archives of Int. Med., 1913, XI, 258.
110 Manual of Vital Function Testing Methods
The normal excretion time for this amount of lac-
tose is four to six hours. The time required for secre-
tion is the main point. Over six hours is delayed ex-
cretion.
(Nylander's reagent consists of Rochelle salts, 4
gins, dissolved in 100 c.c. of 10% NaOH (sp. gr.
1015); warm and saturate with bismuth subnitrate
(about two grams are necessary). When cool, filter
and keep in a dark bottle. The solution remains perma-
nent for years.)
B. ELIMINATION OF DIFFERENT COLORING MATTERS OR
DYES AS A MEASURE OF RENAL FUNCTION. URINARY
CHROMOSCOPY
There are three of these tests used at the present
time, namely:
1. The Methylene Blue Test.
2. The Indigo Carmin Test.
3. The Phenolsulphonphthalein Test.
Other coloring matters such as rosanilin, fuchsin,
etc., have been suggested and employed at different
times for estimating renal function, but with the ex-
ception of the three named they appear to have fallen
into disuse at the present time.
Numbers 2 and 3 are most extensively employed,
namely, indigo carmin which is used particularly in
Europe and phenolsulphonphthalein which is by far
the most popular colorimetric test in this country.
Rosanilin (sodium trisulphate) was introduced by
Lepine.37 One c.c. of a \c/o solution is injected hypo-
dcrmically. The dye appears normally in the urine
in less than half an hour, total elimination requiring
twenty-four hours. The test has never attained wide
37 Lyon Medical, 1898.
Tests of Kidney Function 111
use, probably because of the greater success attending
the use of phenolsulphonphthalein according to the
method of Rowntree and Geraghty (q. v.).
1. The Meihylene Blue Test
The introduction of methylene blue as a test for
renal function is credited to Achard and Castaigne.38
These authors gave the drug intramuscularly, using
1 c.c. of a 5 % solution. Later Czyhlarz and Donoth 39
recommended it by mouth in 1/4 grain dose. The drug
is rapidly eliminated in the urine, appearing therein in
about fifteen minutes as a colorless chromogen, as dem-
onstrated by Voisin,40 which may be shown by boiling
the urine after addition of acetic acid. Normally the
color itself appears in the urine in half an hour. The
excretion of both forms continues for 36-48 hours, but
even in health the time may be very much prolonged.
The authors of the test recommended that the time
of first appearance, time of maximum intensity of
excretion and time required for total excretion should
be noted.
Diminished renal permeability was thought to delay
or prolong all three. Various observers confirmed
these suppositions. In some cases of chronic inter-
stitial nephritis the elimination was found to be pro-
longed for two weeks. Various modifications of the
type of elimination under pathological conditions were
described, consisting of remittances or intermittances of
excretion.
Later observers noted that elimination is not delayed
in all forms of kidney disease and that the elimination
38 Bull, et Mem. Soc. Med. d. hop. de Paris, April, 1897, 63T.
39Wien. klin. Wchnschr., XXIV, 1899, 649.
40Gaz. Hebd. Med., 1897, 493.
Manual of Vital Function Testing Methods
under certain circumstances might be normal or even
accelerated.
Attempts to measure quantitatively the elimination
of methylene blue were made, one of which described
by Rowntree and Geraghty will be mentioned under
technic.
The methylene blue test was later applied to diag-
nosis in surgical diseases of the genitourinary tract
and was at one time considered the best test available
for estimating the functional capacity of one or both
kidneys. Walker showed that the elimination is re-
tarded in lower urinary tract obstructions as in cer-
tain types of prostatic hypertrophy. Casper made
similar observations.
Methylene blue produces some pain when given sub-
cutaneously and occasionally when given intramuscu-
larly. This of course is a drawback though perhaps
not a serious one. Another and greater drawback is its
prolonged elimination necessitating a large number of
urine examinations. A third objection is the difficulty
of accurate colorimetric estimations.
Finally, the methylene blue test is imperfect in that
part of the substance is converted into a colorless
chromogen in the body secreted as a leucobase and
therefore does not contribute to the color results in
the urine. Only 50% of the drug is normally passed
out in the urine.
It has not been demonstrated with certainty in what
part of the kidney the elimination of methylene blue
takes place.
In disease of the tubular epithelium that is in the
chronic parenchymatous nephritis, methylene blue is
quickly and completely eliminated ; in interstitial neph-
ritis, the elimination is delayed.
The duration of elimination is diminished in paren-
Tests of Kidney Function 113
chymatous and increased in interstitial nephritis. A
cyclic, polycyclic or intermittent elimination of the
coloring matter has been said to indicate several dif-
ferent conditions : disturbance of kidney innervation,
hepatic insufficiency, interstitial nephritis and pyo-
hydronephrosis.
Up to the time of the advent of the phenolsul-
phonphthalein test, the methylene blue test was the
most extensively used method of determining renal per-
meability. At the present time, certainly in America,
the Rowntree-Geraghty test has quite superseded it.
Teclinic of Methylene Blue Test.— After urination
1 c.c. of a 5% solution of methylene blue is injected
intramuscularly. A sterile catheter may be introduced
into the bladder or the patient may empty the bladder
in 15 minutes if possible to determine the presence of
the leucobase or chromogen. This is done by boiling
the specimen and adding a few drops of acetic acid.
A greenish color denotes the presence of the chromogen.
At the end of half an hour the bladder should be
emptied spontaneously or by catheter and the urine
examined for color. A greenish blue color denotes the
presence of the dye.
As above mentioned, the time of appearance, time of
maximum intensity and time required for total elimina-
tion (disappearance of color) should be noted.
Quantitative Estimation. — Before administration of
the drug the urine is collected for some time and kept.
The methylene blue is given in the usual manner, the
urine collected for as long a time as necessary, all
chromogen being converted into dye. An equal quan-
tity of urine previously collected is taken, to which
is added from a burette, drop by drop, a sufficient
quantity of a solution of methylene blue of known
strength until the colors are alike. Compare against
114 Manual of Vital Function Testing Methods
a white background. From the quantity of methylenc
blue used, the amount of coloring matter may be esti-
mated.
2. The Indigo Carmm Test — Volcker- Joseph Test 41
The dye was first used by Heidenhain in his famous
investigations into the physiology of the kidney. He
believed that this substance is eliminated exclusively
by the epithelial cells of the convoluted tubules.
Indigo carmin possesses the advantages over methy-
lene blue that the quantity required for the test is
completely eliminated thro the kidney and that no
leucoderivative is formed in the tissues.
After the intramuscular injection of .08 gm. to .16
gm. of indigo carmin, elimination begins in 6 to 8 min-
utes if the kidney is normal. The intensity of the color
will give some idea of the concentrating power of the
kidney, that is to say, the water resorbing power of
the tubules, and consequently, its capacity to produce
a urine of high molecular concentration.
A polycyclic or intermittent elimination is said by
Blum to indicate intermittent hydronephrosis. In
strongly alkaline urine the dye may be discolorized.
In parenchymatous nephritis, the elimination of indigo
carmin may be normal. In interstitial nephritis the
elimination begins later than normal, is diminished in
quantity and much prolonged. The delayed elimination
indicates a diminished reaction power of the kidney, the
diminished elimination a loss of water resorbing power,
a hyposthenuria, a loss of the concentrating function
of the organ, while the long duration of elimination indi-
cates a general loss of secreting power as always accom-
panies the sclerotic kidney.
"Munch, med. Wchnschr., 1903, 2081.
Tests of Kidney Function 115
The indigo carmin test has been quite extensively
used, especially in Europe, in testing the functional
capacity of the single kidney by ureteral catheteriza-
tion and in general functional testing of the kidney. It
is considered superior to methylene blue because of
more rapid elimination. But in this respect, as in
others, it is inferior to phenolsulphonephthalein, which
substance, since its introduction for this purpose by
Rowntree and Geraghty, has become the most exten-
sively used chromoscopic test, at least in this country.
Rowntree and Geraghty consider the indigo carmin
test of more value than the methylene blue test because
of its more rapid appearance in the urine, but that it
is less adapted to functional work than phenolsul-
phonephthalein. This opinion is now shared by most
other workers in this field.
Technic of Indigo Carmin Test. — A 4% solution is
made. Twenty c.c. of this solution are injected into the
muscles usually of the gluteal region. There is some
pain produced by the injection. In 10 to 15 minutes
the urine is collected. In normal persons it is tinged
greenish blue in this time. Excretion is usually com-
plete in 24 hours, but practically the greater portion
escapes in 12 hours. The color of the dye in the urine
does not lend itself well to colorimetric estimation. In
this respect it resembles methylene blue. Purulent
urine decolorizes indigo carmin. It is estimated that
not more than 25% of the amount injected finds its
way out thro the kidneys. The fate of the balance is
unknown.
3. The Phenolsulphonephthalein Test. Rowntree-
Ger eighty Test (The Red Test}
The phenolsulphonephthalein test of kidney function
had its origin in the pharmacological researches of
116 Manual of Vital Function Testing Methods
Abel and Rowntree, upon the phthaleins generally. Of
all the phthaleins studied by these investigators, phe-
nolsulphonephthalein stood out in striking contrast
with all others because of the fact that it is almost ex-
clusively eliminated by the kidney.
This fact suggested its use as a test of renal func-
tion to Rowntree and Geraghty and their first com-
munication upon this subject appeared in July, 1910.42
Phenolsulphonephthalein has the following formula:
'CfiILOH
CGH4OH
C6H4
It was first made by Remsen.43 The substance is a
red crystalline powder, partly soluble in water, the
solution when alkaline being red, becoming more purple
as the alkalinity is increased.
Abel and Rowntree in their pharmacological inves-
tigations of the phthalein group showed that the sub-
stance appears in the urine after administration by
mouth in one to one and a half hours and after sub-
cutaneous injection in about 10 minutes. They found
that after fair-sized doses (1 gm.) the drug appears
in the bile, is passed into the intestine, there reabsorbed,
and, except for a mere trace, is excreted wholly by the
urine.
Phenolsulphonephthalein is practically non-toxic.
43 Jour. Pharmacol. and Exp. Therap., I, 1910, 579.
"Amer. Chem. Jour., VI, 280.
Tests of Kidney Function 117
When a dose of .006 gm. is injected subcutaneously,
40-60% of this quantity is recovered in the urine dur-
ing the first hour after injection. From 15-25% more
is recovered in the second hour, making a total excretion
for the first two hours following the injection of
60-85%.
Normally when the urinary flow is free, the dye ap-
pears in the urine in 5 to 10 minutes after injection.
The maximum excretion appears in 15 to 20 minutes.
This density of excretion continues an hour to an hour
and a half. The elimination then begins to diminish.
At the end of the first hour the pink color on addition
of alkali is slight, and after the expiration of two hours
excretion is practically complete.
In acute nephritis, Rowntree and Geraghty 44 found
a diminished excretion of phthalein in two out of three
cases. In parenchymatous nephritis there was a marked
diminution of excretion in seven out of eight cases. In
chronic interstitial nephritis a low output was encoun-
tered in all the cases experimented upon, 10 in number.
These results concerning the lowered excretion of
phenolsulphonephthalein in the nephritides have as a
general thing been entirely corroborated in subsequent
investigations by many different observers.
In their first researches Rowntree and Geraghty
made over two hundred functional tests in one hundred
and fifty persons. To them the phenolsulphonephtha-
lein seemed to possess advantages over all other func-
tional tests, these advantages consisting chiefly in the
following points:
1. The early appearance of the dye in the urine
and its rapid and complete elimination by the kidney.
2. The accuracy and simplicity of quantitative esti-
mation of the drug in the urine.
"Jour. Phar. and Exp. Ther., I, 1910, 656.
118 Manual of Vital Function Testing Methods
They showed by these researches that the permeabil-
ity of the kidney for phenolsulphonephthalein is de-
creased in both parenchymatous and interstitial nephri-
tis, the decrease being most marked in the latter form.
Further than this they showed that the test is of
value to the surgeon in determining the true condition
of the kidney, in cases with prostatic obstruction. In
such cases the authors believed the phenolsulphoneph-
thalein test to be of greater service than urinalysis or
nitrogen estimations and that the use of the test in
cases of obstruction in the lower urinary tract prior
to operations would disclose the necessity of prelimi-
nary treatment. Finally they pointed out that the
test lends itself and is well adapted to unilateral esti-
mations of the functions of the separate organs in con-
junction with ureteral catheterization.
In surgical cases with urinary obstruction, the
authors contended that when the phthalein excretion
is delayed beyond twenty-five minutes and the output
for the first hour is below 20% the operation may prof-
itably be postponed until treatment by drainage has
improved conditions, such improvement being shown by
an increase in the elimination of phthalein at a subse-
quent time.
Technic of Phenolsulphonephthalein Test. — Twenty
minutes to half an hour before giving the test the
patient is given 200 to 400 c.c. of water to insure
diuresis. The bladder is cathetcrized or completely
emptied. The time being noted, 1 c.c. of a solution of
the drug is injected into the lumbar muscles. The solu-
tion is prepared as follows : .6 gm. phenolsulphoneph-
2
thalein and .84 c.c. - NaOH are added to .75% NaCl
n
2
solution. Add two or three drops of NaOH. The
n
Tests of Kidney Function 119
color becomes Bordeaux red and the solution is non-
irritant.
The urine is passed into a test tube containing a drop
of 25 % NaOH and the time of appearance of the first
pinkish color noted.
If there is no urinary obstruction the catheter is
not necessary after the appearance of the color, and
the patient may then retain the urine and urinate at
the end of one hour in one receptacle and again at the
end of the second hour in another.
A rough estimate of the time of the appearance of
the drug in the urine may be gained by having the
patient urinate, frequently, a small amount without
the catheter. In prostate cases it seems better to keep
a catheter in situ. If this is done the catheter may
be corked and this is removed at the end of the first
and second hours.
Each sample of urine is measured. Twenty-five per
cent. sol. NaOH is added to make the color maximum.
The urine is usually yellow or orange and becomes deep
purple on addition of the alkali. The solution is put in
a liter flask and diluted with distilled water to make a
quart. This is thoroughly mixed and a portion is fil-
tered and compared with a standard in a colorimeter.45
The standard solution consists of .003 gm. phenolsul-
phonephthalein (^ c.c. of solution used for injection)
diluted to 1 liter and made alkaline with a few drops
of 25% NaOH. The test solution retains its fine
purplish color for a week or more.
The colorimeter contains a wedge-shaped cup which
is filled with the standard solution. The rectangular
45 The colorimeter used by Rowntree and Geraghty is a modi-
fication of the Autenrieth-Konigsberger instrument. This can be
obtained from Hynson and Westcott, Balto., Md., who also sup-
ply convenient ampoules containing .006 gm. in each c.c. of phe-
nolsulphonphthalein.
120 Manual of Vital Function Testing Methods
cup is filled with the solution to be tested. The wedge-
shaped cup is manipulated by a screw until the color
fields are identical. The percentage is read off on the
indicator scale.
Technic of the Phenolsulphonephthalein Test as Ap-
plied to Estimation of the Function of the Individual
Kidney. — Twenty minutes previous to the application
of the test the patient is given 600 to 800 c.c. of water
to provide a free flow of urine. The ureters are
catheterized, a special catheter being recommended,
namely, the flute end catheter of Albarran No. 6 or
No. 7. The catheters are passed four inches into the
ureters. The cystoscope is withdrawn, leaving the
catheters in situ. A small urethral catheter is passed
into the bladder to empty that organ and detect later
leakage. The other details of the test are similar to
those of the ordinary technic (q. v. ).
In September, 1911, Geraghty and Rowntree 46 made
a report of their previous experience with the sul-
phonephthalein test and reiterated their first opinion
that the test devised by them appeared to possess dis-
tinct advantages over all other methods of examining
renal function. The reasons upon which this opinion
was based have been given above.
They concluded that the sulphonephthalein test will
enable the clinician to determine quantitatively the
amount of functional derangement of the kidneys in
his nephritis cases, whether of the acute or chronic
type. That in cardiorenal cases the test will show
exactly to what extent the kidney is involved. That
the test is of special value in the diagnosis of uremia
from other conditions which may simulate it and also
to foretell in many cases an impending uremia before
the appearance of indubitable clinical signs.
46 Jour. Arner. Med. Assn., 1911, LVII, 815.
Tests of Kidney Function
The authors reiterated their confidence in the value
of this test in cases of urinary obstruction, it being
in their judgment superior under these circumstances
to measuring the urinary quantity, and to urea or total
nitrogen estimations.
In many surgical cases studied by them in the genito-
urinary clinic of Young at Johns Hopkins, they found
that separate studies of unilateral kidney function re-
vealed more accurate and dependable information than
any other method of examination.
In a third and very complete report of their phenol-
sulphonephthalein test of kidney function published in
1912, Rowntree and Geraghty studied carefully the in-
fluence on the rate of excretion, of the various methods
of administration, subcutaneous, intramuscular and in-
travenous. They concluded from their researches that
intramuscular injection into the lumbar region is the
method of choice.
Studying the influence of various diuretics upon the
excretion of sulphonephthalein the authors found that
while under the conditions of animal experimentation,
some slight increased activity was caused by certain
stimulating diuretics like caffein, yet clinically these
substances do not affect the phthalein output.
The route of phthalein through the kidney was in-
vestigated and it was demonstrated that the drug is
excreted chiefly by the uriniferous tubules and the
smaller remainder by the glomerulus.
In nephritis of all types the output of phthalein
was found diminished, the diminution of excretion be-
ing apparently in proportion to the amount of damage
to the kidney structure. So that the test is of consider-
able value from a diagnostic and prognostic standpoint
since the amount of functional incapacity is revealed.
In those cases in which the heart and kidney are
Manual of Vital Function Testing Methods
both affected (the so-called cardiorenal cases) the test
was found useful in determining just what proportion
of the trouble could be referred to the heart disease and
what to the renal lesions.
In uremia the test proved in the hands of its authors
of value in differentiating uremia from conditions sim-
ulating it. In certain cases, when no clinical evidence
of the imminence of uremia was present, a very low
phthalein output frequently enabled the authors to fore-
see the danger.
From a surgical standpoint the earlier opinions held
by Rowntree and Geraghty as to the utility of the test
in cases of urinary obstruction were completely cor-
roborated by their subsequent work.
According to them the test is more dependable than
estimation of urinary output, total solids, urea or total
nitrogen, in indicating to the surgeon the propriety of
operation or the institution of preliminary treatment,
in contemplated nephrectomy and prostatectomy.
Finally their studies tended to show that no other
test is so adaptable to the examination of unilateral
kidney function to determine the relative amount of
work performed by each organ separately.
The medical and surgical aspects of the phthalein
test will be further developed later on under the general
summary of renal function tests in their medical and
surgical aspects, (v.i. )
Since the introduction of the phenolsulphonephtha-
lein test for kidney function in July, 1910, a very con-
siderable literature upon the subject has appeared. It
is very striking how few are the criticisms and how
numerous are the encomiums which have been passed
upon the test of Rowntree and Geraghty. It might al-
most be said that, in this country at least, the opinion
of those who have used it is unanimously favorable and
Tests of Kidney Function 123
tends to corroborate in every particular the claims
which were advanced for it by its authors.
Clinical and experimental corroboration of the sul-
phonphthalein test have been given by the publications
of Austin and Eisenbrey,47 Boyd,48 Cooke,49 Sehrt,50
Lance,51 Sanford,52 Behrenroth,53 Frank,53 Bonn,54
Erne,55 Mouriquand,56 Lohnstein,57 Frothingham,
Fitz, Folin, Denis,58 Christian, Janeway, Cabot,
Dock,59 Snowden, Thayer,59a and many others.
While many of these reports are extremely illuminat-
ing and important, it cannot be said that they have
added anything noteworthy to the test itself, or to its
indications, which fact is a strong testimony of the thor-
oughness and care with which the work had originally
been performed by Rowntree and Geraghty before its
publication.
From the standpoint of pure experimental corrobora-
tion, the work of Austin and Eisenbrey should be noted.
These authors in 1911 studied the elimination of phe-
nolsulphonephthalein as compared with the elimination
of nitrogen and chlorides, in experimental nephritis in
dogs set up by administering uranium, cantharidin, and
potassium bichromate. They concluded, among other
"Jour. Exper. Med., 1911, XIV, 367; 4,62.
48 Jour. Amer. Med. Assn., 1912, LVIII, 620.
49 Providence Med. Jour., 1912, XIII, 118.
80 Centralbl. f. Ch., 1912, XXXIX, 2, 1121.
51 Gaz. d. hop. de Par., 1912, LXXXV, 32.
62 Cleveland Med. Jour., 1912, XI, 763.
53 Ztsch. f. Exp. Pathol. u. Therap., 1913, XIII, 72.
54 Jour. Ind. State Med. Assn., 1913, VI, 154.
"Munch, med. Wchnschr., 1913, LX, 510.
5SLyon Medicale, 1913, CXXL, 299.
"Allg. Med. Centr. Gtz., 1913, LXXXII, 591.
58 Arch. Int. Med., 1913, vols. XI-XII; also Jour. Exper. Med.,
1911, XIV, 366.
59 Tr. Cong. Amer. Phys. & Surg., 1913, IX, 45.
69aAmer. Journ. Med. Sc., 1914, CXLVIII, 781.
Manual of Vital Function Testing Methods
things, as a result of their researches, that a marked
and early decrease in the elimination of phenolsulpho-
nephthalein takes place in the experimental nephritides
and that the phthalein test is the better indicator of
renal function under the circumstances than total ni-
trogen or chloride elimination, which latter are more
irregular and inconstant.
Other investigators have experimented along the same
line, studying the phthalein elimination in experimental
nephritis and their results have tended to corroborate
the earlier researches. Perhaps the most recent con-
tribution to this phase of the subject is that of Potter
and Bell.60 These authors have studied the phthalein
elimination, also that of lactose and potassium iodide
in experimental tartrate nephritis in rabbits.
It may be recalled that Underbill, Wells and Gold-
schmidt G1 discovered that the injection of racemic tar-
taric acid into rabbits produces a type of acute nephri-
tis in which the great majority of the convoluting tu-
bules become necrotic and the rest are fatty and gran-
ular. The glomeruli may be anatomically intact. In
kidneys of this type it has been found that the excretion
of phenolsulphonephthalein, likewise indigo carmin and
methylene blue, is completely suppressed. The excre-
tion time of lactose is over twice as long as normal,
while that of potassium iodide is four times as long as
normal, but both lactose and potassium iodide are ex-
creted by this type of kidney. Potter and Bell suggest
that their results appear to show that phthalein, indigo
carmin and methylene blue are excreted exclusively by
the tubules, while potassium iodide and lactose are
at least partly excreted by the glomeruli.
In experimental chronic passive congestion of the
°°Amer. Jour. Med. Sci., CXLIX, 1915, 236.
91 Jour. Exper. Mcd., 1913, XVIII, 322.
Tests of Kidney Function 125
kidney in animals, produced by compression of the vena
cava and renal veins, Rowntree, Fitz, and Geraghty 62
found that the functional capacity of the kidney as
judged by the phthalein output is reduced. The reduc-
tion, however, only occurs as the degree of passive
congestion becomes marked.
Goldsborough and Ainley in 1910 63 studied the renal
function in pregnancy and the puerperium by means of
sulphonephthalein, and concluded that even normal fe-
males in pregnancy eliminate less phthalein than non-
pregnant. In the ninth month the power of elimination
may be very low. The exact meaning of these facts is
not known. This result was confirmed by Roth,64
who also claimed that women with diseases of the genital
tract were unsuitable for the test. These investigations
of Goldsborough, Ainley and Roth have not been subse-
quently developed. In this connection it may be men-
tioned that Pepper and Austin 65 reported in 1913 that
occasionally cases of parenchymatous nephritis will
show a quite prompt and fairly normal elimination of
phthalein. In one of their cases there was a phthalein
output of 67% for one hour, strongly suggesting
hyperpermeability. Baetjer has also encountered sim-
ilar cases. Such cases do not appear to represent the
rule, however, and just what meaning is to be attached
to these facts is at present unexplained.
A few suggestions for slight variations of technic in
the test have been published but none of them appears
to have been generally adopted. They are few in num-
ber and the most important may be given.
Fromme and Rubncr GG in 1912 suggested that the
82 Arch, of Int. Med., 1913, XI, 121.
""Journ. Amer. Med. Assn., 1910, LV, 2058.
64Berl. klin. Wchnschr., 1913, L, 1609.
65 Amer. Journ. Med. Sc., 1913 n. s. CXLV, 254.
66Berl. klin. Wchnschr., 1912, XLIX, 1889.
126 Manual of Vital Function Testing Methods
phthalein should always be given hypodermically and
that the observation period should be extended to three
hours. Keyes and Stevens 67 also recommend hypoder-
mic injection when the ureters are to be catheterized.
Bonn,68 as a result of his experience, thought that the
time of appearance of the dye in the urine is not of
much importance except to determine the time for per-
centage estimation. He recommended the intravenous
injection of the phthalein when the ureters are to be
catheterized. He does not believe that the test will
inform the surgeon when the patient can be operated on
safely. He, however, states that, in his belief, the test
is the best one yet devised for studying the renal func-
tion.
Fanz 69 suggests to add a quantity of the patient's
urine obtained just before injecting the indicator, equal
in amount to the first hour's urinary output after the
injection. The standard solution now will have ap-
proximately the same amount of urinary salts as the
specimen solution, and the standard and urinary solu-
tion will equal each other in opaqueness and yellowish
tint, making color comparison easy. To make up the
standard solution, he uses 1 c.c. of the contents of an
ampoule * of the phcnolsulphonephthalein, adds the pa-
tient's urine (obtained before injecting the indicator)
in amount to equal the first hour's urinary output after
appearance of the drug. This mixture is alkalinizcd
with 25 c.c. of 10 % solution of potassium hydroxide,
filtered, and sufficient distilled water is added to make
1000 c.c. This is the standard and contains 100% of
the indicator in 1000 c.c.
67 N. Y. Meet Jour., 1912, XCV, 1134.
88 Indiana State Med. Assn. Jour., 1913, VI, 154.
68 N. Y. Med. Jour., 1915, C, 1193.
* Put on market by Hynson and Westcott.
Tests of Kidney Function 127
The first hour's urinary output, after injection of
2 c.c. phenolsulphonephthalein, is now alkalinized with
25 c.c. of 10% potassium hydroxide and filtered. To
this is added distilled water to make 1000 c.c. This is
the test specimen. By diluting a unit of the standard,
say 50-100 or 200 c.c., with distilled water until it
matches the 1000 c.c. solution of the first hour's urinary
output, the direct percentage of the indicator in the
first hour's specimen can easily be estimated. Say 100
c.c. of the standard had to be diluted up to 500 c.c.
before it matched the first hour's specimen dilution, then
the standard would be five times as strong as the speci-
men, or the specimen would contain 20% of phenolsul-
phonephthalein. The second hour's output of phenol-
sulphonephthalein is estimated precisely like the first.
As a matter of fact it does not appear that any
modifications of the original test method as described
by Rowntree and Geraghty has contributed materially
to its simplification or improvement, and for this reason
the original method is usually followed.
Thayer and Snowden 70 have recently attempted to
compare their results obtained by the test with the
anatomical changes found in the kidneys at autopsy.
They conclude, as the result of their quite extensive
investigations, as follows :
In severe chronic nephritis there is always a low
phthalein output. This rule, in their experience, has
absolutely no exception. The phthalein output in cases
of chronic nephritis diminishes steadily, according to
them, until the terminal uremia, when it approaches
zero, just prior to death.
In the passive renal congestion of heart disease, there
is often a reduction in the elimination of phthalein,
especially when the amount of decompensation of the
70Amer. Jour. Med. Sci., 1914, CXLVIII, 781.
128 Manual of Vital Function Testing Methods
heart is considerable. When these symptoms become
ameliorated, the phthalein output increases.
If passive congestion of the kidney from heart disease
is accompanied by concomitant chronic nephritis the
output of phthalein is lower than in cases of uncom-
plicated congestion.
In one case of acute nephritis and one of amylosis
the phthalein output was reduced.
In acute infectious diseases, during which cloudy
swelling of the renal parenchyma occurred, there was
found a considerable reduction of phthalein output.
From this study Thayer and Snowden concluded
that the phenolsulphonephthalein test of Rowntree and
Geraghty is a procedure of great diagnostic and prog-
nostic value, especially in the study of chronic neph-
ritis.
General Summary of the Application of Renal Func-
tion Tests
This subject may be divided into two parts : 1. Renal
Function Tests and Their Medical Application. 2.
Renal Function Tests and Their Use in Surgery.
1. Medical Application. — A decade ago the burning
question in renal pathology was whether the varying
clinical findings in chronic nephritis could be divided
into classes and each class correlated with certain defi-
nite histopathological findings in a kidney post mortem.
Although no absolute answer to the question was
reached, there is no doubt some progress was made.
During the past decade, in line with the general depar-
ture of interest somewhat away from the anatomical
toward the functional view in pathology, a serious at-
tempt was made to divide the clinical symptomatology
of nephritis into groups ; to correlate these groups with
Tests of Kidney Function 129
certain definite types of renal involvement. Although
this worthy attempt has not been entirely successful any
more than its anatomical prototype which preceded it,
some important facts have been learned which are of
value from a practical as well as a theoretical stand-
point. The method by which these advances have been
gained is none other than the application of functional
tests to a study of the different phases of renal activity.
Different portions of the glomerulo-tubular structure
of the kidney have been supposed to possess selective
secretory activities. It cannot be said that these selec-
tive activities are yet understood, so that at the present
time there is no such thing as an exact topical diagnosis
of the kidney functions.
Nor can it be said in any case that the results of the
most complete and comprehensive functional examina-
tion will reveal with any certainty the anatomical
changes which are present in the kidney. Certainly
they will not reveal the extent to which the vascular,
glomerular or tubular structures are involved in a given
case, and will shed but little light upon the relative
proportions of the changes.
Our chief clinical terms in renal pathology still re-
main as they were in the older tradition ; we still speak
currently of acute nephritis, chronic parenchymatous
nephritis and chronic interstitial nephritis, but we are
not surprised when the autopsy shows the extreme rar-
ity of these arbitrary types and presents us with patho-
logical pictures of such great complexity that it is no
wonder that the intricate problem is not to be solved by
the most skillful and painstaking ante-mortem exami-
nation.
Great hopes were aroused some years ago in this
direction when it was discovered that experimental in-
flammation of the kidney may be set up in animals by
130 Manual of Vital Function Testing Methods
the use of certain poisons 71 which make it possible to
submit some of these difficult problems in renal pathol-
ogy to experimental investigation. Although little has
come of this work so far, it appears to be founded
upon rational and scientific principles and the work
must be enthusiastically encouraged to go on.
Schlayer, Hedinger, and Takayasu, in the Romberg
Clinic at Tubingen, have applied themselves assiduously
to working out the problems of pathological kidney
function from an experimental standpoint. In this
country the purely experimental method has yielded
interesting results in the hands of Rowntree, Fitz,
Geraghty, Christian, O'Hare, Folin, Karsner, Denis,
Frothingham, Austin, Eisenbrey, Potter, Bell, and
others. For an interesting contribution dealing with
the relation of functional tests to pathological diag-
nosis, the reader is referred to an article by Christian,72
to which an excellent bibliography is appended. The
author of this review very justly concludes that tests
of renal function are quite capable of demonstrating
the bare fact (the important fact indeed) that the kid-
neys are diseased but are quite unable to disclose the
exact type of pathological lesion of the kidney, which
exists in any given case.
While no functional test is capable of disclosing the
nature or location of the pathological lesion, it may be
perfectly capable of disclosing the inability of the kid-
ney to perform a given function or set of functions,
such, for example, as the elimination of water, salt,
urea or some foreign substance such as lactose or
phthalein. Such an inability on the part of the kidney
will indicate a depreciation of its functional power if
71 Uranium, ohromates, mercuric chloride, cantharidin, tartrates,
etc.
72 Trans. Cong. Amer. Phys. and Surg., 1913, IX, 1.
Tests of Kidney Function 131
the test is properly carried out, and this particular
information may be important in prognosis and treat-
ment although we shall be unable with certainty to point
to the seat of the disturbance in the kidney or identify
with exactitude the nature of the underlying patho-
logical lesion.
From a medical standpoint renal function tests are
most important in defining the state of kidney activity
in the acute and chronic nephritides, orthostatic and
other albuminurias, arteriosclerosis, uremia, and myo-
cardial insufficiencies.
In some medical clinics renal function tests are ap-
plied as a matter of routine to all these classes of cases.
While they do not in themselves make the diagnosis or
settle the prognosis it is contended, very properly per-
haps, that they will occasionally reveal an unsuspected
latent deterioration of kidney function, just as routine
blood examinations may reveal an unsuspected leukemia.
Renal functional tests will be of prognostic value be-
cause they will often serve to show whether the disease
is stationary, progressing or recovering. Of course it
hardly needs to be said that renal function tests will
always be carried out in the medical clinic in conjunc-
tion with the clinical study of the patient. A progres-
sive lowering of the kidney function in chronic nephri-
tis may indicate impending uremia.
The question of the prognostic value of renal func-
tional tests has been made the theme of a very complete
and excellent article by Rowntree.73 According to this
investigator the prognostic value of renal functional
studies is as great in medical as it is in surgical cases.
In acute nephritis, the prognosis depends upon the eti-
ology more than the result of functional tests. In mild
chronic nephritis with slight albuminuria and cylindru-
73 Trans. Cong. Amer. Physic, and Surg., 1913, IX, 23.
Manual of Vital Function Testing Methods
ria, slight hyperpiesis, moderate arteriosclerosis and
hypertrophy, the tests may show how great the func-
tional deterioration may be, and the regular repetition
of the tests will show whether the disease is progressing
or not. In advanced nephritis also, the proper func-
tional tests will disclose the severity of the disease and
the imminence of uremia. Cases of clinically well-
marked nephritis, even with some uremic signs and a
less marked functional derangement, will be more diffi-
cult to prognose. Head and brain complications cannot
be foreseen by means of renal functional tests. If the
renal function remains fair, say 30% of phthalein out-
put (Rowntree) and the blood tests do not show marked
cumulative phenomena the prognosis is favorable and
vice versa.
In cardiorenal cases it is always difficult to deter-
mine clinically just how much the heart and kidney are
separately responsible for the conditions. Renal func-
tional tests are of considerable service in showing just
how far the kidney is affected. A low phthalein output
with cumulative signs in the blood indicate a severe
degree of renal involvement. In mere passive conges-
tion these signs are not apt to be found.
Moderately advanced nephritis with slight heart fail-
ure may show a fairly good renal function, and, if so,
the prognosis depends more upon the response of the
heart to treatment than on the kidney. If the phthalein
output increases in such cases it is a favorable sign,
and if the phthalein output is fairly good in an appar-
ently severe cardiorenal case, the heart may be judged
the principal offender in the symptom complex. On
the contrary, if the phthalein output is very low and
there are signs of cumulation in the blood tests (-{-urea,
-{-rest nitrogen, low 8*) both kidneys and heart may be
* 5, symbol for freezing point of the blood.
Tests of Kidney Function 133
regarded as failing and the prognosis is grave.
As a rule in cases of pure myocardial insufficiency
the renal function tests give surprisingly good results.
When the kidney is passively congested for long periods,
however, as a result of heart failure, the functional
tests give low results just as they do in severe types
of nephritis. The prognosis becomes proportionately
lugubrious.
2. Surgical Application. — The surgeon is extremely
interested in the problems of renal function. With him
the question often becomes a very vital one in connec-
tion with important and serious operations upon the
genitourinary tract. The proper selection of tests is a
matter of great importance to the surgeon. Geraghty
rightly insists that it is only through familiarity with
the reliability, limitations, and significance of findings
of individual tests of renal function, in their relation to
various types of disease and various kinds of problems
to be solved, that a proper selection can be made.
Certain of the tests or combinations of tests are more
suitable to surgical investigations, while other tests or
combinations of tests may be of greater value to the
internist.
In discussing the utility of renal function tests to the
surgeon, we shall draw freely upon Geraghty, since he
more than any other investigator has identified himself
in his researches with the question of their value and
limitations in surgical practice. Some of the tests of
renal function are adapted to estimating the total func-
tional capacity of the kidneys at any given time. Other
tests are not of great use in this respect but are of
considerable importance in determining the relative
functional capacity of the organs when applied to the
excretion obtained by ureteral catheterization. Finally
there are a few tests which are apparently useful in
134 Manual of Vital Function Testing Methods
both cases. As an illustration, it is known that the
estimation of urea output in a 24-hour specimen is
of no great value by itself in judging the total func-
tional capacity of the kidneys, while comparisons of
urea elimination in catheterized specimens from both
sides may give very important information.
From a purely practical standpoint, as was men-
tioned under preliminary considerations, all tests for
kidney function come under two general heads: 1.
Tests to determine how much of a given substance is
excreted by the kidney or kidneys, comparing the
amount with what is known to be normal. The sub-
stance excreted may be one which is normally found
in the urine or it may be a substance artificially intro-
duced into the circulation to determine the capacity of
the kidneys to eliminate it. 2. The blood may be exam-
ined for substances which are normally passed through
the kidneys, with a view of detecting an accumulation
of such substances in the body, such accumulation being
regarded as an evidence of defective kidney function.
Renal functional tests arc especially valuable to the
genitourinary surgeon in two types of cases : 1st, Sur-
gical diseases of the kidney secondary to obstructions
in the lower urinary tract. 2nd, Unilateral and bilateral
surgical diseases of the kidney not associated with ob-
struction.
Rowntrce and Geraghty, in many of their publica-
tions concerning the phenolsulphonephthalein test, have
called attention to the fact that inasmuch as many cases
with obstruction in the lower tract also have hydroneph-
rosis, pyoncphrosis, pyelonephritis, or pressure at-
rophy, an examination of the total function by means of
the phthalein test will often show diminished functional
activity. In these cases the urinalysis may be mislead-
ing, for urea output and total solids secretion may be
Tests of Kidney Function 135
normal, and yet the kidney function may be so unstable
that the shock of a surgical operation, such, for exam-
ple, as prostatectomy, may be sufficient to inhibit func-
tion altogether, and death will result. In just such cir-
cumstances they believe a total examination by phtha-
lein will serve to differentiate these cases with severe
from those with slight renal involvement.
Experience has shown that cases of obstruction in
which the phthalein output is deficient may be so greatly
improved by preliminary treatment with proper drain-
age that the subsequent radical operation may be per-
formed with greatly diminished risk.
According to Geraghty the phthalein test affords the
truest index of functional capacity of the kidney for
surgical work. The diastase test and urea estimation
are, he thinks, of equal value, but are unreliable indices
of functional capacity ; but when persistently low
values are given by them they may be important from a
prognostic point of view. Estimations of blood urea
and incoagulable blood nitrogen are extremely impor-
tant when they are associated with the phthalein test.
When all three are positive they are extremely sig-
nificant.
The functional tests, one and all, can never be said
to arbitrarily answer the important question, when to
operate and when not to operate, because there are
other factors to consider in a surgical case besides renal
function. If the phthalein test is very low operation
should be postponed until efforts are made by improved
drainage (either by suprapubic cystotomy or by cathe-
ter) to bring about improvement in the functional
capacity of the kidneys.
In the second group of surgical cases above men-
tioned, namely, unilateral surgical kidney diseases, the
relative functional power of the kidneys is a question
136 Manual of Vital Function Testing Methods
of vital importance. It is here that ureteral catheter-
ization is of supreme importance, for by this means the
excretion from each kidney can be separately obtained.
Many functional tests, such as urea estimations, cer-
tain chromoscopic tests, as indigo carmin and methylene
blue, cryoscopy, diastase estimation, phloridzin reac-
tion, and experimental polyuria, give information only
with respect to the relative functional value of the two
kidneys. This is insufficient, since one kidney may be
doing a great deal more work than the other, and yet be
incapable of doing the work of both, and this is the
question of vital importance to the surgeon, requiring
a definite answer in each individual case. The phenol-
sulphonephthalein test has proven of special value here
because by it not only is an idea of the total functional
capacity, but also a quantitative estimate of the work
done by each kidney obtained.
There are two difficulties, however, which belong to
all tests of renal function when carried out with ureteral
catheterization ; one is inhibition of function produced
by the presence of the catheter in the ureter ; the other
is leakage around the catheter. Special catheters are
now used to prevent the latter difficulty. As to the first,
the best method of procedure is to test the total func-
tional capacity with phthalein before ureteral catheter-
ization is performed. If the total phthalein output is
nearly normal one kidney at least is satisfactorily nor-
mal. If, after ureteral catheterization, one kidney is
found with an exceedingly low output, which, together
with the clinical findings, indicates that this is the dis-
eased organ, it may be safely removed even if the output
in the sound side is low, for here the natural inference
is that inhibition has produced the deficiency. Inas-
much, however, as inhibition is not necessarily equal on
the two sides it will always be necessary to combine the
Tests of Kidney Function 137
phthalein test with comparison of diastase and particu-
larly urea percentages, on the two sides. The actual
procedure recommended by Geraghty is, first, the esti-
mation of total output by phthalein and the estimation
of relative function by ureteral catheterization com-
bined with pigment, diastase and urea estimation on the
two sides. If the phthalein output is low, cryoscopy of
the blood serum and estimation of blood urea may
profitably be done.
Geraghty says : "Most of the criticism of functional
tests has come from those who have not used them, and
are unfamiliar with the nature of the information sup-
plied. It is true that in the vast majority of cases a
successful nephrectomy on the diseased side from the
standpoint of renal function, can be performed when
the urine from the opposite kidney is found apparently
normal on analysis. Unfortunately, however, the prob-
lems are not always so simple. Cases of bilateral dis-
ease are encountered in which a knowledge of the renal
function becomes of absolutely vital importance, and in
which every source of information must be called upon
before the proper line of procedure can be employed.
Again, in certain cases, particularly those of tubercu-
losis, it may be possible to introduce a catheter only on
one side. In such cases one must depend to a great
extent upon the information derived from function tests
as to the condition of the opposite kidney since the cys-
toscopic appearance of the ureteral orifice is frequently
deceptive. The recognition of hypoplastic and infantile
kidney is practically impossible without functional esti-
mation. The infantile kidney is a particularly danger-
ous type because the urine which is secreted by the kid-
ney is apparently normal in every respect except that
of quantity. Functional estimation has proved also of
great value in the differentiation between pyelitis and
138 Manual of Vital Function Testing Methods
pyelonephritis. In pyelitis the renal function is prac-
tically normal, while in pyelonephritis there is dimin-
ished function."
Selection and Practicability of Renal Function Tests
In this regard we can do no better than quote the
words of Geraghty, spoken at the conference on Renal
Function Tests at the ninth triennial meeting of the
Congress of American Physicians and Surgeons,
1913 74:
"The number of functional tests has become so
great that it is impracticable to employ all of them in
any individual case ; and, even if not impracticable,
nothing would be gained by employing all of these tests.
The information furnished by many is of the same char-
acter, but more accurately furnished by one test than
by others. For example, there is a parallelism between
the excretion of the different dye substances; but as
phthalein furnishes more accurately all the information
obtainable from this group of substances, no advantage
attaches to the employment of all.
"For chromocystoscopy alone indigo carmin is un-
questionably the test of choice. The estimation of rest
nitrogen and blood urea bear about the same signifi-
cance. Lately we have discarded estimations of resid-
ual nitrogen in the blood and are depending entirely
upon the blood urea determined by Marshall's method
or upon cryoscopy for evidence of cumulative phe-
nomena.
"From a practical standpoint certain tests can be
entirely discarded without loss, such as cryoscopy of
the urine and electrical conductivity of the urine. Total
urea estimations in urine are of doubtful value and dias-
74 See Trans. Cong. Amer. Phys. and Surg., 1913, IX, 45.
Tests of Kidney Function 139
tase determination furnishes only information that is
obtainable more accurately and quickly by other means.
Certain other tests, such as potassium iodide elimina-
tion, can be discarded as furnishing at times unreliable
information. We have seen potassium iodide excretion
delayed in cases with normal function (proven by sub-
sequent history) and excreted within normal limits in
cases of the most severe nephritis. The tests which
we consider of the greatest value in the excretory group,
based upon actual experience, are: Phthalein, lactose,
and chlorides ; and of the tests of retention, blood urea,
rest nitrogen and cryoscopy. The indications for the
specific employments of the individual tests are as fol-
lows:
"Chloride estimation in the urine is useful in all
forms of nephritis and cardiorenal disease, especially
if oedema is present.
"Lactose is indicated for the detection of slight in-
jury to the kidneys and also in severe nephritis, since
its suppression indicates a bad prognosis. It is not par-
ticularly helpful in surgical diseases.
"Of the retention tests either blood urea, rest nitro-
gen or cryoscopy is indicated wherever there is a severe
lesion of the kidneys.
"We consider that one of these tests should be used
as a routine in conjunction with phthalein wherever
functional tests are desirable, particularly if the phtha-
lein function is low.
"Tests in conjunction with ureteral catheterization :
in this connection, phthalein, urea in urine, and urinary
diastase are most serviceable. The diastase and urea
give practically the same information, but only give
relative functional values, while phenolsulphonphthalein
p-ives relative and absolute values. The total function
O
should always be estimated by means of phthalein with-
14»0 Manual of Vital Function Testing Methods
out ureteral catheterization, in order to detect the
amount of catheter inhibition, should this exist. Where
severe bilateral lesions exist, one of the retention tests
should be used.
"As to practicability, the simplest and easiest test is
undoubtedly the phthalein test, as it requires the least
amount of time and apparatus.
"The lactose test, if quantitative determination is re-
quired, necessitates the employment of an expensive po-
lariscope. Furthermore the preparation of the lactose
for injection requires attention and consumes time. Its
use also requires familiarity with the technic of in-
travenous injection.
"Diastase requires the daily quantitative preparation
of soluble starch, accurately graduated pipettes, a large
series of test tubes, a water bath, and one-fiftieth nor-
mal iodine solution. For total estimation it requires
24-hour specimens of urine with preservatives. The
time necessary for a simple determination is scarcely
warranted by the information obtained. Urea estima-
tions of the urine can be accurately and rapidly done
by the Marshall method ; and from the standpoint of
practicability it leaves little to be desired. It is useful
only in conjunction with ureteral catheterization.
"Chloride estimation requires standardized solutions
and carefully graduated apparatus when accurately
done. It consumes considerable time, and, besides, re-
quires daily collections of the urine with the knowledge
of the daily chloride intake.
"All retention tests require, of course, the withdrawal
of blood ; and cryoscopy of the blood is undoubtedly the
simplest, provided that proper apparatus is at hand.
It requires careful attention to the details and con-
sumes considerable time.
"Blood urea can be done by either the Folin or the
Tests of Kidney Function 141
Marshall method, and the total rest nitrogen, preferably
by Folin's method ; but any of these methods is imprac-
ticable for the general practitioner.
"Where only one test can be employed the most value
is unquestionably to be obtained from the use of phtha-
lein ; and this is particularly so from the standpoint of
the surgeon."
CHAPTER III
TESTS OF PANCREATIC FUNCTION
GENERAL CONSIDERATIONS
As Stadmiillcr 1 rightly says, "The recognition in the
living subject of pathological conditions of the pan-
creas belongs, without doubt, to the more difficult prob-
lems of the diagnostic technic of the present day."
Aser 2 also said years ago that there is no organ in
the body in which such a disparity exists between its
known physiological importance and our capacity to
clinically estimate its functioning power.
In other words, the enormous importance of the pan-
creas as a digestive organ and as a gland of internal
secretion is granted, thanks to the work of Von Mering,
Minkowski, Pawlow, Boldyreff, and many others. Our
ability to estimate the functioning power, the physio-
logical capacity, the anatomical condition of the organ
in a given case is limited.
The importance of the pancreas as a digestive gland
is very great. It is the only gland which furnishes an
enzyme for each class of foodstuffs.
The activated proteolytic ferment trypsin breaks up
protein into simpler structures (amino acids) than the
gastric juice. Amylopsin or pancreatic diastase con-
verts starch into sugar. Steapsin or pancreatic lipasc
'Archiv. of Diag., 1911, IV, 20.
"Nothnagel's Spec. Path, und Therap., Wien, 1898, I.
142
Tests of Pancreatic Function 143
splits neutral fats into fatty acids and glycerine.
Glaessner has estimated that the pancreas pours into
the duodenum through the ampoule of Vater perhaps a
pint or more of its mixed secretion per day.
But beyond this great act, which a priori would ap-
pear to be more than sufficient for a single organ, the
pancreas elaborates a mysterious secretion which pre-
sides in an equally mysterious way over the sugar
metabolism of the body.
Von Mering and Minkowski in 1889 discovered that
typical diabetes follows total extirpation of the pan-
creas in dogs. This fact, which has received an immense
amount of experimental corroboration, still stands out
as an epoch-making discovery in the history of pan-
creatic physiology amidst much that is obscure and un-
certain.
It is pretty generally agreed that the internal secre-
tion of the pancreas is elaborated in the islands of
Langerhans, closely crowded groups of polygonal cells
without excretory duct of any kind scattered in the
stroma of the gland between the external secreting
acini and surrounded by a profusely developed net of
blood vessels.
Besides controlling the carbohydrate intermediate
metabolism of the body the internal secretion of the pan-
creas is claimed by Loewi and some others to exert an
inhibitory effect upon the sympathetic.
From the standpoint of the clinician it is an exceed-
ingly difficult matter to determine the existence of
pancreatic disease, and to ascertain its nature. All
the customary methods of examination are, of course,
employed in such a search. Inspection, palpation,
blood examination, and a careful study of the semiology
are made use of. The subjects of pancreatic semiology,
pain, tenderness, tumor, pressure, etc., and pancre-
144 Manual of Vital Function Testing Methods
atic exploration arc extremely important aids and in-
dispensable adjuncts in pancreatic diagnosis. They
do not come, however, under the functional investiga-
tion of pancreatic activity, and will, therefore, be
necessarily left undiscussed in this place.3
In considering the question of studying the functional
capacity of the pancreas, two possibilities become im-
mediately apparent. (A) Functional investigation
may be made of the pancreas regarded as an organ
of external or digestive secretion, and (B) the func-
tional examination may relate to an inquiry concerning
the internal secretory activity of the organ, its power,
in other words, of adequately performing its endocrin-
ous or metabolic function.
A. TESTS FOR PANCREATIC FUNCTION, WHICH CONCERN
THE EXTERNAL OR DIGESTIVE ACTIVITY OF THE ORGAN
These may be divided into four subdivisions, as fol-
lows:
1. Proteid digestion tests.
2. Fat digestion tests.
3. Starch digestion tests.
4. Demonstration of pancreatic ferments in secre-
tions and excretions.
The protcid digestion tests are: Demonstration of
waste muscle fibre in the stools, the so-called azotor-
rhoea or creatorrhoea. Schmidt's test for digestion of
nuclei. Sahli's glutoid capsule test.
The fat digestion tests are: Demonstration of ex-
cess fat in the stools, so-called steatorrhoea. Identifica-
tion of split fat in the stools. Winternitz Diagnosti-
cum.
8 Consult Opie's Text Book, Diseases of the Pancreas.
Tests of Pancreatic Function 145
The starch digestion test consists of identification of
undigested starch in the stools.
Besides the above tests there have been devised special
qualitative and quantitative methods of identifying the
various pancreatic ferments themselves (trypsin, amyl-
ase and lipase) in the stools, urine and gastric contents.
It is somewhat difficult, therefore, to separate the
operations of routine urinalysis and coproanalysis in-
tended to show the presence or absence of pancreatic
enzymes, from certain special tests which have been de-
vised with the express purpose of testing pancreatic
function. For example, Schmidt's test for nuclear di-
gestion, and Sahli's glutoid capsule test might, upon
superficial consideration, be considered as special tests
for pancreatic function, but in reality they are both
merely special methods of demonstrating the presence
or absence in the intestine of pancreatic enzymes.
However, the fact that in the performance of these
tests something particular is done in the way of special
preparation with a certain definite end in view, would,
in all likelihood, insure their inclusion in any scheme or
list of functional pancreatic tests.
All the other tests mentioned in the above synopsis
are, in reality, only special methods of urinalysis or
fecal examination.
The same difficulty of classification becomes apparent
when we come to consider the tests of internal secre-
tion. The Cammidge test, as it is called, is but a
specially technical urinalysis.
The Loewi test for pupillary reaction to adrenalin
and the test for alimentary glycosuria might be said to
answer all the requirements of special functional tests,
but if our knowledge of pancreatic function were lim-
ited to these experimental inquiries alone, it would be
146 Manual of Vital Function Testmg Methods
meagre indeed.
From all these considerations it will be apparent that
any attempt to give a description of methods of study-
ing pancreatic function must include all of the above-
mentioned methods, although a very strict interpreta-
tion might relegate some or many of them to the domain
which is usually covered by books dealing with chemical
or microscopical laboratory methods.
B. TESTS FOR PANCREATIC FUNCTION WHICH CONCERN
THE INTERNAL OR METABOLIC FUNCTION OF
THE ORGAN
There are three tests which belong to this class.
1. The Cammidge reaction.
2. Loewi's pupillary reaction.
3. Provocative alimentary glycosuria.
1. Proteid Digestion Tests. Estimation of Undigested
Protein in the Stools as a Means of Determin-
ing Pancreatic Hypofunction
The patient is placed for three days on Schmidt's
test diet. This diet is as follows: 1.5 liters of milk,
100 gins, of zwieback, 2 eggs, 50 gms. of butter, 125
gms. of beef, 190 gms. of potatoes, and gruel made from
80 gms. of oatmeal. In this diet are contained 102 gms.
of albumen, 111 gms. of fat, 191 gms. of carbohydrates;
making a total of 2234 calories of energy.
According to Schmidt the diet is distributed thro
the day as follows: Morning, .5 liter of milk (if milk
does not agree .5 liter of cocoa made from 20 gms. of
cocoa powder, 10 gms. sugar, 400 gms. water and 100
gms. milk are substituted), 50 gms. zwieback; noon, 125
Tests of Pancreatic Function 147
gms. chopped beef (raw weight) broiled rare, with 20
gms. of butter (the interior of the meat must be raw),
250 gms. of potato broth made of 190 gms. of mashed
potatoes mixed with 100 c.c. of milk and 10 gms. of but-
ter ; afternoon, same as morning ; evening, same as fore-
noon.
It would be well if investigators should agree to use
this well-known standard diet so that comparisons of
results could be made.
Pratt,4 while using the diet, recommends that the
whole quantity be given in three meals instead of five,
conforming thus to American custom.
The Schmidt diet, containing as it does a good
amount of all three varieties of foodstuffs, is adapted to
all tests which involve an examination of the feces.
The presence of azotorrhoea or defective proteid
digestion due to deficiency of trypsin zymase, is deter-
mined by examining the feces microscopically after the
test diet.
Meat consists of connective tissue and muscle fiber.
Connective tissue is digested promptly by the gastric
juice, the muscle fiber by trypsin. Large quantities of
striated muscle fiber in the feces may indicate defective
pancreatic activity.
The bulkiness of the stools should also be noted.
Oser, Musser and others have claimed that there is
no single symptom of pancreatic disease of greater
significance than bulkiness of the stools. The dried
weight of the stools may be rather readily ascertained.
A ventilating hood and scales are all that are necessary.
If the pancreatic juice is not appearing in the bowel the
dried stools will weigh more and the stools themselves
will be more than ordinarily voluminous.
In six healthy individuals on Schmidt's diet Pratt
4 Amer. Jour. Med. Sc., 1913, CXLIII, 313.
148 Manual of Vital Function Testing Methods
found the average weight of the dried feces to be 54.3
gins. The maximum was 62 and the minimum 45 gms.
Schmidt's Cell Nuclei Test for Pancreatic Suf-
ficiency.5— According to Schmidt, the nuclei of animal
cells is digested only by the pancreatic enzymes. Con-
siderable has been written pro and con concerning this
test and, although it does not appear to be extensively
used, there is no doubt that it is not devoid of a certain
value.
Technic. — Raw beef somewhat fibrous is cut into
cubes of .5 c.cm. These are hardened in alcohol and
placed in small bags of coarse silk gauze. Before using
they must be soaked several hours in water. The bags
are recovered in the stools, hardened, paraffined, cut and
stained, and the sections mounted and examined micro-
scopically for nuclei. If the nuclei are not digested,
they will take the stain and become visible. Thymus
gland has been substituted for the meat by Einhorn.
Kashiwado, one of Schmidt's pupils, has altered the
original test by giving a powder consisting of equal
parts of stained thymus nuclei and lycopodium. The
mixture is given in two capsules, each containing .25
gms, at the evening meal. The stool is examined for
stained nuclei.
In performing the Schmidt test the sac must be
recovered in the feces before the expiration of 30 hours
at the latest, otherwise a longer sojourn in the bowel
will enable the bacterial enzymes to dissolve the nuclei.
Sahli's G Glutoid Capsule Test of Pancreatic Func-
tion.— The glutoid capsules used in this test are made
of gelatin and hardened in formalin. This is intended
• Verhandl. d. Kong. f. inn. Med., 1904, XXI, 335.
6 Deutsch. med. Wchnschr., 1897, XXIII, 6; also Deutsch. Ar-
chiv f. klin. Med., 1898, LXI, 475.
Tests of Pancreatic Function 149
to make them resist digestion by the gastric juice, to
permit of their entering the bowel, there to be softened
and disintegrated by the pancreatic juice if it be pres-
ent in sufficient amount. It is the trypsin zymase which
affects this solution.
The capsules may be filled with sodium iodide, iodo-
form or salol. The last is usually employed. Salol is
split up by the pancreatic juice into salicylic acid and
phenol. The former is eliminated by the kidney and
escapes in the urine as salicyluric acid, which is easily
recognized in the excretion by the violet color produced
by adding to the urine a few drops of a solution of
ferric chloride.
Normally the reaction is obtained in five hours.
Sahli did not disclose the precise manner of preparing
the capsules and this, according to Pratt, has hindered
the generalization of the test which is so extremely
simple.
The capsules are made and sold by Hansmann of St.
Gallen, Switzerland.
Sailer 7 says that a satisfactory capsule may be
prepared by placing ordinary gelatin capsules in pure
formalin for three minutes.
This test is practically the same as that introduced
by Sahli for estimating gastric motility. Its extreme
facility of execution would make it an excellent test if
there were unanimity in the findings. Unfortunately
this is not the case and at the present time no one would
pretend to depend upon its results alone, though it
seems undoubtedly true that in the absence of pyloric
spasm or stenosis, the capsule test of Sahli is worth
considering from a corroborative point of view.
7 Amer. Jour. Med. Sc., 1910, CXL, 330.
150 Manual of Vital Function Testing Methods
2. Fat Digestion Tests. Demonstration of Excess of
Fat in the Stools (Steatorrhaea) and Diminu-
tion of Split Fats in the Stools as a Means of
Estimating Pancreatic Insufficiency
Here the insufficiency of pancreatic function, if it
exists, is reflected upon the fat-splitting activity of the
external secretion and shows itself by phenomena which
are due to diminution or absence of the lipasic enzyme
of the juice.
Steatorrhoea, or increased fat in the stools, is known
to be common in pancreatic disease. The light color
of the stools is an important macroscopic feature. They
may be almost white. The stools are often rancid in
smell and bulky. Since the stools in icterus may also
be fatty it may be necessary to test for bile in order to
make sure that the pale color is not due to hydrobiliru-
bin. When fat crystals are present in excess, the stools
have a metallic lustre like aluminum. A white stool,
fluid when passed and becoming solid on cooling, is fatty
and is said to be quite characteristic of defective pan-
creatic secretion. The most important function of pan-
creatic lipase is to split up the neutral fats into fatty
acids and glycerine. The free fatty acids combine with
the alkalies of the pancreatic juice to form readily
assimilable soaps.
If the functional activity of the pancreas is deficient,
one important consequence will be a considerable dim-
inution in the amount of split fats, and hence of
soaps. The fat will remain unsplit, therefore unassirnil-
able and consequently unabsorbed.
Normally from 7 to 11 % of the fat ingested in the
food escapes action by the pancreatic juice and is
passed in the stools. If bile is absent from the intestine
because of occlusion of the duct, there will be incomplete
Tests of Pancreatic Function 151
emulsification of the fats and consequent reduction of
pancreatic effects. Under such circumstances as much
as 45% of the fats ingested may escape in the stools.
If, in a case of icterus, the loss of fat in the stools is less
than 60% the pancreas is probably not implicated. If
the fat loss exceeds 60% it points to pancreatic disease.
Under the microscope, fat appears in the stools as
droplets, needle crystals, or as structureless plates or
flakes. Unfortunately the quantitative estimation of
fat in the stools is not a simple matter. The stools must
first be dried on a water bath; the neutral fats and
fatty acids are extracted in a Soxlet apparatus with
ether, from a known quantity of dried stool ; the residue
is treated with diluted HC1 to convert soaps into fatty
acids and these are extracted in the same manner. The
amount of free acid in the first extract is calculated with
titration with alkali.
Like the Cammidge reaction or test (v.i.) the quanti-
tative estimation of fat in the feces is not to be regarded
as a clinical procedure likely to be carried out by the
physician. The method requires some apparatus and
more chemical experience than the clinician is likely to
possess and, besides, is time consuming. Of course, in
many hospitals, the examination can readily be made.
It is not considered advisable to give the details of the
method for fat extraction and quantitative estimation,
the skeleton of which is above outlined, especially as
the details may readily be obtained from texts on
chemical diagnosis. For convenience two references
may be given — Sahli 8 and Wood.9
Winternitz's Test of Pancreatic Fat-splitting Func-
tion. Winternitz Diagnostikum. The Sajodin Test. —
This test is founded upon the fat-splitting power of
8 Diagnostic Methods, 190,5, p. 446.
9 Chemical and Microscopical Diagnosis, 1909, p. 334.
152 Manual of Vital Function Testmg Methods
steapsin to set free iodine from iodine-containing fats.
Winternitz was the first to apply the artificially iodized
fats to the investigation of the fat-splitting power of
the pancreatic juice. The first substance used was iodi-
pin, but it was found that the iodine present in this sub-
stance is so firmly bound that even normal pancreatic
juice may not split it. He finally selected monoiodo-
behenate of calcium or sajodin as the most suitable sub-
stance.
Sajodin is a thin, oily liquid containing 25 c/o of
iodine. If 3 c.c. of this substance are administered by
mouth to a fasting individual and the urine examined
for iodine 3 to 5 hours later the reaction will be nega-
tive. If the same substance in the same amount is
given with a meal, the iodine reaction will be present.
In the first instance no pancreatic secretion has been
stimulated to appear in the duodenum and consequently
the sajodin is not split. In the second instance the
opposite prevails.
It was further found that in cases of icterus no
splitting of sajodin takes place because of the absence
of bile salts in the intestine, which are necessary to
actuate the fat-splitting ferment and to stimulate ab-
sorption.
Several investigators have investigated the Winter-
nitz test and the question has been made the subject
of a thesis by Stegman.10 He concludes that the failure
to find iodine in the urine 3-5 hours after the ingcstion
of 3-5 c.c. of sajodin with a meal, is indicative of lipo-
lytic pancreatic insufficiency in most cases, and that,
in combination with other well-known tests, the method
of Winternitz may be regarded as of considerable cor-
roborative value.
10 Uebcr eine neue Methode tier Pankreasfunktions prufung
Winternitz Diagnostikum. Dissertation Otto Stegman, 1911.
Tests of Pancreatic Function 153
In a recent inaugural dissertation upon the Winter-
nitz sajodin test by Syring,11 this author finds that
without exception in normal cases iodine appears in the
urine in 3-5 hours after the ingestion, with a meal, of 5
c.c. of calcium monoiodobehenate (sajodin).
The question as to its real value in determining the
existence of pancreatic insufficiency can only be settled
by further investigations.
3. Starch Digestion Test. Identification of Undi-
gested Starches in the Stools as an Evidence of
Pancreatic Insufficiency
The presence of starch in the stools is generally un-
derstood to be of little value in this connection. Nor-
mally scattered granules of undigested starch are to be
found and can be easily identified under the microscope,
when stained with iodine solution (Lugol's), which
colors them blue. Any great excess, however, may
fairly awaken suspicion, especially if there is persistent
diarrhoea and the condition tends to be permanent or
long continued. Under these circumstances it is legiti-
mate to conclude that there is pancreatic insufficiency.
4- Identification of Various Ferments. Tests Where
Examination Is Made for the Pancreatic Fer-
ments Themselves in the Excreta. The Meaning
and Interpretation of These Results in Relation
to Estimating the Pancreatic Function
The different pancreatic ferments can be demon-
strated by proper methods, in the stomach (after an
oil meal), in the urine, and feces.
Einhorn, Gross, and others have devised special ap-
11 Ueber die Funktionsprufung des Pankreas. Leipzig, 1913.
154 Manual of Vital Function Testing Methods
paratus for obtaining the pancreatic juice directly from
the duodenum. Einhorn's duodenal tube is used consid-
erably in this country, but more particularly for thera-
peutic purposes.
One objection which urges against all tests for tryp-
sin in the stools is that erepsin, a ferment coming from
the mucous membrane of the small intestine, may digest
albumen, etc., even in the absence of trypsin. The ob-
jection cannot be urged against the gastric estimations,
and, in fact, seems to be exaggerated even when applied
to fecal analysis methods.
Demonstration of Trypsin in the Stools. — Two
methods are chiefly used ; the Serum Plate method and
the Casein method.
The Serum Plate Method of Mutter and Schlecht.12 —
Trypsin acts upon the surface of serum agar plates.
Method. One drop of the stool obtained by a laxa-
tive (calomel or phenolphthalein) is placed upon a
Loffler serum agar plate and kept at a temperature of
55° to 60° C. for 6-12 hours in an incubator.
If trypsin is found in the stool there will appear a
depression or hole in the serum due to digestion by the
enzyme.
Ordinary diphtheria culture tubes have been sug-
gested by Stadmiiller as being sufficient for the purpose
of the test.
This test is simple and seems to be rather highly re-
garded (Brugsch, Hirshberg, etc.). Other practical
methods of demonstrating trypsin in the feces have been
devised. Arthur and Hubert add a 2% solution of
sodium fluoride to the stools, also fibrin, and incubate
at 40° for 24 hours. Crystals of tyrosin are formed if
trypsin be present. Abderhalden's technique has also
been employed, using glycyl-tyrosin. The Miiller-
12 Munch, med. Wchnschr., 1908, LV, 225.
Tests of Pancreatic Function 155
Schlecht method is sufficient, however, and much more
simple.
For a quantitative variation of the serum agar plate
method of stool examination for trypsin, put in one part
of the plate undiluted feces and in orderly sequence
in other portions of the plate use dilutions of the feces
1:10, 1 : 20, 1 : 100, 1 : 200, and note which still forms
indentation. If the stools are fatty the fat should be
extracted with ether.
The Casein Method of Demonstrating Trypsin in the
Stools. — Casein in alkaline solution is precipitated by
dilute acetic acid. If casein is digested by trypsin it
will no longer give the precipitation reaction with dilute
acetic acid. This is the foundation of a test devised
by Gross.13
In Gross' method the feces are mixed with an alkaline
solution of casein — .1% casein, .1% sodium carbonate.
Various quantities of the filtered feces are added to the
casein solution, incubated for an hour and tested with
dilute acetic acid. (v.i. )
Mette's tubes are also sometimes used in testing for
the presence of trypsin. They consist of glass tubes
about 1 or 2 mm. in diameter, containing coagulated egg
albumen. These are suspended in the dilute feces or
other solution to be tested for a fixed time and the
amount of albumen digested off, measured. The
strength of the ferment will be proportional to the
square of the length digested.
Technic of Gross' Quantitative Test for Trypsin. —
Prepare a .19c solution of casein by adding 1 gm. of
pure casein (Merck) and 1 gm. of sodium carbonate to
1000 c.c. of chloroform water. Place in a flask and
13 Arch. f. Exper. Path, und Pharm., 1907, LVIII, 157; also
Deutsch. med. Wchnschr., 1909, XXXV, 1706.
156 Manual of Vital Function Testing Methods
allow to stand for 24 hours, after which the solution
is shaken vigorously.
Five gms. of feces are placed in a mortar. Add 45
c.c. of \c/o solution of sodium carbonate. Titrate
thoroughly and filter. The first cloudy portion of
filtrate is discarded. The second is used.
To each of six reagent glasses marked for identifica-
tion add 10 c.c. of the casein solution. With graduated
pipette add respectively 1 gm., .5 gm., .25 gm., .2 gm.
and .1 gm. of filtered feces to specimen and mix thor-
oughly. Place all in incubator, adding to each 3 drops
of 1% acetic acid. Specimens in which the casein are
digested (presence of trypsin) will remain clear; others
are cloudy.
In normal stools, glasses 1 to 3 are clear, 4 to 6
cloudy. A trypsin unit equals the amount of feces
which digests 10 c.c. of starch casein solution. If .33
gm. feces which is diluted tenfold digests 10 c.c. of
casein solution, there are 30 trypsin units, which is nor-
mal. In clinical work, disease of the pancreas may be
suspected when no trypsin or, at most, 10 units are
found in examination of the feces.
Demonstration of Trypsin in the Stomach Contents.
Method of Boldyreff-Volhard.14 — Boldyreff noted in
1904 in Pawlow's Institute that feeding olive oil to dogs
caused regurgitation of duodenal contents into the
stomach. Volhard 15 applied the principle to the clinic.
A breakfast of 200 c.c. of olive oil is given by stom-
ach tube or 250 c.c. of cream may be substituted, the
latter being swallowed. Half a tcaspoonful of magnesia
usta are given just prior to the meal and is repeated
"Ccntrbl. f. Physiol., 1904, XVIII. 457; also Zentbl. f. Phys.
und Path. d. Stoffw., 1909, III, 209.
15 Munch, med. Wchnschr., 1907, LIV, 403.
Tests of Pancreatic Function 157
twenty minutes afterward. This is to prevent acidifica-
tion. At the end of 45 to 60 minutes, the stomach con-
tents are removed by tube. Usually a liquid is obtained
which tends to separate into two layers, the lower one
containing the duodenal juice.
The presence of trypsin can be demonstrated by the
Gross casein test described above or by that of Arthur
Hubert,16 previously mentioned, the details of which
follow :
Fresh fibrin obtained from horse blood by whipping
and washing the coagulum is covered with 2% solution
of sodium fluoride and kept for 24 hours at 40° C.,
then filtered.
The fluid to be examined is diluted with equal volume
of 2% sodium fluoride solution, and one volume of this
dilution is added to two or three volumes of the fibrin
solution and digested at 40° for some hours. Crystals
or crusts of tyrosin form on the wall of the vessel.
According to Sahli trypsin can qualitatively be most
easily demonstrated by digesting in alkaline fluid at
incubator temperature a flake of fibrin stained with
magenta red. The fibrin becomes digested and dissolved
and the fluid is colored red.
Stadmiiller mentions a simple qualitative test devised
by Von Oefele. A few drops of Fehling's alkali solution
with a few drops of a 1-1000 solution of casein are
added to a .07% copper sulphate solution and .1%
sodium carbonate. The mixture is incubated at 55°.
To 5 c.c. of this, in a warm test tube, are added five
drops of the fluid (intestinal juice) to be tested, the
whole being shaken. If trypsin is present the solution
which at first is blue or green if bile is present becomes
red-violet or rose color.
"Archiv. de Physiol., 1894, 622.
158 Manual of Vital Function Testing Methods
5. Estimation of Diastatic and Lipolytic Ferment in
the Feces as a Measure of Pancreatic Function
The results obtained by a study of the diastase con-
tent of the stool should, provided all controllable fac-
tors in performing the test are standardized, give val-
uable information concerning the functional integrity
of the pancreas. The physiological basis upon which
the test is founded is, that practically the whole amount
of diastatic ferment found in the feces is of pancreatic
origin, i.e., provided certain possible sources of error
are understood and obviated (Wohlgemuth).
Several satisfactory methods for estimating diastase
in the feces have been devised, chief of which are those
of Wohlgemuth 17 chiefly used in Germany ; that of
Durand,18 chiefly used in France and England, and that
of Brown,19 chiefly used in the United States. The
methods of Durand and Brown will be described.
Durand's Method of Estimating Diastase in the
Feces, — One c.c. of the total diluted feces is added to
50 c.c. of starch solution (1c/c starch and 2% decinor-
mal HC1). The tubes are incubated for half an hour
at 39.5° C. and digestion is then stopped with three
drops of strong soda solution. The sugar formed is
estimated quantitatively with Fehling's solution.
Ten c.c. Fehling's solution is reduced by .0124 gm.
of sugar. If x be the number of c.c. of the incubated
mixture used to reduce the Fehling's solution, the
amount of sugar present in the 51 c.c. of the mixture
.„ , .0124 X 51 .
will be — or, in the whole amount or the
x
"Biochem. Zeitsch., 1908, IX, I; also ibid., 1909, XXX, 432.
"Archiv. des Mai. d. Appnr. Digest., 1911, V, 76.
19 Johns Hopkins Hosp. Bull., 1914, XXXV, 200.
Tests of Pancreatic Function 159
.0124 X 51 X 20,000. „,.
teces (obtained as below), ims
x
figure is multiplied by two to give units of grams of
sugar formed in the hour. The normal limits are 1500-
2000 units.
The feces for the test are obtained as follows: The
patient is well purged 12 hours after his last meal. He
is then given % liter of milk and 45 minutes later 50
grams sodium sulphate in water, and one-half hour after
this a glass of vichy. The feces of the next 3^/2 hours
are passed into a vessel containing ice. They are then
diluted with water to 20 liters and tested as above.
Technic of Brown's Test. — The patient is given a
high enema the night before. The evening meal should
be very light. At 7 A. M. the next morning 750 c.c. of
milk are given. At 7:30 and again at 8:30 A. M. 1/2
ounce of Epsom salts are taken. At 8 :30 a glass of
water containing 1/4 teaspoonful of sodium bicarbonate
is swallowed.
All stools up to 2 P. M. are saved in a vessel con-
taining 2 ounces of toluol which is kept in a cool room
or on ice. If less than 400 c.c. of stool are obtained
an enema of a pint of water is given. The average
quantity collected up to 2 P. M. will be from 400 to
1100 c.c. usually.
The stool should be examined as soon as possible
after 2 P. M. Dilute the amount up to 3000 c.c. with
normal salt solution, stir the whole amount until abso-
lutely homogeneous. Centrifugalize a portion for 5
minutes and use the supernatant fairly clear fluid for
testing.
Diminishing amounts of the fluid are put into a series
of tubes, 1.8 c.c. in the first, 1.6 c.c. in the second, 1.4
c.c. in the third, 1.2 c.c. in the fourth, 1 c.c. in the
160 Manual of Vital Function Testing Methods
fifth, .8 c.c. in the sixth, .6 c.c. in the seventh, .4 c.c. in
the eighth, .2 c.c. in the ninth, .1 c.c. in the tenth, .05
c.c. in the eleventh and .025 c.c. in the twelfth. Bring
the fluid in each of the tubes up to 2 c.c. with normal
salt solution. If the test shows a negative reading in the
first tube or if a low reading is expected, a supplemen-
tary set of tubes is prepared containing respectively, 2
c.c, 3 c.c., 4 c.c., and 5 c.c. centrifugalized mixture.
In each of the tubes is added 2 c.c. of 1% solution
of soluble starch (Kahlbaum) and the tubes are incu-
bated at 38° C. in water bath i/o hour, then cooled by
adding a little tap water and by holding them under
the cool tap. They are then quickly tested with a few
drops of one tenth normal iodine solution. The limit
is held to be that tube before the one in which the first
definite blue color appears.
Demonstration of Lipolytic Ferment. — Two simple
tests for the presence of lipolytic ferment may be men-
tioned. These are the Grutzner-Gamgee 20 method and
the von Oefele 21 method.
The first is as follows : An emulsion of ten parts of
oil, five parts of gum and thirty-five parts of water is
prepared. A neutral solution of litmus is made up
which in test tubes of 12 mm. diameter appears violet
against white paper. Ten c.c. of litmus solution and
5 drops of the emulsion are placed in several of these
tubes and increasing quantities, 2, 4, 8, 16, 32 drops,
of the fluid to be tested are added to the successive test
tubes. These are put in a water bath at 37° C., and af-
ter a short time the tubes are compared. If any fat-
splitting ferment is present the color of the fluid will
have turned redder the larger the amount of solution
added.
20 Quoted by Stadmiiller, loc. cit.
21 From Sahli's Diagnostic Methods, 421.
Tests of Pancreatic Function 161
Von Oefelc's method of demonstrating steapsin is as
follows : sweet butter is melted and the resulting clear
fat mixed with an equal proportion of a \cfo aqueous
solution of potassium carbonate and some phenolphtha-
lein, and then titrated with a soda solution until there is
a red tint. This liquid is heated in the incubator to
55° C. and 5 c.c. of it well shaken in a warm test tube
with 5 drops of intestinal juice. In the presence of a
normal amount of steapsin the red tint will disappear
in from 2 to 5 minutes. According to the rapidity of
the discoloration the quantity of actual steapsin can
be estimated.
C. TESTS FOR PANCREATIC FUNCTION WHICH CONCERN
THE INTERNAL, OR METABOLIC FUNCTION OF
THE ORGAN
All the functional tests of pancreatic activity which
have been described have related to the external secre-
tion with its enzymes, which is poured out thro the
pancreatic duct and the ampoule of Vater into the
small intestine.
There is, however, another phase to the question of
functional deficiency of the pancreas and this relates
to its internal secretion, which in some mysterious or
quite unknown manner presides over the mobilization
and destruction of sugar in the body. When the in-
ternal secreting function of the pancreas is lowered,
the power of assimilation of carbohydrates is reduced
and when a certain limit is reached a hyperglycemia
results which tends at a certain point to manifest itself
by the elimination of sugar in the urine, glycosuria,
diabetes.
There are three functional tests which concern par-
ticularly the internal function of the pancreas. These
are:
162 Manual of Vital Function Testing Methods
1. The Cammidge Reaction.
2. Loewi's Pupillary Reaction.
3. Provocative Alimentary Glycosuria.
1. The Cammidge Pancreatic Reaction
There is still much dispute as to just what position
the reaction holds in the clinical diagnosis of pancreatic
function.
The original Cammidge reaction consisted of two
parts or analyses which were known as A and B tests.
It was held that the presence of a pancreatic lesion
and even its nature could be determined by these tests.
The original theory upon which the test was based was
about as follows. If there is a real pancreatic lesion,
the pancreatic juice will escape into the parenchyma
of the organ and lead to fat necrosis with splitting
of neutral fat into fatty acids and glycerine.
The fatty acids will remain in the necrotic areas
and the glycerine will be absorbed into the blood and
excreted by the urine. The Cammidge test was de-
vised to demonstrate the presence of glycerine in the
urine by the presence of glycerosazone crystals.
The original theory was subsequently modified and
the two original tests were abandoned and replaced
by one process known as the C test. The present
theory of the Cammidge test is this. The crystals
produced in the test when positive, result from the
presence in the urine of a sugar complex which upon
hydrolysis with HC1 yields a substance giving a pentose
reaction. The crystals of a positive reaction are be-
lieved to be pentosazone.
The pancreas contains four or five times as much
pentose as any other organ in the body and conse-
quently when any disintegration of pancreatic tissue
Tests of Pancreatic Function 163
takes place as a result of disturbance or disease of the
organ, crystals of pentosazone, the Cammidge crystals
will be demonstrated in the urine. Neither a mere
blocking of the pancreatic secretions nor a pure fibrosis
of the organ will produce a positive reaction. It is
usually held as Cammidge himself believes that a posi-
tive reaction is evidence of active degeneration such
as occurs in acute or chronic pancreatitis. A negative
reaction contraindicates active degeneration but does
not exclude old pancreatitis nor malignant disease of
the pancreas. In fact, in 75% of malignant cases the
reaction is negative. But the test is not always posi-
tive even in pancreatitis and a negative test does not
definitely exclude pancreatitis. The results must al-
ways be taken in conjunction with clinical, urinary and
fecal findings. The test is not considered generally by
pathologists or clinicians as having great practical
value in diagnosis. But when positive it constitutes an
interesting abnormality which seems to be connected
in some rather cryptic or obscure manner with dis-
turbances of the pancreatic function.
Teclinic of the Cammidge Test. — Filter a portion of
a 24-hour specimen of urine.
Test for Albumen. — If albumen is present in amount
more than a trace, measure out 50 c.c. of filtrate and
add a few drops of acetic acid, boil, cool, filter and
make up to 50 c.c.
Test for Sugar. — Either Fehling's or Nylander's
test is performed. The result must be absolutely nega-
tive. If there is any reduction on standing about 50
c.c. of the albumen free urine must be mixed with
yeast fermented for 12 to 2-1 hours and filtered.
Stage I. Measure 20 c.c. of the clear albumen and
sugar free filtrate into a small flask with an in-
verted filter funnel placed in its mouth as a condenser.
164 Manual of Vital Function Testing Methods
Add 1 c.c. of strong HC1. Boil on sand bath for 10
minutes from commencement of ebullition. The boil-
ing should not be too vigorous and the flame should
be turned low for the greater part of the time.
Stage II. Cool under the tap. Make up contents
to 20 c.c. with distilled water. Slowly add 4 gms. of
lead carbonate ; shake gently at first and more thor-
oughly later. Stand, and shake occasionally until no
more gas comes off. Filter through a paper moistened
with distilled water.
Stage III. Add 4 gms. of powdered tribasic acetate.
Shake thoroughly for some minutes and allow to stand.
Filter thro a moistened filter paper.
Stage IV. To the clear and almost colorless filtrate
add 2 gms. of powdered sodium sulphate, shake thor-
oughly for several minutes. Bring slowly up to the
boiling point on a sand bath, shaking from time to
time. The excess of lead is removed at this stage and
it is important that the shaking and heating should
be done carefully.
Stage V. Cool under the tap and filter. Measure
10 c.c. of clear filtrate. Make up to 18 c.c. with dis-
tilled water. Add 8 gms. of phenyl-hydrazine hydro-
chlorate, 2 gms. powdered sodium acetate and 1 c.c.
of 50^? acetic acid.
Boil in a flask with a funnel condenser on the sand
bath for 10 minutes from the commencement of ebulli-
tion. Do not boil too vigorously. Filter hot through
a filter paper moistened with boiling distilled water
into a 15 c.c. measure. Should the filtrate fail to reach
the 15 c.c. mark make up to 15 c.c. with hot distilled
water. Stand for from 4 to 5 hours or longer at room
temperature or in ice chest.
Examine the filtrate for the appearance solubility
and amount of crystal formation.
Tests of Pancreatic Function 165
The typical crystals examined under the microscope
are of the osazone type and more circular and tuft-like
than glucosazone crystals. Run under the cover slip
33% H2S04; the crystals should dissolve in 10-15 sec-
onds. The crystals have to be distinguished from the
coarse yellow needles which may be deposited if the ex-
cess of lead was not removed in Stage IV. In a strongly
positive reaction the deposit of crystals may occupy
half the bulk of the filtrate. In a completely negative
reaction the filtrate remains clear.
*2. Loewi's Pupillary Symptom or Test of Pancreatic
Insufficiency
In 1908 Loewi 22 made the observation that after re-
moval of the pancreas in certain animals, the instilla-
tion of adrenalin into the eye will cause dilatation of the
pupil. Ordinarily the instillation of adrenalin into the
eye does not cause dilatation altho intravenous injec-
tion will do so. Loewi attributed the mydriasis fol-
lowing instillation to increased excitability of the sym-
pathetic system brought about by the removal of the
inhibitory effect of the pancreatic internal secretion.
From this fact it was thought that a mydriasis in man
following local instillation of adrenalin would indicate
pancreatic internal insufficiency, provided hyperthy-
roidism or Graves disease did not exist.
According to Sladden 23 this test has given interest-
ing and encouraging results and should not be dis-
missed with the comparatively scant attention it has
received lately.
The technic of the test is extremely simple since it
consists merely of dropping into the conjunctional sac
22 Archiv. of Exper. Path, and Pharm., 1908, LIX, 83.
"Quart. Jour, of Med., 1913-14, VII, 455.
166 Manual of Vital Function Testing Methods
a few drops of a 1 : 1000 solution of adrenalin and ob-
serving the effects upon the pupil.
3. Spontaneous and Provocative Alimentary Glyco-
suria in Their Relation to Pancreatic Function
The intimacy of relationship between the pancreas
and control of the carbohydrate metabolism is close
and undisputed, but what particular cells of the pan-
creas are concerned or just what the mechanism of the
control may be is very imperfectly understood. Glyco-
suria is usually present in many of the more serious
pancreatic diseases, but glycosuria is by no means an
infallible index of either the extent or the nature of
the pathological processes. In many cases of pan-
creatic disease, however, glycosuria does not appear
(e.g. many cases of chronic pancreatitis, carcinoma,
etc.). According to Cammidge the presence of glycosu-
ria means only a one to three chance that the pancreas
is diseased, and in case of pancreatic disease about one
in fourteen shows sugar in the urine. The truth is
well expressed by Sladden when he says, "viewed arith-
metically, glycosuria is not a sign of great diagnostic
value." If, however, glycosuria either spontaneous or
provocative be present together with other confirmatory
evidence of pancreatic disease the symptom then ac-
quires a greater value.
It must be remembered that the liver is also concerned
to a very important and intimate extent with carbo-
hydrate metabolism, and tests for alimentary glycosuria
have long been employed with a view of estimating the
functional integrity of that organ. Under a previous
chapter these tests have been given and discussed.
It must be perforce admitted that the question of
applying the so-called carbohydrate tests for provoca-
Tests of Pancreatic Function 167
tive glycosuria to the elucidation of pancreatic func-
tion is one which for the present must be left open.
It would appear rational to assume that the pres-
ence of glycosuria after the provocative tests indicates
either hepatic or pancreatic insufficiency or both, and
such tests are never in themselves sufficient to elucidate
the problems involved, but where they are corroborated
by other more specific evidences of insufficiency, they
assume an importance in diagnosis of no mean value,
an importance which is entirely lacking to them when
interpreted alone.
General Conclusions Concerning the Tests for
Pancreatic Insufficiency
It is certainly true that no one functional test of
pancreatic activity so far devised constitutes an abso-
lute or pathognomonic sign of disease of this organ.
It is quite natural that this should be so in view of the
manifold functions of the pancreas.
In other words it is often a very difficult question
to determine in a given case whether the pancreas
is diseased or insufficient at all, much less to make out
by means of the most complete and comprehensive
semeiological study the precise nature and extent of
the pathological processes.
The whole subject of pancreatic clinical pathology
and diagnosis and that of tests for functional activity
of the organ is in its infancy.
Nevertheless the value of those functional tests so
far devised is considerable and it is the duty of clini-
cians to apply them in practice to such an extent that
their individual and collective worth or lack of worth
may be definitely determined.
CHAPTER IV
TESTS OF HEART FUNCTION
GENERAL, CONSIDERATIONS
IN recent times a considerable transformation has
occurred in the viewpoint of clinicians toward the
cardiopathies and cardiac pathology. A few years
ago all cardiopathology was discussed in terms of
anatomical lesion. The chief interest lay in the exact
localization and delimitation of the lesion. The ten-
dency at the present is to bring more and more into
the foreground the idea of functional capacity of the
organ. Recent studies in cardiac physiology and
pathology have shown that the fundamental factor is
the muscle itself rather than its innervation as was be-
fore believed. It has been likewise shown that the func-
tion of the cardiac muscle is a complex one and that
at least five subdivisions of function may be made
(Englemann). These may be enumerated. The heart
muscle possesses the power of originating contractile
impulses (impulse formation, chronotropic function) ;
it possesses the faculty of susceptibility to the receipt
of these functions (excitability, irritability, bathmo-
tropic function) ; it is endowed by means of the conduct-
ing system of fibers including a histological differenti-
ated tissue, the bundle of His with power to transmit
these impulses from the point of their formation, the
168
Tests of Heart Function 169
sino-auricular node to the cardiac muscular fibers (the
conducting function, conduction, dromotropic func-
tion) ; it possesses the fundamental power of contrac-
tion (contractility, inotropic function), and finally, the
muscle possesses that vital function by which it nor-
mally refuses to dilate beyond a certain point (tonic-
ity).
A perfect method of estimating cardiac function
would be one in which all these five functions of the
cardiac muscle could be separately measured. The
function of the entire organ would be then their arith-
metical sum, if all possessed some degree of integrity
or their algebraic sum if certain of them were below a
normal point which might be diagrammatically repre-
sented by zero. But as a matter of fact we are far
from being able to accomplish such an estimate of
cardiac function at the present time.
There are many instances, however, in which a care-
ful physical examination of the organ together with
the use of special methods of cardiac investigation
which have come into use in recent years (sphygmo-
graph, electrocardiograph, sphygmodynamometer, the
X-ray) will enable the physician to conclude that one
or more of the five functions of the cardiac muscle are
deficient.
When some or all of these functions have become
so insufficient and incompetent that the occurrence of
heart failure (asystole) is imminent, the symptoms
of the condition (cyanosis, decompensation, edema,
signs of venous congestion) become so patent and evi-
dent that the presence of cardiac insufficiency is simple
to recognize from a study of the physical signs and
the symptoms.
This stage of true insufficiency may be called a ter-
minal stage of the cardiopathies no matter what the
170 Manual of Vital Function Testing Methods
anatomical lesion may be in a given case. This stage
is known, however, to be preceded by a long period of
latency in which cardiac insufficiency, if it is present,
cannot be so easily discovered and has to be looked
for in order to be recognized.
It is the desire of the clinician to increase his powers
of observation, to so lengthen as it were his cardiac
vista, that he may be enabled to recognize the earliest
signs of cardiac incompetency. It is for this very evi-
dent reason that tests for estimating the integrity of
heart functions have been devised. As yet no one of
them has succeeded in providing an entirely adequate
means of obtaining this much to be desired end, never-
theless several interesting and valuable methods have
been developed. The work of the past gives promise of
future developments and improvements in this extreme-
ly important domain of functional diagnosis.
To Rosenbach is generally given the credit of in-
sisting upon the necessity of devising proper tests
by which the functional integrity of this most im-
portant organ, the heart, might be measured.
The various methods of testing the cardiac func-
tion may be divided into a few classes. The first and
largest group includes those tests which depend upon
the reaction of the heart muscle to various types of
exertion active or passive. There is a second and
much smaller group based upon the behavior of the
heart to reflex stimulation. A third and extremely
insignificant group includes but one test, based upon
the supposition that sodium chloride elimination is
effected by cardiac insufficiency. A fourth group in-
cludes modern clinical and instrumental methods of
investigating cardiovascular conditions so far as they
are concerned with the question of elucidating heart
functional power.
Tests of Heart Function 171
The following synopsis shows how the various tests
are to be placed in the four categories above men-
tioned.
I. Reaction to muscular exertion active or passive
as a basis for estimating cardiac function.
1. The staircase test.
2. Graupner's test.
3. Mendelsohn's test.
4. Katzenstein's test.
5. Herz's self-checking test.
6. Gymnastic resistance test.
7. The Russian test — "Holding the breath" test.
8. The Venous pressure test.
II. Application of cardiac reflex estimations in de-
termining heart function.
Merklen's test.
III. Estimation of sodium chloride elimination as a test
of cardiac sufficiency.
Vaquez-Digne test.
IV. Modern clinical and instrumental methods of in-
vestigating cardiovascular conditions : their
applicability to estimating cardiac function.
1. The sphygmomanometer as an index of cardiac
function.
2. Rontgenoscopy and Rontgenography as indices
of cardiac function.
3. Sphygmocardiography and electrocardiogra-
phy ; their relation to cardiac functional
capacity.
I. REACTION TO MUSCULAR EXERTION ACTIVE OR PAS-
SIVE AS A BASIS FOR ESTIMATING CARDIAC FUNCTION
The majority of the methods so far suggested for
estimating heart functional power have consisted in
172 Manual of Vital Function Testing Methods
the subjection of the patient to a certain measured
degree of physical exertion followed by the systematic
observation of the phenomena produced by the exer-
tion as compared with conditions carefully ascertained
prior to the beginning of the test.
All these methods, however, have the common objec-
tion that the same work prescribed to different indi-
viduals, will under normal circumstances produce quite
different results, according to certain circumstances,
among which are the size and general muscular
strength of the individual, the usual mode of life with
respect to physical exertion, the condition of the nerv-
ous system, etc. The result, therefore, of exertion
tests may not always be comparable even in healthy
persons. If these factors can be properly estimated
and provided for the exertion tests are rendered more
certain and hence more useful.
Herz has emphasized the fact that the cardiac phe-
nomena produced by exertion tests are varied by the
type of effort attempted: whether, for example, the
movements are rhythmical and gymnastic, whether they
are resisted or not, and especially whether the muscular
groups called into play are weaker or stronger. A
much higher rise of blood pressure is produced by the
effort attempted by a weaker set of muscles than the
same operation performed by a stronger set. Grebner
and Grunbaum have contended that the increase in
blood pressure produced by muscular contractions is
inversely proportional to what may be termed the
specific energy of the muscles employed in the effort.
The influence of psychic factors in varying the re-
sults of muscular tests has always been recognized
(Kornfelds). The same may be said of the nerve
factors. All these facts tend of course to render un-
certain the results obtained by the exertion tests (pulse
Tests of Heart Function 173
rate, blood pressure), but even with these defects this
type of method of estimating cardiac function is of
practical value.
The chief points taken into consideration in this
type of test are the rate of the pulse, the blood pressure
(systolic and diastolic) and the area of cardiac dull-
ness or size of the heart (percussion, rontgenography).
As long ago as 1833 Donnell showed that the pulse
rate is normally slower in the recumbent than in the
semi-erect and erect positions. Christ in 1894 l pro-
posed to register the exact pulse rate after exertion
with the sphygmograph provided with a time marker.
He likewise invented an apparatus by which the patient
could undertake a measured amount of exertion on
a steppage machine. Rosenbach in the same year em-
phasized the importance of noting the condition of
the skin after the performance of the exertion. It
was his belief that the skin remains dry if the heart is
competent but becomes quickly moistened with perspir-
ation if there is cardiac insufficiency. He explained the
increased sudation on the ground that the excretory
function of the skin is called from the list of reserve
forces to compensate as far as possible for the cardiac
inadequacy. This is probably not the correct interpre-
tation of the phenomenon since sweating itself re-
quires the expenditure of force. It is probably due
to vasomotor causes.
1. The Staircase Test. Selig's Test 2
Technique. — Count the pulse and take the systolic
pressure. Have the patient ascend a flight of twenty
steps, rapidly. Count the pulse and take the systolic
pressure after the ascension.
'Archiv. f. klin. Med., 1894, LIU, 1902.
2Prag. med. Wchnschr., 1905, XXX, 418, 432.
174 Manual of Vital Function Testing Methods
Under normal circumstances, there is an increase
in the pulse rate of 20 beats per minute on an average
and a rise of blood pressure of 8 millimeters of Hg.
If the myocardium is insufficient there will be an
increase in the pulse rate of 30 beats per minute or
more. The blood pressure rise will be slower and
average about 6 mm. of Hg. The rise may be fol-
lowed rather suddenly by a fall below normal or the
preliminary rise may be absent.
The length of time required for recovery to the
normal systolic pressure may be taken as a measure
of the amount of cardiac insufficiency present.
The staircase test on account of its simplicity is often
employed by clinicians.
The "hopping test" is a modification of the Selig
test, which has been used for years as a routine method
of eliciting a latent cardiac insufficiency. The patient
is instructed to hop 20 paces on one foot and a com-
parison is then made between the pulse rate before
and after the exertion.
One serious objection to the "hopping test" is that
the actual amount of work performed by the indi-
vidual to be tested cannot be computed. In the method
of climbing stairs, the amount of energy expended can
be approximately known. The amount of work done
in foot pounds will be equal to the product of the
weight of the individual in pounds into the number of
feet ascended.
The advantage of this simple test is that it can be
performed without any special apparatus, which is the
chief objection from a practical standpoint to some
of the functional cardiac tests which have been sug-
gested.
Tests of Heart Function 175
2. Graupner's Test 3
Graupner found at Nauheim in observing the reac-
tion of patients after the exercises carried out as a
part of the treatment of cardiopathies, that persons
with weakened hearts showed a different type of reac-
tion from those with normal or nearly normal myo-
cardia.
Under normal circumstances, as is well known, the
pulse rate and the systolic blood pressure rise after
exertion, returning to normal after a fairly short in-
terval. If the exertion is sufficiently prolonged and
arduous they may fall below the normal. Graupner
discovered that after the pulse rate has risen and again
fallen to normal after an exertion, the systolic pres-
sure rises gradually to a maximum, which is reached
in a few minutes, usually about six, declining to normal
in about 18 to 20 minutes. The rise of blood pressure
following the pulse rise is called the normal erholung.
In weakened hearts, even if the weakness is slight,
Graupner found that the erholung occurs but it is less
in amount than normal and is delayed beyond the
normal interval, usually to about 12 minutes. If the
heart is seriously weakened the erholung may be ab-
sent altogether, the blood pressure declining from the
start then gradually rising to normal. In normal cases
the pulse reaches its normal in 5 to 10 minutes.
Technique of Graupner's Test. — A Zuntz Ergom-
eter 4 of the bicycle or weight and pulley type is used
in conducting the test. The patient turns a wheel
which is supplied with a brake and adjustments for
measuring the amount of work expended. Tests are
3Bcrl. klin. Wchnschr., 1902, 174; Deutsch. med. Wchnschr.,
1906, XXXII, 1029.
* Centrbl. f. Physiol., 1898, 502.
176 Manual of Vital Function Testing Methods
made on successive days at the same hour. The work is
therefore done by the same muscle groups. It is im-
portant not to carry the work to the point of exhaus-
tion or strain. Mental excitement must be absent.
The pulse rate, blood pressure and size of the heart
are noted before and after the test.
Arm muscle work may be substituted for thigh muscle
work on the same machine and this was done by Graup-
ner in his later researches.
Cabot 5 and Bruce have recommended using a meas-
ured amount of stair climbing, which of course is a
more practicable and generally useful method. They
estimate the amount of work in foot pounds which is
readily computed by multiplying the number of pounds
the individual weighs by the number of feet ascended.
Graupner came to the following conclusions as a
result of his rather extensive investigations : If the
blood pressure remains constant after the exercise the
heart muscle is sufficient. If the blood pressure falls
after the exercise there is cardiac insufficiency. If the
blood pressure rises but returns to normal there is com-
pensatory sufficiency. If the blood pressure rises then
rapidly falls without a tendency to subsequent rise the
heart muscle is fatigued.
Graupner stated as his belief that if the pulse is
accelerated and the patient becomes short of breath
after the performance of work equivalent to 45 to 300
kilograms the heart is manifestly insufficient.
Several authors have corroborated Graupner's view
that a persistent tendency toward a fall of blood pres-
sure after the exertion denotes cardiac insufficiency.
According to Graupner's later observation persons
with normal hearts can perform arm muscle work on
the ergometcr equivalent to from 3,000 to 20,000 kilo-
• Amer. Jour. Med. Sc., CXXXIV, 1907, 491.
Tests of Heart Function 177
grams per hour. If the figures fall below 1000 kilo-
grams per hour there is cardiac insufficiency. If meas-
urements are made every half minute after exercise it
was found that the amount of variability in the blood
pressure corresponds with the insufficiency of the heart,
in other words, that the greater the heart weakness
the greater are the variations in the pressure and the
longer 'the time required for the status quo to be
restored. This does not wholly occur for 30 to 35
minutes.
Some observers have found that a lowering of the
blood pressure after exertion may be found in trained
athletes and according to the terms of the test this
should denote cardiac insufficiency. But as Hirsch-
felder 6 states, "the heart of the trained athlete is
habitually throwing out an amount of blood suited
not to the needs of the moment but to the needs of
the periods of exercise to which he has accustomed
himself. The systolic output is above normal when
the exercise (and hence the increased production of
C02) is slight. The heart is then able to take care of
the excess of C02 production in exercise without in-
creasing the output and hence the vasodilatation in
the muscles is the only factor influencing the blood
pressure. When the exercise becomes severe the other
mechanisms begin to play a role.
Also in certain patients with diseased hearts the
blood pressure has been found to rise, it is claimed,
because of high pressure stasis. This rise, however,
comes later than in normal cases.
Cabot and Bruce as a result of their trial of Graup-
ner's test in seventy-five experiments believe that it is
reliable. They say "the main outlines of Graupner's
contention can be easily verified by anyone. Run
6 Hirschfelder, Diseases of the Heart and Aorta, p. 286.
178 Manual of Vital Function Testing Methods
quickly up two flights of stairs and then stop and
count your pulse. After the immediate acceleration
is passed or during the slowing of the pulse following
it you will note that the heart beat and the strength
of the pulse become markedly exaggerated. One feels
the thump thump of the heart against the ribs much
more strongly after the pulse has almost or quite
reached its normal rate than during the period when
the pulse is most accelerated. . . . As regards the
phenomenon designated by Graupner as the normal
erholung we can verify his findings and we likewise
agree with him in the results of our experiments upon
seriously weakened hearts. In some of the cases be-
lieved by us from ordinary examination of the heart
to be normal there was considerable variation from
the ordinary curve of blood-pressure after exertion.
Cases of valvular disease with good compensation
showed, as might have been expected, a normal curve."
Tests like that of Graupner in which blood pres-
sure estimations are used as a criterion of cardiac
function are founded upon observations of Masing 7
and others that the normal blood pressure rises during
exercise and falls immediately afterward.
When a normal individual exercises with chest
weights, for example, the blood pressure may rise 10
to 30 mm. of Hg. If the individual is arteriosclerotic
the rise of blood pressure may go to 40 to 60 mm. of
Hg. and outlast the exercise a variable length of time.
Bauer in employing this test used a stationary bicycle
and this is a good method because the blood pressure
estimations can be easily taken in the arm during the
performance of the exercise. According to Bauer the
bicycle test gives for normal individuals a rise of 5 to 10
mm. of Hg., while in those with cardiac insufficiency
' Dcutsch. Arch. f. klin. Mcd., Lcipz., 1901, LXXI, 253.
Tests of Heart Function 179
there may be a fall of equal degree (5 to 10 mm. of
Hg.)-
The great difficulty with this test is that it has been
found that in trained athletes the blood pressure may
fall instead of rising at the commencement of mild ex-
ercise and the fall may last for a considerable period,
thus making the reaction of the strong man some-
what similar to that of the weak. The proper interpre-
tation of this fact has be.en given.
As Hirschfelder states all functional tests of cardiac
efficiency if based upon mathematical changes in pulse
and blood pressure may lead to ambiguous results.
This is no objection, however, to the application of the
tests but only to their too strict interpretation. The
appearance of the patient after the performance of
the physical tests is of course extremely important.
Accelerated or labored breathing, holding the breath,
dilatation of the nostrils, drawing in of the corners of
the mouth, darkness or pallor of the cheeks, sweating,
palpitation and so forth, all these are signs of cardiac
insufficiency more important perhaps than the mathe-
matical results of individual tests.
According to Hirschfelder the most reliable nu-
merical criterion of cardiac efficiency is whether a given
strain causes the heart to diminish in size (increase
in tonicity) or to dilate (decrease in tonicity — over-
strain).8
3. Mendelsohns Test Q
Teclinic. — -The pulse is carefully counted in the
standing, sitting and recumbent postures and the fig-
ures noted. This may be repeated several times and
8 Hirschfelder, loc. cit., 199.
8 XIX Kongr. f. Inn. Med., 1901.
180 Manual of Vital Function Testing Methods
an average taken. The person to be tested then per-
forms muscular work upon a Gaertner ergostat by
means of which the amount of work may be measured.10
The Gaertner ergostat is an instrument not easily se-
cured and for this reason the original test is not much
employed. By the simple method, however, of Cabot
and Bruce above mentioned of having the patient per-
form a given amount of work in stair climbing which
can be easily calculated, the reaction of the pulse rate
and the return of the latter to normal, which is the
basis of the Mendelsohn tests, can be readily estimated.
After the performance of varying amounts of work the
patient assumes the recumbent posture immediately
and the time is noted which is required for the pulse to
return to the normal figure previously ascertained for
that posture.
The first criterion of the Mendelsohn test is based
on the principle that when the heart is healthy or well
compensated, a transition from vertical to horizontal
position is accompanied by a slowing of the pulse of
10 or 12 beats per minute. If the heart is insufficient
or decompensatcd an opposite condition may prevail,
namely, the pulse becomes quicker in the recumbent
posture or tends to remain constant.
Mendelsohn contended that if there is not a well
marked difference in the pulse rate between the erect
and recumbent postures the heart is incompetent.
The second criterion suggested by Mendelsohn de-
pends upon the principle that the competent heart
is able to return immediately to a normal when rest-
ing after a strain. He suggested, therefore, that an
estimate of the functional capacity of the heart can
be obtained by noting the degree of facility displayed
by the organ to return to normal conditions after meas-
10 Allg. AVien. Med. Zeit., 188T, Nos. 49 and 50.
Tests of Heart Function 181
ured exertion in which extra cardiac energy is called
into play. Mendelsohn found that a normal heart
after performing work equivalent to 100-200 kilograms
returns immediately to the normal with rest in recum-
bent posture. After 500 kilograms of work the normal
heart is accelerated somewhat for a varying period of
time. If, however, there is cardiac insufficiency very
much smaller amounts of muscular exertion prove exces-
sive and disturb the pulse rate. A disturbance of rate
with failure to return immediately to normal following
the expenditure of 25-50 kilograms of work denotes
cardiac insufficiency.
A Variant of Mendelsohn's Test. — When a normal
individual rises from the reclining to the standing posi-
tion the heart rate is accelerated, but it is usually
stated that the increase ought never to be more than
20. Beyond 20 one has the right to assume that the
myocardium is insufficient.
This test from its extreme simplicity has been much
used and is capable of giving some valuable informa-
tion. But nevertheless, under some circumstances it
fails to do so ; for example, under conditions where
the psychic role may play a part. Here the increase
of pulse rate in a normal individual, that is normal so
far as the myocardium is concerned, may be inordinate
and out of proportion. Hirschf elder X1 says that per-
sons with enteroptosis may give a false increase.
4- The Katzenstein Method 12
Katzenstein found from animal experiments that
ligature of peripheral arteries produces an increase in
11 International Clinics, Vol. IV, 1910, p. 39.
12Dcutsch. med. Wchnschr., 1904, Xo. 30, p. 807; also, ibid., 1907,
XLIV, No. 16.
182 Manual of Vital Function Testing Methods
the general blood pressure without change in the pulse
rate. In animals with weakened hearts he demonstrated
that ligature of peripheral arteries produced other re-
sults, namely increased pulse frequency and irregulari-
ties in the blood pressure. He therefore proposed ap-
plying the principle involved to clinical medicine as an
aid to determining the functional capacity of the heart
muscle.
His method consists essentially in making compres-
sion upon peripheral arteries (the femorals) so as to
shut off the circulation in the lower limbs, and observ-
ing the effects upon the pulse and blood pressure. The
author of the test found in cases of cardiac insufficiency
a lowering of the blood pressure and a simultaneous
increase in the pulse rate, both of which deviations
from the normal appeared to maintain a proportionate
relation to the incompetency of the heart muscle.
It has been proposed to substitute an Esmarch band-
age for digital compression of the arteries, thereby
doing away with the necessity of an assistant.
Technic of Kcitzenstein Test. — Sometimes called also
the Marey-Katzenstein-Shapiro Test.
The patient is put in a reclining posture and the
pulse rate and blood pressure taken. Pressure is then
made for two and one-half to five minutes over both
femoral arteries in the groins by means of the fingers
of an assistant or by clastic Esmarch bandage, or
according to Morelli by inflatable rubber stockings.
The pulse rate and blood pressure are again recorded.
In normal individuals with sufficiency of the myocar-
dium the pulse diminishes in number. The blood pres-
sure rises 5 to 15 mm. of Hg. With sufficient but
hypertrophic hearts the pulse diminishes or remains
the same. An increase of 15 to 40 mm. of Hg. takes
place in the blood pressure.
Tests of Heart Function 183
In cases of moderate latent cardiac insufficiency the
blood pressure remains unchanged. The pulse rate
is unchanged or increased. In higher grades of cardiac
insufficiency the blood pressure sinks and the pulse
rate increases.
For practical purposes it may be said that with
sufficiency of the heart muscle the pulse remains un-
changed or diminishes in number and the blood pres-
sure rises. If the pulse increases and the blood pres-
sure remains the same or falls after the Katzenstein
test the heart is insufficient.
Norris 13 made an investigation in 1907 with a view
of determining the adaptability of Katzenstein's test
to clinical use. He found that generally speaking the
results were accurate and confirmatory of its author's
findings but many exceptions were noted. Some of the
cases, many in fact, of cardiac weakness which re-
sponded positively to the test did so in an extremely
equivocal manner, leaving practically the final deter-
mination a matter of personal equation on the part of
the investigator.
As a corroborative test the method of Katzenstein
appears to possess some value, but as an independent
test of cardiac sufficiency or insufficiency no great de-
pendence can be placed upon it. The method should
be used with caution in cases of severe cardiac weak-
ness where it may prove to be actually dangerous.
5. Herz's Self -Che eking Test.14 Selbst-Hemmungs
Probe
Teclinic, — The patient is placed in a sitting posture
and remains so until the pulse rate has become con-
13 Blood Pressure and Clinical Applications, Phila., 1914, p. 145;
International Clinics, 1907, I, 17s, p. 66.
14 Deutsch. med. Wchnschr., 1905, 31, XXXI, 215.
184 Manual of Vital Function Testing Methods
stant. He is then directed to contract all the muscles
of hand and forearm with all his force and to flex and
extend the forearm with all possible force, performing
the motions slowly, paying strict attention to the per-
formance and endeavoring to antagonize his movements
as forcefully as possible.
In healthy persons the pulse rate is unaffected by
this maneuver, whereas in persons with a weak heart
the rate increases 5 to 20 beats per minute.
This test has been found to possess a certain degree
of reliability but it does not possess so absolute a value
as was originally ascribed to it by its author. Some-
times healthy persons give a positive reaction. Hirsch-
felder believes that the vagus plays a part in it and
that the results are not altogether indicative of cardiac
output and vigor.
6. Gymnastic Resistance Test. Herz-Harancliipy
Test
This consists in having the patient execute three
types of resisted movements. First a movement of
flexion-extension of the forearm, the patient being
seated. Second, a movement of separation-approxi-
mation of the thighs in sitting posture. Third, a move-
ment of abduction-adduction of the extended lower
limbs, the patient seated. All of these motions are re-
sisted equally. A slight rest is given between each
series. The whole test lasts 25 to 30 seconds. Prior
to the movements the systolic blood pressure is taken.
While the movements arc being executed and during
repose the blood pressure is retaken. In a normal in-
dividual there should be a variation in the blood pres-
Tests of Heart Function 185
sure as a result of the exercises of not more than 10
to 15 mm. of Hg. In cases of cardiac insufficiency it
reaches 20 to 30 mm. of Hg.
7. The Russian Test.15 "Holding the Breath" Test
This simple test for estimating the integrity of the
cardiac muscle has been in long use empirically. We
have called it the Russian test because Herz has men-
tioned the fact that he could not find any specific men-
tion of it in the literature but knew that it was com-
monly employed by certain Russian physicians.
The test is well called "Holding the Breath" test
since it consists simply in directing the patient to stop
breathing for as long a time as possible. This ma-
neuver puts a considerably added strain upon the
myocardium, particularly the right ventricle. Great
variations of the length of time in which the breath
can be held by different persons are found, but any
marked limitation of the time during which a person
can inhibit the act of respiration indicates cardiac
insufficiency. If the period of voluntary apnoea is
less than 15 seconds the myocardium may be considered
insufficient.
8. The Venous Pressure Test. Schott's 16 Test
The principle of the Schott test depends upon the
fact that in health if the arm is elevated to an angle
of 60 degrees with the patient in a recumbent posture
and making no other exertion the venous pressure in-
creases only .5 cm. of H2O or sometimes may remain
35 Die Herz Krankheiten, \Vien, 1912, p. 12,5.
16Deutsch. Arch. f. klin. Med., 1912, CVIII, 537.
186 Manual of Vital Function Testing Methods
stationary or even fall. If, however, the cardiac muscle
is insufficient, a rise in the venous pressure takes place
which may even be considerable (4 to 7 cm.). Accord-
ing to Schott, any reading above 3 cm. denotes cardiac
insufficiency.
Several methods have been devised to determine ven-
ous blood pressure. Von Frey and Gaertner considered
that the venous pressure can be determined by con-
sidering it equal to the height above the angle of
Ludwig at which the veins of the hand are seen to
collapse when the arm is raised. Von Recklinghausen
used an apparatus whereby the vein could be compressed
by inflating a small rubber capsule provided with a
glass window in the top of a rubber dam floor with an
opening in its center. The dam is coated with glycerine
to insure perfect apposition to the skin. It is placed
over a vein on the back of the hand or wrist and the
system inflated until the vein is seen to disappear, at
which point the pressure is read off on a water ma-
nometer. Eyster and Hooker modified the method by
using an aluminum chamber with a glass top, the two
ends concave to avoid pressure on the vein. The
normal venous pressure obtained by this instrument at
the sterno-xyphoid articulation is 5 to 10 cm. of H2O.
In cardiac cases it may rise to 27 cm. or more. The
pressure in the lip capillaries may be estimated by using
the point of blanching as the criterion. The study of
venous pressure is of some importance as an index of
accumulation of blood in the veins and may therefore
become to some extent an index of heart failure.
Tests of Heart Function 187
II. APPLICATION OF CARDIAC REFLEX ESTIMATIONS IN
DETERMINING HEART FUNCTION
MerTden's Test
The best known cardiac reflexes are those of Abrams
and Livierato.
Abrams' reflex consists of a diminution of the area
of precordial dullness following energetic friction over
the heart. Livierato's reflex consists of an increase of
the area of cardiac dullness following percussion over
the epigastric region. In Abrams' reflex the left ven-
tricle is chiefly affected and in Livierato's reflex the
right ventricle. The heart is so much less meiopragic
(weakened) in proportion to its capacity to give a
positive Abrams and a negative Livierato.
Technique of Reflex Test. — Map out the area of
precordial dullness carefully by light percussion and
mark with dermographic pencil. Make precordial fric-
tion for one minute, using rough cloth or a rubber
eraser. Follow this by rapid percussion of the pre-
cordial area. After three to five minutes wait, map out
the area of dullness by light percussion. If the reflex
is normal, the area will be smaller than before.
In using Livierato's reflex the technique is the same
to determine the area of cardiac dullness. The reflex
is elicited by making a series of rapid rather forceful
strokes for one minute over the median line of the ab-
domen. After three minutes' interim, the area of dull-
ness is again made out and if the reflex is positive the
right border of cardiac dullness will be found increased.
The two reflexes should not be applied to the same pa-
tient on the same day.
188 Manual of Vital Function Testing Methods
III. ESTIMATION OF SODIUM CHLORIDE ELIMINATION AS
A TEST OF CARDIAC SUFFICIENCY
Vaquez-Digne Test
This test is based upon the supposition that the
elimination of sodium chloride is affected by the suf-
ficiency or insufficiency of the heart muscle. In indi-
viduals in whom the integrity of the heart muscle is
normal any excess of salt ingested should be promptly
eliminated from the circulation and passed out thro
the kidney. In cases of cardiac insufficiency even when
latent, it is contended that the salt elimination is defec-
tive.
In applying this test the individual is put for some
days on a fixed sodium chloride ration and when an
equilibrium is established the amount of salt injected
is doubled and a quantitative estimation of sodium
chloride in the urine made. In cases of cardiac in-
sufficiency there will be defective elimination and if
the diminution of function is considerable there may
be oedema and signs of partial decompensation set up.
The integrity of the kidney function must be previously
determined.
IV. MODERN CLINICAL AND INSTRUMENTAL METHODS
OF INVESTIGATING CARDIOVASCULAR CONDITIONS:
THEIR APPLICABILITY TO ESTIMATING CARDIAC FUNC-
TION
1. Spliygmomanometer as an Index of Cardiac Func-
tion. Work-Velocity Ratio
The sphygmomanometer is the instrument in vogue
of our day. The chief value of this instrument is to
Tests of Heart Function 189
register the height of the blood pressure and since its
introduction it has no doubt contributed to a clearer
differentiation of states of hyper- and hypotension.
Like the sphygmograph, it is teaching physicians to
become more expert in the use of their sense of touch
and just . as the latter instrument (sphygmograph)
taught physicians to differentiate the arhythmias with-
out the use of the instrument in many cases, so, too,
it may come to pass that careful comparison of pal-
patory pulse estimations of pressure with instrumental
readings of pressure carried out day by day in cases
of hyper- and hypotension will finally educate the
physician to make correct deductions in many cases
without the use of the instrument. There will always
remain a certain proportion of cases, however, in
which, owing to various physical factors, an accur-
ate digital estimation of the systolic blood pressure
is impossible. A distinguished clinician who has cul-
tivated this perception to a remarkable degree is
quoted as saying: "I can estimate the blood pressure
with the fingers alone quite accurately in about eight
cases out of ten, but those in which it is of real im-
portance are always the other two."
Modern sphygmomanometry will, however, do some-
thing more than show the variations in the systolic
blood pressure. Recently, since the introduction of
the auscultatory or auditory method of using the
sphygmomanometer, the method of Korotkof, a more
accurate means of finding the exact diastolic pressure
has been found than could be had by means of the older
visual method with the vertical mercury or other
manometers.
With accurate data concerning the systolic and
diastolic blood pressure, we are in a better position to
interpret results in terms of cardiovascular function
190 Manual of Vital Function Testing Methods
than we are by means of the systolic pressure alone.
In order to fully appreciate just what may be ex-
pected from such data as the above in the interpretation
of cardiac function, we must bear in mind of course
a few simple physiological facts.
The blood pressure, that is, the systolic blood pres-
sure, depends mainly upon the contractile powers of
the heart muscle which enables it to pump the blood
into the arteries, against the peripheral resistance
caused by the friction of the blood on the vessel walls.
The peripheral resistance depends upon the tonicity
and physical state of the vessel walls. Under normal
circumstances the elasticity of the coats of the arteries
provides for a continuous instead of an intermittent
flow of blood which would be the case if the arteries
were rigid tubes.
The systolic or maximum pressure will approximately
show the actual pressure or work developed by the heart
at the moment of systole. The diastolic or minimum
pressure will show the degree of the peripheral re-
sistance which the heart has been able to overcome
and which is maintained in the peripheral circulation
during the time of heart refilling. The difference be-
tween the highest and the lowest pressures in the larger
arteries, that is the difference between systolic and
diastolic pressures, is known as the pulse pressure. The
pulse pressure, therefore, is the measure of the amount
of force exerted by the heart in maintaining the blood
pressure over and above the arterial or peripheral re-
sistance. To this extent then the pulse pressure is a
measure of the pumping capacity of the heart and
hence is of some importance in estimating the state of
cardiac function.
Gibson has called attention to the fact that there
are certain normal arithmetical relations which are
Tests of Heart Function 191
discoverable in a study of the three factors — systolic
pressure, diastolic pressure and pulse pressure. The
relation of the diastolic pressure to the systolic pres-
sure is normally as two is to three. The relation of the
pulse pressure to the systolic pressure is as one is
to three. If the systolic pressure is 150 the normal
diastolic pressure will be approximately 100. With
a systolic pressure of 150, the normal pulse pressure
will be 50. Of course these figures represent approxi-
mate and not absolute relations. They are of some
service in estimating cardiac function because patho-
logical relations become evident and the presence and
also to a certain extent, the degree, of cardiac overload
may be appreciated.
The normal arteries will apparently withstand a con-
tinual variation in pressure, that is a pulse pressure of
35 to 50 mm. of mercury without deterioration. Any
great increase of the pulse pressure over these figures
is pathological and at least indicates cardiac overload
and consequently justifies suspicion that perhaps the
myocardium may not long succeed in maintaining it.
The heart manages to do so by undergoing hypertrophy
and when this has been accomplished the organ may be
regarded as at least a locus mlnoris reslstentice.
A quantitative idea of the undue stress may be ob-
tained by taking the difference between the normal
and pathological pulse pressure. When this is multi-
plied by the pulse rate and this by 60 (hour) and again
by 24 (day) a concrete idea may be gained of the
enormous excess of energy required to be expended by
the heart in a day to overcome pathological peripheral
resistance.
Such a case, however, may go on and on and we have
no exact method of predicting just when the break will
come, with its attendant consequences of cardiac in-
192 Manual of Vital Function Testing Methods
sufficiency. As it approaches, however, the diastolic
pressure will fall, indicating the approaching collapse
of the cardiovascular mechanism. The larger the
pulse pressure in relation to the diastolic pressure, the
greater the strain on the heart will be and the more
imminent, therefore, is decompensation. A high systolic
pressure with a relatively low diastolic pressure indi-
cates, therefore, impending collapse of the heart muscle.
If after treatment the diastolic pressure rises and the
pulse pressure falls the indication is that recompensa-
tion is taking place.
Technique of Sphygmomanometry. — A brief account
only of the method of obtaining the systolic and dias-
tolic blood pressure will be given. A more complete
description is properly found in texts devoted to the
subject of blood pressure. The arm band is applied
to the bared arm above the elbow by placing the broad
end containing the rubber bag over the region of the
brachial artery. Wrap the band bandagewise around
the arm and tuck in the narrow end. Connect up the
indicator and pump. Apply the sphygmometroscope
or ausculoscope or stethoscope over the brachial artery
at the bend of the elbow. Increase the pressure until
all sounds are gone, then gradually admit air. The
sounds which are heard are divided into four phases.
First, a loud clear-cut snapping tone. This is caused
by the first and the early pulse waves that break
through the constriction and its beginning repre-
sents the systolic pressure. Owing to the greater
sensitiveness of the ear than the fingertip it is usually
heard some 5 to 10 mm. above the point where the
radial pulse is first felt. In normal cases, it is said
to last during the fall of about 14 mm. of pressure.
Second, a murmur or succession of murmurs lasting
during the fall of about 20 mm. This phase is not
Tests of Heart Function 193
always present and if absent the first and third phases
merge into one another. Third, a clear tone resembling
the first, sometimes less well marked but often louder.
This lasts during a fall of about 5 mm. Fourth, a
rather sharp transition from the loud to the dull tone
recently proved to be the time of diastolic pressure.
As there had been considerable discussion as to
whether the diastolic pressure corresponded to the be-
ginning or ending of the fourth phase, Warfield under-
took its investigation some years ago through a series
of animal investigations and comparison of cases with
accurate graphic records. These have been confirmed
by others and the point now seems to be settled, that
diastolic pressure coincides with the beginning of the
fourth phase.
Functional Tests Based on Direct Blood Pressure
Determinations.
I. The Cardiac Efficiency Factor of Tiegerstedt. —
The pulse pressure shows approximately the systolic
output in energy and the velocity of the blood stream
will be the product of this energy into the number of
cardiac cycles (pulse beats) per minute. In other
words, pulse pressure (PP) times pulse rate (PR)
will equal velocity of flow.
Since the interventricular pressure is almost con-
stant throughout systole, it is evident that the work
done by the heart is tolerably constant throughout the
period. The work done by the heart in a unit of time
will be the product of its maximum energy, systolic
pressure (SP), multiplied by the pulse rate (PR), mul-
tiplied by the duration of systole. Since the inter-
ventricular pressure is constant, the factor duration of
systole may be eliminated and work done equals product
194 Manual of Vital Function Testing Methods
of SP X PR (systolic pressure times pulse rate).
The reason why interventricular pressure is constant
is because of the fact that the heart liberates all avail-
able energy at each contraction. A concrete example
will readily show how the velocity work ratio is ob-
tained. If the SP = = 130, DP == 85, then PP == 45.
If PR = 70 then PP 45 X PR 70 = 3,100 (velocity),
and SP 130 X PR 70 == 9,100 (work), the ratio then
becomes
PP
^jjj = Blood pressure coefficient (Tiegerstedt).
fe-i
PP X PR _ Velocity
SP X PR ~ Work
This is the velocity work ratio or coefficient of heart
pumping efficiency. The velocity work relation in this
example is one to three and this is about the normal
ratio. Expressed in percentages, it varies normally
from 25 to 35 f/c . Increase in this ratio may indicate
cardiac insufficiency.
II. The Cardiac Strength, Cardiac Weakness Ratio
of Goodman and Howell. — These authors have studied
the duration of the four-tone phases of auscultatory
blood pressure estimation in a series of normal and
pathological cases. Their test is based on the ratio
of these phases to the pulse pressure and to one an-
other. They set forth their views as follows :
1. "The first phase or tone phase serves principally
as an index as to how far the pressure must fall before
the blood current can be sustained past the obstruction
in the vessel caused by the cuff at a sufficient velocity
and for a sufficient duration to produce the murmur.
Hence the information it affords is of negative rather
than of positive value. In other words, its normal
Tests of Heart Function 195
duration is of no value but an increase or decrease in
length is of importance.
2. The second or the murmur phase seems to be espe-
cially dependent upon cardiac effectiveness, for it is
in this phase alone that the individual sounds possess
a distinct element of duration and this protracted
energy, for so it must be regarded, must evidently come
from the heart.
3. The third phase or second tone phase depends
not alone on cardiac efficiency but also on the character
of the vessel wall. The more sclerotic the vessel and the
greater the cardiac hypertrophy, the more favorable
are the conditions for the production of a clear tone.
4. As the fourth phase or dull tone may be pro-
duced by a resilient vessel, receiving a normal pulse
shock, or by a rigid vessel receiving a weakened shock,
its interpretation is more difficult. If our assumptions
are correct it is evident that increases in the second
and third phases are dependent on cardiac strength
and circulatory deficiency, while the first and fourth
phases suffer increase when there is cardiac weakness.
Furthermore, in dealing with increases or decreases in
any particular phase it is important to know at the
expense of what adjacent phase this has occurred. It is
apparent that an increase in the third phase for example
at the expense of the second has not the same signifi-
cance as an increase of this phase at the expense of
the fourth. In the first instance the unit of cardiac
strength which we obtain by adding the lengths of the
second and third phases has not been materially changed
while in the latter it has been increased. For this rea-
son we recommend that the sum of the second and
third phases be compared with the sum of the first and
fourth phases in order to determine whether the ele-
ments of force or those of weakness are predominating.
196 Manual of Vital Function Testing Methods
Aside from the value of the persistence of the fourth
phase in aortic insufficiency little of diagnostic value has
developed in regard to the length of any individual
phase. Advantage has been derived, however, from
studying the changes in the sequence reading, specially
in decompensating cardiac lesions as the patient im-
proves or not. In these cases changes in the percent-
ages of the various phases are not the only significant
feature but internal peculiarities appear. Or to put
it another way, sequence readings have a functional
rather than an organic significance. Our results uni-
formly show that with decompensation or circulatory
disturbances of lesser degree, the element of heart weak-
ness (the sum of the first and fourth phases) progres-
sively encroaches upon that of heart strength (the sum
of the second and third phases). The second phase
appears to be the one which is with most difficulty
sustained. The fourth phase as weakness gains the
ascendency, is usually the first to lengthen the element
of cardiac weakness by its encroachment on the third
phase, but encroachment of the first phase on the sec-
ond soon adds its share to the total." 17
The average duration in mm., the fall of and per-
centages of the pulse pressure of the different phases
in normal individuals are:
mm. per cent.
First phase 14 31.1
Second phase 20 44.4
Third phase 5 11.1
Fourth phase 6 13.3
The cardiac strength (second and third phases) : car-
diac weakness (first and fourth phases) :: 55.5:44.4.
"Amer. Jour, Med. Sci., 1911, CXLII, 336.
Tests of Heart Function 197
Marked increase of the cardiac weakness factors indi-
cates cardiac inefficiency.
///. Previously to Goodman and Howell's work many
observers 18 have recognized that the duration of the
second phase of the auscultatory tones indicated cardiac
strength. A considerable reduction of its normal per-
centage of the pulse pressure is therefore taken as a
test of cardiac insufficiency.
IV. The Cardiac Overload Factor of Stone. — A
paper on the clinical significance of high and low pulse
pressure with special reference to cardiac load and
overload with a report of 170 cases was presented by
Stone at the meeting of the American Med. Assn. in
1912.19
The ratio of the pulse pressure to the diastolic pres-
sure representing the load of the heart has since been
used as a test for cardiac efficiency. Stone says, "the
pulse pressure measures the energy of the heart in
systole in excess of the diastolic pressure. For clinical
purposes it represents the load of the heart. The
myocardiac load may therefore be expressed by the frac-
pulse pressure
tion —. — — - • and under normal conditions is
diastolic pressure
approximately 50%." Anything in excess of this is an
overload. The average load in these cases was 71, an
overload of 21%. Naturally in this group of cases
some did well and some badly. To quote again, "judg-
ing from this small series of cases it would appear that
when the overload factor exceeds 50% the patient may
be in danger of myocardial exhaustion at any time of
slight overstrain. As a rule the greater the overload
the greater the danger." That is to say, an overload
18Forman, Ztschr. f. diet, und physik. Therap., XIII, 809;
Fisher: Deutsch. med. Wchnschr., 1908, XXXIV, 1141.
19 Jour. Amer. Assn., 1913, LXI, 1256.
198 Manual of Vital Function Testing Methods
of 50% or more (the pulse pressure equal to or greater
than the diastolic pressure) indicates cardiac insuf-
ficiency of a considerable degree with impending de-
compensation. Whereas an overload of 25% would
seem to indicate a mild degree of insufficiency.
Swan 20 has recently published a study of the above
four tests with a series of observations on 40 patho-
logical cases. His conclusions are as follows : "It
appears to me legitimate from the study of the cases
reported to conclude that all four of these factors have
some value in determinkig the efficiency of the myocar-
dium. I am inclined to think' at present that the cardiac
efficiency factor of Tiegerstedt and the percentage of
the pulse pressure formed by the second phase are the
most important. A cardiac efficiency factor of 40%
or over would seem to point out distinct myocardial
inefficiency. A second phase of 30% or under would
seem to indicate the same condition. The CS to CW
(cardiac strength to cardiac weakness) ratio is less
important I think because it so often cannot be deter-
mined and again because a small second phase is very
frequently made up by a large third phase. On the
other hand, CS : CW ratio in which the CW factor is
greater than the CS factor is indicative of disturbance
of the myocardium, functional if not organic. I am in-
clined to think at present that the overload factor of
Stone is indicative more of peripheral resistance than of
myocardial weakness. A cardiac load below 50% as
determined by this method giving a negative overload
may have some significance, but it will require further
study to determine its nature."
20 Archives Int. Med., 1915, XV, 269.
Tests of Heart Function 199
#. Rontgenoscopy and Rontgenography as Indices
of Cardiac Function
The form, position and movements of the heart as a
whole and its different chambers, also the great vessels
at its base, can be very successfully examined by the
X-ray. For several reasons we need only touch upon
this interesting and remarkable method of cardiac ex-
amination. In the first place, the application of the
X-ray to the study of the heart concerns more par-
ticularly the examination of the organ from a diag-
nostic and anatomical point of view. From this stand-
point it constitutes a valuable addition to the older
methods of heart exploration. It cannot be said, how-
ever, that a Rontgen ray examination sheds much
light upon the question of cardiac function. Its chief
use is to denote changes in the shape of the organ,
hypertrophy and dilatation of its cavities, aneurysm,
pericardial effusions, etc. By orthodiagraphy, the posi-
tion and topography of the heart can be accurately
delineated. But neither ordinary Rontgenoscopy nor
orthodiagraphy of the heart shed much light upon the
problem of estimating the exact efficiency of the cardiac
function.
3. Sphygmocardiography and Electrocardiography;
Their Relation to Cardiac Functional Capacity
In many text books the phrase, functional disease of
the heart, is often used synonymously for pulse irregu-
larity. The phrase, cardiac function, is used in an
entirely different sense here. Cardiac function, so far
as the present discussion is concerned, relates to the
capacity of the heart to perform its work, with the
200 Manual of Vital Function Testing Methods
adequate maintenance of its reserve. An irregularity
of the cardiac rhythm does not necessarily mean any
serious deterioration of function. For example, an
individual may have a sinus arhythmia or an occasional
premature contraction and possess an absolutely
normal cardiac reserve.
On the other hand, the discovery of certain other
types of irregular rhythm always indicates a serious
disturbance of heart function. The presence of true
heart block, for example, denotes a lesion of the con-
ducting system and hence a deterioration of function.
The same may be said of auricular fibrillation and to
an even greater extent of pulsus alternans.
But the detection and identification of irregularities
in the cardiac rhythm is a part of the general semi-
ological investigation of that organ and while of great
importance to the clinician who is examining a case
for heart disease, does not properly come within the
scope of an investigation into the methods of estimating
cardiac function.
The study of cardiosphygmography and electro-
cardiography has undergone a tremendous develop-
ment in recent years. The names of Marey, Franck,
Gaskell, Engelmann, Wenkebach, Herring, MacKenzie,
Lewis, Erlanger, His and many others are prominently
identified with the former and those of Waller, Ein-
thoven, Kraus, Nicolai, Edclmann and others with the
latter.
The recognition of nearly all the varieties of arhyth-
mia may be determined by the skilled clinician without
the use of any technical apparatus. Unfortunately,
this is not always the case and there are types of irregu-
larity of the heart rhythm which can only be posi-
tively recognized by the use of some form of instru-
mental registration. Pulsus alternans is the best ex-
Tests of Heart Function 201
ample of this fact. This variety of arhythmia cannot
be recognized with certainty without pulse tracings.
Its importance from a prognostic standpoint, as Lewis
points out, is extremely great. This fact alone will al-
ways make the polygraphic study of the pulse a matter
of necessity in all cases in which there is a suspicion
that pulsus alternans may be present.
The ordinary methods of examining the heart, em-
ployed in clinical diagnosis, are exceedingly well adapted
to disclose diseases of the organ. By inspection, palpa-
tion, percussion and auscultation, properly performed,
not only can it be determined that disease of the heart
is present, but the precise location and often the nature
of the lesion can be made out. As Cabot has well
expressed it, "we are very well satisfied with the ordi-
nary methods of examination, when we find something
such as valvular disease, obstructions, accumulations
and degenerations. But in many cases in which we
fear that the heart is diseased and its functional power
diminished, the ordinary methods of investigation do
not show anything. Even the more technical and re-
fined instrumental methods are negative only too often
in such cases. The heart has passed a good physical
examination and yet may be insufficient. We desire to
know what the heart can do, what is the condition of
its reserve. The necessity of supplemental methods
becomes manifest. This is the proper field for experi-
mental inquiry into the heart function. It is here
that the functional tests become especially useful. We
give the heart some work to do and see how it reacts,
how fast it tires, how slowly it recuperates. We sub-
ject the patient to extra effort and note the general
symptoms produced, particularly dyspnoea. We pro-
ceed to a careful analysis of the history of our patient
with respect to his subjective reaction to all of his
202 Manual of Vital Function Testing Methods
environment. Already we are working with problems
of cardiac function in a fundamental manner."
General Conclusions as to Tests for Cardiac Function
Hirschfelder, speaking upon the importance of func-
tional tests or studies in borderland cases between func-
tional sufficiency and cardiac failure, emphasizes the
importance of careful observation and says : "It must
be admitted that in order to be decisive, all tests have
to be pushed to a point at which the appearance, sensa-
tions and signs of the patient are in themselves per-
fectly characteristic of cardiac insufficiency and at
which, for diagnostic purposes, a little common sense
observation is at least as unambiguous as observation
with elaborate apparatus. This does not mean that
exercise tests are unimportant. On the contrary, they
are of the greatest value and no change in the patient's
mode of living during convalescence or during after
life should be undertaken without them.
"But their importance depends more upon the care
with which the physician watches the general appear-
ance and condition of the patient, the rapidity with
which he recovers from the exercise, his general condi-
tion and whether nervousness, irritability, cough or in-
somnia have set in during the 24 hours following it,
than in the numerical changes which occur at the mo-
ment of exercise. The symptoms to be looked for as
evidence of overwork are well known. These are subtler
manifestations resulting from smaller changes than may
be detected by even the most refined observation by
mechanical methods and which are less easily masked
by ambiguities.
"Moreover, it must be realized that any one form
of exercise furnishes data which may depend as much
Tests of Heart Function 203
upon the condition of the skeletal muscles as upon the
heart. The blacksmith with a diseased heart may be
able to do more work than the bookkeeper with neuras-
thenia and yet under the conditions in which he lives
even if not under the strength test arranged for the
average man, the blacksmith's heart may be failing.
In diagnosis, prognosis and therapy the testing of func-
tional insufficiency is a matter of sociology as well as
physiology. The important question is not what the
person can do in a gymnasium, but what he can do
and what he cannot do in everyday life. Each man
must be fit for his own mode of life or must be made
to change it. His cardiac power must be studied with
reference to that mode of life rather than with refer-
ence to a rigid scheme." 21
21 Diseases of Heart and Aorta, Phila., 1913, 199.
CHAPTER V
THE DUCTLESS GLANDS
GENERAL CONSIDERATIONS
THE clinical examination of function of the endoc-
rinous glands is a subject capable of great future
growth. Only the first steps have so far been taken in
developing this mine of hidden riches. To the physi-
ologist, pathologist and clinician the subject offers a
fertile and tempting field of investigation.
In the following account of the functional diagnosis
of the endocrinopathies but little can be given of the
enormous mass of experimental material, and, as
Barker 1 has said, "the greater mass of theories" pro-
pounded, concerning the physiology, semiology, pathol-
ogy and interrelations of the glands of internal secre-
tion. These subjects with complete bibliographic and
historic references can be found in the classical works
of Biedl,2 Vincent,3 Falta4 Levi-Rothschild,5 Paton,6
Sajous 7 and others.
In these great works little or nothing can be found
concerning the important question of functional diag-
nosis. The material that has been evolved concerning
Southern Med. Jour., 1914, VII, 1.
2 Internal Secretary Organs, London, 1913, Bale Danielsson
(Trans, from German).
8 Ductless Glands, London, 1912, Arnold.
4 Erkrankungen der Blutdriisen Wien, 1913.
5 Endocrinologie, Paris, 1911, Dion.
"Internal Secretions, Phila., 1911, Davis.
7 Regulators of Metabolism, London, 1913, Macmillan.
204
The Ductless Glands 205
the investigation of function of the ductless glands or
glands of internal secretion is not only scant but scat-
tered widely in the literature from whence so far as we
know it has never been gathered. Not a single article
devoted to the general question of the functional diag-
nosis of the endocrine glands exists in the whole medical
literature. In isolated instances where the subject of
functional diagnosis is mentioned, upon investigation
it is found that the question is treated upon an almost
purely semiological basis. The semiological method of
diagnosis of diseases of the ductless glands has there-
fore reached a higher degree of development than the
functional method, among clinicians up to the present
time. Notwithstanding all this, it is admittedly true
that the clinician is much in need of functional tests, to
enable him to discover the various endocrinopathies in
their latent stages, or, as the French say, in the forme
fruste, when the symptomatic picture may be incomplete
or confusing. Therefore, while at the present time it
cannot be said that satisfactory chemical or biological
functional tests have been discovered, capable of dis-
closing with certainty the existence of latent disease of
the thyroid, parathyroid, thymus, pituitary or adrenal
organs, nevertheless some advance has been made in this
direction and the great need for such tests, in this im-
portant tho subtle field of clinical medicine, will always
constitute a sufficient inspiration for further discovery.
As will be developed, the principal tests which have
been devised up to the present time with the object of
testing endocrinous function, relate to the thyroid
gland and particularly with that aspect of thyroidop-
athy which is connected with an increased activity of
the gland. Function testing of the adrenals has received
some attention and development. The other ductless
glands remain so far a terra incognita to the functional
206 Manual of Vital Function Testing Methods
method. When we stop to consider how much remains
to be known concerning the functions and interrelations
of the glands of internal secretions, the fact will not
be surprising that the subject of functional diagnosis
of their diseases has not received a greater develop-
ment.
Regarding the ductless glands as a whole, we cannot
fail to be impressed with their intimate relation to the
processes of body metabolism. The organs of internal
secretion or ductless glands (blutdriisen) are important
regulators of metabolic processes. It is agreed that the
pancreas normally inhibits carbohydrate metabolism
and that on the other hand the thyroid and suprarenals
normally increase carbohydrate metabolism. The thy-
roid has an important effect upon proteid metabolism
not shared by the other glands. Increased function of
the thyroid is accompanied by increased proteid metab-
olism while hypofunction of the thyroid produces a dim-
inution of proteid exchanges. The parathyroids and
thymus are intimately concerned with normal calcium
metabolism but their exact relation to this process is
unknown. The gas exchanges of the organisms are also
fundamentally controlled by the ductless glands. In
hypcrthyrcosis there is an increase, and in myxedema a
decrease, of the oxygen absorption and C02 exchange.
These facts have formed the basis of certain experi-
mental methods of determining the functional capacity
of the ductless glands and were it not for the fact that
the performance of metabolic experiments is so com-
plex and requires so extensive an instrumental equip-
ment, the examination of these processes as aids to the
functional diagnosis of the endocrinopathies would have
a much wider application.
The Ductless Glands 207
THE THYROID GLAND
Tests of Functional Activity
Without entering debatable ground, it may be con-
fidently asserted that there are certain facts regarding
the physiology and pathology of the thyroid gland
which are universally admitted. It is clear, for in-
stance, that the thyroid is of great importance in the
economy of the human organism, and that certain
lesions of this gland give rise to symptoms which when
they are outspoken may be definitely correlated with
thyroid disease.
However, despite all the work, experimental and
clinical, which has been done upon this gland, there is
even now no absolute unanimity of opinion of the pre-
cise function or functions of the thyroid. The thyroid
function is probably not simple but complex. As in
the case of the other ductless glands, many physicians
feel convinced that there is an antitoxic function of the
thyroid, which consists in collecting exogenous iodine
and in some mysterious and unknown way neutralizing
certain hypothetical products of intermediary metab-
olism. Naturally this idea is a pure speculation and
has been arrived at by indirect reasoning. The most
usually accepted theory of thyroid function is, how-
ever, that the thyroid manufactures an internal secre-
tion, possibly an iodine proteid, which is essential to
the proper growth and normal metabolism of the entire
body. Baumann 8 made an epochal discovery in 1895
when he showed that the thyroid gland contains iodine.
Although speculation and investigation have since been
rife in respect to this discovery, its precise meaning
is yet unknown.
8Zeitsch. f. physiol. Chem., 1895, XXI, 319.
208 Manual of Vital Function Testing Methods
The vast amount of work which has been done upon
experimental extirpation of the thyroid can only be
mentioned. If the thyroids are removed from young
animals, growth is retarded. Total extirpation of the
thyroids in human beings is well known to be followed
by serious symptoms, the so-called cachexia strumipriva.
If the individual is young there will be retarded growth,
faulty ossification, mental and metabolic enfeeblement.
Similar symptoms are seen in children with congenital
thyroid aplasia. In human beings who are deprived of
their thyroids, a phenomenon appears which is not seen
in lower animals, namely myxedema.
Spontaneous myxedema also occurs in the adult
human being as a result of thyroid insufficiency — the
so-called Gull's 9 disease. Murray 10 discovered that
the administration of thyroid extract will eliminate the
symptoms of myxedema.
If thyroid substance be fed to normal animals, symp-
toms will develop which are similar to those that occur
in Graves' or Bascdow's disease in the human subject
and are supposed to be due to a hyperthyreosis or
hyperthyroidization — in other words, an increase of
function of the thyroid.
This brings us to the important induction that in
the human subject we may have two different or rather
two opposite pathological states to consider in respect
to the thyroid gland and its functions, namely A. a
hyperthyreosis and B. a hypothyrcosis. In the well
developed state these two symptom groups constitute
definite and tangible clinical syndromes.
A. Hypcrf unction of the Thyroid Gland. — To the
syndrome of hyperthyreosis, the name of Graves' or
Basedow's disease is attached. The symptoms of hyper-
9 Trans. Clin. Soc., London, 18T4, VII, 180.
10 Brit. Med. Jour., 1891, 7.96.
The Ductless Glands 209
thyreosis may be placed in the following categories : 1,
enlargement of the gland; 2, signs of heightened ex-
citability of the vegetative or sympathetic nervous
system ; 3, signs of secondary or concomitant dis-
turbances in other cndocrinous glands ; 4, symptoms
of profound disturbance, usually of excess metabolism ;
5, a variety of disorders of the central nervous system ;
6, a peculiar picture in the blood (leucopenia with
lymphocytosis).
Naturally in this review of functional diagnosis we
cannot go into a detailed account of the semiological
data which might be collected under each of the above
headings. It is interesting to observe that the symp-
toms of Graves' disease which come under the category
of the vegetative nervous system involve both the auto-
nomic and sympathetic portions of that system. The
autonomic and sympathetic innervations are both in-
volved to a certain extent in every case, but in one, the
former, and in another case, the latter, system will be
predominantly affected. The eye, heart, blood vessels,
skin, digestive, respiratory and urogenital apparatus
are all supplied with innervations of both kinds, usually
reciprocally or antagonistically, and consequently in
Graves' disease there are symptoms referable to disturb-
ances in sympathetic or autonomic innervations in sev-
eral or in all these different organs. We shall not enu-
merate all the various symptoms of Graves' disease,
since such an enumeration will be readily found in books
or articles dealing with the semiology of this condition.
Whenever a sufficient number of these symptoms can be
found in a given case the diagnosis of Basedow's disease
can be readily made and a quantitative idea of the se-
verity of the case may be gained by the actual number
or the seriousness of the symptoms. In other words,
when the classical semiology of Graves' disease is pres-
£10 Manual of Vital Function Testing Methods
ent, naturally no functional tests will be needed.
A striking phenomenon of Graves' disease is the ac-
celeration of all the metabolic processes. This accelera-
tion includes the total combustion in calories, the pro-
tein, carbohydrate, fat and mineral metabolism. In
states of hypothyroidism the opposite condition of re-
tardation of metabolic processes occurs.
Functional tests have therefore been proposed as a
criterion of the existence of states of hyper- or hy-
pothyreosis on the basis of increased or diminished oxy-
gen consumption and protein, carbohydrate, fat and
mineral metabolism.
As simpler methods of determining the activity of
these various processes are developed we may hope that
this kind of investigation will gradually enter more and
more into the diagnostic armamentarium of the clinician
who is interested in measuring the functional integrity
of the thyroid. But as was said before, the technical
difficulties which have so far usually surrounded the
methods of determining and measuring the various
processes of metabolism have prevented their general
introduction into clinical medicine.
The symptoms of Graves' disease which are refer-
able to concomitant or reciprocal disturbance of func-
tion of the other endocrinous glands have contributed
somewhat to the diagnosis of hyperthyreosis from a
semiological standpoint. This circumstance we shall
not attempt to develop. But from the standpoint of
functional diagnosis of the thyreopathies, these disturb-
ances acquire a considerable importance since they
open the way, though indirectly, to the development of
means for testing the functional activity of the thyroid.
It is quite generally held that the thyroid and the pan-
creas mutually inhibit one another's activity. The
pancreas and chromaffin system (adrenals) are likewise
The Ductless Glands 211
mutually inhibitory. The thyroid and the adrenals ap-
pear, however, to reciprocally favor each other's activ-
ity, that is, an under function of one leads to an under
function of the other, while an over function of the one
will lead to an over function of the other. This is the
teaching of the present Vienna school of endocrinolo-
gists represented particularly by Eppinger, Hess,
Falta, Rudinger, and others. According to the teaching
of this school, a hyperthyroid function will be accom-
panied by an insufficiency of pancreatic function (inter-
nal secretion) and by an increased activity of the
chromaffin or adrenal system. Hypothyroidism, on the
contrary, will be followed by over function of the inter-
nal secretion of the pancreas and diminution of adrenal
functional activity.
On the basis of these hypotheses, certain tests have
been devised to disclose a hyperfunction of the thyroid
gland. One of these, the so-called adrenalin-mydriasis
test of Loewi, is used to disclose on the one hand an
insufficiency of the internal secretion of the pancreas
and on the other a hyperthyreosis. So far as its ap-
plication to the investigation of pancreatic insufficiency
is concerned, the test has already been described (v. s.)
With reference to the second application, namely that
of disclosing a hyperthyreosis, details will be later
given. Tests founded upon the existence of a glyco-
suria, either spontaneous or following the injection of
adrenalin, will be considered under the heading Adrenal
Glands.
The diagnosis of hyperthyroidism, or Graves' disease,
is easy in typical cases. The enlarged thyroid, the
tachycardia, the disturbances of the sympathetic
nervous system, the tremors, the mental state, the ac-
celerated metabolism and the blood findings make a
definite and indubitable diagnostic picture.
Manual of Vital Ftvnction Testing Methods
There are, however, many cases which elude the clini-
cian because they are atypical. To these latent or
atypical cases the French have given the expressive
title of formes frustes. Barker 1X has warned us very
properly that any one of eight different symptoms
should make the clinician suspicious of Graves' disease.
The symptoms are: (1) persistent tachycardia (pulse
above 85); (2) rapid emaciation without apparent
cause; (3) excessive sweating; (4) persistent watery
diarrhoea; (5) neurasthenic and psychasthenic states;
(6) outspoken lymphocytosis ; (7) fine tremors of the
fingers; (8) one or more of the usual ocular symptoms
of the disease, namely protrusio bulbi or positive Dal-
rymple,12 von Graefe,13 Moebius',14 Stellwag,15 Jelli-
nek 16 and Rosenbach 17 signs.
Suppose, however, one of these suspicious signs be
present. How shall it be determined whether or not
there is actually present a condition of hyperthyroid-
ism? It is here that a satisfactory functional test would
be invaluable. Frederich Miiller first called attention
to its necessity under such circumstances. Several
functional tests are at present available for this pur-
pose. Unfortunately, they have not been found entirely
adequate. Nevertheless, they are of a sufficient degree
of assistance to warrant their retention by the clinician,
especially as they form an important nucleus upon
which future investigators may build new theories and
points of departure for renewed attempts at explora-
tion. Some of them have not been sufficiently developed
as yet to allow a final opinion to be formed.
The tests of hypcrfunctional activity of the thyroid
gland are as follows :
"Widened eye slit; "lagging upper lid; "insufficient converg-
enee; "infrequent incomplete winking; w pigmentcd eyelids;
17 tremor of closed lids.
The Ductless Glands 213
1. Hypophysis Test of Claude, Baudouin, and Porak.
2. The Adrenalin Mydriasis Test of Loewi.
3. Induction of Experimental or Artificial Hyperthy-
roidism as a Functional Test.
4. The Aceto-Nitril Test of Reid Hunt.
5. Metabolic Studies in the Functional Diagnosis of
Hyperthyroidism.
6. The Complement Fixation Reaction as Functional
Test of Hyperthyroidism.
7. The Specific Ferment Reaction of Abderhalden, as
Functional Test of Hyperthyroidism.
1. THE HYPOPHYSIS-EXTRACT TEST FOR HYPERTHYREO-
SIS. CLAUDE, BAUDOUIN, PORAK TEST 18
Recently Claude, Baudouin and Porak have published
some interesting researches upon the use of extract of
the posterior lobe of the hypophysis, in disclosing the
presence of latent hyperthyroidism.
In their experiments they made use of a hypophyseal
extract of posterior lobe of such strength that 1 c.c. of
the substance to be injected was equivalent to 1/2 of a
posterior lobe of a beef's hypophysis. This they say
corresponds to .05 of hypophysis powder.
This extract was obtained by the action of alcohol
at 70° upon the hypophysis powder, dried and freed
from fat. The alcohol is evaporated and the residue re-
dissolved in normal salt solution. They also employed
occasionally a watery extract, obtaining results with
the latter which were practically similar to those ob-
tained from the former.
They found that subcutaneous injections of both
watery and alcoholic extracts of hypophysis produced
a marked reaction, pallor, glycosuria and diarrhoea, the
18 Bull, et Mem. Soc. Med. d. Hop. de Par., 1914, XXX, No.
22, 1904.
214 Manual of Vital Function Testvng Methods
greater effects being produced by the watery solution
and accompanied by greater pain. The alcoholic ex-
tract was found to be less painful and the pain less
lasting.
When a quantity of alcoholic extract corresponding
to one whole lobe (beef) was injected, the authors noted
complex effects, such as an action upon smooth muscle
fiber, a cardiovascular effect and an action on nutrition.
The diuretic effect was doubtful.
The action on nutrition was characterized by gly-
cosuria, the cardiovascular effect consisted of accelera-
tion of the heart. In these complex effects of the
hypophysis there appears to be an excitation of the
sympathetic system shown by the cutaneous vasocon-
striction and the glycosuria. The accelerator fibers of
the heart being likewise of sympathetic origin, one would
naturally expect to find acceleration of the beat. One
would also expect that with concomitant constriction
of the peripheral vessels that the blood pressure would
rise. This phenomenon did not appear. The blood
pressure remained the same or was lowered. The au-
thors attribute this to a depressing effect of hypophysis
extract on the myocardium. They noted occasionally a
galop rhythm following the injections. With normal
individuals the authors noted acceleration of the pulse.
They then proceeded to experiment upon cases of
Graves' disease. The patients were kept under observa-
tion free from emotional excitement until the normal
pulse rate was accurately determined. Injections of
plain salt solution were tried as controls.
Thirteen typical Basedowians were used in their ex-
periments. With the exception of the cardiovascular
effects the results of injections did not differ from
those of normal cases. The symptoms, pallor,
contraction of smooth muscle fiber of intestine
The Ductless Glands 215
and uterus, and glycosuria, the latter fairly well
marked, were noted in the case of Graves' disease.
Alimentary glycosuria was more readily provoked after
the injections than before, showing a diminution of the
already lowered carbohydrate tolerance. The cardio-
vascular effects of the injections were highly significant.
In normal subjects the pulse becomes accelerated. The
acceleration commences two or three minutes after the
injection. It reaches a maximum in 10 or 15 minutes.
Then the frequency rapidly diminishes and in about 20
minutes the pulse is normal.
In the cases of Graves' disease, however, the results
were found to be diametrically opposite. The pulse
which is accelerated before the injection of hypophysis
extract, becomes quickly slowed. In one of these cases
the pulse dropped 42 beats. Usually the diminution
in number of beats is much less, averaging 8 or 10. The
maximum lowering is reached in about 2 minutes, some-
times 4 or 6 and rarely even 10. The bradycardia is
usually ephemeral. Usually in 7 or 8 minutes the
pulse becomes fast again. Sometimes it was found to
return to the original number previous to the injection.
In the majority of instances, however, it remains not-
ably beneath this point.
It would appear as a result of these interesting ex-
periments that extracts of the hypophysis, which nor-
mally produce tachycardia, bring about an opposite
effect, namely, bradycardia in case of Graves' disease.
The authors believe that the extract of hypophysis
contains principles which simultaneously excite the
terminations of both sympathetic and vagus fibers.
This is the only explanation of the complex effects of
the substance, differing from those of adrenalin, which
is a pure sympathetic stimulant. Adrenalin, when in-
jected, causes both glycosuria and tachycardia, but it
216 Manual of Vital Function Testing Methods
does not produce, ordinarily, pallor of the skin, nor
contraction of intestine and uterus, as does hypophysis.
The pneumogastric is generally considered the nerve
which produces intestinal peristalsis. Since hypophysis
produces peristalsis, it must stimulate the 10th pair.
If now we consider the effects of hypophysis in
Graves' disease, we may begin by admitting that in this
condition there is a general state of erethism or hyper-
excitation of the entire sympathetic and parasympa-
thetic (vagus-autonomic) systems. That the sympa-
thetic is excited is proved by the exophthalmus, glyco-
suria and tachycardia. On the contrary, the symptom
diarrhoea which is so constant and characteristic a
symptom in Basedow's disease is explained by Ep-
pinger and Hess and others on the theory of a hyper-
vagotonia.
When one injects into a Basedowian an extract of
hypophysis, with the exception of the pulse rate, the
effects are generally similar to those obtained upon the
normal subject.
The heart slowing phenomenon, however, found by
Claude, Baudouin and Porak, in Basedowians following
injections of hypophysis extract remains to be ac-
counted for. The authors believe that the slow-
ing is due simply to stimulation of the vagus
nerve or 10th pair. They think that hypophysis
extract acts on the cardiac rhythm of the non-
Basedowian by stimulating the accelerator sym-
pathetic. In the Basedowian, however, there is al-
ready a tachycardia which is due to the continual hyper-
excitation of the sympathetics. These nerves being
already in a state of hyperexcitation do not react to
hypophysis extract. The terminations of the vagus
which are not excited, therefore, feel the full effect of
the hypophysis stimulation and the heart is temporarily
The Ductless Glands 217
slowed while the effect lasts.
Naturally, the authors of the "hypophysis test"
mention the possibility of its use in the diagnosis of
latent forms of Graves' disease. If future use of this
test should corroborate the early work of Claude, Bau-
douin and Porak in this regard the "hypophysis test"
will become an important adjunct to the functional
diagnosis of the hyperthyreopathies.
With respect to its applicability to the diagnosis of
the forme fruste or latent form of Graves' disease, the
authors report two very instructive cases. In one a
woman 27 years of age, of neuropathic taint, suffering
from tachycardia, dysmenorrhoea and slight hand
tremor without thyroid enlargement, the injection of
1 c.c. of hypophysis extract produced a slight increase
in the rate from 120 to 126 beats. The test was there-
fore negative. In the second case, that of a nervous
man 47 years old, with a tachycardia (100 to 105
pulsations) with slight exophthalmia and slight hand
tremors, with no apparent thyroid enlargement, the in-
jection of hypophysis extract slowed the pulse from
100 to 84 in four minutes. The test was, therefore,
positive. The case was a true latent form of Graves'
disease, that is, the syndrome of hyperexcitation of the
sympathetic nervous system presented by the patient
was truly connected with and due to a hyperfunctiona-
tion of the thyroid gland.
The authors likewise found, which may be mentioned
for its scientific interest only, that in cases of paroxys-
mal tachycardia the test is negative, as would on a
priori grounds be expected. In this condition the
pathogenesis resides not in the sympathetic nervous sys-
tem as a whole nor in any dysfunction of the thyroid
nor any other endocrinopathy but in changes that have
taken place in the cardiac musculature.
218 Manual of Vital Function Testing Methods
2. THE ADRENALIN MYDRIASIS TEST OF LOEWI 19
In 1907 Loewi found that in pancreatcctomized ani-
mals the instillation of 1-1000 solution of adrenalin pro-
duced marked dilation of the pupil. In human beings
with diabetes Loewi found the same effects. In 30-60
minutes a marked dilation occurred in diabetic cases.
The application of this phenomenon to the detection of
pancreatic insufficiency has already been mentioned.
Loewi also made a simultaneous observation that the
instillation of 1-1000 solution of adrenalin into the con-
junctional sac in cases of Basedow's disease, likewise
resulted in dilation and proposed the method as a test
for hyperfunction of the thyroid gland on the ground
that the internal secretion of the thyroid and suprare-
nal are synergistic, both acting by stimulating the sym-
pathetic nervous system. In cases of hyperthyroidism,
the sympathetic nervous system is in a state of increased
irritability, therefore the dilator fibers of the iris which
are governed by sympathetic nerves respond with ab-
normal alacrity to the instillation of adrenalin. Loewi's
findings were corroborated by Falta 20 and Zak.21
Eppinger, Falta and Rudinger,22 found an increased
adrenalin mydriasis in dogs which had been fed with
thyroid extract. In depancreatized and thyroidectom-
ized animals the reaction was absent. Eppinger and
Hess 23 also reported the test positive in Basedow's
disease.
The interesting and extremely simple test of Loewi
19Wien. klin. Wchnschr., 20, 1907, 782; Archiv f. exper. Path,
und Pharm., 59, 1908, 83.
^Wien. klin. Wchnschr., 20, 1907, 1559.
"Verhandl. d. 25 Kong. f. inner. Med., 1908, 392.
22 Wien. klin. Wchnschr., 21, 1908, 241.
23 Verhandl. des 26 Kong. f. inner. Med., 1909, 385.
The Ductless Glands 219
has not been much discussed in the literature in recent
years. It would be interesting to determine whether an
increased susceptibility of the iris sympathetic as
shown by mydriasis exists in cases of latent Graves'
disease.
3. TEST OF EXPERIMENTAL HYPERTHYROIDISM. ADMIN-
ISTRATION OF THYROID EXTRACT, IODINE AND IODIDE OF
POTASSIUM AS A MEANS OF DISCLOSING FUNCTIONAL
HYPERACTIVITY OF THE THYROID
Fr. v. Mueller, who criticized the metabolism tests
for hyperthyroidism as being too complex, suggested
the administration of iodine as a means of disclosing
hyperthyreosis. But apparently Mueller only made
the general suggestion and did not elaborate any speci-
fied technique. Since no one else has done so, it can-
not be said that an "iodine test" exists for determining
the presence of a latent hyperthyroidism. Patients
with hyperthyreosis often show intolerance to iodine by
developing emaciation and tachycardia, after its admin-
istration.
Many attempts to produce experimental thyroidism
in animals by feeding thyroid substance have been made,
and there seems to be great variation in the resistance
of different genera to thyroid ingestion. When symp-
toms appear in healthy animals the most constant signs
seem to be emaciation and diarrhoea (Carlson,24 Bal-
let25).
Kraus and Friedenthal 2G found that the intravenous
injection of thyroid juice in rabbits also produces en-
largement of the palpebral fissure, projection of eye-
M Prac. Am. Physiol. Soc., 1910-11, XXVII, p. XIII.
25 Limousin Med., 1896, XX, 69.
* Berl. klin. Wchnschr., 1908, 1709.
Manual of Vital Function Testing Methods
balls and enlargement of the pupil. Other authors have
succeeded in obtaining similar results.
Since the introduction of thyroid preparations into
clinical medicine, artificially produced hyperthyroidism
has been observed following their indiscriminate admin-
istration. The continued injection of thyroid extract
is frequently followed by symptoms of intolerance such
as subjective sensations of heat, perspiration, palpita-
tion or tachycardia and occasionally glvcosuria, all of
which denote hyperthyroidism.
A few cases have been reported in which a typical
Graves disease syndrome has been produced by the
administration of thyroid extract. The symptoms dis-
appeared after a suspension of the treatment.
It is a well known fact that the administration of
iodides over long periods to cases of goitre may pro-
duce symptoms of hyperthyroidism (Kocher).27 To
these cases the name iodine-Basedow has been given.
The actual administration of thyroid extract, iodine,
and iodide of potassium to disclose a latent hyperthy-
roidism or Graves' disease is not to be recommended as
a routine procedure. Most writers advise against the
use of iodine, iodides or thyroid extract in any case
where there are signs of emaciation (Krecke 2S).
Taking all the above facts into consideration, it will
no doubt be admitted that there is little, if any, justi-
fication for the administration of either iodides, iodine
or thyroid extract in cases of suspected Graves' syn-
drome with a view of thereby developing indubitable
signs of the disease. It cannot be said to be justifiable
under any circumstances to attempt to convert a latent
or doubtful into an outspoken case of Graves' disease
for purposes of diagnosis.
27 Verhandl. d. Dcutsch. Ges. f. Chir., Berl., 1910, 396.
28 Munch, mod. Wchnschr., 1911, LVIII, 1601 and 1676.
The Ductless Glands
There appears to be among many medical men a
lack of appreciation of the dangers which are attached
to the indiscriminate use of thyroid extract and some
surgeons have stated that many cases of Graves' dis-
ease coming under their observation for operation give
a history of previous ingestion of thyroid extract.
It would certainly seem rational to assume that noth-
ing but harm can come from such a practice.
These facts are, of course, well known and appreci-
ated by a very large majority of medical men, and be-
cause of this knowledge no systematic attempt has
ever been made to develop a test of experimental thy-
roidism. The use of thyroid extract, iodides, or iodine
for such a purpose can only be mentioned to be con-
demned.
4. THE ACETO-NITRIL, TEST OF REID HUNT
It was in the effort to develop a quick and satisfac-
tory method for comparing the physiological activity
of different thyroid preparations that Hunt 29 dis-
covered the remarkable fact that mice when fed upon
thyroids develop an increased resistance to aceto-nitril
or methyl cyanide, CH.CN. This substance produces
toxic effects chiefly through the slow liberation of
hydrocyanic acid in the body. Since thyroid feeding
does not alter the resistance of mice to hydrocyanic
acid, it is probable that its action so far as aceto-nitril
is concerned, is exerted upon the processes by which
the substance is decomposed in the organism.
Hunt found that when small amounts of thyroid
are fed to mice for a few days these animals acquire
^Amer. Jour, of Physiol., 1899, III; Proc. Soc. Exp. Biol., N.
Y., 1905, Oct. 18; Jour. Biol. Chem., I, 33, Oct., 1905; Jour. Amer.
Med. Assn., 1906, XLVII, 790; Hygien. Lab. Bull., No. 47, 1907.
222 Manual of Vital Function Testing Methods
a markedly increased resistance to aceto-nitril. This
is true for both white and gray mice, although most of
his experiments were performed upon the former va-
riety.
A mouse which had received thyroid in the form
of cakes, recovered from 17 times the relative amount
of aceto-nitril fatal to a control. Another mouse
recovered from 16 times the relative dose fatal to con-
trols. A third mouse recovered from 11 times, a fourth
from 6 times and a fifth from 2V2 times the fatal dose
to controls.
Hunt suggested this reaction as a delicate test for
thyroid substance.30 He found no other substance with
an effect upon the resistance of mice to aceto-nitril at
all comparable to thyroid. The test is more delicate
than any chemical test.
Hunt suggested in 1907 that this method is adapted
to throw light on the question as to whether there is
an excessive amount of thyroid secretion in the blood
in cases of Graves' disease. He applied the test in three
cases. In one of these the blood of the patient had a
marked effect in increasing the resistance of mice to
aceto-nitril, indicating thereby an excess of thyroid
secretion. In a second case the results were doubtful
and in a third case, negative.
Hunt suggested that the test might have some diag-
nostic value though he points out that it is not neces-
sary to assume that in Graves' disease there is always
an excess of thyroid secretion present in the blood at
all times.
To carry out the method, he suggested that the
best results might be obtained by administering to
mice 1 or 2 c.c. of blood made up with meal in the
form of cakes, for 9 or 10 days before testing with
30 Jour. Amer. Mecl. Assn., 1907, XLIX, 240.
The Ductless Glands
nitril. Controls are indispensable. One-fourth of a
milligram of aceto-nitril per gram of body weight of
mouse may be fatal to a normal animal in a few hours.
Hunt used doses of a fraction of a milligram, one-
fourth, to several milligrams, 1, 2, 4, in his experi-
ments.
Hunt believed as a result of his researches that the
activity of a given thyroid preparation or substance
is parallel with its iodine content.
The test suggested by Reid Hunt has not been
extensively developed. His findings were, however, sub-
stantially corroborated by several authors, among
whom may be mentioned Trendelcnburg in 1910, 31
Ghedeni in 1911.32
Before a final judgment as to the value of this pro-
cedure can be formed it will be necessary to determine
the resistance of mice to aceto-nitril after feeding with
the blood of patients with Graves' disease in a large
series of cases. Also experiments should be done to
determine the resistance of mice to aceto-nitril after
feeding with normal blood, for perhaps the test is so
delicate that even the amounts of thyroid secretions
present under normal circumstances may be sufficient
to increase the animal's resistance. Of course the vari-
ations in natural resistance of the animals both as
regards species and seasons which have already been
demonstrated as well as the possible variations in the
blood content of thyroid substance even under normal
circumstances may so complicate and obscure results
that the test may be found impracticable.
There is, however, something extremely suggestive
about this type of biological experimentation which
makes it seem probable that some such test will be
31 Biochemische Zeitschr., 1910.
S2Wien. klin. Wchnschr., 1911, XXIV, 736.
Manual of Vital Function Testing Methods
discovered in the future of real value in the diagnosis
of hyperthyreosis.
5. METABOLIC STUDIES AS CRITERIA OF
HYPERTHYROIDISM
In Graves' disease, as was above mentioned, the
metabolic changes are increased. Biedl says they are
so characteristic of the condition as to constitute an
important diagnostic criterion. The metabolism of
Graves' disease is accompanied by a pretty constant
increase in the expenditure of energy.
The respiratory gas interchange shows an increase
of 50%, sometimes 70% to 80% in the amount of oxy-
gen consumed, according to Magnus-Levy,33 Salomon,34
and others.
The increased production of heat is usually accom-
panied by an augmented metabolism of albumen and
fats. The assimilation of carbohydrates is diminished
in Graves' disease and it is for this reason that ali-
mentary glycosuria is readily produced.
Kraus was the first to suggest that determinations
of the respiratory metabolism (increase of CO2 and N),
by the use of the Zuntz-Geppert apparatus may be use-
ful in the functional diagnosis of latent or outspoken
hyperthyroidism. Fr. v. Mueller, however, observed
that the method is too complicated for practical work.
Studies of basal metabolism can be calculated by
indirect calorimetry from the oxygen absorption and
the respiratory quotient, using a Benedict unit appar-
atus (mouthpiece and spiromcter). At least three
ten-minute periods are run and the average taken for
33Bcrl. klin. \Vclmsrhr., 1S95; /cit. f. klin. Mod., 33, 1897; Noor-
di-m's Handbook d. Path. Stoffwi-chs., II, 3.52, 1907.
31 Bcrl. klin. Wdmschr., 1901.
The Ductless Glands 225
that day's basal metabolism.
Means 35 has recently reported some studies of basal
metabolism and its relation to body surface in obesity,
myxedema and pituitary disease. He found a diminu-
tion of 27% below normal in myxedema.
The emaciation which occurs in many cases of Graves'
disease naturally points to an increased katabolism.
Kocher found reduction in weight in 88% of his cases.
As much as 15 to 20 kgs. may be lost in a few
months. The loss of weight is an early symptom. The
basic cause for this loss in weight is the remarkable
increase in the caloric production which occurs in
Graves' disease. By using the Zuntz-Geppert appa-
ratus many authors have made the demonstration of
increased oxygen consumption and increased CO2 elim-
ination. Magnus-Levy and Salomon have already been
mentioned.
Experiments have also been made in the Voit-Petten-
kofer apparatus which give like results. The increase
of caloric production as was before stated may reach
as high as 70% or more above normal. In some cases
(Magnus-Levy) the oxygen consumption has been found
from over 5 to nearly 7 ccm. per kilogram of body
weight. Salomon has shown that these metabolic dis-
turbances occur and may be demonstrated in the latent
cases.
It is to be hoped that the gradual simplification of
methods for studying metabolism will lead to practical
clinical results which will undoubtedly find a rich field
of application in the functional diagnosis of thyroid
diseases.
^Proc. Soc. fr. Exp. Biol. and Med., 1914, XII, 1913.
226 Manual of Vital Function Testing Methods
6. APPLICATION OF THE PRINCIPLE OF COMPLEMENT
DEVIATION TO FUNCTIONAL DIAGNOSIS OF HYPER-
THYROIDISM. MARINESCO-ROSEO TEST
Marinesco 36 in 1911 and Roseo 3T in 1912 appear to
have been the first to suggest that in Graves' disease
there is thrown out into the blood serum sufficient thy-
roid substance (antigen) to give rise to the formation
of antibodies (amboceptors) in the patient's blood.
They therefore proposed to test for the presence of
these antibodies in the blood serum of suspected cases of
Graves' disease by means of an antigen prepared from
thyroid tissue removed at operation from an outspoken
case of Graves' disease.
Both Marinesco and Roseo have studied the reaction
of fixation of alexine (complement) in cases of Base-
dow disease and they believe that the positive results
obtained proved the existence of true specific antibodies
in the blood in this condition. This test may in future
be found useful from the standpoint of functional diag-
nosis.
The principle of the complement fixation or deviation
test depends, as is well known, upon the observation
that the injection of the living organism with bodies of
a proteid nature, cells, bacteria, organ extracts, etc.,
results in the formation by the organism of certain an-
tagonistic bodies called antibodies. Perhaps these anti-
bodies are ferments. The bodies that are inoculated
in order to produce antibodies are called antigens.
Some antibodies such as the agglutinins and precipitins
act directly on the specific agent or antigen which
produces them. Other antibodies, such as cytolysins
and hcmolysins, act only in the presence of a third
^Deutsch. Zeits. f. Nervenheilk., 1911, XLI, 268.
"Biochem e terap. sper., Milan, 1912-13, IV, 1.
The Ductless Glands 227
body, which is always present in blood serum or tissue
juices to which the name complement has been given.
It is upon this state of facts that the now famous Was-
sermann reaction is founded. The performance of the
complement fixation test particularly in the diagnosis of
syphilis has become a part of the routine work of almost
every well equipped pathological clinical laboratory.
Consequently there will be no difficulty in a well
equipped hospital for the clinician to have complement
fixation tests performed.
I have been unable to obtain the exact results of
Roseo's work. Marinesco's 38 observations upon the
reaction of fixation of complement included two series
of experiments. In the first he used an aqueous extract
of goitre from a classical case of Graves' disease as
antigen and the serum of the same patient for antibodies
and that of four other patients with the same disease.
For controls he used the serum of normal persons. The
fixation was complete in the first case whose goitre
furnished the antigen, while in two other Basedowians
there was incomplete hemolysis, and in a fourth the
hemolysis was complete as in a normal control. An
objection which may be urged against this first series as
pointed out by Marinesco is that he did not have at
his disposition an extract of normal thyroid. Later,
with the assistance of Madame Papazol, he repeated
his experiments, this time making an examination of
23 sera, and using different extracts from the thyroid
gland of cases of Graves' disease, also from one goitrous
thyroid and one normal gland.
The extracts were prepared in the usual way for
ether extracts. Eight grams of Basedow goitre were
triturated in a mortar and 100 grams of ether added,
drop by drop. The mixture was put in a sterilized
38 Loco citato.
Manual of Vital Function Testing Methods
glass and placed in a shaker. After shaking and filtra-
tion, the mixture was kept for 48 hours in the thermo-
stat. After the evaporation of the ether a little car-
bolated water was added, 40 c.c. for 8 grams of sub-
stance. The extract was then again shaken and filtered
through cloth. The prepared extract was kept on ice
in a dark bottle. The alcoholic extract was similarly
prepared.
In the two cases in which Marinesco made use of an
autoextract of the thyroid from cases of Graves' dis-
ease, the prevention of hemolysis was complete. He
found the same absolute prevention of hemolysis in six
other cases of Graves' disease when he employed thyroid
tissues obtained from other Basedowians. Aqueous, al-
coholic and ethereal extracts appeared to act about the
same, but sometimes the ether extract seemed more
active.
In most cases of Graves' disease Marinesco obtained
either a total absence of hemolysis or an incomplete or
partial one. On the contrary, the serum of Graves'
disease cases never fixed complement in the presence of
normal thyroid body or extract of ordinary goitre.
He got the same results when he used serum from normal
persons and Basedowian antigen. In one case, however,
Marinesco and Madame Papazol found that the serum
of a syphilitic patient gave a partial hemolysis with
ether and alcoholic extract of normal thyroid. Other
authors have likewise found a more or less complete
fixation with syphilitic serum in the presence of normal
thyroid extract. Mueller, of Vienna, in a personal com-
munication to Marinesco stated that in a case of
Graves' disease he had noted a fixation of complement
in the presence of alcoholic extract of heart.
Marinesco believes that his experiments tend to show
the presence of an antigen in the thyroid gland of Base-
The Ductless Glands 229
dowians and that the reactions of fixation which he
obtained are not simply due to an increase in the active
substance (internal secretion) of the thyroid gland but
to a change in the colloidal state of this substance, due
to the harmful effects of a pathogenic agent. Marines-
co calls attention to the difficulty in penetrating more
deeply into the mechanism of his fixation reaction since
authors in general are by no means in accord in explain-
ing the mechanism of the fixation reaction discovered
by Wassermann in the serum of syphilis. Some authors
like Wassermann himself, seeing in the reaction the
existence of true specific syphilitic antibodies, others on
the contrary considering it as a non-specific physical
chemical phenomenon.
The practical possibilities for functional diagnosis,
of his findings are believed by Marinesco to be worthy
of mention.
For the Marinesco-Roseo test it will of course be
necessary to secure thyroid gland tissue at the time of
operation in a case of outspoken Graves' disease.
As to the clinical value of the test so little work has
been done by investigators subsequent to the reports
of Marinesco and Roseo that no definite opinion can
be formed as to its value. This will become a matter
for future investigation. It is to be hoped that the test
will be carried out in a sufficient number of outspoken
cases of Basedow's disease to determine in just what
percentage it will be positive. If the findings are cor-
roborated, the test should be applied to a series of cases
of suspected latent hyperthyroidism. At the present
time the final decision as to its value remains sub judice.
230 Manual of Vital Function Testing Methods
7. SPECIFIC FERMENT TEST OF ABDERHALDEN APPLIED
TO FUNCTIONAL DIAGNOSIS OF THE THYROID
Lampe 39 has attempted to apply the Abderhalden
dialysis method to the study of the blood serum of
patients with Graves' disease. He believes that in the
blood serum of these patients ferments exist which are
specific for thyroid tissue.
This method has never been applied extensively to
the functional diagnosis of incipient Graves' disease
since its introduction by Lampe. The technical diffi-
culties attached to the carrying out of Abderhalden's
method and the many conflicting results obtained in
its general use have prevented it from becoming popu-
lar in clinical practice. Perhaps in the future when the
method is simplified and its precise limitations defined,
some practical results may be hoped for.
Lampe first demonstrated in 1913 that normal blood
serum obtained from healthy individuals does not con-
tain any ferments capable of splitting the tissues of any
of the organs.
In the same year Lampe and Papazolu 40 examined
the effects of the serum from cases of Graves' disease
to determine the presence or absence of proteolytic fer-
ments specific for thyroid tissue. Lampe thought that
by the results of the dialysis method he might be enabled
to throw some light upon the question as to whether in
Basedow's disease there is a hyperthyroidism or a dys-
thyroidism. He does not mention the possibility of
employing the test as an aid towards the functional
diagnosis of the disease.
Lampe argued that if in Graves' disease there is an
over-production of the normal thyroid secretion, there-
""Miinch. med. Wchnschr., 1913, 26.
40 Munch, med. Wchnschr., 1913, 28.
The Ductless Glands 231
fore a negative result of the Abderhalden reaction
(serum -}- thyroid gland) would be expected because
in this case there would simply be the introduction into
the blood of a purely native protein only in increased
amounts and consequently no development of ferments.
If on the contrary, Graves' disease is a true dysthyroid-
ism, if, in other words, the thyroid gland in Graves'
disease pours into the blood a qualitatively altered pro-
teid secretion, produced by the pathological changes
in the gland, then this secretion acting as a foreign
proteid would be expected to stimulate the produc-
tion of protective ferments, and the Abderhalden reac-
tion would be positive. Lampe hoped also to be able to
throw some light by his method upon the role which the
thymus plays in Graves' disease.
Lampe and Papazolu experimented with the serum
from Basedow cases upon normal thyroid gland, ex-
ophthalmic goitre gland, cystic and parenchymatous
goitre, normal thymus, Basedow thymus and several
other organs and tissues as ovary, testicle, kidney, su-
prarenal, pancreas, etc. In their article they give the
protocols of experiments upon the serum of twenty-five
cases of exophthalmic goitre.
In all cases in which the serum from the Graves'
disease cases was allowed to act upon exophthalmic goi-
tre tissue, the tissue was digested. In very few cases
only was the reaction positive when normal thyroid
tissue was used. In four out of five of the cystic goitre
products, in almost all thymus and ovarian tissues they
found the reaction positive. With all other substrate
kidney, liver, pancreas, etc., the reaction was negative.
Lampe believes that his researches demonstrate that
in Graves' disease there is a true dysthyroidism and
not a simple hyperthyroidism.
Principle of the Abderhalden Method. — The basic
232 Manual of Vital Function Testing Methods
principle underlying the now much discussed method
of Abderhalden 41 is the fact that albumen, being a
colloid does not diffuse through animal membranes,
while, on the other hand, the peptones, which are the
'first products of its decomposition, are diffusible. If
albumen is put in a dialysing tube and the latter placed
in water no albumen appears in the surrounding fluid,
even after a considerable lapse of time. If pepsin and
HC1 are added to the albumen solution peptones are
formed and will appear in the dialysate. If it is desired
to determine whether a liquid contains any proteolytic
substance or ferments, the solution may be placed in a
dialysing tube and peptone will appear in the surround-
ing media.
In this way blood scrum, cerebrospinal fluid, lymph,
extracts of organs, etc., may be tested.
The actual carrying out of the Abderhalden method
is extremely difficult, so much so that the method can-
not be used in the ordinary routine of clinical work. If
the method is to be tried the individual who proposes
to do so will find it advantageous to consult the little
work of Abderhalden himself, which has recently been
translated and to which reference has been given.
B. Hypof unction of the Thyroid Gland (Myx-
edematous States). — The best concrete example of the
loss of function of the thyroid in human beings is met
with in those cases in which the whole gland has been
removed by operation for goitre.
Reverdin 42 in 1882 was the first to describe the
results of goitre extirpation. In 1882 Kocher 43 pub-
lished his classical report on the same condition. The
41 Defensive Ferments of Animal Organism, Abderhalden; tr.
by Gavronsky and Lanchester, Lond. Bale. Co., 1914.
12 Rev. Med. de le Suisse Rom., 1882, 539.
"Archiv f. klin. Chir., 1883, XXIX.
The Ductless Glands 233
names operative myxedema or cachexia strumipriva
were given to this condition. Inasmuch as the thyroid
gland is never completely excised at the present day the
subject has become of historical interest only.
What is of greater practical moment is the fact
that symptoms somewhat similar to the cachexia
strumipriva may spontaneously arise in adult human
beings and give rise to the now well known but only
too often overlooked syndrome of spontaneous or
idiopathic myxedema of adults, Gull's disease. The
symptoms are produced by retrogressive changes in the
thyroid gland. Similarly there may be congenital
states of hypothyroidism and infantile types, develop-
ing after birth, the so-called sporadic cretinism. Fi-
nally in some countries a condition known as endemic
cretinism exists. All the above types of disease are
associated with a diminution of function of the thyroid
gland.
In all states of hypothyroidism the gland itself under-
goes retrogressive changes. There are symptoms refer-
able to the skin and subcutaneous tissues, the nervous
system, metabolism, the bones, blood, etc. We shall not
attempt to go into details in regard to the semiology
of these interesting conditions.
The diagnosis of myxedematous states is easy in
typical cases but even here many cases are overlooked
by the practitioner. In latent cases, however, the
formes frustes, the diagnosis may not be easy. Some-
times the edema is taken to mean Bright's disease. I
have seen the nervous symptoms, speech difficulties and
gait disturbance ascribed to chronic alcoholism.44
Kocher calls the latent form of myxedema, thyropenia.
All authors call attention to the great frequency with
which it is overlooked.
"Jour. Am. Med. Assn., 1915, LX1V, 986.
234 Manual of Vital Function Testing Methods
The functional diagnosis of thyropenia is intimately
bound up with the treatment for there is but one diffi-
culty and that is to suspect the disease. Once sus-
pected there is one infallible test, the therapeutic
test.
Therapeutic Test for Lowered Functional Activity of
the Thyroid Gland
This consists in commencing the administration of
thyroid extract. If the case is one of thyroid insuffi-
ciency the symptoms will magically disappear. If they
are not entirely gone or improved in two weeks, the test
is negative. The condition is not one of hypothyroid-
ism.
Thyroid gland is best given in the form of tablets
of the dried gland. The tablets contain 1% to 5 grains
(.1— .3 gm.) of desiccated thyroid gland. Begin by
administering a small dose after each meal or less often.
The patient should lie down for twenty minutes after
swallowing the tablet.
The dose is gradually increased until 6-10 tablets
per day are given, care being taken not to produce rapid
heart action, sweating, diarrhoea or nervousness, which
are symptoms of intolerance.
From a therapeutic standpoint, which point we can-
not discuss in this place, it is well known that the ad-
ministration of thyroid gland in states of hypothyroid-
ism must be kept up indefinitely, for so soon as the
treatment is stopped, the symptoms will invariably
recur.
THE PARATHYROID GLANDS
The first person to specifically describe the para-
thyroids was the Swedish anatomist, Sandstrom, in
The Ductless Glands 235
1880. Gley practically rediscovered them in 1891.
Since the latter date a very considerable literature has
risen upon these interesting structures.
Following the removal of two or more of the four
parathyroid glands in the human subject, tetany comes
on in from 2 to 5 days. All the special symptoms of
tetany are present. Trousseau's, Erb's, Chvostek's and
Hoffmann's signs, with irritability of the nerves of spe-
cial sense and the sympathetic, irregular pulse, arterial
spasm, angioneurotic oedema, spasm of gastrointestinal
tract, leucocytosis and disturbance of heat regulation
may occur.
After incomplete extirpation or temporary injury
of the glands, milder symptoms occur called tetanoid or
subtetanic hypoparathyreosis (Halsted). Sometimes
the sj'mptoms are latent and come on sometime after
injury or as a result of pregnancy, trauma or infection.
In the well-known infantile tetany lesions of the para-
thyroids have been found.
Other convulsive diseases such as epilepsy, paralysis
agitans, myoclonus, myotonia and myasthenia have
been supposed to be due to disease of the parathyroids,
but the exact facts in this direction are as yet unknown.
The exact relation between the thyroids and para-
thyroids is not known, some thinking that an an-
tagonism, others that a synergism, exists between the
two.
There is no method at present known of experimen-
tally determining the functional activity of the para-
thyroid glands.
THE THYMUS GLAND
The function of the thymus is as yet not definitely
known. It is assumed that inasmuch as the basic
236 Manual of Vital Function Testing Methods
structure of the thymus is that of lymphoid tissue in
general, that there is a related function between the
two, in other words, that lymphocytes and eosinophiles
are formed in the gland.
There are, however, present in the thymus structure
some epithelial elments, the so-called corpuscles of
Hassal. What their function may be is quite unknown.
Many believe that in the epithelial cells an internal se-
cretion is elaborated which has to do with the develop-
ment of the skeleton, nervous system, sexual apparatus
and general metabolism.
There is a general belief that a reciprocal action
exists between the thymus and testes, since castration
delays the involution of the thymus while removal of
the thymus causes rapid development of the testes.
The fullest development of the thymus is reached
at the end of the second year of life. From this time
on to puberty it gradually atrophies and in adults is
represented only by a small mass of fibrous tissue and
fat. Occasionally, however, the thymus gland persists
or undergoes hypertrophy, producing symptoms of
tracheal stenosis with attacks of laryngeal stridor or
asthma, and sometimes there is sudden death, the so-
called mors ihymica. A condition known as the status
lymphaticus may gradually develop in which there is
more or less anemia, with lymphocytosis, together with
rachitic and gastrointestinal symptoms.
No tests have so far been devised for determining the
functional activity of the thymus. The diagnosis of
its diseases is strictly semiological and in the diagnosis,
radiography has recently been of considerable assist-
ance.
The Ductless Glands 237
THE SUPRARENAL GLANDS
Our more intimate knowledge of the suprarenal
glands appears to date from the year 1855, when Addi-
son published his famous work upon the disease which
bears his name, though they have been known since
1564, the date of their discovery by Eustachius.
A tremendous amount of work has been done upon
the adrenals by investigators in the past half century.
Much remains to be learned but certain facts appear to
have been gained. These may be briefly stated as fol-
lows : disease of the glands resulting in their gradual
atrophy or destruction gives rise to a train of symp-
toms characterized chiefly by pigmentation of the skin
and extreme weakness, i. e., Addison's disease. Ex-
tirpation in animals of both adrenals is an extremely
dangerous operation and according to most authori-
ties leads infallibly to death. Extracts obtained from
the medullary or central part of the organ are toxic
when administered to animals, among the symptoms
being glycosuria and arterial degenerations. The
same extracts when injected intravenously produce a
powerful constriction of the blood vessels with rise of
the blood pressure due to stimulation of the sympa-
thetic nervous system.
It is generally accepted by clinicians and patholo-
gists that the adrenal medulla elaborates an internal
secretion and that adrenin is the product of this se-
cretion.
The exact function of the adrenal cortex is still un-
known. The cortex contains a considerable quantity
of lipoid and cholinogen substances, the presence of
which has given rise to the hypothesis that neutraliza-
tion of toxic substances is effected here.
In 1894) Oliver and Schafer noted that extracts of
238 Manual of Vital Function Testing Methods
adrenal medulla produce a rise of blood pressure. In
1897 and 1898 Furth and Abel and in 1901 Takamine
and Aldrich succeeded in gradually separating and
finally isolating in pure form the active substance of
adrenal medulla — adrenin.
From a clinical standpoint the functional activity of
the suprarenal glands may be considered from two
points of view. From the first the functional activity
of the glands may be considered to be lowered and
from the second it may be regarded as raised. To
the first condition, the name hypoepinephria or hypo-
adrcnalism has been given, and to the second hyper-
epincphria or hyperadrenalism.
In our brief discussion of the functional examination
of the suprarenal glands this classification will be found
most practical.
We may properly allude here again to the antago-
nism which is generally considered to exist between the
function of the adrenals (chromaffm system) and the
pancreas. It has already been stated that the adrenals
and thyroid are functional synergists. The adrenal
secretion inhibits carbohydrate catabolism and raises
blood sugar, while the pancreas hormone facilitates
carbohydrate catabolism arid lowers blood sugar. The
effect then of lowering the adrenal function is to raise
that of the pancreas, namely, to facilitate carbohydrate
catabolism and lower blood sugar to lead to hypogly-
cemia, oliguria and hence absence of glycosuria. The
effect of raising the adrenal function will be to inhibit
carbohydrate catabolism and to raise blood sugar,
hence to lead to hyperglycemia diuresis and glycosuria.
A. Hypofunction of the Suprarenal Glands. — Sev-
eral different clinical forms of hypoepinephria or low-
ered adrenal function have been described, but few of
The Ductless Glands 239
them have been generally recognized by clinicians as
separate morbid entities. The essential features of
lowered adrenal function appear to be myasthenia and
hypotension. The systolic blood pressure is usually
below 100 mm. Other features justifying a suspicion
of hypoepinephria are hyperesthesias, lumbar pains,
headache, delirium, coma, digestive disturbances and
sudden death without previous symptoms.
The chief clinical entity which is recognized as be-
ing accompanied by a true persistent hypoadrenalism
is Addison's disease.
Addison's disease, first described by Thomas Addison
in 1855, is a chronic condition usually appearing in the
third or fourth decade of life. It is characterized
clinically by pigmentation of the skin and mucous
membranes, by muscular and vascular weakness, dis-
turbances of the gastrointestinal tract and nervous
system, and final cachexia and death. Anatomically
it is accompanied by disease of both adrenals, usually
a caseous tuberculosis. We shall not attempt a de-
scription of the symptomatology or pathology of the
disease.
The clinical diagnosis of outspoken cases of Addi-
son's disease is sometimes easy. If there is a definite
history of weakness, vomiting, constipation and
diarrhoea, abdominal and lumbar pains and there is
present a pigmentation of the skin and mucous mem-
branes, and when pernicious anaemia and a few other
conditions which might be confused with it can be ex-
cluded, the diagnosis is reasonably certain. In Addi-
son's disease there is a mononucleosis and a hyper-
eosinophilia, in the blood.
Before the pigmentation occurs, however, the diag-
nosis is extremely difficult or impossible. Latent Ad-
dison's disease and other conditions of hypoepinephria
240 Manual of Vital Function Testing Methods
can only be disclosed by the application of principles
of functional diagnostic methods. Unfortunately, the
principles upon which a functional investigation might
be applied, toward the elucidation of adrenal dis-
turbances, have not been as yet developed to an extent
where they can be of great practical assistance to the
clinician.
One very evident possibility, however, suggests it-
self. If states of hypoepinephria are accompanied
by diminished function of the adrenals, there must be
a lessened amount of the substances in the blood which
represent the gland's activity. There is no practical
way at present to make use of such an hypothesis.
Tests for increased amounts of adrenalin in the blood
have been used to discover the opposite state of hyper-
adrenalism and these tests will be described below.
Eppinger, Falta and Rudinger 45 showed that in
cases of Addison's disease (hypoadrenalism) the
sugar tolerance is remarkably high. Polak 46 was un-
able to produce a glycosuria with 2 mg. doses of
adrenalin in a case of Addison's disease. Similar doses
in normal persons invariably produce glycosuria.
Meyer and Kahn corroborated these findings. These
facts form the basis for the application of various kinds
of glycosuria tests to the functional diagnosis of
hypoadrenalism.
Tests of Increased Sugar Tolerance as Evidence of
Hypoadrenal Function
The so-called sugar tests have been described at
length in the chapter on liver function testing. (See
45 See Erkrankungen der Blutdriisen. Falta. Wien, 1913.
"Wien. klin. Wchnschr., 1909.
The Ductless Glands 241
page 14.) In every case of suspected adrenal disease
the sugar tolerance should be investigated.
B. Hyperfunction of the Suprarenal Glands. — Just
what morbid conditions are associated with or produced
by hyperfunction of the adrenals is by no means clearly
understood. Certain tumors of the chromaffin tissues
have been held to produce symptoms connected in some
way with hyperadrenalism. The question as to
whether other conditions besides tumors of the adrenals
can give rise to states of hyperfunction is not de-
cided.
A number of pathological states of the organism
have at one time or another been claimed to owe their
origin to hyperfunction of the suprarenal glands or
chromaffin tissues generally. A school of pathologists
in France has for a long time endeavored to explain the
heightened blood pressure of nephritis on the ground
of an increased function of the suprarenal glands.
Further than this it has been held that the arterio-
sclerosis which accompanies the circulatory hypertonia
is the result of hyperadrenal function. Finally accord-
ing to some pathologists the whole process, of which
circulatory hypertonia and nephritis form important
parts, is to be ascribed to a primary hyperplasia of
the chromaffin tissues.
Certain authors have contended that they were able
in such conditions to demonstrate by means of the
Ehrmann-Meltzer reaction (v.i.) the presence of ex-
cessive amounts of adrenalin in the blood.
The question as to just what pathological processes
and clinical syndromes are to be held related to hyper-
adrenalism as effect to cause, is a question almost
entirely open and undecided at the present time. Fur-
ther than this the subject of functional diagnosis of
24$ Manual of Vital Function Testing Methods
hyperactive states of the adrenal glands has only begun
to be developed.
The principal methods suggested are two in number.
The first depends upon the generally accepted influence
of hyperadrenalism upon the carbohydrate metabolism.
There is said to be always a hyperglycemia. Hence
the demonstration of an excess of sugar in the blood
is one method of diagnosing a hyperepinephria, pro-
vided, of course, that other causes of hyperglycemia can
be excluded. There is unfortunately no simple method
of making the test.
The second class of functional tests for hyperad-
renalism depends upon the demonstration of excess
of adrenalin in the blood and the production of gly-
cosuria following the injection of adrenalin.
1. Tests for Adrenalin in the Blood as an Evidence of
Hyperadrenalism. The Ehrmann-Melt zer Reaction
It has long been known that intravenous injection of
adrenalin produces dilatation of the pupil. This fact
has been utilized as a test for adrenalin in various
fluids, as blood serum, urine, etc.
Meltzer and Auer,47 Wessely 48 and others have
found that when adrenalin is applied to the frog's eye
mydriasis is produced.
Ehrmann 49 studied this phenomenon and suggested
it as a delicate test for adrenalin. He found that
adrenalin acts upon the dilator (sympathetic) fibers
of the iris in a strength of 1 to 20,000,000. Dilatation
of the pupil of the frog's bulbus oculi immersed in salt
solution occurs when excessively minute quantities of
"Centralbl. f. Physiol., 1904, XVIII, 316.
48Zeitsch. f. Augenh., Aug. 13, 1905.
49Archiv f. exper. Path. u. Pharra., 1905, LIII, 96.
The Ductless Glands
adrenalin are present, quantities as small as .000025
mg. Later investigations have shown, however, that
other substances in blood serum will produce the same
reaction and therefore the practical availability of the
reaction as a test for adrenalin in the serum is vitiated.
2. Adrenalin Glycosuria as a Test of Hyper function of
the Chromaffin System
We owe to Blum 50 the discovery that the hypodermic
injection of adrenalin will occasionally produce
glycosuria. The reducing substance found in the urine
has been proved to be glucose and there is always a
hyperglycemia (Metzger51). The action of adrenalin
in thus producing a hyperglycemia is due to its well
known stimulating effect upon the sympathetic nervous
system ( Underbill 52) acting upon sugar storing organs
and causing them to relinquish their supply of dextrose
producing substances as glycogen.
Soon after Blum made this discovery the interesting
fact was tested and confirmed in many directions. It
was discovered that the glycosuria appears after the
exhibition of extract of adrenal substance as well as
after that of its active principle, adrenalin.
Adrenalin glycosuria appears after comparatively
small doses (.Ol-.l mg. ) and is readily provoked by
a subcutaneous injection. The injection of one or two
milligrams of adrenalin is followed in half an hour to
two hours by a glycosuria lasting three hours. The
glycosuria is always accompanied lay a hyperglycemia.
50 Deutsch. Arch, f . klin. Med., 1901, LXXI, 146.
01 Munch, mecl. Wchnschr., 1902, 478.
B2Amer. Jour. Physiol., 1906-07, XVII, 42.
244 Manual of Vital Function Testing Methods
3. Deviation of Complement in Functional Diagnosis of
Suprarenal Disease
Polito and Corelli 53 have attempted to apply the
complement fixation test to the diagnosis of suprarenal
gland disease (hyperf unction), using an alcoholic ex-
tract of suprarenal gland as antigen. Their results
were indeterminate.
THE HYPOPHYSIS
The pituitary gland or hypophysis, as is well known,
is composed of two portions, a larger anterior epithelial,
follicular, glandular portion and a posterior lobe con-
sisting of connective and vascular structures. Between
the two is a partly glandular, partly vascular por-
tion, the pars intermedia. The whole organ is con-
tained in a bony inclosure, the sella turcica, or pituitary
fossa of the sphenoid bone.
Since Marie first described the disease, acromegaly,
and Rogowitch noted hypertrophy of the pituitary
after thyroidectomy, both of which took place in 1886,
a very large literature has sprung into existence con-
cerning the physiology and pathology of the hypoph-
ysis.
The deep situation of this interesting organ at the
base of the brain makes experimental investigation very
difficult. The embryological and histological differ-
ences between the anterior and posterior lobe of the
hypophysis, made it extremely probable early in the
history of these investigations that a different func-
tional activity must be attributed to the two portions.
68 La Nouva Riv. Clin. Terap., 1911, XIV, 482.
The Ductless Glands 245
In this respect there is an analogy with some of the
other ductless glands.
The differentiation of the two systems in the hypoph-
ysis, from the standpoint of pathology, is especially
difficult because of the confined space in which the organ
is lodged, making it almost inevitable that disease of
one portion will affect the other.
The name of Gushing in our country is intimately
associated with our knowledge of the hypophysis on
account of his extensive experimental and clinical in-
vestigations.54
There is still much to be learned concerning the
physiology and pathology of the hypophysis. Certain
facts, are, however, pretty well agreed upon. The
pituitary is probably essential to life, i. e., it is a vital
organ. After its removal, animals soon die with severe
cachexia. The secretion of the posterior lobe is sup-
posed to gain access to the cerebrospinal fluid and the
general circulation. It is concerned in regulating
metabolism, particularly that of carbohydrates. It
affects also the growth of fat. The internal secretion
of the anterior lobe affects the processes of general
metabolism and especially growth.
Oliver and Shafer in 1895 55 discovered that ex-
tracts of the pituitary produce when injected into blood
vessels, a rise of blood pressure, like that of the ad-
renals. Three years later Howell 56 discovered that
only extracts of the posterior lobe have this effect.
As with the thyroid and adrenals there is the same
tendency among clinicians to regard the pathology
of the hypophysis as being manifested by states of
hyper- and hypo-function. Acromegaly or Marie's
54 The Pituitary Body and its Disorders, Phila., 1912.
05 Jour, of Physiol., 1895, 18.
56 Jour. Exper. Med., 1898, 3.
246 Manual of Vital Function Testing Methods
disease is regarded as a typical example of the former,
while adiposo-genital dystrophy or Frohlich's disease
is looked upon as an equally typical example of the
latter.
In our very brief account of the functional diag-
nosis of the pituitaropathies, brief because so little of
importance has been accumulated in medical literature,
we shall consider the two states, opposite and distinct,
producing diametrically opposite effects upon the or-
ganism: hyperpituitarism and hypopituitarism.
A. States of Hyperpituitarism. — The most typical
example is acromegaly or Marie's disease, first described
by him in 1886.57
Acromegaly is a chronic disorder characterized by
an abnormal increase in the size of the nose, lips, tongue,
lower jaw, hands and feet, by hyperplastic changes
in the bones and soft parts, usually accompanied by
considerable enlargement of the hypophysis and widen-
ing of the sella turcica. Symptoms of increased intra-
cranial pressure often occur. The vegetative nervous
system is in a state of hyperirritability. The most
common pathological finding is adenoma of the anterior
part of the hypophysis. There is increase of function
of the glandular hypophysis, a true hyperpituitarism.
We shall not enter into the etiology and pathology
or into the detailed account of the general symp-
tomatology of acromegaly. As to the general diag-
nosis of the disease it may be said to be quite easy in
outspoken and typical cases. It is, however, difficult
in the early stages, the so-called latent period of the
disease. Acromegaly must be differentiated from cer-
tain diseases which may partially resemble it. Brain
tumor, arthritis deformans, Graves' disease, diabetes,
" Revue de Med., 1886, p. 298.
The Ductless Glands 247
progressive muscular atrophy, have all been diagnosed
as present when acromegaly really existed.58
Combinations of acromegaly with Basedowian or
myxedematous symptoms sometimes occur in the earlier
stages. X-ray examination of the sella turcica (Oppen-
heim) is, as is now well known, of extreme diagnostic
value.
Two methods of functionally determining the exist-
ence of hyperpituitarism have been suggested. They
are:
1. Demonstration of Disturbed Metabolism, as shown
by increase of gas exchange.
2. Demonstration of Alimentary or Spontaneous
Glycosuria.
1. Demonstration of Metabolic Disturbance as Shown
by Increase of Gas Exchange as an Aid to the Func-
tional Diagnosis of Hyperpituitarism
Very little work has been done upon the study of
the gas exchanges in acromegaly. That which has
been done has been carried out with the Zuntz-Geppert
apparatus. Cases have been examined by Magnus-
Levy, Salomon, Bernstein and Falta.
In all, there are seven cases reported, in which de-
tails are given as to the clinical facts and the amount
of oxygen consumption and carbon dioxide production
in ccm. per kilogram per minute. The figures vary for
oxygen from 5.19 ccm. down to 3.55 and for C02 from
4.33 down to 2.73. Falta states that the results of
the cases so far investigated do not demonstrate an in-
evitable increase in gas exchanges in acromegaly, as is
the case in hyperthyroidism.
58 Modern Med., Osler-McCrae, 1915, IV, 813. (Dock.)
248 Manual of Vital Function Testing Methods
It appears to be the opinion of Magnus-Levy and
Salomon that if hyperpituitarism is uncomplicated by
disorder of other glands of internal secretion (as thy-
roid) there is no increase in the gas exchanges. In
this opinion Falta 59 concurred.
2. Demonstration of Spontaneous and Provocative
Glycosuria as a Functional Test for Hyperpitui-
tarism
It has been known for some time that acromegaly is
often accompanied by a temporary or permanent glyco-
suria. Marie, who first described the disease, called at-
tention to this fact. Borchard, from a study of 176
cases, from the literature found spontaneous glycosuria
reported in 63, and alimentary glycosuria in 8. In 8
cases studied by Falta there was spontaneous or ali-
mentary glycosuria in 5. Glycosuria appears only in
the early stages of the disease, disappearing towards
the end with the beginning of cachexia.
The test for provocative alimentary glycosuria
should be made in every case of suspected hyper-
pituitarism.
It will be unnecessary here to repeat the details of
technique of the various tests for provocative gly-
cosuria, since they have been fully dealt with under a
previous chapter. The four tests there described are
(1) the Cane Sugar Test, (2) the Glucose Test, (3)
the Levulose Test, (4) the Galactose Test.
The Glucose test has been more frequently used than
the others in testing the carbohydrate powers in cases
of suspected hyperpituitarism.
59 Erkrankungen der BlutdrUsen, Berl., 1913, p. 213.
The Ductless Glands 249
B. States of Hypopituitarism. — This state is typ-
ically represented in the so-called hypophyseal dys-
trophy of Frohlich, or dystrophia adiposo-genitalis.
Frohlich,60 in 1901, first emphasized the connection
between destructive tumors of the hypophysis and
the occurrence of the syndrome which bears his name
and whose chief characteristics are a rapidly developing
adiposity, infantilism of the genitalia and myxedem-
atous degeneration of the subcutaneous tissues. Many
authors have since reported cases.
The opposite conditions with respect to gas ex-
changes and glycosuria obtain in hypopituitarism as
compared with hyperpituitarism. In hypopituitarism,
therefore, the same functional tests are applied as were
discussed above under acromegaly. The results, how-
ever, will be the opposite. The gas exchanges will be
diminished and glycosuria, both spontaneous and pro-
vocative, will be negative.
""Wien. klin. Rundschau, 1901, XLVII, 48.
£<•£
INDEX
PAGE
Abderhalden's test in hyperthyroidism 230
Abderhalden's method and liver function 46
Aceto-nitril test of hyperthyroidism 221
Adrinalinemia and hyperadrenalism 242
Adrenalin glycosuria and hyperadrenalism 243
Adrenalin-mydriasis test of hyperthyroidism 218
Albarran's method of testing kidney function 72
Aminoaciduria, experimental provocative 33
Aminoaciduria and liver function 33
Ammonia nitrogen, estimation in urine (formalin
method) 31
Ammoniuria experimental provocative 32
Antitoxic liver function 36
Biliary liver function 47
Bilirubinuria and liver function 51
Blood coagulation and liver function 41
Blood studies and renal function 90
Cammidge reaction 162
Cane sugar test of liver function 15
Cardiac efficiency factor of Tigerstedt 193
Cardiac overload factor of Stone 197
Cardiac reflex and heart function 187
Cardiac strength, cardiac weakness ratio 194
Cell nuclei test of pancreatic function 148
Claude Baudouin, Porak test of hyperthyroidism. . . 213
Coagulation time and renal function 104
Coagulation time, test for 41
Complement-fixation in adrenal disease 244
Complement- fixation test of hyperthyroidism 226
251
252 Index
PAGE
Cryoscopy of blood and renal function 104
Cryoscopy of urine and renal function 86
Diastase in feces and pancreatic function 158
Diastase in urine and kidney function 85
Diuretic drug tests of kidney function 75
Ehrlich's urobilinogen test 51
Electric conductivity of urine and kidney function. . 89
i
Fat digestion and pancreatic function 150
Ferment identification and pancreatic function 153
Fibrinogen test of liver function 42
Fibrinolysis test of liver function 43
Folin-Denis method of estimating incoagulable nitro-
gen in blood 99
Galactose test of liver function 18
Gas exchange and hyperpituitarism 247
Ghedini's test of liver function 46
Glucose test of liver function 16
Glutoid capsule test of pancreatic function 148
Glycosuria and hyperpituitarism 248
Glycosuria and pancreatic function 166
Goodpasture's test of liver function 43
Graupner's test of heart function 175
Gymnastic test of heart function 184
Heart function tests 1 68
Herz' test of heart function 181
Hippuric acid test of renal function 108
Hohlveg-Meyer method of estimating incoagulable
nitrogen in blood 99
Hunt's test of hyperthyroidism 221
Hyperadrenalism 241
Hyperpituitarism 246
Hyperthyroidism^ experimental 219
Hyperthyroidism tests for 208
Index 253
PAGE
Hypoadrenalism 238
Hypopituitarism 249
Hypophysis cerebri 244
Hypophysis test of hyperthyroidism 213
Hypothyroidism 232
Incoagulable blood nitrogen and renal function 90
Incoagulable nitrogen in blood, estimation of 99
Indicanuria and liver function 38
Indigo carmine test of renal function 114
Katzenstein's test of heart function 181
Kidney function tests 64
Kjeldahl's nitrogen method 28
v. Koranyi's test of renal function 86
Lactose test of renal function 108
Levulose test of liver function 16
Lipase estimation in blood 45
Lipase in blood and liver function 44
Lipase in feces 160
Liver function tests 13
Loewi's test of hyperthyroidism 218
Loewi's pupillary test 165
Lowenhart's lipase estimation method 45
Marshall's method of urea estimation in blood 94
Marshall's method of urea estimation in urine 24
Mendelsohn's test of heart function 179
Metabolism test of hyperthyroidism 224
Methylene blue test of liver function 36
Methylene blue test of renal function Ill
Morris' method of estimating incoagulable nitrogen
in blood 99
Nitrogen coefficient and liver function 22
Nitrogen estimation in urine (Kjeldahl's method) . . 28
Nitrogen in urine and kidney function 80
254 Index
PAGE
Pancreatic function tests 142
Parathyroid glands 234
Phenolsulphonephthalein test of renal function 115
Phenoltetrachlorphthalein test of liver function.... 55
Phloridzin test of renal function 107
Polyuria experimental, and kidney function 72
Potassium iodide test of renal function 105
Protein digestion tests of pancreatic function 146
Residual nitrogen and liver function 34
Rest nitrogen in blood and renal function 90
Roche's test of liver function 37
Rontgenoscopy and cardiac function 199
Rowntree, Geraghty test of renal function 115
Rowntree, Horwitz, Bloomfield test of liver function 55
Russian test of heart function 185
Sahli's test of pancreatic function 148
Sanguinopoietic liver function 40
Schmidt's test of pancreatic function 148
Schott's test of heart function 185
Selig's test of heart function 173
Sodium chloride elimination and cardiac function. . . 188
Sodium chloride elimination and renal function 76
Sodium chloride estimation 79
Sphygmography and cardiac function 199
Sphygmomanometry and cardiac function 188
Staircase test of heart function 173
Starch digestion and pancreatic function 153
Stone's cardiac overload factor 1 97
Straus-Griinwald test of kidney function 75
Sugar tests and adrenal function 240
Sugar tests and liver function 14
Suprarenal glands 237
Thymus gland 237
Thyroid gland 207
Tigerstedt's cardiac efficiency factor 193
Index 255
PAGE
Trypsin estimate in stomach contents 156
Trypsin estimate in stools 154
Urea in blood and renal function 90
Urea elimination and kidney function 82
Urea elimination and liver function 22
Urea estimation in urine (Marshall's method) 24
Urea provocative test of McCaskey 83
Ureagenetic function tests of liver 20
Urinalysis as criterion of kidney function 70
Urinary toxicity and renal function 89
Urobilinogen test of liver function 51
Urobilinuria and liver function. . •. 51
Urobilinuria, tests for 54
Urochrome and kidney function 84
Venous pressure test of heart function 185
Water tests of kidney function 75
Whipple, Horwitz test of liver function 42
Whipple's lipase test of liver function 44
Work-velocity ratio and cardiac function 188
Wright's coagulation-time method 41
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