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Do WEP awtMEN El Or AGRICULTURE,
BUREAU OF CHEMISTRY—BULLETIN No. 94.

H. W. WILEY, Chief of Bureau.

StUDIES ON APPLES.

1. SToRAGE, RESPIRATION, AND GROWTH.
Il. INsoLUBLE CARBOHYDRATES OR Marc.

Ill. Microscopic anp Macroscopic Examt-
NATIONS OF APPLE STARCH.

BS

W. D. BIGELOW, H. C. GORE, AND B. J. HOWARD,
OF THE BUREAU OF CHEMISTRY.

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WASHINGTON :
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GOVERNMENT OFFICE.

FEUER OF SR ANSMIT TAL

U.S. DEPARTMENT OF AGRICULTURE,
BUREAU OF CHEMISTRY,
Washington D. C., April 12, 1905.

Str: I have the honor to transmit herewith the manuscript of a
report on some studies made in the division of foods and in the micro-
chemical laboratory of this Bureau on the development of apples, espe-
cially under different conditions of storage, and to recommend the
publication of the same as Bulletin No. 94 of the Bureau of Chemistry.

This investigation has been in progress for the past two years, and
some preliminary work was done prior to that time by Mr. J. 5. Burd.
The work was originated, planned, and directed by Mr. W. D. Bige-
low, chief of the division of foods. The analytical and bibliographical
data should be, for the most part, accredited to Mr. H. C. Gore,
though Mr. E. M. Chace and Mr. W. L. Dubois participated in the
analytical work. The third section of the report, comprising the
microscopic studies, photographs of the fruit, and the specific gravity
determinations, is entirely the work of Mr. J. B. Howard, chief of
the microchemical laboratory.

The investigation has been made in collaboration with Messrs.
William A. Taylor and G. Harold Powell, of the office of pomological
investigations, Bureau of Plant Industry, who have selected and pro-
cured the samples and furnished facilities for their transportation and
cold storage. They havealso taken an active interest in planning the
work and discussing its results, and have furnished all of the pomo-
logical data.

Very respectfully,
Jal Me \ivisnsie, Clogs

Hon. JAMES WILSON,

Secretary of Agriculture.

CO Nala N GTS:

Page.

I. Storage, respiration, and growth......-.--- To 5 eh Ge a Ee, Sees 9
Review of work on the ripening and respiration of fruits -......---. 9
hemipening olapples im common storagess2=-2. 2 2.2... 22222222 Se 20
Weseripwon-ot the vanieties-analyzed--= Se. 2 22. f ees ee 20
DWeseriptionandsanalyses olsampless 2545. St eS: 72)
Wiscussionuoisvanahyirealedutas ae eee he eee ee he ie 24
Rhewripeninesohapplesaunicoldistorages: =. Sao dl
RESIS Oley SES: et eer he om ee Ge I oe sh 32

Results with samples taken out of cold storage.....-.---------- 39

The respiration of apples in common and cold storage......-------- 40)
Description of apparatus and methods employed.-....-.--.---.- 40

Resuliscal respiration expemments <2. 2. 24 2b 2 ne Pts Se 42

Teiay ss ve O alae Cae 0) DCS ee eS ie i eee ee 44
Results of consecutive analyses of growing apples..-.---------- 45

Discussion of changes occurring during growth -.....---------- 46

CHIC MoeoleamelGiomet tere srt reese eS vole OS So ee 65

Mie Ain soliiple-carhoimycnrates Or MALC. 22-25 ee Bd gaa ee eet en Re 67
IBCeuIne Oelieserm tte. aan a, Seen tense e osme ote Pe oe 67

PAR SEONG NG ROMIE Ware ee ena eR eS e AGS ia i Sw SSL 67

Comments on the problems mvolvedia 2.05.52. 55- 62-5 - - +e 86

PARAS CoO PLE LINER aot: > yep ee Sean mney aie Se cd i 2 kc. ed 87
auc ULONIGO fe SATIN |) LC Near eee eee se ie MeN eee Ny aoe ND) 87

RRCOULES EOSIN LYSIS 2 -Mt=s Ampere meaner eal ok ae 88

111. Microscopic and macroscopic examinations of apple starch.-.....------. 89
LESSORS TOME GT OCs ei gat Se eS ty > 8) Let RIP Se eee a 89

SHAE CHE SUITE Mees EMI ONS poe he So Mo 0 er eR ee ee 89

Veakening of cell walls and increase of intercellular air... ---- 92

11 Lx STPTST To OPT 0 CRS a eae te pa PINE Regn a en ae ey SS 94

‘| OH SLOWING Pe 8 a Aap ed aahaiy us nc 7B A aie en mR me Oc em 94

Grameen, Steve CI COMbeni eee nis sya eee eS 95
TULA STIG) TST eee ye ane SI al. SR sted eg Re le ERSTE, NS he ae 98

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ILLUSTRATIONS.

PLATES.

Piate I. Fig. 1.—Apple tissue, showing starch in apple cells. Fig. 2.—Apple

Fic.

SINGIN Sn SOE SE Ih Se ES Oe eR eee
Il. Location and gradual disappearance of apple starch—Winter Para-
GISen (HOUR eswl= GO) ere ae te ee ene er ON ee 2
III. Location and gradual disappearance of apple starch—Ben Davis (fig-
IBS cig esc Joe rghit eee reer Re Dena eek ely es os ae a |
IV. Location and gradual disappearance of apple starch—Huntsman

“J

15.

14.

15.

(TE GATIRES AON Se eae ie Ee eles ER ea

VY. Location and gradual disappearance of apple starch—Yellow Trans-

PREC Hes (OCR CSS io), ea rere pee GE rey IS SS SS oes Sa the
TEXT FIGURES.

Chart showing results of analyses of Rhode Island Greening apples in
COMIUMOMESLOTAC CE ayers hs eyed we oe dou oe otek
Chart showing results of analyses of Northern Spy apples in common
EOE LO Crs Semen serene irony va A Nera Re Sec SS Soli SS 2 eke ea
Chart showing results of analyses of Winesap apples in common
SUOEAG Cer ety aia eny seer ar oy eee wee ee a UEP Seok
Chart comparing results of analyses of three varieties of apples in
common storage

Chart showing results of analyses by C. A. Brown, jr., of Baldwin

ap PLCSeta COMMON USCOLAG Cee es ae owe ES eS
Chart showing results of analyses by Kulisch of apples stored at cellar
temperature -..-- Brera tns tener MENG ere EL yo Ie es
Chart showing results of analyses of Ben Davis apples picked August
15, 1902, and kept under varying conditions of storage ....--------
Chart showing results of analyses of Ben Dayis apples picked Sep-
tember 15, 1902, and kept under varying conditions of storage -..--
Chart showing results of analyses of Winesap apples picked August
15, 1902, and kept under varying conditions of storage --...-..----
Chart showing results of analyses of Winesap apples picked Septem-
ber 15, 1902, and kept under varying conditions of storage
emetisedsem respirationvexpeniments 2 -82..4.cc > fos Ses 22s | Se
Chart comparing results of analyses of Ben Davis apples used in res-
piration experiments and kept in cold storage and at cellax tem-
perature

Chart showing chemical changes in summer apples ( Early Strawberry )
during growth

Chart showing chemical changes in summer apples (Bough) during
BAUME Be ten eae ee at es 2 ee eS ee ee

Chart showing chemical changes in summer apples (Yellow Trans-
parent) during growth

Fig. 16.

17;

18.

Ligh

25.

ILLUSTRATIONS.

Chart showing average chemical changes during growth in the three
summer yarieties: 2. o2 2225-4. s- eee Ce AE sae a ee
Chart showing chemical changes in winter apples (Ben Davis) during

Chart showing chemical changes in winter apples (Huntsman) during
prowthys so) tect ions. ee oo 2
Chart showing chemical changes in winter apples (Winter Paradise)
during erowth .....--.------- 33222

. Chart showing average chemical changes during growth of the three

winter varieties :.-....2.2-22. 5.2203 SS eee

Page.

Chart showing chemical changes during growth of summer apples ~

(Early Strawberry )—in grams per apple-....- eLearn eee

. Chart showing chemical changes during growth of summer apples

(Bough )—in grams per apple-..-.----- Jot snhes ket Oe ees

. Chart showing chemical changes during growth of summer apples

(Yellow Transparent)—in grams per apple: —2 22252 asso] - ses eee

. Chart showing average chemical changes during growth of the three

varieties of summer apples—in grams perapple-.--.-.-------.----
Chart showing chemical changes during growth of winter apples ( Ben
Davis)—in grams per apple... 2... 32s ass]

. Chart showing chemical changes during growth of winter apples

(Huntsman )—in grams per apple - 2222 22 Sass eee eee ee
Chart showing chemical changes during growth of winter apples
(Winter Paradise)—in grams per:apple: 22 222222 eee
Chart showing average chemical changes in the three varieties of
winter apples examined—in grams per apple..---..--------------

. Chart showing results of analyses of growing apples by Lindet—in

percentages of total solids.........- ud RG Bek pee ee

. Cross section of apple, showing structure and location of parts... .---

STUDIES ON APPLES.

I. STORAGE, RESPIRATION, AND GROWTH.
REVIEW OF WORK ON THE RIPENING AND RESPIRATION OF FRUITS.

In the following résumé of works on this subject, the review of
Gerber” has been used as a guide to the French memoirs on the
subject.

The earliest source available in the original was the paper of Bérard.?
This author carried on elaborate researches on the ripening and res-
piration of fruits, his results apparently contradicting those of De
Saussure,° whose conclusions were considered not to be sufficiently
well established by experiments, De Saussure having put forth the
idea that green fruits act like leaves toward. the atmosphere surround-
ing them, in light or darkness.

Bérard studied the effect produced by unripe strawberries and many
other unripe fruits on the air surrounding them by keeping them for
twenty-four-hour periods in air in large flasks sealed by mercury.
He found in all cases an increase of carbon dioxid at the expense of
oxygen, and in no case the reverse change. Similar experiments were
tried on fruits still attached to the tree (apricots, peaches, and plums),
with the result that the fruit did not mature, but became withered
and browned, showing apparently that oxygen is necessary for the
ripening of fruits. Experiments are also described in which fruits
were kept for long periods of time in inert gases—carbon dioxid,
hydrogen, or nitrogen, and in vacuo. These experiments were not
considered successful, for it was found that the fruit lost its odor and
acquired a disagreeable taste.

In the second contribution” methods for the analysis of fruits are
presented, and analyses given of many fruits when green and when
ripe, viz, apricots, currants, cherries, plums, peaches, and pears.

De Saussure“ confirmed his previous work, published in 1804, by a
report of new experiments, which differ widely in their results from

“ Annales des sciences naturelles, 1896 (8), 4: 1.

6 Ann. chim. phys., 1821 (2), 16: 152, 225.

¢ Recherches chimiques sur la yégétation, Paris, 1804.
@Ibid., p. 225.

¢Ann. chim. phys., 1821 (2), 19: 1438, 225.

10 STUDIES ON APPLES.

those of Bérard.“ The air in which the fruit was kept was examined
at shorter intervals than in the experiments of Bérard, and this may
account for the disagreement of their results, though De Saussure
believed that the glass containers used by Bérard were too small, so
that crowding or overheating may have taken place.

De Saussure reiterated that green (unripe) fruits act on the air like
leaves, differing only in intensity of action, which is less in the ease
of fruits. During the night oxygen disappeared and was replaced by
carbon dioxid, which was partly absorbed by the fruit, absorption
being less in free than in confined air. The fruits consumed, volume
for volume, more oxygen in the dark when they were very green than
when they approached maturity. On exposure to sunlight they gave
off part or all of the oxygen of the carbon dioxid which they had
absorbed in darkness, and used up all the carbon dioxid from the
atmosphere in which they were inclosed. Green fruits could even
remove the carbon dioxid from an atmosphere artificially charged with
several per cent of the gas.

Couverchel’ contributed two papers of much interest, in which the
ideas of several early writers on the subject are given. These ideas
were founded largely on theory rather than on experiment. From
the early work of Sennebier are quoted the following items of interest:

The fruits which have yellowed in ripening are more succulent than green fruits,
are nearer decay, are more gummy than resinous, and are more soluble in water.
Perhaps the phlogiston may have less energy because it is more attenuated, the fiber
loosens, the mass of the fruit increases, etc.

Sennebier supposed that the fruit suffered a loss in phlogiston
(corresponding to a gain in oxygen). Ina later work he says: ** The
taste of fruits, at first bitter, becomes acid, then sweet. The astrin-
gent principle which appears before the formation of vegetable acid
changes to sugar by oxidizing.” This writer considered that galliec
acid was the ** unfinished” vegetable acid, completed by the oxygen
which it appropriated. ** It is certain,” he adds, *‘ that the acids oxi-
dize more and more; for example, citric acid in green grapes passes,
by oxidation, into tartaric acid.” This idea appears to have been cur-
rent among the chemists of the time, since it was specifically denied
by Frémy in 1544 in the case of grapes. (See Frémy, p. 11.)

Other early writers are quoted by Couverchel, showing at what an
early date the functions and fate of the changing constituents of
growing fruits were studied and how varied were the notions con-
cerning them. For example, Lamarck and Decandolle considered
that oxygen arising from the decomposition of carbon dioxid acted
on the mucilage of the fruit, changing it to sugar. Berthollet thought

@ Loc. cit.
’J. pharm. chim., 1821 (2), 7: 249; and Ann. chim. phys., 1831 (2), 46: 147.

STORAGE, RESPIRATION, AND GROWTH. jul

that the acid flavor of green fruits was produced by oxygen, which
was only very teebly held in combination in the fruit.

This first paper by Couverchel gives results of the analysis of juice
of apricots and grapes at different stages of growth, determinations of
density, acidity, dry weight (in vacuo), gum, sugars (weighed as dried
sirup), ash, and soluble ash.

In the second contribution, in 1831,¢ further quotations from early
writings are given. Of much interest is that from the work of
Ingenhouz:?

All fruits, day and night, exhale a mephitic air, and possess the power to render
the surrounding air unwholesome. I have been very much astonished to find a
poison in fruits which are so much eaten, the more since the finest fruits possess
this power in high degree. * * *

Couverchel differed from his predecessors, believing the presence
of air to be only incidental in the ripening of fruits, and that ripening
@oes on by action of principles contained in the fruit. It may be, he
says, that the sap becomes acidified in its passage through the young
branches to the ovary by reason of the decomposition of water and
absorption of the oxygen. The acids so formed may act on the gela-
tin and give rise to sugars, sugars being considered as intermediate
substances between the mucilages and the vegetable acids, containing
more oxygen than the former and less than the latter.

In 1844 Frémy¢ discussed the work of Bérard on the respiration of
fruits. He confirmed the observation that fruits consume oxygen,
giving off carbon dioxid, and carried out an experiment similar to
those of Bérard. In Frémy’s work the unripe fruit, attached to the
tree instead of being kept in a closed jar as in the experiments of
Bérard, was coated with varnish, in this way stopping the normal
respiratory changes. (Growth was found to cease, as in the experi-
ments of Bérard.

The air contained in green and in ripe fruits was analyzed by boiling
slices of the fruit in brine and analyzing the air which separated.
The presence of a ferment causing the respiratory changes was sug-
gested, although a pear, after it was ground, gave off no carbon dioxid
gas, whereas before grinding carbon dioxid was freely evolved.

The old idea that the acids change in ripening (see p. 10) was refuted
in the case of grapes, by the recognition of tartaric acid in the very
young fruit.

Unripe fruit was profoundly altered by soaking in dilute sodium
carbonate, but no conclusions are drawn. Objection is made to the
statement that starch can form the sugar found in fruit, since the sugar

«Loe. cit.
bVersuche mit Pflanzen, 1786, 1: 64; 2: 61, 221, through Gerber, loc. cit.
¢Compt. rend., 1844, 19: 784.

12 STUDIES ON APPLES.

(glucose) formed from starch by the action of acids was found by Biot
to be not the same as the sugar in fruits.

Some of the ideas of Frémy were vigorously combated by Cou-
verchel.“ ,

A-very valuable contribution to the subject of the analysis of fruits
by Buignet,’ which appeared in 1860, is of interest because of the
methods given for the determination of many constituents of the straw-
berry. Water, acid, sugars, fat, soluble and insoluble protein, mare,
nitrogen-free parenchyma, pectin, odoriferous principles, coloring
matters, and ash are worked out. Strawberries containing the least
water are said to contain the most sucrose, and this is explained on
the hypothesis that the acid and sucrose are contained in different cells,
and in the presence of much water these diffuse more readily than in
drier berries, a more rapid inversion ensuing. Sucrose is believed to
be the initial sugar formed. Results for many varieties of strawber-
ries are tabulated. A later paper’ gives results concerning sugars
in fruit in the form of a summary of 14 conclusions, some of which
are as follows: Sucrose is found in many fruits, becoming inverted in
the ripening, sometimes completely, e. g@., in grapes, currants, and
figs: sometimes partially, e. g., in bananas, apricots, peaches, plums,
apples, and pears. No relation exists between sucrose and the acidity
of fruits, but the sugar is probably inverted by a nitrogenous ferment.
This was proved by showing that, after precipitating with alcohol, the
sucrose remained unchanged; that after neutralizing an original sample
of juice with calcium carbonate, the inversion still went on; and that
in bananas, which have no free acid, starch is rapidly changed to
sucrose. Invert sugar prevents the crystallization of sucrose, but the
latter sugar was isolated in crystals from a number of fruits by a pro-
cedure which is described. Starch was not found in fruits (except the
banana), but a tannin-like, astringent principle is described as oceur-
ring in unripe fruits, which decolors an iodin solution, yielding a pre-
cipitate which, when treated with acid, yields a glucose sugar, the same
as that formed from nut galls. The starch and tannin found in the
banana are said to disappear simultaneously, giving rise to cane sugar.
Finally we are told that a difference exists between sugars of fruits
according as they are produced under action of vegetable forces, or
Without it, e. g., bananas contain more sucrose and less invert sugar
when allowed to ripen on the tree than if ripened after picking.

Berthelot and Buignet” worked on the ripening of oranges. Two
samples of green oranges were studied. Several fruits from each

“Compt. rend., 1844, 19: 1) 14.

’J. pharm. chim., 1859 (3), 86: 81-111 and 170-198; résumé in Compt. rend.,
1859, 49: 276-278.

¢Compt. rend., 1860, 41: 894.

@[bid.: 1094.

STORAGE, RESPIRATION, AND GROWTH. 13

sample were analyzed, the remainder being kept for some time in a dry
place at an even temperature, when another analysis was made. No
figures were given, and changes in the sugar only are reported, but
these changes were remarkable, because, while the invert sugar

remained nearly constant, the sucrose increased, calculated either on
the basis of juice or on that of soluble solids. This was considered as
a very curious change to take place in an acid fruit. It is the more
notable when it is remembered that the green fruit contains no starch.

Cahours“” noted that ripe oranges evolved carbon dioxid, consuming
oxygen at the same time, and that when all the oxygen was used up
the carbon dioxid evolution still continued. It also continued in an
atmosphere of nitrogen.

Chatin’ considered that the carbon dioxid evolved by ripening
fruits resulted from the oxidation of the tannin, since, as the fruit
ripened, tannin disappeared.

Frémy° called attention to the three periods in the life Wirore of a
fruit—growth, ripening, and decay. |

Corewinder” gave an analysis of ripe bananas, and later’ studied
bananas from ripeness to decay during a period of eighteen days, dur-
ing which sugars were determined at 10 different times. The sucrose
fell from 15.90 to 2.84 per cent; total sugar from 21.80 to 14.68 per
cent; and invert sugar increased from 3.90 to 11.84 per cent.

Beyer’ studied the growth of gooseberries, analyzing them every
few days from the time when the fruit was very small until it was
ripe. The complete analysis of the fruit at each picking is reported,
including sugar, acid, protein, ash, fat, and nitrogen-free residue. No
determinations of sucrose were made.

A notice of work done by Pasteur in 1866 is found in a review of
literature on the ripening of grapes by Fitz (p. 14). Pasteur cast
doubt upon the idea that the acid in ripening grapes gave rise to sugar,
by discovering that in sour varieties the acid actually increased during
ripening.

Petit? published discussions on the ripening of grapes. He con-
sidered that during ripening the sugar came from the cellulose in the
fruit, the cellulose first changing to acid by oxidation, and the acid
then becoming sugar with the evolution of carbon dioxid. Both
sucrose and reducing sugar were found .in the leaves of the grape,
cherry, and peach, but in the grape itself only reducing sugar was
present.

“Compt. rend., 1864, 58: 495 and 653.

bTbid., 1864, 58: 576.

¢Ibid., p. 656.

@Tbid., 1863, 57: 781.

é Ann. agron., 1876, 2: 429.

fLandw. Versuchs-Stat., 1865, 7: 355.

gCompt. rend., 1869, 69: 760; and ibid., 1873, 77: 944.

14 STUDIES ON APPLES.

Dupré,” however, noted a positive increase in acid in grapes during
ripening, disproving the conclusion of Petit.

Neubauer’ studied the ripening of grapes. The complete analysis
of two varieties of grapes at successive stages of growth is presented.
No satisfactory explanation could be given for the great increase of
sugar that took place on ripening.

Famintzin® published the results of a study of the ripening of grapes,
which is of much interest because it explains the hitherto unknown
source of the sugars which appear in the grape on ripening. He used
both chemical and microscopical methods. ‘Twelve analyses of a
variety of growing grapes are given and the results discussed. The
unripe grape contains no starch, but the stems of the fruit are full of
starch, which disappears as the grape ripens. Tannin was present
throughout the very young fruit, but later occurs only in the outer
laver of the fruit pulp and in the embryo.

This work was confirmed by Hilger,’ who followed the changes of
water, ash, sugar, and acid content in the leaves and fruit of the grape
during growth. The stems of the fruit were found to be full of starch,
which disappeared when the rapid increase of sugar began. The
increase of sugar for a period of six days at the ripening time was
from 3.87 to 7.70 per cent in the case of one variety, and from 5.33
to 7.71 per cent in another, during which time the starch in the stems
disappeared.

Mercadante’ believed that the sugars were formed at the expense of
the gummy matters in the fruit, and also from acids, the fruit being
thus considered to be the seat of the formation of sugar. Macagno’
showed that sugar formation takes place in the leaves, not in the fruit
of the grape.

Saintpierre and Magnien’ held a similar view. These-authors first
give a review of the literature on fruit ripening. They found that
ripening fruits give off carbon dioxid in light or darkness, that they
absorb or give off water according as they are kept in a moist or dry
place, and that acid and sugar come to the fruit through the stem.
The acid is believed to be used up in respiration, the sugar meanwhile
concentrating in the fruit until in its turn it furnishes the carbon dioxid
for combustion.

Pollacci,” studying the after ripening of grapes, found that the sugar
increased and the acid decreased in grapes stored after picking.

“Weinlaube, 1870, p. 274, through A. Fitz, Ann. Oenol. 2: 241.

/Landw. Versuchs-Stat., 1869, 11: 416.

¢ Ann. Oenol., 1871, 2: 242.

7 Landw. Versuchs-Stat., 1874, 17: 245.

‘Gazzetta chimica Italiana, 1875, 5th ser., p. 125, through Gerber, loc. cit.
/ Compt. rend., 1877, 85: 763, 810, through Gerber, loc. cit.

?Ann. agron., 1878, 4: 161.

"Tbid., 1877, 3: 629.

STORAGE, RESPIRATION, AND GROWTH. 15

Lechartier and Bellamy” published two papers which give results
of experiments with apples stored in sealed vessels. Large quanti-
ties of carbon dioxid gas were evolved during the long periods of time
in which the fruit was under observation. Very considerable quanti-
ties of alcohol were found, and the presence of yeast cells in the par-
enchyma cells of the sound apple is noted. These observations have
never been confirmed, neither alcohol nor yeast celis occurring in an
apple whose skin is intact; however, so far as is known no work has
been done since on apples held in sealed chambers.

Pfeiffer’? gives a review of the chemical study of fruits. In the
same paper are given the results and a discussion of the study of the
orowth of apples and pears. Crude fiber, ash, protein, sugar, acid,
water, and pectin and dextrin, are all shown to increase during growth,
the constituents of the apple increasing more rapidly in the three
varieties studied than those of the pear, of which two varieties were
used.

Mach¢ carried on work on the growth and ripening of grapes and
later” extended the work to other fruits, viz, apples, pears, mulber-
ries, strawberries, red and black currants, cherries, and peaches.
With grapes he found that the fruit grows rapidly in size until it
begins to color, then grows more slowly. Sugar develops slowly at
first, but after the coloring of the grape, very rapidly. The percent-
age of acid remains about constant till ripening begins, when it falls.
Tannin is present in the largest quantity at first. Starch is present in
the growing shoots in the leaves and in the grape stems. It disap-
pears, however, as the fruit ripens. Unfortunately, with the other
fruits studied, no determinations of sucrose are made.

Ricciardi’ followed the ripening of bananas, the starch being found
to give rise to sucrose. The author agreed with Buignet/’ that in the
banana ripened on the tree there is no invert sugar, but this point is
not brought out in the analysis given. No alcohol was found in the
over-ripe fruit.

The work of De Luca is used as a basis for further work by Gerber’
on the ripening of olives. In De Luea’s paper” a table showing the
growth of the olive from June to February is given. Thirty-four
examinations were made on different dates to determine the average
weight and density. Picked olives yielded more oil when they were
allowed to stand in oxygen, or in air in daylight, than when analyzed
at once or after they were kept in carbon dioxid. The presence of a
bitter principle was noted in green olives, removable by prolonged
soaking in water. Mannite was found in considerable quantity in the
fruit and leaves of the olive tree and was isolated by extracting with

«Compt. rend., 1869, 69: 356, 466. €Compt. rend., 1882, 96: 393.
6 Ann. Oenol., 1876, 5: 271. Oc elte
¢Ibid., 1877, 6: 409. g Compt. rend., 1897, 125: 658.

@ Ibid., 1879, 8: 46. h Tbid., 1861, 53: 380.

16 STUDIES ON APPLES.

hot alcohol. It was believed by the author to be essential for the
production of oil, though this could not be proved.

In a second contribution” the results of successive analyses of the
pulp of the olives for oil are given. The percentage increased by large
increments, but irregularly (probably because of the small sample used—
three fruits) with the growth. The presence of a chlorophylllike
body in the pulp was observed, becoming less in quantity as the fruit
approached ripeness. The proportion of pulp to pit is given.

The third paper’ discusses mannite. Determinations of mannite in
the fruit during growth are given. It was found to occur in widely
varying amounts up to 1.54 per cent of the dry matter. The mannite
and the chlorophyll-like body disappeared as the fruit ripened and
seemed to be closely connected with the formation of oil.

Upon the results of the above work, and from his own experiments
on the respiration of olives, Gerber ¢ considers that he has established
direct proof of the transformation of sugar-like bodies, especially man-

\

nite, into oils. When the respiratory quotient O “(ratio by volume

of carbon dioxid produced to oxygen consumed) is greater than unity,
and no acids are disappearing to furnish the extra carbon dioxid by
breaking down (which he believes takes place in acid fruits), mannite
is believed to be passing over into olive oil with the evolution of water
and carbon dioxid.

Roussille% presented consecutive analyses of the leaves and fruit of
the olive tree during the growing season. He determined that the
oil did not undergo migration from the leaves to the fruit, but was
formed in the fruit.

Funaro*’ gives a short review of previous work, noting that of De
Luca (see p. 15) and the paper of Harz,’ and concludes that the oil is
formed by special secreting cells containing an unknown material.
The author then gives his results in tabular form, which represent
the growth of the olive, the weight and dimensions of the fruit, the
moisture content of flesh and stone, and the total ether extract in flesh
and stone.

Mannite was found in small quantities in fruits and leaves. Its
presence in fruits at the end of the growing season contradicted the
conclusions of De Luca, who states that it disappears as the fruit
increases in oil. According to Funaro this indicates that mannite prob-
ably has nothing to do with the formation of oil.

This view is in accordance with conclusions reached by Hartwich
and Uhlmann.’ These authors considered @lucose to be the material

“Compt. rend., 1862, 55: 470. ¢ Landw. Versuchs-Stat., 1880, 25: 52.
oT bid., 56: 506. J Ann. prak. Pharm., 19: 161.

¢Tbid., 125: 658. 9 Arch. Pharm., 1902, 240: 471.

d Ann. agron., 1878, 4: 230.

LL

STORAGE, RESPIRATION, AND GROWTH. af

from which oil is formed, and affirm that mannite does not occur in the
fruit of the olive.

Keim“ studied changes in the composition of the flesh of cherries at
intervals of from seven to ten days. As the fruit ripened the percent-
age of water decreased, and both acid and sugar increased with total
dry matter. In the early stages of growth, citric, malic, and succinic
acids were present, but nine days before ripeness the succinic acid
disappeared. Dextrose and jievulose are always present, while inosite,
which at first was present in appreciable quantity, diminished to a mere
trace. Sucrose was present in smallamount. No starch was found in
the fruit save in the outer green layer of the very young fruit, but it
occurred in the parenchyma cells of the fruit stem in increasing
amounts as the fruit ripened.

Kulisch’ presented a valuable discussion of the changes in the con-
stituents of ripening fruits, giving also an account of.a study of the
changes which a variety of apples underwent on storing at cellar tem-
perature. The fruit from two trees of the same variety in the same
orchard were employed and analyzed separately, so that the results
have further interest in showing how the composition of fruit from
different trees of the same variety, grown under the same cultural
conditions, may vary. (See p. 31 and fig. 6, p. 30.)

_ The growth of a single variety of apples was thoroughly studied by

Lindet,° who examined the fruit at fifteen-day intervals through the
growing season (see p. 24 and fig. 29, p. 63). He established the
fact that the acid content gradually became less, and that the starch
increased by degrees until the fruit began to ripen, when it decreased
eradually. Sucrose and invert sugar increased steadily up to the last
analysis. A portion of each sample received was left in darkness and
analyzed at intervals. The starch decreased gradually to about 0.8
per cent, sucrose and invert sugar increased, while total carbohydrates
fell, allowance being made for loss of moisture by the fruit on being
kept. These changes are recognized as taking place: (1) The change
of starch into sucrose; (2) the inversion of sucrose; and (8) the con-
sumption of invert sugar in respiration. The change of starch into
sucrose is said to be a chemical phenomenon whose mechanism escapes
us. ‘The localization and disappearance of starch in apples.is described,
and a description of the starch grains is given.

C. A. Browne, jr.,” noted the starch and sugar changes of a sample
of Baldwin apples, picked green, obtaining results similar to those of
Lindet and those obtained in the Bureau of Chemistry. Otto’ also

«7ts. anal. Chem., 1892, 30: 401, abstr. in Agr. Science, 1892, 6: 387.
bLandw. Jahrb., 1892, 21: 871.

¢Ann. agron., 1894, 20: 5-20.

@Penn. State Dept. Agr., Bul. 58.

eCentrbl. agrikulturchem. (Biedermann), 1902, 31: t07.

27981—Bul. 9405-2

18 STUDIES ON APPLES.

studied the growth and ripening of a variety of apples, noting that
the percentage of water, acid, and starch decreased more or less
uniformly, while dry matter, extract, sucrose, and reducing sugar
increased. In another contribution by Otto” a study of the changes
which go on in common storage is presented. Samples of eight varie-
ties of apples were analyzed before and after cellar storage. A gradual
diminution of all constituents was found except in two cases in which,
while the acid and starch decreased, the total sugar showed a slight
increase.

In further work done by Otto? it was found that the starch in ripe
apples when they were allowed to sweat in piles was entirely converted
into sugar in two or three weeks, the fruit thus becoming more valua-
ble for cider making.

The contribution of Gerber“ to the subject of the ripening of fruits
is of considerable importance. The work is of some length, occupying
280 pages. First, the work of various writers on respiration and car-
bohydrate, acid, and tannin changes in growing and ripening fruits is
reviewed, together with the resulting hypotheses regarding these
functions. The author then describes the methods and apparatus
employed by him in the researches which are described. The relative
intensity of respiration and the value of the respiratory quotient,
CO, RON sath ep : yey, Hos:
“©, > are the chief criteria used. ‘The fruit (a single fruit in case of
apples) is held in a sealed glass container, and the air of the chamber
is analyzed aftera time. The air is then renewed and the experiment
repeated, the temperature being held constant. In this way the inten-
sity of respiration and the respiratory quotient—i. e., the ratio of
carbon dioxid given off by the fruit to the oxygen consumed—may be
observed.

The method is first employed with apples. It is noted that unripe
apples breathe much more rapidly than ripe apples. The respiratory

\

quotient, ~¢ *, is found to be greater than unity in the case of growing

apples, but it is considered that it is impossible to determine to what
constituent this fact is due, because, although the constituent, malic
acid—which is suspected of furnishing the extra carbon dioxid—
decreases relatively to the other constituents (though increasing in
absolute quantity) during growth, it is impossible to determine whether
or not the sap which is supplied to the apple varies in composition
during the period of growth. More definite results are considered to
be obtained with apples during the period of after-ripening—1. e., ripen-
ing after picking—because here no sap is added to the fruit. Apples
“Centrbl. agrikulturchem, (Biedermann), 1902, 31: p. 104.
bLandw. Versuchs-Stat., 1902, 56: 427.

¢ Loc. cit. (see p. 9).

STORAGE, RESPIRATION, AND GROWTH. IL,

were examined during this period. The influence of temperature,
quantity of acid present in the fruit, variety of apple, and the effect
of quartering the apples on the intensity of respiration and on the
respiratory quotient are described. Apples respire more rapidly at
30° and at 33° than at 18° C. Apples relatively high in acid respire
more rapidly and have a higher respiratory quotient than apples low
inacid. The influence of cutting the apple into quarters is to increase
the intensity of respiration and’ respiratory quotient.

Similar researches were carried out with grapes, citrus fruits,
almonds, peaches, plums, and apricots. The existence of a respira-
tory quotient greater than unity due to the combustion of acids is
considered by the author to be established, and numerous conclusions
concerning the effect of various factors on the function are given.

Work with molds is described, the respiratory quotient of these
organisms growing on different media being determined, and the
results are believed to furnish direct support to the conclusions
which the author has drawn from his work with fruits.

Other fruits, Japanese persimmons (Diospyrus khaki zend)1), bananas,
sorbes (Sorbus domestica), and medlars (Jlespilus germanica) were now
examined by the method of the author. In persimmons tannin is
believed to disappear by direct combustion, with no formation of
sugar, this conclusion, however, depending on the results of an analy-
sis of a single persimmon.

The conclusion reached with the acid fruits, that the combustion of
the acids is reponsible for the excess of carbon dioxid given off over
oxygen consumed, possesses a certain probability merely because of the
uniformity of the results obtained with a wide range of varieties of acid
fruit. Gerber’s work is valuable as sugeesting certain auxiliary work
that may be carried on in connection with the chemicai work necessary
to the study of the ripening of fruit. His theories should be considered
as hypotheses which must be confirmed by suitable scientific methods
before they can be seriously regarded. Such data should be consid-
ered as indications of facts only. The number of individual speci-
mens in the samples examined and the methods employed do not
warrant that any greater weight be given to the data. At the same
time it is shown that the examination of the respiratory changes in
fruits is easily made, and that fruits of different varieties and different
degrees of maturity vary materially in the intensity of their respira-
tion. The method would probably be of value in supplementing
chemical work on growing or ripening fruits.

From the above review it will be recognized how varied are the
conclusions reached by workers with ripening fruits. With better
chemical knowledge, particularly concerning sugars and starch, satis-
factory agreements are found between the work of various modern
authors working with apples and grapes. It must be considered that

20) STUDIES ON APPLES.

sach variety of fruit presents a different set of relations between its
constituents, and that each fruit contains a variety of undetermined
substances in considerable amount which make changes between
known constituents difficult to demonstrate. No general conclusions
can therefore be reached until each kind of fruit is worked with as
far as possible through the stages of growth and ripening, and only
as the undetermined material in fruits is gradually worked out will
the chemistry of fruits become clear.

THE RIPENING OF APPLES IN COMMON STORAGE.

In the summer of 1901 the preliminary work on the ripening of
fruit was begun. Eight varieties of apples were selected, and each one
was examined several times, the chief object being the study of meth-
ods of analysis. No consecutive data were obtained, but the experi-
ence gained was valuable in beginning the work in the following year.
This preliminary work on the subject was done principally by Mr.
J. 5. Burd.

DESCRIPTION OF THE VARIETIES ANALYZED.

The following descriptions of the varieties of apples employed in
the investigation is furnished by the pomologist of the Bureau of
Plant Industry:

BEN DAVIS.

Fruit medium to large, yellowish, splashed and striped with red. Flesh somewhat
tough and lacking in juice; flavor mild subacid. A winter variety of excellent keep-
ing quality.

BOUGH (Synonym Sweet Bough).

Fruit above medium in size, pale greenish yellow. Flesh very tender, with a rich,
sweet flavor. An early summer variety ripening overa period of two or three weeks.
Keeps but a few days under ordinary conditions after being picked from the tree.

EARLY STRAWBERRY.

A small roundish apple striped and stained with bright and dark red. Flesh
tender, with a sprightly, brisk, subacid flavor. A midsummer variety in season for
several weeks.

HUNTSMAN (synonym Huntsman Favorite).

Fruit large, pale vellow, sometimes shaded with pale red or deep yellow in the
sun. Flesh rather coarse, tender, of a mild, rich, subacid flavor. A winter variety
of fairly good keeping quality.

NORTHERN SPY.

Fruit large, greenish yellow, covered with light and dark stripes of purplish red.

Mesh tender, with a fine, sprightly subacid flavor. A winter variety of good keep-

ing quality under favorable conditions, but quite subject to decay, if roughly handled, -
on account of thin skin and tender flesh.

STORAGE, RESPIRATION, AND GROWTH. 21

RHODE ISLAND (synonym Rhode Island Greening) .

Fruit medium to large, dark green, becoming greenish yellow when ripe. Flesh
tender, crisp, with a rich, acid flavor. An early winter variety of fair keeping
quality, quite subject to scald after midwinter, except under very favorable condi-
tions.

' WINESAP.

Fruit medium in size, of a fine dark red color. Flesh firm, crisp, with a rich,
subacid flavor. A winter variety of excellent keeping quality, except for being
somewhat subject to scald in late winter and spring.

/ WINTER PARADISE.

Fruit rather large, dull green, with a brownish blush or faint striping. Flesh fine-
grained, sweet, and sprightly, with a distinctive and marked aroma. A winter
apple of good keeping quality.

YELLOW TRANSPARENT.

Fruit medium in size, clear greenish yellow in color.- Flesh very tender; flavor
sprightly, quite acid; one of the earliest of the summer apples. Ripens over a brief
season and keeps but a short time.

DESCRIPTION AND ANALYSES OF SAMPLES.

In the summer of 1902 the study was again taken up, four varieties
being selected from three localities. The description of the samples
of apples studied and such cultural data as were obtained are given in
Table I. The analyses of the apples made from time to time while
the samples were held in common storage are given in Table li. These
results were also calculated to the basis of total solids and platted on
graphic charts (figs. 1-4 and fig. 8), the actual changes in composition
during the growth and development of the fruit being illustrated much
more clearly in this way than by means of the numerical tables of
analyses. In all of these charts the ordinates give the percentage of
the various constituents and the abscisse the dates on which the exami-
nations were made.

“

ON APPLES.

STUDIES

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24 STUDIES ON APPLES.

Discussion oF ANALYTICAL Data.

By referring to the graphic chart of Rhode Island Greening apples
(fig. 1) it will be noted that the examination made on August 25 gave
a sucrose content of 13 per cent on the basis of total solids. The con-
tent of sucrose increased steadily until about November 7, when the
maximum was reached. During this time the content of starch had
decreased, and the curve representing the decrease of starch is almost
the reverse of the curve representing the increase of sucrose. ‘The
sucrose reached the maximum on the date mentioned, November 7,
after which it rapidly decreased, and the date of the disappearance of
the starch was almost, if not quite, coincident with that of the maxi-
mum content of sucrose.

The entire crop of apples was picked between October 6 and Novem-
ber 7, and unfortunately during that period, owing to the congested
condition of the railroads, it was impossible to obtain samples of fruit,
and the changes of composition taking place just at the time of ripen-
ing were not studied in as. great detail as had been planned. It is
probable that after picking the changes of composition may have pro-
gressed more rapidly than would have been the case if they could have
been left on the tree.

The sum of the starch and sucrose (starch calculated as sucrose) is
shown to decrease up to the point where the starch entirely disap-
peared. After this time the sucrose content decreases much more
rapidly. The content of invert sugar increases from the earliest
examination throughout the experiment. The curve representing the
content of invert sugar is approximately the reverse of the curve
representing first the decrease of starch and sucrose, and later the
decrease of sucrose.

It will also be noted on the chart (fig. 1) that the total content of
sugar (calculated as invert sugar) increased from the first examination
to the date of the disappearance of the starch. After this date the
curve representing the total sugar as invert merges with the curve
representing the total carbohydrates as invert sugar. This latter
curve does not include the cellulose. In the Rhode Island Greening
apples the total carbohydrate content decreased to some extent after
the disappearance of the starch. In the cases of the other varieties of
apples previously mentioned, and some varieties to be described later,
the percentage of total carbohydrates did not materially change
during the later stages of the growth and development of the fruit.

At the same time a slight increase of carbohydrates expressed in
terms of total solids seemed to occur during the growth of the apple
before its separation from the tree, and a decrease occurred after
picking, especially during long storage. Up to the point of the
maximum starch content the percentage of total carbohydrates
increases rapidly. This point is discussed in greater detail subse-

25

STORAGE, RESPIRATION, AND GROWTH.

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Fia@. 1.—Chart showing results of analyses of Rhode Island Greening apples in common storage at

room temperature (total solids basis)

26 STUDIES ON APPLES.

SRR EERE REECE BSTeDoess se
BISISETSIT ISte (etc Ie Ap ieee bela
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oS oe Ts i a
a Se / FS [

BREIERY, BERES
a a et EME G2 BREE SRE S Ses,
ERS IT 2s | es ae
SEEsaanGnseeecaasneeseseeeeee
SenUGGREnTap -cauSbErSTeEEEEEeD
ed as ae Pee es [eG
sc

BBS
/ Beane
ES< Ses keeeesake

Bt SS [a

L eae aL
BN BREA
EERE

SHUlSmeawas
NOV. DEC. JAN.

Fie. 2.—Chart showing results of analyses of Northern Spy apples in common storage at room tem-
perature (total solids basis).

SEPT. 1902 OCT.

STORAGE, RESPIRATION, AND GROWTH. 27

sy. SDDS SBaS SS oh SES eea
\ CGE eos ogee ae
eS ae
ee LLLLELLL bad Soe
EEC nee
ES Bene Zoo
SSeS paisalele leet aloo b
Aare sane sence ee
Ge eee
A 2S
HT babe
BER Zaae
eT a
a

19
ocT. 1902 NOV.

Fic. 3.—Chart showing results of analyses of Winesap apples in common storage at room tempera-
ture (total solids basis).

.
Fic,

STUDIES ON APPLES.

|_| | |
ot tt
Bs BERGER RBS ee
BEENGEGE Gees.
oo |

N
DP SERES
fh

alee
SERERER OP Zee Se oa
BRREPNrP See SEwees
Oe Ge
Sie ANS

dal Creal ela
AVé400008ee——— =" HeMeic
2 CECE

HEHEHE Ba pA Eas ee ae a a
H+ op ceca teestestastae

||
| |)
IA,
a
Ea
|

1
y |
ae
A
a
| |

ba pe

(2aeen8
5 JRA
feel tebe

ACEC
PA se eS SUZ el §|
Hees Shap eanee

Oran viim
yl TINT |
1
AG GG if Anes

1
8

Khode Island Greening — ——— Northern Spy — — — — WinNeSAp 4.

1.—Chart comparing results of analyses of three varieties of apples in common storage at room
temperature (total solids basis).

29

STORAGE, RESPIRATION, AND GROWTH.

ae2anm
Baal

AREY Gana eeaeas
SiS) 6 ae

BES Feaeeee

10 20 30
AUG. 1899 SEPT.

__

|_|
(00)
oO

2
%o

(Calculated to total solids basis. )

Fic. 5.—Chart showing results of analyses by C. A. Browne, jr., of Baldwin apples in common storage
at room temperature.

30 STUDIES ON APPLES.

Sey eu

ELT eT
TT eT
TET eA
“ SEE a aALEeeeyy CREPETLRETALEANGURIT

ia
ee
ae
ikem
(aa
es
Blea
oe
+e
ee
eae
fa
oa
ae
iad
=

“
Oo
2

wos

gee
| | eal |

i | HIUTEELEALHAAEEEEY GG00000

PETE eet NT

ESAEH PCCP ENN

OEE EEE ES asa ene aS

PALL LTT

ASV

| seer
~

TESS Ea ae a
PLP
Se Sea Se
rE ETS) e ES ee ap
jf paseo EE
ay eee a ;
EPA APAAATGMEBEAHOTONOTOWOAUOUOTOTOUEE

AAT NITE
pin ao RORERRORARERERRE Se:
90 ASE ENACT TTS oS Lae ar Sranene
SCE CUE AAG PNET EE
INTENT EET EEE EEE
LTTE Tere
BRBRRRSSHERESE SEAL

Gold Parmiine A Gold Parmiine B --------

Fic. 6.—Chart showing results of analyses by Kulisch of apples stored at cellar temperature. (Cal-
culated to an assumed basis of 17 per cent of total solids.)

STORAGE, RESPIRATION, AND GROWTH. 3l

queatly with reference to immature fruit (see page 52). It is greatly
to be regretted that this study was not begun at an earlier date, when
the apples were less mature. Apparently they had reached, or per-
haps passed, the maximum starch content before the first sample was
secured, and a complete study of the life history of the fruit of course
could not be made with the material taken. Certain irregularities
in these results are due to the fact that the apples were grown at some
distance from Washington, and two or three days elapsed from the
time they were picked until they were used in analysis, these condi-
tions not haying been sufliciently understood when the work was
begun. Further information on this subject is found on page 50.
The acid content of the apple (on the percentage basis) decreased per-
ceptibly throughout the experiment.

The foregoing comments on Rhode Island Greening apples are
equally applicable to the results on the Northern Spy and the Wine-
sap apples, platted in figures 2 and 3. The similarity of the curves
representing the content of each ingredient is strikingly brought out
in figure 4, in which the three varieties are compared.

For further comparison with these results those obtained by Dr.
C. A. Browne, jr.,“ on Baldwin apples have been calculated to total
solids and are presented in graphic form in figure 5. The results
obtained by Kulisch,? who worked with apples stored at cellar temper-
ature, have also been recalculated, using an assumed total solid basis
of 17 per cent, and the results are presented graphically in figure 6.

It will be noted that in all respects the results of Browne and of
Kulisch are analogous to those obtained in this Bureau. Doctor
Browne’s experiment, however, was begun with apples in a more
immature state than was the case with the work done in the Bureau of
Chemistry, and for that reason more complete results were obtained,
although his first examination also was made after the point of maxi-
mum starch content was passed. The work of Kulisch was carried on
with two samples of apples of the same variety from the same orchard,
but picked from different trees. His results show what wide varia-
tions in chemical composition may occur in fruit of the same variety
grown under the same cultural conditions but on different trees.

THE RIPENING OF APPLES IN COLD STORAGE.

Late in August, 1902, an experimental export shipment was made
of early-picked Ben Davis and Winesap apples from southern Illinois
for the purpose of determining whether it would be profitable to place
these apples on the London market so early in the season. The fruit
had been picked About August 15. Some weeks later samples of both

—

@ Pa. State Dept. of Agr., Bul. No. 58.
6 Landw. Jahrb., 1892, 21: 871.

32 STUDIES ON APPLES.

varieties were secured from lots from the same locality, picked at the
same time, which had been kept in cold storage in Chicago. Samples
were also secured of the same varieties picked at the usual time, about
September 15, from trees in the same locality and situated similarly
to those from which the export shipments had been taken. Early in
October two barrels of each variety from each picking were shipped
to Washington from Chicago in refrigerator cars. By special arrange-
ment the apples were sent direct from cold storage to the cars and
from the cars to the cold-storage rooms in Washington without expos-
ing them to a higher temperature for more than a few moments at a
time.

It is regretted that samples of the early-picked apples were not
obtained at the time of picking, but owing to the commercial impor-
tance of the shipment in question it seemed advisable to work with this
special lot. The apples were received on October 10 and were exam-
ined then and at varying intervals afterwards.

RESULTS OF ANALYSES.

The analytical results are found in Table III, and the platted data
in figures 7, 8, 9, and 10. The date of the last examination reported
in this bulletin was April 27, 1904. It is improbable, however, even
if they had been made, that continued analyses would have been of
any material value since the apples of all varieties were decaying
rapidly. Owing to this fact it is suggested that the results of such
analyses would jead to incorrect conclusions regarding the change in
composition of apples in storage, because of a probable selective
tendency of the rot, it being possible that apples of a certain compo-
sition may decay more rapidly than those of a different kind. At
the same time no law has been discovered which seems to control or
regulate the development of the decay in fruit. The selective ten-
dency of decay is suggested, therefore, as a possibility which might
lead to erroneous results in the work, and not as a probability indi-
cated by the work itself. At the same time it is worthy of comment
that after October 21, 1903, when decay began, the change of com-
position of the apples in storage which remained sound was noticeably
different from the change occurring prior to that date.

With reference to the solids, the carbohydrates tended to increase
after October 21, in the case of the early-picked Ben Davis and the
late-picked Ben Davis and Winesap apples. The change in content
of carbohydrates was more gradual in the case of the early-picked
W inesap.

33

STORAGE, RESPIRATION, AND GROWTH.

=

s

Hit

7.—Chart chen results of analyses of Ben Davis Or ae August 15, 1902, and under varying conditions of
storage (total solids basis).

981—Bul. 94—05——3

ld
‘

.
-

34 STUDIES ON APPLES.

| He CET
tH I HH HE BOELRRSOE
TTA LUT Ue
LTT Sot EEEEnEEed EE EEE
TRA
bape sMaNoe3007 cs seeeneee et MUUERENEEREELELEE MELLEL
Abi28 pl tee
OMT eee LTT Seat

~<a

Hanne as iiiilare
PTT RRS a
PEE er

Buell

HETEEREETSGGGEEETELELLLEBHOGE>=CUL

HEURAGETLLE Ses caaHOTGTRONOLONONEOLT
nit EHH

ded.

Pe

PO

HNOROGNNGODGKAUURER

POCCCEECCCEEC NSC He HARTRWOWOWOVOTL
AAUAHAHAGH OHAGHGG\ CHAEAONGOUGEUONAUMOGUGUAGAORUONGONGGUORUOTAENOLGENGGyeCqOLUEL
HH aS ee
BAUHEAKUGOULGRHNOMUSAUUEAUERLLGHUSOLUGAUSOAUGOUSOUSOKUOUUUGLUOOLUGLLOULUGILUGLL
PA SUBRUAOONSAENOAGRNUEUROOERRED

aa Hi wali DORGRERRRER REE
a es a Walle SAN ORMAUAGHGHGUAUNGRAONORAGNOAWOWGCOQVGRDOWUCWONONUET
Ree See ee BOGRORRORE
CECE ere Eo Pee cee
oa tn te as ee

6 18 18 1727 16 26 8 18 26 17_ 27
ra 0 a TC a oe Te :

Fic. 8.—Chart showing results of analyses of Ben Davis apples picked Suis 15, 1902, and kept
under varying conditions of storage (total solids basis).

Tasie III.—Analyses of cold-storage apples.
BEN DAVIS, PICKED AUGUST 15, 1902.

Polarization. Suerose. =
Aca ae Ream | Total
Real wate oe ap tal = Starch.) Before By po-| By re- sugar he!
SES eG Ao Seater ciliee inver- | Afterinversion.| lariza-| duc- | a8 | invert
sion. tion. tion. | invert. 7
: | i ae
1902. Per ct. | Per ct. | Per cent.| ° V. Oe STG: Per ct. | Per ct. | Per ct.
1147 | Oct. 10] 17.90] 0.684 0.75 | —3.3 —8.1 at 23.4 3.67 | 3.69 7.32
1903. ‘|
2148 | Jan. 19} 17.02} .689 None. | —4.4 —8.3_ at 24 2,99'| 2.99 8.27 |
2883 | Feb. 16 17.41 | . 592 None. | —4.3 —8.4 at 21 beg Ws Fea dg tY/ 8.27
4516 | Mer. 31 | 16.60] .548 None. | —4.5 —7.37 at 25.7 2. 22 2.33 8.38
5992] May 5 16.83 | .482 None. | —4.8 | 4 75 at 25 2eas 2.04 8.61
6489 | June 15 | 16.92 470 None. | —5.05 —7.7 at 25.7 2.05) 1.81 8. 70
7290 | Oct. 21 16. 80 258 NOn eG: 7s Se eee ee | eee eee joveealeait 8. 34
7829 | Dec. 17 16. 86 . 240 None. | —d.8 —7.9 at 22 1.61 1.42 8. 64 )
1904, | ;
7304 | Jan. 29 15. 97 . 240 None. | —5.8 —7.37 at'22 1.20 1.04 8. 26
1009 | Apr. 27 | 16.21 . 190 None: |< - 222 os satan «is amy et 95 8.33
| a ae Se | | Sein
BEN DAVIS, PICKED SEPTEMBER 15, 1902.
1902.
1148 | Oct. 10 16.99 0,570 0.50 —2.8 —8.3 at 23.4 4,20 4.16 6.96 |
1903.
9150 | Jan. 19 | 16.52 . 43 None, —3.9 —8.6 at 24 3.61 3. 62 7.49
2884 | Feb. 16 17. 25 | 920 None. —3.7 —§.8 at 21 3.87 3. 84 7.72
14517 | Mar. 81 | 16.99 . 459 None. —3.5 —8, 14 at 25.7 3. 59 3.40 7.538
5993 | May 5 16.79 | .899 None; |c2.c oul eee eee 3.11 7.62
6490 | June 15 17.22 | .§99 None. | —4.8 —8.8 at 24.8; 58.10 4, 43 7.05 |
7291 | Oct. 21 17.19 28 NOn6 So) iis ccc hcle tcc co eee le irene 2.05 8.04 |
7330 | Dee. 17 16. 20 180 None. —5,8 —7,81 at22' | 1.91 1.75 8.15 |

aStarch determinations re aka ed by 0.5 per cent. » Not used in calculation.

STORAGE, RESPIRATION, AND GROWTH. 30

Taste II].—Analyses of cold-storage apples—Continued.

WINESAP, PICKED AUGUST 15, 1902.

Polarization. Sucrose. Redu-
| Acid cing Total
seri ate of , : i suge
pore hee | aa as Starch. | Before : ; By po-| Byre- | sugar ae
CC ae ae aa i ” | malic. inver- | After inversion.| lariza-| duc- | as TaeTE
sion. tion. tion. | invert. :

| Per ct. | Per cent. Ory Nie °C | Perct.| Per ct. |. Per ct. | Per ct.

4 :
1143 | Oct. 20 | 17.52 | 0.663 | 1.00 | —3.8 —7.5- at 22 2.82 2.63 8.72 11. 49
1903. |
2149 | Jan. 19) 17.06} .569 | None.| —4.8) —7.25 at 24 1.88 1.74 9. 88 tale Al
98855) Heb. 16) 17208 ) -.572.| None. —5 Ae tral 1. 76 1.68 | 10..07 11. 84
4518 | Mar. 31} 17.07 486 None. —o9.4 —7.4 at 25.7 1.56 | ait. 84 10.28 | a12.22
5994 | May 5 17.35 —. 469 INKS S| |Sn ue noan lone aagoer tected lsoeaeece 123 10. 36 11. 65
6491 | June 15 | 17.62, .459 None. 6.0 —7.6 at 26 1.24 1.01 10. 85 11.91
W292 | Oct. 215). 16.34: .376 INOm ei ree cee |sosaceoeedseccses|seooneda .57 9. 65 10. 35
73al | Dec. 17) 16°31 . 380 None. —6.6 7h CHOP aHET lla enn .64 9.98 10. 65
1904.
730) | Jan. 29 | 16.81 | .310 None. —6.4 | =e AOy ated eal lene ao .52 | 10.08 10. 63
11010 | Apr. 27} 16.34 . 300 NOME. |lcedo5eoe lsoabee aochonounsallesaucsoe \iSoRoeeaS 9:90 sl eases
| |
WINESAP, PICKED SEPTEMBER 15, 1902.
1902. | |
1142 | Oct. 20 16.84 | 0.684 0.86 | —3.1 —7.2 at 22 SHIGE Cet cy 8.01 | @11.66
1903.
2151 | Jan. 19 | 16.65 .610 None. | —4.3 —7.0 at 24 2.46 ONTO e28 11. 92
2886 | Feb. 16 | 17.10 .611 None. | —4.75 —8. 25 at 21 2.68 2.48 9.51 12.12
4519 | Mar. 31 | 17.03 .037 | None. | —o.05 —8. 14. at 25.7 2.39 2.53 | 9.96 12. 62
5995 | May 5! 17.638 0407) ea NOME. | <—5: 2 --8.1 at 25 2.24 2.14 | 10.03 12. 28
6492 | June 15 16.87 .476 | None. | —d.5 —7.6 at 26 LAGS ae aR 1 el ON 46 12.05
7293 | Oct. 21 16. 77 AAD el ararIN ULE ae eee eec|| a Stee op ee ee, PS 74 9.65 10. 43
7302 | Dec. 17 | 16.89 .440 | None. | —6.8 —8.2 at 22 1.07 .97 10.14 11.16
1904. | | |
(BAe | deh, BE NE SB ceil None. | —6.1 —7.37 at 22 97 | SOE eel Olle 10.95
11011 | Apr. 27 16. 60 ey ead eee NOM Casa erates mua ale aoe were yeas 84 84 10. 60 11.49
: | |

a Not used in calculations.

When these four lots of apples were received, only a small amount
of starch was present in the early-picked samples and in the late-
picked Winesap, while the starch had entirely disappeared from the
late-picked Ben Davis. On the date of the second examination, Jan-
uary 19, 1903, the starch in all the samples had entirely disappeared.

RESULTS wirH SAMPLES TAKEN OUT OF CoLD STORAGE.

On this date samples were removed from cold storage and kept in a
dry room in the laboratory at a temperature of about 70° F., where,
owing to the dryness of the atmosphere, they lost their moisture con-
tent rapidly; before they were finally discarded some apples were
evidently drier than others, and in many cases while one side of the
apple was firm and plump the other was dry and shriveled. On this
account great care In sampling was necessary and in spite of all the
precautions taken the variations in composition were greater than was
desirable. It was necessary also to take larger samples for analysis
than would otherwise have been used.

36

STUDIES

ON APPLES.

TasLe 1V.—Analyses of apples removed from cold storage and held in common storage.

BEN DAVIS, PICKED AUGUST 15, 1902.

=.

| Polarization. Sucrose.
Reduc-
Serial| Date of | Total Acid as) Giarchi Berorel Ee By ing
No. |analysis.| solids. malic. inver- | After inversion. | polari- | reduc- pel ce i
sion. zation. | tion. ere
1903. Per cent. | Per cent. OV. ee GF Pervct.\ Per et-.\ Per et
2148 | Jan. 19 17. 02 0.589 | None. | —4.4 — 8.3 at 24 2.99 2.99 8.27
2614 | Jan. 27 16. 91 .600 | None. | —3.9 | — 8.7 at 21 3. 641 2.42 8.71
2715 | Feb. 5 17. 67 .482 | None. | —d. 85 — 8.9 at 22 2.34 2.18 9.51
2826 | Feb. 11 17.91 | .506 | None. |. —5.9 — 8.3 at 25 1.85 1.88 9. 34
2891 | Feb. 17 17. 47 | .372 | None. | —6.95 — 8.8 at21 1.74 ete 9. 64
3531 | Feb. 25 18. 48 .405 | None. | —6.7 — 8.8 at 23.5 R61 s| SS Pepe: 10. 44
3078 | Mar. 2 18. 71 .311 | None.-| —6.5 8.74 at 23.5 1.72 1.45 10. 56
3724 | Mar. 16} 19.55 .250 | None. | —8.4 | —10.2 at 21.3 1.39 1.18 11.69
4464 | Mar. 28 | ~~ 20.26 255 , None. | —8. 25 —10.1 at 22.2 1.41 1.14 11.59
4659 | Apr. 13 | 21.34 918 | None. | —8.5 | —10.12at22.5| 1.23 .93 | 12.12
BEN DAVIS, PICKED SEPTEMBER 15, 1902.
ls 190322 4 |
9150 | Jan. 19| 16.52/ 0.543 | None. | —3.9 | — 8.6 at 24 3.61 | 3.62] 7.49
2615 | Jan. 27| 16.69| .519| None.| —5.0 | — 8.8 at 21 2.88 | 43.48) 7.88
2716 | Fe». 5d FESY/ .426 | None. | —o9.0 — 9.1 at 22 3.14 j- 3.08 8:59
2827 | Feb. 11 17. 29 .401 | None. | —4.9 — 8.3 at 25 2:62) | 1a2248) | = (Shae
2892 | Feb. 17} 18.10 .306 | None. | —o.3 — 9.7 at 2l 204 A OA a EO aot
3532 | Feb. 25 18. 27 . B98 | None. | —6.2 — 9.4 at 23.5 2.45 | 2.52] 9.87
3579 | Mar. 2 18. 53 .320 | None. | —5.0 = FS at 2h 2 228 2 eee 2A Peery
ai2o |) Wars lb js 19-54 .260 | None. | —7.75 | — 9.79 at 21.5 | 1.55 |} 1.78) 11.52
4465 | Mar. 28 20. 92 251 | None. | —7.4 | 10 sat 2252) = 2206) le 85 10.16
4660 | Apr. 13| 20.77 | 228 | None. | —8.5 | —11.0 at 22.5} 1.91 | 1.38 | 11.96
| | |
WINESAP, PICKED AUGUST 15, 1902.
| | |
| 1903. | |
2149 | Jan. 19 17.06 | 0.569 | None. | —4.8 —7.25 at 24 | 1.88 (|) LTE 9588
2616 | Jan. 27 LTT | .o74 ! None. | —5d.5 — Sob able ie Sz 142 10. 70
2717 | Feb. 5 18.35 .413 | None. | —6.45 — S02 ab 22) | ee Lote 11. 69
2828 | Feb. 11 18.13 | .450 | None. | —5.8 —8.0 at25 | 1.69 1.19 PleT5
2893 | Feb. 17| 17.40 .412 | None. | —6.3 —7.9 at 21 TE OS Y reg 18) 10. 99
3533 | Feb. 25 | 18.87 .369 | None. | —6.8 —8.3 at 23.5 LL | -- -22 01 12.19
3080 | Mar. 2| 18.44 375 | None. | —6. 25 | —7.6 at 23.5} 1.11 .89 11.58
3726 | Mar. 16 19. 09 | .340 | None. | —7.05 6582 at 2 On eee eee .74 11. 82
4466 | Mar. 28 20. 88 | .281 | None. | —7.9 —9. 35 at 22.2 LO) 283-[)) tor
4661 | Apr. 13 20.79 250 | None —7, 95 | —9.57 at 22.5 a a Le . 83 13.13
| | |
WINESAP, PICKED SEPTEMBER 15, 1902.
=k See 2s “ a sf : ———.—
1903.
2161 | Jan. 19 16. 65 0.610 | None. | —4.3 —7.5 at 24 2.46 2.01 9, 28
2617 | Jan. 27 17.29 .643 | None. | —4.7 —8. 05 at 21 9.5449 Se )9G8h
2718 | Feb. 5 17. 83 | .543 | None. | —5.4 -—8. 50 at 22 QS Meo oae 10. 63
2829 | Feb. 11 18. 41 514 | None. | —5.8 —8.7 at 25 POA Ys) Wore Bi ga fil 10. 92
2894 | Feb. 17 18. 50 424 | None. | —6.5 —9. 07 at 21 US Ta Reese Bone -f~
3584 | Feb. 25] 18.75| .415 | None. | —6.3 —8.83 at 23.5 | 1.94] 1.59] 11.86
3581 | Mar. 2 18. 29 .38) | None. | —6.1 —8.3 at 23.5 1.68 | 1.35) 11.05
3727 | Mar. 16 19.69 310 | None. | —7.4 —8. 53 at 21.5 . 85 | 1.29 12.19

aNot used in calculations.

11.30
a11.49
11. 83
adi 3s
12.29
12.31
11. 92
13.39
110
13. 44

11.71
12.56
13. 24
12.40
12.18
13. 25
12. 52
12.59
13.38

14.01.

The results of the analyses of the apples removed from cold storage
and kept in the laboratory appear in Table 1V, and they are shown

graphically in figures 7, 8, 9, and 10.

In spite of the irregularity of

the curves it will be noticed that the apples ripened much more rapidly
than those kept in cold storage, and that, before the samples were

ist. #

Ree Me semis oe ems ot ope |

STORAGE, RESPIRATION, AND GROWTH.

=|

PETE EEE EEE %
CT

70
Pe
BARRA ADR R RRA

CCCPEEErn
rE

(Soi aS

KEPT IN COLD STORAGE

b
=

Removed TO Room 70°F.
Removed To Cear 60° F,

CC

vA

10

STUDIES ON APPLES.

38

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anne auatt Pe CoCo
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STORAGE, RESPIRATION, AND GROWTH. ov

destroyed by rot, they had closely approximated ‘the composition
attained by the apples in cold storage some months later. That is,
the percentage of total sugars expressed as invert sugar and of acids
expressed as malic acid reached about the same figure in those kept
in the laboratory as was reached at a later date in those kept in cold
storage. In one respect, however, the composition of the Ben Davis
stored in the laboratory changed in a manner very different from that
of the sample kept in cold storage: In the former the sucrose decreased
before A pril 13, 1903, until it reached a lower figure for sucrose than was
found. for the cold storage apples on April 27, 1904, more than twelve
months later. The apples ripened in the laboratory contained a higher
percentage of invert sugar and a lower percentage of cane sugar than
‘those ripened in cold storage at the time when each of them was ren-
dered useless by decay. In the case of the Winesap apples, however,
this difference in the changes in sucrose and invert sugar does not
obtain. Approximately the same minimum for the sucrose was
reached in the apples stored in the laboratory as in those in cold stor-
age, the latter having reached a minimum sucrose content about Octo-
ber 21, 1903, approximately seven months after the laboratory sample
had been rendered useless by decay.

It appears, so far as can be determined from this work, that the
changes in composition (the content of starch, sugar, and acids) in cold
storage do not greatly differ from those which occur in common stor-
age, the chief difference being in the rapidity with which the changes
take place. At the same time, the fact that the changes which take
place in storage at ordinary temperatures give higher maximum values
for invert sugar and lower minimum values for sucrose in some instances
than those occurring in cold storage is worthy of consideration and
further study. Asan illustration of this may be noted the scalding of
apples in cold storage. Scald is probably caused or accompanied by a
chemical change, but as yet this can not be demonstrated by chemical
analysis.

On May 5, 1903, samples were removed from cold storage and
placed in a cellar temperature at about 60° F. Only one subsequent
analysis of the apples so stored was made. They remained in good
condition during the greater part of the summer, keeping much better
than other cold storage apples bought on the market and removed to
the same cellar and better than some apples removed directly from
the trees to the cellar. The apples of both varieties picked on August
15, 1902, remained firm and in good condition until the latter part of
July, 1903, and the last of them gave way to decay about the Ist of
October, 1903.

It was noticeable that a change in the ripening of all varieties occurred
about October 1, 1903. After that period the ripening progressed
somewhat more slowly than before. At the same time the apples

40 STUDIES ON APPLES. e

which previously had been sound now began to decay. It is suggested
that the two conditions may be due to the same cause; that is, the loss
of vitality of the apples may have exposed them to decay and rot, or
at least made them less resistant to rot, and at the same time may have
led to slower changes in the composition of the fruit. As has been
suggested in other connections, the apparent retarding in the ripening
process may have been due to the greater susceptibility to decay of
the ripest apples, and consequently to an increase in the percentage of
the relatively greener apples on each successive examination.

THE RESPIRATION OF APPLES IN COMMON AND COLD STORAGE.

On October 20, 1902, a barrel of Ben Davis apples, grown at South
Onondaga, N. Y., was secured for respiration experiments.

DESCRIPTION OF APPARATUS AND METHODS EMPLOYED.

Three stone jars of the form shown in figure 11 were secured as
containers. These jars were the ordinary glazed stoneware chlorin

Fic. 11.—Jar used in respiration experiments.

generators with stoneware covers having a ground joint. As shown in
the illustration, the jars were arranged to afford the passage of a slight
current of air. A guard tube of soda lime, then a drying tube of eal-
cium chlorid, and then a tube of moist pumice stone were connected in
series before the apparatus. The last-named tube was for the purpose
of moistening the air which passed through the apparatus, as the pas-
sage of a dry current of air would desiccate the apples and cause them
to shrink abnormally. The tube of calcium chlorid just before the
pumice stone was used in order that by repeated weighings of the latter
the amount of water carried over into the jar with the current of air
might be determined. Connected in series after the generator were

ee ne Ce ee ee

STORAGE, RESPIRATION, AND GROWTH. 41

two caleium-chlorid tubes followed by two soda-lime tubes. The end
of the second soda-lime tube contained calcium chlorid to prevent the
loss of water from the soda lime. Finally there was a guard tube of
soda lime. The current of air for this purpose was afforded by two
bottles, one of which was filled with water and stood on a table, while
the other was empty and was placed on the floor. Each bottle was

28 7 #17 +27 ~+7 ~#17 27 6 6 2 6 16 2 4
[apr] MAY JUNE J yuey TAG SEPT. 1903} J

Cold storage Cellar temperature --------

Fic. 12.—Chart comparing results of analyses of Ben Davis apples used in respiration experiments
and kept in cold storage and-at cellar temperature (total solids basis).

closed with a two-holed stopper. A tube connected the bottom of the
first bottle with the second, and each bottle was connected with one
end of the train of absorption tubes, the full bottle acting as an
aspirator, while the empty one supplied the pressure. The current of
air entered the jar through a delivery tube, passed to the bottom of the
jar, and issued from a delivery tube leading from the top. The three

492 STUDIES ON APPLES.

jars were weighed and filled with apples taken from the same lot.
One of the jars was placed in cold storage at a temperature of about
0° C. (32° F.); the second jar was placed in a cellar temperature of
about 60° F.; the third jar was also placed in cold storage, but, instead
of being supplied-with a current of air, it was sealed and not opened
until September 19, 1903, when an analysis of the contents of the jar,
both the apples and the gas, was made.

In the case of the fruit in cold storage difficulty was experienced
in several instances when the temperature read slightly below the
freezing point, the water freezing in the tube connecting the two
bottles. This difficulty was overcome by dissolving salt in the water
employed to furnish a current of air.

RESULTS OF RESPIRATION EXPERIMENTS.

The results of the examination of the apples employed for the respi-
ration experiment are given in the graphic chart shown in figure 12
and in Table V. These results coincide in a general way with the
experiments in common storage and in cold storage already described.
The fruit kept at a higher temperature ripened much more rapidly
than that kept in cold storage. Moreover, the sucrose reached a
lower minimum and the invert sugar a higher maximum in the experi-
‘ments conducted at the higher temperature. These facts also confirm
those obtained in earlier experiments. As in the case of other experi-
ments previously described (p. 39), the sucrose of the apples kept in
cold storage reached a minimum about the same time (in this case
June 16, 1903) that the invert sugar of the same apples reached a
maximum. After that date the percentage of sucrose increased and
the percentage of invert sugar decreased. It is again suggested that
this may be due to the selective tendency of the rot of the apples,
whether the rot was due to bacteria, fungus growths, or to physio-
logical death; and further, that the more mature apples may be more
subject to decay than the less mature, whatever the cause of the decay.

During the progress of the experiments the carbon dioxid and
water were determined at short intervals, usually daily, up to the
time when the experiments at cellar temperature were discontinued
owing to the exhaustion of the sample. At this time the apples in
cold storage began to rot and the increased amount of carbon dioxid
given out vitiated any conclusions which might have been drawn from
the work had it been longer continued. The amounts of carbon dioxid
given off from both jars are shown graphically in figure 12. On eom-
paring the amount of carbon dioxid eliminated with the malic-acid
curve it hardly seems possible that the carbon dioxid can be accounted
for by the disappearance of malic acid, as several workers, notably
Gerber (see p. 19), have considered to be the case. On the other
hand, the curve representing the content of total carbohydrates,

43

STORAGE, RESPIRATION, AND GROWTH.

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44 STUDIES ON APPLES.

expressed as invert sugar, is approximately the reverse of the curve
representing the evolution of carbon dioxid. The probability that
the carbon dioxid results from the decomposition of carbohydrate
bodies is strengthened by the fact that after protracted storage the
apple has apparently lost vitality, and changes in composition proceed
much more slowly than in the early days of storage. It is probable
that at this time the quantity of carbon dioxid given.off by reason of
the original transpiration of the sound apple would be considerably
less than when chemical processes were more vigorous. <At the same
period of time the changes in the composition of carbohydrate bodies
apparently proceed much more slowly than before, whereas with
apples preserved in cold storage the change in the content of acid
proceeds uniformly. The amounts of water aspirated into the jars
were found to be uniformly larger than the amounts carried out by
the current of air. This may be accounted for by the probable con-
densation of moisture on the interior of the jar, as this has been
observed in later experiments.

THE GROWTH OF APPLES.

In the summer of 1903 the study of the growth of apples was taken
up. It was desired to begin the work with the apples at the earliest
stage practicable, but at the same time it seemed best to wait until the
** June drop” was over, as the apples which did not cling to the trees
might be different in composition from those which remained securely
fastened. For this reason the apples when picked were not so small
as had been planned for the earliest experiments. For the same rea-
son, however, samples taken at the earlier date would probably have
been of little or no value. As in the case of the experiments pre-
viously described, the apples were selected and furnished by the
pomologist in charge of field investigations in the Bureau of Plant
Industry, Mr. William A. Taylor. The fruits chosen were those
which offered the widest possible range of characteristics within the
number of samples which could be examined. (See Table I.)

STORAGE, RESPIRATION, AND GROWTH. 45

RESULTS OF CONSECUTIVE ANALYSES OF GROWING APPLES.

Table VI shows the results of the analyses of apples at different
times during the period of growth in terms of the original sample:

Taste VI.—Analyses of growing apples.

SUMMER APPLES—EARLY STRAWBERRY.

| Polarization. | Sucrose. |
| Elect (Meld * Redue- Loe) ee
Serial | Date o otal | E . ing su-| sugar
= Bus : as 4Starch.a Before -__ {BY po-| By re- er
No. | analysis. | solids. |) atic. linver-| After inver- ljariza-| duc- |, 227288 |. #8 . | apple.
| sion. sion _tion. tion. + EES
1903. | Per ct.| Per et.| Per cent. °V | °V °C | Per ct.| Per ct.| Per ct. | Per ct. | Grams.
6420 | June 11 | 13.07 |. 1.38 2.76 |—3.2 |—3.35at26 | 0.11|,0.29} 3.19] 3.51] 18.56
6519 | June 17 | 14.11 | 1.42 Soh | seo | s06 ae bse le 29°) 269 | — o238 Aliso 2325
b6751 | June 23 | 13.65 1.08 -79 |—2. 85 | -—5.17 at 24 Onl aeledd) es. cose 7.03 22.0
6776 | June 25 | 14.67 | 1.16 | 3.92 _—3.60 | —5.17 at 26 | 1.22 | 1.18) 3.95 5.19 31552
ESTATES cd THA Vntew Sal 1 I bo Jo | Vea .48 |—8.50 | —d.60 at27 | 1.63 1.69 | 6.16 7.94 29. 54
Gay |) dips Ya Beye || siee7 | 2. 913-40 | —or20 at2ne |e 16|> 1.055]. 4.75 5. 85 43.12
b6851 | July 11 | 13.89 j-== ===] -33 —3.30 | —6.35 at 26 g-dl | 2232 | 6232 8.76 40. 54
6842 | July 7 | 13.47 . 98 2.34 |—3.60 | —5. 65 at 30 fe Gleateetl 6451) 574093 6. 66 47. 06
6845 | July 10 13.7 1.02 2.86 |—4.35 | —6.10 at 26 | 1:35] 1.38 5.19 6.64 | 52.92
6856 | July 15 | 13.62 | -.92 POGt 425 6 60 abs. | 6b) 1.620) 531 12 7,01 b- 7 50.62
6863 | July 17 | 14.16 | .88 2.24 |—4.0 | —8.03 at28 |¢3.14) 2.01 5. 56 7.68 63.5
b6907 | Aug. 5 | 13.39 . 61, -00 |—2.8 |—6.16 at 27.6 | 2.63 | 2.40| 5.76 8.38 58. 06
6883 | July 22 | 14.20 =D 1.78 |—3.0 | —7.92at28 |¢3.83} 2.70} 5.05) -7.89 64. 52
6888 | July 24 | 14. 84 76 299) 224, — 6.60 at 29 13029) 1 3-2) 5.15 8.53 67.70
6895 | July 29 | 14.82 | 70 1,19 |—2.4 |—6.7iat32 | 3.42] 3.10) 5.38 8. 64 64. 80
6901 | Aug. 5 | 15.14 | 76 -9f |—1.4 | —6.60 at 27.6-| 4.07 | 4.16 | 5.14 9.52 72.15
| | } |
SUMMER APPLES—BOUGH.
l | l
1903. |
6422 | June 11 | 13.49 | 0.295 2.00 |—3.4 |—3.4 at 25 0.00 0.06 5: Di) 504 37. 88
6520 | June 17 | 13.66 .315 2.47 |—3.75 | —4.4 at 26.2 so 61 5.40 | 6.04 55. 20
66752 | June 23 | 13.74 295 -76 |—3.0 |—4.3 at 2h 99 - 9, 6.03 | 7.02 51. 30
6777 | Jume 25 | 14. 44 248 2.52 |—3.8 | —5. 65 at 26 ee By eo, 5. 83 7.17% | 75.34
668,0 | July 3 | 18.95 |\..-...- 43 |—L.4 | —5.65 at 27 See | SOFT Oe 8. 58 | 64.00
6838 | July 2); 14.27 | -348 2.16 |—3.3 | —4.60 at 27 101s |e 1205 5. 92 7.02 | 96.94
b6852 | July 11 | 13.24 |\....-..: .36 |—4.15 | —6.2 at 26 UN TIA 7.04 | 8.76| 93.24
6843 | July 7 | 14.39 178 1.79 |—4.1 | —5.55 at 30 1.14 1.40 6. 22 7.69 104. 00
6846 | July 10 | 14.95 211 2.00 |—5.5 | —6.85 at 26 1.04] 1.10 7. 06 8.22 97.30
6857 | July 15 | 14.08 194 HAO E64 822) ab 2deo eee 42 | al 29 TAL | 58:00) 1082p
6864 | July 17 | 15.24 275 1.52 |—5.5 | —8.35 at 28 ESs00F Pe ris08) a tele 923s pL
b6908 | Aug. 5 | 14.18 201 | 19 |—3.4 | —5.78 at 27.6 1.86 SO) |e eos 8. 02 119.40
6884 | July 22 | 15.42 181 | 2.13 |—3.5 | —5. 40 at 28 1.48 | 1.69 5.15 6.93 108.60
6889 | July 24 | 16.86 188 1.10 |—3.8 | —7.10 at 29 Yeites |e Baas: 6.88 | 9.54 | 126.30
6896 | July 29 | 16.78 | .164 | 2.43 |—4.3 | —7.26 at 32 2235 1582 |" 6.86 |- 8.78 98. 64
| | | |
SUMMER APPLES—YELLOW TRANSPARENT
1903.

6421 | June 11 | 11.46 EL 1.49 |—3.2 | —3.85 at 25 0.50 0.38 3.70 4.10 | 30. 54
6521 | June 17 | 12.28 1. 44 1.41 |—3.2 | —4.84 at 26.2 0 77 hea kee? 5 4.4l | S91. > 132540
66753 | June 23 | 11.72 123 .30 |—3.4 | —5.06 at 24 1.28 UE! IG snOahda| a groleago
6778 | June 25 | 12.28 1.22 1.49 |—3.05 —5. 65 at 26 2.01 1.98 4.50 6.58 57.95
b6839| July. S$ 11-45 "\.-2...- {- .12 |—3.75 | —5.9 at 27 1. 67 1.63 6. 27 (LE 48. 30
6836 | July 2 | 12.96 ET 1.46 |—3.75 | —6.35at27 | 2.02 PAG 4.45 6. 67 76. 52
BE850' |) July 11) 12-21 |=. 2. 22. 04 |\—4.10 | —6.55 at 26 © 1.89 1.86 6.24 8.20 | 74.950
6844 | July 7 12.65); 1.00 -96 |—4.25 | —6. 75 at 30 1.96 | 1.93 5. 96 8.00 87.34
6847 July 10. 13.18 81 .60 |—4.05 | —7.10 at 26 PRE || DE BY) 6.18 8.67 | 91.78
6858 | July 15 | 13.39 . 96 yt \-4. 30") — 3 Ab 23. ly 2/30, 2.22) o-81 |. 8.15 | 89:60
6865 July 17 | 13.90 | -95 | 1.04 |—3.70 | —8.8 at 28 OSC foe 9in 5.96 |* 29.02} 97500
66909 | Aug. 5 | 13.40 61 09323) || —G-go ab 27.6 2.41 2.76 DeSSuln cane onl s 1Oouae
6885 | July 22 13.85 | °.89 |} . .87-—3.9 | —8.25 at 28 3.39 | 3.30 De 20 Salone eel Qa
6890 | July 24 | 14.62 .94 .39 |—3.2 | —7.6 at 29 Br Eee be 5. 93 QUE ale9

5 |/ —7.04 at 32 2.81 2.74 6.01 8.89 99.36

6394 July 29 | 13.99 | .79 53 |—3.!

aStarch determinations reduced 0.5 per cent.
b All figures in italics give analyses of apples held in the ice box since the preceding date. '
¢ Not used in calculation,

46. -- STUDIES ON APPLES.

TaBLeE VI.—Analyses of growing apples—Continued.

WINTER APPLES—BEN DAVIS.

Polarization. Sucrose.

3 |Reduc-| Total |,,.-

: Acid ; ie Weight

Serial| Date of | Total : Starch. |Before| E ‘By po-| By re-| ing su-| sugar a=
No. | analysis. | solids. shane. ane After inver- joy i7a- duc- | 88% as as : apple

| sion. SOR: | tion. | tion. ea ete ee
| | | }
| | | |

1903. | Perct.| Perct.| Per cent.| °V. | °V °C | Per ct.| Per ct.| Per ct. | Per ct. | Grams
6493 | June 16 | 13.63 | 1.64 2.23 |—1.3 | —2.0 at 25 0.54 | 0.49] 2.35 3.87.| 15.40
6833 | June 30 | 13.37 | 1.27 3.03 |—1.55 | —2.35 at 27 G24 te GF 3. 04 3.74 | 32.47
BESaS uy) AS | 1S. 58u|2 455. = 72 |\—1.8. |—3.4 at 23.5 | 1.23 |, -1.21 3.09 6.36 | 2. 11
6891 | July 28 | 15.71 . 89 3.67 |—2.65 | —3.96 at 32 1.04} 1.13 4,52 Spill 58. 60
6923 | Aug. 18 | 14.92 .78 Si Giese eee aes aces sae 1.46 4.36 5. 90 95.4
7258 | Sept. 24 | 15.05 5 5y. 2.40 "|escc co acei ee ce ee oto eee ban oe 7.56.| 130.2
7285 | Oct. 15 | 14.86 2 1246) Sass 3 a eee eee eee | 3:13) 5230 8.60 | 167.9
W294: |i OCts -23))| J4s82, | eee ee 294 \ooe.. oe] 2-8 ees cee ceeee| seeeer 3:92 | 5.53 9.60 | 149.5
7303 | Oct. 30 | 14.68 | .43, .38 |—1.7_ | —6.87. at 24.1 | 4.30| 3.87 | 5. 84 9.91 | 178.6
7308 | Nov. 5j| 15.73 Schl Boerne —1.95 | —7.15 at 23.7 | 3.99] 3.71] 95.83 9.74 | 147.4

| |
WINTER APPLES—HUNTSMAN.
|
1903. | | | |

6494 | June 16 | 12.75 | 1.43 | 1.55 |—1.2 | —1.76 at25 | 0.43 | 0.47 2.54 3. 04 | 15.
6834 | June 30 | 13.46 | 1.12 | 2.69 |—1.1 |—1.90at27 | .62 OOM eros 4.04} 31.70
a6s5, | July 13 | 12°96) Sj ose -69 |—1.7 —3.00 at 23.5 1.00 LOO n| Siroseen 6.26 | ~ 25.69
6892 | July 28 | 15.33 80 3.69 |--2.0 | —3.74 at 32 1538 -|) 124))_ 4530") = 526i) |= ogee
6924 | Aug. 18 | 14.07 60 Pe Peery mre RE Im cots eS Sek 1.78 4.08 5.95 | 98.50
7259 | Sept. 24 | 14. 03 Sie TOO: 42 25 SAS Se See ee a es | 3.09 5. 08 8.33) 118.9
7286 | Oct. 15 | 13.50 40 | SS UE Geeseed Iporeeeseaacaecelocescas | 3.65 5.56 9.40 , 163.6
B2ODE FOG. Zon las2osles ences Det) chs oe ali ees ee ee | ee | 4.82 4.98 | 10.05 166.8
7304 | Oct. 30 | 13.08 41 19 |+ .5 |—4.9 at24.1 | 4.49 | 4.26 4.82 pa a re ED
7309 | Nov. 5 | 13.39 OBO) |i ccece see + .3 | —5.55 at 23.7 | 4.49 | 4. 4.83 9.36 163.0

| { | |
1903. | | | |
6495 | June 16 | 13.57 | 0.38 1.98 |-1.9 |—2.2 at25 | 0.93] 0.32] 412|° 4.45) 18.35
6835 | June 30 | 12.60 164.» (2193 | 23 5) Sa ae a7 16 22} 4.00| 4.23| 29.63
a6855 | July 13 | 13.16 |...-.-. 61 |—-3.2 |—3.8 at 28.5 46| .41| 6.40| 6.88 | 25:41
6893 | July 28/14.30| .12| 2.42 |-2.9 |—3.74 at 32 67| .77| 5.53| 6.34| 55.19
6925 | Aug. 18 | 14.07 12, 159-15 69: [2 ha ee eee ea 1.09| 5.58] 6.68| 97.56
7260 |.Sépt: 24 | 13:88:| G09] | Sketg) | soe ele ee ee eee 1.93] 5.44| 7.47] 144.8
7287 | Oct. 15 | 14.40} .10 Mite Pe MM Seedy 2.35| 6.37] 8.84] 94.96
7296 | Oct. 23 | 14.22 |....... 1 SS Blame ho Sb ie aaa rim 2.90} 6.58} 9.63| 140.9
7305 | Oct. 30|14.22| .12| .28 |—3.2 |—7.1 at24.1| 3.99] 2197|° G74) 9.871 deers
7310 | Nov. 5|13.93| .09| None. |—4.1 |—8.2 at23.7| 3.14] 2.51| 6.72| 9.36] 105.4
| | | |

a All figures in italics give analyses of apples held in the ice box since the preceding date.
Discussion OF CHANGBS OCCURRING DURING GROWTH.

In terms of total solids the composition of the summer apples is
shown graphically in figures 13 to 15, inclusive. The results of this
work give a much more complete account of the life history of the
apple than was obtained in the previous year. By referring to the
graphic chart for the Early Strawberry (fig. 18) as an illustration of
the summer apples, it is seen that the starch content increased from
June 11, 1903, the date of the first examination, until June 25, when
the maximum starch content was attained. From that time until
August 5 the starch steadily decreased. and on that date the sample
examined still contained 6 per cent of starch based on the total solid
content of the apple.

The apples were not in a good state of preservation, however, for
by this time all had fallen from the trees and none in a condition suit-

STORAGE, RESPIRATION, AND GROWTH. 47

able for analysis could be obtained. ‘The last stages of ripening,
therefore, could not be followed. ‘This same observation is true of
the other varieties of summer apples. No sample could be obtained
which became fully ripe in the chemical sense—that is, in which the
starch had completely disappeared.

In the first sample of summer apples examined very little sucrose

CAE ce ae
Lie te

Y

*

| Net TS ee
NUTTY
HTN
N
BERN

NS,

HE NT ‘
Ht Hh be HUE
HL EELS | nH bee d
TTT Z| a

(xem eal oa ea SS Ea Ee SE (a Be J ee

EN VR da eT el PS a a a
>< Se

Refrigerated subsamples ---------

Fic. 13.—Chart showing chemical changes in summer apples (Early Strawberry) during growth
(total solids basis).

was found, showing that the work was begun early in the life history

of the fruit. The content of sucrose increased steadily, however, until

the end of the experiment. It is specially noteworthy that the per-

centage of sucrose increased even while the starch was forming. At

the beginning of this work the sample contained a larger percentage

48 STUDIES ON APPLES.

of invert sugar than starch and.the percentage of invert sugar
increased steadily until the sample was examined on July 4. Later
the percentage of invert sugar decreased. This does not mean, how-
ever, that an actual loss of invert sugar is indicated, as will be seen
by consulting figure 21, in which the composition of the fruit in grams

TETHER ELT TTT Tg
ag SU UAGEEATATTTHEERD
Hite HEU ARSE ELEC
GNOIOHOMONGWGUONBNGAE
ci HH RRIF ARNG REAR RRR RN DZ

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See

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if ace \ GAB RGRED S|

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=

aa i
au |
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ails

hic. 14.—Chart showing chemical changes in summer apples (Bough) during growth (total solids
basis).

per apple is given. The reason for the apparent loss of invert sugar—-
that is, for the decrease in the percentage of invert sugar—is due to
the rapid growth of the seh s after the date mentioned. This growth
of the apples after July 2 amounted, in the case of the Karly Straw-
berry apple, to 29 grams, the weight increasing from 43 grams on
July 2 to 72 grams on August 5.

STORAGE, RESPIRATION, AND GROWTH. 49

. In figure 16 the average composition of the three varieties of sum-
mer apples during growth is shown graphically in the form of com-
posite curves. In this it is seen that the conclusions already drawn
from the Early Strawberry apple were true of each of the varieties.
The curves representing the content of each of the substances deter-

SSS) (SS SSS2/ 9S] ASS] SSS SS SSS SSeS nS

|
SS Bee
Al

Refrigerated subsamples - -------

Fig. 15.—Chart showing chemical changes in summer apples (Yellow Transparent) during growth
(total solids basis).

| mined from time to time in the summer apples are very irregular.
| This is apparently due to the fact that summer apples usually do not
ripen with the uniformity of later varieties. An effort was made at
the beginning to secure apples of uniform ripeness, but in spite of
| 27981—Bul. 94—05 —4

50

STUDIES

ON APPLES.

this it doubtless happened that in some cases a larger percentage
of relatively green apples was obtained than in others. In addition to
this, the chemical composition. of the apples changes very rapidly

after they are picked from the trees.

This is a matter which was not

well understood at the beginning of the work and the error caused

FLETHETEEEEEEE

it rh

riGg. lo.—Chart showing

thereby

fe

was not guarded

average chemical changes during growth in the three summer varieties |

(total solids basis).

against :

is fully as it might have been.

Owing to the fact that the apples were grown in West Virginia, from
twenty-four to thirty-six hours elapsed from the time they were picked

until analysis was begun in the laboratory. It was supposed that this

lapse of time would not cause great changes in the composition of the

pees meer vig yt re

!

—R SS SS ss :hLlhUre Sh

STORAGE, RESPIRATION, AND GROWTH. Sul

apples, but it was found that such was not the case. At each picking
the sample was divided into two portions, one of which was placed in
a refrigerator at a temperature of from 12° to 15° C. (58.6° to 59° F.),
and the apples so kept were examined after the lapse of several days.
The results of the examinations of these subsamples which were kept
in the refrigerator are printed in italics in Table VI, and they are also
expressed on the charts (figs. 13 to 16) by dotted lines.

Referring to figure 13, in which the composition of the Karly Straw-
berry apples, based on total solids, is shown graphically, it is seen
that the composition of the sample of apples received June 17, 1903,
changed with remarkable rapidity. The sample at that time contained
24 per cent of starch, 4.6 per cent of sucrose, and 24 per cent of invert
sugar. After being kept in the refrigerator six days, it was examined
and found to contain 5.9 per cent of starch, 12.9 per cent of sucrose,
and 38.2 per cent of invert sugar. Thus, in six days the apples which
were picked from the trees and kept in the dark at a temperature con-
siderably lower than that to which those remaining on the trees were
exposed, contained less starch than the apples which ripened fully on
the trees forty-three days later, and almost as high a content of invert
sugar. At the same time it must be borne in mind that the apples
remaining on the trees during this period continued to grow the whole
time, whereas the transformation of starch in case of the apples
stored in the ice box was limited to a few days. For this reason the
parallel drawn is not entirely applicable.

On several succeeding dates the samples drawn from the trees were
preserved in the refrigerator for a few days with similar results. It
was found with each successive picking that apples which were stored
in the refrigerator developed somewhat more slowly than on the pre-
ceding occasion. Thus, each succeeding curve representing the change
of the apples kept in the refrigerator is a little less vertical than that
preceding it. This demonstrates that the less mature the fruit is when
gathered the more rapid are the changes tending to maturity after
picking. It would seem, therefore, that, from a commercial stand-
point, apples which are fairly mature may be expected to retain a
more constant composition than those picked in an immature state.
The same generalization also applies to the charts representing the
changes in composition in other varieties of summer apples, and are
especially borne out in the composite chart (fig. 16), which gives the
average of the results obtained with the three varieties of summer
apples. ;

The work on the winter apples was much more satisfactory than
that on the summer appies, because of the fact that they ripened more
simultaneously and the problem of securing a representative sample
was not so difficult. For this reason the curves representing the
changes of composition of the winter apples (as shown by figures 17

5Y STUDIES ON APPLES.

to 20, inclusive) are much more uniform than in the charts just preced-
ing, which represent the composition of the summer apples. The
work on the winter apples began on June 16, 1903, and extended until
November 5, 1903. These curves illustrate much better the early life
history of the fruit than those representing the work of the previous
year. As in that year, the sucrose curve is almost exactly the reverse
of the starch curve. This is only true, however, after the maximum
content of starch has been reached, which was between June 30 and
July 28 with the Ben Davis apples, on July 28 with the Huntsman
apples, and on June 30 with the Winter Paradise apples. It must be
understood that no one of these dates is suggested as the exact date of
the maximum content of starch in the apple. It is only intended to
represent the maximum cortent of starch on the various dates when
the apples were examined. On the whole, however, the maximum
content can not have varied greatly from the date given, and the
maximum percentage determined must also be approximately correct.

The observation as to the uniformity of results applies equally to
all of the determinations made. On the date of the first examination—
June 16—the content of sucrose based on total solids was 4 per cent.
The percentage of sucrose increased regularly until the last examina-
tion, which was made on November 5, when it amounted to 25.4 per
cent of the total solid content of the.apple, the rate of increase being
apparently no greater before the maximum content of the starch than
afterwards. It would appear that during its own growth and accumu-
lation a portion of the starch is converted into sucrose. Unlike the
summer apples, the percentage of invert sugar here increased from
the date of the first examination to approximately the date of the last,
so that even in percentage composition the amount of invert sugar
present did not reach its maximum until the maturity of the fruit.

In all three of the varieties of winter apples studied the percentage
of malic acid decreased from the first examination to the full maturity
of the fruit. The percentage of total sugar estimated as invert sugar
increased steadily from the first examination to full maturity. It is
a notable fact that after the maximum content of starch is reached
the percentage of starch and invert sugar taken together remains
approximately constant. As in the case of the preceding studies, the
average composition of the three varieties of winter apples has been
expressed in the form of a composite chart which is given in figure 20.

In many respects it was considered that a graphic statement showing
the actual increase in weight of the various constituents of the apple
determined would have a more definite meaning than the changes in_
chemical composition on the percentage basis. Before examination
each sample was weighed so that the data were secured for this caleu-
lation. In figures 21 to 28 the changes of the apples just considered,
both summer and winter varieties, are represented in terms of grams
per apple,

53

STORAGE, RESPIRATION, AND GROWTH.

SRHORUUSLIOOLE 7
Hit | HE
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STORAGE. RESPIRATION, AND GROWTH. 5

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Refrigerated subsamples --------

Fic. 21.—Chart showing chemical changes during growth of summer apples (Early Strawberry )—in
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Refrigerated subsamples --------
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STORAGE, RESPIRATION, AND GROWTH. 63

In the case of the summer apples the curves representing the changes
of composition in terms of grams per apple, as in the preceding charts
in which percentage composition alone was represented, are very
irregular, owing to the fact, as previously stated, that it was very dif-
ficult to secure a representative sample, the season of ripening being
so irregular that it was impossible to secure samples having an equal
number of relatively ripe apples. Irregularities of the curves in fig-
ures 21 to 24, representing the summer apples, are due to this fact.

In the charts just mentioned the actual weight of starch increased
from the date of the first analysis until July 17 in the case of the Karly

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Subsamples stored in dark --------

Fig. 29.—Chart showing results of analyses of growing apples by Lindet (calculated to an assumed
basis of 20 per cent of total solids).

Strawberry apples, and until July 2 in the case of the Yellow Trans-
parent. The percentage of starch in the Yellow Transparent apples
remained almost constant until July 23. The irregularities in the
curves representing the starch content in the Bough apples were so
great as to render conclusions from this curve impracticable. It is
probable that the maximum starch content of this variety was attained
on July 2, the same date as in the case of the Yellow Transparent. On
two subsequent occasions, however (July 22 and 29), a higher content
of starch was noted, but on each of these dates the content of sucrose

64 STUDIES ON APPLES.

was correspondingly lower than would have been expected from pre-
ceding and succeeding analyses. On account of the irregularities
in the starch content of the Bough apples, the last of the curve
representing the starch content of the composite sample is of little
value, and the same is true of the curve representing sucrose. On
the whole it is apparent that the starch content increased until the
early part of July, after which it steadily decreased. On the other
hand, the sucrose content increased regularly from the first analysis to
the full maturity of the fruit. The same is true of the invert sugar.

Turning now to the succeeding charts (figs. 25 to 28) the curves
representing the changes of composition of the winter apples are
again found to be more satisfactory than in the case of the summer
apples. Owing to the more uniform ripening of this fruit and the
consequent greater uniformity attained in the selection of samples,
the curves are much more regular. The maximum content of starch
in winter apples was attained on August 18, at least a month later than
in the case of the summer apples just considered. After reaching the
maximum starch content the sucrose increased. more rapidly than
before and maintained a fairly rapid increase until the apples were
fully matured. The content of invert sugar, total sugar, total carbo-
hydrate, and total solids remained reasonably constant from the first
analysis to the full maturity of the fruit.

In the series of plattings indicated by the star after the date of
analysis (figs. 25, 27, and 28), the analyses were calculated to the
weight of the largest average weight of apples received up to the time
when analyzed, as the sample received consisted of smaller apples.

As in the case of the summer apples duplicate samples were taken,
one of which was subjected to analysis immediately on its receipt in
the laboratory, the other being placed in a refrigerator and allowed to
stand for some time before it was examined. Owing to the press of
other work, however, examinations of these subsamples were not made.

In figure 29 are given representative results obtained by Lindet
which illustrate somewhat more clearly the change of composition of
apples during ripening than does the tabular form in which they were
given by the author. The apples were of cider varieties. The results
as given by him were entirely on the original composition of the apples.
The content of water was not given, and for the calculation of the fig-
ures here given a total solid content of 20 per cent was assumed.
While this is probably not entirely correct, it can not be greatly dif-
ferent from the true amount. One interesting feature of Lindet’s
work was the storage in the dark of a portion of the samples analyzed
at each picking. The temperature at which the apples were stored is
not given. Lindet did not appear to observe the more rapid ripening
of the immature fruit, and that fact was not apparent in the table of
composition which he published. The graphic representation here

STORAGE, RESPIRATION, AND GROWTH. 65

given, constructed from his figures, makes itevident. This article was
found subsequent to the completion of the writers’ work on the same
subject, but it is confirmed by the latter in all respects.

As indicated by the results of Lindet and those obtained by the
writers (as given in figures 13 to 16), the green apples ordinarily
mature much more rapidly when stored than when left on the tree.
It is also equally true that the apples which were picked earliest
matured more rapidly than those picked at a later date, and the invert
sugar content of the earliest picked apples reached a higher figure
after picking than that of those picked at a later time. ** Maturing”
in this connection is regarded from a chemical point of view only,
especially with regard to the decrease of malic acid and starch, and the
increase of sucrose and invert sugar.

Germination tests, conducted at 20° C. on the seeds of all samples of
the six varieties of growing apples studied in 1903, were made by the
seed laboratory of the Bureau of Plant Industry, Department of Agri-
culture, under the direction of Dr. J. W. T. Duvel. In all cases nega-
tive results were obtained, except in two instances—Huntsman (serial
No. 7286), in which case a germination of 2 per cent was noted, and
Winter Paradise (serial No. 7260), which gave 6 per cent.

METHODS OF ANALYSIS.
PREPARATION OF SAMPLE.

The sample was quartered, cores removed, twice passed through a meat grinder,
and received in a fruit jar (provided with a cover) from which the various portions
were weighed out.

The methods of the Association of Official Agricultural Chemists were followed in
general, but on account of various changes introduced as the work progressed, they
are given here in detail.

DETERMINATION OF TOTAL SOLIDS.

A sample of about 10 grams was weighed into a tared flat-bottomed lead dish,
stirred with a little water, and evaporated to nearly constant weight in a vacuum
oven at a temperature not higher than 70° C., and in a vacuum of 16 to 20 inches.

DETERMINATION OF TOTAL ACID.

Samples weighing 10 or 20 grams were weighed in a counterpoised sugar dish, and
washed into a beaker of about 400 ce capacity with about 300 cc of water, brought to
a boil, cooled somewhat (while the beaker remained covered with a watch glass),
titrated with tenth normal sodium hydroxid, using phenolphthalein as indicator,
and the result expressed as malic acid.

DETERMINATION OF SUGARS.

Five times the normal weight for the Schmidt and Haensch polariscope, 130.24
grams, was weighed in a counterpoised sugar dish, washed into a 500-ce flask, and
25 ce of basic lead acetate” was added, the mixture made nearly up to the mark,

« Prepared according to the directions given in Bul. 65, p. 84, Bureau of Chemistry,
U.S. Dept. of Agr.
27981—Bul. 94—05——5

66 STUDIES ON APPLES.

shaken, and allowed to stand over night. The solution was then filtered, the lead
nearly all removed by dry sodium sulphate, followed by dry sodium carbonate, the
solution again filtered, and the filtrate used in the determination of sugars by polar-
imetric and gravimetric methods. The Clerget method was used for the polari-
metric work, calculating by the formula as modified by Tolman, ?¢ viz:

a=)

- =

=

T
141.85-+ .062b—>

Soxhlet’s method was used in the determination of reducing sugar as invert before
and after inversion. The tables of Meissl and Wein were used, and the cuprous
oxid was filtered off on Gooch crucibles and weighed as such, as described by
Munson. ?

DETERMINATION OF STARCH.

Fifty grams of pulp were weighed into a clean cloth bag, the mouth of the bag
closed with a rubber band, and the contents squeezed with the hand or lemon
squeezer, and then washed with portions of about 25 ce of water until the washings
amounted to about 250 ec. The last washings were in all cases neutral to litmus
and free from reducing sugar. The starch was settled from the washings by means
of a centrifugal machine, repeatedly washed by stirring with fresh portions of water
and settling, and transferred to a 300-cc flask, roughly graduated at 200 cc. The
mare from the cloth bag, separated as completely as possible by scraping with a
spatula, was washed into the flask, and the whole made up to 200 cc. Twenty cubic
centimeters of approximately 25 per cent hydrochloric acid (sp. gr. 1.125) was added,
and the whole heated for three hours in, not on, the steam bath under an air con-
denser. The mixture was then cooled, almost neutralized by sodium hydroxid,
cooled again, made up to 300 cc, filtered, and 25 ce portions employed for the
determination of dextrose by Allihn’s method. ¢

DISCUSSION OF METHOD FOR STARCH.

Determinations by the above method are all 0.5 to 0.8 per cent too
high, owing to bodies not starch which remain in the mare and become
partially hydrolyzed to reducing bodies by the acid treatment.

The supernatant liquors from the centrifugal machine always give
a test for starch, but the quantity of starch present is very minute.

Satisfactory results could not be obtained by the diastase method,
because it was impossible to break all the cells. The diastase did not
have ready access to the swollen starch grains, and the complete
washing out of dextrin was not possible. The direct extraction of the
sugars by alcohol was tried, but was found less convenient than the
water extraction.

Other methods for the determination of starch in fruits have been
deseribed, among which the methods of Browne and Lindet are given
as worthy of special consideration.

“ Bul. 73, p. 70, Bureau of Chemistry, U.S. Dept. of Agr.
b Ibid, p. 65.
¢ Bul. 65, p. 49, Bureau of Chemistry, U.S. Dept. of Agr.
@ [bid, p. 58.

0

INSOLUBLE CARBOHYDRATES OR MARC. 67

C. A. Browne, jr.,¢ determined starch by the following method:
One hundred grams of finely grated pulp were washed upon a muslin
filter with repeated quantities of cold water until the filtrate amounted
to two liters, the muslin being squeezed after each addition of water.
The washed out starch was allowed to settle, the supernatant liquid
poured off, and the starch collected on a hardened filter and washed
with water. The starch was determined by the official diastase method
with the exception that sodium hydroxid instead of sodium carbonate
was used for neutralization after the hydrolysis with hydrochloric acid.
The residue on the filter was also run by the diastase method, in case
it showed any reaction with iodin. Not more than 0.1 to 0.2 per cent
of starch, calculated on the original bulk, was ever found in this residue.

Lindet? used the following procedure, depending on the solvent
action of salicylic acid on starch: One hundred grains of pulp were
placed on a filter and washed with 5 to 6 liters of water, to which a
little mustard oil was added to prevent fermentation. The filter and
contents were introduced into a flask with 250 cc of water, 2 grams of
salicylic acid, and 100 grams of salt, and boiled for three hours under
a return condenser. The volume of the resulting solution was meas-
ured and the solution polarized, corrections being made for the volume
of solid constituents. The percentage of starch was calculated from
the rotation observed, the rotatory power of the dextrin being taken
as (@)p=+177. The method was checked by hydrolyzing the dextrin
obtained to dextrose and determining the latter by means of Fehling’s
solution.

II. INSOLUBLE CARBOHYDRATES OR MARC.

PECTIN BODIES.
HistoricaAL REvIEw.

Good reviews of the literature and descriptions of pectins have been
made by von Lipmann,’ Tollens,? Hebert,’ and by Mangin,/ the latter
having taken up the work from the botanist’s point of view. In
order to bring the subject before American investigators the present
review is presented.

The pioneer work in the field was done by Braconnot,’ who found
pectins very widely distributed in plants, occurring in the dahlia, in
Jerusalem ‘artichokes, celery, carrots, onions, in stems and leaves of

«J. Am. Chem. Soc., 1901, 23: 869.
b Ann. agron., 1894, 20: 5.

¢ Chemie der Zuckerarten, 1895, p. 924-936.

@ Handbuch der Kohlenhydrate, 1888 ed., p. 242-246, 1895 ed., p. 242-247.
é Ann. agron., 1900, 26: 34-50.

J J. bot., 1891, 5: 400, 440; 1892, 6: 12.

g Ann. chim. phys., 1825 [2], 28: 173.

68 STUDIES ON APPLES.

herbaceous plants, in the cortical layers of trees, and in fruits. So con-
stant was its occurrence in his researches that this author regarded
it as one of the principal constituents of all plants. He considered it
to be similar, if not entirely identical, with the principle in plants
vaguely described as jelly. The method given by Braconnot for
obtaining pectin (as pectic acid) from roots containing starch is to
pulp the roots, wash out with water to remove sugar, then boil the
mare with dilute hydrochloric acid, wash, and heat the starch-free and
sugar-free mare with very dilute potash or soda. There results a
mucilaginous slightly alkaline liquid, from which hydrochloric acid
separates pectic acid as a jelly. So prepared, pectic acid had a feebly
acid reaction to litmus and was slightly soluble in hot water, but the
filtered liquor did not precipitate on cooling, and barely reddened
litmus. It was, however, coagulated by alcohol, by metallic salts, and
even by sugar. Dried on a capsule it appeared as transparent leaflets
which loosened from the capsule as they dried. These were slightly
swelled by cold water, and dissolved slightly in boiling water. <A
potassium salt was separated by precipitating its water solution with
alcohol, excess of alcohol extracting coloring matter and excess of
alkali. This salt was very soluble in water, possessed a flat insipid
taste, and yielded 15 per cent of potassium, calculated from the sul-
phated ash. Its water solution was coagulated by salts, and by alcohol
and sugar, and gave precipitates with solutions of salts of heavy
metals. An ammonium salt was prepared which possessed properties
similar to those of the potassium salt. The use of these salts for the
preparation of jellies was suggested, and experiments are described in
which beautiful jellies were obtained.

The pectic acid was attacked only slightly by concentrated sulphuric
acid in the cold. Nitric acid yielded oxalic acid and a white powder
which was treated with ammonia to separate from calcium oxalate.
The ammonia solution, acidulated, gave a granular crystalline sub-
stance which he believed to be mucic acid. Braconnot concluded the
article by proposing for this acid the name pectin, from the Greek
word 77xKTis, meaning coagulum.

Vauquelin“ worked on the carrot, obtaining pectin from the juice
by boiling in order to clarify, then precipitating with alcohol; and
from the mare (as pectic acid) by boiling with dilute caustic potash and
precipitating with calcium chlorid, or better, barium hydroxid, filter-
ing, treating the filtrate with sulphuric acid, and then with potassium
hydroxid, finally precipitating with hydrochloric acid. Distilled or
filtered rain water is stated to be necessary.

Braconnot’ in 1833 described the pectin separated from oak bark by
solution in alkali. It was not precipitated from solution in alkali by

“Ann. chim. phys., 1829 [2], 41: 46.
b Ann. Chem. (Liebig), 1833, 5: 275.

INSOLUBLE CARBOHYDRATES OR MARC. 69

organic acids, but was readily thrown out of solution asa jelly by a
trace of mineral acid or alkaline earth salts. The jelly was more solu-
ble in water than the pectin (pectic acid) from currants.

Mulder“ and Regnault? reported combustions of the metallic salts
of pectins and pectic acid. Mulder concluded that pectic acid prepared
by dissolving in alkali and precipitating with acid differed from pectin
only in its higher ash content.

Frémy ¢ contributed an important article treating of the difficulties
involved in this field of work and of the mutations of the pectin bodies.
Pectin was prepared from fruit juices by first boiling to coagulate
albuminous matters, filtering, and then repeatedly purifying by pre-
cipitating with alcohol and dissolving in water. The resulting body
was white and soluble in water. It proved to be difficult to burn
quantitatively on account of its ash, which retained carbon dioxid; so
its lead salt was prepared and burned, the results indicating the formula
C,,H,,0,,. Boiling this pectin with water increased the amount of
lead which would combine with it, an increase of ** saturation capacity.”

The author states that pectins do not yield sugar on hydrolysis.
Pectic acid dissolved in dilute potassium hydroxid would no longer
precipitate on adding acid, the salt of metapectic acid having been
formed. From the free meta acid, neutral lead acetate precipitated a
salt much richer in lead than the lead salt of pectic acid. The free
acid had an acid taste and was deliquescent. Long standing with
caustic potash reduced an acid of still greater ‘‘ saturation capacity ”
and more acid taste. Dilute acids effected a similar change in pectin
and pectic acid (see Chodnew, p. 71). The author considered that
the original pectin became hydrated in the above treatments and
could in this way combine with more lead.

A discussion is given concerning the changes in the cell wall as
fruits ripen, the wall becoming thinner and the fruit less acid. The
presence of an insoluble mother substance of pectin-forming material,
later named pectose, residing in the cell walls of unripe fruits, is sug-
gested, as a result of experiments on a fruit mare, which with boiling
water yielded only small quantities of pectin, but with dilute acid
gave it in abundance.

Poumarede “ denied the existence of pectic acid in plants. He con-
sidered pectin to be an organized tissue, and pectic acid a reaction
product.

Poumarede and Figuier® considered pectin and ‘* lignin” (cellulose)
to be identical.

«Ann. Chem. (Liebig), 1838, 28: 280.

J. pharm. chim., 1838, 24: 201; J. prak. Chem., 1838, 14: 270.
¢Ibid., 1840 [2], 26: 368.

“Comptes rend., 1839, 9: 660.

€Ibid., 1846, 28: 918.

70 STUDIES ON APPLES.

Fromberg“ corroborated some of Frémy’s work. Beet mare was
boiled with dilute sodium carbonate in order to extract the pectin as
the sodium salt of pectic acid, and the acid was precipitated by addi-
tion of excess of hydrochloric acid to the solution. On boiling a
solution of pectic acid made faintly alkaline, a solution of the salt of
metapectic acid was usually formed, from which no precipitate formed
on acidifying.

Chodnew” added materially to this literature of the pectin bodies.
He considered that little was established in regard to their properties
or constitution.

Pectin was prepared from pears, which were ground up, boiled, and
the juice filtered first through linen, then through paper. . The bright
filtrate was precipitated with alcohol, and the filtrate washed with
alcohol and ether and squeezed out, whereby it lost its gelatinous
character and became lke woody fiber (Holztfaser). This pectin was
soluble, easily powdered, neutral in reaction, and gave no precipitate
with calcium or barium chlorid, or with ammonium hydroxid. Lead
acetate, basic lead acetate, copper sulphate, limewater, and potassium
hydroxid gave the gelatinous precipitates. This pectin contained
from 8.5 to 8.76 per cent ash.

Pectin from apple juice was studied in a similar way. It was also
high in ash, but was largely freed from ash by precipitating by
alcohol from an acid solution. This pectin gave the combustion
figures: Carbon, 43.70 and 43.79; hydrogen, 5.63 and 5.41; oxygen,
50.67 and 50.80; and differed from pear pectin by not dissolving clear
in water. Pectin is given as C,,H,,O,,.

Pectic acid was studied, the author first giving a résumé of previous
work, particularly that of Frémy. It was prepared as follows, mostly
from white beets: The mare was boiled with dilute potassium hydroxid
for a quarter to half an hour, and filtered first through cloth and then
through paper, the last filtration being very slow. It was considered
better, after filtering through cloth, to precipitate the free acid and
dissolye in ammonium hydroxid, which yields easily filterable solu-
tions. The pectic acid was precipitated by hydrochloric or nitric
acid washed with acidulated water, then with water containing alcohol,
and finally with alcohol, and pressed out by hand. Alcohol is neces-
sary in the wash water, for as the mineral acid diminishes in concen-
tration, the pectic acid begins to stop up the pores of the filter. With
uleohol the precipitate gradually lost its gelatinous property and
became like woody fiber. It was colorless and easily powdered. By
long drying at 120° C. it became yellow-colored. It was obtained
nearly free from ash by repeatedly precipitating with alcohol from
water solution made slightly acid with nitrie acid. So prepared it
burned without swelling, yielding 1 per cent of ash, which consisted

“Ann. Chem. (Lrebig), 1843, 48 : 56. )Ibid., 1844, 51: 355,

INSOLUBLE CARBOHYDRATES OR MARC. (Ek

chiefly of iron phosphate. It was insoluble or only slightly soluble in
hot water, but readily dissolved in alkali to a clear solution.

The author discussed the difficulties in preparing pure metallic
derivatives for analysis, which he considered to be best prepared from
the alkali salt of pectic acid and a soluble salt of the metal. It is dif-
ficult to avoid inclosing excess of alkali or salt in the resulting jellies.
This source of error was removed by squeezing out the salts and wash-
ing with water. The author so prepared and analyzed the calcium,
barium, sodium, and silver salts, obtaining the formula C,,H,,O,,.

Pectinic acid was named and studied by Chodnew, who obtained it
by boiling beet mare with hydrochloric acid, precipitating by alcohol,
and washing with alcohol and ether. It differed from pectin in having
a slight acid reaction. This author discussed the question of whether
pectic acid exists as such in plants. Beet mare yielded no pectic acid
toammonium hydroxid, so that pectic acid may exist here in combined
form, perhaps joined to calcium. But the mare after twice boiling
out with hydrochloric acid (removal of pectinic acid?) still yielded to
potassium hydroxid a body which on precipitating with acid showed
all the properties of pectic acid. The author considered then that
there existed in beet marc, not pectic acid, but a mixture of pectinic
acid and a newly found acid—iiberpectic acid—which he stated.was
changed by potassium hydroxid into pectic acid, and was insoluble in
ammonia. These two pectin bodies, pectinic acid and iiberpectic acid,
he considered to be the sources of pectic acid as usually prepared.
Similar results were obtained from apples, pears, and red and yellow
beets. It was suggested that Frémy’s acid, obtained from unripe
gooseberries by boiling with acid, might be pectinic acid.

In support of the theory that pectin bodies exist combined with cal-
cium in beet marc, the author described an experiment in which beet
mare was allowed to stand with dilute hydrochloric acid. It became
soft and translucent. On washing and adding a very little limewater,
it regained its opacity and harshness.

Metapectic acid (cf. Frémy, p. 69), prepared by Chodnew by boiling
pectin with potassium hydroxid, gave precipitates with acids and salts
on standing, anda jelly with alcohol. Its lead salt treated with hydro-
gen sulphid gave a black colloidal solution from which it was impossible
to separate the lead sulphid. Its properties differed from those given by
Frémy in being not a penta-valent acid, not deliquescent in the air, and
forming no soluble salts with caleium or barium. Pectic acid boiled for

_two hours with dilute mineral acid did not dissolve entirely to metapec-

tie acid as Frémy states. The author carefully described the phenom-
ena which occurred as the acid was boiled with dilute sulphuric acid
salts. The solution soon became colored red, and reducing with silyer
and copper; formic acid and carbon dioxid were formed in small quan-
tity, the former recognized by itsodor, and the latter by limewater. As

fi. STUDIES ON APPLES.

the action progressed and the concentration increased, the odor of cara-
mel appeared, but if the volume was kept constant the solution re-
mained nearly colorless. There was nearly always left a black residue
which consisted in part of pectic acid. This was soluble in alkali, from
which acid threw it down in brownish flocks. From the reaction prod-
uct, sulphuric acid was removed by barium carbonate. The filtrate was
concentrated to a sirup and treated with alcohol, which yielded an
abundant precipitate. The filtrate from the alcohol precipitate, evap-
orated to dryness, consisted largely of sugar, recognized by the erys-
talline form of the compound with sodium chlorid, reducing property,
and by a weak fermentation. The presence of sugar explained the
formation of the black substance, a material of the nature of humic
acid. Boiling with sulphuric acid also gave rise to a peculiar odor,
like that of benzoie acid.

This author notes that pectins disappear during the ripening of
fruits; for example, pears containing much pectin when stored in the
fall yielded only 0.5 gram from 100 pears in the spring. It is sug-
gested that the pectins may form malic acid when they disappear as
pectins.

Frémy“ now published an elaborate paper on the pectin bodies.
Pectose was considered to be a substance analogous to starch. Pectin
was prepared by boiling a fruit mare or a root mare with malice or
citric acid, and also by precipitating a ripe fruit juice with alcohol.
Its properties are given—neutral, soluble in water, insoluble in alcohol,
not colored by iodin, not precipitated by neutral Jead acetate, and
inactive to polarized light.

Boiling pectin with water gave parapectin, which only differed
from pectin in being precipitated by neutral lead acetate. Boiling
pectin with dilute acid gave metapectin, like pectin and parapectin
except that it was slightly acid and precipitable by barium ehlorid.
Prolonged boiling with acids gave parapectic acid; the action of pectase
(an enzym analogous to diastase) on pectin gave soluble pectosie acid -
which gelatinized. Alkalis converted pectin into pectic acid, soluble
in alkalis and precipitating on adding acid. Heating pectic acid to
200° C. gave pyropectic acid. The author noted great difficulty in
obtaining pure pectin. Further changes and other pectin bodies were
described in an unsatisfactory way. The methods employed were not
described and practically no experimental data were given. The work
of Chodnew received no notice. The formation of sugar by the acid
hydrolysis of pectins is denied, since, though the solutions are reducing,
they are not active to polarized light and are not fermentable.’ A list
of pectins with proposed formulas is contained in the paper.

“J, pharm. chim., 1847, [8] 12: 13; Comptes rend., 1847, 24: 1046; Ann. Chem.
(Liebig), 1847, 64: 383.

’ The fact that pentoses undergo alcoholic fermentation with difficulty was discoy-
ered later. See W. E. Stone, Amer. Chem. J., 1891, 18: 73.

INSOLUBLE CARBOHYDRATES OR MARC. ~ (3

Another paper by Frémy“ appeared in 1848. The facts contained
in the previous paper are retold, and new onesare given. ‘The article
ignores the work of every other chemist working in the field, and
appears thirty-five years later, practically without change, in the
Encyclopédie Chimique. ae

Neubauer? reported concerning arabin, precipitated by alcohol from
eum arabic solution (found by Scheibler to be identical with metapec-
tic acid). |

Frémy° obtained metapectic acid by boiling beet mare with milk of
lime. |

Stude” published work which was largely confirmatory of work
already done by others. The existence of pectose was denied, the
author believing that the pectin bodies in tissues existed as calcium
pectate. No data of any kind are given in the paper.

Scheibler’ published two papers of much interest in which the for-
mation of sugar from pectin is described. Metapectic acid was pre-
pared from beet marc, and this by acid hydrolysis yielded a crystal-
lizable sugar which he called arabinose. This was followed in 1873/7
by a report of further work dealing with arabinose and metapectic
acid. ‘The sugar was the same whether prepared from beet mare or
from gum arabic. The method of preparing metapectic acid used by
him was as follows:

Beet pulp was repeatedly washed and macerated in cold 85 per cent
alcohol. The pressed-out residue was thrown into boiling water,
alcohol boiled out, potassium hydroxid added to a strong alkaline
reaction, and the solution heated for a long time on a water bath.
The product was then filtered, saturated with carbon dioxid, eoncen-
trated by evaporation, filtered, and the filtrate acidulated with acetic
or hydrochloric acid. The pectic acid was then precipitated with
alcohol and the crude product purified by repeated solution in water and
precipitation with alcohol. Finally, the concentrated aqueous solu-
tion was poured into a small high cylinder, a little alcohol was added,
and the mixture was allowed to stand several weeks. During this
time there was formed a precipitate which carries nearly all the ash,
and the filtrate yielded arabic acid of fine quality. |

Girard’ determined the pectin in gum tragacanth.

Kirchner and Tollens” hydrolyzed quince gum by boiling with 1.25
per cent of sulphuric acid for varying periods of time. In one exper-

« Ann. chim. phys., 1848, [3] 24: 5; J. f. prak. Chem., 1848, 45: 385; Ann. Chem.
(Liebig), 1848, 67: 257.

bJ. prak. Chem., 1854, 62: 193.

¢Compt. rend., 1859, 49: 561.

@ Ann. Chem. (Liebig), 1864, 131: 241.

¢ Ber. d. chem. Ges., 1868, 1: 58, 108.

Ff Ibid., 1873, 6: 612.

9 Ibid., 1875, 8: 340.

h Ann. Chem. (Liebig), 1875, 175: 205.

74 STUDIES ON APPLES.

iment the time of boiling was seven hours, and the reaction product
was examined as follows: It was filtered and the insoluble residue on
the filter weighed. Sulphuric acid was removed from the filtrate with
barium carbonate, and sugar and soluble solids were determined.
They found 28.3 per cent of nonhydrolyzed material, 39.8 per cent of
sugar (calculated as dextrose), and 46.36 per cent of gum; in all, 114.46
per cent.

Reichardt“ prepared from carrots pararabin, a body somewhat dif-
ferent from Scheibler’s arabie acid, but readily converted into it, and
showing a carbohydrate percentage composition of C,,H,,O,,.

Barfoed’ studied changes of various arabic acids from soluble into
insoluble form by standing in alcohol or on drying.

Scheibler* found that beet marc, previously extracted with alcohol,
yielded to water a body which could be obtained ash free, and which
possessed a rotatory power of over +200°.

Von Lippman“ described a body, y-galactan, of high rotatory power
from the scums of sugar manufactured from unripe beets, possibly
very similar to the metapectic acid obtained by Scheibler.

Muntz* suggested that the galactose complexes so widely distributed
in plants were probably the sources of the galactose radical in lactose
in milk.

Battut,’ Chevron,’ and Pellet” discussed the influence on the polari-
zation of sugar of the dextrorotatory pectin dissolyed on boiling
sugar-free beet mare with water. Basie lead acetate precipitates it
completely, thus disposing of the question of its interference with
polarimetric measurements.

Weisberg ‘ confirms the view of Scheibler that gelatinous hot water
extract from beet mare exhausted by alcohol was a pectin. On heat-
ing with water the acidity of the solution gradually increased. By
sulphuric acid hydrolysis of the pectin, he also obtained a furfurol
yielding sugar (arabinose).

Wohl and Van Niessen“ discuss the work of Scheibler. Pectins of
beet mare were considered to be insoluble in water, unless rendered
soluble by hydrolysis, and to be slowly changed to soluble forms by
hot water, and much more rapidly by acids and alkalis. For example,

@ Ber. d. chem. Ges., 1875, 8: 807.

bJ. prak. Chem., 1875, [2] 11: 186.

¢ Neue Zts. f. Riibenz., 1879, 3: 341.

d Ber. d. chem. Ges., 1887, 20: 1001.

Ann. chim, phys., 1887 [6], 10: 566.

J Suere indigéne, 1888, 32: 285, 311, 333, 357, 415, 456.

9 Neue Ztg. f. Rubenz., 1588, 20: 169. Sucre belge, 1888, 13: through Chem.
Ztg. R., 1888, 12: 82.

/ Sucre indigéne, 1888, $2: 390.

t Neue Zts. f. Riibenz., 1888, 21: 325.

k Zts. Ver. d. Zucker-Ind., 1889, 39: 924.

Pee Meo.

INSOLUBLE CARBOHYDRATES OR MARG. 75

water dissolved about one-third of beet mare in about thirty hours,
while dilute oxalic acid dissolved nearly 45 per cent in four hours,
heating on the water bath in each case. The method used for obtain-
ing mucic acid from marc is as follows: 385 grams of beet pulp con-
taining 4.3 per cent of mare were extracted with alcohol, the alcohol
removed, and the mare thoroughly dried. The mare was then heated
with 17 ce of nitric acid (sp. gr. 1.15) for two hours in a boiling water
bath in a 100-ce flask. The reaction product was cooled, diluted to
mark, filtered, and 50 ce evaporated to 3 cc. After standing forty-eight
hours, 94.5 mg of mucie acid had separated, equal to 0.53 per cent of
original pulp, or 12.4 per cent, calculated to marc, which equals 16.2
per cent of galactose or 14.6 per cent of galactan.

Herzfeld” considered pectins as combinations of araban and galac-
tan, not separable by known means, and recognized by yielding fur-
furol on the one hand and mucic acid on the other. He was able to
concentrate the furfurol yielding a complex by precipitating the
ammonia solution of parapectin with calcium chlorid, the calcium salt:
yielding as high as 40 per cent of furfurol. From ripe oranges he
obtained an inactive pectin which was, however, precipitable by neu-
tral lead acetate, distinguishing it from Frémy’s pectin.

Bertrand and Mallévre ? discussed the effect of the ferment pectase
on pectin, and the wide distribution of that enzym in plants. It is
especially abundant in leaves, and it has been possible to prepare it
from this source. It can only coagulate pectins in the presence of
alkaline earth salts, forming salts of pectic acid. In acid fruits it is
present in soluble form, but its action is inhibited by the free acid.

Ullik* found pectic acid to have a high rotatory power (about | a@])>=
+ 186° to 300°) and to form easily soluble, alkali salts dialysing
readily, while the other salts are insoluble and gelatinous. He sepa-
‘ated up to 80 per cent of mucie acid, but noted that pectic acid from
different sources and prepared by different methods behaved very
differently in regard to yield of mucic acid; some giving the above
high percentage, while others gave very little, and still others no
mucic acid whatever. Those yielding high percentages of mucic acid
showed the highest rotatory power (up to [@],=+300°), and on
hydrolysis passed over principally into galactose, while those showing
low percentages of mucic acid gave exclusively or almost exclusively
arabinose. Such characteristics are indicated by the author in case
of pectin from beet mare.

Sugar-free beet mare was allowed to stand several days with 1 per
cent of hydrochloric acid, then pressed out, and the filtrate concen-

“7Zts. Ver. d. Zucker-Ind., 1891, 41: 295, 667.

Compt. rend., 1894, 119: 1012; 1895, 120: 110; 1895, 121: 726.

eOsterr.-Ung. Zts. Zucker-Ind. Landw. 21: 546; 23: 268, through Chemie der
Zuckerarten, yon Lippman, 1895, p. 927, 928.

76 STUDIES ON APPLES.

trated at the lowest possible temperature. Alcohol was added and
resulting gelatinous precipitate after filtering off was dissolved in
water. After digesting for one hour with 1 per cent hydrochloric acid
at 60° C., and filtering, two precipitates were formed on fractional
precipitation of the filtrate with alcohol. The first, after repeated
solution and precipitation with water and alcohol respectively, was a
white weakly acid mass, precipitated by barium chlorid and lead ace-
tate, of strongly dextro-rotatory power ([@]p=-+ 167.4°), and yielding
20 per cent of mucic acid on oxidation. The second, after purifica-
tion in the same way, was an amorphous white substance, precipitated
by lead acetate, but not by barium chlorid, showing less dextro-
rotation ([@]p»>=+ 123.8°), and gave no muciec acid on oxidation, but
yielded with phloroglucin and hydrochloric acid an intense color
reaction denoting pentoses or pentosans.

Tromp de Haas and Tollens“” gave the ultimate composition and
products of hydrolysis by sulphuric acid of pectins from many
sources. Their results showed that the relation of hydrogen and
oxygen in these bodies was nearly 1 to 8, as required by the carbohy-
drate formula, and that the pectin bodies which they-studied contained
no complex which gave rise to dextrose, but that complexes were pres-
ent which yielded pentose sugars and galactose on hydrolyzing with
acids.

Tollens’ in a later paper says that pectin bodies may probably be
regarded as glucosides, since the acid reaction, combination with bases,
and slightly higher oxygen-hydrogen ratio of extracted pectins indi-
cate the presence of carboxyl groups. The pectin in the plant may
not have acid properties, but may exist as a lactone.

Andrlik® discusses the action of dilute hydrochloric acid in the cold
on beet mare. A pectin of specific rotatory power ([@]p=+214.4° to
220) was extracted, and purified by repeated precipitation with alco-
hol. The longer the acid acted on the beet marc, the more insoluble
in water was the pectin dissolved.

Bourquellot and Hérissey,/ and later Bourquellot’ alone, studied the
extraction of pectins with hot water, and the effect of two enzyms on
the dissolved bodies. A 1 per cent solution of pectins from gentian
root was gelatinized by a solution of pectase within 40 minutes, and
also by limewater, sodium hydroxid followed by hydrochloric acid,
neutral and basic lead acetates, ferric chlorid, magnesium sulphate, and
ammonium sulphates, but not by sodium sulphate. The pectin was
not reducing and was dextro-rotatory ([@]p>=+82.3°). Acidified water
extracted a more dextro-rotatory body ([a@],)=+145.3'

“#Ann. Chem. (Liebig), 1895, 286: 278.

bIbid., 1895, 286: 292.

¢Zts. Gacker-Ind. Bohm., 1894, 19: 101, throngh Chem. Centrbl. 1895, 66: 1, 833.

a@J, pharm. chim., 1898, [6] 7: 473; 1898, [6] 8: 145; 1899, [6] 9: 281.

‘Compt. rend., 1899, 128: 1241.

INSOLUBLE CARBOHYDRATES OR MARC. ae

They found that the soluble ferments produced by Aspergillus niger
partially hydrolyzed the pectose of gentian root, converting it into
pectin. Pectin was hydrolyzed (rendered noncoagulable) by diastase
from malt, but not by saliva or by emulsin. The pectin was said to
yield mucic acid and arabinose like that from beet mare. Bourquellot
defined pectins as substances which dissolve in water, yield mucic acid,
and are coagulated by limewater, baryta water, and by pectase. The
solutions were optically active, contrary to the results of Frémy ({a@]p=
82.3° to 194°). The ferment in malt which dissolves pectin he called
pectinase.. Pectase and pectinase, added together to a pectin solution,
‘aused coagulation, then solution, similar to the effects of rennet and
trypsin on casein.

Javillier,“ using the methods of Bourquellot and Hérissey, obtained
corroborating results with quince pectin. It was strongly dextro-
rotatory ([a@],=188.2°), gave arabinose and mucic acid, and behaved
toward malt diastase like the pectins obtained by Bourquellot.

Votocék' and Sebor? obtained from beets, by the treatment with
alkali, an arabic acid which they determined to be not a homogeneous
compound, since different preparations showed varying rotatory pow-
ers. Other evidence that the arabic acid was a mixture of similar
substances was that varying quantities of arabinose and galactose were
formed on hydrolysis of different preparations, and by hydrolyzing
the acetylation product the original acid was not regained, but the
products possessed different rotatory powers and contained different
proportions of the groups which give rise to arabinose and galactose.
A very pure glucosazone was obtained from the hydrolysis product of
the arabic acid, so that three complexes may be present
tan, and glucose.

Bauer“ obtained various sugars from pectins from different sources—
galactose from pear pectin, xylose from apple pectin, and other sugars
not positively identified from orange peel.

Widtsoe and Tollens” reported arabinose, xylose, and fucose in gum
tragacanth, which Girard’ had found to be 60 per cent pectin.

Cross’ considers that pectin may be ligno-cellulose free from
incrusting materials, and suggests that it would be well to try the
methods used for separating cellulose bodies on the pectins.

araban, galac-

@J. pharm. chim., 1899, [6] 9:163 and 513.

b Zts. Zucker-Ind. Bohm., 1899, 24: through Chem. Centrbl., 1899, [2] 70: 1022,
through J. Chem. Soc., 1900, 78:1, 208.

¢J. prak. Chem., 1891, [2] 48: 112; Landw. Versuchs-Stat., 1892, 41: 477; 1894,
43: 191; Verh. Vers. Deutsch, Ntf. u. Arzte, 1900, II, 1, Halfte, 99, Aachen, through
Chem. Centrbl., 1901, [2], 72: 196.

d Ber. d. chem. Ges., 1900, 33: 132.

eIbid., 1875, 8: 340.

J Ibid.. 1895, 28: 2609.

78 STUDIES ON APPLES.

Mangin“” contributed a comprehensive review, referred to at the
beginning of this paper, and studies which appear to be worth cor-
roborating at least by the microchemist, and if proved to be reliable,
they may be used in connection with chemical investigation. Mangin
considers that pectic compounds are constant constituents of cell
membranes; that pectose itself may be a mixture of several similar
compounds or a single body; that pectose seems to be elaborated
earlier than cellulose in young tissue, and forms the intermedullary
layer in mature tissue; that it is not elaborated from cellulose; and
that soft parenchymatous tissue is essentially characterized by a very
close association of cellulose and of pectins. |

Many coloring agents are stated to dye the pectins in plant tissues
and differentiate them from cellulose but not from nitrogenous matters
nor from lignin, suberin, or cutin, so that the number of suitable dyes
issmall. These dyes are the safranines, methylene blue, ** bleu de nuit,”
and naphthalene blue R in crystals. Safranine is said to color the
nitrogenous bodies and lignin a cerise red, while the pectins are colored
a yellow orange. Methylene and ‘‘bleu de nuit” color nitrogenous
bodies and lignin a beautiful blue, while the pectins are colored a
violet blue best seen by lamplight. A mixture of naphthalene blue R-
in crystals and acid green J. E. E. E. (poirier) (equal parts of a 1 per
cent solution of each) colors pectins violet and other bodies green.
Ruthenium red is also recommended.

The above review does not nearly do justice to Mangin’s contribu-
tions. Since the methods used are distinctly microchemical, however,
a more extended notice does not seem necessary in this connection.
Much of the material found in Mangin’s review is also found in an
article by Reynolds Green,’ who discusses the pectin bodies from the
botanist’s point of view.

Most of the facts brought out in the above review are tabulated, for
convenience of reference, in Table VII (see p. 80).

“Compt. rend. 1888, 107: 144; 1889, 109: 579; 1890, 110: 295; 1893, 116: 653;
J. bot. 1891, 5: 400-440; 1892, 6: 12, 206, 235, 363; 1898, 7: 37, 121, 325.
Science Progress, 1896, 6: 344.

i

80 STUDIES ON APPLES.
TABLE
a | - | |
| Page
in
Name. Author. Date. this Source.
| | re-
view.
ReChinee=s sane: Braconnot...-.-.-- 1825 67 | A very wide range of
plant tissues.
|
DOrsae.. ese Vauquelin ..--..- 1829 68 | (a) From carrot juice
and (b) from the
mare of earrot.
DoRex = hse IN HUY KG Kes ee = eee 1838 697) Sicha eee eee
|
Doses as ia gee aA oa 1840) 69 |, Fruit juices\/--- = 5-25 -
DORs a Chodnewes----- 1844 70 | Pears, apples ........-
WO stoctes Poumarede and | 1846 691 a 2 ce ee eee are een
Figuier.
Dos ssnerse. lnysh Aegean 1847 72 | Fruit maresand fruit
juices.
DO. tee | Studev< csc eee 1864 TS NGS Le coe a See remnes
Dors-se | Giverdy once ee 1875 | 73 | Gum tragacanth....-.
DORs cee Scheibler......... 1879 fd) || BECUINATC 2 eeeeseeeee
Doves. ...- Weisberg: ........- 188831) 74s eee GO o28 eer ee
|
Divas: Wohl and Van | 1888| 74 |..... do. 2 ees |
| Niessen,
| | |
1) fo ee | Herzfeld. ......... sv. | 75} Beet mare and or-
| anges.

Made

VIL.— Tabular résumé of

Preparation.

The mare of the tissues is
extracted with hot dilute
alkali after exhaustion
with dilute hydroehlorie
acid. The product is pre-
cipitated by acidifying
the solution.

(a) By precipitating with
alcohol; (6) by extract-
ing with dilute alkaliand
acidifying.

The juice is clarified by
boiling, then precipitated
by alcohol. The product
is then purified by repeat-
ed solution in water and
precipiation with alco-

ol.

The crushed fruit is boiled
with water, the product
filtered, and the filtrate
precipitated with alcohol.

From mares by boiling with
solutions of organic acids;
from juices by precipi-
tating with alcohol.

soluble by boiling
with water.

From beet mare by heating
with water; from oranges |.
by precipitating the clear
juice with alcohol.

INSOLUBLE CARBOHYDRATES

the literature of the pectin bodies.

OR

MARC,

81

Properties.
: | . Remarks.

Solvents. | Precipitants. poe | As Renee Other properties.

é

Alkaline | Metallic salts; |.......-.-.-.- geet em Ser sane Mucie and oxalic | The product obtain-

solutions. aleohol;: acid formed on ed by Braconnot is
sugar. oxidation with probably the pee-
nitric acid. tic acid of later

workers.

(GawWaiter sec) PAU cools | sae ye Si os (eames cra slayvalaall se terte wimialesatojars aoe The product from

(b) al- () acids. the juice (4) was

kali so- probably the pec-

lutions. tin and that from
the mare (6) the
pectic acid of later
workers,

6 ee Aye RSS aE ee tee ere pee el pean hae ape ee Combustion figures | Peetie acid was
given for pectin believed to differ
and pectic acid. from pectin only

in its higher ash
content.

WIGNER ie ee a a Does not yield | Alkali hydrolysis | The existence of a
sugar; meta- yielded the salt of mother substance
pectic acid metapectic acid. in the cell walls
formed. Combustion fig- | of fruits, which

ures of lead salt | slowly passes into
given. Boiling pectin as the fruits
with water in- ripen, issuggested.
creased the
amount of lead
which would
combine with the
pectin.
naar GOR lea Glacetatern asses ee ee eae ee ea ees oe | eNeuLralimreaction:
basic lead easily powdered.
acetate; Combustion — fig-
copper sul- ures given. Ap- t
phate; lime ple pectin differs
water; po- from pear pectin
tassium hy- | in not dissolving
droxid. in water to form
a clear solution. |
Pectins disappear
as fruits ripen.
= 2 pst eth se Selb cca SSE Sood ae Oo Se | OOS Aa reese tea ar ante fener aaa ea | Pectin was consid-
ered to be identical
| with cellulose. No
| other authors
agree with these.

Walter ses |ATCOhol as = 2 Inactive..| Does not yield | Neutralin reaction, |
sugar, but not precipitated
gives meta- by lead acetate.
pectin and The enzym pec-
patapectice tase produces pec-
acid. tosic acid. Solu-

tions of alkalis

change pectin to |
pecticacid. Pyro- |
pectic acid is pro- |
duced on heating |
to 200° C. |

ool pe Sere DE SIS Se ces REE HSA | Se Bee eres ee Pe Ree as ee ed fc ae ae ah De A discussion only;

| no data given.

\\! CICS cet | Fe Sea er eee el DN (Se a ee nee A Rene oni eee ae ate peli ae |

200°, | |

BR eau do ....| Alcohol; ba- |............| Arabinose ......| On heating the wa- |

sic lead ace- | ter solution its |
tate. | acidity gradually |
__ increased. |

2 oS Reag 2 aE ee ee ee ees do ..........| Mucic acid formed |

| from the beet

| mare indicating

| presence of galac-

| Gehan
Water ....} Alcohol; lead | From or- |.-..-- Goes eso | INIGIO EVO Bee ea ane Pectin considered to
acetate. anges in- be a mixture of
active. arabans and galac-
tans.

27981— Bul. 94—05——6

STUDIES ON APPLES. .

Tapie VII.—Tabular résumé of the .

Bertrand and

| Tromp de -Haas
and Tollens.

Bourquellot and

Pectic acid ....

Pectinie acid..

Page
in :
Date.} this Source. Preparation,
re-
view.

Td. |e J cl entke es oe sab SSeS sini ee eee ele eee ee

16 | uc sSees aot b= oes 552 Set ee eee ee

16 | .02- sods - 22 e See oaae tee ee eee

76 |"Beet miareseeeo eee Cold dilute” hydrochloric

- acid. ~

76 | Gentian root -.--- _...| Extraction with hot water -

11\- QU CCl oes See eee | eee do.23525) 5:1 Se ee

77.| Pears, apples, OTAD C= |/3)) 5 Se ee a

peel.

17 | weceed sale ee ee | ee

18>) - s-02 52. Sse ode one se] eee ee eee

60 |ics- oe eo |

70; Beetimareece-ee eee Boiling with sodium car-
bonate.

VOSA see GOs Fs Seat Boiling with dilute potassi-
um hydroxid, then fil-
tering and precipitating

~ with hydrochloric acid.

a Pectines. <. osasaeeeee Treatment with alkali......

Old MSc nee tt ee ot | i oe ec em »

of

TL | Beet mare... -.ceerees Boiling with dilute hydro- |

chlorie acid and precipi- |

tating with alcohol, |

7)
44

iy
i,

INSOLUBLE CARBOHYDRATES OR

literature of the pectin bodies—Continued.

MARC.

83

| Properties.
. Spe te Polariza- | Results of acid
| Solvents. | Precipitants. fans hydrolysis.
| |
| 2
BS eet Bae cio y nS: al sherallia a = Neuere ae Pentose sugars
| and galactose.
| | =
"Ss roe Mone eae mera Re en ne) Se
Water ....| Alcohol ...--. fe. ca) a ess Nahe eee oe
| | 214.4°to |
| pon BRIS
==. 40 Coagulated [a]pD = + | Arabinose-..-
by pectase, | 82.3°and
| limewater, | -188.2°.
| sodium, hy- c
| droxid fol-
lowed by
hydroc hlo-
EVeG 1a C1 a:
neutraland
basic lead
acetate, fer-
ric chlorid,
magnesium
and ammo- |
| nium sul- |
phate.
Se ORCS EOC aero ae [a] D = JSG Oe seer es
188.2°.
2 OER E Fe PES Dc RIN bate [ng Sear oe te Galactose and
xylose.
ohne pees eee a Sa ee 2 eS ae ae See ie ee
}
|
PAM HEA CIOSOlUtIONS)|--- 52 .22-2-|--2c2 2 s22 7 nce
solutions.
SAUIEAMIS SE Sth 7 ACIG FSR SS tl aa ea | na ag
Alkaline |....- IO pre emia] a Tans | ate ee ee he
solutions.
Petes era | fal Dp = + |e as sess
186° to |
300°.
: |
Meera eANCONO] x2. .|.°..--.-..2- ese ine eee ets

| Other properties.

Remarks.

Coagulated by pec-
| tase in presence
| of salts of alka-
line earth metals.
Hydrogen to oxy-
-genratioisnearly
1 to 8. Combus-
| tion figuresgiven.
| Author considers
that pectin may
be regarded as a
glucoside.

Mucie acid formed.
Pectin hydrol-
yzed by malt
diastase, not by
saliva or emul-
sin.

Mucie acid formed.
Hydrolyzed by
malt diastase.

| Prolonged treat-
ment with alkali
gave metapectic
acid. Noprecipi-
tate would form
on acidifying.

Prolonged boiling

in dilute alkali
gave a solution
from which noth-
ing would precip-
itate on acidify-
ing, metapectic
acid having been
formed.

Forms easily solu-
ble alkali salts.
Yields up to 80
per cent of mucic
acid.

Possesses a_ slight
acid reaction.

Pectins defined as
bodies which dis-
solve in water,
yield mucie acid,
coagulated by
limewater, baryta
water, and by pec-
tase, and which
are dextro rota-
tory.

| Discussion only; pec-

tins may be ligno-
cellulose free from
incrusting mate-
rials.

| Discussion and mi-

crosecopical work.
Pectin a constant
fundamental con-
stituent in young
tissues.

84

STUDIES ON APPLES. ; a

Taste VII.—Tabular résumé of the

Name.

Uberpectic
acid.

Pectose

Parapectin
Metapectin

Parapectic acid

Pectosic acid ..|..

. Metapecticacid

Do
Metapecticacid
(arabic acid).

Pararabin

Author. Date
Chodnew.-....-.--- 1844
Bremniviess sean 1§47

Bee he COM See ees S| LSAT,
Baa ee CO ee 1847
eat CO ee Sas,
See Ona ees | 1847
Ghodnewenss- 5. 1844
WR ON faunas 1859
Scheibler......--- 1868
Reichardt ---- == 1875

~l
i

Source. Preparation.

Beet marGeesese ee Beet marc, after exhausting
with dilute hydrochloric
acid, was treated with al-
Kali.

Plant:tisSues>: 22.555 | Se eee

Pectin == ee Boiling with water .........

ees do -...-.----------| Boiling with dilute acid -.--

Pace (Oe soses | Prolonged _ boiling with
acids.

Spe €02352- 5255-52222 -| breatnents.with she owen—
zym pectase.

Eee GO 223222 -26-25--e| Bollingawith al kel aes

Beetimarers sos seere Boiling with milk of lime..

peas do ..-.....-....-.:| Heated with potassium hy-

droxid, filtered, acidun-
lated, and precipitated
with alcohol.

| | Carrots scowle alstsceca cs | eee ae See Se eee ee

INSOLUBLE CARBOHYDRATES OR MARC. 85

literature of the pectin bodies—Continued.

Properties.
F F Remarks
: eke Pol - | Results of acid - ;
Solvents. | Precipitants. yes hydrolysis. Other properties.
Ee Se te eine A een ee a In ye eens ee | seers Soe eee ame aats Mu. NM RES ATaed as the'sub-
; F stance which gives
rise to the pectin
bodies.
sae h eae 1 TRG TUUUGT RET Reese, Se a pee pees ae | ict a me | LE a eee
Be exe ae 2a L@EKG! AEGUIGIES |e as te cececllecoucenacooscescos|| Ile jOSCinha AypnKGl
[eee Damrete User parapectin, save
— ehlorid. slightly acid.
Sa ne Wicanol acids: pee eee a ANSI A TL OLMNC GA Sere! ieee seen
|: salts.
Been elie hese a eee el Acr a Din Ose lece ces ek SS
formed.
Bi geet Spates ea at iit |e ok eee Se Verysimilar to Schei-

| bler’s arabie acid.

86 STUDIES ON APPLES.
COMMENTS ON THE PROBLEMS INVOLVED.

From the foregoing review pectin bodies would be defined as sub-
stances of undetermined function, very widely distributed in plant tis-
sues. Pectin bodies occur both in the juice and mare, i. e., in soluble and
insoluble forms. The latter form, according to the work of Scheibler,
Wohl, Van Niessen, and others, and from the work done in the Bureau
of Chemistry, seems to be resolved into soluble forms by boiling with
water. The solutions possess considerable viscosity, the property of
forming jellies with precipitants—such as alcohol, sugars, solutions of
salts, and pectase—and usually rotate polarized light to the right.
The rotating power of the different pectin bodies appears to vary con-
siderably. The distinctions laid down by Frémy as existing between
the pectin bodies seem to be based on uncertain physical properties “@
and on the amount of lead with which they will combine. Criteria
adopted by later workers are based mainly on chemical behavior and
action on polarized light. -

Chemically, pectin bodies are characterized by yielding reducing
sugars, furfurol, and mucic acid in widely varying amounts, according
to the source of the pectins and the method employed in isolating
them. These variations may be largely due to varying degrees of
hydration and to impurities in the pectin bodies examined. Dif-
ferences in the pectin bodies themselves, however, are indicated
by the results of Ullik and Herzfeld. Pectins are profoundly
changed by alkalis with the formation of salts of the so-called
pectic acid, the free acid being insoluble in water. Acid groups
appear to be formed even by treatment with very dilute alkali fora
very short time, so that extraction with alkali of pectins from plants
does not recommend itself to the writers when a study of the dissolved
material is desired.

The most important problem appears to be the quantitative deter-
mination of the pectin bodies occurring in a given tissue, because
such a method could be used to determine the function of the material
in plants—whether, for example, it is a reserve material, a by-product,
is used for structural purposes, or has all three functions or two of
them; whether the nature of the pectin body changes with the
growth or age of the tissue, or possesses a practically constant com-
position; whether the pectin bodies obtained from different sources
are identical, are mixtures of the same substances (such as araban and
galactan) in varying proportions, or are inherently different.

It is of interest to note that all pectin bodies thus far studied have
been derived from the softer tissues. The harder woody material has
not been considered.

aj. prak. Chem., 1884, 30: 370. Neue Ztg. Zucker-Ind., 1885, 14: 151.

SO PO RE PEE ET

in

fo Sty tanta ls

UY)

(

INSOLUBLE CARBOHYDRATES OR MARC. (

if

In this connection the following quotation from Cross and Bevan“
is of interest:

It appears, therefore, generally, that the pectic group are compounds of carbo-
hydrates of varied constitution with acid groups of undetermined constitution, asso-
ciated together to form molecular complexes, more or less homogeneous, but entirely
resolved by the continued action of simple hydrolytic agencies; and the pectocellu-
loses are substances of similar character in which the carbohydrates are in part
replaced by nonhydrolyzable celluloses. The general characteristics of the pecto-
celluloses are therefore these: they are resolved by boiling with dilute alkaline solu-
tions into cellulose (insoluble) and soluble derivatives of the noncellulose (pectin,
pectic acid, metapectic acid); they are gelatinized under the alkaline treatment;
they are ‘‘saturated compounds,’’ not reacting with the halogens, nor containing
any groups immediately. allied to the aromatic series.

Later in the same volume, page 221, Cross and Bevan refer to the
parenchymatous tissue of fruits, Fesliey roots, etc., as being typical
pectocelluloses. ,

This suggests the possibility that all insoluble pectin bodies occur-
ring in the vegetable world are really in combination with cellulose
and belong to the group of pectocelluloses. This idea is in part borne
out by the results previously published by several writers and con-
firmed by the results given on page 88, in which it is seen that iasolu-
ble pectin bodies are changed to soluble form by boiling with water.
If such insoluble pectin bodies, usually called pectoses, are really
pectocelluloses, this action by which they are converted into soluble
form Is really a splitting off of the pectin group from the cellulose
complex.

In this connection should be noted the recent work by Mangin,? in
which attention is called to deposits which that writer considered to
be caleium pectate between the cell walls of plant tissues. His con-
clusions, however, have recently been disputed by Devaux,’ who
asserts that this insoluble deposit was not really calcium pectate but
true pectose. If it is found that the substance now known as pectose
is really a pectocellulose, it is suggested by the writers that the latter
term be employed to designate it and that the use of the term pectose,
which is a misnomer and altogether misleading, be discontinued.

ANALYSES OF APPLE MARC.

PREPARATION OF SAMPLE.

Fully ripe Rhode island Greening apples were taken from cold
storage, wiped off, quartered, cores and bruised places removed, and
passed through a meat grinder. The pulp was then exhausted with
water by pressing out by hand in cloth bas wel successive jsouenoins

a Gomes 2d ed., 1903, p. 217,

6 Loe. cit. (see p. 78).

“Mémoires de la société des sciences physiques et naturelles de Bordeaux, 1903
[6 ]2 5-90:

S88 STUDIES ON APPLES.

of water until the wash water gave no test for reducing sugar. The
resulting mare, amounting to 2.745 per cent of the ground fresh pulp,
was spread out in shallow pans and placed on a steam radiator, and,
after drying, was ground to a fine powder, exhausted with alcohol, and
then with ether in a Soxhlet’s extractor. The alcohol removed any
sugar remaining in the marc, and the extractions which occur on the
skins of apples, namely, apple wax and a white solid, apple vitin.¢
Ether then removed very little from the marc, chiefly green color-
ing matter. The ether was evaporated spontaneously and the mare .
bottled.

RESULTS OF ANALYSIS.
Prepared as above, the mare was analyzed, with the following results:

Taste VIII.—Analysis of apple mare.

Water-free

Determinations. |Air-dry basis. Bea
b ePerncent: Per cent.
WiStCr Grice hwo 2 tinkinwentictie nee ee es RE CES See oe eee eee eee 14.04" | so eee
GellwloseGy = 2-3-2856 solo ke bce ee bR Saeko eae oa ns Sa eee eee eee 33. 82 39.33
ES Ea gS Ai ae ee es eee PEPE SRE Se os assoccck se seee cero: 27.68 32. 20
Rentosans @ i505 2022s Seen Ba tk Sea et See a ee ee Perea he) 26.51
Proven @. soso St ce. 3 ee ee ee ee ee | 3. 50 4.07
ASR Son oe ewes s ciinons oak ets be deka ke ae re, eat Rea ea ee | 96 Jt?
Soluble in boiling water: f
Hirst, HOULS: so. 22 ssa yee See he ee ee ee Se eee ee eee 20. 83 24.23
SECONG*MOUE se. =. ees Beek eae ae ee a ee 8.52 9.91
Dhird Howre Ss .2 2.3 os Melee 2s | Stee eee eee 4. 37 5.08
ROUEEAINOUT oe oc ah oo st ora Se noe ee ee 2.67 3.10
Bifthshour 2.222252 Se ee as eS ee ee Boa) 2.94
SEXUH HOUT 25.25 22S oe Sao eS Se ee ny Ti ee ap 1. 92 2.23
Total. 2.2.55 24 5. coe eet Se: Oe eee 40. 84 47.49
Soluble in 1 per cent sodium hydroxid by Gifferenceg .................-.- 62. 94 56. 80
Residue from boiling-water treatmenth -_...... 2.22. .2 222 eee eee 46. 60 54. 22
Residue from 1 per cent sodium-hydroxid treatment @...............-.--- 37. 06 43. 20

a Dried at 100° to constant weight.

b Method of Cross and Bevan, ‘‘ Cellulose,”’ p. 95.

¢ No starch was present, but the mare hydrolyzed by hydrochlorie acid, as in the method for starch
(see p. 66), gave the above percentage of reducing sugar calculated as starch.

@ Bul. 65, Bureau of Chemistry, U. 8. Dept. of Agr., p. 173; Cir. 7, Bureau of Chemistry, U.S. Dept.
Agr., revised edition, p. 1.

eN x 6.25.

J Twenty grams of mare were boiled with 400 ce of water under a return condenser for successive
periods of 1 hour each; the product was filtered after each boiling through a cloth bag, the residue
washed and returned to flask, and the filtrate evaporated and weighed.

9 The actual amount dissolyed may be somewhat higher than is indicated here, owing to hydration
(see footnote ‘‘h).”’

AIt will be noted that the sum of the residue and material extracted by boiling water is greater
than 100 per cent (101.71 per cent dry basis), probably indicating hydration of one or both of the
products.

‘ Five grams of the air-dry mare were boiled with 200 ce of 1 per cent sodium hydroxid for one-half
hour, this being the first treatment described for the determination of cellulose by the chlorination
method deseribed by Cross and Bevan.

It will be noted that 11.02 per cent (dry basis) is soluble in sodium
hydroxid, and not dissolved by the hot-water treatment. But it is
probable that longer treatment with boiling water would have removed
larger amounts. It seems impossible to make sharp separations. It
will also be noted that the lignin (removed by chlorin and sodium sul-

“Seifert, Landw. Versuchs. Stat., 1894, 45:29.

EXAMINATIONS OF APPLE STARCH. 89

phite treatments in method for cellulose) amounts to but 3.87 per cent
of the dry marc.@

The pectin removed during the first hour of boiling was soluble in
water. It yielded 46.58 per cent of pentosan and 54.6 per cent
‘‘ starch” (°) and showed a rotatory power of [@|p =71.2° to 78.7°; i. e.,
a 1.851 per cent solution polarized at 3.8° to 4.2° in a Schmidt and
Haensch polariscope in a 10-cm tube. The pectin removed during
the second. hour gave 41.46 per cent of pentosan and 30.02 per cent
‘* starch.” It was impossible to get a solution clear enough to polarize.
No mucic acid determinations were made.

Whether pectin in apple marc is a series of bodies of similar consti-
tution but of different degrees of hydration, or whether it is a mixture
of bodies of different kinds, was not determined. This could be found
out by examining, by the criteria of yields of reducing sugar on hydro-
lysis with acid, pentosans, and mucic acid, the portions obtained by
fractional extraction with boiling water from the tissue and by a simi-
lar examination of the products resulting from fractional precipitation
of the water extract with alcohol. It is hoped that such studies may
be made.

III. MICROSCOPIC AND MACROSCOPIC EXAMINATIONS OF APPLE
STARCH.

MICROSCOPIC STUDIES.
S1zE oF STARCH GRAINS.

The presence of starch in apples is in itself no new consideration.
Buignet® in his work upon the apple failed to find starch present,
although the filtered juice from green fruit was colored blue with iodin,
which phenomenon was accredited to the presence of tannin. Accord-
ing to Lindet,” the earliest mention of starch in apples was made by
Grignon’ in 1887. Lindet himself describes the general features con-
cerning the disappearance of the starch, and gives as the size of the
erains 6 to 20 microns.

An examination of the starch as it normally occurs in the tissues
will show that the grains are generally compounded, being composed
of from 2 to 5 simple grains. In preparing samples of apple starch,
the grating of the flesh usually results in tearing these compound
grains apart, so that grains having a truncated form very greatly pre-
dominate, as is shown in Plate I.

“That is, 43.20 per cent was insoluble in 1 per cent sodium bydroxid and 39.33
per cent of cellulose was obtained, so that 3.87 per cent was removed by the treat-
ments with chlorin and sodium sulphite.

bSee footnote—c, page 88.

¢J. phar. chim., 1859, (3) 36: 81-111.

@ Ann. agron., 1894, 20: 5-20.

¢Le Cidre Doin.

90 STUDIES ON APPLES.

The individual grains are small and vary in size from 1 to 16 microns,
though generally they are between 4and 10 microns. Browne“ found
the average size of the grains to be about 9 microns, but the measure-
ments made in the Bureau of Chemistry show that the average varies
widely, according to the condition of maturity of the fruit. In Table
IX are given the results of measurements of grains from several
varieties. An examination of these results will show that, generally
speaking, the starch grains from within the core line are smaller than
those from the torus flesh. In only three specimens, out of the 47
which were examined with this point in mind, did the reverse condition
occur.

It is true, however, that in nearly ripe apples the figures might give
a slightly erroneous idea, inasmuch as there does not appear to be so
much decrease in the average size of the grains as might be expected.
In this connection, however, it must be remembered that the number
of starch grains has decreased very much, and the great majority of
those which figured in earlier measurements have disappeared, leaving
only a few stray grains behind. The smallest-sized grains which the
writer has been able to identify and measure are those slightly less than
1 micron in diameter. The writer has also occasionally found what
seemed to be stray starch grains remaining in apparently normal cells
of apples which appeared fully ripe. Such grains are rare, however,
and the cause for their persistence is not at all clear, though it is pos-
sible that it is due to a condition within the cells themselves, resulting
in a vitality too low to carry on the ordinary metabolic change.

All the specimens of each variety were taken from the same orchard
so that the variations arising from differences in climate and soil have
been eliminated.

TaBLeE IX.—Starch-grain dimensions.

PRELIMINARY OBSERVATIONS (GROWING FRUIT).

=A Ar. date oF i. : : = - es See
Serial Date of | give of grains in torus Size of grains within —

' Variety. examina- P
. - aS b . . >
No. rari flesh. core line.}
1902. Microns. Microns.
eet = York Imperial. .25 225 eC EE oe el ee ee 7. 3 (2.8 tO. 2) 6.0 (2.4 to 11.2)
sackoee GTIMES ose = ask Sack Sore ee ee ee Ores 10. 2 (6.0 to 16.0) 6.0 (2.4 to 11. 23%
areas | be Ey Aer >
COMMON-STORAGE APPLES. 4
1120 | Rhode Island Greening: -
AS pee ut eS ee ened 5.5 (3.2 to 8.8) 2.9(1.6to 6.4)
Bie sm = FSET aoe se mel hee don. 6.0 (2.4 to 9.6) 4.2(1.6to 8.0)
ES is te My Se eae CPR SS | ae do 4.9(1.6to 9.6) 4.2(1.6to 8.0)7
LD SR Se AL Se eee. eee t sf ee do 4.2(1.6to 9.6) | 3.1(0.8to 6.4%
A VONR OG. oe 0G oe ao 8 i Sh ee 5.1(1.6to 9.6) 3.6(0.8 to 8. 0)
a Pa. Dept. of Agr., bulletin 5s. -

6The mean diameter of the starch grains in each specimen is given, followed by the minimum and —
maximum diameters in parenthesis; then the average of the mean diameters for all specimens exam-_
ined on the same date is given, followed by the lowest and highest diameters found in the group.

Se ree a siete

EXAMINATIONS OF APPLE

STARCH.

Taste 1X.—Starch-grain dimensions—Continued.

COMMON-STORAGE APPLES—Continued.

91

KS YSN —r——— —S—SEO SS

: Date of | a; eet d : Neiee
Serial Sy Gn -amina.| 5ize Of grains in torus | Size of grains within
No Variety. gunna flesh. oe ees
1127 | Rhode Island Greening: Microns. Microns.
| 1 Naa TE Pe ae RES eee i arelooa he See acca Oct. 7 4.7(2.4to 8.0) 3.8(1.6 to 7.2)
Bee on re ey eee ayes 200 4.6(1.6to 8.0) 3.7(1.6 to 8.0)
AOS Ree he eee Sane Ce kc ea eae ..do 5.7(1.6 to 9.6) 4.4(0.8 to 8.0)
| NCTA Orso ee See eee ree aciens siaicie |Weisie aie aes 5.0 (1.6 to 9.6) 3.9 (0.8 to -8.0)
1121 | Agree Spy:
1s NU Sa ee ee ha a moe Sept. 17 7.0 (2.4 to 14. 4) 4.0(1.6 to 7.2
| = IN A ets Ae i ee Oe Ree et an ..do 6.7 (2.4 to 138.6) 4,2(1.6to 7.2
| Ss erly le 5 eco aga a eee ..do 9.5 (1.6 to 12.0) 3.7(1.6 to 5.6
| 1D) ona banter ae aCe en ee ree nei ..do 8.1 (4.0 to 13. 6) 4.8 (1.6 to 8.8
| IAW CLAD Cie areata te ote tyne a erasure [imme ee erciee 7.8 (1.6 to 14, 4) 4.2(1.6to 8.8
- 1128 | Northern Spy:
YN EBs aes Sips en A Ne Ba Oct. 7 5.3 (2.4 to 9.6) 3.7(1.6 to 9.6
SB er ees ieee A rear hao areata do 5.5 (1.6 to 11. 2) 4.2 (1.6 to 11.2
Cee a neers ce wie Satis wise --do 6.4 (1.6 to 9.6) 4.7 (2.4 to 11.2
AWERNGO asad occosdgadsconesSuea|loouscosade 5. 7 (1.6 to 11. 2) 4,2(1.5 to 11.2
1154 | Northern Spy+
SI NSE 5 ARE: ake oie ae Re RGN a Oa Nov. 8 5.1 (2.6 to 10.5) 5x4" (35d toON e749
1D See OScons Ge cca ane ee AR ae HO) 5.2 (1.7 to 8.8) 5.3 (2.6 to 9.6
i AY erage BE Bre COREE Reacts GDS neCeror 5.1 (1.7 to 10.5) 5.3(2.6 to 9.6
1122 | Ben Davis:
Te Seta fe Nai 5 rier age ee eS Sept. 19 9.2 (3.2 to 15. 2) 5.3 (1.6 to 12.8
Bi eee pa ares apron ee --do. 9.0 (3.2 to 16.8) 3.9(1.6to 9.6
apa eee eet kar ee Cae oe a (0K) q 8.6 (4.0 to 13.6) 5.9 (2.4 to 10.4
INGLES es 3 eo Ba Sige el CCIE 8.9 (3.2 to 16.8) 4.8 (1.6 to 12.8
1145 | Ben Davis:
| Nee Peet eg ate Sa aN os fa RD Oct. 23 3.6 (0.8 to 8.0) No starch present.
133 UoresGed ose CaSe eS Sea eee Oo 4.6 (1.6 to 8.0) No starch present.
IAS VCMT A Crete Sees Rete Ses, ie, atae | wie weraiste Soe 4.1(0.8to 8.0)
ld Gal SBemeD avis ates sae ee eae sre een Oct. 23 No starch present. No starch present.
ftase Bem Daryi sete ese ee Nov. -7 3.5(0.8 to 5.3) 3.1(0.8 to 6.0)
HMG4S IB Enel aS eee soe eee a Noy. 18 No starch present. No starch present.
1123 | Winesap:
| ope aetiologies eee Sept. 19 9.5 (4.8 to 12.8) 5.6 (2.4 to 14. 4)
TBS ie ee cae cae Se ana a ofr .-do 9.6 (4.0 to 14.4) 8.8 (5.6 to 18.6)
Caer ee es ee earn aes wae o ..do 0.0 (2.4 to 16. 0) 5. 4 (2.4 to 11. 2)
LTO RO ACES Ae asec 6 ee eee |e ame 9.7 (2.4 to 16.0) 6.6 (2:4 to 14. 4)
1126 | Winesap:
| ON ps ot SERS IEE eT ea Oct 7 7.3 (4.0 to 11.0) 4:8(1.6 to 8.0)
| 13) oon oa Se tyne ohne do 7.1 (2.4 to 12.0) 4.9(1.6 to 9.6)
(Os dose He Boos one De See Been aaee ee .-do 6.9 (2.4 to 12,0) 3.0(1.6to 8.0)
IMCS Soin a aRe ARO Oe OE ns aeons 7.1 (2.4 to 12.0) 4.2 (1.6 to 9.6)
1144. Winesap:
IRS Sai see UAE Teg tat ae Re Oct.. 23 5.8 (2.4 to 8.0) No starch present.
1B) e258 JE ee Se eee ee ...do 5.9 (2.4 to 9.6) No starch present.
IN WETS Osis a Sea ae Sone ae ees eat Deemer 5.8 (2.4 to 9.6)
1156 | Winesap:
| PAW eee mre Oe mid de A A ls Nov. 8 7.2 (1.7 to 15. 8) 6.4 (2.6 to 11.4)
| 5s stele se i 2 A lee ee eal ais) ere do 5.5 (0.8 to 11. 4) 7.0 (3.5 to 11.4)
| IN GORASE SS Soe Se ee ee [eee 6.3 (0.8 to 15.8) 6.7 (2.6 to 11.4)
1165 | VN Ta PERSE ga See Oe le an ee Noy. 18 starch present No starch present.
COLD-STORAGE APPLES.
1142 | Winesap:
Wal Mise Pe AS AN a Sy ee Se Oats 23 5.9 (3.0 to 12.0) No starch present.
lene Li eae ile ee ee ee edo 6.0 (2.4 to 11.0) No starch present.
INSET ERO ee ae Oo og TCR | (eee 5.9 (2.4 to 12.0) No starch present.
1143 | Winesap: 4
Ja SS Seis ith ae ace oe OE Oct. 23 5.6 (1.6 to 9.6) No starch present.
1B Se Smee CoS Sa Cane eer ts P| (ee eS 5.7 (2.4 to 11.0) No starch present.
(WOM SE cas Gea as cts aoast epee oecllesteesseer 5.6 (1.6 to 11.0)
1147 Ben Davis: 0 ;
IN ESSE poh BOR See ER ae see emer ees Oct. 23 4-3 (1.7 to 8.0) No starch present.
Ra ta eet eM Se Nn eo eats ota Edo 3.8 (1.7 to 7.0) No starch present.
Pee tS 8 Loreto ee yet tse ere Sees Se do. 4.9 (1.7 to 10.0) No starch present.
1D) SBE0 oe BoE SE ae eee ener do. 5.0 (1.7 to 8.8) No starch present.
PEL AR CM eee cea) Ase Sec sta ieee meee 4.5 (1.7 to 10.0) No starch present.
1148 | Ben Davis: «
RU ee eet riic Sn cine ns ae Scsee seas Oct. 23 6.3 (1.7 to 10.5) No starch present.
1b 2a Gat Re Oe eee tae .-do 4.8 (1.7 to 9.6) No starch present.
ORE er see teed ears oats So Selec ..do 5.3 (1. 7 to 10: 5) No starch present.
TD) soe oie On RO ee in eA ve Semen ..do 6.0 (2.6 to 8.8) No starch present.
AST CLV YO? Bs Sts AAS A | do 6.6) (1. 7 to 10:5) No starch present.

a Picked September 15.

6 Picked August 15.

<©
bo

STUDIES ON APPLES.

WEAKENING OF CELL WALLS AND INCREASE OF INTERCELLULAR AIR.

During the fall of 1901, in the course of the examination of certain
varieties, the intercellular air in the flesh of the fruit seemed to
increase constantly. This led to the making of some determinations
of the specific gravity of apples at different periods. The only tests,
however, which were made during the first season’s work, were carried
out in December. The next summer and autumn a number of esti-
mations were made at different stages in the growth and maturity of
the fruit, the results showing that the specific gravity diminishes to
the extent of 2 to 5 per cent. The results given, however, are the
average specific gravities of from two to four specimens, and toward
the latter part of the experiment it became apparent that the results
would have possessed more value if a larger number of specimens had
been used. In determining this factor the entire apple was used, and
hence is not to be considered as representing the exact specific gravity
of the flesh, though comparisons made for the purpose of this work
showed that the results were only shghtly below that of the flesh when
taken alone. The results of this work are tabulated in Table X.

TABLE X.—WSpecific gravity of whole fruit.
L yO :

Serial :

Se ey ra Date of Specifie
mee Variety. analysis. gravity.
23667 | Baldwin FE te aah Se a Te te ee os Os Hat ouoas Dec. 10,1901 0. 823
536704|' Tonathan 2s. . 2. 2UtUs ole os Pec a ee ee eee ee | ene doe ckewcs 799
290221A\ York Tmperial)/: Cats Susteis. soci on eine ne ere eee ee July 22, 1902 938
D5219 NW MGriNIes (2b a NE a tes REE Sh sy ee |e GOES G Se 8506
23069 4 NOrth ern: Sp yrs 8 ee eee cae cee cee See eeiceee See Ee eee eee eee eas Dee. 10,1901 802
23668") Rhode Island: Gmreempip a. 32 0 a. ee cases te eee oe ee ae ee ees dossces 823
23658"|) ROMND VISES ee ee eS eae RN DO Vg plats at Ry ea done 753
QOZLSn eee ne (6 Ka yee ee Ie a eerste OS Na ee ee aN ee od oe July 22,1902 . 823

LT26" | WWAMOSAID icin hohe OSE SA See Scare re SAS e era tare ar So ee a Oct.” “751902 8529
L658 |e oe oe CC (a Re ne Ree I Ue eR Ee MR OS A eh Sa eh ry Ses Nov. 18, 1902 . 8284
U2149y 22 eee GO SEF se Be eS Se EAD aE Fo ee a Jan. 20,1903 . 8726
DPA a GOs see oA tee Re eo See Se Eee UO ea . 8480
11.20|-RhodeAlsland, Greenimgys,...252:52 se; oe oe see oe ee eee ee eee eee Sept. 15, 1902 . 8337
MD Tiel eel GO isc aa hs SRE Se I ee SE Se Rd Sa Oe eee One Oct 7, 1902 . 8297
NIN epee ONS ae en a Se eR EES ea Np RH lg eg ane ee Noy. 18, 1902 . 8272
20210 | Base GO) oie ee ee EE Re SR hla ae ne Jan. 15,1908 S083
PA ai Koj. | te te (0 V0 Yeeeah Oe a ere ee ere Oni eee Li Mads Dek Lorie: Ge Re ate Jan. 28,1903 7994
LI22 |) Northern! S py sac nec Sk oe ee 5 ee ee ee ee Sept. 15, 1902 8513
L239 eee CO: scien cet oe Se ok BS ten ee Oct 7, 1902 $275
Ua eee OO). conn eek ee eee ee aes Nov. 18, 1902 S092
©1146 OM DAVIS: So jeco epee ee ee eee ae a ee Oct. 23,1902 . 7588
Dot Re ee (1 ee rey ones ae ete See Per Per eo oe nee eke Novy. 18, 1902 7832
II7{0) | | ee O25 Fe ei od ot hon, ee Rr ee doce 7890
rs Bi: 7 Ft Nee GO). ve c8 cones cd Be ae no aes Laren Oct. 23,1902 7T7D4
PANG oS See GO wis scn Se SS ed es BAe ee ee Jan. 20,1903 | . 7722
DNAS ee eee GO oF eee ote eee eee ee een oe | Oct. 23,1902 . 7919
DOU! sears 6 (0 nn a ee mneneee CRU Se aise oo AR dt Jone aos oe seo oe ote Jan. 20,1908 . 7961
iD | sos OO oe iciene bin So ies oe Se ee ae ce Nov. 18, 1902 | 7799
QOD |= cm ae CLO wae mjattinsln oe ab see spice cle e oe ee ae eee are ee Jan. 28,1903 | 7698
887 |....- 6 (0 nie et eevee tals Spcte lS Sa Gin OP ee pee oie aha he tact es oie Feb. 19, 1903 | 7120
FO eee (6 (0 nee Sa eee ene ROR REM le Ee St ee eS ah eg ree a Ge Mar. 3, 19038 7680
@:3803>'|\. 32... GOs. kee oe teee Ee ee ee ere Mar. 18, 1903 7A31
44494 |..... Oran Bsc PEE a os ae eye eatin wren Ce tee ee oe eee re Mar. 30,1903 | 7736
1 Ay pl aa Ge osc: od See is le Ss Oe Fe ee | Apr. 15,1903 | 7684
M5915 |...--. Oe wale aioe shape onus 08 uelal erence cre cen tere Oe eT ee ane Apr. 29, 1903 | 7685
rea Eel lan eae DOs. ean Sean sae ee ese tl ce fr oe ee ene eee eee May 12,1903 7732
Les | ees rhe CLO Sete nia wean Sha tli mle hwo ws MINS leone OO eee eT Jan. 28,1903 7507
aa a ee LL Ss cise a talc a orcas AMIN alge ae Re es ee na nn Mar. 38,1903 | TADS
CABZO a: so CLO oaks cate Sosre sc pve whe Cal's 6 lay te IE Re PO nce et nee ee Mar. 31, 1903 | 7645
e6914 |..4.. 0 a AR pe pee ey Se eee Oe ieet eee ers pee ee Apr. 29,1903 | 7703
62500 |e 0 ae GO Sree Ste Me wise nw Wie Sin wk ie a elke ok De eR May 26,1908 | 7529

a Pieked August 15.
» Picked september 15,
¢ Pieked September 15-20,

¢Used in respiration experiment, cellar temperature,
e Used in respiration experiment, cold-storage temperature,

EXAMINATIONS OF APPLE STARCH. 93

The appearance of more intercellular air in the tissues and the con-
sequent lessening of the specific gravity of the fruit are also correlated
with the *‘mealiness” of the fruit. It is a matter of common knowl-
edge that certain apples when ripe are more ** mealy” than others, and
that in some varieties this quality increases greatly just before decay.
If a small piece of a mealy apple is carefully crushed and then exam-
ined under the microscope, it is found that the cells, for the most part,
have not been ruptured, but have simply been separated from each
other. This has been made possible by the softening of the middle
lamelle of the cell walls, which occurs in the last stages of ripening.
The middle lamella becomes so very weak that under a slight shearing
force it gives way, thus allowing the adjacent cells to separate without
rupturing their walls.

In the earlier stages of the ripening of the fruit a very different
condition exists, because then the middle lamelle are strong, and the
cells rupture instead of splitting apart when subjected to pressure.
The softening of the middle lamella may also be artificially accom-
plished by boiling. <A striking illustration of this is found in the
process of making apple butter, in which boiling is continued until the
cells fall apart when stirred. The readiness with which this occurs
varies with the different varieties and the decree of ripeness attained.
This difference in the behavior of the middle lamella in the readiness
with which it is dissolved by heat the housewife recognizes under the
common expression of difference of ‘‘ cooking qualities,” and she selects
apples with regard to this property as well as that of flavor. The
softening of the middle lamella, whatever its cause, serves, in part at
least, to explain the difference in the mealy texture of apples which
may have the same percentage of juice, since in one case the process of
mastication does not accomplish the rupturing of the cells, which, like
little bags or capsules, retain the juice. Freezing also appears to
have a similar softening effect upon the lamella, inasmuch as apples
which have been frozen have a decidedly mealy texture and soft
consistency.

In normal ripening the softening of the middle lamella results in a
slight change in the contour of the cell wall, so that the cell is more
regular in outline. The intercellular space is increased, and, as there
is usually but little liquid to fill the cavity, the amount of intercellular
air is increased. When viewed under the microscope, the presence of
air in some varieties is very noticeable indeed when nearly ripe, while
in the greener condition it is but little in evidence.

An experiment was conducted to show the difference in the readi-
ness of disintegration (dependent on the softening of the middle
lameila in the same fruits) under different conditions of storage.
Cubes of as nearly the same size as possible were cut from two sam-
ples and placed in test tubes with a given amount of water. One set

94 STUDIES ON APPLES.

of samples used was taken from Ben Davis apples (No. 4494) which
had been kept at cellar temperature and contained 14.7 per cent of
solids. The other set was of the same variety, but had been kept in
cold storage and contained 14.3 per cent of solids. These samples were
placed under as nearly similar conditions as regards containers and
amount of water as possible and were shaken alike for four minutes.
The solids of the nondisintegrated portions were then determined.
While shaking the specimens they took up more water, so that the
solids were reduced to about 12 per cent. This made it necessary to
make the calculations on the basis of the solids present, and not on the
weight as found before and after shaking. The results showed that of
those specimens taken from cellar storage over 50 per cent disinte- --
grated in the shaking process, while with the others less than 20 per
cent of loss occurred. It seemed probable that the former apples would
be more mealy to the taste than the latter, even though they contained
an equal amount of juice, and this was found to be the case.

MACROSCOPIC STUDIES.
TERMINOLOGY.

The difficulty experienced in recording the changes occurring during
the disappearance of starch led to the free use of photography to
record these results. The cut sections were exposed for a few moments
to fumes of sulphur dioxid, rinsed in clear water, and then dipped for
about 10 minutes in a deep straw-colored solution of iodin in potas-
sium jodid solution. The starchy regions were of course strongly
differentiated by this treatment, and from the sections thus prepared
the making of suitable photographs was easily accomplished.

Before taking up the discussion of the macroscopic features of the
occurrence of starch it is necessary to locate in the fruit certain zones
which are of more or less importance in understanding the changes
which occur in the ripening. It is not our purpose, however, to go
into a discussion of the morphological features of the apple which do
not seem to be of vital importance in this connection. For the more per-
fect understanding of this part of our work it has seemed necessary to
devise a few new names, since in botanical literature no terms have been
found which exactly designate certain parts of the tissue considered.

If an apple is cut in half, along the equator balf way between the
stem and the calyx ‘Seye,” and the cut ends are examined, certain fea-
tures concerning the fruit are presented very clearly, as shown in
figure 30. The most prominent among these are the oval or elliptical
shaped seed cells with the sharp points clustered at the center. The
number of these cells is usually five, though specimens having either
four or six are not infrequently found.

RTP eer

EXAMINATIONS OF APPLE STARCH. 25

Some distance out in the fleshy portion is a line more or less dis-
tinctly shown, and commonly of a greenish color, called the core line.
This line is divisible into the same number of sections as the seed cells,
the ends of each section bending in somewhat toward the seed cells.
The points where two of these portions, usually considered the carpels,@
meet may for convenience be called the ‘*earpel seams.” Midway
between the carpel seams and on the core line are greenish dots—
the cut ends of fibrovascular bundles which might be called ‘* carpel
ribs,” since they are the midribs of the carpel leaves. Extending
down between the seed cells are wedge-shaped portions of flesh, which
may be called *‘ core wedges.”

Outside the core line in the torus flesh are other zones which figure
in the course of ripening, though probably they are of little other
interest. Their prominence
varies in different varieties,
though there is a general
similarity for all the varie-
ties so far examined. The
most important of these are
oval-shaped regions, as seen
in cross section, situated in
the midst of the torus flesh,
and having the small ends
in juxtaposition to the fibro-
vascular bundles.

Flanking the sides of each

oval region, and extending Fic. 30.—Diagrammatic cross section of apple showing lo-

toward the fibrovascular cation of various regions and structures: T, torus flesh;
S. me C, core line; §, seed cells; O, oval regions; CS, core
bundles, are two fan-shaped seam; W, core wedge; R, carpel rib; V, V-zone.

portions which meet at the

carpel seams and ribs, and form what might be termed ** V-zones.’
With these various zones in mind the photographs illustrating the
rarious stages in the ripening of apples may be clearly described.

5

CHANGES IN STARCH CONTENT.

The changes in starch content, and its location, for 4 varieties of
apples at various stages of development are shown in Plates II to V,
2 other varieties, not illustrated, being discussed from this point of -
view.

“Transactions of Ill. Horticultural Society, 1894, new series, 28: 125, Possibilities
of Improvement, T. J. Burrill.

96 STUDIES ON APPLES.
WINTER PARADISE (PLATE II).

The samples of this variety examined August 17 indicated by the
amount of starch present that ripening had already begun. The starch
had nearly all gone from the core wedges, but the torus flesh was heavily
charged with it. On September 24 the starch had practically disap-
peared from within the core line, and there was a marked diminution
in that of the torus flesh, especially in the oval zones. On October 15
most of the starch had disappeared, though some still persisted in the
region of the V-zones and fibrovascular bundles of the outer portion.
On October 23 only a trace of starch could be found, except in bruised
spots, and by the end of the month it had all disappeared.

From the starch content it would appear that the ripening period of
the Winter Paradise begins about August 15, and is ended by the last
week of October, unless kept under special conditions of storage.
Since the ripening of the core region is practically completed before
the changes in the starchy portions of the torus flesh begin, the ripen-
ing may be divided into two fairly distinct steps. In the Winter
Paradise the first period of ripening extended from about August 15
to about the middle of September, and the second period from that
time to the last part of October.

BEN DAVIS (PLATE III).

On August 17 the fruit presented an even dense deposit of starch
throughout all parts of the flesh. On September 24 the starch had
begun to decrease. It was most pronounced in the core wedges, and
a very slight decrease in the density of color produced with iodin was
noticeable in certain parts of the torus flesh. When examined again,
on October 23, a decided change was found to have occurred during
the preceding month, and only small amounts of starch remained in the
V-zones and the periphery, and a week later, on October 30, no starch
at all remained.

In the Ben Davis the ripening season is rather short, setting in
about the middle of September and being completed about the last of
October.

HUNTSMAN (PLATE Iv).

The first fruit in this series was examined August 17, at which time
the ripening apparently had not begun, unless perhaps very slightly,
as there seemed to be small amounts of starch in the inner portions
of the ‘‘core wedges.” However, it is usually true that there is less
starch in the core wedges than elsewhere, even in the green fruit.

On September 24 the iodin test showed the starch to be nearly all
gone from the core region. ‘Traces still remained in the tissues flank-
ing the carpel seams within the core flesh. A decrease was also

EXAMINATIONS OF APPLE STARCH. a

apparent in the outer portions, producing a slightly streaked appear-
ance. The beginning of the decrease in the amount of starch in the
oval zones is not so plain in this as in some other varieties. 3

On October 15 the starch was all gone and the fruit had every
appearance of being fully ripe. Specimens were also examined on
October 23, October 30, and November 5, but no material change was
noticeable except an increasing mealiness in the texture of the tissue.

From these facts it would appear that the ripening of the Hunts-
man begins about the last of August. The first period extends over
about three or four weeks and the second stage is completed by the
middle of October.

YELLOW TRANSPARENT (PLATE V).

On July 2+ this variety was found to have reached the second stage
in the course of ripening. It is true, however, that considerable
starch still remained within the core line while a decided loss of starch
seemed to be occurring in the torus flesh. The specimens examined
on July 29 showed but little change from this condition, and nearly a
month elapsed before specimens were again examined. The photo-
graphs made showed how slowly the second stage of ripening. some-
times progresses.- At this time it was found that the starch was all
gone from within the core line, while a considerable amount. still
remained in the torus flesh. Specimens examined again August 31
showed that the starch had completely disappeared.

It seems that the ripening period for the Yellow Transparent. is
approximately from the middle of June to the latter part of August.
This variety also illustrates how irregularly many of the summer
varieties mature, many specimens being much in advance of others on
the same tree as regards ripeness. It is also true that this variety
does not strictly follow a given order of ripening, as is the case with
most winter varieties.

EARLY STRAWBERRY (NOT ILLUSTRATED).

These specimens had apparently ripened considerably while en route
to the laboratory. In this variety the starch had very largely disap-
peared from within the core line by June 25. Some streaking was
also appearing in the torus flesh. On July 24 the starch had decreased
very much in the outer portion. The V-zones were very well marked
in this variety. The specimens of July 29 showed but little advance
over those of the 24th, except that there seemed to be a slight weak-
ening in the intensity of the iodin reaction. Unfortunately this set
is Incomplete both at the beginning and the ending of the maturing
period.

27981— Bul: 94—05 1

he a : STUDIES ON APPLES.

BOUGH (NOT ILLUSTRATED).

The apples of this variety ripen so rapidly that the time necessary
to bring them from the orchards gave considerable opportunity for
marked changes to occur before the specimens reached the laboratory.

The apples used were obtained from two orchards, one in Delaware
and the other at Geneva, N. Y. In both cases the samples had reached
an advanced stage of ripening before they arrived at the laboratory.
Those arriving from Geneva, August 24, showed the starch to be
nearly gone from within the core line—only a little remaining near
the carpel seams. Outside this line a slight tendency to streak was
becoming evident.

In the specimens arriving one week later, on August 31, the starch
within the core limit had entirely disappeared. Outside the core line
strong streaking had occurred; at each of the V-zones were regions
which stained very black with iodin. On September 8 the starch had
very much decreased, the greater part appearing in the V-zones.

Of the several varieties observed the Bough is one of the most strik-
ing illustrations of the changes in the quantity of starch in the various
zones of the tissues, though the location of the oval zone perhaps is
not so well defined as in some others.

CONCLUSIONS.

The macroscopic studies on the localization of starch in the process
of ripening led to the following conclusions:

The ripening takes place in two more or less well-defined stages.
The first of these occurs within the core line, the second in the torus
flesh. The first ordinarily begins by a decrease in the starch content
in the tips of the core wedges. This decrease extends outward to the
core line, and the last regions within this zone to lose their starch are
the fan-shaped portions flanking the carpel seams of the core region.

_The second stage ordinarily sets in somewhat before the first is fully
completed, and is usually heralded by starch-free streaks appearing
in the midst of the torus flesh. Though not at all contined to them,
the first indication of the disappearance of the starch is commonly
most pronounced in the oval zones. These zones increase in size and
other radial streaks appear in the fleshy portion until in the middle
part of the torus flesh there exists but little starch. Usually, extend-
ing outward from the fibrovascular bundles of the carpel ribs and
seams, there are fan-shaped (or V-shaped) portions which may best be
termed V-zones, and which, together with the peripheral 1.5 to 5 mm
of the flesh, are among the last to lose their starch.

Another fact shown is one well recognized by horticulturists,
namely, that summer apples ripen much more irregularly than the

.
i
‘
t
i

EXAMINATIONS OF APPLE STARCH. 99

winter varieties. Hence, although a longer time is necessary to com-
plete the work, for the purpose of studying the chemical changes
occurring in apples it would seem that the winter varieties are to be
preferred. It must be borne in mind, however, that much of the
ripening in these varieties occurs after they are picked, while the sum-
mer varieties are more nearly ripened before picking time.

It is also demonstrated that although there is a general similarity
between different varieties in their ripening, there are also well-defined
differences.

DESCRIPTION OF PLATES.

(The serial numbers refer to the analyses of the same s}«cimens
\7 1ich appear in the tables in the earlier part of the bulletin. |

Puate 1.—Photomicrographs of apple starch.

Fig. 1. Apple tissue, showing starch in apple cells (x 40).
g. 2. Apple starch (xX 200).

ie9|
—_—-
9

bo

Bieaale
Fig. 2.
Fig. 3:
Fig. 4.
Fig. 5.
Fig. 6.
ioe
Fig. 2.
Bions:
Fig. 4.
Fig. 5.
Fig.

Fig.

Biyous
Biggs.
Figs. ;

Fig. 5.

100

Serial No.
Serial No.
Serial No.
Serial No.
Serial No.
Serial No.

Serial No.
Serial No.
Serial No.
Serial No.
Serial No.

1. Serial No.
2. Serial No.
Fig. 3. Serial No.
4. Serial No. 7:
5. Serial No.
}. Serial No.

Serial No.

3 and 4.

Per cent.
6920; August 18, 1903; starehkcontents.2 s-5e5 eee Ber i 1. 69
7260; September 25, 1905:-starch content —=45 S25 e== =e <2 SAS
12873. October 15; 1903-sstarchycontent==-- == ae eee seul
7296:;..October:23; 1903;sstarchicontente = see = .50
7300; October305 1903- starch content=. 5s armen ORs
(310; Novemberio..1903- starchiconkem te: sass see None.

Serial No.
Serial No. 7083;

* Pirate Il.— Winter Paradise.

Puare II1.—Ben Davis.

6923; Aueust 18.1903: starehecontents= ae ee 3. £6
(208; september 'Z5, 1903: starch content: a: eas == =n are 2.40
(294: October 23, 1903: starch content = s4— eee . 94
7303;7October 30,1903. Starch comtenta === a= eee .38
73808: “November 5, 1903 starch content. 4.255042 oee ee None.

PLateE 1V.—HAuntsman.

6924: Aucustl8: 1903: -starchycontent= sae: a= eee 69
7259; September 25, 1903; starch content 225. = =e ane 00
7286; October. 15,1903: starchtcontenive: = ese fo eee 30
7295; October 23,, 1903; ‘starchicontent aoe eee mee
7304: October 30; 1903 starchycontentss2 22 =2see anaes ee .19
7310; November 5,1903> starchecontent) ses. =a aaa eae E None.
Pirate V.— Yellow Transparent.
6890: July 24, 1903: staréhycomtents Je == eee eran ene eee 0. 35
Serial No. 6894; July 29, 1903; .starch:contente 32 =~ 2 pee s= eee .53

7054; August 24, 1903; starch content.....--...-..- None.
August $1,'1903;-starehcontent =ss2ee 2-55. eae None.

O

a” ae

pan te i ee |

—

Bul. 94, Bureau of Chemistry. U. S. Dept. of Agriculture. PiPAnerle

FiG. 1.—APPLE TISSUE, SHOWING STARCH IN CELLS. (ENLARGED
40 DIAMETERS.)

FiG. 2.—APPLE STARCH. (ENLARGED 200 DIAMETERS).

aw

”

Bul. 94, Bureau of Chemistry, U. S. Dept. of Agriculture. PLATE Il.

1

| |

| |

| |

|

|

, |

| |

| Fig. 1.—August 18, 1903. Fig. 2.—September 25, 1903.

Fig. 3.—October 15, 1903. Fig. 4.—October 23, 1903.

Fig. 5.—October 30, 1903. Fig. 6.—November 5, 1903.

LOCATION AND GRADUAL DISAPPEARANCE OF APPLE STARCH, WINTER PARADISE.

Bul. 94, Bureau of Chemistry, U. S. Dept. of Agriculture.

PLATE III.

\|
i
i

Fig. 1.—August 18, 1903. Fig. 2.—September 25, 1903.

Fig. 3.—October 23, 1903. Fig. 4.—October 80, 1903.

| —<— |

POONER cre g

Fig. 5.—November 5, 1903.

LOCATION AND GRADUAL DISAPPEARANCE OF APPLE STARCH, BEN DAVIS.

Bul. $4, Bureau of Chemistry, U. S. Dept. of Agriculture. PLATE IV.

Fig. 4.—October 23, 1903.

ee a — — =

Fig. 5.—October 30, 1903. Fig. 6.—November 5, 1903.

LOCATION AND GRADUAL DISAPPEARANCE OF APPLE STARCH, HUNTSMAN.

Bul. 94, Bureau of Chemistry, U. S. Dept. of Agriculture.

PLATE V.

Fig. 1.—July 24, 1903. Fig. 2.—July 29, 1903.

Fig. 3.—August 24, 1903. Fig. 4.—August 24, 1903,

Oh ae

\

Fig. 5.—August 31, 1903.

LOCATION AND GRADUAL DISAPPEARANCE OF APPLE STARCH, YELLOW TRANSPARENT.