ways; either by König’s new process as employed by Filsinger for cacao
or by the older method of Weender’s[185] as follows:
3 grammes of the defatted and atmospherically dried substance are
boiled for ½ hour with 200 ccm of a 1·24 percent solution of
sulphuric acid. It is allowed to settle, then decanted, and the residue
boiled twice with the same volume of water. The decanted liquids are
allowed to settle in cylinders and the sediment added to the rest of
the substance, which is then boiled half an hour with 200 ccm of a 1·25
percent solution of caustic potash, filtered through a weighed filter
and the residue twice boiled with 200 ccm of water. The cellulose-like
substance collected on the filter is washed first with hot water, then
with cold, afterwards with alcohol, and finally with ether.
After being dried and weighed, it is incinerated and the necessary
corrections made for ash.
The process worked out by Henneberg is the one usually adopted for the
determination of crude fibre in vegetable matter. Recently H. Suringer
and B. Tollens[186] and more particularly König[187] have pointed out
that in Weender’s process the so-called pentosan (sugar derivative)
of the composition C_{5} H_{10} O_{5}, which comprises a not
inconsiderable portion of crude fibre, would undergo a disproportionate
alteration, so that the analytical results thus obtained would not
represent the amount of cellulose correctly. The crude fibre must
therefore be treated in such a manner as to eliminate the pentosan.
König attains that result by treating 3 grammes of the defatted
substance with 200 ccm of glycerine (1·23 sp. gr.) containing per litre
20 grammes of concentrated sulphuric acid, under a pressure of three
atmospheres, for one hour. It is then filtered through an asbestos
filter whilst hot, and after being successively washed with hot water,
alcohol and ether, it is weighed, incinerated and the ash weighed. The
difference between the two weighings expresses the amount of ash-free
crude fibre.
Filsinger has determined by König’s method the amount of crude fibre
in a series of different varieties of bean, the results of which have
already been given on page 72. Which process is the better has yet to
be established, and in issuing results as data the method employed has
always to be indicated owing to the many variations which arise.
9. ~The determination of cacao husk~, which will be for the
most part a matter of ascertaining the amount of raw or crude fibre,
could formerly only be effected by means of the microscope. In 1899
Filsinger[188] proposed a method of levigation which according to P.
Welman’s[189] gives trustworthy results. Manifold treatises have been
devoted to the subject, and it would be advisable to turn a few of
these up and compare the details of the accounts.[190] In this method,
which works best with the modifications suggested by Drawe (see below)
5 grammes of cocoa or chocolate are defatted with ether and dried, then
ground in a mortar after a little water has been added, and levigated
with about 100 ccm of water in a cylinder. The liquid is allowed to
rest for some time and the suspended matter poured off almost to the
sediment, which is again shaken up with fresh water, allowed to settle,
and the operation repeated until all the fine particles have been
floated off and the water over the sediment no longer becomes cloudy,
but remains clear after the coarse and heavy particles have settled
down.
The powdery sediment is collected on a watch glass, dried in the water
bath, and after being cooled down in a desiccator, weighed. The weighed
residue is then softened with caustic soda and glycerine and examined
under the microscope. The presence of any cotyledon particles must be
carefully observed, such as have escaped separation in the grinding
and levigation, and whether particles of husk or epidermis or germ
preponderate. With proper levigation only traces of cacao substance,
especially here recognisable by the cacao starch, should be present.
The sand, which always adheres to the shells in the fermenting and
drying operations, is also easily recognised and many indications as to
the nature of the article under investigation can be noted by the use
of a simple magnifying glass applied to the washed residue on the watch
glass before drying.
Examined in that way, a sample of so-called Cocoas from unshelled beans
gave from 6 to 8 percent of husk; usually good cacao powder shows
a maximum of 2·5% husk. It is true that from this Filsinger-Drawe
procedure the correct percentage of shell can only be estimated in
very rare instances, for when it is necessary to be absolutely fair to
all concerned in the manufacture, the cacao must be so often washed
until no grains of cacao starch are visible under the microscope; and
so the result is often too small, more especially in the case of the
finer qualities. But when all particles of starch have been removed,
the finer particles of shell have often been taken along with them. Yet
when the residue certainly exceeds the standard percentage of shell, it
may be taken for granted that adulteration with husks has been carried
to excess, or that the cleansing processes have not been effectively
carried out. There is no other method which yields the same degree of
certainty.
The result obtained by the levigation method can be controlled by the
previously mentioned methods of Weender or Filsinger, as well as by the
determination of any silica in the ash (page 256).
Latterly the admixture of cacao husk with the cheaper kinds of cocoa
powder has largely increased, therefore the determination of the amount
of husk in cacao preparations has become of special importance.
10. ~Determination of sugar.~ There are three methods for the
quick determination of sugar, two of them polarimetric and the third
consisting of taking the specific gravity of the solution obtained
by shaking up the cacao with water. It is as well to note that in
all these methods the result includes the normal amount of sugar in
cacao, which Welmans[191] gives at 0·75-2 in cocoa and 0·4-1·0
percent in chocolate. That source of error is of no special significance,
for, as Welmans has shown, it is compensated for in the course of the
succeeding operations, so that these methods are of service.
For official investigations under this head the statutes of May 31st
1891 and May 27th 1896 respectively together with the instructions
issued by the council concerning the carrying out of the process
(Berlin, July 9th 1896, and Nov. 8th 1897, E) constitute a standard.
They read as follows: “Half the normal weight (13·024 g) of chocolate
is damped with alcohol and then warmed for 15 minutes with 30 ccm
of water on the water bath. While still hot, it is poured on to a
wet filter, the residue again treated with hot water, and until the
filtrate nearly amounts to 100 ccm. The filtrate is to be mixed with 5
ccm of basic lead acetate solution, allowed to stand for a quarter of
an hour, then clarified with alum and a little alumina, made up to a
definite volume (110 ccm) and polarised.” But it is to be noted that
these instructions are not exhaustive enough, and prove particularly
deficient as regards the employment of water, also through their
non-observation of the errors which can arise in using basic lead
acetate, though it is true that these are only of a minor character.
The Berlin chemist Jeserich (ex officio) had a rather hot dispute with
the official over the matter, who declared that his results were false
in spite of all protest, until he finally proved that it was not these
results but the process advised by law which lacked correctness. He
described the rencontre in very lucid if drastic detail to an assembly
of official chemists.
Something similar happened to the present editor, who in his office
of sworn chemist was called upon to determine the amount of sugar and
starch present in certain crumb chocolates on the one hand, and the
amount of cacao material on the other. As the official inspectors
insist on their prescriptions being carried out with scrupulous
exactitude, he found it necessary to give a double result, the one in
accordance with these prescriptions, and the other when double the
amount of water was used, taking care to explain the whole matter
at length. But it occasioned some surprise, and finally the task of
investigating and testing was withdrawn and given to another.
Another polarimetric method, recommended by Woy[192], is carried out
as follows. Two portions of half the normal weight (13·024 grammes) of
rasped or shaved chocolate are placed in 100 ccm and 200 ccm flasks
respectively, moistened with alcohol, then treated with hot water and
stirred up till the sugar is dissolved. 4 ccm of basic lead acetate
solution are added to each flask, by which means the chocolate in
suspension loses its viscosity. After being cooled, the solutions are
made up to the marks, well mixed and filtered. Two quickly filtering
liquids are thus obtained, which are then polarised in 200 mm tubes.
With chocolate containing meal, the temperature must not exceed 50° C.
From the two polarisations, the following equation results: a (100-x)
= b(200-x), in which _a_ and _b_ are the results of polarising, and x
the volume of the insoluble substances, including the lead precipitate,
contained in the half normal weight. The product of the equation gives
the amount of sugar present. Woy’s method has the great advantage of
avoiding the error due to the volume of the undissolved cacao and lead
precipitate.
The third method, as adopted by Zipperer[193], is as follows: 50
grammes of chocolate, finely divided with an iron grater or rasp, are
treated with exactly 200 ccm of cold water, frequently stirred for 4
hours, then poured on to a previously moistened and well wrung pointed
bag. The specific gravity of the filtrate is taken in an araeometer,
specially constructed for the purpose by Greiner of Munich on lines
suggested by Zipperer himself. On the scale of the araeometer is given
the percentage amount of sugar in the chocolate, from 5 to 5 percent,
with subdivisions of one percent, so that the reading can be quickly
taken, without correction.
In the determination of sugar by weight, the chocolate is first
defatted with ether, the sugar extracted with alcohol, then inverted,
the inverted solution treated with Fehling’s solution and the copper
precipitate weighed. The process has little to recommend it, being
troublesome and admitting of a large margin of errors.
Here again much has been written of late[194] concerning the two former
methods, their liabilities to error and the avoidance of these, yet
without bringing to light anything which calls for a specially detailed
treatment in this book.
11. ~Determination of Albuminates.~ The determination of albumin
is frequently required in the analysis of cacao powder and is necessary
to the ascertainment of its nutritive value. The determination of
nitrogen is determined by mixing 0·5 grammes of finely powdered bean
with soda lime and burning the mixture in a tube. (This determination
of nitrogen is a necessary part of the process.) Thus ammonia is
formed, which is passed through a known quantity of sulphuric acid.
When the combustion is finished, the acid solution is titrated with a
standard solution of barium hydroxide, and from the quantity consumed
the percentage of nitrogen is calculated. But as the diureides also
contain nitrogen (31·1 % of the theobromine and caffeine present) the
nitrogen corresponding to this amount must be deducted from the total
quantity of nitrogen yielded by combustion and the remainder multiplied
by 6·25 will indicate the amount of albumen present as a constituent.
Another and better method of determining the nitrogen is by
Kjeldahl’s[195] process. It has been frequently subjected to
modifications, but was originally carried out as follows. 0·25 grammes
of the nitrogenous substance (cacao preparation) is heated on the
sand bath together with 20 ccm of concentrated sulphuric acid and a
little quicksilver, till the solution becomes colourless or only of a
very pale yellow. After diluting with about 200 ccm of water, it is
made alkaline by the addition of soda lye (which must of course be
entirely free from nitrogen, the same remark applying to the sulphuric
acid used) and, potassium or sodium sulphide being added, it is then
distilled, and the ammonia given off collected and determined as above
described. As this method also determines the total amount of nitrogen,
an allowance must be made for the nitrogen in the theobromine and
caffeine before multiplying the result by 6·25. This modification is
still to be recommended as the best and most reliable.
In rare cases an excessive amount of albumen may be due to the
admixture of earth-nut cake or gelatine. As to the detection of the
latter adulteration, see page 254. Bileryst[196] says that earth-nut
cake can be recognised by its high percentage of albumen content,
amounting to between 45 and 47 percent.
12. ~Investigation of Milk and Cream Chocolate.~ The tests bearing
on these products really constitute a chapter in themselves, which has
acquired special importance owing to the great popularity they enjoy
and the consequently greatly increased production. According to the
unanimous opinion of the Association of German Chocolate Manufacturers
and the Free Union of German Food Chemists, expressed when considering
the respective claims of such chocolates, it is chiefly if not
exclusively a matter of determining the percentage of milk or cream,
which ought not to be below 12·5 or 10%, always supposing the milk
or cream to be a substitute for sugar, and this means therefore that
the quantity of cacao material in the chocolate product should on
no account sink below 32%. (Cf. p. 283 No. 3. Abs. 5.) The method
employed in the investigation is generally the same as that suggested
by Laxa in his treatise on “Milk Chocolates”[197] although it has been
considerably improved by Baier and his colleagues.[198] It is here a
matter of working backwards from the determination of the fatty and
nitrogenous components (or caseine) to the amount of milk or cream in
the chocolate. This presents a certain amount of difficultly as it
is not only necessary to determine the milk, but also to establish
that neither skimmed or whipped material (either in part or entirely)
has been employed. Yet it is possible here to proceed with absolute
certainty, as Baier[199] convincingly demonstrates, by taking into
consideration the relative proportion of milk fat, called caseum or
caseine.
If it is desired finally to characterise the respective chocolates,
determinations of the ~quantity~ of milk fat present and the
amount of milk product used become essential. Baier gives both as
calculable (cf. footnote 1)[200], the Reichert-Meissl number of the
total fat being ascertained, and from this, subtracting the R.-M.
number of the cacao fat present[201] the quantity of milk fat, finally
the amount of caseine, milk sugar, mineral matter and other factors.
No details of this somewhat extensive calculation are proved in the
original.[202] We give the following regulations (Laxa-Baier) for
carrying out the determination of the caseine, together with the
necessary formula.
20 grammes of fine divided chocolate are loosely introduced into a
Soxhlet’s extracting apparatus, and there extracted with ether for a
period of 16 hours. Of the residue, 10 grammes are used for testing in
connection with caseine, and this after the ether has evaporated. These
are mixed up in mortar with gradual and even addition of a 1% solution
of sodium oxalate, so that no lump formations occur, and then brought
into a marked carboy of 250 ccm capacity, until 200 ccm of the sodium
oxalate solution have been used. The carboy is then provided with an
asbestos net, and heated by means of a flame from the under side, until
its contents are brought to boil. The mouth of the carboy is covered
with a small funnel which has been hermetically sealed at its narrower
end. Then boiling oxalate solution is poured into the vessel up the
bend, and it is then allowed to stand over till another day, shaking
however being often repeated, then filled with sodium oxalate solution
up to the mark, agitated with a regular motion, and then filtered
through an ordinary filter. To 100 ccm of this solution 5 ccm of an
uranous acetate solution (5% strong) and drop by drop and with repeated
stirring a 30% solution of acetic are added until there is a deposit.
(This will require from 30 to 120 drops, according to the amount of
caseine present.) Then an extra 5 drops of acetic acid can be added.
This causes the deposit to stand out clearly from the liquid matter
and it can be readily separated by centrifugalising. Afterwards it can
be washed out with 100 ccm of solution, of which 5 ccm are uranous
acetate and 3 ccm acetic acid 3 % strong, until the sodium oxalate
can no longer be seen on adding calcium chloride (i. e. after about
three repeated centrifugalisations). The contents of the tube are then
rinsed on to the small filter by means of the wash fluid, stirred in a
Kjeldahl carboy with concentrated sulphuric acid and copper oxide, and
the quantity of nitrogen found converted into caseine by multiplying
with the factor k = 6·37.—Bearing in mind the quantity of fat, the
percentage of caseine in the original chocolate is calculated.[203]
In the following:
b = signifies the total of fatty content of the chocolate[204],
a = the Reichert-Meissl number of the total fat,
and K = the amount of caseine as established by the Laxa-Baier method
(nitrogen contents times 6·37).
(a - 1) b Further: 1. The desired quantity of fat
1. F = ————————- equal is to the R.M.N. a, of the total fat
27 less that of the cacao fat (1·0) multiplied
by the total amount of fat and divided by the
average R.M.N. for butter fats = 27.
2. a) E = 1·11 K 2. The total amount of albumen E is
equal to the amount of caseine K multiplied
1·11 K · 132 by 1·11, as this constitutes about 90% of
b) M = ———————————— the former; and as the albumen E, milk sugar
100 M and the mineral constituents A (Ash V)
are present in the milk in the proportion of
1·11 K · 21·4 100 : 132 : 21·4, this yields the formula
c) A = ————————————- given in b & c.
100
3. T = F + E + M + A 3. The total quantity of milk stuff T is
equal to the total of fatty contents,
albumen, milk, sugar and ash.
4. x = Q · k 4. The fatty constituency of the original
milk or cream to be calculated from the
F formula x = Q times k, where Q is the
Q = ——- quotient resulting when the amount of fat
K
k = const.
In the case of milk: F is divided by that of caseine K, and k
k_{1} = 3·15 the normal caseine consistency of average
k_{2} = 3·05 milk preparation. Or it varies as the
k_{3} = 2·7 numbers k_{1} etc. indicate in the case of
k_{4} = 2·5 cream and so forth. Higher percentages
10% than those given do not come into
consideration.
c. Microscopic-botanical investigation.
[Illustration: Fig. 100.
_A._ Parenchyma of the cotyledon after removal of fat and treatment
with Iodine chloral hydrate, a: parenchyma cells with starch, b: with
cacao red.
_B._ Aleuron particles with globois (Molisch) from parenchyma cells.]
[Illustration: Fig. 101.
_A._ Mitscherlich particles.
_B._ Seed cells, above with starch bodies, underneath with violet
colouring matter (cacao red) lying in chloral.
_C._ Series of yeast germs.
_D._ Threads of extraneous growth.
_E._ Epidermis and layer of cells occurring on the outer shell
(enlarged 340 times).]
Cacao is to no great extent particularly characterised anatomically.
The parenchyma cells fig. 100 are chiefly to be noticed, containing
either fat, albumin (protoplasm) aleuron granules, pigment, or
cacao starch. The ~starch~, as already remarked, consists of
especially small globular granules, mostly separate, but also two
or three adherent. It is somewhat more difficult to gelatinise than
other kinds of starch, and it is coloured blue by iodine somewhat more
slowly than many other starch granules, especially in the preparations
containing fat. Cacao preparations which have been disintegrated by
fixed alcalis, differ in this respect; according to Welmans, iodine
first forms colourless iodine compounds, and not until the alkali has
been saturated, is the blue colour developed. In such cases, care
must be taken, that an excess of iodine is present. In estimating the
amount of foreign starch, great care must be taken that the conspicuous
bluish-black granules of the foreign starch, which immediately strike
the eye, are not over estimated, which may easily occur. For control
observations, mixtures containing various known amounts of starch
should be tried comparatively. The pigment cells and the epidermis
with the Mitscherlich’s particles (figs 101 and 102) should be noticed
as well as the characteristic globoids, which occur in the ash of the
cotyledon tissues (compare page 67). The ~outside shell~, more
or less woody according to the origin of the bean, consists of four
layers of cells; this is best recognised by the large cells of the
principal tissue, which are distinguished by their form as well as by
their thickened side walls from the tissue of the cotyledon. Another
characteristic of this layer consists of the large number of coarse
spiral vessels, which exceed those of the seed lobes in size, and
finally, the inner elements of the stone cell layer, which, however,
on account of their limited development are seldom to be discovered.
The smooth, fine brown coloured, and light refracting fragments, which
frequently appear quite structureless and have their fibrous character
made perceptible only after treatment with caustic alkali, must be
regarded as characteristic of the inner part of the husk or the seed
membrane. The best observing medium is a solution of chloral hydrate or
almond oil, as well as dilute sulphuric acid and glycerine.[205] The
substance is always to be defatted with ether, before the microscopical
examination. A complete extraction of the fat, according to Welmans,
can occur only with exceedingly thin cuttings, in which every cell of
the section would be operated on, or in powdered preparations, when the
cells have been completely torn asunder by mechanical pulverisation.
The fat is not extracted by solvents from intact cells, as the cell
walls are impermeable by them.[206]
The detection of ~foreign starch~ is possible only by use of
the microscope; by means of standard preparations an approximate
estimate may be made as to the amount and kind of meal added.[207] The
examination of starch is especially facilitated by H. Leffmann and
W. Beam’s[208] centrifugal method: the sample suspended in water is
subjected to rotation for a short time in the centrifugal apparatus.
The presence of foreign starch is shown by a white layer in the
resulting sediment. This layer can be collected and microscopically
examined for foreign starch and husk. In the case of cacao
preparations, it is always well to distinguish between unimportant
traces and quantities that justify objection.[209]
[Illustration: Fig. 102.
_A._ Silver membrane with the hairs (Mitscherlich particles) _tr_, and
the crystals _f_ and _K_.
_B._ Cocoa powder: _c_ Cotyledon tissue with cells of fat and
colouring matter, _p_ shell parenchyma, _sp_ speriods, _d_ layer of
dry cells.]