Cane sugar, plus the water of hydrolysis, will yield theoretically 53.8
per cent of alcohol and 51.5 per cent of carbon dioxid.
In practice the theoretical proportions of alcohol and carbon dioxid
are not obtained because of the difficulty of excluding other
fermentative action, resulting in the formation especially of succinic
acid and glycerol. Moreover, a part of the sugar is consumed by the
yeast cells to secure their proper growth and development. In all
only about ninety-five per cent of the sugar can be safely assumed as
entering into the production of alcohol. About 48.5 per cent of alcohol
are all that may be expected of the weight of dextrose or invert sugar
used. Only sugars containing three molecules of carbon or some multiple
thereof are fermentable. Thus the trioses, hexoses, nonoses, etc., are
susceptible of fermentation, while the tetroses, pentoses, etc., are
not.
=159. Estimating Alcohol.=—In the determination of sugar by
fermentation, a rather dilute solution not exceeding ten per cent
should be used. A quantity of pure yeast, equivalent to four or five
per cent of the sugar used, is added, and the contents of the vessel,
after being well shaken, exposed to a temperature of from 25° to 30°
until the fermentation has ceased, which will be usually in from
twenty-four to thirty-six hours. The alcohol is then determined in the
residue by the methods given hereafter.
The weight of the alcohol obtained multiplied by 100 and divided by
48.5, will give the weight of the hexose reducing sugar which has been
fermented. Ninety-five parts of sucrose will give 100 parts of invert
sugar.
_Example._—Let the weight of alcohol obtained be 0.625 gram. Then 0.625
× 100 ÷ 48.5 = 1.289 grams, the weight of the hexose, which has been
fermented; 1.289 grams of dextrose or levulose correspond to 1.225 of
sucrose.
=160. Estimating Carbon Dioxid.=—The sugar may also be determined
by estimating the amount of carbon dioxid produced during the
fermentation. For this purpose the mixture of sugar solution and yeast,
prepared as above mentioned, is placed in a flask whose stopper carries
two tubes, one of which introduces air free of carbon dioxid into the
contents of the flask, and the other conducts the evolved carbon dioxid
into the absorption bulbs. In passing to the absorption bulbs the
carbon dioxid is freed of moisture by passing through another set of
bulbs filled with strong sulfuric acid. During the fermentation, the
carbon dioxid is forced through the bulbs by the pressure produced, or
better, a slow current of air is aspirated through the whole apparatus.
The aspiration is continued after the fermentation, has ceased, until
all the carbon dioxid is expelled. Towards the end, the contents of
the flask may be heated to near the boiling-point. The increase of
the weight of the potash bulbs will give the weight of carbon dioxid
obtained. A hexose reducing sugar will yield about 46.5 per cent of its
weight of carbon dioxid. The calculation is made as suggested for the
alcohol process.
The fermentation process has little practical value save in determining
sucrose in presence of lactose, as will be described in another place.
=161. Precipitation of Sugars in Combination with the Earthy
Bases.=—The sugars combine in varying proportions with the oxids
and hydroxids of calcium, strontium and barium. Sucrose especially,
furnishes definite crystalline aggregates with these bases in such
a way as to form the groundwork of several technical processes in
the separation of that substance from its normally and abnormally
associated compounds. These processes have little use as analytical
methods, but are of great value, as mentioned, from a technical point
of view.
=162. Barium Saccharate.=—This compound is formed by mixing the aqueous
solutions of barium hydroxid and sugar. The saccharate separates
in bright crystalline plates or needles from the warm solution,
as C₁₂H₂₂O₁₁BaO. One part of this precipitate is soluble in about
forty-five parts of water, both at 15° and 100°.
=163. Strontium Saccharates.=—Both the mono- and distrontium
saccharates are known, _viz._, C₁₂H₂₂O₁₁SrO + 5H₂O and C₁₂H₂₂O₁₁2SrO.
The monosalt may be easily secured by adding a few of its crystals to a
mixture of sugar and strontium hydroxid solutions.
The disaccharate is precipitated as a granular substance when from two
to three molecules of strontium hydroxid are added to a boiling sugar
solution. The reaction is extensively used in separating the sugar from
beet molasses.
=164. Calcium Saccharates.=—Three calcium saccharates are known in
which one molecule of sugar is combined with one, two and three
molecules of lime respectively.
The monosaccharate is obtained by mixing the sugar and lime in the
proper proportion and precipitating by adding alcohol.
The precipitate is partly granular and partly jelly-like, and is
soluble in cold water. The dicalcium compound is obtained in the same
way and has similar properties. Both, on boiling, with water, form the
trisaccharate and free sugar.
The tricalcium saccharate is the most important of these compounds, and
may be obtained directly by mixing freshly burned and finely ground
lime (CaO) with a very cold dilute solution of sugar.
The compound crystallizes with three or four molecules of water.
When precipitated as described above, however, it has a granular,
nearly amorphous structure, and the process is frequently used in the
separation of sugar from beet molasses.
In the laboratory but little success has been had in using even the
barium hydroxid as a chemical reagent, and therefore the reactions
mentioned above are of little value for analytical purposes. In
separating sugar from vegetable fibers and seeds, however, the
treatment with strontium hydroxid is especially valuable the sugar
being subsequently recovered in a free state by breaking up the
saccharate with carbon dioxid. The technical use of these reactions
also is of great importance in the beet sugar industry.
=165. Qualitive Tests for the Different Sugars.=—The analyst will
often be aided in examining an unknown substance by the application
of qualitive tests, which will disclose to him the nature of the
saccharine bodies with which he has to deal.
=166. Optical Test for Sucrose.=—The simplest test for the presence of
sucrose is made with the polariscope. A small quantity of the sample
under examination is dissolved in water, clarified by any of the usual
methods, best with alumina cream, and polarized. A portion of the
liquor is diluted with one-tenth its volume of strong hydrochloric acid
and heated to just 68°, consuming about fifteen minutes time in the
operation. The mixture is quickly cooled and again polarized in a tube
one-tenth longer than before used; or the same tube may be used and the
observed reading of the scale increased by one-tenth. If sucrose be
present the second reading will be much lower than the first, or may
even be to the left.
=167. Cobaltous Nitrate Test for Sucrose.=—Sucrose in solution may be
distinguished from other sugars by the amethyst violet color which it
imparts to a solution of cobaltous nitrate. This reaction was first
described by Reich, in 1856, but has only lately been worked out in
detail. The test is applied as follows:
To about fifteen cubic centimeters of the sugar solution add five cubic
centimeters of a five per cent solution of cobaltous nitrate. After
thoroughly mixing the two solutions, add two cubic centimeters of a
fifty per cent solution of sodium hydroxid. Pure sucrose gives by this
treatment an amethyst violet color, which is permanent. Pure dextrose
gives a turquoise blue color which soon passes into a light green. When
the two sugars are mixed the coloration produced by the sucrose is the
predominant one, and one part of sucrose in nine parts of dextrose can
be distinguished. If the sucrose be mixed with impurities such as gum
arabic or dextrin, they should be precipitated by alcohol or basic lead
acetate, before the application of the test. Dextrin may be thrown
out by treatment of the solution with barium hydroxid and ammoniacal
lead acetate. The reaction may also be applied to the detection of
cane sugar in wines, after they are thoroughly decolorized by means of
lead acetate and bone-black. The presence of added sucrose to milk,
either in the fresh or condensed state, may also be detected after the
disturbing matters are thrown out with lead acetate. The presence of
sucrose in honey may also be detected by this process. The reaction
has been tried in this laboratory with very satisfactory results. The
amethyst violet coloration with sucrose is practically permanent. On
boiling the color is made slightly bluish, but is restored to the
original tint on cooling. Dextrose gives at first a fine blue color
which in the course of two hours passes into a pale green. A slight
flocculent precipitate is noticed in the tube containing the dextrose.
Maltose and lactose act very much as dextrose, but in the end do not
give so fine a green color. If the solutions containing dextrose,
lactose and maltose be boiled, the original color is destroyed and a
yellow-green color takes its place. The reaction is one which promises
to be of considerable practical value to analysts, as it may be applied
for the qualitive detection of sucrose in seeds and other vegetable
products.[133]
=168. The Dextrose Group.=—In case the carbohydrate in question shows a
right-handed rotation and the absence of sucrose is established by the
polariscopic observation described above, the presence of the dextrose
group may be determined by the following test.[134]
Five grams of the carbohydrate are oxidized by boiling with from twenty
to thirty cubic centimeters of nitric acid of 1.15 specific gravity,
and then at gentle heat evaporated to dryness with stirring. If much
mucic acid be present, as will be the case if the original matter
contained lactose some water is added and the mixture well stirred,
and again evaporated to dryness to expel all nitric acid. The residue
should be of a brown color. The mass is again mixed with a little
water and the acid reaction neutralized by rubbing with fine-ground
potassium carbonate. The carbonate should be added in slight excess and
acetic acid added to the alkaline mixture, which is concentrated by
evaporation and allowed to stand a few days. At the end of this time
potassium saccharate has formed and is separated from the mother liquid
by pouring on a porous porcelain plate. The residue is collected,
dissolved in a little water and again allowed to crystallize, when it
is collected on a porous plate, as before, and washed by means of an
atomizer with a little aqueous spray until it is pure white and free of
any oxalic acid. The residual acid potassium saccharate may be weighed
after drying and then converted into the silver salt. The potash salt
for this purpose is dissolved in water, neutralized with ammonia and
precipitated with a solution of silver nitrate. The precipitate is
well stirred, collected on a gooch and washed and dried in a dark
place. It contains 50.94 percent of silver. All sugars which contain
the dextrose group yield silver saccharate when treated as above
described. Inulin, sorbose, arabinose and galactose yield no saccharic
acid under this treatment, and thus it is shown that they contain
no dextrose group. Milk sugar, maltose, the dextrins, raffinose and
sucrose yield saccharic acid when treated as above and therefore all
contain the dextrose group.
=169. Levulose.=—The levulose group of sugars, wherever it occurs, when
oxidized with nitric acid, gives rise to tartaric, racemic, glycolic
and oxalic acids, which are not characteristic, being produced also
by the oxidation of other carbohydrates. A more distinguishing test
is afforded by the color reactions produced with resorcin.[135] The
reagent is prepared by dissolving half a gram of resorcin in thirty
cubic centimeters each of water and strong hydrochloric acid. To the
sugar solution under examination an equal volume of strong hydrochloric
acid is added, and then a few drops of the reagent. The mixture is
gently warmed, and in the presence of levulose develops a fire-red
color. Dextrose, lactose, mannose and the pentoses do not give the
coloration, but it is produced by sorbose in a striking degree, and
also by sucrose and raffinose since these sugars contain the levulose
group.
=170. Galactose.=—The galactose which arises from the hydrolysis of
milk sugar is readily recognized by the mucic acid which it gives on
oxidation with nitric acid.[136] The analytical work is conducted
as follows: The body containing galactose or galactan is placed in
a beaker with about sixty cubic centimeters of nitric acid of 1.15
specific gravity for each five grams of the sample used. The beaker
is placed on a steam-bath and heated, with frequent stirring, until
two-thirds of the nitric acid have been evaporated. The residual
mixture is allowed to stand over night and the following morning is
treated with ten cubic centimeters of water, allowed to stand for
twenty-four hours, filtered through a gooch, and the collected matter
washed with twenty-five cubic centimeters of water, dried at 100° and
weighed. The mucic acid collected in this way will amount to about
thirty-seven per cent of the milk sugar or seventy-five per cent of the
galactose oxidized. Raffinose yields under similar treatment, about
twenty-three per cent of mucic acid, which proves that the galactose
group is contained in that sugar. Raffinose, therefore, is composed of
one molecule each of dextrose, levulose, and galactose.
=171. Invert Sugar.=—The presence of a trace of invert sugar
accompanying sucrose can be determined by Soldaini’s solution,
paragraph =124=, or by boiling with methyl blue.[137] Methyl blue is
the hydrochlorate of an ammonium base, which, under the influence of
a reducing agent, loses two atoms of hydrogen and becomes a colorless
compound. The test for invert sugar is made as follows: The reagent
is prepared by dissolving one gram of methyl blue in water. If the
sugar solution is not clear, twenty grams of the sugar are dissolved
in water clarified by lead subacetate, the volume completed to 100
cubic centimeters, and the solid matters separated by filtration. The
filtrate is made slightly alkaline with sodium carbonate to remove the
lead. A few drops of soda lye solution are then added and the mixture
thrown on a filter. To twenty-five cubic centimeters of the filtrate a
drop of the methyl blue solution is added, and a portion of the liquor
heated in a test tube over the naked flame. If, after boiling for one
minute, the coloration disappear, the sample contains at least 0.01
per cent of invert sugar; if the solution remain blue it contains none
at all or less than 0.01 per cent. The test may also be made with the
dilute copper carbonate solution of Ost described further on.
=172. Compounds with Phenylhydrazin.=—Many sugars may also be
qualitively distinguished by the character of their compounds with
phenylhydrazin. In general, it may be said that those sugars which
reduce fehling solution form definite crystalline compounds with the
reagent named. If a moderately dilute hot solution of a reducing sugar
be brought into contact with phenylhydrazin acetate, a crystalline
osazone is separated. The reaction takes place between one molecule of
the sugar and two molecules of the hydrazin compound, according to the
following formula: