Part 14

Copper. Invert Sugar. Dextrose. Levulose. Galactose. Arabinose. Milli- Milli- Milli- Milli- Milli- Milli- grams. grams. grams. grams. grams. grams. 50 15.2 15.6 14.7 17.4 17.0 55 16.6 17.0 16.1 19.1 18.6 60 18.0 18.5 17.5 20.8 20.3 65 19.4 19.9 18.9 22.5 21.9 70 20.8 21.4 20.3 24.2 23.5 75 22.3 22.9 21.7 25.9 25.1 80 23.7 24.4 23.0 27.7 26.7 85 25.2 25.8 24.3 29.3 28.3 90 26.6 27.3 25.7 31.1 29.9 95 28.1 28.8 27.1 32.8 31.5 100 29.5 30.3 28.5 34.5 33.1 105 31.0 31.8 29.9 36.2 34.7 110 32.4 33.3 31.2 38.0 36.3 115 33.9 34.8 32.6 39.7 37.9 120 35.3 36.3 34.0 41.4 39.5 125 36.8 37.8 35.4 43.1 41.1 130 38.2 39.3 36.8 44.8 42.8 135 39.7 40.8 38.2 46.5 44.4 140 41.1 42.3 39.6 48.3 46.0 145 42.6 43.8 41.0 50.0 47.6 150 44.0 45.3 42.5 51.8 49.3 155 45.5 46.8 43.9 53.6 50.9 160 47.0 48.3 45.3 55.4 52.6 165 48.5 49.8 46.7 57.2 54.3 170 50.0 51.4 48.1 59.0 55.9 175 51.5 52.9 49.5 60.8 57.5 180 53.0 54.5 51.0 62.7 59.2 185 54.5 56.0 52.5 64.5 60.9 190 56.0 57.6 54.0 66.4 62.7 195 57.5 59.2 55.5 68.3 64.4 200 59.1 60.8 57.0 70.3 66.2 205 60.7 62.4 58.6 72.3 68.0 210 62.4 64.1 60.2 74.3 69.8 215 64.1 65.8 61.8 76.3 71.6 220 65.8 67.5 63.5 78.3 73.5 225 67.5 69.2 65.2 80.3 75.4 230 69.3 70.9 66.9 82.4 77.3 235 71.1 72.7 68.7 84.5 79.3 240 72.9 74.5 70.6 86.6 81.3 245 74.8 76.4 72.5 88.9 83.4 250 76.7 78.4 74.4 91.2 85.5 255 78.6 80.5 76.5 93.5 87.6 260 80.5 82.8 78.8 95.9 89.8 265 82.5 85.1 81.1 98.3 92.2 270 84.7 87.5 83.5 100.7 94.6 275 87.1 89.9 85.9 103.3 97.1 280 89.7 92.4 88.6 106.1 99.6 285 92.3 94.9 91.3 109.0 102.3 290 95.1 97.6 94.2 112.0 105.1 295 98.0 100.4 97.2 115.1 107.9 298 100.0 102.5 99.0 117.0 109.5

VOLUMETRIC METHODS BASED UPON THE USE OF AN AMMONIACAL COPPER SOLUTION.

=129. Pavy’s Process.=—The well-known solubility of cuprous oxid in ammonia led Pavy to adopt a copper reagent containing ammonia in the volumetric determination of reducing sugars.[95] In Pavy’s process an alkaline copper solution is employed made up in the usual way, to which a sufficient quantity of ammonia is added to hold in solution all the copper when precipitated as cuprous oxid. The solution used by Pavy has the following composition: One liter contains

Crystallized copper sulfate 34.65 grams Potassium-sodium tartrate 173.00 ” Caustic potash 160.00 ”

For use 120 cubic centimeters of the above reagent are mixed with 300 of ammonia of specific gravity 0.88, and the volume completed to one liter with distilled water. Twenty cubic centimeters of this reagent are equivalent to ten milligrams of dextrose or invert sugar when added in a one per cent solution.

In the use of ammoniacal copper solution, care must be taken that all the liquids employed be entirely free of oxygen and that the contents of the flask in which the reduction takes place be in some way excluded from contact with the air. Pavy secured this by conducting the reduction in a flask closed with a stopper carrying two holes; one of these served for the introduction of the burette carrying the sugar solution and the other carried a tube dipping into a water seal by means of a slit rubber tube, which would permit of the exit of the vapors of steam and ammonia, but prevent the regurgitation of the water into the flask.

The complete decoloration of the copper solution marks the end of the reaction. The usual precautions in regard to the length of the time of boiling must be observed.

It is easy to see that in the Pavy process the quantity of ammonia in the solution is rapidly diminished during the boiling and this has led to the suggestion of other methods to exclude the air. Among these have been recommended the introduction of a current of hydrogen or carbon dioxid. One of the best methods of procedure is that proposed by Allen, who recommends covering the copper solution by a layer of paraffin oil (kerosene).[96]

=130. Process Of Peska.=—Peska has also independently made use of Allen’s method of covering the solutions with a layer of paraffin oil and finds it reliable.[97] The copper reagent employed by him has the following composition:

Crystallized copper sulfate 6.927 grams Ammonia, twenty-five per cent strength 160.00 cc.

The copper sulfate is dissolved in water, the ammonia added, and the volume completed to half a liter with distilled water. A second solution containing half a liter is made by dissolving 34.5 grams of potassium-sodium tartrate and ten grams of sodium hydroxid and completing the cool solution to half a liter with distilled water. In all cases the water used in making up the above solutions must be freshly boiled to exclude the air.

For the titration, fifty cubic centimeters of each of the above solutions are taken, mixed and covered with a layer of paraffin oil half a centimeter in depth. The reduction is not accomplished at a boiling temperature, but at from 80° to 85°. The manipulation is conducted as follows:

The mixed solutions are placed in a beaker, covered with oil, and heated to 80°. The temperature is measured by a thermometer which also serves as a stirring rod. The sugar solution is run down the sides of the beaker from a burette of such a shape as to be protected from the heat. After each addition of the sugar solution the mixture is carefully stirred, keeping the temperature at from 80° to 85°. The first titration is made to determine approximately the quantity of sugar solution necessary to decolorize the copper. This done, the actual titration is accomplished by adding at once the total amount of sugar solution necessary to decolorize, less about one cubic centimeter. Any sugar solution adhering to the side of the beaker is washed down by distilled water, the contents of the beaker well stirred, and the temperature kept at 85° for two minutes. The rest of the sugar solution is then added in quantities of one-tenth of a cubic centimeter until the decoloration is completed. The total time of the final titration should not exceed five minutes. The sugar solution should be as nearly as possible of one per cent strength. If a lower degree of strength be employed a larger quantity of the sugar is necessary to reduce a given quantity of copper.

In the case of dextrose, when a one per cent solution is used, eight and two-tenths cubic centimeters, corresponding to 80.2 milligrams of dextrose, are required to reduce 100 cubic centimeters of the mixed reagent. On the other hand, when the sugar solution is diluted to one-tenth of a per cent strength 82.1 milligrams are required.

With invert sugar slightly larger quantities are necessary, the reducing power being as 94.9 to 100 as compared with dextrose. With a one per cent strength of invert sugar it is found that eighty-four milligrams are required to reduce 100 cubic centimeters of the mixed reagent and when the strength of the invert sugar is reduced to one-tenth per cent 87.03 milligrams are required.

=131. Method Of Allein and Gaud.=—Allein and Gaud have proposed a further modification of the ammonia process which consists essentially in the suppression of rochelle salt and fixed caustic alkali and the entire substitution therefor of ammonia. Ammonia acts with much less vigor upon sugars than the caustic alkalies, and it is therefore claimed that the decomposition of the sugar due to the alkali is reduced to a minimum when ammonia is employed.[98] The copper solution is made as follows:

Dissolve 8.7916 grams of electrolytic copper in ninety-three grams of concentrated sulfuric acid diluted with an equal volume of water. Complete the resulting solution to one liter with concentrated ammonia. Ten cubic centimeters of this solution are equal to fifty grams of dextrose.

It is recommended that the reduction be accomplished in an atmosphere of hydrogen, but it is apparent that the use of kerosene is permissible in this case, and on account of its greater simplicity it is to be recommended as the best means of excluding the oxygen. The reduction is accomplished at a temperature of about 80°.

It is also proposed to reoxidize the copper by substituting a current of air for the hydrogen at the end of the reaction, and thus use the same copper a number of times. The danger of loss of ammonia, and the difficulty of determining when the oxidation is complete, render this regeneration of the reagent undesirable.

=132. Method of Gerrard.=—The method of Gerrard does not depend upon the use of ammonia, but the principle involved is the same, _viz._, the holding of the separated cuprous oxid in solution and the determination of the end of the reaction by the disappearance of the blue color. As first proposed by Gerrard, the copper sulfate solution is made of double the strength usually employed and to each 100 cubic centimeters thereof, before use, three and three-tenths grams of potassium cyanid are added. This is sufficient to hold the precipitated cuprous oxid in solution.[99]

The original method of Gerrard is found difficult of execution and the author, in conjunction with Allen, has lately modified it and reduced it to a practical working basis.[100]

In the new method the ordinary fehling solution is employed and it is prepared for use in the following way: Ten cubic centimeters of the fehling solution, or half that quantity of each of the component parts kept in separate bottles, are placed in a porcelain dish with forty cubic centimeters of water and brought to the boiling-point. To the boiling liquid is added, from a pipette, a five per cent solution of potassium cyanid until the blue color just disappears, or only a very faint tint of blue remains, avoiding any excess of the cyanid. A second portion of the fehling solution equal to that first employed is added, and to the boiling mixture the solution of sugar is added, from a burette, until the blue color disappears. The contents of the dish should be kept boiling during the addition of the sugar solution. The volume used will contain fifty milligrams of dextrose. The sugar solution should be of such a strength as to contain no more than half a per cent of reducing sugar.

The principle of the preparation of the solution may be stated as follows: If to a solution of copper sulfate, potassium be added until the blue color disappears, a double cyanid of copper and potassium cyanid is formed according to the following reaction: CuSO₄ + 4KCN = Cu(CN)₂.2KCN + K₂SO₄. This double cyanid is a salt of considerable stability. It is not decomposed by alkalies, hydrogen or ammonium, sulfid. With mineral acids it gives a whitish, curdy precipitate. With fehling solution the same double cyanid is formed as that described above. If, however, fehling solution be present in excess of the amount necessary to form the double cyanid of copper, this excess can be used in the oxidation of reducing sugar and the colorless condition of the solution will be restored as soon as the excess of the fehling is destroyed. The double cyanid holds in solution the cuprous oxid formed and thus complete decoloration is secured.

=133. Sidersky’s Modification of Soldaini’s Process.=—In all cases where the sugar solutions are not too highly colored, Sidersky finds that the method of reduction in a large test tube, as practiced by Violette, is applicable with the copper carbonate solution.[101] For more exact work it is preferred to determine the quantity of copper reduced by an indirect volumetric method. The sugar solution, properly clarified and the lead removed if subacetates have been used, is made of such a volume as to contain less than one per cent of reducing sugars. In a flask or large test tube are placed 100 cubic centimeters of the copper solution, which is boiled for a short time and the sugar solution added, little by little, from a pipette, at such a rate as not to stop the ebullition. The boiling is continued for five minutes after the last addition of the sugar. The vessel is taken from the flame and 100 cubic centimeters of cold water added, the whole brought on an asbestos felt and the cuprous oxid washed with hot water until the alkaline reaction has disappeared. The residual cuprous oxid is dissolved in a measured quantity of set sulfuric acid, semi- or fifth-normal, a few particles of potassium chlorate added, and the mixture boiled to convert any cuprous into cupric sulfate. The reaction is represented by the following formula:

3Cu₂O + 6H₂SO₄ + KClO₃ = 6CuSO₄ + KCl + 6H₂O.

The residual sulfuric acid is titrated with a set alkali in excess, ammonia being preferred.

The solution of ammonia is made by diluting 200 cubic centimeters of commercial aqua ammonia with 800 of water. Its strength is determined by adding a little copper sulfate solution as indicator and then the set solution of sulfuric acid until the blue color disappears. The copper sulfate secured from the cuprous sulfate as described above is cooled, and a quantity of the ammonia, equal to twenty-five cubic centimeters of the set sulfuric acid, added. The excess of the ammonia is then determined by titration with the sulfuric acid, the disappearance of the blue color being the indication of the end of the reaction. The number of cubic centimeters of the set sulfuric acid required to saturate the ammonia represents the equivalent of cuprous oxid originally present. One cubic centimeter of normal sulfuric acid is equivalent to 0.0317 gram of metallic copper.

To determine the weight of invert sugar oxidized, multiply the weight of copper, calculated as above described, by the factor 0.3546.[102] For a general application of this method of analysis the relative quantities of copper reduced by different quantities of sugar must be taken into consideration.

While, as has already been stated, the copper carbonate process has heretofore been applied chiefly to the detection of invert sugar, it has merits which justify the expectation that it may some time supplant the fehling liquor both for volumetric and gravimetric work. Large volumes of the reagent can be prepared at once and without danger of subsequent change. The action of the reagent on the hexobioses and trioses is far less vigorous than that of the alkaline copper tartrate, and the end reactions for volumetric work are, at least, as easily determined in the one case as the other.

=134. Method Depending on Titration of Excess of Copper.=—Instead of measuring the quantity of copper reduced, either by its disappearance or by reducing the cuprous oxid to a metallic state, Politis has proposed a method of analysis depending on the titration of the residual copper.[103] The reagents employed are:

(1) A copper solution containing 24.95 grams of crystallized copper sulfate, 140 grams of sodium and potassium tartrate, and twenty-five grams of sodium hydroxid in one liter:

(2) A solution of sodium thiosulfate containing 24.8 grams of the salt in one liter:

(3) A solution of potassium iodid containing 12.7 grams of iodin in one liter.

The reaction is represented by the formula

2CuCl₂ + 4KI = Cu₂I₂ + 4KCl + I₂.

The analytical process is carried out as follows: In a 100 cubic centimeter flask are boiled fifty cubic centimeters of the copper solution, ten cubic centimeters of about one-tenth per cent reducing sugar solution are added, the boiling continued for five minutes, the flask filled to the mark with boiling water and its contents filtered. Fifty cubic centimeters of the hot filtrate are cooled, slightly acidified, potassium iodid solution added in slight excess; and the iodin set free determined by titration with sodium thiosulfate. The quantity of iodin obtained corresponds to the unreduced copper remaining after treatment with the reducing sugar. The number of cubic centimeters of thiosulfate used subtracted from twenty-five will give the number of cubic centimeters of the copper solution which would be reduced by five cubic centimeters of the sugar solution used.

_Example._—In the proportions given above it was found that eleven cubic centimeters of thiosulfate were required to saturate the iodin set free. Then 25 - 11 = 14 cubic centimeters of copper solution reduced by five cubic centimeters of the sugar solution. Since one cubic centimeter of the copper solution is reduced by 0.0036 gram of dextrose the total dextrose in the five cubic centimeters = 0.0036 × 5 = 0.0180 gram.

The above method does not seem to have any practical advantage over those based on noting the disappearance of the copper and is given only to illustrate the principle of the process. While the titration of the iodin by sodium thiosulfate is easily accomplished in the absence of organic matter, it becomes difficult, as shown by Ewell, when organic matters are present, as they always are in the oxidation of a sugar solution. Ewell has therefore proposed to determine the residual copper by a standard solution of potassium cyanid, but the method has not yet been developed.[104]

GRAVIMETRIC COPPER METHODS.

=135. General Principles.=—In the preceding pages the principles of the volumetric methods of sugar analysis by means of alkaline copper solution have been set forth. They depend either on the total decomposition of the copper solution employed by the reducing sugar, or else on the collection and titration of the cuprous oxid formed in the reaction. In the gravimetric methods the general principle of the process rests upon the collection of the cuprous oxid formed and its reduction to metallic copper, the weight of which serves as a starting point in the calculations of the weight of reducing sugar, which has been oxidized in the solution.

The factors which affect the weight of copper obtained are essentially those which influence the results in the volumetric method. The composition of the copper solution, the temperature at which the reduction is accomplished, the time of heating, the strength of the sugar solution and the details of the manipulation, all affect more or less the quantity of copper obtained. As in the volumetric method also, the kind of reducing sugar must be taken in consideration, dextrose, levulose, invert sugar, maltose and other sugars having each a definite factor for reduction in given conditions. It follows, therefore, that only those results are of value which are obtained under definite conditions, rigidly controlled.

=136. Gravimetric Methods of the Department of Agriculture Laboratory.=—The process used in this laboratory is based essentially on the methods of Maercker, Behrend, Morgen, Meissl, Hiller and Allihn.[105] Where dextrose alone is present, the table of factors proposed by Allihn is used and also the copper solution corresponding thereto.

For pure invert sugar, the tables and solutions of Meissl are used. For invert sugar in the presence of sucrose, the table and process proposed by Hiller are used.

The reduction of the copper solution and the electrolytic deposition of the copper are accomplished as follows:

The copper and alkali solutions are kept in separate bottles. After mixing the equivalent volume of the two solutions in a beaker, heat is applied and the mixture boiled. To the boiling liquid the proper volume of the cold sugar solution is added. This must always be less than the amount required for complete reduction. The solution is again brought into ebullition and kept boiling exactly two minutes. A two-minute sand glass is conveniently used to determine the time of boiling. At the end of this time an equal volume of freshly boiled cold water is added, and the supernatant liquor at once passed through a gooch under pressure. The residual cuprous oxid is covered with boiling water and washed by decantation until the wash water is no longer alkaline. It is more convenient to wash in such a way that, at the end, the greater part of the cuprous oxid is in the gooch. The felt and cuprous oxid are then returned to the beaker in which the reduction is made. The gooch is moistened with nitric acid to dissolve any adhering oxid and then is washed into the beaker. Enough nitric acid is added to bring all the oxid into solution, an excess being avoided, and a small amount of water added. The mixture is again passed under pressure through a gooch having a thin felt, to remove the asbestos and the filtrate collected in a flask of about 150 cubic centimeters capacity. The washing is continued until the gooch is free of copper, when the volume of the filtrate should be about 100 cubic centimeters. The liquid is transferred to a platinum dish holding about 175 cubic centimeters and the flask washed with about twenty-five cubic centimeters of water. From three to five cubic centimeters of strong sulfuric acid are added and the copper deposited by an electric current.