mother liquors, removing the gummy impurities by precipitation with hot ethyl alcohol and evaporating the alcoholic extract.7

Recrystallization.-Crude arabinose which contains gums may be purified in the following manner: 300 g of the sugar is dissolved by heating with 100 ml of water; the solution is mixed with 750 ml of hot ethyl alcohol and 15 g of a decolorizing carbon. The hot solution is filtered, and the carbon residue is washed with 150 ml of hot ethyl alcohol. The hot filtrate is allowed to cool, and accidental seeding with crystalline arabinose is avoided. As the solution cools it separates into two phases, the lower more sirupy layer of which contains most of the impurities. After the solution reaches room temperature and the sirupy phase has settled, the alcoholic layer is decanted. The residue is extracted as before with 300 ml of hot methyl alcohol, followed by 300 ml of hot ethyl alcohol. The combined alcoholic extract is evaporated in vacuo to a volume of approximately 600 ml. Sometime before this volume is reached, crystallization takes place in the distillation flask. When the evaporation has reached the desired stage, the sirup is seeded with crystalline arabinose, placed in the refrigerator, and allowed to stand until a satisfactory crystal growth is obtained. The first crystals are usually about 60 percent of the arabinose content. By concentrating the mother liquors, additional crystalline sugar is obtained.

Relatively pure arabinose can also be recrystallized in the following manner: 300 g of arabinose is dissolved with 180 ml of water. The hot solution is mixed with 200 ml of hot methyl alcohol, and after the addition of 50 g of a decolorizing carbon the mixture is filtered rapidly while hot. The filtrate is diluted with 400 ml of methyl alcohol and 300 ml of ethyl alcohol, cooled to room temperature, seeded, and set aside to crystallize. After several days the resulting crystals are collected on a filter and washed with methyl alcohol. The yield is about 150 g. The sugar in the mother liquors is reclaimed by evaporating the liquors to a sirup (n=1.465), which, after mixing with about 5 volumes of methyl alcohol, yields additional crystalline sugar.

In 4-percent aqueous solution, B-l-arabinose gives [a]=+190.6° initially, which changes in the course of several hours to +104.5°.

NOTES

1 The method is that of Anderson and Sands [1] as modified by Isbell (heretofore unpublished work).

Mesquite gum is obtained from a plant (Prosopis juliflora and related species) widely distributed through the southwest. It may be purchased from the Martin Drug Co., Tucson, Ariz. Cherry gum may also be used with essentially the same procedure.

3 The gum requires about 24 hours to dissolve. Before starting the hydrolysis, it is well to filter the solution through cheese cloth in order to remove bark and other foreign matter.

This must be done slowly with small portions of calcium carbonate. Foaming may be reduced by the addition of a commercial antifoam agent or capryl alcohol. The exceptionally large quantity of decolorizing carbon is added to reduce foaming, which can be further reduced by the use of an antifoam agent such as capryl alcohol.

A mechanical stirrer is of considerable help in this procedure.

1l-Arabinose has been prepared from a number of other sources: Cherry gum [2], wheat and rye bran [3, 4], peach gum [5], Australian black wattle gun [6], and beet pulp [7]. Harding [8] gives a review of the sources.

REFERENCES

[1] E. Anderson and L. Sands, J. Am. Chem. Soc. 48, 3172 (1926), also, Organic Syntheses 8, 18 (1928).

[2] H. Kiliani, Ber. deut. chem. Ges. 19, 3029 (1886).

[3] W. Stone and B. Tollens, Liebigs Ann. Chem. 249, 238 (1888).

[4] E. Steiger and E. Schulze, Ber. deut. chem. Ges. 23, 3110 (1890).

[5] W. Stone, Am. Chem. J. 12, 435 (1890).

[6] W. Stone, Am. Chem. J. 17, 196 (1895).

[7] E. Zitkowski, Am. Sugar Ind. 13, 98 (1911).

[8] T. S. Harding, Sugar 24, 656 (1922).

3. CELLOBIOSE (4-(8-d-GLUCOPYRANOSIDO)-d-GLUCOSE)

Method. The octaacetyl cellobiose is prepared by the action of acetic anhydride and sulfuric acid on cotton or filter paper according to the method of Klein [1]. Fifty grams of pure cotton is put in a precipitating jar (about 12 cm in diameter and 25 cm high), which is cooled in an ice-and-salt bath. Acetic anhydride, 140 ml, is added and the jar covered with a watch glass. Sixty milliliters of acetic anhydride is cooled in an ice-and-salt bath and 28 ml. of sulfuric acid slowly added. When this solution has cooled, it is poured onto the cold mixture of cotton and acetic anhydride and the whole mixture worked to a homogeneous mass by stirring with a glass rod. The cotton gradually disintegrates. The mass is transferred to a 2-liter Erlenmeyer flask and after vigorous shaking and occasional heating on the steam bath, a reddish solution is obtained." Several such batches combined are allowed to stand at room temperature for 2 or 3 days while crystallization takes place. The crystalline mass is not poured into water, as described by Klein, but filtered on a large-size Büchner filter having small holes, and fitted with a double layer of ordinary filter paper. It is filtered dry and washed with ether. The product is now stirred in a dish with 95-percent alcohol, again filtered by suction, and finally recrystallized from 95-percent alcohol. The yield may reach 40 to 50 percent of the theoretical. In a 2.5-percent concentration in chloroform the specific rotation of the pure substance is [a]2+41.5°, and the melting point is 229.5° C.

3

The cellobiose octaacetate is deacetylated by dissolving it in dry methyl alcohol and adding 1 ml of 0.5 N barium methylate solution for each gram of sugar. After standing overnight in the refrigerator, the barium is removed by precipitation with an equivalent quantity of sulfuric acid and the barium-free solution is concentrated in vacuo to a thin sirup which on standing yields crystalline cellobiose.

Anhydrous B-cellobiose melts at 225° C and gives [a]=+14.2° initially, which changes in the course of several hours to +34.6°.

NOTES

1 The simultaneous hydrolysis and acetylation of cellulose was described first by Franchimont [2]. Modiications of the original method are described in references [3, 4, 5, and 6].

2 This part of the preparation requires some practice in order to obtain uniform results. Several batches may be prepared in this way and combined in the next step, but it is not advisable to work with larger amounts up to this point, as charring may result.

The preparation of the barium methylate solution and additional details for the method are given on page 493,

REFERENCES

[1] F. Klein, Z. angew. Chem. 25, 1409 (1912).

[2] A. P. N. Franchimont, Ber. deut. chem. Ges. 12, 1941 (1879). [3] Z. H. Skraup, Ber. deut. chem. Ges. 32, 2413 (1899).

[4] G. Zemplén, Ber. deut. chem. Ges. 59, 1258 (1926).

[5] G. Braun, Organic Syntheses 17, 36 (1937).

[6] C. C. Spencer, Cellulosechemie 10, 61 (1929).

4. 2-DESOXYGALACTOSE

Method.-[1, 2, 3] Fifty grams of recrystallized galactal' is dissolved in 700 ml of 5-percent cold sulfuric acid, and the solution is kept in the refrigerator at 0° C for 24 hours, after which it is neutralized with 120 g of barium carbonate and the esters are saponified by heating to 60° C. The saponification usually requires about 48 hours of heating. Eight- or ten-gram portions of barium carbonate are added at intervals during this time. The solid material is separated by filtration, and the filtrate is evaporated in vacuo to a thick sirup which is taken up with 50 ml of absolute alcohol and allowed to crystallize. About 40 g of material is obtained."

The crystals are purified by dissolving in water and evaporating the solution to a sirup, which is taken up with three volumes of methyl alcohol and allowed to crystallize, preferably in a slowly rotating flask.

Desoxygalactose melts at 120° to 121° C. In 4-percent aqueous solution, desoxygalactose gives [a]=+40.8° initially, which decreases in 5 minutes to a minimum and then increases to +60.5° in about 30 minutes.

NOTES

The preparation of galactal is described on page 532.

2 The preparation of 2-desoxyglucose and 2-desoxyrhamnose is given by Bergmann, Schotte, and Leschinsky [3], while that of 2-desoxyarabinose and desoxyxylose is described by Levene and Mori [4]. Several natural-occurring desoxy sugars are reported [5, 6, 7].

REFERENCES

[1] H. S. Isbell and W. W. Pigman, J. Research NBS 22, 397 (1939) RP1190. [2] P. A. Levene and R. S. Tipson, J. Biol. Chem. 93, 644 (1931).

[3] M. Bergmann, H. Schotte, and W. Leschinsky, Ber. deut. chem. Ges. 55, 158 (1922); 56, 1052 (1923).

[4] P. A. Levene and T. Mori, J. Biol. Chem. 83, 803 (1929).

[5] H. Kiliani, Ber. deut. chem. Ges. 38, 4040 (1905).

[6] A. Windaus and L. Hermanns, Ber. deut. chem. Ges. 48, 88 (1915). [7] W. A. Jacobs and N. W. Bigelow, J. Biol. Chem. 96, 355 (1932).

5. I-FUCOSE (1-GALACTOMETHYLOSE)

Method.'-[1, 2, 3] The seaweed, ascophyllum nodosum, is washed with tap water and any large shells are removed. It is then air-dried and ground to a fine powder. One kilogram of the dried seaweed is added to 8 liters of hot 4-percent sulfuric acid, and the solution is allowed to simmer for 3 hours. The liquid is separated by filtration and the residue is washed with hot water. The filtrate is neutralized with an excess of calcium carbonate, refiltered, and the calcium sulfate washed with water. The mixture is cooled to room temperature and fermented with baker's yeast acclimatized to ferment galactose. When the fermentation is complete, about 100 g of a decolorizing carbon is added; and after standing for several hours, the

2

The

solution is filtered and concentrated in vacuo to about 2.5 liters. precipitated calcium sulfate is separated, and the filtered solution is concentrated to a sirup (n=1.48). The sirup is mixed with 1 liter of hot methyl alcohol; the insoluble residue is separated and washed with three 100-ml portions of hot methyl alcohol. The residue is discarded, and the alcoholic extract is purified by adding 600 ml of ether and separating the precipitate by filtration. The solution is concentrated to a thick sirup (n=1.50), which is taken up in 200 ml of hot absolute alcohol. The alcoholic solution is filtered and then mixed with 75 ml of phenylhydrazine. After standing for 1 or more days in the refrigerator, the crystalline hydrazone is collected on a filter, washed with absolute alcohol, and dried at 50° C. The hydrazone (100 g) is transferred to a flask containing 50 ml of benzaldehyde and 2 liters of hot water. The mixture is stirred and heated to about 90° C for about 1 hour, after which it is cooled to room temperature. After the addition of 10 g of a decolorizing carbon, the benzaldehyde phenylhydrazone is separated by filtration. The filtrate is extracted three times with 100-ml portions of ether to remove all of the benzaldehyde. The solution is then made acid with acetic acid and concentrated to a sirup of about 80 percent of total solids. This sirup is diluted with about 50 ml of absolute alcohol and seeded with crystalline fucose. After this solution has stood for 1 or more days in the refrigerator, the resulting crystals are separated and washed with absolute alcohol. The yield is about 30 g.

Recrystallization. The sugar is dissolved in approximately an equal quantity of hot water, and after treatment with a decolorizing carbon, is filtered. The solution is made acid with acetic acid and concentrated in vacuo to a sirup of about 80 percent of total solids (n=1.490). This sirup is diluted with 2 ml of absolute alcohol for each gram of sugar and placed in the refrigerator for crystallization to take place. The resulting crystals are collected on a filter and washed with 95-percent alcohol. The mother liquors are concentrated and additional crystals are separated so that nearly all of the sugar is reclaimed. Better purification but lower yields are obtained by crystallizing the sugar from an aqueous solution contained in a flask which is slowly rotated. A solution containing approximately 78 percent of sugar by weight gives a satisfactory crystallization.

In 4-percent aqueous solution l-fucose gives [a]=-152.6° initially, which changes in the course of several hours to -75.9°.

NOTES

The method is essentially that described by Tollens and coworkers [1] as modified by Clark [2] and by Hockett, Phelps, and Hudson [3].

? The fermentation removes mannose and galactose, which are present in certain seaweeds [4].

REFERENCES

[1] J. A. Widtsoe and B. Tollens, Ber. deut. chem. Ges. 33, 132 (1900); A. Günther and B. Tollens, Liebigs Ann. Chem. 271, 86 (1892).

[2] E. P. Clark, BS Sci. Pap. 18, 527 (1922) S459; J. Biol. Chem. 54, 65 (1922). [3] R. C. Hockett, F. P. Phelps, and C. S. Hudson, J. Am. Chem. Soc. 61, 1658 (1939).

[4] R. H. F. Manske, J. Biol. Chem. 86, 571 (1930).

903232 - 50 - 31

6. d-GALACTOSE

Method. [1] Fifteen hundred grams of lactose is dissolved in 3,750 ml of hot water containing 75 g of concentrated sulfuric acid. The solution is brought to a boil and then simmered for 2 hours. A thin paste of barium carbonate is then added to the hot solution until it reacts neutral to congo-red paper. The precipitate of barium sulfate is allowed to settle, after which as much as possible of the supernatant liquid is drawn off and filtered. Then the precipitate is placed on the filter and washed with water. The filtrate is concentrated under diminished pressure until it has a weight of about 1,650 g (n between 1.5120 and 1.5125). The very thick sirup is warmed to between 60° and 70° C, and 250 ml of ethyl alcohol is dissolved in it by vigorous shaking. The solution is then poured into a beaker or jar and the remaining sirup is washed from the flask with 500 ml of methyl alcohol. This is best done by adding the methyl alcohol to the flask portionwise and warming and shaking in a water bath. The whole solution is thoroughly mixed, seeded with some pure galactose crystals,1 and allowed to crystallize, preferably in a slowly rotating flask.

The crystallization is generally complete in about 4 days, after which the crystals are filtered off, washed with a little methyl alcohol, and dried. The yield of the crude sugar is about 27 percent of the lactose taken.2,3

Recrystallization.-The crude galactose is dissolved in an equal weight of hot water, and after the addition of a small quantity of a decolorizing carbon the solution is filtered. A few drops of acetic acid are added, and the solution is evaporated in vacuo to a sirup of 60 percent of total solids (n=1.442). The sirup is then seeded with a-d-galactose and kept in motion while crystallization proceeds. When a satisfactory crystal growth is obtained (in about 1 day), the crystals are separated either by filtration or by means of a centrifuge. The crystals are washed thoroughly with methyl alcohol. About 50 percent of the sugar is obtained in the first crop, and the remainder may be separated by concentration of the mother liquor.

A more rapid recrystallization can be made by dissolving 200 g of galactose in 100 ml of water, filtering the solution with the addition of about 5 g of a decolorizing carbon, and adding 100 ml of methyl alcohol. The mixture is seeded and stirred while crystallization takes place. After several hours the crystalline galactose is separated by filtration and washed with methyl alcohol. About 75 percent of the original sugar is obtained in the first crop of crystals; the rest of the sugar is reclaimed by concentrating the mother liquor.

In 5-percent aqueous solution, pure a-d-galactose has an initial specific rotation, [a]=+150.7°, which changes after a number of hours to the equilibrium value, +80.2°.

NOTES

1 Contamination by dextrose seed must be avoided.

If the hydrolysis is not complete, unchanged lactose may be obtained. Also dextrose may occasionally crystallize simultaneously with the galactose.

Galactose may be prepared from plant gums, such as those from the western larch [2] and by partial fermentation of the hydrolysis products of lactose [3]. A review of methods for the preparation of galactose is given by Harding [4]. If the solution is not kept in motion, the crystals form a solid mass.