September 15,1933
INDUSTRIAL AND ENGINEERING
LITERATURECITED Fales, “Inorganic Quantitative Analysis,” p. 236, Century, 1925. Hillebrand, W. F., and Lundell, G . E. F., “Applied Inorganic Analysis,” p. 502, Wiley, 1929. Kolthoff, I. M., and Mensel, H., ti-. by Furman, “Volumetric Analysis,” p. 279, Chapman & Hall, 1929. Low, “Technical Methods of Ore Analysis,” 6th ed., p. 67, Wiley, 1922. McBride and Sherer, J. Am. Chem. Soc., 39, 928 (1917).
CHEMISTRY
319
(6) Scott, “Standard Methods in Chemical Analysis,” 4th ed., Vol. I, p. 108, Van Nostrand, 1927. (7) Talbot, “Quantitative Chemical Analysis,” 7th ed., p. 191, Macmillan, 1931. (8) Treadwell, F. P., and Hall, W. T., “Quantitative Analysis,’’ Vol. 11, p. 530, Wiley, 1924. (9) Washington, H. S., “Chemical Analysis of Rocks,” 4th ed., p. 294, Wiley, 1930. RECEIVEDMay 4, 1933.
Estimation of Total and Bound (D) Gossypol in Cottonseed Meal A Modified Method F. H. SMITH AND J. 0. HALVERSON, Agricultural Experiment Station, Raleigh, N. C.
G
OSSYPOL in the cottonseed is readily soluble in ethyl ether. However, probably because of the influence of heat, pressure, and moisture, when expressing the oil from the crushed seed in the commercial manufacture of the cottonseed meal, the greater part of the gossypol left in the meal becomes insoluble in ethyl ether. This insoluble part is known as bound or (D) gossypol (I, 2 ) . The method used by Carruth (4) for the estimation of bound gossypol follows: Fifty grams of meal, having been previously extracted with ethyl ether, are saturated with 100 cc. of aniline, heated for 5 minutes a t 110” C., with constant stirring; then as much of the aniline as possible is removed immediately by a Biichner funnel and, after cooling sufficiently, the residue is washed with ethyl ether in order to remove the remaining aniline. The filtrate and washings are concentrated by distillation to a volume of 5 to 10 cc. The residue is washed with ethyl ether into a small beaker and allowed to stand one week in order to crystallize. The precipitate of dianiline gossypol is then filtered through a weighed Gooch crucible, and dried a t 100” C. to constant weight. The dianiline gossypol is converted to gossypol by the factor 0.775. TABLEI. THE ESTIMATION OF TOTAL GOSSYPOL (CHIEFLY BOUND)IN COTTONSEED MEAL1578 CARRUTH METHOD % 0.920 0.812 0.929
... I
.
.
... ... Av. 0.881 a
MODIFIED CARRUTH METHOD % 0.941 0.961 0.972
... ... .
.
I
... 0.958
PROPOSED RECOVERY OF GOSSYPOL METHOD BY PROPOBED METHODO % Mg. % 1.115 1.060 1.140 1,090 1.080 1.110 1.120 1,102
19.6 19.4 18.8 19.0 19.0 19.3
98.0 97.0 94.0 96.0 95.0 96.5
19.2
95.9
..
..
20 mg. gossypol uaed.
From a study of the factors concerned in the determination of gossypol by a modified method (S),certain improvements in the method of Carruth for bound gossypol were apparent. These were a more complete removal of the gossypol from the meal; the elimination of some decomposition of probably dianilirie gossypol by distilling off the excess aniline under reduced pressure, in which the greater part was distilled under 110” C. (just before completion, however, the maximum temperature of 140” C. was reached); a more complete precipitation of dianiline gossypol was also possible by the elimination of the ether used in the precipitation and washing since the precipitate is somewhat soluble in this
reagent, The precipitation of dianiline gossypol was facilitated by the use of 75 cc. of petroleum ether and 10 cc. of ethylene glycol in which dianiline gossypol is quite insoluble. The precipitate was washed with petroleum ether, then with a small amount of alcohol, and finally with water. With these modifications an average of 0.958 per cent gossypol was obtained from meal 1578, compared with 0.881 per cent by the method of Carruth, or an increase of 0.077 per cent. For the convenience of using smaller charges than 50 grams and 100 cc. of aniline the following method was worked out for total and bound gossypol. In order to obtain the total or the bound gossypol, the meal is first extracted on the Pickel extraction apparatus with petroleum ether and with ordinary ethyl ether, respectively, then dried, and stirred. All results reported in Table I are for total gossypol, inasmuch as the amount of ether-soluble gossypol in a charge of 2 grams is small, being less than 2 mg., and since the bound gossypol constitutes 90 per cent or more of the total. The accuracy of the proposed method for the determination of bound gossypol is also better shown by the determination of total gossypol, since it has been found that the amount of ether-soluble gossypol is variable, depending upon the moisture content of the meal (in press). MODIFIEDMETHODIN DETAIL Place a charge of 2 grams, ground to pass through a 40-mesh sieve, in a medium or porous alundum filter crucible of 25 cc. capacity, extract in the Pickel apparatus 3 hours with petroleum ether in order to remove the crude oil, or 72 hours with ethyl ether for the ether-soluble gossypol in the estimation of total and bound gossypol, respectively. Dry the crucible and contents, stir the meal, and place the crucible with contents in a clean Pickel flask. Pipet (by suction) 5 cc. of aniline onto the meal. Then add 15 cc. of alcohol (90 per cent) to the flask and extract for 24 hours on the Pickel apparatus. Remove the flask, and transfer the extract to a 200-cc. Erlenmeyer flask with the aid of ethyl ether and a funnel. (The raffinose precipitated by the ether goes into solution during the process of precipitation. Any crystals of gossypol that may be adhering to the Pickel flask are transferred to a weighed Gooch crucible with the aid of water.) Remove the alcohol and ether by reduced pressure, then add 7 cc. of ethylene glycol and 10 cc. of ethyl ether to the residue. Digest on the steam bath. After the ether begins to boil, add water with shaking, a t 3-minute intervals
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Vol. 5 , No. 5
ANALYTICAL EDITION
as follou7s: 0.5, 0.5, 1.5, and 2.5 cc. (This procedure prevents the production of a fine precipitate which is difficult to transfer, filter, and wash.) Digest gently 15 to 20 minutes and let stand overnight. Filter through the weighed Go.och crucible, wash with alcohol (50 per cent), then with a small amount of petroleum ether, followed by 5 cc. of alcohol (95 per cent), then four times with water. Dry a t 100' C. to constant weight. The factor 0.775 times the weight of dinniline gossypol gives the weight of gossypol. The results of eight determinations on meal 1578 (Table I) average 1.102 per cent total gossypol with a maximum variation of 4.99 per cent. The recovery from 20 mg. of purified gossypol by this method is given in the last column. The average recovery is 19.2 mg., or 95.9 per cent with a variation of 4.2. By the Carruth method, meal 1578 yielded 0.881 per cent gossypol and 0.958 per cent by the modified Carruth method, whereas the method proposed gave 1.102 per cent gossypol. The difference in results between the proposed method and the Carruth method is 0.221 per cent. Part of this difference is probably due to the gossypol remaining in the meal as it still retains a faint yellow color, and small
additional amounts can be extracted from the residue from Carruth's method. After the treatment with aniline and 90 per cent alcohol on the Pickel apparatus by the proposed method, all residues are devoid of any yellow color. SUMMARY A method for total and bound gossypol in cottonseed meal has been outlined which gives results of an accuracy of 95.9 per cent on known ambunts of gossypol. The method is readily performed and is economical, yielding somewhat higher results than the Carruth method. LITERATURE CITED (1) Carruth, F. E., J . Am. Chem. Soc., 40, 647-63 (1918). (2) Clark, E. P., J . Bid. Chem., 76, 229-35 (1928). (3) Halverson, J. O., and Smith, F. H., IND. ENG.CHEM.,Anal. Ed., 5, 29-33 (1933). (4)Sherwood, F. W., J . Agr. Research, 32, 793-800 (1926). RBCEWEDMay 15, 1933. Published with the approval of the Director of the North Carolina Agricultural Experiment Station &a Paper 71 of the Journal Series.
Relation of Moisture to Extraction of Gossypol from Cottonseed Meal with Ether J. 0. HALVERSON AND F. H. SMITH, Agricultural Experiment Station, Raleigh, N. C.
T
HAT swine do not thrive Isolated gossypol is readily soluble in anhyEFFECT OF METHODS OF DRYING drous or ordinary ethyl ether. However, anhywhen f e d relatively Cottonseed meal, dried over large amounts Of certain drous ether extraots little or no gossypol from dried phosphorus pentoxide at 2,-inch c o t t o n s e e d meals over a long period is ascribed to the small Or air-dry whether the be vacuum for 2 and 6 hours a t 50' amounts of gossypol present in dried with or without heat. Moisture is the C.,wasextractedwith anhydrous important factor in extracting gossypol from and with ordinary ether. Little t h e s e m e a l s . Therefore the if a n y gossypol could be exfactors affecting the estimation cottonseed meal with ether, the highest yields tracted by the anhydrous ether Of gossypol are Of importance' occurring when the charge of meal contains about (an average of o.oo50 per cent), In the usual method of estima2o per cent Of water and when water is added while the ordinary grade gave tion (8) gossypol is extracted to the ether in the receiving flask. Meals SO s o m e w h a t higher results (an from air-dry cottonseed meal treated have yielded from twice to ten times more average of 0.0127 per cent). with ordinary ethyl ether. Algossypol than the Same samples of aipdry meal. TOascertain whether the heat t h o u g h gossypol, purified, or e m p l o y e d in drying the meal isolated from the seed, is readily had rendered the gossypol unsoluble in anhydrous ethyl ether, it was found that anhydrous ether extracts materially less extractable, charges were dried in a desiccator over calcium chloride without any heat except that of room temperature. gossypol from moisture-free cottonseed meal. In the quantitative estimation of gossypol, a recently The results were no different from those obtained by drypublished modified method (1) was used in which a portion ing with heat and vacuum. The average amount of gossypol extracted with anhydrous of the oil was removed from charges of 75 grams of meal by washing with petroleum ether and the meal allowed t o dry ether from meals dried as above and also dried for 48 hours overnight in trays by exposure to the air. During this ex- a t room temperature in a desiccator over calcium chloride is posure, the meal readily absorbed or lost moisture, being 0.0069 per cent, compared to 0.0130 per cent with ordinary ether. responsive t o the humidity of the air. The Soxhlet apparatus was employed for the extraction of EFFECT OF MOISTURE IN ETHER gossypol by ether. The extraction was usually continued for Since ordinary ether extracted slightly more gossypol, 48 hours, but later was extended to 72 hours because of a slightly increased yield. Both ordinary ethyl ether (not for other charges of meal dried in a vacuum oven but a t 30" C. anesthesia or medicinal use) and anhydrous ether, which were extracted with anhydrous ether and with ether saturated with water. The results of determinations with anhydrous was distilled over sodium, were employed.
