14 Fractionation of Phosphotungstic Acid Precipitate 



SOLUTIONS OF DiFFERENT coNCENTRATioNS. In prcliminary study 

 of this problem, phosphotungstates of several naturally occurring 

 nitrogenous bases were prepared and their solubility in 25 percent, 

 50 percent, 75 percent, and 100 percent acetone sol. estimated.* The 

 f ollowing bases were used : cholin, betain, stachydrin, guanin, adenin, 

 guanidin, Creatinin and nicotinic acid. It was found that the phos- 

 photungstates were increasingly soluble in the direction of the arrow : 

 25 percent->ioo percent— »50 percent^75 percent (acetone). The 

 solubihty in 75 percent acetone sol. was many times greater than 

 that in pure acetone. Ahhough mixtures of phosphotungstates may 

 exhibit a behavior difTerent from that of the individual substances — 

 a point which has not as yet been determined — it seemed promising 

 to adopt this method for the investigation of the phosphotungstates 

 obtained from autolyzed yeast; in which case, as we saw from the 

 data in the preceding section, a large fraction is insoluble in pure 

 acetone. By applying this method to the mixture of phosphotung- 

 states from autolyzed yeast, five fractions were obtained, the last 

 insoluble fraction having been much smaller in bulk, than that with 

 pure acetone. It is not known, at present, in which of the five frac- 

 tions the Vitamine is contained but, from previous findings, one is 

 inclined to assume its presence in the insoluble fraction, which is 

 f reed from the bulk of inactive substances without sustaining loss by 

 decomposition. 



The fractionation was conducted as f ollows : 



Ten Ib. of wet brewer's yeast were autolyzed for three days. 

 To the semi-liquid mixture a third of its volume of alcohol was 

 added and the liquid was filtered. The filtrate was concentrated in 

 vacuo and the residue shaken with i 1. of 10 percent sulfuric acid 

 sol. for 24 hr. The residue was obtained by filtration and washed 

 on the filter with sufficient water to make 2 1. of filtrate and wash- 

 ings. This liquid was treated with 50 percent phosphotungstic 

 acid sol. After Standing for 24 hr. the precipitate was separated 

 by filtration and washed four times with 200 cc. portions of 5 per- 

 cent sulfuric acid sol. and finally dried. The dry precipitate weighed 

 1,125 gm. 



For fractionation, the precipitate was powdered, passed through 



* The estimations were made by my assistant, Mr. J. C. Drummond. 



