2 Fractionation of Phosphotungstic Acid Precipitate 



first investigation the author noticed that the active substance seemed 

 to diminish rather than increase. Our usual methods proved to 

 be inadequate for its isolation. In this respect the substance re- 

 minded one very strongly of the behavior of certain hormones and 

 products of the glands of internal secretion, whose isolation in the 

 active State has not yet been effected. 



The first attempt to isolate the vitamine, by the fractionation of 

 rice-polishings, was made by the author by the following method : 

 Rice-polishings were extracted with cold abs. alcohol, which was 

 partially saturated with gaseous hydrochloric acid. The extracts 

 were evaporated in vacuo at a low temp., and the fatty residue 

 melted and extracted with water. These aqueous extracts were 

 precipitated with 50 percent phosphotungstic acid sol., after addition 

 of sulfuric acid to the extent of 5 percent, and the precipitate de- 

 composed with baryta in the usual manner. The Solution, entirely 

 freed from baryta and sulfuric acid, was filtered, and the filtrate 

 neutralized with hydrochloric acid and evaporated in vacuo. The 

 residue was extracted with alcohol and the Solution freed by filtra- 

 tion from inorganic chlorids. The alcoholic sol. was then precipi- 

 tated with alcoholic mercuric chlorid sol. The active substance 

 was found, to a small extent, in this precipitate but the bulk was in 

 the filtrate. From each of these fractions the vitamine could be 

 completely thrown down by silver nitrate and baryta. From this 

 fraction, after decomposition with hydrogen sulfid, there was iso- 

 lated a very small quantity of crystalline substance, with a melting 

 point of 233 °C. This was not recrystallized and possessed very 

 marked curative power. The only substances isolated from 54 k. of 

 rice-polishings were this crystallin material which, from our present 

 knowledge, was undoul^tedly impure nicotinic acid; and a large 

 quantity of cholin. This work proved that vitamine is quanti- 

 tatively precipitated by phosphotungstic acid, and by silver nitrate 

 and baryta, partially by mercuric chlorid in alcoholic sol., but not by 

 platinic chlorid or picric acid. 



Shortly afterwards (1912) there appeared a paper on this sub- 

 ject by Edie, Evans, Moore, Simpson and Webster (2), who used a 

 slightly modified method. The source of their material was yeast. 

 This was extracted with alcohol, and the alc. extract evaporated. 



