VITAMIN G (Bo) 127 



of adding 50 cubic centimeters water, 5 cubic centimeters of a 30 per 

 cent solution of sodium nitrite and 10 cubic centimeters of 10 per cent 

 sulfuric acid to 50 cubic centimeters of a concentrate of vitamin Bg (G). 

 The mixture was shaken 5 minutes, allowed to stand overnight, heated 

 on a water bath until foaming ceased, and then boiled for 10 minutes. 



In 1930 Narayanan and Drummond reported success in separating 

 bios from vitamin Ba (G). This they did by concentrating in vacuo a 

 yeast extract prepared from brewers' yeast at 60° to 70° C. with alcohol 

 of 50 per cent concentration, hydrolyzing the product at 15 pounds 

 pressure with 20 per cent barium hydroxide, freeing the filtrate from 

 barium with dilute sulfuric acid, and treating with an excess of lead 

 acetate solution (110 grams lead acetate to the extract from 7 pounds 

 yeast). The resulting filtrate and precipitate were both treated with 

 hydrogen sulfide, and the lead-free liquids were brought to pH 6.8. 

 Vitamin G (Bo) was found to be concentrated in the fraction prepared 

 from the lead acetate precipitate whereas bios remained in the fraction 

 made from the filtrate. 



These investigators attempted to concentrate vitamin B, (G) from 

 the lead acetate precipitate obtained as above with or without the alka- 

 line hydrolysis step. The precipitate was decomposed by suspending in 

 warm water and slowly adding 10 per cent sulfuric acid with constant 

 stirring until acid to Congo red, whereupon the lead sulfate was re- 

 moved by filtration and the filtrate neutralized with sodium hydroxide. 

 The filtrate and subsequent fractions were tested for the vitamin by 

 finding the minimal daily doses which, when fed to rats depleted of 

 their bodily reserves of vitamin Bg (G) (vitamin B, furnished by the 

 Kinnersley and Peters concentrate), permitted a gain of at least 10 

 grams weekly during a three weeks test period. 



The filtrate, active in daily doses of 10 to 15 milligrams organic 

 matter, was subjected to adsorption at various hydrogen-ion activities 

 by fuller's earth and by norite, and to fractional precipitation with 

 alcohol. Between pH 6.8 and pH 0.1 adsorption of vitamin B2 (G) on 

 fuller's earth increased as the acidity increased, being almost complete 

 at pH 0.1. Using 3 grams fuller's earth per 100 cubic centimeters liquid 

 (0.9 A'' sulfuric acid), stirring for ^ hour, and washing with 0.9 A^ 

 sulfuric acid resulted in removing from the filtrate 40 per cent of the 

 total solids and organic material and producing a fuller's earth active 

 in (Jaily doses of 40 milligrams, carrying 6 milligrams adsorbed organic 

 matter. No satisfactory method was found for extracting the vitamin 

 from the fuller's earth. Several reagents were found inefifective, among 

 which were cold barium hydroxide, alkaline to bromocresol purple; 50 



