330 J. HOWARD MUELLER 



to contain the greater part of the active material. It was found 

 that the filtrate from the HgS04 precipitate was no longer active, 

 after precipitating the Hg with H2S, and remo\'ing the H2SO4 

 with Ba (OH) 2, while the mercury precipitate, after freeing from 

 Hg and H2SO4 in the same way, was quite active. 



Following this observation, preparations of the amino acids 

 known to be precipitated by this reagent, namely, tryptophane, 

 tyrosine, cystine and liistidine, were obtained and their ability 

 to reactivate the decolorized infusion either singly or in combina- 

 tion with each other was tested and found negative. 



It was then found that mercuric sulphate would precipitate 

 the active material directly from the casein hydrolysate, without 

 resorting to the preliminary separation of the latter by means of 

 butyl alcohol, using Dakin's method. T^Ioreover, the sulphuric 

 acid used in hydrolj^sis did not have to be removed with baryta, 

 but could be neutralized by sodium hydroxide and the precipi- 

 tation carried out in the resulting strong solution of sodium 

 sulphate equally as well as in a solution free from salts. It is 

 rather difficult to determine the optimal conditions of precipita- 

 tion, but a considerable excess of HgS04, and not too high a 

 concentration of H2SO4 in the mixture seem to give the best 

 results. As a standard procedure a weight of HgS04 equal to 

 that of protein taken, make up in 5 per cent H2SO4, and added 

 to a hydrolysate which contains from 5 to 10 per cent amino 

 acids and is nearly neutral in reaction, has been used. Pre- 

 cipitation is complete in about twenty-four hours. The filtrate, 

 after freeing from Hg and H2SO4, may still show a slight ability 

 to reactivate the decolorized infusion, but the precipitate is 

 always strongly active; whether one of the two active materials 

 to be described is removed more completely than the other by 

 this method has not been definitely determined, but it is quite 

 possible. 



Attempt to purify the active fraction by fractional precipitation 

 with mercuric sulphate 



In the preparation of tryptophane by the method of Hopkins 

 and Cole, advantage is taken of the fact that cystine is precipi- 



