THE SYNTHESIS OF THE PROTEINS 55 



It resulted by the hydrolysis with 70 per cent, sulphuric acid followed 

 by trypsin, and by the hydrolysis with hydrochloric acid. 



This diketopiperazine could arise both from glycyl-d-alanine and 

 d-alanyl-glycine by the above method. It could also arise by synthesis 

 from glycine and d-alanine under the conditions of the experiment ; 

 control experiments were carried out to determine this possibility, and 

 they showed that this was impossible so that there was no doubt con- 

 cerning the presence of a dipeptide amongst the products of hydrolysis. 

 Since this dipeptide was resistant to trypsin it was most probably glycyl- 

 d-alanine and not d-alanyl-glycine which is easily hydrolysed by this 

 enzyme. 



At the same time another diketopiperazine, glycyl-1-tyrosine anhy- 

 dride, was also obtained ; its nature was established a little later by 

 identification with synthetical glycyl-1-tyrosine anhydride prepared from 

 the ester of chloracetyl tyrosine and ammonia. In one experiment its 

 yield amounted to 4*2 per cent, of the silk-fibroin employed. 



In the same way by the hydrolysis of elastin with 70 per cent, 

 sulphuric acid and by the action of ammonia upon the esters, a product 

 was isolated which was composed of glycine and 1-leucine and which 

 was identical with synthetical glycyl-1-leucine anhydride. 



In 1907 Fischer and Abderhalden definitely showed that the first 

 diketopiperazine isolated from the hydrolysis products of silk-fibroin 

 was derived from glycyl-d-alanine. Silk-fibroin was partially hydro- 

 lysed by hydrochloric acid and then precipitated by phosphotungstic 

 acid. A portion of the filtrate from this precipitate, after removal of 

 the excess of phosphotungstic acid, was treated in alkaline solution with 

 /3-naphthalene-sulphochloride and a product was obtained which was 

 identical with /3-naphthalene-sulpho-glycyl-d-alanine ; further proof of 

 this was given by its careful hydrolysis with dilute hydrochloric acid 

 when the dipeptide chain was split, but the naphthalene-sulpho radical 

 not removed ; /3-naphthalene-sulpho-glycine and d-alanine were obtained 

 according to the equation : 



C 10 H 7 S0 2 NH . CH a . CO NHCH(CH,)COOH + H 2 O = 

 C 19 H 7 SO 2 NH . CH 2 . COOH + NH 2 . CH(CH 3 ) . COOH 



From the remainder of the filtrate, glycyl-d-alanine anhydride and a 

 small quantity of glycyl-1-tyrosine anhydride were obtained by the 

 action of ammonia upon the esters, as well as another product which 

 was most probably d-alanyl-1-serine anhydride. 



An examination of the phosphotungstic acid precipitate showed that 

 it contained several products of a complex nature. From them a sub- 

 stance was isolated which consisted of two molecules glycine, one 



