36 THE CHEMICAL CONSTITUTION OF THE PROTEINS 



dry gaseous hydrochloric acid. In some cases it may be necessary to 

 repeat this operation once more. 



The esterification, according to Osborne and Jones [1910, 2], is 

 more advantageously effected by the method of Phelps and Tillotson. 

 The concentrated solution of amino acid hydrochlorides is dissolved 

 in alcoholic hydrochloric acid and zinc chloride is added as a catalyst. 

 The solution is maintained at a temperature of 100 and the vapours 

 of absolute alcohol containing some hydrochloric acid are passed into 

 the solution. The water arising during the process is removed by the 

 alcohol vapours as fast as it is formed, and complete esterification 

 results in a shorter time. 



Since the completion of the esterification of the amino acids necessitates at least two 

 evaporations and two treatments with absolute alcohol and hydrogen chloride, Foreman 

 [1913 and unpublished J ] proposed that the esterification be effected with the dry lead or 

 copper salts of the amino acids. The solution of amino acids is evaporated in vacua and 

 steam is passed into the remainder to remove hydrochloric acid as completely as possible. 

 It is diluted to about 2000 c.c. (with 320 grams protein) and heated with 100 grams of pre- 

 cipitated lead hydroxide. The undissolved matter, which is dark in colour and contains 

 humin, is filtered off. The filtrate is boiled for forty-five minutes with 400-500 grams of 

 litharge. It is best to treat first with precipitated lead hydroxide, as under these conditions 

 the humin separates in a flocculent state. The excess of litharge is filtered off and washed, 

 and the solution is evaporated to about 500 c.c. On cooling it sets to a semi-solid mass, 

 but it is stirred up on a water-bath until it becomes too viscous and dried in a steam oven. 

 The dry mass, so obtained, can be powdered, but this is not necessary. The lead salts are 

 suspended in about 1500 c.c. of absolute alcohol and dry hydrogen chloride passed in until 

 the liquid is saturated. The solution is allowed to cool and again saturated with hydrogen 

 chloride. The lead chloride is filtered off and washed with absolute alcohol ; the filtrate 

 and washings are evaporated to half their volume at 40 and 15 mm. The excess of hydro- 

 chloric acid is removed by slowly adding at o alcohol saturated with dry ammonia until 

 the liquid is only faintly acid, thus avoiding liberation of the esters from their salts. The 

 ammonium chloride, thus formed, is filtered off and washed with absolute alcohol and the 

 alcohol evaporated off in vacua at 40 and 15 mm. (the further treatment is described 

 on p. 40). 



(3) Isolation of Glycine as Ester Hydrochloride. 



At this stage, glycine, if it occurs in the protein, e.g., in gelatin, in 

 any considerable amount, is separated as glycine ester hydrochloride 

 by concentrating in vacua at 40 to two-thirds and seeding the solu- 

 tion with a crystal of this compound and allowing to stand for twenty- 

 four hours at o. The precipitate is filtered off while the liquid is kept 

 cold and is washed with ice-cold alcohol ; the mother liquor, on further 

 concentration in vacua and saturation again with hydrochloric acid, 

 may give another crop of glycine ester hydrochloride, which is treated 



1 Mr. Foreman has kindly informed me of the full details of his process, which gives as 

 good, or even better, yields than the direct esterification process. 



