28 THE CHEMICAL CONSTITUTION OF THE PROTEINS 



Isolation and Characterisation of Valine. 



The filtrate is freed from lead by means of hydrogen sulphide, and 

 the solution, filtered from lead sulphide, is evaporated to dryness. The 

 dry residue is treated with an alcohol-ether mixture (3 : i) to extract 

 acetic acid and ammonium acetate. The small amount of valine 

 which dissolves is recovered by again evaporating to dryness and 

 extracting with alcohol and ether. 



Pure valine generally remains ; it is identified by recrystallisation 

 from water, elementary analysis, and rotation in 20 per cent, hydro- 

 chloric acid [a]gj = +28*8; a rotation of 26-28 is generally found 

 as racemisation occurs in the process. 



If analysis shows leucine to be present the above treatment must 

 be repeated. 



Treatment of the Lead Salts of Leucine and Isoleucine. 



The purity of the lead salt, as obtained above, is tested by analysis. 

 This is performed by dissolving about 0-3 gm. in 5 c.c. of normal nitric 

 acid in a 100 c.c. beaker and precipitating the lead with 5 c.c. of normal 

 sulphuric acid followed by 50 c.c. of absolute alcohol. The lead sul- 

 phate, which settles rapidly in a granular form, is collected after about 

 fifteen minutes in a Gooch crucible and washed with 95 per cent 

 alcohol acidified with sulphuric acid. The crucibleas placed in another 

 crucible, heated gently until the alcohol is driven off and then with the 

 full heat of the Bunsen burner for ten minutes. 



If the lead content be too high, due to contamination with the lead 

 salt of valine, the mixture is purified by dissolving it, after thorough 

 pulverisation, in 5 parts of hot water + J part of glacial acetic acid and 

 reprecipitating by adding 0-5 c.c. of concentrated ammonia solution 

 for every gramme of salt. The precipitate is collected and treated as 

 above under the conditions where the amount of valine present is 

 small. 



The last portions of the mixed leucines are recovered by repeating 

 the entire process. 



