348 PRACTICAL ORGANIC AND BIO-CHEMISTRY 



Tryptophan. 



Tryptophan was shown to 



CH(NH 2 ) COOH be a constituent of proteins by 

 . Hopkins and Cole in 1902, who 

 isolated it from the mixture of 

 amino acids which results from 

 the digestion of proteins with the enzyme, trypsin. Its discovery gave 

 the clue to the well-known Adamkiewicz reaction of proteins and the 

 proteinochrome reaction of tryptic digests. 



Preparation. 



i kilo, of caseinogen is dissolved in about 8 litres of '8 per cent, crystallised 

 sodium carbonate solution and digested with 2-4 gm. of a trypsin prepara- 

 tion for 5-6 days at 37 in the presence of toluene or chloroform as an 

 antiseptic. The digestion is allowed to proceed until a sample of 5-10 c.c. 

 withdrawn at intervals gives a maximum colour reaction with bromine water 

 after acidifying with acetic acid. The digestion is stopped by heating the 

 solution to 80. The insoluble portion, consisting of undigested proteins, 

 calcium phosphate and tyrosine, is filtered off and the clear nitrate, better after 

 concentration in vacuo to a volume of about i litre and filtration from tyro- 

 sine, is acidified with sulphuric acid until it contains 5 per cent. The acid 

 solution is precipitated with a 10 per cent, solution of mercuric sulphate in 

 5 per cent, sulphuric acid. After 12 hours the precipitate, which contains 

 tryptophan, tyrosine and cystine, is filtered off and washed with 5 per cent, 

 sulphuric acid to remove tyrosine until the washings show only a faint reaction 

 with Millons' reagent for tyrosine. 



The precipitate is suspended in water, warmed and decomposed with 

 hydrogen suiphide. The filtrate from mercuric sulphide is freed from hydrogen 

 sulphide by a current of air, acidified to 5 per cent, with sulphuric acid 

 and again precipitated with the acid mercuric sulphate solution. The 

 cystine comes down first and is removed ; the tryptophan is thrown out on 

 the addition of more mercuric sulphate. The precipitate is washed with 5 

 per cent, sulphuric acid, decomposed with hydrogen sulphide and the solution, 

 freed from sulphuric acid by baryta, is evaporated in vacuo to a small volume. 

 Tryptophan separates out and is recrystallised from a mixture of water and 

 alcohol containing animal charcoal. The yield of tryptophan is from 5-10 gm. 



Properties. 



Tryptophan crystallises in colourless glistening platelets which 

 are not easily soluble in cold water, but readily in hot. It is 

 insoluble in absolute alcohol and ether. It dissolves easily in hot 

 pyridine, less easily in cold. On heating in a capillary tube it 

 changes colour at 220, becomes brown at 240 and melts at '2 5 2. 

 If heated quickly it turns yellow at 260 and melts at 289. Tryp- 

 tophan is a weak base and forms salts with acids. As an amino 

 acid it forms salts and forms acyl derivatives with acid chlorides, some 

 of which serve for its isolation and characterisation. 



