ISOLATION AND ESTIMATION OF TRYPTOPHAN 29 



ing way : Solutions of tryptophan of a concentration of I in 1000, 

 I in 2000, i in 3000 and so on up to I in 10,000 are prepared and 

 also I in 20,000 up to I in 50,000. To I c.c. of these solutions in 

 test tubes of equal size and bore are added 2 c.c. of glyoxylic acid 

 solution l arid 6 c.c. of concentrated sulphuric acid. A series of 

 coloured solutions from deep blue to red-violet result. A known 

 weight (about O'l gram) of previously purified and dried protein is 

 placed in another test tube and treated with 2 c.c. of glyoxylic acid 

 and 6 c.c. of concentrated sulphuric acid. The colour so produced is 

 compared with the other colours. The one showing the closest com- 

 parison is then matched with the one from the protein in a Duboscq 

 colorimeter, the cups of which are entirely made of glass. Readings 

 are taken from both above and below in the usual manner. Usually 

 the comparison was made with the tryptophan solution I in 4000, 

 z.e., I c.c. = 0*00025 gram tryptophan. If the colorimeter readings 

 were for example 22 : 18 (standard) the protein contains I -23 times as 

 much tryptophan as the solution, z.e. t O'l gram protein contains 

 0*00025 v 1-23 gram tryptophan or O'3 per cent. 



In most cases the colours could be matched fairly well, but in some 

 cases the comparison was difficult owing to brown shades in the pro- 

 tein solution. This could be sometimes overcome by taking different 

 amounts of protein. In these cases Fasal [1913] found that a com- 

 parison was possible if the colours were compared in the apparatus 

 with colour filters described by Sorensen. 2 Numerous estimations 

 were made by Fasal ; they are given in the following table in which 

 the figures are the percentage of tryptophan : 



Caseinogen 0*65 Epidermis . . . . 0*30 



Edestin 0*38 Cutis o-o 



Lactalbumin 3*07 Psoriasis scales 0-41 



Liver tissue I'oo Carcinoma (epithelial) . . . 1-7 



Horn (ox) 0-17 (liver) . . . .1*7 



Nail (human) + (mouse) . . . .1-6 



Hair (human) o Sarcoma (rat) 1-4 



Wool (sheep) + Carcinoma (breast) . o 



The different kinds of keratin, for which the method was particu- 

 larly devised on account of their insolubility by pepsin and trypsin, 

 show very striking differences. The value for caseinogen is higher 

 than that found by the gravimetric method by Abderhalden and 

 Kempe, but lower than that found by Hopkins and Cole. The large 



1 Prepared by reducing a saturated aqueous oxalic acid solution with 3 per cent, sodium 

 amalgam. 



2 Biochemische Zeitschrift, 1909, 21, 201. 



