98 THE CHEMICAL CONSTITUTION OF THE PROTEINS 



diluted to 200 c.c. ; 100 c.c. of alcohol are added to prevent frothing 

 and then an excess of a 10 per cent, suspension of calcium hydrate, 

 as shown by the turbidity and alkaline reaction of the solution. Air 

 may be introduced through the stop-cock, if distillation starts too 

 rapidly ; the stop-cock serves to release the vacuum when the distilla- 

 tion is finished. The decinormal acid in both flasks is mixed and 

 titrated with decinormal soda using alizarin sulphonate as indicator, 

 which may be added previous to the distillation. 



4. Humin Nitrogen. The black colouring matter is absorbed by 

 the lime, which is filtered off on a folded filter paper and washed with 

 water until the washings are free from chlorides. The precipitate 



FIG. 5. 

 From Biol. Chem., 1911, 10, 21. 



and paper are *th en submitted to Kjeldahl analysis, using 35 c.c. of 

 sulphuric acid. 



5. Precipitation and Washing of the Phosphotungstate Precipitate. 

 The filtrate from the humin nitrogen is neutralised with hydrochloric 

 acid, returned to the vacuum distillation apparatus, and concentrated 

 to about 100 c.c. 



It is then washed into a 250 c.c. conical flask, and 18 c.c. of con- 

 centrated hydrochloric acid and 1 5 grams of phosphotungstic acid l in 

 aqueous solution are added. 



The entire solution is diluted to 200 c.c. with water and heated in 

 a water-bath, until the precipitate has nearly, or quite, redissolved v 

 On cooling, the phosphotungstates separate in a crystalline form. 



1 This should be purified by Winterstein's method by shaking its acid solution with 

 ether;* from the ethereal layer containing the substance the acid is recovered by recrystal- 

 lisation. * 



