102 TH? CHEMICAL CONSTITUTION OF THE PROTEINS 



to the solution in the flask, and the solution is boiled gently for exactly 

 six hours. Nearly all the evolved ammonia diffuses into the bulbs. 



The bulbs are disconnected and 100 c.c. of water are poured 

 through the condenser into the flask. The flask is connected with the 

 condenser of a Kjeldahl distilling apparatus, and the remainder of the 

 ammonia driven off and collected in the acid from the bulbs, which has 

 been transferred to a suitable receiver. Not more than 100 c.c. of the 

 solution must be distilled over as the strong potash may destroy the 

 other substances in the solution. 



The excess of acid in the receiver is titrated. Each c.c. of acid 

 neutralised by the ammonia corresponds to 0*0028 gram arginine- 

 nitrogen, or 0*0056 gram in the total solution. 



Plimmer [1916] showed that arginine was decomposed by boiling 

 with 20 per cent, caustic soda for six hours. 1 The estimation is carried 

 out preferably by adding an equal volume of 40 per cent, caustic soda 

 to the arginine solution and boiling for six hours. A larger volume 

 of liquid is thus present in the flask and the lower concentration of the 

 alkali minimises breakage of the glass flasks through the action of the 

 alkali. Instead of disconnecting the bulbs and distilling as recom- 

 mended by Van Slyke so as to collect the remainder of the ammonia, 

 it is only necessary to run the water out of the condenser and continue 

 boiling for another fifteen to twenty minutes so as to drive the am- 

 monia into the bulbs. 20 c.c. instead of 25 c.c. of the solution may 

 be used. 



Cystine is usually not present in sufficient amount in proteins to 

 cause any considerable error in this determination, but if a keratin be 

 under investigation a correction must be made ; 1 8 per cent, of its 

 nitrogen is evolved when it is boiled with alkali ; the nitrogen figure 

 of cystine is obtained later in the process. 



8. Total Nitrogen of the Hexone Bases and Cystine. The solution 

 remaining from the arginine determination is transferred to a Kjeldahl 

 flask of 500 c.c. ; 35 c.c. of sulphuric acid are added and 0*25 gram 

 of copper sulphate. The nitrogen is then estimated in the usual way. 

 This estimation can be performed in duplicate, if the solution be 

 diluted to 100 or 200 c.c. and divided into two halves, each half being 

 oxidised with 20 c.c. of sulphuric acid. 



On account of persistent bumping from silica dissolved from the 

 glass in the arginine estimation and consequent loss, Plimmer [1916] 



1 Histidine is decomposed to a slight extent by this treatment, but the error introduced 

 is negligible. 



