104 METHODS FOR THE ANALYSIS OF BLOOD 



acid to make an approximately 3 per cent solution (either the 

 solid acid or a corresponding amount of 50 per cent solution is 

 used). Dilute to volume, shake, and let stand thirty minutes. 

 Add 2 gms. of kaolin, shake well, centrifuge, and filter. Transfer 

 an aliquot part (15 to 40 c.c.) of the filtrate to a small flask (50 to 

 100 c.c.), add two or three beads, one drop of alizarin indicator, 

 bring to a boil and keep slowly boiling (simmering) until the 

 indicator turns. Add 1 to 2 c.c. of N potassium hydroxide, and 

 boil one to two minutes to remove the ammonia. Make acid with 

 a few drops of acetic acid and evaporate to a small volume. The 

 whole amount may be used in the micro Van Slyke apparatus or 

 the liquid transferred to a small graduated flask, diluted to a defi- 

 nite volume, and aliquot portions measured out by means of the 

 burette of the Van Slyke apparatus. 



If the blood is older than twenty-four hours, or if it has been 

 laked or frozen, the proteins are precipitated according to the 

 following procedure: A measured volume of blood (30 to 50 c.c.) 

 is introduced into a flask which contains 0.4 gm. of ground soy 

 bean, 3 to 5 c.c. of water, and 1 c.c. of a 3 per cent solution of 

 NaH 2 PO4, and the mixture gently agitated. After standing for 

 one-half hour at room temperature the mixture is precipitated by 

 diluting to ten times its original volume with a 2.5 per cent solu- 

 tion of trichloroacetic acid. After standing for thirty minutes 

 2 gms. of kaolin are added, the mixture shaken vigorously, and 

 filtered. An aliquot part of the filtrate is evaporated to a small 

 volume, transferred to a small flask (50 to 100 c.c.) and treated 

 from this point on as above. 



THE QUANTITATIVE DETERMINATION OF 

 ALIPHATIC AMINO GROUPS 



Method of Van Slyke l 



Principle. This method for the determination of aliphatic 

 amino nitrogen is based on the measurement of the nitrogen gas 

 evolved in the reaction, 



RNH 2 +HNO 2 = ROH+N 2 -fH 2 O. 



1 Van Slyke: Jour. Biol. Chem., 1912, 12, 275; 1913, 16, 121 and 125. 



