TITRATION OF THE BICARBONATE CONTENT 135 



neutral red (dissolved in 50 per cent alcohol) is added. 0.02 N 

 carbonate-free sodium hydroxide is then run in from a burette 

 (preferably but not necessarily a micro-burette) until the color 

 of the solution matches that of 29 c.c. of a standard phosphate, 

 solution, of pH 7.4, contained in a similar 50 c.c. flask. 



In place of neutral red 0.3 c.c. of a 0.04 per cent solution of 

 phenolsulfonephthalein may be used as indicator, and gives an 

 end-point slightly more easy to distinguish than that of neutral 

 red. When phenolsulfonephthalein is used, however, the standard 

 solution must be of pH 7.2 instead of 7.4. The reason for this is 

 that when phenolsulfonephthalein is added to a solution contain- 

 ing serum proteins, the pH is considerably higher than that esti- 

 mated by colorimetric comparison with standard phosphate 

 solutions. The indicator suffers from a protein error. Under the 

 conditions of the analysis here described, however, the error is 

 quite constant at 0.2 to 0.3 pH, i.e., if the color matches that of 

 a standard solution of pH 7.4, the actual pH in the solution under 

 examination is about 7.7. If a standard solution of pH 7.2 is used, 

 however, the titration runs to the same point obtained with 

 neutral red. 



For the titrations there is used a 3 c.c. micro-burette of the 

 model devised for use with the blood sugar method of Bang. 

 Such a burette is desirable but not absolutely necessary. An 

 ordinary 50 c.c. burette divided into 0.1 c.c. divisions may be 

 employed. 



The End-point. With both indicators, a peculiar phenomenon 

 occurs as the end-point is approached. Each drop appears to 

 change the color past the end-point, but within a few seconds 

 the color shifts back, and it is seen that at least another drop 

 is needed before the genuine end-point is reached. Consequently, 

 the final color comparison should not be made until at least 

 thirty seconds after the last drop of 0.02 N sodium hydroxide 

 has been added. Because of this behavior, as well as the character 

 of the color change, it is well, particularly with neutral red, to 

 overrun the end-point by a drop, rather than stop short of it when 

 in doubt. 



Calculation of Results. The number of c.c. of 0.02 N sodium 

 hydroxide used in the titration is subtracted from the number 

 required to neutralize to the same indicator 5 c.c. of the 0.02 N 

 hydrochloric acid used. This number is, of course, approximately 



