Clinical Chemistry, Vol 7, 646-654, Copyright © 1961 by the American Association for Clinical Chemistry

Apparent Dissociation Constant of Phenol Red as Determined by Spectrophotometry and by Visual Colorimetry

¹ Division of Chemistry, Naval Medical Research Institute, National Naval Medical Center, Bethesda, Md.

The apparent pK' of phenol red was determined in buffered NaCl solutions of known composition at 25° and 37°, by both spectrophotometric and visual colorimetric measurement of the concentration ratios of the alkaline to acid form of the indicator. A small but statistically significant difference was obtained by the two procedures. The pK' values obtained in 0.15M NaCl spectrophotometrically (7.67 at 25° and 7.57 at 37°) were approximately 0.03 unit lower than those obtained colorimetrically. Measurements made as a function of NaCl concentration at 25° were in accord with the Debye-Hückel theory. Small variation was observed between solutions buffered with phosphate or tris at the higher NaCl concentrations. The results indicate that in addition to ionic strength and temperature, variations in method of measurement and in the source of the preparation are important factors in the determination and use of pK' values for phenol red. Attention is called to the pertinence of these results to the measurement of blood pH by colorimetric and spectrophotometric methods.