Apparent stability constants and purity of Ca-chelating agents evaluated using Ca-selective electrodes by the double-log optimization method

Apparent calcium stability constants and the purity of Ca-chelating agents were evaluated using Ca(2+)-selective electrodes by the double-log optimization method [Oiki S. Yamamoto T. Okada Y. (1994) Cell Calcium, 15, 199-208]. The method was amended to allow evaluation of the free Ca2+ concentrations contaminating electrolyte solutions. The value thus estimated (3.7 microM) was not significantly different from the total contaminating Ca content measured by atomic absorption spectroscopy. The purity of EGTA of different commercial brands was found to be in the range from 95.5-98.0% and was almost stable over several years of storage. The impurity of EGTA was completely eliminated by baking at 150 degrees C for 3 h. The purity of BAPTA decreased from 85.8 to 77.2% after storing for 3 months at -20 degrees C. The impurity of BAPTA was also abolished by the same drying procedure. At physiological pH (7.30), the apparent stability constants (K's) of EGTA were determined to be 7.13 and 6.97 in KCl-based solutions of 0.10 M and 0.16 M ionic strength, respectively, at 25 degrees C in the absence of Mg. At pH 7.30 and 0.20 M ionic strength K' values of BAPTA were 6.50 at 22 degrees C and 6.69 at 37 degrees C. The K' value increased with decreasing ionic strength.