Estimation of pKa values using microchip capillary electrophoresis and indirect fluorescence detection

Microchip capillary electrophoresis (CE), coupled with indirect fluorescence detection was investigated for estimating the pKa values of non-fluorescent compounds. The CE method is based on the differences in electrophoretic mobility of the analyte as a function of the pH of the running buffer. Nine compounds were tested, including several of pharmaceutical importance, with pKa values from 10.3 to 4.6. All buffers contained 5-TAMRA as the fluorescent probe for indirect detection. Calculated pKa values agreed well with literature values obtained by traditional methods, differing not more than 0.2 from the literature value. The current work on single lane chips demonstrates the principle of microchip CE with indirect detection as a viable method for estimating pKa values. However, increased throughput will be required using a multilane chip to enable the approach to be used practically.