The application of gradient reversed-phase high-performance liquid chromatography to the pKa and determination of polyprotic analytes

The purpose of this work was to propose a theoretical model of the combined pH/organic modifier gradient in reversed-phase high-performance liquid chromatography (RP HPLC) with special emphasis on its applicability to polyprotic analytes. The model was developed and approximated to be useful for a data set comprising organic modifier gradients obtained at different pH changes and different gradient durations. It was evaluated regarding its ability to describe experimental data. The chromatographic pKa and lipophilicity parameter, log   k w , were obtained by fitting to the proposed model and comparing to the literature values.