pka Estimation Of Ambrisentan By Reversed-Phase High-Performance Liquid Chromatography

The characterization of ionization behavior in pharmaceutical active ingredients is fundamental to understanding their bioavailability, pharmacokinetics, and formulation development. Ionization constants (pKa) are critical parameters that directly influence the solubility, stability, and membrane permeability of drugs, especially those with weakly acidic or basic characters. Ambrisentan, a selective endothelin receptor antagonist used in the treatment of pulmonary arterial hypertension, is a lipophilic compound whose ionization behavior requires detailed investigation. In this study, a reversed-phase high-performance liquid chromatography (RP-HPLC) method was employed to determine the pKa value of Ambrisentan using a C18 column at 37 °C across a mobile phase pH range of 2.5–7.5. To examine the effect of solvent composition on ionization, pKa determinations were performed in 40%, 45%, and 50% (v/v) acetonitrile-water binary mixtures. The experimental data were evaluated using the dielectric constant method and mole fraction (XACN– pKa) extrapolation methods. The calculated pKa value of Ambrisentan in aqueous medium was successfully estimated as 4.145±0.156 and 4.134±0.156 for the respective models. The results demonstrate that the developed chromatographic approach provides a reliable and reproducible tool for the pKa determination of drugs with limited aqueous solubility, which is strongly supported by the excellent agreement between the two extrapolation methods and high linear correlation coefficients (r > 0.999). This study contributes to the literature by providing essential physicochemical characterization data for Ambrisentan, which is vital for future pharmaceutical applications and analytical method development.