High-throughput and reliable assessments of the ionization constant of monoprotic organic acids through an arginine based mixed mode HPLC

The charge state of a molecule is the single most prominent attribute ruling out its interactions with the surrounding environment. In a previous study, the retention of acids on the new Celeris™ Arginine (ARG) column was found to be predominantly driven by electrostatics and, specifically, their charge state. Therefore, we analysed 41 compounds in liquid chromatography with ultraviolet detection to study possible relationships between the analytical retention on this phase and the pKa of the acidic solutes. Highly significant relationships were observed indicating either a linear (r2 = 0.86) or a quadratic (r2= 0.89) trend. To improve the throughput of the method, this was transferred to LC mass spectrometry, allowing the analysis of a molecule every 3 mins. The developed method was found to be fast, reliable, accurate, easily automatable and simple to set up. Finally, the analytical column’s being industrially manufactured and commercially available offers broad applicability.