pH-Dependent Retention Time Measurement for the Identification of Unknown Substances with LC-HRMS

The charge state of molecules containing an ionizable functional group has a large effect on their retention in reversed-phase liquid chromatography. The pKa values of many acidic or basic functional groups typically cover characteristic ranges. Therefore, differences in retention time could yield information about the presence/absence of functional groups in an unknown compound if analyzed with eluents of different pH, making it a promising approach for confirming or rejecting molecular structures for a given molecular formula in nontarget screening.