Computer simulation for the prediction of separation as a function of pH for reversed-phase high-performance liquid chromatography. I. Accuracy of a theory-based model.

Computer simulation software (DryLab I/mp) is described for predicting high-performance liquid chromatographic separation as a function of changes in mobile phase pH. Three experimental runs with pH (only) varied are used to derive values of pKa plus capacity factors (k') for the ionized and non-ionized form of each ionizable solute. Various tests of the experimental data then allow classification of each solute as acidic, basic, neutral (including strong or weak acids or bases) and amphoteric. Experimental data are reported for the separation of several substituted anilines as a function of pH and solvent composition (%B). Experimental requirements for the accurate prediction of separation (ca. +/- 2-4% in alpha) as a function of pH are discussed. The reliability of the software is demonstrated for three different samples: mixtures of (a) substituted benzoic acids, (b) substituted anilines and (c) catecholamine-related compounds.