Interactions of basic compounds in reversed-phase high-performance liquid chromatography - Influence of sorbent character, mobile phase composition, and pH on retention of basic compounds

Differences in a degree of base deactivation of selected reversed-phase sorbents were evaluated. Five silica-based sorbents (SGX C-18, LiChrosorb RP-18, LiChrospher RP-select B, Purospher RP-18, Symmetry C-18), a polybutadiene-coated alumina (Aluspher RP-select B) and 2-hydroxyethylmethacrylate-based stationary phase (HEMA-BIO 1000 C-18) were compared. The best results were obtained for Symmetry C-18 and HEMA-BIO 1000 C-18. For testing purposes a set of nineteen basic compounds differing in pK(a) constants and hydrophobicity was used. The bases possessing high pK(a) values (pK(a)>9.0) and/or low hydrophobicity proved to be the best indicators of sorbent surface deactivation. Dependencies of capacity factor on pH of the mobile phase were measured. Shape deviations between the sigmoidal theoretically predicted and experimentally obtained curves evidence for a complex retention mechanism in which not only hydrophobic but also other interactions, for example, ion-exchange, participate. The effect of methanol content in the mobile phase on the dissociation of buffer and a basic analyte used was studied. The characteristic shifts of k versus pH of the mobile phase dependencies caused by the addition of methanol were found to be in a good agreement with theory.