Liquid chromatographic separations with mobile-phase additives: influence of pressure on coupled equilibria

On the basis of equilibrium thermodynamics, pressure can cause a shift in equilibrium for any interaction that exhibits a change in partial molar volume. This shift in equilibrium can be observed in liquid chromatography as a pressure-dependent shift in solute retention. In this paper, the impact of pressure on liquid chromatographic separations with mobile-phase additives is examined from both theoretical and experimental perspectives. The theoretical development for coupled-equilibria separations shown here is general and can be applied to any separation using mobile-phase additives. Predictions indicate that the coupled nature of these equilibria leads to pressure-induced perturbations in partitioning and complexation that can either compete with or complement one another. Using positional isomers and enantiomers as model solutes, experimental retention observations are fully consistent with these predictions, showing the diminution of individual pressure effects for competing cases and enhanced pressure effects for complementary cases. When pressure-induced changes in capacity or retention factor differ between individual solutes, changes in solute selectivity are predicted and observed. Using a C18 stationary phase with beta-cyclodextrin as the mobile-phase additive, solutes studied here exhibit changes in selectivity ranging from - 7 to + 10% for a change in average pressure of approximately 215 bar. Perhaps the most dramatic change in selectivity is observed for the separation of positional isomers where pressure-induced changes in selectivity actually reverse solute elution order.