Theoretical and Experimental Studies of the Effect of Pressure on Solute Retention in Liquid Chromatography

Pressure is often assumed to have a negligible influence on solute retention in liquid chromatography because of the small compressibility of the mobile and stationary phases. The range of pressures commonly encountered in reversed-phase separations is considerable, however, and may give rise to significant changes in solute capacity factor. In this study, the retention of model solutes is measured directly along the chromatographic column as a function of the local pressure. The model solutes, a homologous series of derivatized fatty acids, exhibit a significant increase in capacity factor ranging from +9.3% for n-C10 to +24.4% for n-C20 for inlet pressures from 1500 to 5000 psi. These experimental results are compared with a thermodynamic model derived from regular solution theory. This model suggests that state effects alone are not sufficient to describe the measured change in solute retention and that variations in interaction energy with density must also be considered. By using the simple relationship of van der Waals for the interaction energy (E ∝ 1/V), the change in capacity factor with density is slightly underestimated. However, by using an extended relationship that better describes polar fluids (E ∝ 1/V2), good agreement is observed. Finally, the correlation of experimental results with this thermodynamic model reveals that all components in the chromatographic system, including the solute, mobile phase, and stationary phase, must be considered compressible. The results of this study have clear implications for the determination of fundamental physicochemical parameters, as well as for the everyday practice of liquid chromatography.