Analysis of Group Retention Contributions for Peptides Separated by Reversed Phase High Performance Liquid Chromatography

Abstract

Within the framework provided by solvophobic theory, selectivities for unprotected peptides separated on fully porous, microparticulate, chemically bonded alkylsilicas can be ascribed to differences between the effective hydrophobic contact areas of the solutes. Furthermore, this theoretical treatment predicts that retention behaviour differences can be evaluated from topological parameters which accomodate the influence of amino acid side chain and end group contributions in the retention process. With data obtained for 57 peptides, including a variety of peptide hormones, eluted under the same conditions from a μBondapak C₁₈ column, these predictions have been rigorously tested using two methods of numerical analysis. The results provide further evidence that the hydrophobic group retention contributions of the amino acid residues in small peptides have an essentially additive effect on peptide retention with alkylsilicas. Divergences in retention behaviour are interpreted in terms of specific silanophilic and solvation interactions.