Factors Affecting the Separation of Arginine Vasopressin Peptide Diastereoisomers by HPLC

Abstract

Experimental conditions and parameters involved in HPLC separations of the peptide hormone arginine vasopressin and some of its diastereoisomers on several reverse phase columns were investigated. The effects of percent carbon loading on an octadecyl reverse phase column, carbon chain length of the bonded phase, concentration of the buffer, and organic solvent were examined. Using the appropriate solvent systems, arginine vasopressin was separated from each of its diastereoisomers with most solvent systems studied, but the order of elution of the diastereoisomers was dependent on the column employed. Separation of the peptides was only part of the goal. A continuing study to understand the interactions of the peptides with the stationary phase as a function of structure was also undertaken. This lead to the conclusion that the choice of column as well as solvent affect the separation of peptide diastereoisomers and that both the eluting strength of the mobile phase and the stationary phase chemical composition must be considered.