Preparative Separation of Bioactive Protein on High Performance Hydrophobic Interaction Chromatography

Abstract

The separation by hydrophobic interaction chromatography (HIC) firstly described by Hjert6n (Hjerten 1973), is based on the differential hydrophobic surface interaction. In comparison with reversed phase chromatography (RG), a weak hydrophobic group is coupled onto the chromatographic support as stationary phase and a descending salt gradient is used as mobile phase in HIC. During the separation process by HIC the bioactive protein molecules are in an aqueous salt environment and a fractionation route is applied to isolate the bioactive protein which is similar to salting out procedures used since long for protein purification. As a result, HIC might overcome the major limitations of reversed phase chromatography, which usually induces losses of the bioactivity of the sample molecule, and keeps the separating ability depending on the hydrophobicity.