PROTEIN RETENTION IN HYDROPHOBIC INTERACTION CHROMATOGRAPHY - MODELING VARIATION WITH BUFFER IONIC STRENGTH AND COLUMN HYDROPHOBICITY

摘要

The variation in protein retention times with protein and surface hydrophobicity and mobile phase composition and concentration has been described with a simple thermodynamic model. Column capacity factors for two proteins have been measured as a function of mobile phase ionic strength for a series of columns with varying levels of hydrophobicity. Application of the model to these data suggests that the protein retention is dominated by the release of water molecules upon adsorption, which is consistent with the entropically driven nature of hydrophobic interactions. The calculated number of water molecules released agrees with estimates based on the reduction in hydrophobic surface area for adsorption.