Specific and nonspecific adsorption in affinity chromatography.: Part I.: Preliminary and equilibrium studies

The amount of enzyme adsorbed at equilibrium by hydrophobic and hydrophilic bonds or retained inside the adsorbent was studied for asparaginase on Sepharose 4B, Sepharose 4B activated with CNBr, activated Sepharose 4B with hexamethylenediamine as a spacer arm and activated Sepharose 4B with hexamethylenediamine and L-(+)-chlorosuccinamic acid as the spacer arm and ligand, respectively, in a batch reactor for the range of temperatures 298- 302 K, the range of pH 7.5-8.6, and for different ionic strengths (0.0-1.5 M NaCl). Adsorption increased with pH and decreased with temperature. With respect to ionic strength, adsorption increased until an I value of 0.05 M NaCl. A change in the adsorption process was observed when the ionic strength was steadily decreased. The equilibrium data were correlated using a semiquantitative theory in which electrostatic and hydrophobic interactions between enzyme and ligand were considered. This correlation shows that hydrophobic effects increase with temperature, in very good agreement with the experimental data.