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Abstract
The adsorption behaviors of hydrophobic charge induction chromatography (HCIC) adsorbents with different functional ligands were investigated with immunoglobulin of egg yolk (IgY) as a model antibody. The adsorption isotherm and retention behavior in the column were studied, and the influences of the ligand structure and the pH on the adsorption were discussed. The results indicated that the pI of the target protein and pKa of HCIC ligand are the important parameter to determine the maximum adsorption pH of HCIC adsorbent, and high adsorption of IgY was found at pH 5 for all five adsorbents tested. Some differences could be found for different HCIC adsorbents, and the ligand structure influenced pH effect on the binding/elution of target protein. 2-mercapto-1-methyl-imidazole (MMI) ligand with a sulfone group showed a high adsorption capacity and strong pH-sensitivity, which would be more suitable for antibody purification. Moreover, the retention experiments indicated that IgY could be efficiently eluted from the adsorbents with 4-mercapto-ethyl-pyridine (MEP) or MMI as the ligand at acid conditions, while 2-mercapto-benzimidazole (MBI) ligand showed some difficulties on the elution. The retention study would help in defining not only the effective pH of elution for a given protein but also the elution efficiency of a given adsorbent.
ACKNOWLEDGEMENTS
This work was supported by the National Natural Science Foundation of China, the Program for New Century Excellent Talents in University (NCET), and the Fundamental Research Funds for the Central Universities.