Abstract: Mixed-mode chromatography is a new technology for the purification of immunoglobulin (IgG). Previous researches indicated that a certain concentration of sodium caprylate (NaCA) could significantly reduce the adsorption of serum albumins and improve the efficiency of mixed-mode chromatography. With MEP HyperCel resin and bovine serum albumin (BSA) as the model, the interactions between NaCA and BSA were investigated by static adsorption and isothermal titration calorimetry (ITC) in the present work. The influences of concentration of NaCA, pH, salt addition and temperature were discussed. The static adsorption experiments showed that the adsorption capacities of BSA and IgG firstly decreased and then increased with increasing NaCA concentration. The influence of NaCA on BSA was much obvious than IgG. ITC analysis indicated that the interactions between NaCA and BSA were quite stronger than those for IgG, and the hydrophobic force dominated the NaCA-BSA interactions. With NaCA addition, the adsorption of BSA decreased with increasing pH value or temperature, and different salts showed varying impacts. ITC analysis also demonstrated that the NaCA-BSA interactions were influenced by pH, salt addition and temperature. In general, ITC results were consistent with those of adsorption experiments, which demonstrated that ITC could be used to evaluate the interactions between small molecule and protein, then study the mechanism of specific additive on protein adsorption.

Key words: mixed-mode adsorption, sodium caprylate, bovine serum albumin, isothermal titration calorimetry

摘要： 混合模式吸附是一种抗体分离新方法，适量添加辛酸钠，可以减弱血清白蛋白等杂质吸附，提高抗体结合的选择性。本文以MEP HyperCel介质和牛血清白蛋白（BSA）为模型，结合静态吸附平衡和等温滴定量热（ITC）法，考察了辛酸钠浓度、pH、盐和温度等影响，探讨了辛酸钠的作用机制。随辛酸钠浓度增大，BSA和牛血免疫球蛋白的饱和吸附量均呈现先降低后升高的趋势，不过辛酸钠对BSA影响更显著，ITC分析表明辛酸钠与BSA之间存在较强的相互作用，且以疏水作用为主导。添加辛酸钠后，BSA吸附量随pH或温度升高而降低，不同盐具有不同效果，ITC分析表明pH、盐和温度均不同程度影响辛酸钠-BSA相互作用，从而影响MEP HyperCel吸附BSA。结果表明，等温滴定量热分析与宏观吸附现象相一致，可量化分析小分子-蛋白间相互作用，为研究蛋白吸附过程中小分子添加物的作用机制提供了新思路。

关键词: 混合模式吸附, 辛酸钠, 牛血清白蛋白, 等温滴定量热