Recent developments have shown that enhanced selectivities can be achieved in multimodal chromatography as compared to traditional single-mode chromatography. Multimodal interactions in these systems exist between four key components: proteins, multimodal ligands immobilized on a resin, fluid phase modifiers and water. The complex interplay of these interactions, multimodal of otherwise, can result in unique selectivities, reflected in retention behavior of proteins. While protein-ligand interactions are dependent on the chemistry of the ligands and the structural arrangement of their constituent modes, these interactions can be further modulated by fluid phase modifiers. Previous work in our group in multimodal cation exchange chromatography of a library of proteins had suggested protein-specific effects of these cosolvents. This was further probed by performing a series of simulations with fluid phase modifiers to evaluate the interactions of these cosolvents with proteins and how they enhance/reduce protein-ligand binding. Protein surface characterization techniques were also employed to evaluate the strength and importance of electrostatic and hydrophobic interactions in these systems. The results presented here can be used to not only explain protein retention behavior in multimodal chromatographic systems, but also to design specific ligand-modifier conditions for challenging bioseparations and facilitate downstream process development.

Modulating protein-ligand interactions in chromatography using ligand chemistry and fluid phase conditions

247th National Meeting (2014)

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