Adsorption Equilibria of Proteins in Ion-Exchange Chromatography

The design and optimization of ion-exchange processes for the separation of biomolecules of interest (e.g. monoclonal antibodies) can be speeded up and fastidious screening experiments can be avoided by using accurate simulation models. In this research project, a model for the protein adsorption isotherm as a function of the pH and of the salt concentration is developed. It is based on a fundamental description of the colloidal interactions in ion-exchange chromatography and, in particular, of the effect of the pH and of the salt concentration on those interactions.

This kind of interactions has been successfully modelled for colloid particles, especially for the determination of the aggregation conditions. Proteins, by their size and their charged surfaces (polypeptide molecule with ionizable surface groups) are similar to small colloids (colloids' sizes are in the order of 10 - 1000 nm). Thus, this colloidal approach is also suitable to characterise the electrostatic interactions between a charged protein and the surface of an ion-exchange material. Using this model, the adsorption equilibrium is directly related to the structure of the protein and of the ion-exchange surface and it explicitly takes into account the pH and the salt concentration of the eluent. Experimentally, the retention time of a protein in diluted conditions provides a direct measurement of its adsorption equilibrium constant.

Furthermore, the interactions between loaded proteins become important as the concentration on the ion-exchanger surface is increasing. Following the same approach as for the protein-surface interactions, the effect of the pH and of the salt concentration on the saturation capacity is studied.

The model offers a basis for the rational optimization of ion-exchange processes. By including the adsorption isotherm in a simulation model, it is possible to predict the output of the chromatographic separation. The explicit pH and salt dependence of the adsorption equilibrium model allows the in silico exploration of a broad range of operating conditions. The experimental work can thus be focused on the most promising conditions. Furthermore, such a physically-based model can be used in a 'Quality by Design' approach, as encouraged by pharmaceutical regulatory agencies.

Contact Person: Rushd Khalaf