Model Development for Hydrophobic Interaction Chromatography

Understanding of water structure on hydrophobic surfaces forward model-based process development for Hydrophobic Interaction Chromatography

Figure: Equilibrium adsorption data points from high-throughput experimentation and resulting isotherms. Plot shows case study for glucose oxidase.

Hydrophobic interaction chromatography (HIC) bases on a separation of molecules with respect to their hydrophobicity. It is a commonly used method for intermediate and polishing steps in protein purification.

HIC process development generally relies on experimentation. Different to ion exchange chromatography, HIC inheres an intricate chromatographic behavior. The slow and volatile binding as well as the complex salt-dependent protein-ligand interaction challenges modeling, in particular the derivation of suitable adsorption isotherm models.

New insights into water structure on hydrophobic surfaces. Various works in the area of hydrophobic surfaces were considered to derive a new mechanistic isotherm for modeling HIC. This model approach assumes an equilibrium between well-ordered and bulk-like ordered water molecules on the hydrophobic surface. Additionally, the hydration number of ions was considered to be a function of the ionic strength to incorporate salt effects. This novel isotherm was implemented in ChromX by means of its plug-in interface and is also available in ChromX' enterprise version.

Prediction of protein adsorption and desorption behavior. With the new mechanistic isotherm, accurate HIC modeling could be realized. Using this concept, protein adsorption and desorption behavior for any given binding and elution salt concentration can now be predicted.

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