Responsive hydrophobic interaction membrane chromatography (HIMC) is distinguished by its high recovery and high resolution in protein purification and fractionation processes. Poly (N-vinylcaprolactam) (PVCL) is used as the thermo-responsive ligand. The lower critical solution temperature (LCST) of PVCL ligands in solution is sensitive to different salt types and salt concentrations. At temperatures above its LCST or high ionic strength, PVCL adopts a collapsed hydrophobic conformation leading to strong protein adsorption. At temperatures below its LCST or low ionic strength, PVCL adopts a coil-like hydrated conformation with low protein affinity. Here PVCL ligands have been grafted from micro-porous regenerated cellulose membrane (0.45μm) surfaces via atom transfer radical polymerization. Optimization of this novel HIMC system requires systematic investigations varying the salt type and salt concentration, which is also critical to understand the protein adsorption and desorption mechanisms in HIC. Therefore, dynamic binding capacities of our PVCL modified membranes were determined at different pH, salt type and concentration at room temperature with two model proteins (a human immunoglobulin4 (IgG4) and bovine serum albumin (BSA)). In addition, BSA adsorption isotherm in monovalent (Na+, NH4+), divalent (Mg2+, Zn2+) and trivalent (Al3+) sulfate salt solutions were determined. Finally, the responsiveness of the PVCL ligand synthesized in solution as well as grafted on silicon wafer was investigated and correlated with the performance of our HIMC.

Understanding the effect of salt ions on protein adsorption in novel responsive hydrophobic interaction membrane chromatography

249th National Meeting (2015)

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