A novel polymer-grafted hydrophobic charge-induction chromatographic resin for enhancing protein adsorption capacity

Abstract

Hydrophobic charge-induction chromatography (HCIC) is a developing technology for antibody purification. To enhance the protein adsorption capacity, a novel polymer-grafted HCIC resin was developed, in which the surface-initiated activator generated by electron transfer (AGET) atom transfer radical polymerization (ATRP) was explored as a controlled polymerization technique to reconstruct matrix structure and ligand distribution. Using poly(glycidyl methacrylate, GMA) as grafting polymer and 2-mercapto-1-methyl-imidazole (MMI) as functional ligand, poly(GMA)-grafted HCIC resins were prepared with series of grafting and ligand densities. Adsorption behaviors of human immunoglobulin G (hIgG) on the prepared resins demonstrated the necessity of controlling grafting and ligand density. Saturated adsorption capacity (Qm) and effective pore diffusivity (De) reached the maximum under medium ligand density when the grafting density was kept constant. The highest Qm and De values were found under the highest grafting density, which were 73% and 7.17times higher than the non-grafted resin, respectively. Column breakthrough tests indicated that the dynamic binding capacity of the resin with optimized grafting density and ligand density was up to 34.6mg/g when linear velocity was 300cm/h, which was 86.3% higher than dextran-grafted resin. The resin was then used to separate hIgG from a protein mixture (hIgG/human serum albumin=1:4), high purity (>99%) and recovery (>90%) of hIgG were found with 50-cycle reuses, which verified the selectivity and robustness of G-MMI resin prepared. In general, the surface-initiated AGET ATRP provides a controlled grafting strategy to improve protein binding capacity for chromatographic separation, and new resins developed have great potential in large-scale protein purification applications.