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Abstract
In this study, monolithic high performance liquid chromatography (HPLC) composite columns were synthesized for immunoglobulin G (IgG) separation by boronate affinity chromatography. 4-Vinyl phenyl boronic acid (VPBA) was polymerized with 2-hydroxyethyl methacrylate (HEMA). The poly(HEMA-VPBA) monoliths were crushed into fine particles by using ball-milling. Then, the crushed particles were embedded into poly(2-hydroxyethyl methacrylate) (PHEMA) cryogels to prepare monolithic HPLC composite columns. The PHEMA cryogel was also synthesized to evaluate the efficiency of the embedding process. The monolithic HPLC composite columns were characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), surface area measurements, boron content analysis, swelling studies, and flow rate-back pressure relation. The performance of IgG separation of monolithic composite columns was determined with HPLC and the parameters such as pH, IgG concentration, temperature, flow rate, and ionic strength were investigated. The maximum adsorption was observed at pH 8.0 phosphate buffer. The IgG separation from human plasma was also performed and selectivity experiments were carried out with human serum albumin (HSA) and hemoglobin (Hb) as competitors.