Abstract:

A novel packing for high performance gel filtration chromatography (GFC) was synthesized and characterized. High porosity silica prepared by base-dissolving method was used as the matrix. γ-(2,3-Epoxy propoxyl) propyltrimethoxysilane was used as the ligand and covalently bonded onto the silica matrix. After acidic hydrolysis, the epoxy groups were converted to the diol groups. Because a condensation tube filled with water at 70 ℃ was used in the grafting reaction, the resulting methanol could easily be discharged from the reaction system to shift the reaction equilibrium to achieve high ligand density. The hydrolysis condition greatly affects ligand density and column efficiency. The high column efficiency is observed when the ligand density is between 2.6 and 3.5 μmol/m2. Several proteins, such as cytochrome C, chymotrypsin, ovalbumin, bovine serum albumin, aldolase, ferritin, insulin, γ-globulin, phosphorylase, actin, carbonic anhydrase, were used to characterize the separation properties of the resulting high performance GFC column. It was shown that the excluded limit of relative molecular mass for the separation of bio-molecules was 300000. The recovery yield of bovine serum albumin was 99%.

Key words: diol group, high porosity silica, protein separation , packing for high performance gel filtration chromatography