New method for preparation of hydrophobic interaction chromatographic stationary phases based on polymer-grafted silica and their chromatographic properties



A new “grafting to” method for the preparation of polymer-grafted silica hydrophobic interaction (HIC) packings was used in following steps. Firstly, 3-mercaptopropyltrimethoxysilane was bonded on silica to obtain SH-group modified silica. Secondly, glycidylmethacrylate (GMA) was coated on the surface of SH-silica to obtain polymer-grafted epoxide-silica. Finally, the polymer-grafted epoxide-silica was dispersed in polyethylene glycol of dioxane solution to produce HIC stationary phase. The prepared HIC packings have advantages for biopolymer separation, high column efficiency and good resolution for proteins. The dynamic protein loading capacity of the synthesized packings was 36.0 mg/g. Six proteins were fast separated in less than 12.0 minutes using the synthesized HIC stationary phases. Then the stationary phase was evaluated in detail to determine the effects of pH of mobile phase, concentration of ammonium sulfate and the temperature on the retention of proteins. The thermodynamic parameters of these proteins are consistent with the entropically driven nature of hydrophobic interactions, The HIC column was used for purification of trypsin from a crude extract solution with only one step. The purity of the purified trypsin was more than 95%. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2010