Preparation of reversibly immobilized lysozyme onto Procion Green H-E4BD-attached poly(hydroxyethylmethacrylate) film for hydrolysis of bacterial cells

The flat sheet support for enzyme immobilization was prepared by UV-initiated photopolymerization of 2-hydroxyethyl-methacrylate (HEMA) in the presence of an initiator (αα′-azoisobutyronitrile; AIBN). An affinity dye, Procion Green H-E4BD, was attached covalently under alkaline conditions and the pHEMA-Procion Green H-E4BD-attached film was used for the immobilization of lysozyme via adsorption. The amount of attached dye on the pHEMA film was 160 μmol m2 and the water content of the dye-attached pHEMA film was 69%. The lysozyme adsorption capacity of the dye-attached pHEMA film was determined under conditions of different pH and with different initial concentrations of enzyme in the medium. The maximum lysozyme adsorption capacity of the dye-attached pHEMA film, under the specified experimental conditions was 3.92 g m−2. Non-specific adsorption of the lyzozyme on the pHEMA film was negligible. Optimum reaction pH was 6.0 for the free and 7.0 for adsorbed enzyme. The free enzyme had an optimum temperature of 35°C, whereas it shifted to 40°C for the immobilized enzyme system. The enzyme could be repeatedly adsorbed and desorbed from the dye-attached pHEMA film without any significant loss in adsorption capacity.