Study on Papain Immobilization on a Macroporous Polymer Carrier

Macroporous resin microbeads of methyl methacrylate-divinyl benzene copolymer were synthesized by radical suspension polymerization of acrolein with divinylbenzene in the presence of a pore-creating agent, petroleum ether. The microbeads had a large specific surface area and large pores covered the entire surface of the resin. This macroporous polymer carrier was aminated by hydrazine hydrate that produced a large number of amino groups on the carrier. Papain was immobilized on the porous polymer carrier by glutaraldehyde cross-linking or diazo-coupling. The factors involved in the activity recovery of the immobilized papain and the enzymic properties of the resulting immobilized papain were studied in comparison with free papain, for which casein was chosen as the substrate. The results show that the activity recovery of immobilized papain can reach 60.1%. At the same time, the stability, resistance against inhibition, Michaelis constant Km, and reusability of the immobilized enzyme were also investigated. The proteolytic residual activity of free enzyme, using casein as substrate, was 100-50% at 37-60 °C. The proteolytic residual activity of immobilized papain, reached 100-50% although the temperature increased to 70-90 °C. The optimum range of pH was 5-10 for immobilized papain and 5-7.8 for free enzyme at 37 °C. Its residual activity was 98% when immobilized enzyme had been stored at 0-4 °C for 30 days. The results indicated that the immobilized papain by this method had not only higher activity recovery, but also remarkable stability, better reusability and environmental adaptability than free papain.