Epoxy-derived pHEMA membrane for use bioactive macromolecules immobilization: Covalently bound urease in a continuous model system

Poly(2-hydroxyethylmethacrylate) (pHEMA) membranes were prepared by UV-initiated photopolymerization of HEMA in the presence of an initiator (α-α′-azobisisobutyronitrile, AIBN). The epoxy group, i.e., epichlorohydrin, was incorporated covalently, and the urease was immobilized onto pHEMA membranes by covalent bonding through the epoxy group. The retained activity of the immobilized enzyme was found to be 27%. The Km values were 18 and 34 mM for the free and the immobilized enzymes, respectively, and the Vmax values were found to be 59.7 and 16.2 U mg−1 for the free and the immobilized enzyme. The optimum pHs was 7.2 for both forms, and the optimum temperature for the free and the immobilized enzymes were determined to be 45 and 50°C, respectively. The immobilized urease was characterized in a continuous system and during urea degradation the operational stability rate constant for the immobilized enzyme was found to be 5.83 × 10−5 min−1.