Invertase immobilized on spacer-arm attached poly(hydroxyethyl methacrylate) membrane: Preparation and properties

Abstract

Microporous poly(2-hydroxyethyl methacrylate) (pHEMA) membrane was prepared by UV-initiated photopolymerization. The spacer arm (i.e., hexamethylene diamine) was attached covalently and then invertase was immobilized by the condensation reaction of the amino groups of the spacer arm with carboxyl groups of the enzyme in the presence of carbodiimides. The values of the Michael's constant K-m of invertase were significantly larger (ca. 2.5 times) upon immobilization, indicating decreased affinity by the enzyme for its substrate, whereas V-max was smaller for the immobilized invertase. Immobilization improved the pH stability of the enzyme as well as its temperature stability. Thermal stability was found to increase with immobilization and at 70 degrees C the half times for the activity decay were 12 min for the free enzyme and 41 min for the immobilized enzyme. The immobilized enzyme activity was found to be quite stable in repeated experiments. (C) 2000 John Wiley & Sons, Inc.