Abstract
Summary: Glucose oxidase (GOD) and catalase (CAT) were covalently immobilized onto three types of polyacrylonitrile (PAN 1, PAN 2, and PAN 3) ultrafiltration (UF) membranes with different pore sizes and one type of polyamide (PA) microfiltration (MF) membrane by the bifunctional reagent, glutaraldehyde. The initial membranes were pre‐modified to generate active amide groups in the PAN membranes and active amino groups in the PA membranes. The PAN 3 membrane contained the highest amount of active groups, and the membrane PA the lowest. The modified membranes were enzyme‐loaded by diffusion and convection (UF). The effect of membrane pore size and immobilization methods on enzymatic activity and bound protein were studied. The most effective immobilized system was prepared by diffusion using a PAN 3 membrane as a carrier (bound protein: 0.055 mg/cm^2^, relative activity: 87.6%). This membrane had the highest pore size of all the PAN membranes. Despite the highest pore size of PA membrane, the enzyme PA membranes prepared by diffusion showed the lowest amount of bound protein (0.03 mg/cm^2^) and the lowest relative activity (35.38%). This correlates with the lowest amount of active groups found in these membranes. The relative activity was higher for all the enzyme systems loaded by diffusion. The systems prepared by convection of the enzyme solution contained higher amounts of enzymes (0.035–0.13 mg/cm^2^ protein), which led to internal substrate diffusion resistance and a decrease in the GOD relative activity (21.55–68.5%) in these systems. The kinetic parameters (V~max~ and K~m~) and the glucose conversion of the immobilized systems prepared by diffusion were also studied.
Relative activity of GOD in immobilized GOD plus CAT membranes prepared by different methods of enzyme loading: diffusion and convection (UF). The bars represent the difference between relative activities of immobilized GOD prepared by two methods for all the membranes. The enzyme activity was determined under static (a) and dynamic (b) conditions; 1 M glucose, pH 5.8, and 28 °C.
magnified imageRelative activity of GOD in immobilized GOD plus CAT membranes prepared by different methods of enzyme loading: diffusion and convection (UF). The bars represent the difference between relative activities of immobilized GOD prepared by two methods for all the membranes. The enzyme activity was determined under static (a) and dynamic (b) conditions; 1 M glucose, pH 5.8, and 28 °C.