Immobilization of enzymes on porous membranes

In this thesis, the results from a theoretical and experimental investigation of enzyme immobilization in porous membranes is reported. A theoretical model of the immobilization process which accounts for restricted diffusion of enzyme in the pores of the membrane has been developed. The model predicts the effect of immobilization kinetics and time of immobilization on the enzyme distribution in the pores of the membrane.; Experiments were performed using alumina membranes obtained from Anotec Separation Inc. Characterization experiments revealed an ideal porous structure in these membranes. The pore density and the pore length of the membranes were determined using scanning electron microscopy, while the pore radius was accurately measured by independent solvent flow and solute diffusion experiments. The immobilization of glucose oxidase and glucose oxidase-biotin conjugate on porous alumina membranes was experimentally investigated. Enzyme uptake data was correlated to the theory to determine the rate constant of immobilization. Immobilization studies were carried out for enzyme adsorption and covalent coupling.; The distribution of enzyme was experimentally studied by assembling five membranes together in the diffusion cell. Following immobilization, the membranes were separated and each membrane was assayed for activity. The amount of enzyme present in each membrane yielded a discrete distribution that compared well with that predicted by theory.; Preliminary activity studies of the immobilized enzyme membrane were conducted using a flow through reactor configuration and a crushed particle reactor configuration. A 30% loss in activity was observed on comparing the immobilized enzyme activity with that of the free enzyme. The effect of temperature on the immobilized enzyme activity was studied to examine the effect of immobilization on enzyme stability. Comparing the free enzyme activity with that of the immobilized enzyme indicated that immobilization apparently imparts some resistance to thermal deactivation.