Development of Immobilized 8 -Galactosidase Bioreactor for Processing Lactose in Dairy Fluids

The aim of this study was to develop an immobilized 8 - galactosidase bioreactor for processing lactose in dairy fluids. The role of low lactose milk and whey in diet and preventive health care has been discussed. The recycling of lactose containing dairy fluids to food manufacture has been strongly recommended. A literature survey of research on immobilization of 8 -galactosidase and immobilized enzyme bioreactor indicate diffusion as one of the main problems in reactor modelling. In this study diffusion was minimized by immobilizing the enzyme on an impervious surface by covalent method. Tricel, a cellulose triacetate fibre has been used as the support. A derivative fibre was obtained by controlled hydrolysis of surface acetate groups. The surface hydroxyl groups were activated by the method given by Mirion and Wilchik (1987). The method was modified to circumvent the solubility of the fibres in acetone (Shukla et al, 1987). Steric hindrance by direct attachment of the enzyme to the fibres resulted in very low yields. Spacers of various lengths were incorporated to reduce the steric hindrance (Chaplin et al, 1987). Studies on the inactivation of 8 - galactosidase by the coupling agent (EDC) showed that the inactivation was probably due to binding at the active site. The protection of the active site by the substrate during the immobilization gave higher yields. The immobilization method was optimized and a protocol for immobilizing the enzyme on the cellulose triacetate fibres has been given. Physical and chemical methods to monitor the immobilization process were studied. FTIR spectroscopy allowed rapid monitoring of the immobilization process and further research on this aspect has been suggested. The kinetics of the free and immobilized enzyme were studied. The kinetic constants for the free and immobilized enzyme were found to be similar. The optimum pH was dependent on the type of spacer used. Studies on the inhibition of B-galactosidase by galactose showed non-competitive type of Inhibition (Shukla and Chaplin, 1993). This was contradictory to the studies reported previously in the literature. A laboratory scale fibre reactor was developed. The residence time distribution at different flow rates showed that an ideal plug flow was approached at a flow rate of 82 ml minute-'. The immobilization was carried out in-situ. Lactose hydrolysis in a recycle reactor showed that the external diffusion was minimized at a flow rate of 54 ml minute-1. The reactor performance fitted the non-competitive inhibition model. Coimmobilization of galactose-ioxidase to remove galactose has been suggested to increase the reactor performance and stability.