Immobilization of Pectinases by Sequential Layering on Chitosan Beads

Abstract

Chitosan beads were prepared as a support material for immobilization of pectinases using a sequential layering approach. Glutaraldehyde (GA) was used as an activating agent for covalent binding. Different concentrations of cross-linking agent were examined and the highest specific activity of 7.8 U/mg was observed at 3% GA concentration, which was used in the multilayered immobilization studies. Three layers of pectinases were covalently immobilized on the support. As the layer number increased the total bound protein content and the activity of the chitosan beads increased; however, their specific activity significantly decreased from 9.1 U/mg in the first layer to 2.9 U/mg in the 3-layered structure. The results suggested that increasing the density of pectinases on chitosan support might have decreased the catalytic ability of enzymes due to either restriction to the protein backbone or due to substrate accessibility limitations. Therefore, optimal protein loading should be pursued for utilization of immobilized pectinases with highest specific activity.

Keywords: Enzyme Immobilization, Pectinase, Chitosan Support, Glutaraldehyde