Immobilization of fungal cellulase on chitosan beads and its optimization implementing response surface methodology

Abstract

The present study deals with the immobilization of cellulase produced from our isolated fungus Aspergillus niger ISSFR-019 on different matrices of which chitosan beads has been found to be the most suitable one. The optimization of the amount of cellulase, amount of beads, temperature and amount of glutaraldehyde used as a cross-linking agent for the procurement of maximum immobilization efficiency has been done on the basis of statistical approach BBD (Box-Behnken Design). Cellulase is found to be successfully immobilized on the chitosan bead surface, with 94.2% efficiency and immobilization yield is found to be 54.1% after statistical optimization. Significant enzymatic activity is recorded after 12 cycles of reuse of the beads. The kinetic study revealed that the Michael’s Menten constant (K_m) is found to be 1.3 mM and 2 mM for free enzyme and immobilized enzyme respectively, the higher Km value for immobilized enzyme indicated conformational change in the enzyme that altered the accessibility of substrate towards the active site of the enzyme. The V_max is found to be 2.7 and 4.4 U/mg-1 respectively for the immobilized and free enzyme. Fourier Transform Infrared Spectroscopy studies revealed the change in absorbance of functional groups when the enzyme is bound to chitosan beads.