Immobilization of laccase onto polyethyleneimine grafted chitosan films: Effect of system parameters

Chitosan films were prepared via phase-inversion technique. The polyethyleneimine (PEI) was grafted onto chitosan films after activation with various agents, such as epichlorohydrin, genipin and glutaraldehyde. The PEI grafted activated chitosan films (Ch-g-PEI) were used for immobilization of laccase by adsorption. The effects of adsorption parameters on immobilization efficiency were researched. Activity and stability of immobilized laccase on Ch-g-PEI films were investigated under desired optimum conditions. Characterization studies of the supports were also performed. The effect of pH on the activity of free and immobilized laccase was carried out in the pH range 4.0-8.0, and the optimum pH was determined as 5.5. Kinetic constants of the free and immobilized laccase were calculated. The maximum activity (V (max) ) and Michealis constant (K (m) ) of immobilized laccase on the films were found to be 71.2 U/mg and 0.51 mM, respectively. Thermal stability of the enzyme increased together with immobilization on the support. Free laccase retained 39% of its original activity at 55 A degrees C for a 120 min incubation period. Immobilized laccase retained 72% of its original activity for the same period at 55 A degrees C. The free enzyme lost all of its activity during the 7 weeks storage period. Conversely, immobilized laccase lost 36% of its activity during the 7 weeks storage period.