Laccase immobilization via cross-linked aggregate preparation: Characterization, thermodynamic/kinetic properties and application in removal of bisphenol A from solutions

Abstract Fungal laccase from Aureobasidium pullulans was immobilized via cross-linked enzyme aggregate (CLEA) preparation under statistically optimized conditions. The stability of the CLEA to heat inactivation was studied via investigation of its thermodynamic and kinetic parameters. The immobilized enzyme was then deployed in the biodegradation of a bisphenol-A (BPA). The optimum conditions for CLEA preparation resulting in the highest immobilization yield were ammonium sulphate (60% v/v), glutaraldehyde (30 mM), pH (4.5), time (6 h) and temperature (55ºC). The CLEA retained about 51% of its activity after eight catalytic cycles. The optimum pH and temperature of the laccase CLEA were 5.5 and 60ºC respectively. The SEM indicated that the laccase CLEA was type II (unstructured). The data obtained from the heat inactivation kinetics and thermodynamic characterization indicated that the CLEA was stable to heat denaturation than the free enzyme. The kinetic parameters obtained for the CLEA with ABTS as substrate were 101.3 µM, 2.94 µmols-1mg-1 and 0.03 dm3s-1mg-1 for the Km, Kcat and Kcat/Km respectively. The optimum conditions for BPA biodegradation using the CLEA were temperature (55ºC), time (2 h), CLEA (1.0 mg) and BPA concentration (40 mg/L). After the 7th cycle, laccase CLEA retained about 63 ± 2.3% biodegradation efficiency. A heat-resistant laccase CLEA was able to remove BPA from solutions under statistically optimized conditions. The laccase CLEA has properties for other futuristic applications.