Covalent Immobilization of Multi-Enzyme Biocatalysts on Nano-Silica for Efficient p,p'-DDT Degradation

Abstract

Abstract Growing concerns over environmental pollution have necessitated the development of new-generation environmental protection technologies. Multi-enzyme biocatalysts offer a promising approach for reducing pollution caused by organic wastes. However, the recovery of free-form multienzymes is challenging, resulting in high costs and low production efficiency, limiting their application in bioremediation. Immobilization provides a solution by stabilizing and retaining enzymes. In this study, we devised a sensitive and stable enzyme biocatalyst by covalently immobilizing multi-enzymes onto nano-silica using glutaraldehyde. Ligninolytic enzymes (laccase, aryl alcohol oxidase, lignin peroxidase, and manganese peroxidase) were produced from Pleurotus ostreatus (NRRL-2366) under submerged fermentation. Enzymes were partially purified through ammonium sulfate precipitation and dialysis. These purified enzymes were immobilized on nano-silica. The resulting immobilized enzymes biocatalyst exhibited stability and activity across a pH range of 4 to 9 and a temperature range of 20 to 55°C. Immobilization of laccase, lignin peroxidase, manganese peroxidase, and aryl-alcohol oxidase achieved residual activities of 77%, 62.5%, 41.59%, and 28.21%, respectively, after three consecutive batches. Immobilized enzymes biocatalyst effectively degraded p,p'-DDT, and its complete degradation was achieved after incubation at pH 5 and 30°C for 12 hours, as confirmed by HPLC analysis. The GC-MS analysis revealed the detection of eleven major metabolites during the degradation process, which were utilized to predict the degradation pathway.