An efficient chitinase from Bacillus amyloliquefaciens M6: Immobilization to ZnO/RGO nanocomposite and its application in biocontrol

In this study, the chitinase enzyme purified from Bacillus amyloliquefaciens M6 was immobilized on Zinc oxide/reduced graphene oxide nanocomposites (ZnO/RGO NCs), and its activity on Rhyzopertha dominica and Oryzaephilus surinamensis was tested. To optimize the culture medium, the M6 strain was developed under varying conditions, and the optimum substrate concentration, temperature, pH, and incubation time for chitinase production were determined as 5 g/L chitin, 35 degrees C, 6.0 pH, and 24 h. Compared to its activity prior to optimization (10.42 U/mL), the chitinase activity of the strain was observed to increase by 2.2 times. Under these conditions, chitinase was purified 6.75-fold from strain M6 with a specific activity of 228.90 U/mg (activity: 11,48 U/mL) and a yield of 1.84 %, and its molecular weight was determined to be similar to 50 kDa. Chitinase was immobilized on ZnO/RGO NCs with 99.68 % immobilization efficiency and 94.30 % recovered activity. The optimum pH and temperature values of free and immobilized chitinase were determined as 6.0 and 45 degrees C. The activity of immobilized chitinase against R. dominica and O. surinamensis was tested and the LC50 and LC90 values for R. dominica were calculated as 1.8 mg/mL (32.8 U/mL) and 3.8 mg/mL (69.2 U/mL) and 4.8 mg/mL (87.4 U/mL) and 10.0 mg/mL (182.0 U/mL) for O. surinamensis. Thus, the obtained results concluded that the immobilized chitinase proved an alternative biocontrol agent to hazardous synthetic pesticides, the application of this biocontrol agent to make biopesticides will prove more reliable for sustaining the crop productivity and in crop protection.