Cross-linking of fungal β-glucosidase on Al₂O₃ nanocrystals synthesized using Cajanus cajan L. Millsp. extracts for in suit genistein manufacture

This present study deals with the cross-linking of fungal β-glucosidase on Al₂O₃ nanocrystals (NCs) synthesized in C. cajan for in-suit genistein production. The Cajanus cajan leaves were dried and used to prepare their extract at 65 °C by agitation for 30 min. For enzyme production under submerged culture, 50 mL of medium at pH 8.6 with an inoculum volume of 2 mL; was incubated for 72 h with optimized parameters at 30 °C. The Al₂O₃ NCs were synthesized by adding 30 mM Al₂NO₃ to 25 mL of leaf extract with NaOH at 65 °C for 50 min which enhanced the β-glucosidase specific activity when immobilized. Genistein by biotransformation was obtained using both free (0.67 ± 0.42 mg/mL) and Al₂O₃ immobilized β-glucosidase (1.3 ± 0.66 mg/mL) for 48 h. The substrate level and enzyme concentration were 2.5 and 1 mL respectively. The UV visible spectra for leaf extract; free and cross-linked β-glucosidase and Al₂O₃ NCs were at 225, 235, 300, and 210 nm. The bands for Al₂O₃ NCs were achieved at 500–750 cm^{− 1} which showed the FTIR analysis to check the change in functional groups of free and Al₂O₃ cross-linked β-glucosidase. In XRD analysis, peaks depicted the crystalline structure of Al₂O₃ NCs ranging from 10–50°. The size of NCs was confirmed by using different magnifications (1.01, 2.00, 3.00, 5.00, 7.02, and 10 K X) of SEM images obtained. For zeta potential measurements, the peak was obtained at ˗21.0 mV.