Preparation of Chitosan/Magnetic Porous Biochar as Support for Cellulase Immobilization by Using Glutaraldehyde

In this work, porous biochar was obtained from sugarcane bagasse by alkali activation and pyrolysis and then magnetized with γ-Fe₂O₃ by calcination. After functionalization with chitosan and activation with glutaraldehyde, the as-prepared chitosan/magnetic porous biochar served as a support to immobilize cellulase by covalent bonds. The immobilization amount of cellulase was 80.5 mg cellulase/g support at pH 5 and 25 °C for 12 h of immobilization. To determine the enzymatic properties, 1% carboxymethyl cellulose sodium (CMC) (dissolved in 0.1 M buffer) was considered as a substrate for hydrolysis at different pH values (3-7) and temperatures (30-70 °C) for 30 min. The results showed that the optimum pH and temperature of the free and immobilized cellulase did not change, which were pH 4 and 60 °C, respectively. The immobilized cellulase had a relatively high activity recovery of 73.0%. However, it also exhibited a higher Michaelis-Menten constant (K_m) value and a slower maximum reaction velocity (V_max) value compared to the free enzyme. In the reusability assay, the immobilized cellulase showed initial glucose productivity of 330.9 mg glucose/g CMC and remained at 86.0% after 10 uses. In conclusion, the chitosan/magnetic porous biochar has great potential applications as a support for enzyme immobilization.