The Stability Improvement of Aspergillus fumigatus α-Amylase by Immobilization onto Chitin-Bentonite Hybrid.

Abstract

Enzyme immobilization is a powerful method to improve the stability, reuse, and enzymatic properties of enzymes. The immobilization of the α-amylase enzyme from Aspergillus fumigatus on a chitin-bentonite (CB) hybrid has been studied to improve its stability. Therefore, this study aims to obtain the higher stability of α-amylase enzyme to reduce industrial costs. The procedures were performed as follows: production, isolation, partial purification, immobilization, and characterization of the free and immobilized enzymes. The CB hybrid was synthesized by bentonite, chitin, and glutaraldehyde as a cross-linker. The free enzyme was immobilized onto CB hybrid using 0.1 M phosphate buffer pH 7.5. The free and immobilized enzymes were characterized by optimum temperature, Michaelis constant (K _M), maximum velocity (V _max), thermal inactivation rate constant (k _i ), half-life (t _1/2), and transformation of free energy because of denaturation (ΔG _i ). The free enzyme has optimum temperature of 55°C, K _M  = 3.04 mg mL^-1 substrate, V _max=10.90 μmolemL^-1min^-1, k _i  = 0.0171 min^-1, t _1/2 = 40.53 min, and ΔG _i  = 104.47 kJ mole^-1. Meanwhile, the immobilized enzyme has optimum temperature of 60°C, K _M  = 11.57 mg mL^-1 substrate, V _max=3.37 μmolemL^-1min^-1, k _i  = 0.0045 min^-1, t _1/2 = 154.00 min, and ΔG _i  = 108.17 kJ mole^-1. After sixth cycle of reuse, the residual activity of the immobilized enzyme was 38%. The improvement in the stability of α-amylase immobilized on the CB hybrid based on the increase in half-life was four times of the free enzyme.