The Stability Improvement of α-Amylase Enzyme from Aspergillus fumigatus by Immobilization on a Bentonite Matrix

Abstract

The stability of the α-amylase enzyme has been improved from Aspergillus fumigatus using the immobilization method on a bentonite matrix. Therefore, this study aims to obtain the higher stability of α-amylase enzyme from A. fumigatus; hence, it is used repeatedly to reduce industrial costs. The procedures involved enzyme production, isolation, partial purification, immobilization, and characterization. Furthermore, the soluble enzyme was immobilized using 0.1 M phosphate buffer of pH 7.5 on a bentonite matrix, after which it was characterized with the following parameters such as optimum temperature, Michaelis constant (KM), maximum velocity V max , thermal inactivation rate constant (ki), half-life (t1/2), and the change of energy due to denaturation (ΔGi). The results showed that the soluble enzyme has an optimum temperature of 55°C, KM of 3.04 mg mL−1 substrate, V max of 10.90 μmole mL−1 min−1, ki of 0.0171 min−1, t1/2 of 40.53 min, and ΔGi of 104.47 kJ mole−1, while the immobilized enzyme has an optimum temperature of 70°C, KM of 8.31 mg mL−1 substrate, V max of 1.44 μmole mL−1 min−1, ki of 0.0060 min−1, t1/2 of 115.50 min, and ΔGi of 107.37 kJ mole−1. Considering the results, the immobilized enzyme retained 42% of its residual activity after six reuse cycles. Additionally, the stability improvement of the α-amylase enzyme by immobilization on a bentonite matrix, based on the increase in half-life, was three times greater than the soluble enzyme.

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

“…The phases of production, isolation, partial purification, and characterization of the soluble and immobilized enzymes were based on a previous study [ 16 ]. The crude enzyme was partially purified by fractionation using ammonium sulphate and dialysis [ 17 ].…”

“…The crude enzyme was brought to 0–20% saturation with ammonium sulphate. The precipitate was removed by centrifugation at 5,000 rpm for 15–20 min, and then, ammonium sulphate was added to the supernatant to 20–85% saturation [ 16 , 17 ]. The precipitated protein was collected by centrifugation at 5000 rpm for 15 min at 4°C and dissolved in a minimum volume of phosphate buffer (0.025 M; pH 6.5).…”

“…The precipitated protein was collected by centrifugation at 5000 rpm for 15 min at 4°C and dissolved in a minimum volume of phosphate buffer (0.025 M; pH 6.5). Then, the enzyme suspension was dialyzed in a dialysis bag (8000 Da) for 24 h at 4°C against phosphate buffer (0.01 M; pH 6.5) [ 16 , 17 ]. Meanwhile, the immobilization method was based on Yandri et al [ 18 ].…”

“…en, the enzyme suspension was dialyzed in a dialysis bag (8000 Da) for 24 h at 4 °C against phosphate buffer (0.01 M; pH 6.5) [16,17]. Meanwhile, the immobilization method was based on Yandri et al [18].…”

Immobilization and Stabilization of Aspergillus Fumigatus α-Amylase by Adsorption on a Chitin

“…According to a previous research, the α-glucosidase enzyme from Thermatoga maritime which immobilized on a chitin can be used repeatedly up to 10 times and retained 66% of its initial activity [21]. Likewise, the A. fumigatus α-amylase immobilized on a chitin/bentonite hybrid matrix has greater thermal stability about 3.8 times than the native form, and maintained 72% of its initial activity after incubating at 60 o C for 80 min [22].…”