IDR - IIT Kharagpur

Production, Purification, Characterization of α-Amylase and Its Application With Other Carbohydratases as Food Additives

Production, Purification, Characterization of α-Amylase and Its Application With Other Carbohydratases as Food Additives


GROWTEK bioreactor and a newly configured bioreactor (NB) were utilized to circumvent many problems associated with conventional metallic tray reactors for solid-state fermentation (SSF). Among these bioreactor systems, NB showed most efficiency and produced highest α-amylase activity of 22,316.67 U/gds from Aspergillus oryzae IFO 30103 with the 1% glucose supplementation within a very short incubation period (48 h) at 2 cm bed height with air flow rate of 0.1 L min-1 g-1 wheat bran. Polygalacturonase was produced by Aspergillus awamori Nakazawa MTCC 6652 utilizing mosambi orange (Citrus sinensis var mosambi) peels. α-Amylase and polygalacturonase were decolourized and partially purified by activated charcoal in a single step. In addition, for biochemical characterization, α-amylase was highly purified through Sephadex G-100 and DEAE-cellulose columns sequentially. Enzyme kinetics of purified α-amylase revealed Km and Vmax values of 0.5% and 1000 U/mg protein for soluble starch. Total phenolic content and antioxidant property of wheat (54% ethanolic extract) enhanced drastically when fermented with A. oryzae and A. awamori. It was proved that α-amylase along with other two carbohydratases (xylanase and β-glucosidase) of A. oryzae was mainly responsible for the release of higher amount of phenolics compared to A. awamori, where only xylanase and β-glucosidase were involved during fermentation process. Purified α-amylase was also utilized for the extraction of phenolics from wheat. Performance of NB for the improvement of antioxidant activity in rice koji was studied and the results were compared to the conventional SSF conducted in Erlenmeyer flask. Interestingly, rice fermented by A. oryzae in NB resulted in higher yield of phenolics and antioxidant potential in each of the extraction media (water, methanol and ethyl acetate) as compared to SSF and non-fermented rice. This was due to the higher titre values of β-glucosidase (62.7%) and α-amylase (40.7%) in the NB compared to SSF. The decolourized and partially purified α-amylase along with polygalacturonase was utilized for apple juice clarification. In presence of 1% polygalacturonase (9.87 U/ml) and 0.4% α-amylase (899 U/ml), maximum clarity (%T660nm = 97.0%) of juice was attained after 2 h of incubation at 50 oC in presence of 10 mM CaCl2. Keywords α-Amylase; GROWTEK; SSF; NB; Polygalacturonase; Activated charcoal; Carbohydratases; Phenolic; Antioxidant; Rice; Wheat; Apple juice.

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