Isolation and identification of α-amylase producing soil microorganisms and partial characterization of α-Amylase enzyme

Abstract

Alpha (α)-amylase is an enzyme responsible for the breakdown of starch molecules. It hydrolyzes starch into reducing and fermentable sugars like maltose. The enzyme has various industrial applications; namely in food, detergent, leather, paper, chemical, and pharmaceutical industries. In this study, α-amylase producing bacteria were isolated from two soil samples taken from two different places of Dhanmondi, Dhaka. Out of 23 isolates from the soil samples, 10 could hydrolyze starch and were selected for identification and characterization. The selected isolates were gram-positive Bacillus and Solibacillus; and gram-negative Raoultella and Klebsiella. Antibiotic susceptibility of the 10 isolates was determined by the Kirby-Bauer test. Gram-negative isolates were more resistant to antibiotics than the gram-positive isolates. Among the selected isolates, Bacillus cibi showed the highest antimicrobial activity against pathogenic strains. The effects of temperature, pH and salinity on the growth of isolates were determined. The optimum growth temperature for the isolates ranged from 370C to 400C, and optimal pH for growth ranged from 7.0 to 7.5. The isolates were subjected to submerged fermentation for the production of α-amylase and the activities of extracted amylase enzymes were determined by the DNS test. Maximum amylase activity were displayed by Bacillus subtilis (0.49 unit/ml), Bacillus amyloliquefaciens (0.46 unit/ml), and Bacillus sporothermodurans (0.44 unit/ml). The isolates were further characterized by studying the effects of temperature, pH, and presence of various chemical agents and metallic salts on the activity of bacterial amylase. The enzymes showed maximum activity at temperature 370C~400C, and pH 6.0~6.7. Enzyme activity was increased in the presence of calcium. However, it was decreased in the presence of chemical agents like urea, EDTA and acetic acid.