Introduction Enzymes involved in starch conversion technology are of major importance and considerable interest exists in obtaining new enzymes having improved properties or new applications 1-3 . Enzymatic processes now produce over 75% of syrup and solid dextrose in the USA. New developments have taken place in the area of starch-degrading enzymes. Enzymes have several advantages. First, the specificity of enzymes allows the production of sugar syrups with well-defined physical and chemical properties. Second, the milder enzymatic hydrolysis results in few side reactions and less browning 4 . α-Amylases are group of enzymes classified as hydrolases that catalyze the hydrolysis of ortho-glycosyl compounds of starch and glycogen. They catalyze the hydrolysis of starchy materials into smaller glucose subunits that in turn are acted upon by other amylases to produce glucose 5 . Amylases, starch-degrading enzymes, have numerous biotechnical applications. These enzymes are used in textile and garments, paper industries, starch liquefaction, food, adhesive and sugar production and pharmaceuticals 6 . Carbohydrates in the form of sugar and starch represent a major part of total caloric intake for humans and for most animal life as well as for many microorganisms. The vast amount of starch and other carbohydrates made by photosynthesis become the ultimate energy and carbon sources for non-photosynthetic cells of the animal, plant and microbial worlds 7 . Most agricultural biomass containing starch can be used as a potential substrate for the production of gaseous or liquid fuels, feed proteins and chemicals by microbial processes. These substrates include corn (maize), wheat, oats, rice, potato, cassava etc. α-Amylases are produced by fermentation of such starchy substrates. There are several sources of α-amylases, which include bacteria, fungi, animals and high starch containing plants. However, industrial α-amylases is produced through fermentation using bacteria and fungi 4 . Because of the commercial and industrial uses, α-amylases from many sources has been studied in great detail. The genus Bacillus is the single most important bacterial source of this enzyme. Due to the thermostability of the enzyme produced by genus Bacillus, they have commercial significance 4 . This paper reports on the α-amylase from Bacillus subtilis, Bacillus amyloliquefaciens and a newly isolated Bacillus sp. under various conditions. Materials and Methods Microorganisms Bacillus subtilis and Bacillus amyloliquefaciens were obtained from Department of Microbiology, University of Dhaka, Bangladesh. In addition, a soil organism identified as the member of Bacillus was also included. The organisms were maintained on nutrient agar (NA) medium. Plate assay method The Bacillus isolates were tested for amylase activity by employing zone clearing technique 8-9 using starch agar medium. The inoculated plates were incubated at 37°C for three days. After incubation, the zone of hydrolysis of starch was detected by flooding the plates with iodine solution. The development of blue Comparative Studies on the Production of Extracellular α-Amylase by Three Mesophilic Bacillus Isolates Arifa Nusrat and Sabita Rezwanan Rahman * Department of Microbiology, University of Dhaka, Dhaka 1000, Bangladesh [Received 22 November 2007; Accepted 08 December 2007] In the present study three mesophilic Bacillus isolates were analyzed for their α-amylase activity in shake- flask cultures. The organisms were capable to produce hydrolysis zone around their colonies on starch agar medium. The effect of various fermentation conditions on α-amylase production was investigated, and in every case it was found that B. subtilis was the best producer of the enzyme, which was followed by the newly isolated Bacillus sp. and B. amyloliquefaciens. The synthesis of extracellular α-amylase by the bacteria was repressed by the presence of readily metabolizable carbon source like glucose in the culture medium. Maximum α-amylase activity by the Bacillus isolates was obtained at 37°C with an initial medium pH 7.0 under agitation at 160-180 rpm for 72 h of growth. Keywords: Bacillus species, α-Amylase, Enzyme production, Shake-flask culture Original Article Bangladesh J Microbiol, Volume 24, Number 2, December 2007, pp 129-132 * Corresponding author: Dr. Sabita Rezwana Rahman, Associate Professor, Department of Microbiology, University of Dhaka, Dhaka 1000, Bangladesh Tel (Office): (02) 9661920-73, Ext 7746; Tel (Home): (02) 9351014; Fax: +880 (02) 8615583; E-mail: sabita_rahman@hotmail.com 129