ISSN 1330-9862 original scientific paper (FTB-1685) Solid Culturing of Bacillus amyloliquefaciens for Alpha Amylase Production Dhanya Gangadharan, Swetha Sivaramakrishnan, Kesavan Madhavan Nampoothiri and Ashok Pandey* Biotechnology Division, Regional Research Laboratory, CSIR, Trivandrum 695019, India Received: November 30, 2005 Accepted: March 22, 2006 Summary Fourteen different agroresidues were screened for alpha amylase production using Ba- cillus amyloliquefaciens ATCC 23842. Among them, wheat bran (WB) and groundnut oil cake (GOC) in mass ratio of 1:1 was proved as the best substrate source. Supplementation with 0.01 M KH 2 PO 4 and 1 % soluble starch enhanced the enzyme yield considerably. Maximum enzyme recovery from the solid mass was obtained when extracted with 0.1 M acetate buffer, pH=5.0. Maximum enzyme titer expressed as units per mass of dry sub- strate obtained was 62 470 U/g after 72 hours of fermentation at 37 °C by using the above solid substrate mixture (5 g) with the initial moisture of 85 % and inoculated with Bacillus amyloliquefaciens of 2·10 9 CFU/mL. Key words: alpha amylase, Bacillus amyloliquefaciens, solid-state fermentation, agroresidues Introduction Alpha amylases (endo-1,4-a-D-glucan glucanohy- drolase, E.C. 3.2.1.1) are extracellular endo enzymes that randomly cleave the 1,4-a linkage between adjacent glu- cose units in the linear amylose chain and ultimately ge- nerate glucose, maltose and maltotriose units. Enzyma- tic hydrolysis of starch has now replaced acid hydrolysis in over 75 % of starch hydrolysing processes due to many advantages, not least its highest yields (1). Alpha amy- lase has been derived from several fungi, yeasts, bacte- ria and actinomycetes. However, enzymes from fungal and bacterial sources have dominated applications in in- dustrial sectors (2). The most abundantly used bacterial a-amylases were derived from Bacillus amyloliquefaciens, B. licheniformis and B. stearothermophilus (3). The hydro- lyzed products are widely applied in the food, paper, and textile industries (4). They are used for starch hy- drolysis in the starch liquefaction process that converts starch into fructose and glucose syrups. They are also used as a partial replacement for the expensive malt in the brewing industry, to improve flour in the baking in- dustry, and to produce modified starches for the paper industry. In addition to this, they are used to remove starch in the manufacture of textiles (desizing) and as additives to detergents for both washing machines and automated dishwashers. Industrially important enzymes have traditionally been obtained from submerged fermentation (SmF) be- cause of the ease of handling and greater control of envi- ronmental factors such as temperature and pH. However, solid-state fermentation (SSF) constitutes an interesting alternative since the metabolites so produced are con- centrated and purification procedures are less costly (2,4–6). SSF is preferred to SmF because of simple tech- nique, low capital investment, lower levels of catabolite repression and end product inhibition, low waste water output, better product recovery, and high quality pro- duction (7). On a dry basis, agricultural substrates like corn, wheat, sorghum and other cereal grains contain around 60–75 % (by mass) starch, hydrolysable to glu- 269 D. GANGADHARAN et al.: a-Amylase Production by B. amyloliquefaciens, Food Technol. Biotechnol. 44 (2) 269–274 (2006) *Corresponding author; Phone: ++91 471 25 15 279; Fax: ++91 471 24 91 712; E-mail: ashokpandey56@yahoo.co.in, pandey@csrrltrd.ren.nic.in