تولید آمیلاز از B. amyloliqufaciens در تخمیر مستغرق با استفاده از فرآورده های فرعی صنعتی کشاورزی Production of amylases from Bacillus amyloliquefaciens under submerged fermentation using some agro-industrial by-products

Enzymes are defined as biocatalysts protein in nature, produced by living cells to bring about specific biochemical reactions, generally forming parts of the metabolic processes of the cell. Enzymes are highly specific in their action on substrates. The production of amylases is overshadowing all other enzymes; hence, amylases account 65% of enzyme market in world (van der Maarel et al., 2002; Balkan and Figen, 2007). a-amylase is an endo-acting enzyme and hydrolyzes linkages in a random fashion that hydrolyzes a-1,4 bonds and bypass a-1, 6 linkages and leads to the formation of linear and branched oligosaccharides and limit dextrins. b-amylase is an exo-acting enzyme that attacks the substrate from the nonreducing end and hydrolyzes a-1, 4 and cannot bypass a-1, 6 linkages thus producing oligosaccharide maltose (2 units of adjacent glucose) as a major end product. c-Amylase (glucoamylase) is an exo-acting enzyme that attacks the substrate from the non-reducing end and hydrolyzes a-1, 4 and a-1, 6 linkages thus producing monosaccharides (1 unit of glucose) as a major end product (Gupta et al., 2002). Amylases are used commercially for starch liquefaction, paper, desizing of textile fabrics, in preparing starch coatings of paints, in removing wall paper, food in the brewing industry, sugar induction by production of sugar syrups from starch which consist of glucose, maltose and higher oligosaccharides, pharmaceutical and in preparing cold water dispersible laundry starches. To meet the demands of these industries low cost medium is required for the production of amylases (Balkan and Figen, 2007). Nowadays the potential of using microorganisms as a biological source of industrially economic enzymes has stimulated interest in the exploitation of extracellular enzymatic activity in several microorganisms. Amylases can be obtained from several sources such as plant, animal and microbes such as bacteria and fungi (Murakami et al., 2008). The microbial source of amylases is preferred to other sources because of its plasticity and vast availability. Until now all commercial enzymes have been derived from cultivated bacteria or fungi. Bacterial amylases are generally preferred for starch processing. Among bacteria, Bacillus species such as B. subtilis, B. stearothermophilus, B. macerans, B. megaterium and B. amyloliquefaciens were the best producers of thermostable a-amylase using submerged fermentation and these have been widely used for commercial production of the enzyme for various applications (Enhasy, 2007; Bozˇic´ et al., 2011), Clostridium thermosacharolyticum, Cl. thermohydrosulfuricum and Pseudomonas sp. (Mrudula et al., 2011). Biosynthesis of amylases was performed on agro-industrial wastes and by-products such as starchy materials to solve pollution problems and obtain a low cost medium (Haq et al., 2005; Djekrif-Dakhmouche et al., 2006; Anto et al., 2006; Mukherjee et al., 2009). Rice husk, wheat bran and potato starchy waste were used as a low cost carbon substrate for amylase activity by B. subtilis (Baysal et al., 2003; Shukla and Kar, 2006; Asgher et al., 2007). The objective of this study was to investigate the agroindustrial residues as an alternative carbon source to produce amylases by Egyptian local bacteria in order to reduce environmental pollution and product cost. Materials and methods Samples collection from soil environment Rhizosphere samples were collected from the fertile fields planted with Egyptian clover (Trifolium alexandrinum), in Qalyubia governorate. Soil samples were taken from 3 to 5 cm depth after removing 5 cm from the ground surface. These samples were collected into sterilized plastic bags and stored in ice-boxes during their transport to the laboratory. In the laboratory all samples were kept refrigerated until isolation.