Production, Purification and Optimisation of Amylase by Submerged Fermentation Using Bacillus Subtilis

Abstract

Amylase is abundantly present in nature. The main source of this enzyme is the microbial origin. It is known that about two-third of the industrial enzymes (amylase, protease, cellulose, penicillinase, chitinase, etc,) are produced by the Bacillus spp. The present work comprised in the identification of amylase producing Bacillus spp and exposure of the producers to various parameters for the maximum yield of the enzyme. To isolate and identify the amylase and protease producing strain, soil samples were collected from different vegetation from the altitude at 4367.35 feet above sea level. The isolates were screened and various biochemical tests and morphological observations were done to identify the isolates. The enzymes were produced by the submerged state fermentation (SmF) from the isolates and purified by dialysis. Effects of temperature, pH, and different carbon and nitrogen sources of the medium using SmF were optimized. Among 95 isolates, 36 were identified. Among the identified isolates, Bacillus subtilis produced the maximum yield and thus, it was optimized for the amylase production. The maximum amylase production was found at 42 C temperature, in fructose as a carbon sugar, peptone as a nitrogen source and at pH 7. Almost all the enzyme producers inhabited the roots of leguminous plants. In the present study, starch is used with the nutrient agar medium to help in cell immobilization for maximum production of amylase by strains of Bacillus. More sophisticated process of purification such as chromatography and electrophoresis will yield more enzyme as compared to the dialysis.