Isolation, mutation of bacillus sp. And production of nattokinase: a fibrinolytic enzyme

Abstract

Present study was undertaken to isolate the potential microorganisms and to modify the isolate for Nattokinase production, using batch wise submerged fermentation process. For screening, five different locations in and around Kanpur had been selected and after exhaustive screening, isolates were mutated and identified by morphological and biochemical examination. Thirty colonies from primary screening were selected for further investigation and PM2 found best performer with 11mm zone of clearance during secondary screening. Hence PM2 selected for further modification by mutation. After three successive mutation of PM2 under UV radiation, PM2713 found most efficient among all the mutated isolates. Various combinations of carbon and nitrogen sources examined for production of Nattokinase in suspension culture. Best combination with lactose and peptone giving the yield of 48 units/ml. Further experiments were designed to optimize production process of Nattokinase using RSM taking five parameters namely lactose concentration, peptone concentration, pH, temperature and fermentation time. Analysis of the results obtained from 32 experiments reveled the maximum yield 49.01 unit/ml of Nattokinase under the condition of lactose concentration 2.04 %, peptone concentration 3.45 %, pH 6.94, temperature 32.92oC and fermentation time 42.06 hours. A wet experiment performed taking the optimized parameters for production of Nattokinase which verify the result obtained from RSM.