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Enzyme Research
Volume 2012, Article ID 905804, 13 pages
Research Article

Statistical Approach for Optimization of Physiochemical Requirements on Alkaline Protease Production from Bacillus licheniformis NCIM 2042

Department of Biotechnology, National Institute of Technology, Mahatma Gandhi Avenue, Durgapur 713209, India

Received 26 April 2011; Revised 6 September 2011; Accepted 21 September 2011

Academic Editor: Alane Beatriz Vermelho

Copyright © 2012 Biswanath Bhunia and Apurba Dey. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


The optimization of physiochemical parameters for alkaline protease production using Bacillus licheniformis NCIM 2042 were carried out by Plackett-Burman design and response surface methodology (RSM). The model was validated experimentally and the maximum protease production was found 315.28 U using optimum culture conditions. The protease was purified using ammonium sulphate (60%) precipitation technique. The HPLC analysis of dialyzed sample showed that the retention time is 1.84 min with 73.5% purity. This enzyme retained more than 92% of its initial activity after preincubation for 30 min at 37C in the presence of 25% v/v DMSO, methanol, ethanol, ACN, 2-propanol, benzene, toluene, and hexane. In addition, partially purified enzyme showed remarkable stability for 60 min at room temperature, in the presence of anionic detergent (Tween-80 and Triton X-100), surfactant (SDS), bleaching agent (sodium perborate and hydrogen peroxide), and anti-redeposition agents (Na2CMC, Na2CO3). Purified enzyme containing 10% w/v PEG 4000 showed better thermal, surfactant, and local detergent stability.