Protease production using Bacillus licheniformis by submerged fermentation

Document Type : Research Paper



2 Pilot of Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran.


     Enzymes, such as protease, occupy a pivotal position in the world of enzymes with respect to their applications in both physiological and commercial applications. Bacillus, like Bacillus licheniformis, produce protease and was used throughout the present study. The Bacillus licheniformis is a potential microorganism for producing proteins with various functions, including protease with substantial activity, effective pH, and temperature tolerance, at laboratory and industrial scales. This study investigated the effect of several temperatures ranging from 25-50 οC, pH ranging from 7-10.5, shaker speeds of 100-200, and inoculum concentrations of 3-12 %V/V on enzyme production. Results showed that cultivating the medium at about pH (8.5), a temperature of 35 οC, and a shaker gyratory speed of 180 rpm influenced protease production. Firstly, it was found that using yeast extract, soybean meal corn, and steep liquor (powder) as nitrogen sources also enhanced protease production. Furthermore, cornstarch and glucose of 3 %W/V were found to be effective as a carbon source for protease production. The protease stability was retained for almost 35 minutes, heating at 60 οC and pH 8.5, and as heating continued at the same temperature and pH, the enzyme had 40% of its activity. Secondly, the effect of different metal ions types on the stability of the partially purified protease is reported. It was found that the protease activity at 60 ± 0.5 οC, pH 9.5, and the presence of 15 mM CaCl2 and 10 mM NaCl during an hour of treatment retained approximately 40 percent of its activity. Since a batch period of 24 protease production showed similar proteases activity as a  48 h operation,  the following fermentation was conducted within 24 h. The partially purified protease had sound stability in the alkaline pH of 9-9.5 and temperature range of 60 ±0.5 οC in the presence of certain ions like calcium and sodium salt.


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