Optimization of Culture Conditions for Protease Production using Three Strains of Bacillus

The aim of this work was to determine the effect of a few external factors on bacterial growth and the production of enzymes with a proteolytic effect in three strains of Bacillus: CMS5 (Bacillus subtilis), CMS4 (Bacillus sp.) and SPo5 14′ (Bacillus velenzensis) isolated from squashes packed in traditionally prepared cassava leaves, but also to determine the best source of carbon and nitrogen. All three strains have the ability to actively degrade milk casein. The strains were grown in Luria Bertani medium and the suspension from the cell culture was used to measure optical density and demonstrate enzyme activity on a petri dish containing skim milk. Several parameters were verified including the influence of temperature, pH, and carbon and nitrogen source on growth and enzyme production. Growth was possible from 25 to 60°C with an optimal temperature of 30°C after 24 hours. Enzyme production was observed from 25 to 55°C with an optimum at 37°C. For pH, growth and enzyme production was possible from pH 5.7 and 9 with an optimum of 7 in all three strains. Among the sources of carbons used, galactose is the best source for growth after 24 h in all three strains, and starch for production. Among nitrogen sources, Bacto-peptone is best for growth as well as production

Production, Partial Purification and Based SDS-PAGE Profiles of Caseinolytic Enzyme in two Bacillus Strains Isolated from Fermented Cassava leaves "Ntoba mbodi" in Congo Brazzaville

Two Bacillus strains isolated from Ntoba mbodi : Bacillus megaterium (B.me NM 02) Bacillus licheniformis (B.li NM01), has shown a significant caseinolytic enzyme activity. We set optimization of growth and enzyme production conditions. Several parameters have been optimized: temperature, pH, various types of media, carbon and nitrogen sources. In both strains, growth is possible from 25 to 60 ° C with an optimum temperature at 37°C for B.li and at 35°C for B.me. Enzyme production was observed from 25 to 55 °C with an optimum temperature of 30 °C for B.li. Enzyme production was observed from 25 to 50 ° C with an optimum temperature of 35 ° C in B.me. For pH, growth and enzyme production can be at 5, 7 and 9 with an optimum at 7 for both strains. The LB medium is better for growth and enzyme production than TSB for B.li and B.me. Among the carbon sources used, fructose is better for growth after 48 hours of incubation in both strains (B.li: 0,93±0,001, B.me: 0,928±0,002), but for enzyme production fructose remains the best carbon source for B.li (14,33±1,24) , while starch is the best for B.me (14.66±1,24). Concerning nitrogen sources, in both strains the best source of growth is the yeast extracts (B.li: 0,969±0,015, B.me: 0,952±0,01). For enzyme production, the two sources can be used for B.li.(14,333±1,247) but for B .me (14,333±0,471) only the yeast extracts is the best as well as for growth and enzyme production. Furthermore, in both strains the produced enzyme was partially purified by using ammonium sulfate precipitation, and SDS-PAGE has been hold, profiles of specific bands are useful to give more information and differentiate the two strains