Optimization of protease and lipase production by Bacillus pumilus SG 2 isolated from an industrial effluent

摘要

This investigation was aimed at isolating a potent bacterial strain which can produce both protease and lipase. Effluents of meat processing industry, dairy industry, food processing industry and oil industry were collected. Among the four positive strains screened, a bacterial strain identified as Bacillus pumilus SG2 isolated from food processing industry effluent was found to be a comparatively potent producer of both protease and lipase (52U/ml and 38U/ml respectively). The process parameters for the optimal production of both enzymes using this strain were studied. The optimum pH for protease production was 8.0 and that of lipase was 9.0. The optimum temperature was 37oC for production of both enzymes. The optimum incubation time was found to be 36 hours for protease and 63 hours for lipase. The best carbon source for enzyme production was glucose and the best nitrogen sources were yeast extract and casein. Addition of additives like SDS, Triton X-100 and Tween-20 influenced enzyme production. Whey was the best crude substrate that could be used for protease production and castor oil, gingelly oil and olive oil were the best substrates for lipase production. This study has indicated the possibility of producing both protease and lipase simultaneously on a common production medium by a single bacterial strain, Bacillus pumilus. Introduction Enzymes have a great industrial potential and are widely found in various sources like plants, animals and microbes. Microbes have undermined plants and animals as sources of enzymes due to their broad biochemical diversity, ease of mass culture and also due to the ease with which they can be genetically modified. Of all the industrially important enzymes, proteases and lipases are exploited maximally due to their various applications. Proteases are hydrolytic enzymes and find their utmost application in laundry detergents (Banerjee et al., 1999). They also find applications in food industry for cheese making and baking, in pharmaceutical industry in combination with antibiotics to treat ulcers and wounds (Mala Rao et al., 1998). Proteases are used effectively for dehairing in leather processing industry (Nilegoankar et al., 2006). Lipases too have considerable industrial potential and find promising applications as additives in detergent (Gerritse et al., 1998) and food additives for flavour enhancement in cheese ripening, baking etc (Kauzlauskas and Bornscheuer, 1998). Lipases are used to synthesise chiral building blocks for pharmaceuticals (Patel, 2000) and as component of personal care products (Maugard et al., 2002).Protease and lipase have many common applications and are best used in a mixture in various industries like tanneries and also in detergents. Hence organisms which can produce both these enzymes simultaneously can be best exploited. There are very few reports on the concomitant production of lipase and protease by Pseudomonas species but there are no such reports with Bacillus species. This present report has described the isolation of a Bacillus species which produces both protease and lipase and optimization of the production parameters for both the enzymes. Materials and methods Isolation and screeningEffluents from various industries like oil industry, food processing industry, dairy industry and meat processing industry were collected, serially diluted and plated on sterile nutrient agar (0.5% peptone, 0.3% yeast extract. 0.5% sodium chloride and 2% agar) plates. The isolated pure colonies were then screened for extracellular protease production using casein agar (0.5% casein in nutrient agar) and nutrient gelatin (0.5% gelatin in nutrient agar) and lipase production was screened using tween agar (0.5% tween in nutrient agar). The strains which produced both the enzymes were then analysed for their enzyme production potential. The most potent strain was considered for future study.Enzyme ProductionThe production medium for protease consisted of (w/v) 0.04% CaCl2,