SHARMA, NIVEDITASHARMA, RANJANA2019-08-052019-08-052019-04http://krishikosh.egranth.ac.in/handle/1/5810119266ABSTRACT Enzymes are considered as nature’s catalysts. are a class of enzymes which catalyze the hydrolysis of long chain triglycerides into polar lipids. Soil contaminated with oil from different sites of Himachal Pradeshbeing a highly probablesource for lipolytic microorganisms was utilized for isolation of lipase producing microorganisms. In total, 53 lipolytic bacteria have been screened on tributyrin agar medium. Among them hyper lipolytic strains C6 and G7 were selected and identified asBacillus siamensis C6MF446911 and Paenibacillus sp.G7. Cultural conditions and process parameters viz. media types, pH, temperature, inoculum size, incubation time, substrate concentration, divalent ions and surfactants etc. were optimized firstly through classical onevariable at a time (OVAT)followed by statistical optimization by employing central composite design of response surface methodology. The enzymes obtained from both the strains were purified to homogeneity by following a sequential purification approach. B. siamensis C6 lipase was purified to a final purification fold of 1.91 and had a molecular weight of 32kDA whereas P. sp.G7 lipase was purified to a final purification fold of 2.01 and had a molecular weight of 66 kDa. Lipase activity was found to be maximum at 40oC and pH 8.0 for B. siamensis C6 and at 35 oC and pH 8.0 for P.sp.G7. Lipase from both the strains was quite thermostable with retention of more than 50% activity after incubation of 120 min at 30-45 oC. Shelf life of lipase from both the strains showed that the enzyme was very efficient qualitatively as well as quantitatively. Press cake of Brassica nigra showed maximum stability for both isolates i.e. 1213.06 IU/mg specific activity for B. siamensis C6 lipase and 1236.84 IU/mg specific activity for P. sp.G7 lipase. Chitosan beads and κ-carrageenan chips at a concentration of 1.5% and 4 % respectively, were found to be most efficient for immobilization of purified lipase of P. sp.G7. Purified P. sp.G7 lipase showed a considerably good stability in κ-carrageenan chips followed by immobilized chitosan beads (1.5%) giving an outstanding immobilization efficiency of 80.99 and 76.96% at the end of 20thcycle. Applicability of purified lipase from P. sp.G7 (20 ppm) in order to enhance product quality and earn a natural and clean label status to bread which is highly sought after by health conscious consumers of modern age. In addition pseudocereal grains. Therefore incorporation of these grains in bread dough formulation enhances significantly the nutritional quality of baked product.ennullThesis