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Production and purification of an alkaline lipase from Bacillus sp. for enantioselective resolution of (±)-Ketoprofen butyl ester

Saraswat, Rashmi, Bhushan, Indu, Gupta, Pankaj, Kumar, Vivek, Verma, Vijeshwar
3 Biotech 2018 v.8 no.12 pp. 491
Bacillus subtilis, Pseudozyma antarctica, agitation, biomass, calcium chloride, carboxylic ester hydrolases, enantioselectivity, ketoprofen, metal ions, molecular weight, olive oil, pH, peptones, pharmaceutical industry, response surface methodology, solvents, surfactants, temperature
The present study was conducted to purify lipase from indigenous Bacillus subtilis strain Kakrayal_1 (BSK-L) for enantioselective resolution of racemic-ketoprofen. The production of lipase (BSK-L) was optimized using Plackett–Burman and central composite design of response surface methodology (RSM). The optimized media containing olive oil (3.5%), MnSO₄ (8 mM), CaCl₂ (5 mM), peptone (20 g/l), pH (8), agitation (180 rpm) and temperature (37 °C) resulted in maximum lipase production of 7500 U/g of cell biomass. The lipase was purified using sequential method to an overall purification fold of 13% with 20% recovery, 882 U/mg specific activity and a molecular weight of 45 kDa. Optimal pH and temperature of purified lipase were found to be 8 and 37 °C, respectively. Furthermore, BSK-L displayed good stability with various organic solvents, surfactants and metal ions. Kₘ and Vₘₐₓ values of lipase were observed to be 2.2 mM and 6.67 mmoles of product formed/min/mg, respectively. The racemic ketoprofen butyl ester was hydrolyzed using lipase with 49% conversion efficiency and 69% enantiomeric excess (ee) which was superior to the commercially procured lipase (Candida antarctica lipase). Thus, this enzyme could be considered as a promising candidate for the pharmaceutical industry.