Properties and application of poly(methacrylic acid‐ co ‐dodecyl methacrylate‐ cl ‐ N,N ‐methylene bisacrylamide) hydrogel immobilized Bacillus cereus MTCC 8372 lipase for the synthesis of geranyl acetate

A range of fatty acid esters is now being produced commercially with immobilized microbial lipases (glycerol ester hydrolases; EC) in nonaqueous solvents. In this study, a synthetic hydrogel was prepared by the copolymerization of methacrylic acid and dodecyl methacrylate in the presence of a crosslinker, N,N ‐methylene bisacrylamide. A purified alkaline thermotolerant bacterial lipase from Bacillus cereus MTCC 8372 was immobilized on a poly(methacrylic acid‐ co ‐dodecyl methacrylate‐ cl ‐ N,N ‐methylene bisacrylamide) hydrogel by an adsorption method. The hydrogel showed a 95% binding efficiency for the lipase. The bound lipase was evaluated for its hydrolytic potential toward various p ‐nitrophenyl acyl esters with various C chain lengths. The bound lipase showed optimal hydrolytic activity toward p ‐nitrophenyl palmitate at a pH of 8.5 and a temperature of 55°C. The hydrolytic activity of the hydrogel‐bound lipase was enhanced by Hg 2+ , Fe 3+ , and NH   4 +ions at a concentration of 1 m M . The hydrogel‐bound lipase was used to synthesize geranyl acetate from geraniol and acetic acid in n ‐heptane. The optimization of the reaction conditions, such as catalyst loading, effect of substrate concentration, solvent ( n ‐pentane, n ‐hexane, n ‐heptane, n ‐octane, and n ‐nonane), reaction time, temperature, molecular sieve (3 Å × 1.5 mm) and scale up (at 50‐mL level), was studied. The immobilized lipase (25 mg/mL) was used to perform an esterification in n ‐alkane(s) that resulted in the synthesis of approximately 82.8 m M geranyl acetate at 55°C in n ‐heptane under continuous shaking (160 rpm) after 15 h when geraniol and acetic acid were used in a ratio of 100 : 100 m M. The addition of a molecular sieve (3 Å × 1.5 mm) to the reaction system at a concentration of 40 mg/mL in reaction volume (2 mL) resulted in an increase in the conversion of reactants into geranyl acetate (90.0 m M ). During the repetitive esterification under optimum conditions, the hydrogel‐bound lipase produced ester (37.0 m M ) after the eighth cycle of reuse. When the reaction volume was scaled up to 50 mL, the ester synthesized was 58.7 m M under optimized conditions. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008