Production, purification, characterization of selected microbial lipases and their application for interesterification of butter fat

• Main Author: Pabai, Franknel Sandi Kouvea.
• Other Authors: Kermasha, S. (advisor)
• Format: Others
• Language: en
• Published: McGill University 1997
• Subjects: Milkfat.; Esterification.; Microbial enzymes.
• Online Access: http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=42112

The screening, biomass production of lipase-producing microorganisms from several sources, as well as the purification, characterization and utilization of the enzymatic extracts for the interesterification of butter fat were investigated. Pseudomonas fragi CRDA 323 and Aspergillus niger CBS 131.52 were considered to be good lipase producers, whereas, those from Pseudomonas putida ATCC 795 and Rhizopus oryzae ATCC 34612 as weak ones; all four microorganisms produced maximal amount of extracellular lipases by batch fermentation after three-four days of incubation in a continuously stirred tank reactor. The lipases were partially purified by ammonium sulfate precipitation and characterized with respect to pH, kinetic parameters and molecular size. The lipases from P. fragi and P. putida were optimal at pH 8.5 and 8.0, respectively, whereas those from A. niger and R. oryzae were optimal at pH 7.5. The A. niger lipase had the lowest V$ sb{max}$ value $ rm(0.51 times 10 sp3 U min sp{-1});$ R. oryzae the highest $ rm (1.86 times 10 sp3 U min sp{-1}).$ The K$ sb{m}$ values for P. fragi, P. putida, A. niger and R. oryzae lipases were 0.70, 1.18, 0.97 and 0.98 mg ml$ sp{-1},$ respectively. Interesterification of butter fat by the partially purified enzymatic extracts in a microemulsion free co-surfactant system containing sorbitol monostearate and polyoxyethylene sorbitan monostearate in the ratio 48:52 (V/V) decreased the water activity as well as the hydrolytic activity. The P. fragi lipase had the highest interesterification yield value (43%) and the R. oryzae lipase the lowest (4%). In addition, P. fragi lipase exhibited the highest decrease (18%) in long-chain hypercholesterolemic fatty acids (C12:0, C14:0 and C16:0) at the sn-2-position; the P. putida lipase demonstrated the least favorable changes in specificity at the same position. Continuous cultivation technique was developed to investigate the screening for lipase-producing microorganisms from four commercial