Purification and partial characterization of Rhizomucor miehei lipase for ester synthesis
A commercial Rhizomucor miehei lipase was purified by ammonium sulfate precipitation. Phenyl Sepharose 6 Fast Flow hydrophobic interaction chromatography, and DEAE Sepharose Fast Flow anion-exchange chromatography. The recovery of lipase activity was 32% with a 42-fold purification. The molecular si...
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Veröffentlicht in: | Applied biochemistry and biotechnology 1996-05, Vol.59 (2), p.145-158 |
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Sprache: | eng |
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Zusammenfassung: | A commercial Rhizomucor miehei lipase was purified by ammonium sulfate precipitation. Phenyl Sepharose 6 Fast Flow hydrophobic interaction chromatography, and DEAE Sepharose Fast Flow anion-exchange chromatography. The recovery of lipase activity was 32% with a 42-fold purification. The molecular size of the purified enzyme was 31,600 Dalton and the pI 3.8. The enzyme was stable for at least 24 h within a pH range of 7.0-10.0, and 96.8% of the enzyme activity remained when kept at 30 degrees C for 24 h. Further, about 10-30% of the lipase activity was inhibited by K+, Li+, Ni+, Co2+, Zn2+, Mg2+, Sn2+, Cu2+, Ba2+, Ca2+, and Fe2+ ions and by SDS, but EDTA had no effect. Under the experimental conditions, the optimum temperature for the hydrolysis of olive oil was 50 degrees C (pH 8.0), and for the synthesis of 1-butyl oleate, 37 degrees C. It was concluded that hydrolytic activity of lipase alone is not a sufficient criterion for its synthetic potential. The optimal molar ratio of oleic acid and 1-butanol was 2:1 for 1-butyl oleate synthesis. The 1-butyl oleate yield was unaffected by purification of the enzyme after 12 h |
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ISSN: | 0273-2289 1559-0291 |
DOI: | 10.1007/BF02787816 |