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United States Department of Agriculture

Agricultural Research Service

Title: Substrate Specificity of Membrane-Bound Oat Seed Peroxygenase: Analyses of Lipid Epoxides by Liquid Chromatography Coupled with Electron Impact and Atmospheric Pressure Ionization Mass Spectrometry

Authors
item Piazza, George
item Nunez, Alberto
item Foglia, Thomas

Submitted to: Journal of Molecular Catalysis B: Enzymatic
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: December 20, 2002
Publication Date: April 20, 2003
Citation: PIAZZA, G.J., NUNEZ, A., FOGLIA, T.A. EPOXIDATION OF FATTY ACIDS, FATTY METHYL ESTERS, AND ALKENES BY IMMOBILIZED OAT SEED PEROXYGENASE. JOURNAL MOLECULAR CATALYSIS B: ENZYMATIC. 2003. v. 21. p. 143-151.

Interpretive Summary: The chemical introduction of oxygen into fats and oils can produce a class of compounds called epoxides, which are valuable materials used in the manufacture of plastics. Epoxides from vegetable oils, particularly soybean oil, are widely used to stabilize plastics and make them more flexible. The present procedure for making epoxides uses acetic acid and hydrogen peroxide in the presence of a strong acid catalyst, which results in the formation of a chemically unstable intermediate that is potentially explosive. In addition, exposure of the epoxides to the acid catalyst for a prolonged period causes the epoxides to degrade, and the direct discharge of the acid catalyst into the environment must be avoided. Improved catalysts and safer procedures are therefore needed for manufacture of epoxides. Prior research has shown that an enzyme termed peroxygenase can promote the formation of epoxides in the presence of a mild oxidizing agent. In the current study, reactions using peroxygenase were investigated. Conditions were identified for achieving high yields of epoxides. Those fat and oil fractions preferred for epoxide formation were determined. This research will contribute to the development of safer, more environmentally friendly methods for making epoxides from renewable agricultural materials.

Technical Abstract: The ability of immobilized oat (Avena sativa) seed peroxygenase to catalyze the epoxidation of a variety of unsaturated fatty acids, fatty methyl esters, and hydrocarbons was tested using t-butyl hydroperoxide as the oxygen donor. Fatty epoxide formation was followed by high performance liquid chromatography with evaporative light-scattering detection, and formation of the hydrocarbon epoxides was measured by gas-liquid chromatography-mass spectrometry. Fatty epoxides were further characterized using high performance liquid chromatography with electron impact (HPLC-EI-MS) and atmospheric pressure chemical ionization mass spectrometry (HPLC-APCI-MS). Results showed that peroxygenase exhibited good specificity for epoxidation, producing negligible amounts of byproducts. In particular, allylic oxidation was not evident, which distinguishes peroxygenase-catalyzed oxidations from those catalyzed by chloroperoxidase. The absence of di- and polyhydroxylated materials indicated that epoxide hydrolase was not present. Peroxygenase showed substantial discrimination against trans double bonds, and terminal unsubstituted aliphatic double bonds were not reactive. Chiral analysis of the epoxide derived from oleic acid showed that 9(R),10(S)- epoxyoctadecanoic acid was the predominant stereoisomer.

Last Modified: 10/24/2014
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