- Lipoxygenase-catalyzed transformation of epoxy fatty acids to hydroxy-endoperoxides: A potential P450 and lipoxygenase interaction
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Herein, we characterize a generally applicable transformation of fatty acid epoxides by lipoxygenase (LOX) enzymes that results in the formation of a five-membered endoperoxide ring in the end product. We demonstrated this transformation using soybean LOX-1 in the metabolism of 15,16-epoxy-α-linolenic acid, and murine platelet-type 12-LOX and human 15-LOX-1 in the metabolism of 14,15-epoxyeicosatrienoic acid (14,15-EET). A detailed examination of the transformation of the two enantiomers of 15,16-epoxy-α-linolenic acid by soybean LOX-1 revealed that the expected primary product, a 13 S-hydroperoxy-15,16-epoxide, underwent a nonenzymatic transformation in buffer into a new derivative that was purifi ed by HPLC and identified by UV, LC-MS, and 1H-NMR as a 13,15-endoperoxy-16-hydroxy-octadeca-9,11-dienoic acid. The configuration of the endoperoxide (cis or trans side chains) depended on the steric relationship of the new hydroperoxy moiety to the enantiomeric configuration of the fatty acid epoxide. The reaction mechanism involves intramolecular nucleophilic substitution (SNi) between the hydroperoxy (nucleophile) and epoxy group (electrophile). Equivalent transformations were documented in metabolism of the enantiomers of 14,15-EET by the two mammalian LOX enzymes, 15-LOX-1 and platelet-type 12-LOX. We conclude that this type of transformation could occur naturally with the co-occurrence of LOX and cytochrome P450 or peroxygenase enzymes, and it could also contribute to the complexity of products formed in the autoxidation reactions of polyunsaturated fatty acids.
- Teder, Tarvi,Boeglin, William E.,Brash, Alan R.
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- Analysis of epoxyeicosatrienoic acids by chiral liquid chromatography/ electron capture atmospheric pressure chemical ionization mass spectrometry using [13C]-analog internal standards
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The metabolism of arachidonic acid (AA) to epoxyeicosatrienoic acids (EETs) is thought to be mediated primarily by the cytochromes P450 (P450s) from the 2 family (2C9, 2C19, 2D6, and 2J2). In contrast, P450s of the 4 family are primarily involved in omega oxidation of AA (4A11 and 4A22). The ability to determine enantioselective formation of the regioisomeric EETs is important in order to establish their potential biological activities and to asses which P450 isoforms are involved in their formation. It has been extremely difficult to analyze individual EET enantiomers in biological fluids because they are present in only trace amounts and they are extremely difficult to separate from each other. In addition, the deuterium-labeled internal standards that are commonly used for stable isotope dilution liquid chromatography/mass spectrometry (LC/MS) analyses have different LC retention times when compared with the corresponding protium forms. Therefore, quantification by LC/MS-based methodology can be compromised by differential suppression of ionization of the closely eluting isomers. We report the preparation of [13C20]-EET analog internal standards and the use of a validated high-sensitivity chiral LC/electron capture atmospheric pressure chemical ionization (ECAPCI)-MS method for the trace analysis of endogenous EETs as their pentafluorobenzyl (PFB) ester derivatives. The assay was then used to show the exquisite enantioselectivity of P4502C19-, P4502D6-, P4501A1-, and P4501B1-mediated conversion of AA into EETs and to quantify the enantioselective formation of EETs produced by AA metabolism in a mouse epithelial hepatoma (Hepa) cell line. Copyright
- Mesaros, Clementina,Lee, Seon Hwa,Blair, Ian A.
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- Enzymatic synthesis and chemical inversion provide both enantiomers of bioactive epoxydocosapentaenoic acids
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Epoxy PUFAs are endogenous cytochrome P450 (P450) metabolites of dietary PUFAs. Although these metabolites exert numerous biological effects, attempts to study their complex biology have been hampered by difficulty in obtaining the epoxides as pure regioi
- Cinelli, Maris A.,Yang, Jun,Scharmen, Amy,Woodman, Joey,Karchalla, Lalitha M.,Lee, Kin Sing Stephen
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p. 2237 - 2252
(2018/11/30)
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- Practical, enantiospecific syntheses of 14,15-EET and leukotoxin B (vernolic acid)
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Cytochrome P450BM3 and its F87V mutant were exploited for a convenient, laboratory scale (1 mmol) preparation of 14(S),15(R)-epoxyeicosatrienoic acid [14(S),15(R)-EET] from arachidonic acid and (+)-leukotoxin B [(+)-12(S),13(R)-vernolic acid] from linoleic acid, respectively. Their enantiomers were accessed via a four-step chemical inversion.
- Falck,Reddy,Haines, Donovan C.,Reddy, Komandla Malla,Krishna,Graham, Sandra,Murry, Barbara,Peterson, Julian A.
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p. 4131 - 4133
(2007/10/03)
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- ASYMMETRIC TOTAL SYNTHESIS OF 14(R),15(S)-, 14(S),15(R)-, 14(R),15(R)-, AND 14(S),15(S)-EPOXYEICOSATRIENOIC ACIDS
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The four chiral stereoisomers of 14,15-oxido-5Z,8Z,11Z-eicosatrienoic acid were synthesized.The absolute stereochemistry in each case was derived from an asymmetric epoxidation of E-2-octen-1-ol.
- Ennis, Michael D.,Baze, Mark E.
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p. 6031 - 6034
(2007/10/02)
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