Identification of all classes of spin-trapped carbon-centered radicals in soybean lipoxygenase-dependent lipid peroxidations of omega-6 polyunsaturated fatty acids via LC/ESR, LC/MS, and tandem MS.
Using the combined techniques of on-line high performance liquid chromatography/electron spin resonance (LC/ESR) and mass spectrometry (MS), we previously identified spin-trapped adducts of all expected carbon-centered lipid-derived radicals ((*)L(d)) formed in linoleic acid peroxidation. In the present study, spin trapped lipid-derived carbon-centered radicals formed from the reactions of two omega-6 polyunsaturated fatty acids (PUFAs: linoleic and arachidonic acids) with soybean lipoxygenase in the presence of alpha-[4-pyridyl 1-oxide]-N-tert-butyl nitrone (POBN) were identified using a combination of LC/ESR and LC/MS. All expected lipid-derived carbon-centered radicals in lipoxygenase-dependent peroxidations of linoleic acid and arachidonic acid were detected and identified by the combination of LC/ESR and LC/MS with confirmation by tandem mass spectrometry (MS/MS). The five classes of (*)L(d) formed from both omega-6 PUFAs including lipid alkyl radicals (L(*)), epoxyallyic radicals (OL(*)), dihydroxyallyic radicals ((*)L(OH)(2)), and a variety of R(*) and (*)RCOOH from beta-scission of lipid alkoxyl radicals, gave distinct retention times: POBN/(*)L(OH)(2) approximately 4-6 min, POBN/R(*) and POBN/(*)RCOOH approximately 8-22 min, POBN/L(*) and PBON/OL(*) approximately 25-36 min. The major beta-scission products in peroxidations of omega-6 PUFAs were the pentyl radicals. The ratio of beta-scission products, however, varied significantly depending on pH, [PUFA], as well as [O(2)].
Qian
Steven Yue
SY
Laboratory of Pharmacology and Chemistry, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709, USA. qian 1@niehs.nih.gov
Yue
Gui-Hua
GH
Tomer
Kenneth B
KB
Mason
Ronald P
RP
D016718:Arachidonic Acid / Q000737:chemistry / Q000378:metabolism; D002244:Carbon / Q000737:chemistry*; D002853:Chromatography, Liquid; D004578:Electron Spin Resonance Spectroscopy; D043371:Fatty Acids, Omega-6 / Q000378:metabolism*; D005609:Free Radicals; D019787:Linoleic Acid / Q000378:metabolism; D050356:Lipid Metabolism; D015227:Lipid Peroxidation; D008084:Lipoxygenase / Q000737:chemistry*; D013058:Mass Spectrometry; D008956:Models, Chemical; D010100:Oxygen / Q000378:metabolism; D013997:Time Factors
D043371:Fatty Acids, Omega-6; D005609:Free Radicals; D016718:Arachidonic Acid; D002244:Carbon; D019787:Linoleic Acid; C101084:lipoxygenase L-1; D008084:Lipoxygenase; D010100:Oxygen