Determination of singlet oxygen-specific versus radical-mediated lipid peroxidation in photosensitized oxidation of lipid bilayers: effect of beta-carotene and alpha-tocopherol.
Photosensitized oxidation reactions damage tissue by catalyzing the formation of oxyradicals and singlet oxygen. beta-Carotene is hypothesized to exert photoprotective effects by quenching singlet oxygen formed by Type II reactions and by scavenging free radicals formed by Type I reactions. beta-Carotene antioxidant mechanisms were studied in a phospholipid membrane model of photooxidation with a new isotope dilution gas chromatography-mass spectrometry (GC-MS) assay that quantitatively distinguishes singlet oxygen-mediated and radical-mediated lipid peroxidation. This assay measures 9- and 10-hydroxylinoleate methyl esters and was used to generate photooxidation profiles for the photosensitizers methylene blue, Rose Bengal, and tetraphenylporphine. These profiles indicate a shift from Type II to Type I photooxidation mechanisms in later stages of photooxidation. beta-Carotene (0.45 mol %) inhibited singlet oxygen-mediated lipid peroxidation at early stages of methylene blue-sensitized photooxidation. Production of radical-mediated products increased faster than singlet oxygen-mediated products at later stages. beta-Carotene-5,8-endoperoxide, a specific marker for singlet oxygen oxidation of beta-carotene in solution, was unstable under the incubation conditions and was not detected in this system. alpha-Tocopherol (0.45 mol %) was ineffective in inhibiting photosensitized lipid peroxidation, whereas 4.5 mol % alpha-tocopherol inhibited almost all radical-mediated lipid peroxidation as well as early-stage singlet oxygen-mediated lipid peroxidation. Cumene hydroperoxide stimulated radical-mediated lipid peroxidation, indicating that accumulation of hydroperoxides from Type II photooxidation may enhance Type I reactions. These data suggest that singlet oxygen quenching, rather than radical scavenging reactions, accounts for the photoprotective actions of beta-carotene.
Stratton
S P
SP
Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson, Arizona 85721, USA.
Liebler
D C
DC
D002851:Chromatography, High Pressure Liquid; D005609:Free Radicals; D008401:Gas Chromatography-Mass Spectrometry; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry*; D015227:Lipid Peroxidation; D010084:Oxidation-Reduction; D010100:Oxygen; D010777:Photochemistry; D026082:Singlet Oxygen; D014810:Vitamin E; D019207:beta Carotene
D005609:Free Radicals; D008051:Lipid Bilayers; D019207:beta Carotene; D014810:Vitamin E; D026082:Singlet Oxygen; D010100:Oxygen