Kinetic study of aroxyl radical-scavenging action of vitamin E in membranes of egg yolk phosphatidylcholine vesicles.

Vitamin E is localized in membranes and functions as an efficient inhibitor of lipid peroxidation in biological systems. In this study, we measured the reaction rates of vitamin E (α-, β-, γ-, δ-tocopherols, TocH) and tocol with aroxyl radical (ArO·) as model lipid peroxyl radicals in membranes by stopped-flow spectrophotometry. Egg yolk phosphatidylcholine (EYPC) vesicles were used as a membrane model. EYPC vesicles were prepared in the aqueous methanol solution (MeOH:H(2)O=7:3, v/v) that gave the lowest turbidity in samples. The second-order rate constants (k(s)) for α-TocH in MeOH/H(2)O solution with EYPC vesicles were apparently 3.45×10(5)M(-1)s(-1), which was about 8 times higher than that (4.50×10(4)M(-1)s(-1)) in MeOH/H(2)O solution without EYPC vesicles. The corrected k(s) of α-TocH in vesicles, which was calculated assuming that the concentration of α-TocH was 133 times higher in membranes of 10mM EYPC vesicles than in the bulk MeOH/H(2)O solution, was 2.60×10(3)M(-1)s(-1), which was one-seventeenth that in MeOH/H(2)O solution because of the lower mobility of α-TocH in membranes. Similar analyses were performed for other vitamin E analogues. The k(s) of vitamin E in membranes increased in the order of tocol<δ-TocH<γ-TocH∼β-TocH<α-TocH. There was not much difference in the ratios of reaction rates in vesicles and MeOH/H(2)O solution among vitamin E analogues [k(s)(vesicle)/k(s) (MeOH/H(2)O)=7.7, 10.0, 9.5, 7.4, and 5.1 for α-, β-, γ-, δ-TocH, and tocol, respectively], but their reported ratios in solutions of micelles and ethanol were quite different [k(s)(micelle)/k(s)(EtOH)=100, 47, 41, 15, and 6.3 for α-, β-, γ-, δ-TocH, and tocol, respectively]. These results indicate that the reaction sites of vitamin E analogues were similar in vesicle membranes but depended on hydrophobicity in micelle membranes, which increased in the order of tocol<δ-TocH<γ-TocH∼β-TocH<α-TocH.