tocopherylquinone and linoleic-acid-hydroperoxide

The turnover rate of vitamin E is slow in nerve tissue. Therefore, we have developed in vitro techniques to study the biochemical reactions of this nutrient in brain. Subcellular fractions were isolated from the cerebral hemispheres of 4-month-old, male, Fisher 344 rats. Aliquots of fractions (500 micrograms protein) were suspended in 50 mM phosphate buffer at pH 7.4 and incubated at room temperature (20-22 degrees) or 37 degrees for 2 hr in the presence or absence of the following oxidizing agents: 1 mM tertiary butyl hydroperoxide, 10 microM linoleic acid hydroperoxide, 0.5 to 50 mM 2,2'-azobis (2-amidinopropane) dihydrochloride (ABAPH) or 0.1 to 2 mM 2,2'-azobis (2,4-dimethyl) valeronitrile (ABDVN). The latter two compounds generate free radicals upon heating. After oxidation, the subcellular fractions were sedimented, saponified and assayed for tocopherol by liquid chromatography. Linoleic acid hydroperoxide was the most potent oxidizing agent, suggesting that endogenous fatty acid peroxides (e.g. eicosanoid intermediates) are very powerful oxidizing agents. Vitamin E may play an important role in providing antioxidant protection for membranes against excessive oxidation induced by these peroxides. Tocopherol in mitochondria and microsomes was much more susceptible to oxidation than synaptosomal tocopherol. The possible reasons for this observation are: (a) mitochondria and microsomes may contain less of the other reducing agents such as sulfhydryl compounds than synaptosomes, and/or (b) the electron transport structures in the former two subcellular fractions may be facilitating oxidation of tocopherol induced by free radicals. A portion of tocopherol remained unoxidized in all subcellular fractions even at high concentrations of ABAPH, suggesting that tocopherol exists in labile and nonlabile biochemical compartments or complexes.

Topics: Amidines; Animals; Brain Chemistry; Cell Membrane; Linoleic Acids; Lipid Peroxides; Male; Oxidation-Reduction; Rats; Rats, Inbred F344; Subcellular Fractions; Vitamin E