OXIDATION AND ANTIOXIDATION OF HUMAN LOW-DENSITY-LIPOPROTEIN AND PLASMA EXPOSED TO 3-MORPHOLINOSYDNONIMINE AND REAGENT PEROXYNITRITE

As peroxynitrite is implicated as an oxidant for low-density lipoprote in (LDL) in atherogenesis, we investigated this process using reagent peroxynitrite (ONOO-) and 3-morpholinosydnonimine (SIN-1, which produc es peroxynitrite via generation of NO. and O-2(.-)). LDL oxidation was assessed by the consumption of ubiquinol-10 (CoQ(10)H(2)) and alpha-t ocopherol (alpha-TOH), the accumulation of cholesteryl ester hydro(per o)xides, the loss of lysine (Lys) and tryptophan (Trp) residues, and t he change in relative electrophoretic mobility. Exposure to ONOO- or S IN-1 resulted in rapid (<1 min) and time-dependent oxidation, respecti vely, of LDL's lipids and protein. Manipulating the alpha-TOH content by in vivo or in vitro means showed that when ONOO- or SIN-1 was used at oxidant-to-LDL ratios of (100:1 the extent of LDL lipid peroxidatio n increased with increasing initial alpha-TOH content. In contrast, in vivo enrichment with the co-antioxidant CoQ(10)H(2) decreased LDL lip id peroxidation induced by SIN-1. At oxidant-to-LDL ratios of 1200:1, alpha-TOH enrichment decreased LDL lipid peroxidation for both SIN-1 a nd ONOO-. In contrast to Lipid peroxidation, altering the alpha-TOH co ntent of LDL did not affect Trp or Lys loss, independent of the amount s of either oxidant added. Aqueous antioxidants inhibited ONOO--induce d lipid and protein oxidation with the order of efficacy: 3-hydroxyant hranilate (3-HAA) > mate > ascorbate. With SIN-1, these antioxidants i nhibited Trp consumption, while only the co-antioxidants ascorbate and 3-HAA prevented alpha-TOH consumption and lipid peroxidation. Exposur e of human plasma to SIN-1 resulted in the loss of ascorbate followed by loss of CoQ(10)H(2) and bilirubin. Lipid peroxidation was inhibited during this period, though proceeded as a radical-chain process after depletion of these antioxidants and in the presence of alpha-TOH and mate. Bicarbonate at physiological concentrations decreased ONOO--indu ced lipid and protein oxidation, whereas it enhanced SIN-1-induced lip id peroxidation, Trp consumption, and alpha-tocopheroxyl radical forma tion in LDL. These results indicate an important role for tocopherol-m ediated peroxidation and co-antioxidation in peroxynitrite-induced lip oprotein Lipid peroxidation, especially when peroxynitrite is formed t ime-dependently by SIN-1. The studies also highlight differences betwe en ONOO-- and SIN-1-induced LDL oxidation with regards to the effects of bicarbonate; ascorbate, and urate.