Hydroxyl radical, formation

McCormick ML, GR Buettner, BE Britigan (1998) Endogenous superoxide dismutase levels regulate iron-dependent hydroxyl radical formation in Escherichia coli exposed to hydrogen peroxide. J Bacterial 180 622-625. 

Grootveld, M. and Halliwell, B. (1986b). Aromatic hydroxyla-tion as a potential measure of hydroxyl radical formation in vivo. Identification of hydroxylated derivatives of salicylate in human body fluids. Biochem. J. 237, 499-504. 

Beckman, J.S., Beckman, T.W., Chen, J., Marshall, P.A. and Freeman, B.A. (1990). Apparent hydroxyl radical formation by peroxynitrite implications for endothelial cell injury from nitric oxide and superoxide anion. Proc. Natl Acad. Sci. USA 87, 1620-1624. 

For a long time one question remained unanswered the efficiency of the Fenton reaction as the in vivo producer of hydroxyl radicals due to the low rate of Reaction (2) (the rate constant is equal to 42.11 mol 1 s 1 [18]). It is known that under in vitro conditions the rate of Fenton reaction can be sharply enhanced by chelators such as EDTA, but for a long time no effective in vivo chelators have been found. From this point of view new findings obtained by Chen and Schopfer [19] who found that peroxidases catalyze hydroxyl radical formation in plants deserve consideration. These authors showed that horseradish peroxidase (HRP) compound III is a catalyst of the Fenton reaction and that this compound is one to two orders of magnitude more active than Fe EDTA. 

Of course, superoxide may reduce ferric to ferrous ions and by this again catalyze hydroxyl radical formation. Thus, the oxidation of ferrous ions could be just a futile cycle, leading to the same Fenton reaction. However, the competition between the reduction of ferric ions by superoxide and the oxidation of ferrous ions by dioxygen depends on the one-electron reduction potential of the [Fe3+/Fe2+] pair, which varied from +0.6 to —0.4 V in biological systems [173] and which is difficult to predict.)... 

Iron-stimulated free radical-mediated processes are not limited to the promotion of peroxidative reactions. For example, Pratico et al. [188] demonstrated that erythrocytes are able to modulate platelet reactivity in response to collagen via the release of free iron, which supposedly catalyzes hydroxyl radical formation by the Fenton reaction. This process resulted in an irreversible blood aggregation and could be relevant to the stimulation by iron overload of atherosclerosis and coronary artery disease. 

Recent studies suggest that many factors may affect hydroxyl radical generation by microsomes. Reinke et al. [34] demonstrated that the hydroxyl radical-mediated oxidation of ethanol in rat liver microsomes depended on phosphate or Tris buffer. Cytochrome bs can also participate in the microsomal production of hydroxyl radicals catalyzed by NADH-cytochrome bs reductase [35,36]. Considering the numerous demonstrations of hydroxyl radical formation in microsomes, it becomes obvious that this is not a genuine enzymatic process because it depends on the presence or absence of free iron. Consequently, in vitro experiments in buffers containing iron ions can significantly differ from real biological systems. 

However, subsequent studies demonstrated that the formation of hydroxyl radicals, even if it takes place during lipid peroxidation, is of no real importance. Beloqui and Cederbaum [11] have found that although the glutathione-glutathione peroxidase system suppressed hydroxyl radical generation during the oxidation of 4-methylmercapto-2-oxo-butyrate, it exhibited a much smaller effect on microsomal lipid peroxidation. Therefore, hydroxyl radical formation is apparently unimportant in this process. Other authors also pointed out at an unimportant role of hydroxyl radicals in the initiation of microsomal lipid peroxidation [12 14], For example, it has been shown that Fe(EDTA), a most efficient catalyst of hydroxyl radical formation by the Fenton reaction, inhibited microsomal and liposomal lipid peroxidation, while the weak catalysts of this reaction Fe(ADP) and Fe(ATP) enhanced it [13]. 

Xanthine oxidase, a widely used source of superoxide, has been frequently applied for the study of the effects of superoxide on DNA oxidation. Rozenberg-Arska et al. [30] have shown that xanthine oxidase plus excess iron induced chromosomal and plasmid DNA injury, which was supposedly mediated by hydroxyl radicals. Ito et al. [31] compared the inactivation of Bacillus subtilis transforming DNA by potassium superoxide and the xanthine xanthine oxidase system. It was found that xanthine oxidase but not K02 was a source of free radical mediated DNA inactivation apparently due to the conversion of superoxide to hydroxyl radicals in the presence of iron ions. Deno and Fridovich [32] also supposed that the single strand scission formation after exposure of DNA plasmid to xanthine oxidase was mediated by hydroxyl radical formation. Oxygen radicals produced by xanthine oxidase induced DNA strand breakage in promotable and nonpromotable JB6 mouse epidermal cells [33]. 

Conflicting data were also received for the reactions of LA and DHLA with hydroxyl radicals and superoxide. Suzuki et al. [206] found that both LA and DHLA inhibited the formation of DMPO-OH adducts formed in the Fenton reaction. However, Scott et al. [207] concluded that only LA is a powerful scavenger of hydroxyl radicals while DHLA accelerated iron-catalyzed hydroxyl radical formation and lipid peroxidation. 

Chelators of transition metals, mainly iron and copper, are usually considered as antioxidants because of their ability to inhibit free radical-mediated damaging processes. Actually, the so-called chelating therapy has been in the use probably even earlier than antioxidant therapy because it is an obvious pathway to treat the development of pathologies depending on metal overload (such as calcium overload in atherosclerosis or iron overload in thalassemia) with compounds capable of removing metals from an organism. Understanding of chelators as antioxidants came later when much attention was drawn to the possibility of in vivo hydroxyl radical formation via the Fenton reaction ... 

It was found that various chelators are able to affect the hydroxyl radical formation in a different way, namely, some chelators (EDTA) can increase the rate of the Fenton reaction, while others (rutin) can inhibit it through the formation of inactive iron-chelator complexes. 

Marston, G., C. D. McGill, and A. R. Rickard, Hydroxyl-Radical Formation in the Gas-Phase Ozonolysis of 2-Methylbut-2-ene, Geophys. Res. Lett, 25, 2177-2180 (1998). 

Zepp, R. G., B. C. Faust, and J. Hoigne, Hydroxyl Radical Formation in Aqueous Reactions (pH 3-8) of Iron(II) with Hydrogen Peroxide The Photo-Fenton Reaction, Environ. Sci. Technoi, 26, 313-319 (1992). 

Thus, antioxidant effects of nitrite in cured meats appear to be due to the formation of NO. Kanner et al. (1991) also demonstrated antioxidant effects of NO in systems where reactive hydroxyl radicals ( OH) are produced by the iron-catalyzed decomposition of hydrogen peroxide (Fenton reaction). Hydroxyl radical formation was measured as the rate of benzoate hydtoxylation to salicylic acid. Benzoate hydtoxylation catalyzed by cysteine-Fe +, ascorbate - EDTA-Fe, or Fe was significantly decreased by flushing of the reaction mixture with NO. They proposed that NO liganded to ferrous complexes reacted with H2O2 to form nitrous acid, hydroxyl ion, and ferric iron complexes, preventing generation of hydroxyl radicals. 

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