Dehydroascorbate radical

Pulse radiolysis was also used to elucidate the mechanism of catalytic action of monodehydroascorbate reductase, an enzyme containing FAD and using Nicotinaminde atjenine dinucleotide (NADH) as reductant. The substrate is dehydroascorbate radical produced by pulse radiolysis (130). The authors show that this radical reacts with the protein to give the FADH radical and that the... 

It has long been recognized that ascorbate levels are low in patients with RA (Lunec and Blake, 1985) and ascorbate is predominantly found in the dehydro form. The presence of increased dehydroascorbate has been suggested to indicate its rapid oxidation by stimulated PMNs (Halliwell and Gutteridge, 1990). When ascorbate concentrations are lower than about 20 /tmol/1, as can occur in rheumatoid synovial fluid, the Fe(III) reducing effects of ascorbate outweigh its radical-scavenging effects. Ascorbate then causes increased OH formation and promotes lipid peroxidation (Blake et al., 1981). 

Ascorbate is known to act as a water-soluble antioxidant, reacting rapidly with superoxide, hydroxyl and peroxyl radicals. However, reduced ascorbate can react non-enzymatically with molecular oxygen to produce dehydroascorbate and hydrogen peroxide. Also, ascorbate in the presence of light, hydrogen peroxide and riboflavin, or transition metals (e.g. Fe, Cu " ), can give rise to hydroxyl radicals (Delaye and Tardieu, 1983 Ueno et al., 1987). These phenomena may also be important in oxidative damage to the lens and subsequent cataract formation. 

In contrast, antioxidant enzymes can efficiently counteract all UV-induced ROS (Aguilera et al. 2002). These enzymes are represented by superoxide dismutase (SOD), catalase and glutathione peroxidase as well as those involved in the ascorbate-glutathione cycle, such as ascorbate peroxidase, mono-dehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase. One of the most important classes of antioxidant enzymes is the SOD family, which eliminate noxious superoxide radical anions. Different metalloforms of SOD exist (Fe, Mn, CuZn and Ni), which due to their intracellular localisation protect different cellular proteins (Lesser and Stochaj 1990). 

The biological functions of vitamin C appear to be related principally to its well-established reducing properties and easy one-electron oxidation to a free radical or two-electron reduction to dehydroascorbic acid. The latter is in equilibrium with the hydrated hemiacetal shown at the beginning of this box as well as with other chemical species.1 Vitamin C is a weak acid which also has metal complexing properties. 

Two ascorbate radicals can react with each other in a disproportionation reaction to give ascorbate plus dehydroascorbate. However, most cells can reduce the radicals more directly. In many plants this is accomplished by NADH + H+ using a flavoprotein monodehydroascorbate reductase.0 Animal cells may also utilize NADH or may reduce dehydroascorbate with reduced glutathione.CC/ff Plant cells also contain a very active blue copper ascorbate oxidase (Chapter 16, Section D,5), which catalyzes the opposite reaction, formation of dehydroascorbate. A heme ascorbate oxidase has been purified from a fungus. 11 1 Action of these enzymes initiates an oxidative degradation of ascorbate, perhaps through the pathway of Fig. 20-2. 

Compounds that can scavenge radicals are also referred to as antioxidants. The best known anti-oxidants are vitamin C and vitamin E. Vitamin C is L-ascorbate (7.8), a good reducing agent that prevents oxidation of other molecules. The oxidized form of L-ascorbate is L-dehydroascorbic acid (7.9). Vitamin E is a mixture of a-, [3-, y-, and 8-tocopherol (7.10a-d). Of these four compounds, a-tocopherol is the most effective. Vitamin E is lipid-soluble and has the ability to disrupt the chain reaction during lipid peroxidation (see Chapter 2, Section 1.9). 

Vitamin C (ascorbate) (Fig. 9.5) has the ability to act as a reducing agent, i.e. it will tend to reduce more reactive species. This ability to reduce Fe3+ to Fe2+may be important in promoting iron uptake in the gut. Oxidation of ascorbate by reaction with reactive oxygen species or reactive nitrogen species seems to lead to its depletion. In vitro, vitamin C can also exert pro-oxidant properties. Fe3+ can react with ascorbate to form Fe2+ and the semi-dehydroascorbate or ascorbyl radical. The latter can react with hydrogen peroxide to form Fe3+, the hydroxyl radical and a hydroxide anion. A key question with regard to the pro- or anti- oxidant effects of ascorbate may therefore be the availability of transition metal ions. Neurons main-... 

In its reduced form, a-lipoic acid (Fig. 9.5) is a powerful antioxidant. It can reduce GSSG to GSH, dehydroascorbate to ascorbate, and regenerate a-tocopherol from the cy-tocopheryl radical either directly or via ascorbate. Supplementation with a-lipoic acid decreases oxidative stress and restores reduced levels of other antioxidants in vivo. However, a-lipoic acid and dihydrolipoic acid may exert pro-oxidant properties in vitro. a-Lipoic acid and dihydrolipoic acid promote the permeability... 

The radical form 9.4 has an unpaired electron and may undergo fast reactions with redox partners that also undergo one-electron processes. Such a redox partner is the triplet radical, dioxygen. The copper complex of ascorbic acid undergoes rapid aerial oxidation to give the dione, dehydroascorbic acid, which may be viewed as being derived by electron loss from the radical (Fig. 9-4). 

Figure 12.4 illustrates the electrochemical characteristics of ascorbic acid and its anion (H2A and HA-) and of dehydroascorbic acid (A) and its anion radical (A ) in DMF at a platinum electrode. The hydrogens of H2A are moderately acidic (in H20 p=4.1, pK2 = 11.8), which accounts for the cathodic voltammograms (Figure 12.4a) ...

In contrast to H2A, electrochemical reduction of A yields the anion radical of dehydroascorbic (A- -) in an irreversible, complex electron-transfer process (Figure 12.4c, d) ... 

FIGURE 29.1 Pathways of the chain-breaking action of vitamin E in lipid peroxidation and its subsequent regeneration. LOOH lipid hydroperoxide, LOO lipid peroxyl radical, vitamin C ascorbate radical (semi-dehydroascorbate), vitamin E a-tocopheroxyl radical. The lipid peroxyl radical is reduced to lipid hydroperoxide by tocopherol. The resulting tocopheroxyl radical can be re-reduced by ascorbate. The thus formed ascorbate radical can be reduced to ascorbate by the NADH-dependent semidehydroascorbate reductase. 

Antioxidant compounds are an important defense for immediate detoxication of highly reactive intermediates. They act as competing nucleophiles and can bind to the intermediate, forming a less reactive species. Glutathione is one antioxidant molecule that can directly interact with free radicals. Other chemical antioxidants include vitamins A (retinol), C (ascorbic acid), and E. Ascorbic acid, for example, may react directly with reactive intermediates by hydrogen abstraction, resulting in the formation of dehydroascorbic acid. 

Donation of one electron by ascorbate gives the semidehydroascorbate radical which on further oxidation gives dehydroascorbate, according to the scheme in Fig. 7. Dehydroascorbate is unstable and breaks down by a... 

Delocalisation onto oxygen stabilizes radicals considerably. An important example is the ascorbate radical (Scheme 1.3) formed by electron-loss from the ascorbate anion, or electron-capture by dehydroascorbate. This is remarkably stable, and is characterized by an ESR doublet (1.7 G) which is quite distinctive. Because of the high sensitivity of ESR spectroscopy, and the fact that opaque samples can be used, ascorbate radical intermediates have been widely studied (Liu et al., 1988a). The most probable structure is shown in Scheme 1.3 but this is still a matter of some controversy (Liu et al., 1988a). A key factor in the formation of ascorbate radicals is that ascorbate anions... 

This contrasts with the ascorbate system in which two ascorbate radicals give ascorbate + dehydroascorbate. 

By reaction with ascorbate to yield the monodehydroascorbate radical, which in turn can either be reduced to ascorbate or can undergo dis-mutation to yield dehydroascorbate and ascorbate (Section 13.4.7.1). In vitro, the formation of the tocopheroxyl radical can be demonstrated by the appearance of its characteristic absorbance peak, which normally has a decay time of 3 msec in the presence of ascorbate, the tocopheroxyl peak has a decay time of 10 /rsec, and its disappearance is accompanied by the appearance of the monodehydroascorbate peak. There is an integral membrane oxidoreductase that uses ascorbate as the preferred electron donor, linked either directly to reduction of tocopheroxyl radical or via an electron transport chain involving ubiquinone (see no. 4 below May, 1999). 

As shown in Figure 13.3, oxidation of ascorbic acid, for example, by the reduction of superoxide to hydrogen peroxide or Fe + to Fe +, and similar reduction of other transition metal ions, proceeds by a one-electron process, forming the monodehydroascorbate radical. The radical rapidly disproportionates into ascorbate and dehydroascorbate. Most tissues also have both nicotinamide adenine dinucleotide phosphate (NADPH) and glutathione-dependent monodehydroascorbate reductases, which reduce the radical back to ascorbate. Ascorbate is thus an effective quencher of singlet oxygen and other radicals. 

The antioxidant activity of ascorbate is variable. From consideration of the chemistry involved, it would be expected that, overall, 2 moles of peroxyl radical would be trapped per mole of ascorbate, because of the reaction of 2 moles of monodehydroascorbate to regenerate ascorbate and yield dehydroascorbate (see Figure 13.3). However, as the concentration of ascorbate increases, so the molar ratio decreases, and it is only at very low concentrations of ascorbate that it tends toward the theoretical 2 1. 

The immediate product of the oxidative reaction, the monodehydroascorbate radical (Eq. [1]), is a fairly reactive and unstable species which, in the presence of a suitable reductase system, is reduced back to ascorbate. Monodehydroascorbate reductases have been identified and purified in a few cases (Ushimara et al., 1997 Dalton et al., 1992 Shigeoka et al., 1987 Borraccino et al., 1986 Hossain et al., 1984) and cDNA sequences have been published for the pea (Murthy and Zilinskas, 1994) and cucumber enzymes (Sano et al., 1995). A bacterial expression system is also available for the cucumber enzyme (Sano et al., 1995). In the absence of a suitably efficient reductase system, the monodehydroascorbate radicals disproportionate to dehydroascorbate and ascorbate in this case, ascorbate is regenerated using a glutathione-dependent dehydroascorbate reductase enzyme (Foyer and Mullineaux, 1998 and references therein). Under non-... 

The term vtiamin C refers to ascorbic acid (the fully reduced form of the vitamin) and to dchydroascorbic acid- Removal of one electron from ascorbic acid yields semidehydroascorbic acid (ascorbate radical). This form of the vitamin is a free radical it contains an unpaired electron- The structures of free radicals are written with large dots. The removal of a second electron yields dehydroascorbic acid. Conversion of ascorbate to dehydroascorbate, via the removal of two electrons, can occur under two conditions (1) with use of ascorbic add by ascorbate-dependent enzymes and (2) with the spontaneous reaction of ascorbate with oxygen. Semidehydroascorbate is an intermediate in this conversion palhway... 

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