Dehydroascorbate reduction

Nardai G, Braun L, Csala M, Mile V, Csermely P, Benedetti A, Mandl J, Banhegyi G. Protein-disulfide isomerase- and protein thiol-dependent dehydroascorbate reduction and ascorbate accumulation in the lumen of the endoplasmic reticulum. J. Biol. Chem. 2001 276 8825-8828. 

Ascorbic acid, for example, is oxidized to dehydroascorbic acid with reduction of the iron(III) ions. The Fe(II) ions so produced react with 2,2 -bipyridine with formation of a colored complex. 

In a recent study, serum ascorbate concentrations were significantly reduced in a group of elderly diabetic patients (w = 40, mean age 69 years) in comparison with an age-matched group of non-diabetic controls ( = 22, mean age 71 years), and this reduction was more pronounced in those patients with microangiopathy (Sinclair et al., 1991). Diabetic patients were shown to have a high serum dehydroascorbate/ascorbate ratio indicative of increased oxidative stress. Ascorbate deficiency was partially corrected by vitamin C supplementation, 1 g daily by mouth, but the obvious disturbance in ascorbate metabolism in the diabetic patients was accentuated, since serum ascorbate concentrations fell (after the initial rise) despite continued vitamin C supplementation (Fig. 12.3). 

BOROHYDRIDE REDUCTION DEHYDROASCORBATE REDUCTASE DEHYDROCHOLESTEROL REDUCTASE 2-DEHYDRO-3-DEOXY-D-GLUCONATE... 

Solution Abbreviating dehydroascorbic acid as D, and ascorbic acid as H2A, we rewrite the reduction as... 

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. 

Analysis of vitamin C has been the topic of numerous papers. Most of the methods analyze L-ascorbic acid by HPLC, before and after reduction of the dehydroascorbic acid present.338 The concentration of the dehydroascorbic acid is calculated by subtraction. A later work describes a method, which combines iodometry with a voltammetric technique to detect the endpoint of the titration.339 The results are comparable to those obtained by HPLC and can be applied to vegetable and fruit samples. 

The enzyme catalyses the oxidation of L-ascorbic acid, 4, to dehydroascorbic, 5. The blue protein belongs to the group of blue oxidases together with laccase and ceruloplasmin. These are multicopper enzymes catalysing the electron reduction of molecular oxygen to water with concomitant one electron oxidation of the substrate. 

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) ... 

The first step in the mechanism involves the reduction of Cu(II) to Cu(I) by ascorbyl-6-hexadeeanoate giving dehydroascorbic acid and a weak acid HY benzoic acid). In fact this stage of the process has no importance since Cu(I) benzoate may directly be used to initiate the polymerization by reducing the pyridinium salt. The strong Bronsted acid formed attacks the monomer and initiates the polymerization. Notably, lower polymer yields were obtained by using pyridium salt rather than iodonium salt. 

Cu2(L )(NCS)2] (66), which has been shown (c.f. Fig. 8) [43] to contain two tetrahedrally coordinated Cu(I) ions held 2.796(8) A apart and linked inter-molecularly via the sulfur atoms of the thiocyanate ions. For the complexes 63 and 64, the reduction product is the diamagnetic complex [Cu2(L )(MeCN)2](Y)2 (67) (Y = CIO4 or BPh4) in which each three coordinate Cu(I) ion is bonded to two of the four macrocyclic nitrogen atoms and to the nitrogen of one of the two MeCN molecules (Fig, 9) [43]. In the presence of certain substrates the reduction of 63 or 64 is accompanied by substrate oxidation. For example, PhSH, PhC=CH, hydrazobenzene, catechols, hydroquinone, and ascorbic acid afford PhSSPh, PhC=CCsPh, azobenzene, o-quinones, p-quinone, and dehydroascorbic acid, respectively, together with the reduced species 67 and/or other copper complexes... 

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). 

Vitamin C can also be determined colorimetrically, after oxidation to dehydroascorbate, by reaction with dinitrophenylhydrazine. Under appropriate conditions, neither ascorbic acid itself nor potentially interfering sugars react with dinitrophenylhydrazine. However, diketogulonate, which has no vitamin activity, also reacts with dinitrophenylhydrazine under the same conditions. Unless diketogulonate is determined separately after reduction of dehydroascorbate to ascorbate, this method overestimates the vitamin. 

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. 

Luminal thiol oxidation is facilitated by ascorbate (vitamin C) (45) or FAD (12, 13), so the physiologic role of their transport has been proposed. ER membrane is permeable selectively to dehydroascorbate, the oxidized form of ascorbate (10, 11). Luminal reduction of dehydroascorbate to ascorbate is associated with thiol oxidation and leads to ascorbate entrapment (46). 

Figure 12. Ascorbic acid-dehydroascorbic acid redox system (a) oxidation of ascorbate to semidehydroascorbic acid, (b) disproportionation of semidehydroascorbic acid, and (c) reduction of dehydroascorbic acid [From (100), with permission].

These differences are correlated with differing functions. The intracellular reduced thioredoxins are thermodynamically the best reductants of disulfide linkages in proteins and they help keep intracellular proteins reduced. Glutaredoxin can be reduced efficiently by reduced glutathione or by NADPH and glutathione reductase and can, in turn, reduce cysteine and the oxidized form of vitamin C, dehydroascorbic acid (Box The peri-... 

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