2.3- Diketogulonic acid

Chromatographic methods, notably hplc, are available for the simultaneous deterrnination of ascorbic acid as weU as dehydroascorbic acid. Some of these methods result in the separation of ascorbic acid from its isomers, eg, erythorbic acid and oxidation products such as diketogulonic acid. Detection has been by fluorescence, uv absorption, or electrochemical methods (83—85). Polarographic methods have been used because of their accuracy and their ease of operation. Ion exclusion (86) and ion suppression (87) chromatography methods have recently been reported. Other methods for ascorbic acid deterrnination include enzymatic, spectroscopic, paper, thin layer, and gas chromatographic methods. ExceUent reviews of these methods have been pubHshed (73,88,89). 

A chemical reaction subsequent to a fast (reversible) electrode reaction (Eq. 5.6.1, case b) can consume the product of the electrode reaction, whose concentration in solution thus decreases. This decreases the overpotential of the overall electrode process. This mechanism was proposed by R. Brdicka and D. H. M. Kern for the oxidation of ascorbic acid, converted by a fast electrode reaction at the mercury electrode to form dehydro-ascorbic acid. An equilibrium described by the Nernst equation is established at the electrode between the initial substance and this intermediate product. Dehydroascorbic acid is then deactivated by a fast chemical reaction with water to form diketogulonic acid, which is electroinactive. 

Since ascorbate reduces photooxidation of lipid emulsions and multivitamin preparations (see Figure 4) [19], Lavoie et al. [34] studied the formation of oxidative by-products of vitamin C in multivitamins exposed to light. They found that the loss of ascorbic acid in photoexposed multivitamin preparations was associated with the generation of products other than dehydroascorbate and 2,3-diketogulonic acid, which are the usual products of vitamin C oxidation. The authors showed that hydrogen peroxide at concentrations found in TPN solutions induced the transformation of dehydroascorbate into new, biologically active compounds that had the potential to affect lipid metabolism. They believe that these species have peroxide and aldehyde functions [35]. 

The concentration of ascorbic acid in milk (11.2-17.2mgl-1) is sufficient to influence its redox potential. In freshly drawn milk, all ascorbic acid is in the reduced form but can be oxidized reversibly to dehydroascorbate, which is present as a hydrated hemiketal in aqueous systems. Hydrolysis of the lactone ring of dehydroascorbate, which results in the formation of 2,3-diketogulonic acid, is irreversible (Figure 11.2). 

Both forms are biologically active. Ascorbic acid (the reduced form) is relatively stable to heat however, dehydroascorbic acid (the oxidized form) is unstable. The lactone ring is easily hydrolyzed to diketogulonic acid, which has no antiscurvy activity. When fruits, vegetables, and other foods are heated, there is some loss of active vitamin C by conversion to diketogulonic acid. 

According to the scheme, ascorbic acid is readily converted into pentoses, but this does not explain the differences in behaviour between ascorbic acid and pentoses. 2,3-Diketogulonic acid (and the corresponding enol) is very unstable and readily develops brown coloration even at low temperatures.550 It can also undergo oxidative fission. 

Dehydroascorbate is unstable in solution, undergoing hydrolytic ring opening to yield diketogulonic acid. However, in vivo, it is normally reduced to... 

Disposition in the Body. Readily absorbed after oral administration the proportion of a dose absorbed tends to decrease with increasing dose it is widely distributed in the body tissues. The concentration of ascorbic acid is higher in leucocytes and platelets than in erythrocytes and plasma. Ascorbic acid is metabolised to dehydroascorbic acid, 2,3-diketogulonic acid, oxalate, and carbon dioxide some conjugation with sulphate occurs to form ascorbate-3-sulphate. Ascorbic acid in excess of the body s requirements is rapidly eliminated in the urine. About 85% of an intravenous dose, given to subjects previously saturated with the vitamin, is excreted in the urine in 24 hours, with about 70% of the dose excreted unchanged and 15% as dehydroascorbic acid and diketogulonic acid. The amount normally present in the body is in excess of 1.5 g. 

The DHA dimer is converted to the monomer when it is dissolved in water. The chemistry of DHA is reviewed, including the hydrolysis to diketogulonic acid and the reactions of the 2- and 3-oxo groups. DHA readily forms Schiff bases and undergoes a Strecker reaction with amino acids. 

The main metabolites of ascorbate in the human body are oxalate, dehydroascorbic acid, 2,3-diketogulonic acid, and ascorbic acid 2-sulfate. Of these, oxalate has attracted the most attention because of the potential hazard for renal complications by precipitation of oxalate stones. The above-mentioned metabolic turnover for ascorbate, which appears saturable, indicates that in normal humans the amount of oxalate that can be formed is limited. Therefore, the overall risk of inducing oxalate precipitation by increasing the intake of ascorbate probably is minute. 

Liquid Dosage Forms. In dry form and at very low moisture content, L-ascorbic acid is very stable, but in solution exposed to air or oxygen it is subject to oxidation accelerated by dissolved trace minerals (copper and iron) and light exposure, l-Ascorbic acid is a reducing agent and is subject to oxidative decomposition in solution. This proceeds first to dehydroascorbic acid, which has full vitamin C activity, but continues to diketogulonic acid and various other breakdown products. The degradation reactions are complex and vary with aerobic or anaerobic... 

Loss in Potency (%) Diketogulonic Acid (%) Oxalic Acid Dehydroascorbic (%) Acid(%) ... 

DHA can be reduced to RAA by chemical agents, such as hydrogen sulfide or enzymatically, by dehydroascorbic acid reductase. The conversion of DHA to diketogulonic acid (DKG) is irreversible and occurs both aerobically and anaerobically, particularly during heating. This reaction results in loss of biological activity. The total oxidation of RAA may result in the formation of furfural by decarboxylation and dehydration. With subsequent polymerization, the formation of dark-colored pigments results. These compounds affect the color and flavor of certain foods, such as citrus juices, and decrease nutritive value. 

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