Although tea catechins in green tea and green tea beverages must be stable to deliver good sensory quality and healthy benefits, they are always unstable during processing and storage. Ascorbic acid ...(AA) is often used to protect catechins in green tea beverages, and AA is easily oxidized to form dehydroascorbic acid (DHAA). However, the function of DHAA on the stability of catechins is not clear. The objective of this study was to determine the effects of DHAA on the stability of catechins and clarify the mechanism of effects by conducting a series of experiments that incubate DHAA with epigallocatechin gallate (EGCG) or catechins. Results showed that DHAA had a dual function on EGCG stability, protecting its stability by inhibiting hydrolysis and promoting EGCG consumption by forming ascorbyl adducts. DHAA also reacted with (-)-epicatechin (EC), (-)-epicatechin gallate (ECG), and (-)-epigallocatechin (EGC) to form ascorbyl adducts, which destabilized them. After 9 h of reaction with DHAA, the depletion rates of EGCG, ECG, EC, and EGC were 30.08%, 22.78%, 21.45%, and 13.55%, respectively. The ability of DHAA to promote catechins depletion went from high to low: EGCG, ECG, EGC, and EC. The results are important for the processing and storage of tea and tea beverages, as well as the general exploration of synergistic functions of AA and catechins.
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•One pot co-precipitation synthesis method for DHA coated Fe3O4 nanoparticles under atmospheric air is reported.•An antioxidant ascorbic (AA) acid is used to cease the oxidative ...impact of air.•DHA coating stabilizes the Fe3O4 nanoparticles and effectively enhances their colloidal dispersibility in water.•DHA coated Fe3O4 nanoparticles show a negative MRI contrast nature (T2 and T2∗) and human blood cell compatibility.
Water dispersible and biologically important molecule dehydroascorbic acid (DHA, capable to cross the blood brain barrier) coated Fe3O4 superparamagnetic nanoparticles having an average size of ∼6nm were synthesized through one pot aqueous coprecipitation method under atmospheric air. An antioxidant ascorbic acid (AA) used in the synthesis oxidized itself to dehydroascorbic acid (DHA) to consume dissolved or available oxygen in reaction mixture which died away the oxidative impact of atmospheric air and formed DHA encapsulated the Fe3O4 nanoparticles which stabilized the Fe3O4 nanoparticles and significantly enhanced their colloidal solubility in water. Fe3O4 phase, superparamagnetic property, DHA coating and stable colloidal solubility in water were confirmed by means of XPS, VSM, IR and zeta potential analysis respectively. T1, T2 and T2∗ weighted magnetic resonance imaging (MRI) and corresponding relaxivity (r1=0.416, r2=50.28 and r2∗=123.65mM−1 and r2/r1=120.86, r2∗r1=297.23) of colloidally dispersed DHA-coated nanoparticle water phantom revealed a strong contrast enhancement in T2 and T2∗ weighted images. The compatibility of DHA-coated Fe3O4 nanoparticles toward human blood cells was examined by means of cell counting and cell morphological analysis with the use of optical microscope and scanning electron microscope imaging.
For a long time, the effect of vitamin C on cancer cells has been a controversial concept. From Linus Pauling's studies in 1976, it was proposed that ascorbic acid (AA) could selectively kill tumor ...cells. However, further research suggested that vitamin C has no effect on tumor survival. In the last decade, new and emerging functions for vitamin C have been discovered using the reduced form, AA, and the oxidized form, dehydroascorbic acid (DHA), independently. In this review, we summarized the latest findings related to the effects of DHA on the survival and metabolism of tumor cells. At the same time, we put special emphasis on the bystander effect and the recycling capacity of vitamin C in various cellular models, and how these concepts can affect the experimentation with vitamin C and its therapeutic application in the treatment against cancer.
We summarized the latest findings related to the effects of dehydroascorbic acid (DHA) on the survival and metabolism of tumor cells. DHA alters glucose metabolism and induces cell death in normal and tumor cells. The complex relationship between ascorbic acid and DHA will continue to be a subject of controversy with respect to vitamin C use as an antitumor drug.
A validated silver nanoparticle assay (SNaP-C) for quantitation of Vitamin C, as ascorbic acid (AA) and total AA (TAA), was applied to 31 beverages. SNaP-C assay results (LOD of 2.2 mg/L AA) were ...compared to AA and TAA determined by high-performance liquid chromatography with UV/Vis (LOD = 0.4 mg/L AA), and two well-known assays. All approaches were calibrated using meta-phosphoric acid stabilized AA, where the reducing agent tris(2-carboxyethyl) phosphine hydrochloride was added to convert dehydroascorbic acid to AA for determination of TAA. Statistical comparisons of these four resulting datasets were completed. SNaP-C and HPLC were not statistically significantly different (P > 0.05) for comparison of AA and TAA (mg/L) in these samples, whereas the CUPRAC and Folin-Ciocalteu assays statistically significantly overestimated values of AA and TAA content, respectively. The SNaP-C method is a novel assay that has high specificity for AA capable of quantifying TAA with addition of TCEP.
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•Ascorbic acid is selectively quantified using silver nanoparticles (SNaP-C).•SNaP-C assay results were compared to HPLC, CUPRAC, and Folin-Ciocalteu.•SNaP-C assay results were statistically comparable to HPLC results.•Low LOD and LOQ were observed using SNaP-C for AA and DHAA.
Rationale
Oxidative stress is an imbalance between reactive free radical oxygen species and antioxidant defenses. Its consequences can lead to numerous pathologies. Regulating oxidative stress is the ...complex interplay between antioxidant recycling and thiol‐containing regulatory proteins. Understanding these regulatory mechanisms is important for preventing onset of oxidative stress. The aim of this study was to investigae S‐thiol protein chemistry associated with oxidized vitamin C (dehydroascorbate, DHA), homocysteine (HcySH) and glutathione (GSH) using mass spectrometry.
Methods
Glutaredoxin‐1 (Grx‐1) was incubated with DHA, with and without GSH and HcySH. Disulfide formation was followed by electrospray ionization mass spectrometry (ESI‐MS) of intact proteins and by LC/ESI‐MS/MS of peptides from protein tryptic digestions. The mechanism of DHA‐mediated S‐thiolation was investigated using two synthetic peptides: AcFHACAAK and AcFHACE. Three proteins, i.e. human hemoglobin (HHb), recombinant peroxiredoxin 2 (Prdx2) and Grx‐1, were S‐homocysteinylated followed by S‐transthiolyation with GSH and investigated by ESI‐MS and ESI‐MS/MS.
Results
ESI‐MS analysis reveals that DHA mediates disulfide formation and S‐thiolation by HcySH as well as GSH of Grx‐1. LC/ESI‐MS/MS analysis allows identification of Grx‐1 S‐thiolated cysteine adducts. The mechanism by which DHA mediates S‐thiolation of heptapeptide AcFHACAAK is shown to be via initial formation of a thiohemiketal adduct. In addition, ESI‐MS of intact proteins shows that GSH can S‐transthiolate S‐homocysteinylated Grx‐1_ HHb and Prdx2. The GS‐S‐protein adducts over time dominate the ESI‐MS spectrum profile.
Conclusions
Mass spectrometry is a unique analytical technique for probing complex reaction mechanisms associated with oxidative stress. Using model proteins, ESI‐MS reveals the mechanism of DHA‐facilitated S‐thiolation, which consists of thiohemiketal formation, disulfide formation or S‐thiolation. Furthermore, protein S‐thiolation by HcySH can be reversed by reversible GSH thiol exchange. The use of mass spectrometry with in vitro models of protein S‐thiolation in oxidative stress may provide significant insight into possible mechanisms of action occurring in vivo.
Intestinal vitamin C (Asc) absorption was believed to be mediated by the Na+-dependent ascorbic acid transporter SVCT1. However, Asc transport across the intestines of SVCT1 knock-out mice is normal ...indicating that alternative ascorbic acid transport mechanisms exist. To investigate these mechanisms, rodents were gavaged with Asc or its oxidized form dehydroascorbic acid (DHA), and plasma Asc concentrations were measured. Asc concentrations doubled following DHA but not Asc gavage. We hypothesized that the transporters responsible were facilitated glucose transporters (GLUTs). Using Xenopus oocyte expression, we investigated whether facilitative glucose transporters GLUT2 and GLUT5–12 transported DHA. Only GLUT2 and GLUT8, known to be expressed in intestines, transported DHA with apparent transport affinities (Km) of 2.33 and 3.23 mm and maximal transport rates (Vmax) of 25.9 and 10.1 pmol/min/oocyte, respectively. Maximal rates for DHA transport mediated by GLUT2 and GLUT8 in oocytes were lower than maximal rates for 2-deoxy-d-glucose (Vmax of 224 and 32 pmol/min/oocyte for GLUT2 and GLUT8, respectively) and fructose (Vmax of 406 and 116 pmol/min/oocyte for GLUT2 and GLUT8, respectively). These findings may be explained by differences in the exofacial binding of substrates, as shown by inhibition studies with ethylidine glucose. DHA transport activity in GLUT2- and GLUT8-expressing oocytes was inhibited by glucose, fructose, and by the flavonoids phloretin and quercetin. These studies indicate intestinal DHA transport may be mediated by the facilitative sugar transporters GLUT2 and GLUT8. Furthermore, dietary sugars and flavonoids in fruits and vegetables may modulate Asc bioavailability via inhibition of small intestinal GLUT2 and GLUT8.
Background: The molecular identity of the intestinal vitamin C transporters is incomplete.
Results: Facilitative sugar transporters, GLUT2 and GLUT8, transport dehydroascorbic acid, the oxidized form of vitamin C.
Conclusion: Intestinal vitamin C absorption can occur via facilitative sugar transporters.
Significance: Vitamin C bioavailability may be inhibited by dietary factors, such as glucose and phytochemicals.
During vitamin C catabolism, H2O2 non-enzymically oxidised dehydroascorbic acid to yield cyclic-oxalyl-threonate and, simultaneously, an oxalyl-threonate (OxT) pool composed of an interconverting ...mixture of 3-OxT and 4-OxT. Other oxidation products were partially characterised. Display omitted
► Catabolism of vitamin C non-enzymically under conditions mimicking the apoplast. ► Dehydroascorbic acid+H2O2 yield two interconvertible oxalyl-threonate (OxT) isomers. ► 3-OxT and 4-OxT differ in acidity and are separable by paper electrophoresis. ► Diketogulonate+H2O2 yield two partially characterised acidic products. ► Possible signalling or defence roles of these metabolites await exploration.
The rate of l-ascorbate catabolism in plants often correlates positively with the rate of cell expansion. The reason for this correlation is difficult to explore because of our incomplete knowledge of ascorbate catabolism pathways. These involve enzymic and/or non-enzymic oxidation to dehydroascorbic acid (DHA), which may then be hydrolysed to 2,3-diketogulonate (DKG). Both DHA and DKG were susceptible to further oxidation under conditions of pH and H2O2 concentration comparable with the plant apoplast. The kinetics of their oxidation and the identity of some of the products have been investigated here. DHA, whether added in pure form or generated in situ by ascorbate oxidation, was oxidised non-enzymically to yield, almost simultaneously, a monoanion (cyclic-oxalyl-threonate; cOxT) and a dianion (oxalyl-threonate; OxT). The monoanion was resistant to periodate oxidation, showing that it was not oxalic threonic anhydride. The OxT population was shown to be an interconverting mixture of 3-OxT and 4-OxT, differing in pKa. The 3-OxT appeared to be formed earlier than 4-OxT, but the latter predominated at equilibrium. DKG was oxidised by H2O2 to two partially characterised products, one of which was itself further oxidised by H2O2 to yield threonate. The possible occurrence of these reactions in the apoplast in vivo and the biological roles of vitamin C catabolites are discussed.
Although l-ascorbate (vitamin C) is an important biological antioxidant, its degradation pathways in vivo remain incompletely characterised. Ascorbate is oxidised to dehydroascorbic acid, which can ...be either hydrolysed to diketogulonate (DKG) or further oxidised. DKG can be further degraded, oxidatively or non-oxidatively. Here we characterise DKG products formed non-enzymically and non-oxidatively at 20 °C and at a slightly acidic pH typical of the plant apoplast.
High-voltage electrophoresis revealed at least five products, including two novel CPLs (epimers of 2-carboxy-l-threo-pentonolactone), which slowly interconverted with CPA (2-carboxy-l-threo-pentonate). One of the two CPLs has an exceptionally low pKa. The CPL structures were supported by MS (C6H7O7)– and by 1H and 13C NMR spectroscopy. Xylonate and its lactone also appeared.
Experiments with 1–14CDKG showed that all five products (including the 5-carbon xylonate and its lactone) retained DKG's carbon-1; therefore, most xylonate arose by decarboxylation of CPLs or CPA, one of whose –COOH groups originates from C-2 or C-3 of DKG after a ‘benzilic acid rearrangement’. Since CPLs appeared before CPA, a DKG lactone is probably the main species undergoing this rearrangement.
CPA and CPL also form non-enzymically in vivo, where they may be useful to researchers as ‘fingerprints’, or to organisms as ‘signals’, indicating a non-oxidative, slightly acidic biological compartment.
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•Vitamin C is degraded to diketogulonate, whose own breakdown products are enigmatic.•Here we characterise 5 such products, formed non-oxidatively at slightly acidic pH.•Novel products include two epimers of 2-carboxy-l-threo-pentonolactone (CPL).•CPLs and their free acid (CPA) are decarboxylated to xylonate and/or lyxonate.•CPLs and CPA are ‘fingerprints’ and signals, reporting a cell's redox and pH status.
Vitamin C (Ascorbic Acid), the antiscorbutic vitamin, cannot be synthesized by humans and other primates, and has to be obtained from diet. Ascorbic acid is an electron donor and acts as a cofactor ...for fifteen mammalian enzymes. Two sodium‐dependent transporters are specific for ascorbic acid, and its oxidation product dehydroascorbic acid is transported by glucose transporters. Ascorbic acid is differentially accumulated by most tissues and body fluids. Plasma and tissue vitamin C concentrations are dependent on amount consumed, bioavailability, renal excretion, and utilization. To be biologically meaningful or to be clinically relevant, in vitro and in vivo studies of vitamin C actions have to take into account physiologic concentrations of the vitamin. In this paper, we review vitamin C physiology; the many phenomena involving vitamin C where new knowledge has accrued or where understanding remains limited; raise questions about the vitamin that remain to be answered; and explore lines of investigations that are likely to be fruitful.
Monocopper lytic polysaccharide monooxygenases (LPMOs) catalyse oxidative cleavage of glycosidic bonds in a reductant‐dependent reaction. Recent studies indicate that LPMOs, rather than being ...O2‐dependent monooxygenases, are H2O2‐dependent peroxygenases. Here, we describe SscLPMO10B, a novel LPMO from the phytopathogenic bacterium Streptomyces scabies and address links between this enzyme’s catalytic rate and in situ hydrogen peroxide production in the presence of ascorbic acid, gallic acid and l‐cysteine. Studies of Avicel degradation showed a clear correlation between the catalytic rate of SscLPMO10B and the rate of H2O2 generation in the reaction mixture. We also assessed the impact of oxidised ascorbic acid, dehydroascorbic acid (DHA), on LPMO activity, since DHA, which is not considered a reductant, was recently reported to drive LPMO reactions. Kinetic studies, combined with NMR analysis, showed that DHA is unstable and converts into multiple derivatives, some of which are redox active and can fuel the LPMO reaction by reducing the active site copper and promoting H2O2 production. These results show that the apparent monooxygenase activity observed in SscLPMO10B reactions without exogenously added H2O2 reflects a peroxygenase reaction.
Lytic polysaccharide monooxygenases (LPMOs) catalyze oxidation of glycosidic bonds in a reductant‐dependent reaction. While it is well known that the nature of the reductant affects LPMO activity, the basis for this connection has remained unclear. Here, we address this issue in detail and investigate LPMO activation by various compounds, including dehydroacrobic acid, which is not considered a reducant but nevertheless fuels LPMO catalysis.
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