PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “Dithionite”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1995

Dithionite increases radical formation and decreases vasoconstriction in the lung. Evidence that dithionite does not mimic alveolar hypoxia.

Dithionite is a powerful reducing agent used to deoxygenate hemoglobin and create anaerobic conditions in vitro. Recently, dithionite has been used as a convenient means of creating "hypoxia" in experiments studying the O2 sensor in the pulmonary circulation and carotid body. We evaluated the hypothesis that hypoxia created by hypoxic ventilation and that created by dithionite have different effects on the pulmonary circulation. In vitro, dithionite (10(-5) to 10(-3) mol/L), added to oxygenated Krebs' solution, rapidly created superoxide anion in a dose-dependent manner. Dithionite consumed O2 in parallel with the generation of superoxide radical, with both processes peaking within seconds. Anoxia was sustained only if resupply of O2 was prevented. In isolated rat lungs (whether perfused with autologous blood or Krebs' solution), hypoxic ventilation alone lowered perfusate PO2 from approximately 140 to 40 mm Hg and decreased lung levels of activated oxygen species (AOS), measured by luminol-enhanced chemiluminescence, before the onset of hypoxic pulmonary vasoconstriction. Constrictor responses to angiotensin II and KCl were not impaired by intermittent hypoxic challenges, and lung weight did not increase. In contrast, dithionite impaired constrictor responses of the Krebs' solution-perfused lungs to all vasoconstrictors tested and increased lung weight. When given as a bolus (5 x 10(-3) mol/L) into the pulmonary artery during normoxic ventilation, dithionite caused no vasoconstriction and only briefly lowered PO2 (because of constant resupply of O2 from the alveoli). When superimposed on hypoxic ventilation, dithionite further lowered PO2 from approximately 40 to approximately 0 mm Hg and caused additional constriction. Unlike hypoxic ventilation, dithionite increased AOS production. Antioxidant enzymes diminished dithionite-induced radical production and diminished the loss of vascular reactivity and lung edema. In conclusion, unlike hypoxic ventilation, dithionite causes edema and loss of vascular reactivity in the lung by generating superoxide anion and hydrogen peroxide. Hypoxia elicited by dithionite is not equivalent to authentic hypoxia because of the obligatory associated generation of AOS. Dithionite usage should not be substituted for authentic hypoxia in studies of O2 sensing.

Animals↗
PubMed1991

A method for preparing analytically pure sodium dithionite. Dithionite quality and observed nitrogenase-specific activities.

Sodium dithionite (Na2S2O4) is widely used as a reductant in biochemical studies, but has not been available in its pure form. A convenient, detailed procedure is given for the recrystallization of commercial dithionite from 0.1 M NaOH-methanol under anaerobic conditions. Twice-recrystallized dithionite had a purity of 99 +/- 1% by UV spectroscopy (A315) and elemental analysis. The influence of dithionite quality on the apparent reduction activities of the nitrogenase components (Av1 and Av2) from Azotobacter vinelandii was investigated.

Crystallization↗
PubMed1978

The redox potential of dithionite and SO-2 from equilibrium reactions with flavodoxins, methyl viologen and hydrogen plus hydrogenase.

1. It has been shown that redox equilibria can be formed between dithionite ion (plus SO-2) and (bi)sulphite, and the low-potential electron carriers flavodoxin and methyl viologen. The equilibria were established either by treating the oxidized electron carriers with dithionite, or by treating flavodoxin hydroquinone or methyl viologen semiquinone with (bi)sulphite. Similar redox equilibria were established between dithionite/(bi)sulphite and hydrogen using catalytic amounts of hydrogenase in the presence of a low-potential electron carrier. The effects of pH and temperature on the equilibria were determined. 2. The equilibria were analyzed to determine the redox potential of the dithionite/(bi)sulphite system. In accordance with the results of earlier kinetic studies, it was assumed that the reductant in dithionite solutions is the dissociation product SO-2. The calculated midpoint redox potential E' for the couple SO-2/HSO-3 at pH 7 and 25 degrees C was -0.66 V. The reductant is present largely as the dimer at concentrations of dithionite above about 10nM. Consequently, the midpoint potential, Em, of dithionite solutions becomes less negative as the concentration of dithionite is increased (deltaEm/deltalog S2O2-4 = 29 mV). The theoretical potential of a solution of 1 M S2O2-4 and 2 M (bi)sulphite at pH 7 was calculated to be -0.386V. This value is 59 mV more negative than that determined in 1911 by potentiometry, but considerably more positive than other values in the literature. The effects of pH on the equilibria showed that E' is controlled by the pK of (bi)sulphite at 6.9; the slope deltaE'/deltapH was -59 mV below the pK and -118 mV above the pK. The effects of temperature on the equilibria suggested that Em for dithionite changed by -1.6 mV/degrees C for a rise in temperature between 2 degrees C and 40 degrees C. If sodium dithionite is contaminated with small amounts of (bi)sulphite, its addition in large excess to a low potential electron carrier can cause oxidation of the carrier.

Anaerobiosis↗
PubMed1981

Tetramer-dimer dissociation of carboxyhemoglobin in the absence of dithionite.

The generally accepted value for the tetramer-dimer dissociation constant KL4,2 of carboxyhemoglobin in pH 7.0 phosphate buffer lies in the range 1--2 micrometers. Previous determinations of the quantity have generally involved addition of dithionite to samples to exclude oxygen. We report flash photolysis experiments on carboxyhemoglobin in the absence of dithionite which suggest that KL4,2 is 0.2 +/- 0.05 micrometer. Addition of dithionite to our samples resulted in an order of magnitude increase in KL4,2 in good agreement with previously published results. The mechanism of this increase in dissociation has not been determined with certainty. However, impurities, possibly metal ions, are required in addition to dithionite to produce this effect. Dithionite did not increase KL4,2 for phosphate buffer solutions treated with Chelex 100 analytical grade chelating resin. Addition of bovine serum albumin to untreated buffer solutions before addition of dithionite was found to prevent increased dissociation. The sulfhydryl-reducing agents dithiothreitol and beta-mercaptoethanol were found to protect against the effect of dithionite and to reverse its effect on KL4,2 if they were added after the dithionite. The interaction of the unknown impurities with dithionite to produce increased values of KL4,2 could be mimicked by addition of CU2+ ions in concentrations of less than 1 micrometer to buffer treated with Chelex 100 resin.

Carboxyhemoglobin↗
PubMed1992

The crystal state binding of dithionite to deoxy-hemoglobin.

The crystal state binding of sodium dithionite to deoxyhemoglobin is reported. Dithionite has been used extensively to deoxygenate hemoglobin and myoglobin and there has been considerable interest among users of dithionite about its effect on protein structure and binding site(s). We have determined that dithionite binds to deoxygenated hemoglobin crystals at the interface of two molecules in the crystal lattice. Specific residues involved in hydrogen bonds or salt interactions with dithionite include His116 and His117 of the beta 2 subunit and Lys16 of the alpha 1 subunit of the adjacent hemoglobin molecule. No binding was observed at the symmetry related His116 and 117 beta 1 residues. We have shown that dithionite does not affect the native hemoglobin structure or the binding of several allosteric inhibitors to hemoglobin and can be used to mount T state crystals in the air.

Binding Sites↗
PubMed1975

Reduced diphosphopyridine nucleotide peroxidase. Intermediates formed on reduction of the enzyme with dithionite or reduced diphosphopyridine nucleotide.

DPNH peroxidase is a flavin adenine dinucleotide-containing flavoprotein. Anaerobic titration of enzyme with dithionite has shown that the active site of the enzyme contains 2 mol of flavin and in addition 1 mol of a non-flavin electron acceptor that is tentatively identified as a disulfide group. Thus complete reduction of the enzyme requires 3 mol of dithionite per mole of active site. The first mole of dithionite reduces the non-flavin acceptor; complex formation between the reduced acceptor and one of the bound flavin molecules causes the formation of a long wavelength absorption band between 500 and 670 nm. The second mole of dithionite reduces the flavin that interacts with the reduced non-flavin group, and the long wavelength band disappears. The third mole of dithionite reduces the second mole of flavin. All groups are reoxidized in the presence of air. DPNH reacts with only two of the enzyme-bound electron acceptors. The first mole of DPNH reduces the non-flavin group to form an intermediate (I) that is almost identical with that formed by dithionite. The second mole of DPNH complexes with the second flavin of Intermediate I to form Intermediate II. This reaction causes a further absorbance increase in the long wavelength region; the tail of the absorption band now extends to 960 nm. The titration data (potassium phosphate, 0.05 M, pH 7.0) can be fitted with dissociation constants of 1 times 10-7 M for the formation of I, and 3 times 10-6 M for the conversion of I to II. In air, species II is oxidized to I; I is stable in air, but is oxidized stoichiometrically to oxidized enzyme by H2O2. Present evidence suggests that bound DPN-plus is responsible for the air stability of species I. Intermediate I, but not oxidized enzyme, reacts slowly with phenylmercuric acetate. This reaction causes loss of the air-stable intermediate and parallel loss in enzyme activity. The inactive enzyme cannot be reduced by DPNH to Species I; DPNH can, however, still react with the second flavin to form the autoxidizable complex. With other methods of enzyme inactivation there is also a direct correlation between residual enzyme activity and the ability of enzyme to form the air-stable intermediate. It is concluded that the air-stable intermediate is an important catalytic species.

Anaerobiosis↗
PubMed1980

Fluorescence emission spectra of cells and subcellular preparations of a green photosynthetic bacterium. Effects of dithionite on the intensity of the emission bands.

Fluorescence emission spectra were measured of intact cells and subcellular preparations of the green photosynthetic bacterium Prosthecochloris aestuarii in the presence and in the absence of dithionite. A 3--5-fold increase in bacteriochlorophyll a fluorescence at 816 nm occurred upon addition of dithionite in a membrane vesicle preparation (Complex I), in a photochemically active pigment-protein complex and in a bacteriochlorophyll a protein complex free from reaction centers. The pigment-protein complex showed a relatively strong long-wave emission band (835 nm) of bacteriochlorophyll a, which was preferentially excited by light absorbed at 670 nm and was not stimulated by dithionite. With Complex I, which contains some bacteriochlorophyll c in addition to bacteriochlorophyll a, a 3--4-fold stimulation of bacteriochlorophyll c emission was also observed. Emission bands at shorter wavelengths, probably due to artefacts, were quenched by dithionite. With intact cells, the effect of dithionite was smaller, and consisted mainly of an increase of bacteriochlorophyll a emission. The results indicate that the strong increase in the yield of bacteriochlorophyll emission that occurred upon generating reducing conditions is, at least mainly, due to a direct effect on the light-harvesting systems, and does not involve the reaction center as had been earlier postulated.

Bacteriochlorophylls↗
PubMed1985

Kinetics of reduction of cytochrome c oxidase by dithionite and the effect of hydrogen peroxide.

The reduction of cytochrome c oxidase by dithionite was reinvestigated with a flow-flash technique and with varied enzyme preparations. Since cytochrome a3 may be defined as the heme in oxidase which can form a photolabile CO adduct in the reduced state, it is possible to follow the time course of cytochrome a3 reduction by monitoring the onset of photosensitivity. The onset of photosensitivity and the overall rate of heme reduction were compared for Yonetani and Hartzell-Beinert preparations of cytochrome c oxidase and for the enzyme isolated from blue marlin and hammerhead shark. For all of these preparations the faster phase of heme reduction, which is dithionite concentration-dependent, is almost completed when the fraction of photosensitive material is still small. We conclude that cytochrome a3 in the resting enzyme is consistently reduced by an intramolecular electron transfer mechanism. To determine if this is true also for the pulsed enzyme, we examined the time course of dithionite reduction of the peroxide complex of the pulsed enzyme. It has been previously shown that pulsed cytochrome c oxidase can interact with H2O2 and form a stable room temperature peroxide adduct (Bickar, D., Bonaventura, J., and Bonaventura, C. (1982) Biochemistry 21, 2661-2666). Rather complex kinetics of heme reduction are observed when dithionite is added to enzyme preparations that contain H2O2. The time courses observed provide unequivocal evidence that H2O2 can, under these conditions, be used by cytochrome c oxidase as an electron acceptor. Experiments carried out in the presence of CO show that a direct dithionite reduction of cytochrome a3 in the peroxide complex of the pulsed enzyme does not occur.

Animals↗
PubMed1992

Evidence of dithionite contribution to the low-frequency resonance Raman spectrum of reduced and mixed-valence cytochrome c oxidase.

The resonance Raman spectra of deoxygenated solutions of mixed-valence cyanide-bound and fully reduced cytochrome oxidase derivatives that have been reduced in the presence of aqueous or solid sodium dithionite exhibit two new low-frequency lines centered at 474 and 590 cm-1. These lines were not observed when the reductant system was changed to a solution containing ascorbate and N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD). Under enzyme turnover conditions, the addition of dithionite to the reoxidized protein (the 428-nm or "oxygenated" form) increases the intensity of these lines, while reoxidation and rereduction of the enzyme in the presence of ascorbate/TMPD resulted in the absence of both lines. Our data suggest that both lines must have contributions from species formed from aqueous dithionite, presumably the SO2 species, since these two lines are also observed in the Raman spectrum of a solution of aqueous dithionite, but not in the spectrum of an ascorbate/TMPD solution. Since heme metal-ligand stretch vibrations are expected to appear in the low-frequency region from 215 to 670 cm-1, our results indicate that special care should be exercised during the interpretation of the cytochrome a3 resonance Raman spectrum.

Animals↗
PubMed1986

Kinetic studies of the reduction of neutrophil cytochrome b-558 by dithionite.

The reduction with dithionite of neutrophil cytochrome b-558, implicated in superoxide generation by activated neutrophils, was investigated by a stopped-flow technique in non-ionic-detergent extracts of the membranes and in crude membrane particles. The dependence of the pseudo-first-order rate constants on the concentration of dithionite was consistent with a mechanism of reduction that involves the dithionite anion monomer SO2.- as the reactive species. The estimated second-order rate constant was 7.8 X 10(6) M-1 X S-1 for Lubrol PX-solubilized cytochrome b-558 and 5.1 X 10(6) M-1 X S-1 for the membrane-bound protein. The similarity of the kinetic constants suggests that solubilization did not introduce gross changes in the reactive site. Imidazole and p-chloromercuribenzoate, known as inhibitors of NADPH oxidase, did not affect significantly cytochrome b-558 reduction rates. The reaction rate of cytochrome b-558 with dithionite exhibited a near-zero activation energy. The first-order rate constant for reduction decreased with increasing ionic strength, indicating a positive effective charge on the reacting protein.

Chloromercuribenzoates↗
PubMed1990

Dithionite treatment of flavins: spectral evidence for covalent adduct formation and effect on in vitro bacterial bioluminescence.

Intrigued by the apparent requirement of dithionite for FMN reduction (as opposed to photoreduction or catalytic hydrogenation) in the H2O2-initiated bacterial bioluminescence reaction, we chose 5-ethyl-3-methyllumiflavinium cation I as a model to investigate possible flavin adduct formation by treatment with dithionite or (bi)sulfite. In the range of pH 5-8, the reaction of dithionite with 5-ethyl-3-methyllumiflavinium cation, which is in equilibrium with the 5-ethyl-4a-hydroxy-3-methyl-4a, 5-dihydrolumiflavin pseudobase II (X = OH), is not limited to the formation of flavosemiquinone and dihydroflavin following two one-electron steps. Several parallel and sequential reactions may take place involving the intermediacy of covalent flavin adducts. Addition of (bi)sulfite gave a 4a-sulfiteflavin adduct II (X = SO3-). Consistent with the S2O4(2-) in equilibrium with 2 SO2-. equilibrium, the reaction of dithionite and II (X = OH; SO3-) gave rise to two flavin adducts in competitive nucleophilic displacements: a 4a-sulfoxylate-flavin radical (II, X = SO2.) and a 4a-dithioniteflavin adduct (II, X = S2O4-), respectively. On increasing the (S2O4(2-), SO2.-)/flavin ratio under N2, the formation of the 4a-sulfoxylate-flavin radical became predominant. The II (X = SO2.) so formed was in equilibrium with the flavosemiquinone and bisulfate and can be trapped by reacting with hydroxylamine. In the initial presence of oxygen, II (X = SO2.) was highly reactive toward O2, giving a fast oxidation to II (X = SO3-) and effectively suppressing the formation of the flavosemiquinone.(ABSTRACT TRUNCATED AT 250 WORDS)

Dithionite↗
PubMed1983

Inhibition by dithionite and reactivation by iron of the tartrate-resistant acid phosphatase in bone of osteopetrotic (ia) rats.

The staining intensity and inhibitor sensitivity of acid phosphatase activity was determined histochemically in various tissues of normal and ia rat pups by the use of freeze-dried whole body sections. Activity was determined using alpha-naphthylphosphate as substrate and hexazonium pararosaniline as coupler. Sections from ia rats (6 and 24 days old) showed markedly higher enzyme activity in bone than sections from normal littermates. However, there were no differences between ia and normal pups in acid phosphatase activity in soft tissues and developing teeth. Preincubation of sections with 1-100 mM sodium dithionite (an iron-binding agent) caused a dose-related inhibition of enzyme activity in bone of ia and normal pups, but only slight inhibition of activity in soft tissues. Partial restoration of the dithionite-inhibited activity in bone was achieved by subsequent preincubation in 1 mM FeCl2. Addition of 100 mM sodium tartrate to the staining solution of non-preincubated sections caused almost complete inhibition of activity in soft tissues and the developing teeth but no inhibition of the activity in bone that was sensitive to sodium dithionite. These data indicate a) that sodium dithionite can be used as a specific histochemical inhibitor of the tartrate-resistant acid phosphatase and b) that the source of increased acid phosphatase activity in bone from ia rats is mostly from the tartrate-resistant acid phosphatase.

Acid Phosphatase↗
PubMed1984

Chemically coupled spectrophotometric assays based on flow injection analysis: determination of nitrogenase by assays for creatine, ammonia, hydrazine, phosphate, and dithionite.

Micromethods of direct chemical coupling have been developed for several different enzyme reactions, using the principles of flow injection analysis. Samples of 1-25 microliters are injected into a flowing stream of color-forming reagents and the peak of color change is measured after about 1 min. Alternatively, continuous slow infusion of a reacting system (5-100 microliters/min) gives a continuous change of color which can be monitored to derive enzyme reaction rates. These techniques are highly sensitive, requiring a few nanomoles of the substance being detected. Phosphate, ammonia, dithionite, creatine, and hydrazine have been measured. Consumption of reagents is less than 75 ml per hour; typical sample throughout is 30-40 samples per hour by the injection method, and 5 samples per hour by continuous infusion. The procedure has been applied to nitrogenase, continuously monitoring creatine produced from creatine phosphate by creatine kinase which is used to supply a constant level of ATP for nitrogenase. In this way nitrogenase activity can be determined over a wide range of enzyme concentrations. Production of inorganic phosphate directly from ATP, by injection of formaldehyde-quenched samples, was used when coupling to creatine kinase was not possible. Both injection of aliquots and continuous infusion were used for detection of hydrazine during nitrogenase reduction of azide, and the injection method has been used for ammonia assay during dinitrogen reduction. Dithionite oxidation was measured directly from decolorization of iodine, after trapping both dithionite and bisulfite with formaldehyde.

Ammonia↗
PubMed1988

Mechanism of reductive activation of a 5-nitroimidazole by flavoproteins: model studies with dithionite.

The flavoprotein nitroreductases NADPH:cytochrome P-450 reductase and xanthine oxidase catalyzed the cofactor-dependent anaerobic nitro group reduction and covalent binding to protein sulfhydryl groups of the 5-nitroimidazole substrate ronidazole [1-methyl-5-nitroimidazole-2-yl)-methyl carbamate). Studies with variously radiolabeled ronidazole molecules demonstrated that the imidazole ring was intact while greater than 80% of the C-4 3H and 2-carbamoyl group were lost from the covalently bound product. The stoichiometry of cofactor consumption during the enzyme-catalyzed reduction of the substrate could not be determined, so a model nitroreductase system which utilized dithionite as the reductant and agarose-immobilized cysteine as the target for alkylation was developed. Two moles of dithionite was consumed per mole of substrate for maximal reduction of uv absorbance due to the nitro group, for maximal release of C-4 3H, and for maximal covalent binding to agarose-immobilized cysteine. These results indicate that four electrons are required for the reductive activation of the substrate, consistent with formation of a hydroxylamine reactive intermediate. Covalent binding of variously radiolabeled substrate molecules after dithionite reduction exhibited the same labeling pattern as flavoprotein-catalyzed covalent binding, suggesting that covalent binding is mediated by the same species in both chemical and biological systems. The data are consistent with a mechanism where the substrate undergoes four-electron reduction to form a hydroxylamine, which is susceptible to nucleophilic attack at C-4. When water attacks C-4, the 2-carbamoyl group can eliminate to form a Michael-like acceptor which adds thiols at the 2-methylene position.

Animals↗
PubMed1990

Formation and decay of cytochrome c peroxidase compound ES during aerobic reduction with dithionite.

Stopped-flow and rapid scanning studies have clearly demonstrated that mixing of an oxygen-saturated solution of yeast cytochrome c peroxidase with sodium dithionite yields compound ES, indicating generation of H2O2. The formation of compound ES was most pronounced when [Na2S2O4]/[O2] approximately 1, and it reverted to the ferric form while standing. Even in the presence of an excess of dithionite ([Na2S2O4]/[O2] = 3.4) compound ES was formed immediately, but was soon replaced by the ferric form, followed by its final reduction to the ferrous state. The apparent first order rate constant for the decay of compound ES to the ferric form increased linearly with the square root of the dithionite concentration, thus involvement of SO2- in that process being suggested.

Aerobiosis↗
PubMed1985

Calorimetric studies of oxyhemoglobin dissociation. II. Erythrocytic oxygen depletion by sodium dithionite.

Dithionite causes the depletion of dioxygen from suspensions of erythrocytes by reduction of the external dioxygen and not by diffusion into the cell. The molar enthalpy for the reduction shows a small difference with respect to the values found for free hemoglobin; and the normal stoichiometry of 2 moles dithionite/mole dioxygen found there is not observed with erythrocytes. At low hematocrit, the stoichiometry is 2.6:1 and decreases to 1.5:1 at high hematocrit. The change is not due to differences in the hemoglobin saturation or to an inability of dithionite to reduce all dioxygen present at the higher hematocrit. Neither catalase nor peroxidase added to the extracellular volume significantly alters the stoichiometry or the enthalpy of dioxygen reduction by dithionite. Addition of superoxide dismutase, however, restores the normal stoichiometry at high hematocrit and further increases the stoichiometry at low hematocrit. The calorimetrical signal of hydrogen peroxide, clearly seen with free dioxygen, is not present with erythrocytes. In all these cases the total heat evolved is the same.

Animals↗
PubMed1985

Kinetics of dithionite reduction of the heme nonapeptide of cytochrome c.

The kinetics of dithionite reduction of the oxidized heme nonapeptide fragment of horse heart cytochrome c have been measured as a function of ionic strength at pH 7 and pH 9 by the stopped-flow technique. Dithionite concentration dependences indicate that the radical anion monomer, SO2-., is the active reductant. The pH 7 ionic strength dependence suggests that the heme peptide is reacting as a negatively charged molecule (its overall charge is calculated to be -1). Comparison of these results with the known rate of dithionite reduction of cytochrome c indicates that the heme nonapeptide has substantially greater inherent reactivity than cytochrome c, perhaps due to the greater accessibility of the heme.

Animals↗