fumarates and dihydroxyfumarate

In the context of a "glyoxylate scenario" of primordial metabolism, the reactions of dihydroxyfumarate (DHF) with reactive small molecule aldehydes (e.g., glyoxylate, formaldehyde, glycolaldehyde, and glyceraldehyde) in water were investigated and shown to form dihydroxyacetone, tetrulose, and the two pentuloses, with almost quantitative conversion. The practically clean and selective formation of ketoses in these reactions, with no detectable admixture of aldoses, stands in stark contrast to the formose reaction, where a complex mixture of linear and branched aldoses and ketoses are produced. These results suggest that the reaction of DHF with aldehydes could constitute a reasonable pathway for the formation of carbohydrates and allow for alternative potential prebiotic scenarios to the formose reaction to be considered.

Topics: Aldehydes; Fumarates; Glyoxylates; Ketoses

A solid base nanocatalyst was prepared by ZrO(2) loaded with C(4)H(4)O(6)HK and investigated for transesterification of soybean oil with methanol to biodiesel. The obtained nanocatalyst was characterized by means of XRD, FTIR, TEM, TGA, N(2) adsorption-desorption measurements and the Hammett indicator method. TEM photograph showed that the nanocatalyst had granular and porous structures with particle sizes of 10-40 nm. The nanocatalyst had longer lifetime and maintained sustained activity after being used for five cycles. The separate effects of the molar ratio of methanol to oil, reaction temperature, nanocatalyst amount and reaction time were investigated. The experimental results showed that a 16:1M ratio of methanol to oil, 6.0% catalyst, 60°C reaction temperature and 2.0 h reaction time gave the best results and the biodiesel yield of 98.03% was achieved. Production of biodiesel has positive impact on the utilization of agricultural and forestry products.

Topics: Biofuels; Catalysis; Fumarates; Methanol; Microscopy, Electron, Transmission; Nanoparticles; Soybean Oil; Spectroscopy, Fourier Transform Infrared; Temperature; Time Factors; X-Ray Diffraction; Zirconium

Jack of all trades: water-soluble salts of DHF underwent self-condensation to afford the threo diastereomer of pentulosonic acid, through differing reaction pathways contingent on the metal salt used. This transformation exemplifies the diverging roles of DHF as a nucleophile (a synthon for α-hydroxyacetyl anion) and an electrophile (an α-carboxyglycolaldehyde equivalent).

Topics: Carbohydrates; Dimerization; Fumarates; Metals; Salts; Stereoisomerism; Water

This work studied the possibilities for quantitative determination of iron mobilization in connection with ferritin reduction by ascorbic acid (vitamin C) and sodium dithionite in vitro. The iron storage protein was incubated with an excess of reductant in aerobic conditions in the absence of complexing agents in the medium. The release of Fe(2+) was let to go to completion, and the overall content of Fe(2+) in the solution was evaluated with the aid of potentiometric titration using Ce(4+) as an oxidizing titrant. Results suggest a moderate iron efflux under the influence of the chosen reducing agents. Although such a reduction of the protein mineral core by dihydroxyfumarate contributes greatly to the iron mobilization, ferritin behavior with vitamin C and dithionite seems to be different. Although redox properties of dihydroxyfumarate are determined by hydroxyl groups similar to those of ascorbic acid, the two compounds differ significantly in structure, and this could be the basis for an explanation of the specificities in their interaction with ferritin. As revealed by the study, potentiometric titration promises to be a reliable tool for evaluation of the amount of Fe(2+) present in the solution as a result of the reduction of the ferritin's mineral core.

Topics: Ascorbic Acid; Cerium; Dithionite; Ferritins; Fumarates; Iron; Oxidation-Reduction; Potentiometry

The kinetics of the hexacyanoferrate (III) oxidation of dihydroxyfumaric acid to hexacyanoferrate (II) and diketosuccinic acid was looked into within the 0.04 to 5.3 M HCl acidity range under different temperatures, ionic strengths, and solvent permittivity conditions. The kinetic effect of alkali metal ions, transition metal impurities, and substrate concentrations have also been analyzed. The observed inhibition effect brought about by addition of the reaction product, hexacyanoferrate (II), is a sign of a complex mechanism. The rate constants remained essentially unchanged up to 1 M HCl, diminished between 1.0 and 3.0 M HCl, and rose above 3.0 M HCl. Depending on the medium acidity, three mechanisms can be put forward, which involve different kinetically active forms. At low acidity, the rate-determining step involves a radical cation and both the neutral and the anion substrate forms are equally reactive ( k 1 = k 2 = 2.18 +/- 0.05 M (-1) s (-1), k -1 = 0.2 +/- 0.03). When the medium acidity is boosted, the rate-determining step involves the neutral dihydroxyfumaric acid and two hexacyanoferrate (III) forms. In the intermediate region the rate constant diminished with rising [H (+)] ( k' 1 = 0.141 +/- 0.01 and k' 2 = 6.80 +/- 0.05). Specific catalytic effect by binding of alkali metal ions to oxidant has not been observed. In all instances it was assessed that the substrate decomposition is slow compared to the redox reaction.

Topics: Absorption; Anions; Binding Sites; Catalysis; Cations; Chemistry, Physical; Ferrocyanides; Fumarates; Ions; Kinetics; Models, Chemical; Oxidation-Reduction; Oxygen; Temperature; X-Ray Diffraction

Oscillatory kinetics in the peroxidase-oxidase reaction catalyzed by structurally different peroxidases were investigated using NADH as a substrate. For horseradish peroxidase, lactoperoxidase, and soybean peroxidase the oscillatory waveforms of their dominating enzyme intermediates, ferric peroxidase and compound III, are similar. Coprinus peroxidase, on the other hand, has ferrous peroxidase and compound III as the dominating intermediates. The oscillatory waveform of its compound III differs from the waveforms of compound III of the three other peroxidases. Also, the phase plot of the signal for compound III versus the oxygen concentration for Coprinus peroxidase differs from the corresponding phase plots obtained using other peroxidases. A detailed model of the reaction mechanism is proposed, which is able to simulate these different kinds of behaviour. Substituting NADH with dihydroxyfumaric acid as a substrate, oscillations in the oxygen concentration were observed for about 1.5 h when a concentrated solution of this substrate was continuously fed to a solution containing horseradish peroxidase. This is the first demonstration of sustained oscillations with this substrate.

Topics: Animals; Cattle; Coprinus; Fumarates; Glycine max; Horseradish Peroxidase; In Vitro Techniques; Kinetics; Lactoperoxidase; Milk; Models, Biological; NAD; Oscillometry; Peroxidases; Substrate Specificity

1. We used electrophysiological and binding techniques to determine the effects of oxygen free radicals (OFRs) generated by dihydroxyfumaric acid (DHF, 5 mM) on calcium current and dihydropyridine binding sites in guinea-pig isolated ventricular myocytes. 2. Binding of [3H]-PN200-110 to isolated ventricular myocytes revealed one population of binding sites with a KD of 0.11 +/- 0.01 nM and Bmax of 139.1 +/- 6.9 fmol mg-1 protein (n = 24). After 15 min of exposure to DHF, the density, but not the affinity of [3H]-PN200-110 binding sites was significantly (P < 0.01) reduced to 35% of the control value (Bmax = 49.4 +/- 3.7 fmol mg-1 protein, KD = 0.11 +/- 0.01 nM, n = 15). In the presence of superoxide dismutase (SOD) and catalase (CAT) the reduction in [3H]-PN200-110 binding sites was almost completely prevented (Bmax = 120.5 +/- 7.4 in control, n = 4 and 98.8 +/- 7.4 fmol mg-1 protein in DHF plus SOD and CAT, n = 4). KD values were not modified (0.08 +/- 0.01 in control and 0.09 +/- 0.01 nM in DHF plus SOD and CAT). 3. The time-course of the reduction of [3H]-PN200-110 binding sites by OFRs was paralleled by the decrease in L-type calcium current (Ica,L) measured in patch-clamped guinea-pig ventricular myocytes either in the absence or in the presence of EGTA in the patch pipette. In the former conditions OFRs induced the appearance of calcium-dependent alterations, i.e. the transient inward current, within 10 min. After 30 min of incubation with DHF, [3H]-PN200-110 binding sites were reduced to 25% of the control value. 4. In myocytes incubated with the antilipoperoxidant agent, butylated hydroxytoluene (BHT, 50 microM), the decrease in [3H]-PN200-110 binding sites caused by DHF was partially prevented (Bmax values after 30 min exposure to DHF were 55.5 +/- 1.9 and 23.7 +/- 5.9 fmol mg-1 protein in the presence and in the absence of BHT respectively, P < 0.05). BHT did not affect the decrease in [3H]-PN200-110 binding sites during the first 15 min of exposure to DHF, but was able to prevent completely the further decrease occurring during the following 15 min of incubation with OFRs. 5. Our results demonstrate that the OFR-induced decrease in calcium current is associated with a reduction in DHP binding sites. The decrease in calcium current and in calcium channels may be implicated in the mechanical dysfunction associated with oxidative stress.

Topics: Action Potentials; Animals; Calcium; Calcium Channel Blockers; Calcium Channels; Dihydropyridines; Electrophysiology; Free Radicals; Fumarates; Guinea Pigs; Heart Ventricles; Isradipine; Myocardium; Patch-Clamp Techniques; Ventricular Function

We studied the antioxidant effects of nine calcium antagonists (nisoldipine, benidipine, nilvadipine, felodipine, nicardipine, nitrendipine, nifedipine, verapamil, and diltiazem) by means of rat myocardial membrane lipid peroxidation with a nonenzymatic active oxygen-generating system (DHF/FeCl3-ADP). The order of antioxidant potency of these agents was nilvadipine > nisoldipine > felodipine > nicardipine > verapamil > benidipine. Their IC50 values (microM) were 25.1, 28.2, 42.0, 150.0, 266.1, and 420.0, respectively. In contrast, nitrendipine, nifedipine, and diltiazem had little inhibitory effect on lipid peroxidation. These six calcium antagonists could be divided into four types on the basis of their antioxidant mechanisms. Nilvadipine, nisoldipine, and verapamil, which showed antioxidant effects both before and after the addition of active oxygen, and reduced the dihydroxyfumarate (DHF) auto-oxidation rate, were chain-breaking and preventive antioxidants. Felodipine, which showed antioxidant effects both before and after exposure to active oxygen and increased the DHF auto-oxidation rate, was only a chain-breaking antioxidant. Nicardipine, which showed an antioxidant effect only before exposure to active oxygen and reduced the DHF auto-oxidation rate, was mainly a preventive antioxidant. Benidipine, which showed an antioxidant effect only before exposure to active oxygen and had no appreciable effect on the DHF auto-oxidation rate, could interrupt the chain reaction of lipid peroxidation at the initial step alone. Although these results suggest that the antioxidant properties of some calcium antagonists may be beneficial clinically in protecting against cellular damage caused by lipid peroxidation, further studies are required to establish the antioxidant effects of these agents in vivo.

Topics: Adenosine Diphosphate; Animals; Antioxidants; Calcium Channel Blockers; Cell Membrane; Dihydropyridines; Diltiazem; Felodipine; Fumarates; Heart; Iron; Lipid Peroxidation; Male; Myocardium; Nicardipine; Nifedipine; Nisoldipine; Nitrendipine; Oxidation-Reduction; Rats; Rats, Sprague-Dawley; Verapamil

1. The possible mechanisms of action of the inhibitory effect of gomisin C on the respiratory burst of rat neutrophils in vitro was investigated. 2. The peptide formyl-Met-Leu-Phe (FMLP) induced superoxide anion (O2-) formation and O2 consumption, which was inhibited by gomisin C in a concentration-dependent manner (IC50 21.5 +/- 4.2 micrograms ml-1 for O2- formation). Gomisin C also suppressed O2- formation and consumption at low concentrations of phorbol myristate acetate (PMA) with an IC50 value of 26.9 +/- 2.1 micrograms ml-1 for O2- formation. However, gomisin C did not affect the responses induced by a high concentration of PMA. 3. Gomisin C had no effect on O2- generation and uric acid formation in the xanthine-xanthine oxidase system, and failed to alter O2- generation during dihydroxyfumaric acid (DHF) autoxidation, indicating that it does not scavenge superoxide. 4. Like trifluoperazine (TFP), gomisin C attenuated the activity of PMA-activated neutrophil particulate NADPH oxidase in a concentration-dependent manner. 5. Gomisin C reduced the elevations of cytosolic free Ca2+ in neutrophils stimulated by FMLP in the presence or absence of EDTA. Cyclopiazonic acid (CPA) induced the release of Ca2+ from intracellular stores and this was also reduced by gomisin C. However, the Ca2+ influx pathway activated by CPA was not affected by gomisin C. 6. The cellular cyclic AMP level was markedly increased by forskolin, but not by gomisin C. Moreover, the inositol phosphate levels in FMLP-activated neutrophils were not affected by gomisin C. 7. These results show that the inhibitory action of gomisin C on the respiratory burst is not mediated by changes in cellular cyclic AMP or in inositol phosphates, or by scavenging O2- released from neutrophils, but may be mediated partly by the suppression of NADPH oxidase and partly by the decrease of cytosolic Ca2+ released from an agonist-sensitive intracellular store.

Topics: Animals; Calcium; Cyclic AMP; Cyclooctanes; Dioxoles; Fumarates; Inositol Phosphates; Lignans; N-Formylmethionine Leucyl-Phenylalanine; NADH, NADPH Oxidoreductases; NADPH Oxidases; Neutrophils; Oxygen Consumption; Plant Extracts; Plants, Medicinal; Rats; Rats, Sprague-Dawley; Respiratory Burst; Superoxides

Both the production of active oxygen species and cellular damage due to concurrent lipid peroxidation are believed to be important factors in the pathogenesis of cardiovascular diseases and the ageing process. Since cardiovascular drugs are often administered over a long term, it might be advantageous if they reduced lipid peroxidation. There have been conflicting reports concerning the antiperoxidant effect of nifedipine. Therefore, we investigated whether nifedipine could inhibit lipid peroxidation in a nonenzymatic active oxygen-generating system, utilizing rat crude myocardial membranes, and compared its effect with those of propranolol, nisoldipine, and diltiazem. Nifedipine and diltiazem had no inhibitory effects on the lipid peroxidation of myocardial membranes. In contrast, nisoldipine and propranolol had a concentration-dependent antiperoxidant effect, with IC50 values of 28.2 and 50.1 microM, respectively. In addition, nisoldipine appeared to possess dual antiperoxidant mechanisms, involving both preventive and chain-breaking properties.

Topics: Animals; Antioxidants; Diltiazem; Fumarates; Heart; In Vitro Techniques; Lipid Peroxidation; Male; Myocardium; Nifedipine; Nisoldipine; Oxidation-Reduction; Oxygen; Propranolol; Rats; Rats, Sprague-Dawley

The antioxidant effects of SB 211475, a metabolite of carvedilol, a novel antihypertensive agent, were studied and compared with carvedilol and other antioxidants such as U78517F, U74500A and probucol. SB 211475 inhibited Fe(2+)-vitamin C-initiated lipid peroxidation, assessed as thiobarbituric acid reactive substance, in brain-homogenate with an IC50 of 0.28 microM. Under the same conditions, the IC50s of probucol, carvedilol, U74500A and U78517F were 50, 8.1, 0.71 and 0.16 microM, respectively. SB 211475 inhibited oxidation of human low density lipoprotein by mouse macrophages with an IC50 of 0.043 microM. In the same model, the IC50s of carvedilol, U78517F and probucol were 3.8, 0.15, and 0.80 microM, respectively. SB 211475 protected cultured bovine pulmonary artery endothelial cells against hydroxyl radical-initiated lipid peroxidation (IC50 = 0.15 microM) and cell damage (lactate dehydrogenase release, IC50 = 0.16 microM), and promoted cell survival with an EC50 of 0.13 microM. SB 211475 also protected endothelial cells against xanthine/xanthine oxidase-initiated cytotoxicity and protected rat cerebellar neurons from hydroxyl radical-mediated cell death (EC50 = 0.19 microM). Moreover, SB 211475 inhibited superoxide (O2-) release from human neutrophils stimulated by phorbol myristate acetate. These observations indicate that SB 211475 is a potent antioxidant and may potentially contribute to the therapeutic effects of carvedilol in vivo.

Topics: Adenosine Diphosphate; Animals; Antihypertensive Agents; Antioxidants; Ascorbic Acid; Brain; Carbazoles; Cattle; Cell Death; Cells, Cultured; Chromans; Endothelium, Vascular; Fumarates; Humans; Iron; Lipid Peroxidation; Lipoproteins, LDL; Macrophages; Male; Mice; Neurons; Neutrophils; Nicardipine; Oxidation-Reduction; Piperazines; Propanolamines; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Superoxides; Tetradecanoylphorbol Acetate; Xanthine; Xanthine Oxidase; Xanthines

Several studies established the role of oxygen radicals in cardiac alterations occurring during ischemia and reperfusion. Recently, platelet-activating factor (PAF), a phospholipid mediator of inflammation, was also implicated in ischemia-reperfusion injury. The present study was performed to evaluate whether biosynthesis of PAF may mediate the mechanical and electrical alterations induced by perfusion with dihydroxyfumaric acid (DHF; 1 mM), a free radical-generating compound, in guinea pig isolated perfused heart and isolated atrium. The results obtained indicate that DHF induces an intracoronary production of PAF (DHF-perfused hearts = 43.1 +/- 3.9 pg PAF; saline-perfused control hearts = PAF undetectable) and electrical and mechanical alterations in both isolated heart and atrium. These effects were shown to be dependent on superoxide (O2-) generation, because they were completely prevented by superoxide dismutase (10 mM), and were absent when DHF's ability to produce O2- was exhausted in solution. The role of PAF in mediating oxygen radical-induced electrical and mechanical alterations was established by pretreatment of cardiac preparations with WEB 2170 (0.1-10 microM), a specific PAF-receptor antagonist devoid of any direct antioxidant activity. At the concentration of 3 microM, WEB 2170 abrogated almost completely all the cardiac effect of DHF. These results suggest that PAF may act as secondary mediator of oxygen radicals in the heart.

Topics: Animals; Azepines; Free Radicals; Fumarates; Guinea Pigs; Heart; In Vitro Techniques; Myocardium; Platelet Activating Factor; Reactive Oxygen Species; Superoxide Dismutase; Triazoles

Although superoxide anions are a well-known mediator of cytotoxicity, their mechanism of target cell lysis is not clearly understood. In the present study we have used an exogenous source of superoxide to study erythrocyte cytolysis. RBC lysis was studied in buffers containing the cations Li+, Na+, K+, Rb+, and Cs+; superoxide anions were produced and available in these buffers. During this model superoxide-dependent cytolytic process, erythrocytes underwent a shape change from biconcave disk to sphere as shown by scanning electron microscopy. Soret band transmitted light microscopy has confirmed this shape change and shown that it precedes cytosolic oxidation. This evidence is consistent with a colloid-osmotic type lytic mechanism. Erythrocyte lysis was studied by 51Cr-release and light scattering methods. Superoxide-mediated target cytolysis was characterized by: 1) a sigmoidal dose-response curve and 2) a lag time in cytolysis after superoxide addition in kinetic light scattering experiments. The efficacy of cytolysis followed the rank order Cs+ > Rb+ > Na+, Li+ > sucrose = raffinose, which provides additional support for a colloid-osmotic lytic mechanism. Furthermore, the rank order potency correlates with the cations' hydration numbers. We suggest that oxidative events trigger the formation of colloid-osmotic pores approximately 1 nm in diameter.

Topics: Buffers; Cesium; Colloids; Erythrocytes; Fumarates; Hemolysis; Humans; Lithium; Microscopy, Electron, Scanning; Osmotic Pressure; Potassium; Rubidium; Scattering, Radiation; Sodium; Superoxides

To understand the direct involvement of hydroxyl radical (.OH) in the modification of functional reactivity in isolated rabbit lingual artery ring preparations, this study was undertaken to examine the effect of .OH generated from dihydroxy fumarate (DHF) plus Fe(3+)-ADP or from H2O2 plus FeSO4. When vasodilators (acetylcholine and nitroglycerin) were given after the .OH-generating system was removed from the organ chamber, the earlier .OH exposure produced an attenuation of the ring relaxation induced by acetylcholine but not that by nitroglycerin. Moreover, the earlier .OH exposure attenuated caffeine-induced contraction and depressed the phasic response, but potently enhanced the tonic response of norepinephrine-induced contraction. Both the enhanced tonic response of KCl-induced contraction produced by earlier .OH exposure and norepinephrine-induced contraction was inhibited by nisoldipine. These results are consistent with the view that .OH radicals can potentiate the voltage-dependent influx of Ca. It is also postulated that .OH may damage sarcoplasmic reticulum (SR) function in the smooth muscle cells, thus reducing Ca release from the SR (this may be reflected by the attenuation of the phasic response), and may selectively attenuate endothelium-dependent relaxation as opposed to endothelium-independent relaxation.

Topics: Animals; Arteries; Caffeine; Endothelium, Vascular; Ferric Compounds; Ferrous Compounds; Fumarates; Hydrogen Peroxide; Hydroxyl Radical; In Vitro Techniques; Male; Muscle Contraction; Muscle Relaxation; Muscle, Smooth, Vascular; Nisoldipine; Norepinephrine; Rabbits; Sarcoplasmic Reticulum; Tongue; Vasodilator Agents

In an effort to understand the damaging actions of free radicals to neuronal electrophysiology, the superoxide generator, dihydroxyfumarate (DHF), was evaluated in slices of guinea pig hippocampus. Using field potential recording techniques, population spikes and population synaptic potentials were recorded in field CA1. Slices were exposed to 3 mM DHF either alone or in the presence of a protectant. DHF did not alter the ability of the afferent volley to generate a synaptic potential, but it did impair the ability of the synaptic potential to elicit a population spike. In addition, DHF induced lipid peroxidation as measured by the thiobarbituric acid assay. Superoxide dismutase (SOD) provided no protection. Instead, SOD treatment promoted DHF damage to synaptic potentials. Catalase alone mitigated the actions of DHF, but only in SOD plus catalase was the DHF-induced electrophysiological deficit and lipid peroxidation completely antagonized. The iron chelator, Desferal, did not protect but promoted synaptic damage. Desferal may be ineffective because of the nitroxide radical formed upon its reaction with DHF. The hydroxyl radical scavenger, dimethylsulfoxide, prevented lipid peroxidation and reduced the DHF-induced deficit but did not completely prevent the impairment of spike generation. These data suggest that DHF exerts its actions through generation of hydrogen peroxide which would further react with tissue iron to produce hydroxyl radicals.

Topics: Animals; Catalase; Deferoxamine; Dimethyl Sulfoxide; Electrophysiology; Evoked Potentials; Fumarates; Guinea Pigs; Hippocampus; In Vitro Techniques; Male; Malondialdehyde; Neurons; Reference Values; Superoxide Dismutase; Superoxides; Synapses

To understand the direct involvement of free radicals causing reduction in endothelium-dependent relaxation of isolated canine coronary ring preparations, this study was undertaken to examine the effect of free radicals generated from dihydroxy fumarate (DHF) plus Fe(3+)-ADP or from H2O2 plus FeSO4. The vasodilators (acetylcholine, bradykinin, A23187, and nitroglycerin) were given after DHF/Fe(3+)-ADP or H2O2/FeSO4 was removed from the organ chamber. The earlier DHF/Fe(3+)-ADP exposure produced an attenuation of the relaxation of the rings induced by acetylcholine, bradykinin, or A23187 but not of the relaxation induced by nitroglycerin. The observed effect of previous DHF/Fe(3+)-ADP exposure was significantly protected in the vessels isolated from the dogs treated with alpha-tocopherol. In the experiments for assessing the effect of various scavengers, 1O2 scavenger histidine or iron chelator deferoxamine effectively protected the attenuation induced by DHF/Fe(3+)-ADP exposure of the relaxation elicited by acetylcholine; superoxide dismutase (SOD), catalase, or dimethyl sulfoxide (DMSO) had no effect on this system. Furthermore, the relaxation elicited by acetylcholine, but not nitroglycerin, was significantly attenuated by the earlier exposure to .OH generated by Fenton's reagent (H2O2+FeSO4); the attenuation was significantly protected by DMSO. These results are consistent with the view that .OH, 1O2, and/or iron-dependent reactive species selectively damage endothelium-dependent relaxation as opposed to endothelium-independent relaxation in endothelium-intact coronary ring preparations. It is also postulated that lipid peroxidation may be responsible for this effect.

Topics: Acetylcholine; Animals; Bradykinin; Calcimycin; Coronary Vessels; Cyclic N-Oxides; Dogs; Electron Spin Resonance Spectroscopy; Endothelium, Vascular; Female; Free Radicals; Fumarates; In Vitro Techniques; Iron; Male; Muscle Relaxation; Muscle, Smooth, Vascular; Nitroglycerin; Oxygen; Spin Labels; Superoxides; Vasodilation; Vasodilator Agents

Previous studies in our laboratory have indicated a role for free radical participation in magnesium deficiency cardiomyopathy. We have demonstrated the ability of various antioxidant drugs and nutrients to protect against magnesium deficiency-induced myocardial injury. In this study, we have examined erythrocytes from normal and magnesium-deficient animals and compared their susceptibility to an in vitro oxidative stress. Syrian male hamsters were placed on either magnesium-deficient or magnesium-supplemented diets. Animals from each group also received vitamin E in doses of 10 and 25 mg as subcutaneous implants. Erythrocytes obtained after 14 days on the diet were exposed to an exogenous hydroxyl (.OH) radical generating system (dihydroxyfumarate not equal to Fe3+ ADP) at 37 degrees C for 20 min. Erythrocyte crenation was observed and quantified by scanning electron microscopy. Lipid peroxidation, hemolysis (%), and intracellular glutathione levels were determined. In addition, serum lipid changes and membrane phospholipids were characterized. Our data demonstrate that erythrocytes from magnesium-deficient animals are more susceptible to free radical injury, supporting our hypothesis that magnesium deficiency reduces the threshold antioxidant capacity.

Topics: Adenosine Diphosphate; Animals; Chlorides; Cholesterol; Cricetinae; Erythrocyte Membrane; Erythrocytes; Ferric Compounds; Fumarates; Glutathione; Hemolysis; Hydroxides; Hydroxyl Radical; In Vitro Techniques; Iron Chelating Agents; Lipid Peroxidation; Magnesium Deficiency; Male; Malondialdehyde; Membrane Lipids; Mesocricetus; Microscopy, Electron, Scanning; Phospholipids; Reference Values; Superoxides; Triglycerides

Carvedilol produced a dose-dependent inhibition of superoxide (O2-) release from human neutrophils (PMNs) (IC50 = 28 microM) and scavenged O2- generated during dihydroxyfumaric acid (DHF) autooxidation (IC50 = 41 microM). Other beta-blockers, such as celiprolol, labetalol and atenolol, or the antioxidant, 'lazaroid', U74500A had no effect on O2- either released from PMNs or generated during DHF autooxidation. Propranolol, at 0.3 mM, inhibited O2- release from PMNs (73%) but failed to scavenge O2- generated from DHF. The novel free radical-scavenging effect of carvedilol may contribute to the cardioprotective activity of the compound.

Topics: Adrenergic beta-Antagonists; Adult; Antihypertensive Agents; Carbazoles; Carvedilol; Free Radical Scavengers; Fumarates; Humans; In Vitro Techniques; Lipid Peroxidation; Neutrophils; Propanolamines; Superoxides; Tetradecanoylphorbol Acetate; Vasodilator Agents

Secreting and producing capacities of alpha-fetoprotein (AFP) and albumin (ALB) by human hepatoma cell line (HH2-6) exposed to oxygen free radicals generated by dihydroxyfumarate (DHF) were studied in vitro. It was found that cell number were declined in proportion to DHF concentrations for 48 hrs culture and cell proliferations were inhibited by DHF in growth curve. The amount of AFP secreted per cell (secreting capacity) was decreased at high concentration of DHF (50 micrograms/ml) for 48 hrs culture, and remarkable elevation of AFP-secreting capacity for growth stage was inhibited even by DHF (10 micrograms/ml). On the other hand, ALB-secreting capacity was not affected with DHF. Producing capacities of AFP and ALB were correlated with secreting capacities. Cu, Zn-superoxide dismutase (Cu, Zn-SOD) interfered these reactions. These results suggest that oxygen free radicals inhibit cell proliferations and suppress AFP-secreting and producing capacities of HH2-6 cells selectively.

Topics: Albumins; alpha-Fetoproteins; Carcinoma, Hepatocellular; Cell Division; Free Radicals; Fumarates; Humans; Liver Neoplasms; Oxygen; Tumor Cells, Cultured

The effect of magnesium (Mg)-deficient culture on endothelial cell susceptibility to oxidative stress was examined. Bovine endothelial cells were cultured in either control sufficient (0.8 mM) or deficient (0.4 mM) levels of MgCl2. Oxygen radicals were produced extracellularly by the addition of dihydroxyfumarate and Fe(3+)-ADP. Isolated Mg-deficient endothelial cells produced 2- to 3-fold higher levels of thiobarbituric acid (TBA)-reactive materials when incubated with this free radical system. Additional studies were performed using digitized video microscopy and 2',7'-dichlorofluorescein diacetate (DCFDA) as an intracellular indicator for oxidative events at the single cell level. In response to the exogenous oxidative stress, endothelial cells exhibited a time-dependent increase in fluorescence, suggestive of intracellular lipid peroxidation. The increase in cellular fluorescence began within 1 min of free radical addition; the Mg-deficient cells exhibited a more rapid increase in fluorescence than that of Mg-sufficient cells. In separate experiments, cellular viability was assessed using the Trypan blue exclusion assay. Mg deficiency increased cytotoxicity of the added oxyradicals, but the loss of cellular viability began to occur only after 15 min of free radical exposure, lagging behind the detection of intracellular oxidation products. These results suggest that increased oxidative endothelial cell injury may contribute to vascular injury during Mg deficiency.

Topics: Adenosine Diphosphate; Animals; Aorta; Cattle; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Endothelium, Vascular; Free Radicals; Fumarates; Kinetics; Lipid Peroxidation; Magnesium; Magnesium Deficiency; Oxidation-Reduction; Thiobarbituric Acid Reactive Substances

The vascular endothelium is a significant site for tissue injury following exposure to reactive oxygen species derived from a number of sources. In order to develop a better understanding of the mechanism(s) of oxidative damage, monolayer cultures of endothelial cells obtained from bovine pulmonary arteries were exposed to reactive oxygen species generated from the oxidation of dihydroxyfumarate (DHF) to diketosuccinate. Exposure to oxidizing DHF caused a loss of cell membrane integrity that was delayed in onset; that is, it did not begin until 2 h after the addition of DHF although reactive oxygen species are produced immediately by DHF in solution. Endothelial cell lysis by DHF was prevented by the simultaneous addition of superoxide dismutase (SOD), catalase (CAT), or deferoximine (DFX). This oxidant-induced lysis was unaffected by N,N,-diphenyl-p-phenylenediamine (DPPD), a potent inhibitor of lipid peroxidation. However, simultaneous addition of 3-aminobenzamide (3AB) and nicotinamide (NA), inhibitors of poly(ADP-ribose) polymerase, prevented cell lysis. Oxidant-induced loss of membrane integrity was preceded by the early appearance of DNA strand breaks, by increased levels of poly(ADP-ribose), the product of polymerase activity, and by depletion of NAD+ and ATP, followed by a decline in the energy charge ratio of the cells. None of these intracellular changes occurred when either SOD, CAT, or DFX were added at the same time as DHF, suggesting that O2-., H2O2, and HO. mediated these changes. The O2-. appears to be important in the autoxidation reaction of DHF. The latter two reactive oxygen species may be part of cellular-catalyzed Fenton chemistry. The increase in poly(ADP-ribose), depletion of NAD+, and the decline in ATP were also prevented by the addition of 3AB. The oxidant-induced DNA strand breakage was, however, unaffected by either 3AB or NA. Addition of 3AB immediately prior to the onset of cell lysis (2 h after the addition of DHF), prevented cell lysis, i.e., "rescued" the cells when neither SOD, CAT, nor DFX addition were effective. Concurrent with the "rescue" from lysis by 3AB, there was an increase in NAD+ content and a return of the energy charge ratio to control levels. The data presented in this study suggests that in endothelial cells, DNA is a very sensitive target for reactive oxygen species and HO. is the likely proximal damaging species.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Adenine Nucleotides; Animals; Antioxidants; Catalase; Cattle; Cell Survival; Cells, Cultured; Deferoxamine; Endothelium, Vascular; Energy Metabolism; Enzyme Activation; Free Radicals; Fumarates; Hydrogen Peroxide; Kinetics; NAD; Oxygen; Phenylenediamines; Poly(ADP-ribose) Polymerases; Pulmonary Artery; Superoxide Dismutase; Superoxides

Oxygen radicals have been implicated in the pathogenesis of reperfusion arrhythmias. However, the basic electrophysiological alterations accompanying the effects of oxygen radicals on action potential (AP) are poorly understood.. We investigated the effects of oxygen radicals generated by dihydroxyfumarate (DHF, 5 mM) on AP parameters and on ionic currents in patch-clamped guinea pig ventricular myocytes. DHF consistently caused a marked prolongation of AP duration, which was already significant after 60 seconds of exposure and continued to increase over time. Within 5 minutes, the majority of cells developed early afterdepolarizations (EADs) or became unexcitable. Both AP prolongation and occurrence of EADs were completely prevented in the presence of the oxygen radical scavengers superoxide dismutase (SOD) and catalase (CAT). Prolongation of AP duration was accompanied by a marked decreased in time-dependent potassium current (IK) and calcium current (ICa). The inward rectifier K current (IK1) was unaffected, suggesting no widespread changes in membrane properties. IK and ICa alterations were also significantly reduced by SOD and CAT. In additional experiments, intracellular calcium levels were kept constantly low by addition of 200 microM ethyleneglycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetra-acetic acid (EGTA) to the pipette solution. Under these conditions, the effects of DHF on AP duration and the occurrence of EADs were largely prevented. However, EGTA did not prevent cells from becoming unexcitable, nor did it affect the decrease in both IK and ICa upon exposure to DHF.. Exposure to an exogenous source of oxygen radicals may induce major electrophysiological alterations in isolated myocytes, which might be related to changes in specific ionic currents and in level of intracellular calcium. These alterations occur with a time course consistent with the rapid onset of ventricular arrhythmias in reperfused hearts.

Topics: Action Potentials; Animals; Arrhythmias, Cardiac; Free Radical Scavengers; Free Radicals; Fumarates; Guinea Pigs; In Vitro Techniques; Ion Pumps; Myocardial Reperfusion Injury; Myocardium; Oxygen; Time Factors

Exposure of diploid fetal human fibroblasts (IMR-90) to superoxide generated by dihydroxyfumarate resulted in increased collagen synthesis. The synthesis of type III collagen was stimulated to a greater extent than the synthesis of type I collagen. The stimulation of collagen synthesis was abolished by superoxide dismutase. Our observations suggest that superoxide may play a role in the regulation of collagen synthesis and may modulate differential collagen gene expression. These observations may explain the increased synthesis of collagen in tissues following inflammation or exposure to oxidant conditions.

Topics: Cells, Cultured; Collagen; Fibroblasts; Fumarates; Humans; Superoxides

A relatively pure and stable compound III of bovine spleen myeloperoxidase was prepared from native enzyme using the aerobic oxidation of dihydroxyfumarate to generate O2-(.). Spectral scans show well defined peaks at 450 and 625 nm and an isosbestic point between compound III and native enzyme at 440 nm. Compound III decayed to native enzyme without any detectable intermediate. The rate of decay was faster at alkaline pH values and also in the presence of superoxide dismutase. Ascorbic acid reduces compound III to native enzyme with a second order rate constant of (4.0 +/- 0.1) x 10(2) M-1 s-1. The ascorbic acid reduction of compound III has potential physiological relevance since it could help maintain the catalytic cycle of myeloperoxidase to generate the bactericidal agent hypochlorous acid.

Topics: Animals; Ascorbic Acid; Cattle; Enzyme Stability; Fumarates; Hydrogen-Ion Concentration; Kinetics; Peroxidase; Spectrophotometry; Spleen; Superoxide Dismutase

The ability of captopril and enalaprilat, 2 angiotensin-converting enzyme (ACE) inhibitors, to scavenge superoxide anion radical was examined. With use of a number of superoxide-generating systems, such as xanthine-xanthine oxidase, phorbol myristate acetate-activated neutrophils, auto-oxidizing dihydroxyfumarate, and auto-oxidation of epinephrine to adrenochrome, captopril was seen not to scavenge superoxide directly, because it did not inhibit superoxide-dependent cytochrome c or nitro-blue tetrazolium reduction. Superoxide-dependent cytochrome c reduction was inhibited only when captopril was preincubated with a lower concentration of cytochrome c (22 microM). This effect was due to a decrease in the concentration of cytochrome c, because captopril reduced cytochrome c directly. When this effect was compensated for, no cytochrome c reduction induced by superoxide was observed. Captopril inhibited the auto-oxidation of epinephrine to adrenochrome at pH 10.2 where this auto-oxidation is superoxide-dependent, and at pH 7.8 where it is superoxide-independent and superoxide dismutase insensitive. It appears that captopril, in this respect, acted as a nonspecific antioxidant, probably by reducing an intermediate in the complex oxidation of epinephrine to adrenochrome. Therefore, caution may be used in interpreting the role of captopril in the attenuation of reperfusion-induced myocardial dysfunction and in attributing this effect to the inhibition of free radical mechanism.

Topics: Captopril; Cytochrome c Group; Enalaprilat; Epinephrine; Fumarates; Humans; Nitroblue Tetrazolium; Oxidation-Reduction; Superoxides

Stopped-flow techniques were used to investigate the kinetics of the reaction of lignin peroxidase compounds II and III (LiPII and LiPIII) with peroxides. Rate data were obtained from single-turnover experiments under pseudo-first-order conditions. LiPII reacts with H2O2 or peracetic acid (AcOOH) to form a modified LiPIII, designated as LiPIII*, via a biphasic reaction. During the first phase, LiPIII is formed as an intermediate. Kinetic analysis also indicates a LiPII-peroxide complex. The first-order dissociation rate constants for the reaction of LiPII with H2O2 and AcOOH are 7.9 +/- 0.5 and 4.9 +/- 0.6 s-1, respectively. The rate of the H2O2 reaction is approximately 500 times the rate of the comparable reaction with horseradish peroxidase, suggesting it is physiologically significant. The activation energy for the formation of LiPIII is 23 kJ mol-1. During the second phase, the intermediate LiPIII is converted to LiPIII*, confirmed by analyzing the reaction of exogenously prepared LiPIII with peroxides. The second-order rate constants for the reaction of LiPIII with H2O2 and AcOOH are (3.7 +/- 0.2) x 10(2) M-1 s-1 and (2.9 +/- 0.2) x 10(2) M-1 s-1, respectively. The conversion of LiPIII to LiPIII* is reversible; the first-order rate constant for the reverse reaction is approximately (6.6 +/- 0.6) x 10(-2) s-1. The rates of both LiPIII and LiPIII* formation decrease markedly above pH 4.0. The pH dependence of these reactions is controlled by a heme-linked ionizable group of pK alpha congruent to 4.2.

Topics: Basidiomycota; Fumarates; Hydrogen Peroxide; Hydrogen-Ion Concentration; Kinetics; Peracetic Acid; Peroxidases; Peroxides; Superoxides; Thermodynamics

Topics: Animals; Cataract; Cattle; Ethylenes; Free Radicals; Fumarates; Glutathione; In Vitro Techniques; Light; Oxidation-Reduction; Oxygen; Potassium Iodide; Riboflavin

Oxygen radicals have been implicated as important mediators of myocardial ischemic and reperfusion injury. A major product of oxygen radical formation is the highly reactive hydroxyl radical via a biological Fenton reaction. The sarcoplasmic reticulum is one of the major target organelles injured by this process. Using a oxygen radical generating system consisting of dihydroxyfumarate and Fe3+-ADP, we studied lipid peroxidation and Ca2+-ATPase of cardiac sarcoplasmic reticulum. Incubation of sarcoplasmic reticulum with dihydroxyfumarate plus Fe3+-ADP significantly inhibited enzyme activity. Addition of superoxide dismutase, superoxide dismutase plus catalase (15 micrograms/ml) or iron chelator, deferoxamine (1.25-1000 microM) protected Ca2+-ATPase activity. Time course studies showed that this system inhibited enzyme activity in 7.5 to 10 min. Similar exposure of sarcoplasmic reticulum to dihydroxyfumarate plus Fe3+-ADP stimulated malondialdehyde formation. This effect was inhibited by superoxide dismutase, catalase, singlet oxygen, and hydroxyl radical scavengers. EPR spin-trapping with 5,5-dimethyl-1-pyrroline-N-oxide verified production of the hydroxyl radical. The combination of dihydroxyfumarate and Fe3+-ADP resulted in a spectrum of hydroxyl radical spin trap adduct, which was abolished by ethanol, catalase, mannitol, and superoxide dismutase. The results demonstrate the role of oxygen radicals in causing inactivation of Ca2+-ATPase and inhibition of lipid peroxidation of the sarcoplasmic reticulum which could possibly be one of the important mechanisms of oxygen radical-mediated myocardial injury.

Topics: Adenosine Diphosphate; Animals; Calcium-Transporting ATPases; Dogs; Electron Spin Resonance Spectroscopy; Ferric Compounds; Fumarates; In Vitro Techniques; Lipid Peroxides; Myocardium; Oxygen; Sarcoplasmic Reticulum

Ferritin iron release, a process of considerable interest in biology and medicine, occurs most readily in the presence of reducing agents. Here is described a kinetic assay for measuring the rate of ferritin iron removal promoted by various reductants. The new procedure uses ferrozine as a chromophoric, high-affinity chelator for the product, Fe(II). The initial rate of iron release is quantified by continuous spectrophotometric measurement of the Fe(ferrozine)2/3+ complex which absorbs maximally at 562 nm. The initial rate of iron mobilization is dependent on reductant concentration, but not on the concentration of the chelating agent, ferrozine. Saturation kinetics are observed for all reductants, including dihydroxyfumarate, cysteine, caffeic acid, ascorbate, and glutathione. Superoxide dismutase greatly inhibits ferritin iron release by ascorbate, but has little or no effect on the reducing action of dihydroxyfumarate, cysteine, caffeic acid, or glutathione. Ferritin iron removal by dihydroxyfumarate was inhibited by various metal ions. This new assay may be used for rapid screening of test compounds for treatment of iron overload and for investigation of the mechanistic aspects of ferritin iron reduction.

Topics: Animals; Ascorbic Acid; Ferritins; Ferrozine; Fumarates; In Vitro Techniques; Iron; Kinetics; Oxidation-Reduction

Oxygen free radicals may participate in a variety of pathological cardiac conditions which are associated with an increased incidence of arrhythmias. However, evidence that free radicals per se can alter the electrical function of the myocardium is not convincing. Physiological solutions containing 3 mM dihydroxyfumaric acid (DHF), a compound known to generate free radicals, were superfused over calcium-tolerant cells isolated from the adult canine ventricle. The time course for changes in transmembrane action potentials was monitored using conventional microelectrode techniques. Changes were observed which could be conveniently segregated into three stages. Initially during superfusion with DHF, the voltage of the action potential plateau became more positive and the action potential duration increased (stage 1). Continued superfusion was associated with the development of both early and delayed afterdepolarizations (stage 2), which occasionally produced triggered beats. Subsequently, some cells failed to repolarize beyond -40 mV following an action potential upstroke. In cells which maintained normal levels of resting membrane potential, early and delayed afterdepolarizations ceased concomitant with the development of an increasingly more negative plateau voltage. Action potential duration decreased and plateau potential "collapsed", eventually merging with the resting level of the membrane potential. Resting membrane potential then gradually depolarized to less than -40 mV and all cells became inexcitable within 6 to 20 min (stages 3). Exposure of cells to xanthine (2 mM): xanthine oxidase (0.01 U/ml), another system known to generate free radicals, produced similar results. Superfusion with DHF solutions containing either superoxide dismutase or catalase delayed the appearance and attenuated the development of the changes in the cardiocyte action potential. The results demonstrate that isolated cardiocytes exposed to free radical generating solutions can undergo changes in their electrophysiological activity that resemble those said to underlie disturbances of cardiac rate and rhythm in the clinical setting.

Topics: Action Potentials; Animals; Catalase; Dogs; Electric Stimulation; Electrophysiology; Free Radicals; Fumarates; Heart; Myocardium; Superoxide Dismutase; Time Factors; Xanthine; Xanthine Oxidase; Xanthines

To delineate the active free radical species mediating the toxic effects of autoxidizing dihydroxyfumarate (DHF), isolated rabbit right ventricular papillary muscles were exposed to 4.5 mM DHF in the presence of FeCl3, ADP and bovine albumin. In the absence of free radical scavengers a 47.3 +/- 11.5% (mean +/- standard deviation) depression in contractile force was noted over 60 minutes. Neither the combination of superoxide dismutase (SOD) 3,200 u/cc and catalase (CAT) 2,950 u/cc nor mannitol 0.1 M provided statistically significant protection. Deferoxamine mesylate (DFX) 10 mg/cc (15 mM) did provide significant protection of muscle function both in the presence and absence of SOD and CAT (p less than 0.01). The degree of protection conferred by DFX alone was statistically similar to that of DFX with SOD and CAT. This data suggests the involvement of an iron-oxygen complex not dependent on superoxide or hydrogen peroxide for its formation and not readily scavenged by mannitol. The perferryl ion may be representative of such a species. Alternatively, a reactive complex similar to the 'Crypto-OH' radical proposed by Youngman may be formed by the reaction of DHF with iron and oxygen.

Topics: Animals; Catalase; Deferoxamine; Free Radicals; Fumarates; In Vitro Techniques; Iron; Oxygen; Papillary Muscles; Rabbits; Superoxide Dismutase

The effects of naturally occurring lipid amphiphiles on free radical-mediated peroxidative injury in isolated canine sarcolemma were studied. Highly enriched canine myocytic sarcolemmal membranes were preincubated for 10 min at 37 degrees C with or without different amphiphilic lipids before the addition of a free radical-generating system consisting of dihydroxyfumarate and Fe3+-ADP. Lipid peroxidation, assayed as malondialdehyde formation, was catalyzed linearly up to 40 min in the control samples. Pretreatment of the sarcolemma with palmitoyl-CoA, palmitoylcarnitine, or lysophosphatidylcholine accelerated the initial rates (20 min) of peroxidation in a concentration-dependent manner (10-100 microM) and achieved maximal stimulation (240%, 160%, and 210%, respectively, of controls) at 50 microM concentrations of each of these amphiphiles. However, free fatty acids, CoA, and carnitine were without effect. These promoting effects of the amphiphiles persisted over a wide pH range (pH 6.0-7.8) and exhibited additive effects when lower levels of different amphiphiles were combined together. Associated with the accelerated rates of peroxidation produced by palmitoyl-CoA and palmitoylcarnitine were greater losses in the activity of sarcolemmal (Na,K)-ATPase. Since all three kinds of amphiphilic lipids accumulate during ischemia, this study suggests a novel mechanism of potentiation of sacolemmal membrane injury when free radicals are present.

Topics: Animals; Dogs; Free Radicals; Fumarates; Lipid Metabolism; Lipid Peroxides; Myocardium; Oxidation-Reduction; Palmitoyl Coenzyme A; Palmitoylcarnitine; Sarcolemma; Sodium-Potassium-Exchanging ATPase

Superoxide dismutase and catalase, which catalytically remove O-2 and H2O2, respectively, each separately protected cultured pulmonary artery endothelial cells from loss of membrane integrity after exposure to oxygen radicals generated either cellularly (polymorphonuclear leukocytes) or chemically (dihydroxyfumarate). Nicotinamide, a precursor of nicotinamide-adenine dinucleotide (NAD) and an inhibitor of ADP-ribose synthetase, also protected cultured endothelial cells from loss of membrane integrity in a concentration-dependent manner after exposure to DHF.

Topics: Animals; Catalase; Cattle; Drug Interactions; Endothelium; Fumarates; In Vitro Techniques; L-Lactate Dehydrogenase; Lung Diseases; Niacinamide; Pulmonary Artery; Superoxide Dismutase

Topics: Aniline Compounds; Ascorbic Acid; Cytochrome P-450 Enzyme System; Fumarates; Hemoglobins; Humans; Hydroxylation; Kinetics; Methemoglobin; NAD; Oxyhemoglobins; Riboflavin

Topics: Animals; Anions; Azides; Bile Pigments; Carbon Monoxide; Erythrocytes; Fumarates; Heme; Hemeproteins; Horses; Hydrogen Peroxide; Oxygen; Potassium Cyanide; Sodium Azide; Superoxides; Xanthine Oxidase

Autooxidizing dihydroxyfumarate (DHF) is a known generator of superoxide anions and by hydroxyl free radicals. Isolated rat pancreatic islets were exposed for 5 min to 0.4 mg/ml DHF in an oxygenated buffer solution. As a result of this exposure, the islets exhibited a 60% reduction in glucose-stimulated insulin release. The presence of superoxide dismutase, catalase, or diethylenetriaminepentaacetic acid, a metal chelator, during the exposure period protected the islets from the effects of autooxidizing DHF. This suggests that superoxide anions, hydrogen peroxide, and, ultimately, hydroxyl free radicals play a role in the insulin inhibitory effects caused by DHF exposure. Glucose (5 mg/ml), but not galactose, was also capable of protecting islets from DHF. The effects of DHF exposure on isolated islets appear to be similar to those previously reported for alloxan and lend support to the concept that hydroxyl free radicals generated during the oxidation of certain compounds can alter endocrine cell function.

Topics: Animals; Catalase; Dose-Response Relationship, Drug; Fumarates; Glucose; Insulin; Islets of Langerhans; Male; Pentetic Acid; Rats; Superoxide Dismutase

Topics: Alloxan; Animals; Dose-Response Relationship, Drug; Free Radicals; Fumarates; Glucose; Insulin; Insulin Secretion; Islets of Langerhans; Male; Methylurea Compounds; Rats

Topics: Animals; Carbon Monoxide; Chromatography; Cyanides; Enzyme Induction; Fumarates; Hydrogen Peroxide; Kinetics; Lipid Peroxides; Lipids; Male; Mitochondria, Liver; Molecular Weight; Peroxidases; Rats; Spectrum Analysis

Topics: Animals; Catalase; Copper; Cytochrome P-450 Enzyme System; Fumarates; Iron; Kinetics; Lipid Peroxides; Male; Manganese; Microsomes, Liver; Oxidation-Reduction; Rats; Superoxide Dismutase

1. Dihydroxyfumarate slowly autoxidizes at pH6. This reaction is inhibited by superoxide dismutase but not by EDTA. Mn2+ catalyses dihydroxyfumarate oxidation by reacting with O2 leads to to form Mn3+, which seems to oxidize dihydrofumarate rapidly. Cu2+ also catalyses dihydroxyfumarate oxidation, but by a mechanism that does not involve O2 leads to. 2. Peroxidase catalyses oxidation of dihydroxyfumarate at pH6; addition of H2O2 does not increase the rate. Experiments with superoxide dismutase and catalase suggest that there are two types of oxidation taking place: an enzymic, H2O2-dependent oxidation of dihydroxyfumarate by peroxidase, and a non-enzymic reaction involving oxidation of dihydroxyfumarate by O2 leads to. The latter accounts for most of the observed oxidation of dihydroxyfumarate. 3. During dihydroxyfumarate oxidation, most peroxidase is present as compound III, and the enzymic oxidation may be limited by the low rate of breakdown of this compound. 4. Addition of p-coumaric acid to the peroxidase/dihydroxyfumarate system increases the rate of dihydroxyfumarate oxidation, which is now stimulated by addition of H2O2, and is more sensitive to inhibition by catalase but less sensitive to superoxide dismutase. Compound III is decomposed in the presence of p-coumaric acid. p-Hydroxybenzoate has similar, but much smaller, effects on dihydroxyfumarate oxidation. However, salicylate affects neither the rate nor the mechanism of dihydroxyfumarate oxidation. 5. p-Hydroxybenzoate, salicylate and p-coumarate are hydroxylated by the peroxidase/dihydroxyfumarate system. Experiments using scavengers of hydroxyl radicals shown that OH is required. Ability to increase dihydroxyfumarate oxidation is not necessary for hydroxylation to occur.

Topics: Coumaric Acids; Free Radicals; Fumarates; Horseradish Peroxidase; Hydrogen Peroxide; Hydroxylation; Oxidation-Reduction; Oxygen; Peroxidases; Phenols; Superoxide Dismutase; Superoxides

Topics: Animals; Catechol Oxidase; Chromatography; Dihydroxyphenylalanine; Fumarates; Hydrogen-Ion Concentration; Melanoma; Methods; Mice; Monophenol Monooxygenase; Neoplasm Transplantation; Neoplasms, Experimental; Oxidation-Reduction; Radioisotopes; Spectrophotometry; Time Factors; Tyrosine

Topics: Citrates; Citric Acid; Enzyme Inhibitors; Fumarates; Oxidation-Reduction; Oxidoreductases; Peroxidase; Peroxidases; Research

Topics: Coloring Agents; Fumarates; Oxidoreductases; Oxygen Isotopes; Peroxidase; Peroxidases

Topics: Acids; Animals; Fumarates; Liver; Maleates; Metabolic Networks and Pathways; Pentoses; Pyruvates; Rats

Topics: Decarboxylation; Fumarates; Maleates; Succinates

Topics: Aspartic Acid; Fumarates; Glutamates; Glutamic Acid; Maleates

Topics: Acids; Coenzymes; Fumarates; Oxidation-Reduction; Succinates