Showing papers in "Free Radical Research in 1996"
TL;DR: The methods available for measuring steady-state damage and the actual rate of damage to DNA, proteins and lipids are reviewed, highlighting areas in which further methodological development is urgently required.
Abstract: Reactive oxygen species and reactive nitrogen species are formed in the human body. Endogenous antioxidant defences are inadequate to scavenge them completely, so that ongoing oxidative damage to DNA, lipids, proteins and other molecules can be demonstrated and may contribute to the development of cancer, cardiovascular disease and possibly neurodegenerative disease. Hence diet-derived antioxidants may be particularly important in protecting against these diseases. Some antioxidants (e.g. ascorbate, certain flavonoids) can exert pro-oxidant actions in vitro, often by interaction with transition metal ions. The physiological relevance of these effects is uncertain, as is the optimal intake of most diet-derived antioxidants. In principle, these questions could be addressed by examining the effects of dietary composition and/or antioxidant supplementation upon parameters of oxidative damage in vivo. The methods available for measuring steady-state damage (i.e. the balance between damage and repair or replacement of damaged molecules) and the actual rate of damage to DNA, proteins and lipids are reviewed, highlighting areas in which further methodological development is urgently required.
595 citations
TL;DR: Ascorbic acid has a multiplicity of antioxidant properties, but it can exert pro-oxidant effects in vitro, usually by interaction with transition metal ions, and it is as yet uncertain that these pro-oxide effects have any biological relevance.
Abstract: Ascorbic acid has a multiplicity of antioxidant properties, but it can exert pro-oxidant effects in vitro, usually by interaction with transition metal ions. It is as yet uncertain that these pro-oxidant effects have any biological relevance: some of the available data are summarized.
533 citations
TL;DR: The abilities of several biological antioxidants to protect against peroxynitrite-dependent inactivation of alpha 1-antiproteinase, and to inhibit tyrosine nitration upon addition of peroxlynitrite are compared.
Abstract: Peroxynitrite, formed by reaction of superoxide and nitric oxide, appears to be an important tissue-damaging species generated at sites of inflammation. In this paper, we compare the abilities of several biological antioxidants to protect against peroxynitrite-dependent inactivation of α1-antiproteinase, and to inhibit tyrosine nitration upon addition of peroxynitrite. GSH and ascorbate protected efficiently in both systems. Uric acid inhibited tyrosine nitration but not α1-antiproteinase inactivation. The possibility that ascorbic acid is an important scavenger of reactive nitrogen species in vivo is discussed.
190 citations
TL;DR: Results showed that peroxynitrite exhibits oxidizing properties toward purine moieties both in nucleosides and isolated DNA, and questions the involvement of hydroxyl radicals as the main oxidizing species released by decomposition of per oxynitrous acid.
Abstract: Reaction of nitric oxide with superoxide anion produces the highly reactive species peroxynitrite (ONOO−). This compound has been shown to be a strong oxidant of lipids and proteins. However, no data are available on its effect on DNA, with the exception of the induction of strand breaks. We report the result of studies on the reactions of peroxynitrite with the adenine and guanine moieties of nucleosides and isolated DNA. The samples were analyzed for 8-oxo-7,8-dihydro-2′-deoxyguano-sine (8-oxo-dGuo), 2,2-diamino-4–[(2-deoxy-β-D-erythro-pentofuranosyl)amino]-5–(2H)-oxazolone (oxazolone) and 8-oxo-7,8-dihydro-2′-deoxyadenosine (8-oxo-dAdo). The effects of peroxynitrite treatment were compared with those of ionizing radiation in aerated aqueous solution, chosen as a source of hydroxyl radicals. At the nucleoside level, both oxidizing conditions led to the formation of oxazolone and 8-oxo-dAdo. In addition, evidence was provided for the formation of the 4R* and 4S* diastereoisomers of 4-hydroxy-8-oxo-4,8-di...
163 citations
TL;DR: The oxidative inactivation of rabbit skeletal muscle Ca(2+)-ATPase in sarcoplasmic reticulum (SR) vesicles by peroxynitrite (ONOO-) was investigated and possibly the oxidation of other amino acids contributes to enzyme inactivation, corroborated by amino acid analysis.
Abstract: The oxidative inactivation of rabbit skeletal muscle Ca 2+-ATPase in sarcoplasmic reticulum (SR) vesicles by peroxynitrite (ONOO−) was investigated. The exposure of SR vesicles (10 mg/ml protein) to low peroxynitrite concentrations (≤0.2 mM) resulted in a decrease of Ca2+-ATPase activity primarily through oxidation of sulf-hydryl groups. Most of this deactivation (ca. 70%) could be chemically reversed by subsequent reduction of the enzyme with either dithiothreitol (DTT) or sodium borohydride (NaBH4), indicating that free cysteine groups were oxidized to disulfides. The initial presence of 5 mM glutathione failed to protect the SR Ca2+-ATPase activity. However, as long as peroxynitrite concentrations were kept ≤0.45 mM, the efficacy of DTT to reverse Ca 2+-ATPase inactivation was enhanced for re action mixtures which initially contained 5 mM gluta thione. At least part of the disulfides were formed intermolecularly since gel electrophoresis revealed protein aggregation which could be reduced under reducin...
159 citations
TL;DR: The characterization of several monoclonal antibodies which recognize 4-hydroxynonenal (HNE) modified proteins revealed that the two antibodies are highly selective for HNE bound to histidine with only some cross reaction to H NE bound to lysine and cysteine.
Abstract: A promising approach to study lipid peroxidation pathology is antibodies recognizing aldehydes which react with and became bound to amino acid side chains of proteins. We present in this study the characterization of several monoclonal antibodies which recognize 4-hydroxynonenal (HNE) modified proteins. Six out of 20 antibodies recognizing HNE modified BSA were able to detect HNE-protein adducts in peroxidized liver microsomes. Two of these antibodies were selected and characterized. Both antibodies could also detect HNE-protein adducts in oxidized low density lipoprotein. They exhibit no detectable cross reaction with proteins modified by malonaldehyde, nonanal, nonenal and 4-hydroxyhexenal. Protein bound 4-hydroxyoctenal and 4-hydroxydecenal were recognized to some extent. Further characterization revealed that the two antibodies are highly selective for HNE bound to histidine with only some cross reaction to HNE bound to lysine and cysteine. Preliminary quantitative ELISA-analysis showed that oxidized ...
154 citations
TL;DR: The rate constants for the spontaneous dimerization and decomposition of the protonated ferrates are orders of magnitude slower than their corresponding reduction reduction by superoxide indicating an outer-sphere mode of electron transfer for the latter process.
Abstract: The reduction of ferrate(VI) to ferrate(V) by superoxide ions was studied over the pH range 2.6-13.0 using the premix pulse radiolysis technique. The pH dependence indicates that only the unstable protonated forms of ferrate, H2FeO4 (pKa3.5) and HFeO4- (pKa7.3) are reactive, k(HFeO4(-) + O2) = (1.7 +/- 0.2) x 10(7) M-1 s-1. The stable ferrate ion, FeO4(2-), showed no significant reactivity towards either hydrogen peroxide or superoxide anion. The rate constants for the spontaneous dimerization and decomposition of the protonated ferrates, e.g. k(HFeO4(-) + HFe04) approximately 250 M-1s-1, are orders of magnitude slower than their corresponding reduction reduction by superoxide indicating an outer-sphere mode of electron transfer for the latter process. In contrast the ferrate(VI) species H3FeO4+ (pKa = 1.6 +/- 0.2), H2FeO4, and HFeO4- oxidize hydrogen peroxide, e.g. k(HFeO4(-) + H2O2) = 170 M-1 s-1), at rates which correspond closely to their dimerization rates suggesting an inner-sphere controlled mechanism.
144 citations
TL;DR: Profiles of the local nitric oxide (.NO) diffusion-concentration product across the egg yolk phosphatidylcholine membrane in the absence and presence of 30 mol% cholesterol were obtained using line-broadening electron paramagnetic resonance (EPR) and lipid-soluble nitroxide spin labels.
Abstract: Profiles of the local nitric oxide (.NO) diffusion-concentration product across the egg yolk phosphatidylcholine membrane in the absence and presence of 30 mol% cholesterol were obtained using line-broadening electron paramagnetic resonance (EPR) and lipid-soluble nitroxide spin labels. Membrane .NO permeability coefficients were calculated from these profiles. At 20 degrees C, values of 93 and 77 cm/s for membranes in the absence and presence of cholesterol were obtained, compared with 73 and 66 cm/s for water layers of the same thickness as the membranes. Fluid-phase membranes are not barriers to .NO transport. Cholesterol significantly increases .NO transport in the center of the lipid bilayer.
144 citations
TL;DR: It is demonstrated that H. pylori induces enhanced production of ROS in GC, and enhances membrane damage, and the dose-dependent protective abilities of selected ROS scavengers on LDH leakage were determined.
Abstract: Reactive oxygen species (ROS) and Helicobacter pylori have been identified as pathogenic factors in several gastrointestinal disorders. Since little information is available regarding the mechanistic pathways of H. pylori-induced gastric injury, the potential role of ROS was investigated. The induction of ROS in gastric cells (GC) by H. pylori was assessed using chemiluminescence, cytochrome c reduction, nitrobluetetrazolium (NBT) reduction and lactate dehydrogenase (LDH) leakage. The dose-dependent protective abilities of selected ROS scavengers on LDH leakage were determined. Following incubation of GC with three strains of H. pylori (1:1), approximately 5.7–8.0 and 3.8–4.3 fold increases were observed in cytochrome c and NBT reduction, respectively, demonstrating production of ROS. Enhanced chemiluminescence responses of 2.1– and 3.7-fold were observed following incubation of GC with H. pylori (ATCC 43504) at ratios of 1:1 and 1:10, respectively. Approximately 2.2– and 3.5-fold increases in LDH leakage...
140 citations
TL;DR: The results suggest that L-DOPA and dopamine might have a complex mixture of pro- and anti- oxidant effects, which could contribute to tissue damage due to oxidative stress in Parkinson's disease and other neurological disorders.
Abstract: The antioxidant and pro-oxidant properties of L-DOPA and dopamine were investigated in vitro. Both compounds inhibited the peroxidation of ox-brain phospholipids, with IC50 values of 8.5 microM for dopamine and 450 microM for L-DOPA. Dopamine and L-DOPA reacted with trichloromethyl peroxyl radicals (CCl3O2.) with rate constants of 2.1 x 10(7)M-1s-1 and 1.3 x 10(7)M-1s-1 respectively. The effects of dopamine and L-DOPA on iron ion-dependent hydroxyl radical generation from H2O2 were complex. In general, low concentrations stimulated OH. formation in the presence of ferric-EDTA and, in the case of L-DOPA, ferric-ADP and ferric citrate chelates. Both compounds also reacted with superoxide radical and hypochlorous acid. The products of the reaction with HOCl could still inhibit alpha 1-antiproteinase and appear to be 'long lived' chloramine-type oxidizing species. Our results suggest that L-DOPA and dopamine might have a complex mixture of pro- and anti- oxidant effects, which could contribute to tissue damage due to oxidative stress in Parkinson's disease and other neurological disorders.
137 citations
TL;DR: Findings support the notion of enhanced oxidative stress in the embryo as an etiologic agent in diabetic teratogenesis as well as the regulation of these enzymes occurs primarily at the pretranslational level.
Abstract: Maternal diabetes during pregnancy is associated with an increased rate of congenital malformations in the offspring. The exact molecular etiology of the disturbed embryogenesis is unknown, but an involvement of radical oxygen species in the teratological process has been suggested. Oxidative damage presupposes an imbalance between the activity of the free oxygen radicals and the antioxidant defence mechanisms on the cellular level. The aim of the present study was to investigate if maternal diabetes in vivo, or high glucose in vitro alters the expression of the free oxygen radical scavenging enzymes superoxide dismutase (CuZnSOD and MnSOD), catalase and glutathione peroxidase in rat embryos during late organogenesis. We studied offspring of normal and diabetic rats on gestational days 11 and 12, and also evaluated day-11 embryos after a 48 hour culture period in 10 m M or 50 mM glucose concentration. Both maternal diabetes and high glucose culture caused growth retardation and increased rate of congenita...
TL;DR: The purpose of this review is to bring together the different approaches for studying the oxidation of low density lipoproteins and try to identify some critical factors which will permit greater comparability between laboratories.
Abstract: The purpose of this review is to bring together the different approaches for studying the oxidation of low density lipoproteins and try to identify some critical factors which will permit greater comparability between laboratories. These issues are discussed both in terms of the variety of exogenous mediators of oxidation applied (transition metal ions, haem proteins, azo initiators, peroxynitrite, cells etc.) and their raisons d'etre, as well as the methodologies (formation of conjugated dienes, hydroperoxides, decomposition products of lipid peroxidation, altered surface charge, macrophage uptake) applicable to the different stages of the oxidation and the factors underlying their accurate execution and interpretation.
TL;DR: It is concluded that sunlight influences the beta-carotene and alpha-tocopherol content of blood and tissues.
Abstract: We conducted a randomized placebo-controlled double-blind study in 20 healthy young female students (skin type II + III, body mass index 18–22) in order to evaluate the efficacy of 10 weeks of mode
TL;DR: The biological role of the soxRS regulon of Escherichia coli, which is involved in the adaptation toward oxidative stress, is presumably to counteract the oxidative inactivation of the iron clusters and the subsequent release of iron with consequent increased rate of production of HO.
Abstract: The in vivo production of HO- requires iron ions, H2O2 and O2- or other oxidants but probably does not occur through the Haber-Weiss reaction. Instead oxidants, such as O2-, increase free iron by releasing Fe(II) from the iron-sulfur clusters of dehydratases and by interfering with the iron-sulfur clusters reassembly. Fe(II) then reduces H2O2, and in turn Fe(III) and the oxidized cluster are re-reduced by cellular reductants such as NADPH and glutathione. In this way, SOD cooperates with cellular reductants in keeping the iron-sulfur clusters intact and the rate of HO- production to a minimum.O2- and other oxidants can release iron from Fe(II)-containing enzymes as well as copper from thionein. The released Fe(III) and Cu(II) are then reduced to Fe(II) and Cu(I) and can then participate in the Fenton reaction.In mammalian cells oxidants are able to convert cytosolic aconitase into active IRE-BP, which increases the “free” iron concentration intracellularly both by decreasing the biosynthesis of ferritin a...
TL;DR: This paper examined the free radical-scavenging properties of representative extracts and of purified glucosinolates from cruciferous vegetables, by measuring their effect on ascorbate- or NADPH/iron-induced peroxidation of human liver microsomes, and showed that the glucosinsolate content appeared to play only a minor role in these effects.
Abstract: Fruits and vegetables contain several classes of compounds that can potentially contribute to antioxidant activity, including vitamins, simple and complex phenolics, sulphur-containing compounds and glucosinolates. The glucosinolates are found in high concentration in many cruciferous vegetables, and it is well established that their breakdown products induce endogenous antioxidant defences such as quinone reductase and glutathione S-transferase in cells and in vivo. Despite the anticarcinogenic effect of these compounds in animal models, the direct antioxidant properties of this class of compounds have not been systematically studied. We therefore examined the free radical-scavenging properties of representative extracts and of purified glucosinolates from cruciferous vegetables, by measuring their effect on ascorbate- or NADPH/iron-induced peroxidation of human liver microsomes, ascorbate/iron-induced peroxidation on phospholipid liposomes, iron chelation and hydroxyl radical scavenging using the deoxyribose assay, total antioxidant potential using ABTS (2,2'-azinobis(3-ethyl-benzothiazoline-6-sulphonate)) and the bleomycin assay. Most of the extracts from cruciferous vegetables exhibited some antioxidant properties, although extracts from cooked Brussels sprouts increased the rate of microsomal lipid peroxidation. The effects in these assays were dependent upon processing and species of crucifer, and the glucosinolate content appeared to play a minor role in these effects, since purified glucosinolates exhibited only weak antioxidant properties. The total antioxidant activities of extracts from cooked and autolysed Brussels sprouts were identical within experimental error. This is probably due to the content of phenolics which is unaltered by autolysis, despite the differences between these samples in other assays especially NADPH-iron-induced lipid peroxidation of human liver microsomes. The results demonstrate that glucosinolates are unlikely to account for the direct antioxidant effects of extracts from cruciferous vegetables.
TL;DR: This work has studied the reaction between hydrogen peroxide and purified (catalase free) human metHbA, and found that maximum ferryl levels could be obtained at a 1:1 stoichiometric ratio of haem to H2O2.
Abstract: The formation and reactivity of ferryl haemoglobin (and myoglobin), which occurs on addition of H2O2, has been proposed as a mechanism contributing to oxidative stress associated with human diseases. However, relatively little is known of the reaction between hydrogen peroxide and human haemoglobin. We have studied the reaction between hydrogen peroxide and purified (catalase free) human metHbA. Addition of H2O2 resulted in production of both ferryl haem iron (detected by optical spectroscopy) and an associated protein radical (detected by EPR spectroscopy). Titrating metHbA with H2O2 showed that maximum ferryl levels could be obtained at a 1:1 stoichiometric ratio of haem to H2O2. No oxygen was evolved during the reaction, indicating that human metHbA does itself not possess catalatic activity. The protein radicals obtained in this reaction reached a steady state concentration, during hydrogen peroxide decomposition, but started to decay once the hydrogen peroxide had been completely exhausted. The prese...
TL;DR: There was a five-fold increase in mitochondrial reactive oxygen species production in whole blood and monocytes from patients with rheumatoid arthritis, when compared to healthy subjects or patients with non-rheumatic diseases.
Abstract: Mitochondrial dysfunction contributes to cell damage in a number of human diseases. One significant mechanism by which mitochondria damage cells is by producing reactive oxygen species from the respiratory chain. In this study we measured the production of reactive oxygen species by leukocyte mitochondria in blood from rheumatoid arthritis patients. To do this we used the chemiluminescence of lucigenin, which is accumulated by mitochondria within cells and reacts with superoxide to form a chemiluminescent product. By using specific inhibitors we could distinguish between the production of reactive oxygen species by mitochondria and by NADPH oxidase. There was a five-fold increase in mitochondrial reactive oxygen species production in whole blood and monocytes from patients with rheumatoid arthritis, when compared to healthy subjects or patients with non-rheumatic diseases. There was no increases in mitochondrial reactive oxygen species production by neutrophils from rheumatoid arthritis patients. The enhanced mitochondrial radical production in rheumatoid arthritis patients correlated significantly with increased levels of tumor necrosis factor alpha in plasma (p < 0.0001). As tumor necrosis factor alpha is known to increase mitochondrial reactive oxygen species production the elevated mitochondrial radical formation seen in rheumatoid arthritis patients may be due to activation of the mitochondrial radical production. These data suggest that elevated mitochondrial oxidative stress contributes to the pathology of rheumatoid arthritis.
TL;DR: In this article, the authors found that the presence of H2O2 at 100 μM or 1 mM to the culture medium of BHK-21 fibroblasts results in increased intracellular levels of H 2O2, leading to an increase in the appearance of apoptotic-like cells in the cultures.
Abstract: Addition of H2O2 at 100 μM, or 1 mM, to the culture medium of BHK-21 fibroblasts results in increased intracellular levels of H2O2. Whilst exposure of BHK-21 cells to lower levels of H2O2 (1 μM) actually stimulates proliferation, these higher oxidant concentrations not only depress proliferation rates but also lead to an increase in the appearance of apoptotic-like cells in the cultures. Other agents such as inhibitors of glutathione peroxidase and catalase, or mimics of superoxide dismutase, which also bring about elevated cellular levels of H2O2 in BHK-21 cells, similarly lead to decreased proliferation and an apparent increase in cells with apoptopic features. Thus intracellular conditions which are considered more prooxidant than normal, appear to favour apoptosis over proliferation in BHK-21 fibroblasts. Additionally these abnormal cellular conditions also appear to favour excessive DNA replication, in remaining non-apoptotic cells.
TL;DR: It is suggested that NO is beneficial to sperm viability and motility in both fertile and infertile individuals, and that reduction of lipid peroxidative damage to sperm membranes and increase of intracellular cGMP may be involved in these benefits.
Abstract: The capacity of human sperm fertilization is principally dependent on sperm motility and membrane integrity. Oxygen-derived free radicals, such as superoxide anion, are known to impair sperm motility and membrane integrity by inducing membrane lipid peroxidation (LPO). Nitric oxide (NO), a biologically active free radical, has recently been shown to inactivate superoxide and increase intracellular guanosine-3', 5'-cyclic monophosphate (cGMP). The aim of this study is to investigate the effects of NO on human sperm motility, viability, lipid peroxidation and cGMP in fertile and asthenozoospermic infertile individuals in vitro. Semen samples were obtained from 10 fertile volunteers and 10 asthenozoospermic infertile patients. Washed spermatozoa were incubated at 37 degrees C in Ham's F-10 medium with 0, 25, 50, 100, 200, or 400nM sodium nitroprusside (SNP, Na2 [Fe(CN) 5NO] 2H2O), a nitric oxide releaser. Samples were analyzed for viability, determined by eosin-Y dye exclusion method at 0, 1, 2, 3, 5 and 6 h of incubation; motility, determined by the trans-membrane migration method within 2 h of incubation; LPO determined by malondialdehyde (MDA)-thiobarbituric acid method at 3 h of incubation; and the intracellular cGMP, determined by 125I-cGMP radioimmunoassay at 3 h of incubation. The results showed: in both fertile and infertile samples, viability, trans-membrane migration ratio and the levels of intracellular cGMP in 25-100nM SNP-treated spermatozoa were significantly higher than those in control groups, while MDA contents in treated groups were significantly lower than those in controls. However, when concentrations of SNP increased to 200-400nM, the opposite effects were exhibited. The effects of SNP on these processes were biphasic within 25-400nM. The most effective concentration was 100nM. These data suggested that NO is beneficial to sperm viability and motility in both fertile and infertile individuals, and that reduction of lipid peroxidative damage to sperm membranes and increase of intracellular cGMP may be involved in these benefits.
TL;DR: In this article, the authors show that the actual reported levels of brain protein carbonyls vary over a wide range, probably due to the use of different protocols for the carbonyl assay; results differ depending on when the dinitrophenylhydrazine reagent is added and at what stage in the procedure protein is assayed for the calculation of carbs on a unit protein basis.
Abstract: It has been suggested in the literature that elevated oxidative protein damage, measured as protein carbonyls, is present in the nervous system of patients with sporadic motor neurone disease (MND). However, the actual reported levels of brain protein carbonyls vary over a wide range. We show here that this is probably due to the use of different protocols for the carbonyl assay; results differ depending on when the dinitrophenylhydrazine reagent is added and at what stage in the procedure protein is assayed for the calculation of carbonyls on a unit protein basis. Using a range of different procedures, we were unable to confirm reports of elevated protein carbonyls in motor cortex from brains of patients with MND. We also measured thiobarbituric acid-reactive material in the brain samples using an HPLC-based TBA test in the presence of butylated hydroxytoluene. In general, there was no significant elevation of TBARS in MND motor cortex. However, four patients showed values higher than any of the control ...
TL;DR: It was found that the efficacy of radical scavenging by alpha-tocopherol became smaller as the radical went deeper into the interior of LDL, and the tocopherol mediated propagation was observed notably at low rate of radical flux, but this was suppressed by reductant such as ascorbic acid and ubiquinol.
Abstract: The oxidation of low density lipoprotein (LDL) was carried out aiming specifically at elucidating the anti-oxidant action of alpha-tocopherol. Lipophilic and hydrophilic azo compounds and copper induced the oxidation of LDL similarly to give cholesterol ester and phosphatidylcholine hydroperoxides as major products. The antioxidant potency of alpha-tocopherol in LDL was much poorer than in homogeneous solution. Doxyl stearic acids were used as spin probe and incorporated in LDL. The rate of reduction of doxyl nitroxide in LDL by ascorbate decreased with increasing distance from the LDL surface. From the competition between the spin probe and alpha-tocopherol in scavenging radical, it was found that the efficacy of radical scavenging by alpha-tocopherol became smaller as the radical went deeper into the interior of LDL. On the other hand, 2,2,5,7,8-pentamethyl-6-chromal spared the spin label regardless of the position of nitroxide. The antioxidant activity of chromanols against LDL oxidation increased with decreasing length of isoprenoid side chain at the 2-position. All these results were interpreted by location and low mobility of alpha-tocopherol in LDL. The tocopherol mediated propagation was observed notably at low rate of radical flux, but this was suppressed by reductant such as ascorbic acid and ubiquinol.
TL;DR: DDC is a powerful reductant and antioxidant since it scavenges hypochlorous acid, hydroxyl radical and peroxynitrite, which may provide an explanation for the apparent beneficial effects of DDC against oxidative stress-related diseases that have been observed in experimental and clinical studies.
Abstract: Diethyldithiocarbamate (DDC), a potent copper chelating agent, has long been used for the treatment of oxygen toxicity to the central nervous system, as an immunomodulator to treat cancer, and in HIV-infected patients. We evaluated the antioxidant properties of DDC, including its scavenging of reactive oxygen species, its reducing properties, its iron-chelating properties, and its protective effects on oxidant-induced damage to brain tissue, protein, human LDL, and DNA. It is found that DDC is a powerful reductant and antioxidant since it scavenges hypochlorous acid, hydroxyl radical and peroxynitrite; it chelates, then oxidizes ferrous ions; it blocks the generation of hydroxyl radicals and inhibits oxidative damage to deoxyribose, protein, DNA, and human LDL. These findings may provide an explanation for the apparent beneficial effects of DDC against oxidative stress-related diseases that have been observed in experimental and clinical studies.
TL;DR: Reduction of iron (IV) in ferrylmyoglobin in the presence of beta-lactoglobulin in aqueous solution is the result of two parallel reactions: a so-called autoreduction, and a second-order-reaction resulting in bityrosine formation in beta- lactoglOBulin.
Abstract: Reduction of iron (IV) in ferrylmyoglobin in the presence of P-lactoglobulin in aqueous solution is the result of two parallel reactions: (i) a so-called autoreduction, and (ii) reduction by β-lactoglobulin in a second-order-reaction resulting in bityrosine formation in β lactoglobulin. In the pH-region investigated (5.4–7.4), the rate of reduction increased for both reactions with decreasing pH. The second order-reaction had for non-denatured β-lactoglobulin the activation parameters: AH* = 45 kJ.mol-1 and AS* = -93J mol-1.K-1 at pH = 7.0 and ionic strength 0.16 (NaCl). Reduction of ferrylmyoglobin by β-lactoglobulin denatured by heat (86°C for 3 min) or by hydrostatic pressure (300 MPa for 15 min) resulted in formation of higher molecular weight species as detected by size-exclusion chromatography and by SDS-PAGE. No molecular weight changes were observed for reduction of ferrylmyoglobin by native P-lactoglobulin. Detection of bityrosine in the native β-lactoglobulin fraction after oxidation with ferryl...
TL;DR: Data indicate that the more heat-tolerant cultivars had an enhanced capacity for scavenging active oxygen species and a more active ascorbate-glutathione cycle and suggest a strong correlation between the ability to up-regulate the antioxidant defense system and the ability of tomatoes to produce greater yields when grown under heat stress.
Abstract: Four putative heat-tolerant tomato (Lycopersicum esculentum) cultivars (Tamasabro, Heat Wave, LHT-24, and Solar Set) and one putative heat-sensitive tomato cultivar (Floradade) were grown in the field under non-stress (average daily temperature of 26 degrees C) and heat-stress (average daily temperature of 34 degrees C) conditions. At anthesis, approximately five weeks after being transplanted to the field, leaf samples were collected for antioxidant analyses. Yield was determined by harvesting ripe fruit seven weeks after the collection of leaf samples. Heat stress resulted in a 79.1% decrease in yield for the heat-sensitive Floradade, while the fruit yield in the heat-tolerant cultivars Heat Wave, LHT-24, Solar Set, and Tamasabro was reduced 51.5%, 22.1%, 43.8%, and 34.8% respectively. When grown under heat stress, antioxidant activities were also greater in the heat-tolerant cultivars. Superoxide dismutase (SOD) activity increased up to 9-fold in the heat-tolerant cultivars but decreased 83.1% in the heat-sensitive Floradade. Catalase, peroxidase, and ascorbate peroxidase activity increased significantly in all cultivars. Only Heat Wave showed a significant increase in glutathione reductase in response to heat stress but all heat-tolerant cultivars exhibited significantly lower oxidized ascorbate/reduced ascorbate ratios, greater reduced glutathione/oxidized glutathione rations, and greater alpha-tocopherol concentrations compared to the heat-sensitive cultivar Floridade. These data indicate that the more heat-tolerant cultivars had an enhanced capacity for scavenging active oxygen species and a more active ascorbate-glutathione cycle and suggest a strong correlation between the ability to up-regulate the antioxidant defense system and the ability of tomatoes to produce greater yields when grown under heat stress.
TL;DR: The results argue against the participation of Fe3+ in the initiation of LOOH-independent lipid peroxidation and suggest its possible involvement in LooH-dependent lipidperoxidation.
Abstract: A study conducted on Fe2+ autoxidation showed that its rate was extremely slow at acidic pH values and increased by increasing the pH; it was stimulated by Fe3+ addition but the stimulation did not present a maximum at a Fe2+/Fe3+ ratio approaching 1:1. The species generated during Fe3+-catalyzed Fe2+ autoxidation was able to oxidize deoxyribose; the increased Fe2+ oxidation observed at higher pHs was paralleled by increased deoxyribose degradation. The species generated during Fe3+-catalyzed Fe2+ autoxidation could not initiate lipid peroxidation in phosphatidylcholine liposomes from which lipid hydroperoxides (LOOH) had been removed by treatment with triph-enylphosphine. Neither Fe2+ oxidation nor changes in the oxidation index of the liposomes due to lipid peroxidation were observed at pHs where the Fe3+ effect on Fe2+ autoxidation and on deoxyribose degradation was evident. In our experimental system, a Fe2+/Fe3+ ratio ranging from 1:3 to 2:1 was unable to initiate lipid peroxidation in LOOH-free phos...
TL;DR: Formation and decay of the near infrared absorbing species and bleaching of beta-carotene are independent of whether oxygen is present or absent in the solutions.
Abstract: Upon laser flash photolysis of β–carotene in chloroform instantaneous bleaching of β–carotene and concomitant formation of near infrared absorbing species are observed. One species, absorbing with maximum at 920 nm, is formed during the laser pulse (10 ns) and is practically gone in one millisecond, the decay showing a bi-exponential behaviour. The second species, absorbing with maximum at 1000 nm, is formed from the species absorbing at 920 nm by first order kinetics with a rate constant of 4.9·104 s−1 at 20°C. This second species decays by second order kinetics and is gone within a few milliseconds. An additional slow bleaching of β–carotene and formation of the species absorbing at 920 nm is observed. This slow bleaching/formation of transient absorption is probably due to processes involving free radicals generated during the instantaneous bleaching. The species absorbing at 920 nm is suggested to be either (i) a free radical adduct formed from β–carotene and chloroform or (ii) β–carotene after abstra...
TL;DR: The results suggested that only capacitated spermatozoa themselves are able to generate O2- which stimulated their capacitation in turn, and it may be possible to utilize the inhibitory effect of SOD on sperm capacitation so as to regulate fertilization.
Abstract: Capacitation of spermatozoa is an essential procedure for fertilization. Capacitated spermatozoa have an in crease in the intracellular cAMP and acrosome reaction (AR) occurs immediately. The effect of exogenous superoxide anion (O−2) on the level of intracellular c AMP and the percentages of both spontaneous AR and lysophosphatidylcholine-induced AR (LPC-AR) were studied using semen samples collected from 10 healthy and fertile volunteers working or studying in Lanzhou Medical College. Spermatozoa were separated by Percoll and incubated at 37°C in Ham's F-10 medium with O−2 generation system: xanthine + xanthine oxidase + catalase + diethylenetriaminepentaacetic acid + sodium formate. The intracellular cAMP was deter mined by (3H)— cAMP radioimmunoassay at 3 h of incubation, and the percentages of AR and LPC-AR were evaluated by the triple-stain technique at 3.5 h of incubation. The effects of SOD with different concentration were also determined. The results showed: the level of intracellular CAMP (pmol...
TL;DR: It is indicated that MCI-186 reduces reperfusion injury in isolated perfused rat hearts with prolonged ischemia and the effect is more closely related to the reduction of myocyte damage than the preservation of the coronary circulation.
Abstract: MCI-186 (3-methyl-1-phenyl-2-pyrazolin-5-one) is a newly developed antioxidant which has been shown to reduce brain edema in cerebral ischemia through inhibition of the lipoxygenase pathway of arachidonic acid. However, its effect on myocardial reperfusion injury after prolonged ischemia has not yet been demonstrated. We compared the mode of the effect of MCI-186 and recombinant human CuZn superoxide dismutase (rh-SOD) in isolated perfused rat hearts subjected to 60-min ischemia followed by 60-min reperfusion. Left ventricular developed pressure (LVDP), necrotic area and the release of creatine phosphokinase (CPK) and endogenous CuZn superoxide dismutase (endoge-SOD) were measured to evaluate myocardial damage. The decrease in left coronary flow (CBF) was measured as an index of the damage of left coronary circulation. MCI-186 (17.5 mg/L) was perfused for 10 min in the MCI group and rh-SOD (70 mg/L) was perfused during the reperfusion period in the SOD group starting 5 min prior to reperfusion. The releas...
TL;DR: Phenoxyl radicals generated by laser flash photolysis were found to react with beta-carotene with concomitant beta-Carotene bleaching in two parallel reactions with similar rates, and may prove important for an understanding of how beta- carotene acts as an antioxidant.
Abstract: Phenoxyl radicals generated by laser flash photolysis were found to react with beta-carotene with concomitant beta-carotene bleaching in two parallel reactions with similar rates: (i) formation of a beta-carotene adduct with a (pseudo) first order rate constant of 1-15 x 10(4) s-1 with absorption maximum around 800 nm, and (ii) formation of a beta-carotene radical cation with a (pseudo) first order rate constant of 2-3 x 10(4) s-1 with absorption maximum around 920 nm Both beta-carotene radicals decay on a similar time scale and have virtually disappeared after 100 ms, the beta-carotene adduct by a second order process Oxygen had no effect on beta-carotene bleaching or radical formation and decay The reduction of phenoxyl radicals by beta-carotene may prove important for an understanding of how beta-carotene acts as an antioxidant
TL;DR: The results suggest that isoprenoids of VE and CoQ participate in the inhibition of the NADPH oxidase activation system through modulation of the neutrophil membrane probably by the inhibitory activity of PKC.
Abstract: Effects of various derivatives of α-tocopherol (VE) and coenzyme Q (CoQ) on superoxide (O2*-) generation of neutrophils and protein kinase C (PKC) activity were examined. VE and CoQ8 inhibited O2*- generation of neutrophils stimulated by a protein kinase C mediated process monitored by cytochrome c reduction and spin trapping methods. The inhibitory action was observed not only with α-tocopherol, but also with β-, γ-, dL-tocopherols and with tocol which is a chemical similar to VE but lacking methyl groups on the chromanol ring structure and which is not a radical scavenger. By con trast, no inhibition was observed with 2-carboxy-2,5,7,8-tetramethyl-6-chromanol (CTMC, trolox) or 2,2,5,7,8,-pentamethyl-6-chromanol (PMC) which are water soluble VE derivatives having radical scavenging activity. Compounds having a similar isoprenoid chain, such as CoQ, also have inhibitory activity on PKC-dependent O2*- generation of neutrophils. The inhibitory activity of CoQ derivatives is dependent on the length of the un...