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Gliclazide: a general free radical scavenger.

Free radical mechanisms have been implicated in diabetic microangiopathy. Agents that scavenge free radicals may be beneficial. We assessed the scavenging ability of two sulphonylureas, gliclazide and glibenclamide, in vitro. The assay which employs o-dianisidine sensitised by riboflavin can be used to distinguish between superoxide scavengers and general scavengers. The former species lead to an augmentation while the latter has an inhibitory effect. The drugs were added in final concentrations of 0.5, 1.0, 2.5 and 5.0 micrograms/ml. The percentage inhibition (mean +/- S.D.) for each concentration of gliclazide respectively was 11.0 +/- 2.5%, 20.8 +/- 2.9%, 31.4 +/- 2.2% and 47.2 +/- 0.8%. Glibenclamide had no scavenging effects. The results demonstrate that gliclazide is a powerful general free radical scavenger in vitro. We postulate that this scavenging quality of gliclazide may be important in diabetes.

Dianisidine↗

Photooxidation of epinephrine sensitized by methylene blue as an assay for the evaluation of alpha-mercaptopropionylglycine as a free radical scavenger.

Free radicals, which are instable intermediates of some biochemical reactions, are produced everywhere in the living matter. They may induce cell membrane lipid peroxidation, with non reversible letal effects. Alpha-mercaptopropionylglycine capability of scavenging free radicals has been tested. H2O2 has been used as a free radical donor under actinic light, epinephrine oxidation to adrenochrome as a revealing system and methylene blue as sensitizing agent. alpha-MPG at 6 mM concentration showed the maximal inhibition of free radical formation in our system. Higher concentrations were unable to produce further inhibition.

Amino Acids, Sulfur↗

Protection of the kidney after temporary ischemia: free radical scavengers.

Free radicals have been implicated in the damage caused by tissue ischemia and reperfusion. Canine kidneys were subjected to a 60-minute period of normothermic ischemia. One group of animals received intra-arterial superoxide dismutase (SOD, 17.6 mg/50 ml, 50,000 units) at the end of the ischemic period, whereas a second group received albumin (17.6 mg/50 ml). The kidneys treated with SOD demonstrated significantly less edema formation (1.0 +/- 0.3% wet weight vs. 1.8 +/- 0.2%, mean +/- SEM, p less than 0.05) and lower renovascular resistance (44.0 +/- 5.6 dynes-sec/cm5 vs. 64.0 +/- 12.0, p less than 0.05). The SOD group displayed greater preservation of both glomerular filtration rate (45.9 +/- 6.1% of baseline vs. 23.7 +/- 5.8%, p less than 0.05) and urine flow (1.3 +/- 0.4 ml/min vs. 0.3 +/- 0.1, p less than 0.05). We conclude that the free radical scavenger SOD provides significant protection of the kidney during ischemia and reperfusion.

Albumins↗

Intravitreal U75412E: a new free radical scavenger.

Free-radical-mediated tissue damage is thought to be involved in a multitude of ophthalmic pathologies. Therefore, drugs that scavenge free radicals may find wide clinical application. A new chemical class of antiinflammatory agents, the 21-aminosteroids, has been developed as free radical scavengers. These agents are thought to inhibit lipid peroxidation and prevent the release of free arachidonic acid from injured cell membranes. In the present study, we investigated the intraocular toxicity of one of the 21-aminosteroids, U75412E. Intravitreal doses of up to 0.1 mg of U75412E were nontoxic to rabbit ocular tissues.

Animals↗

In vitro free radical scavenging activity of hepatic metallothionein induced in an Indian freshwater fish, Channa punctata Bloch.

Mammalian metallothioneins (MT) have been reported to scavenge free radicals. There is no experimental evidence to show that fish MT has a similar property. In the present study cadmium-induced MT (Cd-MT) from the liver of an Indian freshwater fish Channa punctata Bloch was investigated for its free radical scavenging activity using three different in vitro assays. Exposure to cadmium chloride (0.2 mg/kg body weight; three doses on alternate days) resulted in a marked induction of Cd-MT in liver. Only a single isoform of Cd-MT was found to be induced. Molecular weight of Cd-MT was found to be 14 kDa as deduced by SDS-PAGE analysis. The purified Cd-MT effectively scavenged the following free radicals: superoxide radical (O2*-), 2,2'-azinobis 3-ethylbenzothiazoline-6-sulfonic acid (ABTS*+) and 1,1-diphenyl-picrylhydrazyl radical (DPPH*). The radical scavenging effect was found to be concentration-dependent. Also, the purified MT exhibited an inhibitory effect on ferric nitrilotriacetate (Fe-NTA) induced oxidative DNA damage in vitro. The cysteine residues of MT are proposed to be the main candidate for its radical scavenging activity. Findings of the present study strongly suggest a free radical scavenging role for fish MT. Present study adds to the little existing knowledge about fish MT and its possible biological functions.

Animals↗

The anticonvulsant zonisamide scavenges free radicals.

Zonisamide (ZNS) is effective in animal models of epilepsy and epileptic patients. The free radical scavenging activities of ZNS were analyzed by using electron spin resonance. ZNS, in the mmolar range, scavenged hydroxyl and nitric oxide radicals in a dose-dependent manner. This suggests that the mechanism of antiepileptic effect of ZNS may involve protection of neurons from free radical damage and stabilization of neuronal membranes.

Anticonvulsants↗

Antioxidant activity of South African red and white cultivar wines: free radical scavenging.

The free radical scavenging activity of South African red (n = 46) and white (n = 40) cultivar wines was determined using 2,2'-azinobis(3-ethylbenzothialozinesulfonic acid) radical cations (ABTS(.+)) and 2,2-diphenyl-1-picrylhydrazyl radicals (DPPH.). The total antioxidant activities (TAA) of red and white wines using ABTS(.+) were 14.916 and 0.939 mM Trolox, respectively, at corresponding total phenol (TP) contents of 2339.0 and 273.8 mg of gallic acid equiv/L. Ruby Cabernet wines had the lowest TAA(ABTS) (13.177 mM Trolox) of the red wines, whereas the TAA(ABTS) values of Chardonnay and Chenin blanc wines were the highest (1.060 mM Trolox) and lowest (0.800 mM Trolox) of the white wines. The TAA(DPPH) values were of the same magnitude as the TAA(ABTS) values, and similar trends were observed. TAA correlated (P < 0.001) with total phenol content of red (r = 0.935) and white (r = 0.907) wines, as well as flavanol content of red wines (r = 0.866) and tartaric acid ester content of white wines (r = 0.767). Canonical discriminant analysis using phenolic composition and antioxidant activity was applied to differentiate between red and white cultivar wines.

Anthocyanins↗

Free radical scavenging by brain homogenate: implication to free radical damage and antioxidant defense in brain.

To study the mechanisms of free radical-induced brain damage and the antioxidant defense in the brain, we quantified the superoxide and hydroxyl radical scavenging effects of brain homogenate using electron spin resonance spectrometry. Brain homogenate was found to scavenge both superoxide and hydroxyl radicals in concentration-dependent fashion. Heat denaturation significantly decreased these scavenging effects. The ability of brain homogenate to scavenge free radicals implies that brain damage can be induced by free radicals since they are known to react virtually with any type of molecule such as nucleic acids, membrane lipids, and proteins in the brain. On the other hand, some molecules which can be regenerated or repaired after free radical scavenging are considered to be antioxidants which include both enzymatic and non-enzymatic antioxidants. Measurement of the decrease in antioxidant activity following heat denaturation suggests that the contribution of enzymatic antioxidants is about 20-40% in scavenging superoxide radicals and about 10-20% in scavenging hydroxyl radicals.

Animals↗

Free radical scavenging potential of L-proline: evidence from in vitro assays.

An assessment of the potential of proline to scavenge free radicals was made in a couple of in vitro assay systems, namely graft co-polymerization and autooxidation of pyrogallol. Both these assays are essentially dependent upon free radical mechanisms. Graft co-polymerization involved a ceric (Ce(4+)) ion- or gamma-radiation-induced grafting of methyl acrylate (MA) onto a cellulose backbone. The degree of grafting, measured gravimetrically, was taken as a measure of free radical generation. The gamma-radiation-dependent grafting was far greater than that due to Ce(4+) ions. Inclusion of proline in the assay, irrespective of the initiator used, led to suppression of grafting in a concentration-dependent manner indicating the ability of proline to scavenge free radicals. The gamma-radiation-dependent grafting was also suppressed by hydroquinone and glutathione but not by ascorbate, glycine and spermine. In contrast to graft co-polymerization, proline did not inhibit the autooxidation of pyrogallol, a reaction involving superoxide radical generation. A subset of data constitutes an evidence for the ability of proline to scavenge free radicals in vitro. It is implied by extension that free proline, known to accumulate in plant tissues during abiotic stresses, would contribute to scavenging of surplus free radicals produced under a variety of abiotic stresses.

Acrylates↗

beta-Amyloid toxicity in organotypic hippocampal cultures: protection by EUK-8, a synthetic catalytic free radical scavenger.

Oxygen free radicals have been proposed to mediate amyloid peptide (beta-AP)-induced neurotoxicity. To test this hypothesis, we evaluated the effects of EUK-8, a synthetic catalytic superoxide and hydrogen peroxide scavenger, on neuronal injury produced by beta-AP in organotypic hippocampal slice cultures. Cultures of equivalent postnatal day 35 (defined as mature) and 14 (defined as immature) were exposed to various concentrations of beta-AP (1-42 or 1-40) in the absence or presence of 25 microM EUK-8 for up to 72 hours. Neuronal injury was assessed by lactate dehydrogenase release and semiquantitative analysis of propidium iodide uptake at various times after the initiation of beta-AP exposure. Free radical production was inferred from the relative increase in dichlorofluorescein fluorescence, and the degree of lipid peroxidation was determined by assaying thiobarbituric acid-reactive substances. Treatment of mature cultures with beta-AP (50-250 microg/ml) in serum-free conditions resulted in a reproducible pattern of damage, causing a time-dependent increase in neuronal injury accompanied with formation of reactive oxygen species. However, immature cultures were entirely resistant to beta-AP-induced neurotoxicity and also demonstrated no dichlorofluorescein fluorescence or increased lipid peroxidation after beta-AP treatment. Moreover, mature slices exposed to beta-AP in the presence of 25 microM EUK-8 were significantly protected from beta-AP-induced neurotoxicity. EUK-8 also completely blocked beta-AP-induced free radical accumulation and lipid peroxidation. These results not only support a role for oxygen free radicals in beta-AP toxicity but also highlight the therapeutic potential of synthetic radical scavengers in Alzheimer disease.

Amyloid beta-Protein Precursor↗

Antioxidant and analgesic activities of turpentine of Pinus nigra Arn. subsp. pallsiana (Lamb.) Holmboe.

The aim of this study is to examine possible antioxidant and analgesic activities of turpentine exudes from Pinus nigra Arn. subsp. pallsiana (Lamb.) Holmboe (TPN). Total antioxidant activity, reducing power, superoxide anion radical scavenging, free radical scavenging, metal chelating, and hydrogen peroxide scavenging activities were studied. The total antioxidant activity increased with the increasing amount of extracts (100, 300, and 500 microg) added to linoleic acid emulsion. All of the doses of TPN showed higher antioxidant activity than alpha-tocopherol. The samples showed 49, 70, and 91% inhibition on peroxidation of linoleic acid emulsion, respectively. On the other hand, the 300 microg of alpha-tocopherol showed 40% inhibition on peroxidation of linoleic acid emulsion. There is correlation between antioxidant activity and the reducing power, superoxide anion radical scavenging, free radical scavenging, metal chelating, and hydrogen peroxide scavenging activities. Like antioxidant activity, the reducing power, superoxide anion radical scavenging, free radical scavenging, metal chelating, and hydrogen peroxide scavenging activities of TPN depending on concentration and increasing with increased concentration of TPN. These properties may be the major reasons for the inhibition of lipid peroxidation. The results obtained in the present study indicate that the TPN has a potential source of natural antioxidant. In addition, analgesic effect of TPN was investigated in present study and TPN had strong analgesic effect. The analgesic effect of TPN compared with metamizol as a standard analgesic compound.

Analgesics↗

Specificity of oxygen radical scavengers and assessment of free radical scavenger efficiency using luminol enhanced chemiluminescence.

The selective scavenging capacities of 19 important oxygen radical scavengers were determined by adding them individually to each of the four oxy radical standards, (superoxide, hydroxy, alkoxy and hydroperoxy, and singlet O2), calculating the percent chemiluminescence inhibited, and extrapolating O2 equivalents neutralized from baseline. The sensitivity (0.01 nm/ml) and selectivity of this method not only allows identification of individual oxygen free radical species but also quantitates the efficiency of free radical scavengers.

Benzofurans↗

Vitamin E analogues reduce the incidence of ventricular fibrillations and scavenge free radicals.

The aim of our study was to analyse the protective effects of different alpha-tocopherol analogues 1) against fibrillations induced by an ischemia-reperfusion sequence, and 2) to further investigate in vitro the radical scavenging properties of these analogues by two sensitive methods. Concerning 1: isolated rat hearts underwent 10 min of coronary ligation followed by reperfusion and the alpha-tocopherol analogues were infused 15 min before occlusion. Functional parameters including heart rate and fibrillations were recorded. Concerning 2: the beta-phycoerythrin assay was utilised to determine the oxygen radical absorbing capacity (ORAC) of these vitamin E analogues against peroxyl radicals. Electron paramagnetic resonance (EPR) was used to measure their scavenger abilities on hydroxyl radical and superoxide anion production. Concerning 1: ventricular fibrillation times were reduced for all analogues treated hearts at concentrations of 1 microM and 5 microM, with Trolox being the most efficacious. Concerning 2: in our experimental conditions of intense production of free radicals, scavenging IC50 values for hydroxyl radical were 1.15, 2.17 and 4.04 mM for Trolox, MDL 74270 and MDL 74366 respectively. Superoxide anion IC50 values were 1.0 and 6.75 mM for Trolox and MDL 74270. Our results show that water-soluble analogues of vitamin E are effective in the prevention of coronary ligation induced reperfusion arrhythmia, under our experimental conditions. Moreover, our data demonstrate that these vitamin E analogues are effective scavengers for a variety of radicals. Our studies support the view that compounds that can either inhibit the formation or scavenge free radicals can protect the heart against arrhythmia associated with ischemia-reperfusion.

Animals↗

Characterization of the total free radical scavenger capacity of vegetable oils and oil fractions using 2,2-diphenyl-1-picrylhydrazyl radical.

The total free radical scavenger capacity (RSC) of 57 edible oils from different sources was studied: olive (24 brands of oils), sunflower (6), safflower (2), rapeseed (3), soybean (3), linseed (2), corn (3), hazelnut (2), walnut (2), sesame (2), almond (2), mixture of oils for salad (2), "dietetic" oil (2), and peanut (2). Olive oils were also studied according to their geographical origins (France, Greece, Italy, Morocco, Spain, and Turkey). RSC was determined spectrophotometrically by measuring the disappearance of the radical 2,2-diphenyl-1-picrylhydrazyl radical (DPPH(*)) at 515 nm. The disappearance of the radical followed a double-exponential equation in the presence of oils and oil fractions, which suggested the presence of two (fast and slow) groups of antioxidants. RSC was studied for the methanol-soluble phase ("methanolic fraction", MF) of the oil, the fraction nonsoluble in methanol ("lipidic fraction", LF), and the nonfractionated oil ("total oil"; TF = MF + LF). Only olive, linseed, rapeseed, safflower, sesame, and walnut oils showed significant RSC in the MF due to the presence of phenolic compounds. No significant differences were found in the RSC of olive oils from different geographical origins. Upon heating at 180 degrees C the apparent constant for the disappearance of RSC (k(T)) and the half-life (t1/2) of RSC for MF, LF, and TF were calculated. The second-order rate constants (k2) for the antiradical activity of some phenolic compounds present in oils are also reported.

Antioxidants↗

Free radical scavenging activity of fullerenol on the ischemia-reperfusion intestine in dogs.

Fullerenol, a water-soluble C(60)-fullerene derivative, has been demonstrated to have the capability to scavenge free radicals in vitro and in vivo. The purpose of this study was to investigate whether fullerenol can scavenge the free radicals that are massively induced during ischemia-reperfusion (I/R) injury of the small intestine, either preventively or therapeutically. Clamping the superior mesenteric artery and vein for 60 minutes to induce I/R injury was performed on male mongrel dogs. Thirty dogs were divided into three groups (10 in each): The control (C) group received no medication; the preventive (P) group received fullerenol (1 mg/kg) intravenously 30 minutes before ischemia; the therapeutic (T) group received the same dose of fullerenol immediately after reperfusion. This study was an experimental randomized trial. Intestinal segments were obtained 10, 20, 30, and 60 minutes after reperfusion; and blood samples and specimens of major organs were taken 60 minutes after reperfusion. Concentrations of lipid peroxidation products, including conjugated diene (CD) and malondialdehyde (MDA), and the level of glutathione (GSH) in intestinal tissue were determined. Serum indicators of liver and renal function were measured. Histologic examination of the small intestine and major organs were also performed. A significant increase in intestinal MDA and CD contents was detected at 30 and 60 minutes after reperfusion. The tissue GSH content, in contrast, was decreased 60 minutes after reperfusion. Administration of fullerenol diminished these changes both preventively and therapeutically. Liver and renal functions were within normal limits in all groups. Moreover no obvious histopathologic additional damage could be found in either the P or the T group. It is suggested that fullerenol can be considered a powerful scavenger for the free radicals induced by I/R injury of the small intestine.

Alanine Transaminase↗

Hispidin analogs from the mushroom Inonotus xeranticus and their free radical scavenging activity.

Three new free radical scavengers were isolated from the methanolic extract of the fruiting bodies of Inonotus xeranticus (Hymenochaetaceae), along with the known compound davallialactone. Their structures were established as hispidin analogs by extensive NMR spectral data. Compounds 3 and 4 displayed significant scavenging activity against the superoxide radical anion, ABTS radical cation, and DPPH radical, while 1 and 2 exhibited potent antioxidative activity only against ABTS radical cation.

Agaricales↗

Reduction of traumatic brain injury-induced cerebral oedema by a free radical scavenger.

Oxygen derived free radicals have been proposed to be in part responsible for the cerebral oedema resulting from head injury. In the present study the effects of free radical suppression with MDL 74,180 (2,3-dihydro-2,2,4,6,7-pentamethyl-3-(4-methylpiperazino)-methyl-1 - benzofuran-5-ol dihydrochloride), an alpha-tocopherol analogue free radical scavenger, on the development of cerebral oedema resulting from head injury has been assessed. Fluid percussion head injury in rats caused a regional oedema 48 h after injury. Infusion of MDL 74,180 for 2 h after the injury significantly attenuated oedema development in a dose-related manner. Using magnetic resonance imaging, cerebral oedema development was monitored in head injured mice. Oedema was apparent 4 h after head injury and was greatest in the vicinity of the olfactory bulb and surrounding the ventricles. Treatment with MDL 74,180 (1-10 micrograms/kg intravenously, administered 3-5 min after the injury) significantly reduced the oedema development. MDL 74,180 is a potential treatment for the oedema caused as a result of head injury.

Animals↗

Delayed radioprotection by nuclear transcription factor kappaB -mediated induction of manganese superoxide dismutase in human microvascular endothelial cells after exposure to the free radical scavenger WR1065.

The free radical scavenger WR1065 (SH) is the active thiol form of the clinically approved cytoprotector amifostine. At doses of 40 microM and 4 mM it can activate the redox-sensitive nuclear transcription factor kappaB (NFkappaB) and elevate the expression of the antioxidant gene manganese superoxide dismutase (MnSOD) in human microvascular endothelial cells (HMEC). MnSOD contains binding motifs for a number of transcription factors other than NFkappaB and codes for a potent antioxidant enzyme localized in the mitochondria that is known to confer enhanced radiation resistance to cells. The purpose of this study was to determine the effect of WR1065 exposure on the various transcription factors known to affect MnSOD expression along with the subsequent kinetics of intracellular elevation of MnSOD protein levels and associated change in sensitivity to ionizing radiation in HMEC. Cells were grown to confluence and exposed to WR1065 for 30 min. Affects on the transcription factors AP1, AP2, CREB, NFkappaB, and Sp1 were monitored as a function of time ranging from 30 min to 4 h after drug exposure using a gel-shift assay. Only NFkappaB exhibited a marked activation and that occurred 30 min following the cessation of drug exposure. MnSOD protein levels, as determined by Western blot analysis, increased up to 16-fold over background control levels by 16 h following drug treatment, and remained at 10-fold or higher levels for an additional 32 h. MnSOD activity was evaluated using a gel-based assay and was found to be active throughout this time period. HMEC were irradiated with X-rays either in the presence of 40 microM or 4 mM WR1065 or 24 h after its removal when MnSOD levels were most elevated. No protection was observed for cells irradiated in the presence of 40 microM WR1065. In contrast, a 4 mM dose of WR1065 afforded an increase in cell survival by a factor of 2. A "delayed radioprotective" effect was, however, observed when cells were irradiated 24 h later, regardless of the concentration of WR1065 used. This effect is characterized as an increase in survival at the 2 Gy dose point, i.e., a 40% increase in survival, and an increase in the initial slope of the survival curve by a factor of 2. The anti-inflammatory sesquiterpene lactone, Helenalin, is an effective inhibitor of NFkappaB activation. HMEC were exposed to Helenalin for 2 h at a nontoxic concentration of 5 microM prior to exposure to WR1065. This treatment not only inhibited activation of NFkappaB by WR1065, but also inhibited the subsequent elevation of MnSOD and the delayed radioprotective effect. A persistent marked elevation of MnSOD in cells following their exposure to a thiol-containing reducing agent such as WR1065 can result in an elevated resistance to the cytotoxic effects of ionizing radiation and represents a novel radioprotection paradigm.

Blotting, Western↗