Scavenging of Superoxide Anion by Phosphorylethanolamine: Studies in Human Neutrophils and in a Cell Free System

On the basis of previous observations, we attempted to characterize the effects of various products of phospholipid hydrolysis on neutrophil (PMN) respiratory burst activity. We studied the effects of phos- phorylcholine (PC) and phosphorylethanoline (PE) on superoxide anion production in PMN and in a cell free system. We found that PE but not PC inhibited measured superoxide anion, but that this was not due to inhibition of cellular superoxide generation but to scavenging of generated superoxide anion. Further, utilizing a system based upon the photo-oxidation of O-dianisidine sensitized by riboflavin, we were able to determine that the scavenging effect of PE was not superoxide dismutase (SOD)-like but rather a general scavenging or glutathione (GSH)-like effect. These data underscore the importance of identifying the mechanism of inhibition of superoxide generation by putative inhibitors as being due to a direct cellular effect or to a scavenging property.     

高效抗氧化大型真菌的筛选

为获得高效抗氧化菌株,采用直接提取方式从20个大型真菌菌株菌丝体培养液中提取抗氧化活性物质,采用邻苯三酚自氧化法、水杨酸法、DPPH法测定各菌株菌丝体培养粗提液对超氧阴离子自由基(O2)、羟基自由基.(OH)及1,1-二苯代苦味酰基自由基(DPPH-)的清除作用。结果表明:各菌株菌丝体培养粗提液对(O2)、.OH及DPPH-均有一定的清除作用,其中菌株NG菌丝体培养粗提液对OH-的清除效果最好,清除率为75.56%;菌株02菌丝体培养粗提液对(O2-)的清除效果最好,清除率为37.51%;菌株EG菌丝体培养粗提液对DPPH-清除效果最好,清除率为66.91%。     

Development of pyridine-containing macrocyclic copper(II) complexes: potential role in the redox modulation of oxaliplatin toxicity in human breast cells

Abstract

The unique redox and catalytic chemistry of Cu has justified the development of novel Cu complexes for different therapeutic uses including cancer therapy. In this work, four pyridine-containing aza-macrocyclic copper(II) complexes were prepared (CuL1–CuL4) varying in ring size and/or substituents and their superoxide scavenging activity evaluated. CuL3, the most active superoxide scavenger, was further studied as a modulator of the cytotoxicity of oxaliplatin in epithelial breast MCF10A cells and in MCF7 breast cancer cells. Our results show that CuL3 enhances the therapeutic window of oxaliplatin, by both protecting non-tumour cells and increasing its cytotoxic effect in breast carcinoma cells. CuL3 is thus a promising complex to be further studied and to be used as a lead compound for the optimization of novel chemotherapy sensitizers.     

Superoxide scavenging activity of BG-104 before and after sonication-determined by ESR spin trapping method

BG-104 is a compound of Chinese herbs which were treated with infrared ray-heating and brewed with microorganisms resulting in activation of actual principles contained in the herbs, and its effectiveness has been reported on various inflammatory disorders such as Behcet's disease, rheumatoid arthritis, bacterial infections and so on. In addition, it is noted as an effective agent to retard cancer progress. This report shows a superoxide scavenging activity of BG-104 using an ESR spin trapping method. For the measurement of superoxide scavenging activity, recombinant human superoxide dismutase is used as a standard. Superoxide scavenging activity of BG-104 increases in a dose-dependent manner and activity is significantly augmented by its sonication.     

Attenuation of oxidative stress by Allylpyrocatechol in synovial cellular infiltrate of patients with Rheumatoid Arthritis

Abstract

Free radicals are involved in the pathogenesis of Rheumatoid arthritis, a systemic autoimmune disorder characterized by unchecked synovial inflammation. Allylpyrocatechol, a phytoconstituent of Piper betle leaves, has potent anti-inflammatory activity and this study evaluated its anti-oxidant effect on the synovial infiltrate of patients with Rheumatoid arthritis. The ex vivo effect of allylpyrocatechol upon generation of reactive oxygen species in neutrophils, macrophages and lymphocytes was measured by flow cytometry using dichlorodihydrofluorescein diacetate, wherein it significantly decreased basal levels as also scavenged phorbol myristate acetate generated reactive oxygen species. Furthermore, its effect on generation of superoxide and hydroxyl radicals produced within infiltrated neutrophils was measured by cytochrome c and deoxyribose assay, respectively. Allylpyrocatechol significantly scavenged superoxide and hydroxyl radicals in infiltrated neutrophils. The effect of allylpyrocatechol on nitric oxide was measured in macrophages using 4,5-diaminofluorescein diacetate by flow cytometry wherein it decreased production of nitric oxide in infiltrated macrophages, which correlated with its in vitro nitric oxide scavenging activity. Taken together, this ex vivo study has established that allylpyrocatechol has potent scavenging activity and could be considered as an add-on therapy in the treatment of inflammation-associated disorders like Rheumatoid Arthritis.     

Ginseng extract exhibits antimutagenic activity against induced mutagenesis in various strains of Salmonella typhimurium

Ginseng has been reported to exhibit antioxidant and antimutagenic activity. The present study was undertaken with a view to confirm whether the antioxidant activity of Ginseng is responsible for its antimutagenic action. The concentrated root extract of Panax ginseng (Ginseng extract I) and its lyophilized powder (Ginseng extract II) obtained from two different manufacturing houses, were tested against mutagenesis using the well-standardized Ames microsomal test system. The extracts exhibited antimutagenic effect against hydrogen peroxide induced mutagenesis in TA100 strain, and against mutagenesis produced by 4-nitroquinoline-N-oxide in both TA98 and TA100 strains of Salmonella typhimurium. Both the extracts failed to show any antimutagenic potential against tert-butyl hydroperoxide (an oxidative mutagen) in TA102 strain, a strain highly sensitive to active oxygen species. The extracts also indicated a weak antioxidant activity in a series of in vitro test systems viz., 1,1-diphenyl picryl hydrazyl (DPPH) assay, hydrogen peroxide scavenging and superoxide anion scavenging. The results indicate that the protective effects shown by ginseng extract(s) against 4-nitroquinoline-n-oxide and hydrogen peroxide induced mutagenesis in TA98 and TA100 could mainly be due to its property to initiate and promote DNA repair rather than free radical scavenging action.     

杨桃提取物体外清除氧自由基作用

从杨桃果中提取得到三种提取物为匀浆提取物、蛋白提取物和多糖提取物。采用化学发光法测定这三种提取物清除氧自由基的活性,实验结果:匀浆提取物清除羟自由基(·OH)和H2O2的活性大小相近,而清除超氧阴离子自由基(O2–·)的活性较小,其IC50约为前两者的4倍。蛋白提取物清除O2–·和·OH的活性大小相近,而清除H2O2的活性明显小于前两者,IC50约为前两者的9倍。多糖提取物清除.OH的活性明显大于清除O2–·和H2O2的活性,其IC50约为O2–·的1/22,约为H2O2的1/65。结果表明,杨桃果具有清除O2–·、·OH和H2O2的作用,不同提取物对这些活性氧自由基的清除能力有所不同。     

Radical and Superoxide Scavenging Activities of Matairesinol and Oxidized Matairesinol

The radical and superoxide scavenging activities of oxidized matairesinols were examined. It could be assumed that the free benzylic position was important for higher radical scavenging activity. The different level of activity was observed between 7′-oxomatairesinol (Mat 2) and 7-oxomatairesinol (Mat 3). The activity of 8-hydroxymatairesinol was lower than that of matairesinol (Mat 1). The superoxide scavenging activity of the oxidized matairesinols was also demonstrated for the first time. It is assumed that the pKa value of phenol in the oxidized matairesinols affected this activity.     

迎春花色素提取及抗氧化性研究

目的:提取迎春花黄色素研究其抗氧化性能。方法:利用对羟基自由基(OH-·)和超氧阴离子自由基(O2-·)的清除能力研究迎春花色素的抗氧化性能。结果:随着迎春花色素量的增加,其清除OH-·和O2-·的能力逐渐提高,最高分别可达31.5%和91.0%。结论:迎春花色素对超氧阴离子和羟基自由基均具有较强的清除能力。     

Evaluation of Superoxide Scavenging Activities of Hamamelis Extract and Hamamelitannin

Hamamelitannin, which is a component of bark extract of hamamelis (Hamamelis virginior L.), was found to be a potent scavenger of superoxide anion radicals. Superoxide anion scavenging activity of the compound was evaluated by ESR-spin trap method using DMPO (5,5′-dimethyl-1-pyrroline-N-oxide) as a spin trapping agent. The IC₅₀ value (the concentration producing 50% inhibition of superoxide anion radicals) of hamamelitannin was found to be 1.38 ± 0.06 μM much lower than that of ascorbic acid (23.31 ± 2.23 μM). Supporting the superoxide scavenging activity of hamamelitannin, the compound showed both suppresive ability against depolymelization of hyaluronic acid and protective ability against cytotoxicity induced by superoxide anion radicals. Hamamelitannin increased the survival rate of fibroblast to 85.5 ± 3.3%, compared with that of control (27.2 ± 4.3%).     

Evaluation of Superoxide Scavenging Activities of Hamamelis Extract and Hamamelitannin

Hamamelitannin, which is a component of bark extract of hamamelis (Hamamelis virginior L.), was found to be a potent scavenger of superoxide anion radicals. Superoxide anion scavenging activity of the compound was evaluated by ESR-spin trap method using DMPO (5,5'-dimethyl-1-pyrroline-N-oxide) as a spin trapping agent. The IC₅₀ value (the concentration producing 50% inhibition of superoxide anion radicals) of hamamelitannin was found to be 1.38 ± 0.06 μM much lower than that of ascorbic acid (23.31 ± 2.23 μM). Supporting the superoxide scavenging activity of hamamelitannin, the compound showed both suppresive ability against depolymelization of hyaluronic acid and protective ability against cytotoxicity induced by superoxide anion radicals. Hamamelitannin increased the survival rate of fibroblast to 85.5 ± 3.3%, compared with that of control (27.2 ± 4.3%).     

Extracellular superoxide dismutase

The extracellular space is protected from oxidant stress by the antioxidant enzyme extracellular superoxide dismutase (EC-SOD), which is highly expressed in selected tissues including blood vessels, heart, lungs, kidney and placenta. EC-SOD contains a unique heparin-binding domain at its carboxy-terminus that establishes localization to the extracellular matrix where the enzyme scavenges superoxide anion. The EC-SOD heparin-binding domain can be removed by proteolytic cleavage, releasing active enzyme into the extracellular fluid. In addition to protecting against extracellular oxidative damage, EC-SOD, by scavenging superoxide, preserves nitric oxide bioactivity and facilitates hypoxia-induced gene expression. Loss of EC-SOD activity contributes to the pathogenesis of a number of diseases involving tissues with high levels of constitutive extracellular superoxide dismutase expression. A thorough understanding of the biological role of EC-SOD will be invaluable for developing novel therapies to prevent stress by extracellular oxidants.     

Scavenging Effect of Various Tea Extracts on Superoxide Derived from the Metabolism of Mutagens

We determine the superoxide formed in the self-degradation of mutagens activated by cytochrome enzymes and evaluated the scavenging effect of various tea extracts. Benzo[a]pyrene (B[a]P) and 2-amino-6-methyldipyrido(1,2-a:3′,2′-d)imidazole (Glu-P-1) each produced a large amount of superoxide after activation by cytochrome enzymes. However, 2-amino-3-methyl-imidazo(4,5-f)quinoline (IQ), 3-amino-1,4-dimethyl-5H-pyridol(4,3-b)indole (Trp-P-1) and alfatoxin B₁ (AFB₁) failed to generate a significant amount of superoxide. The addition of a tea extract to the reaction system marked inhibited the derivation of superoxide from Glu-P-1. However, the tea extracts showed weaker inhibition of the B[a]P-mediated formation of superoxide. Among the four teas tested, the oolong tea extract tended to exhibit the strongest inhibitory effect. Our results suggest that the chemopreventive efficacy of a tea extract is partly associated with its antioxidative activity.     

Detoxification of SO2 derivatives in chloroplasts ofHardwickia binata

Intact chloroplasts isolated from sulphur dioxide fumigatedHardwickia binata leaves showed inhibition of PS II electron transport activity without any significant effect on photosystem I. Sulphur dioxide exposed leaves accumulated more hydrogen peroxide than those from non-fumigated plants and this was caused by increase in superoxide radical production. Hydrogen peroxide formation was inhibited by addition of cytochrome C and superoxide disrnutase. In sulphur dioxide fumigated leaves, increase in superoxide dismutase activity showed resistance to sulphite toxicity. The localization of ascorbate peroxidase, glutathione reductase and dehydroascorbate reductase activities in chloroplasts provide evidence for the photogeneration of ascorbate. The scavenging of hydrogen peroxide in chloroplast due to ascorbate regenerated from DHA by the system: PS I → Fd → NADP → glutathione. The system can be considered as a means for preliminary detoxification of sulphur dioxide by chloroplasts     

Development of pyridine-containing macrocyclic copper(II) complexes: potential role in the redox modulation of oxaliplatin toxicity in human breast cells

The unique redox and catalytic chemistry of Cu has justified the development of novel Cu complexes for different therapeutic uses including cancer therapy. In this work, four pyridine-containing aza-macrocyclic copper(II) complexes were prepared (CuL1-CuL4) varying in ring size and/or substituents and their superoxide scavenging activity evaluated. CuL3, the most active superoxide scavenger, was further studied as a modulator of the cytotoxicity of oxaliplatin in epithelial breast MCF10A cells and in MCF7 breast cancer cells. Our results show that CuL3 enhances the therapeutic window of oxaliplatin, by both protecting non-tumour cells and increasing its cytotoxic effect in breast carcinoma cells. CuL3 is thus a promising complex to be further studied and to be used as a lead compound for the optimization of novel chemotherapy sensitizers.     

芥子碱硫氰酸盐清除超氧阴离子自由基作用的研究

目的:研究芥子碱硫氰酸盐清除超氧阴离子自由基的作用。方法:用化学发光法测定芥子碱硫氰酸盐清除超氧阴离子自由基的能力,并以抗坏血酸的清除能力做为对照,以IC50值(清除率为50%时的浓度值)作为评价指标。结果:芥子碱硫氰酸盐和抗坏血酸的IC50值分别为0.135 mmol/L和18.74 mmol/L,后者的IC50值约为前者的140倍。结论:芥子碱硫氰酸盐具有良好的抗氧化作用,可以作为天然抗氧化剂进行开发。     

A discrepancy in superoxide scavenging activity between the ESR-spin trapping method and the luminol chemiluminescence method

In a previous study of ours, the superoxide scavenging activity of aqueous extracts from dinophycean red tide flagellates was detected by an electron spin resonance (ESR)-spin trapping method, but not by an L-012 (luminol analog)-dependent chemiluminescence (CL) method. To investigate the discrepancy between the two methods, the effect of ferric-protein complexes on superoxide scavenging activity was examined. The reduced signal intensity of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO)-OOH due to superoxide dismutase (SOD) did not change with the addition of horseradish peroxidase (HRP), while the reduced CL response due to SOD was restored by the addition of different concentrations of HRP. Since HRP is a ferric-protein complex, the effects of other ferric-protein complexes, catalase and hemoglobin, on the reduced CL response due to SOD were examined, and similar results were obtained. As is the case with SOD, the reduced CL response activity due to an aqueous extract from a raphidophycean red tide flagellate, Chattonella ovata, was also enhanced by HRP, catalase, and hemoglobin. ESR spectra analyzed at 77 K indicated that aqueous extracts of Gymnodinium impudicum and Alexandrium affine, both of which are dinophycean red tide flagellates, contained a ferric-protein complex, and that an extract of C. ovata did not. These results suggest that the presence of such a ferric-protein complex is a causative factor in the discrepancy between the ESR and luminol CL methods when determining superoxide scavenging activity.     

Preparation and in vitro antioxidant activities of 6‐amino‐6‐deoxychitosan and its sulfonated derivatives

The 6‐amino‐6‐deoxychitosan (NC) and their 2, 6‐di‐N‐sulfonated derivatives were prepared via N‐phthaloylation, tosylation, azidation, hydrazinolysis, reduction of azide groups and N‐sulfonation, and their structures were systematically characterized by FT‐IR, 2D HSQC NMR, XRD, gel permeation chromatography (GPC), and elemental analysis. The 6‐amino‐6‐deoxychitosan showed effect in three selected antioxidant essays, including reducing power, superoxide anion radical scavenging ability, and hydroxyl radical scavenging effect. But the factors affecting each activity were different. The reducing power and the superoxide anion radical scavenging ability of NC were strong and closely related to the amino groups in the molecular chains. Both introducing N‐sulfonated groups into NC and the concentration reduction of NC and its sulfonated derivatives decreased these activities. For the superoxide anion radical, the molecular charge property was also a significant influence factor. For the hydroxyl radical, NC only showed weak scavenging activity in a special inverse concentration‐dependent manner. However, the incorporation of N‐sulfonated groups significantly improved the scavenging activity, and the more N‐sulfonated groups, the higher the concentrations, the stronger the activity was. The results could be due to the different conformations of NC and its sulfonated derivatives in aqueous solution. © 2015 Wiley Periodicals, Inc. Biopolymers 103: 539–549, 2015.     

EPR studies on the superoxide-scavenging capacity of the nutraceutical resveratrol

Resveratrol (3,4',5-trihydroxystilbene), a polyphenolic compound found in mulberries, grapes, and red wine, has received considerable attention because of its apparent protective effects against various degenerative diseases due to its potential antioxidant activities. However, direct evidence for the superoxide-scavenging capacity of resveratrol is lacking in literature. In this study, electron paramagnetic resonance spectroscopy in combination with 5-(diethoxyphosphoryl)-5-methylpyrroline-N-oxide (DEPMPO)-spin trapping technique was utilized to determine the ability of resveratrol in scavenging superoxide anions generated from both potassium superoxide and the xanthine oxidase/xanthine system. We have demonstrated here for the first time that the presence of resveratrol resulted in decreased formation of DEPMPO-superoxide adduct (DEPMPO-OOH) in both the potassium superoxide and xanthine oxidase/xanthine systems, indicating that resveratrol could directly scavenge superoxide anions. The inhibition of DEPMPO-OOH in the xanthine oxidase/xanthine system, however, was found to be much potent as compared to that observed in potassium superoxide system. It was further shown that resveratrol could also directly inhibit xanthine oxidase activity as assessed by oxygen consumption and formation of uric acid. Taken together, the dual role of resveratrol in directly scavenging superoxide and inhibiting its generation via xanthine oxidase reported in this study may explain, at least in part, the protective role of this compound against oxidative injury in various disease processes.     

Bisphenol-A Can Inhibit the Enzymatic Activity of Human Superoxide Dismutase

Bisphenol-A (BPA), a synthetic xenoestrogen, is currently being used to produce a wide variety of consumer products. Humans as well as animals are exposed to this ubiquitous compound through ingestion, inhalation, and dermal exposure. The effect of this compound on superoxide dismutase (SOD), an antioxidant enzyme, isolated from human blood was studied using an enzyme inhibition assay. The mode of interaction of BPA on SOD was investigated using modeling and docking studies. Purified human SOD from erythrocytes was used to study the enzyme inhibition assay of BPA. Molecular level interactions of BPA on SOD were also analyzed by modeling and docking studies. Our study demonstrates that BPA has an inhibitory effect on SOD. The docking results showed that it could bind to the active site residues of SOD and could interfere with the catalytic activity of the enzyme. Our study reveals for the first time that BPA can directly inhibit the enzymatic activity of human SOD and thus impairs the free radical scavenging mechanism.