Flavonoids protect U-937 macrophages against tert-butylhydroperoxide induced oxidative injury.

The study was carried out to determine the relative efficacies of polyphenolic flavonoids, quercetin, catechin and epicatechin against tert-BOOH induced oxidative stress in human macrophage, U-937 cell line. Exposure of the cells to tert-BOOH oxidative stress resulted in a significant increase in cytotoxicity and reactive oxygen species (ROS) generation. Further, a significant decrease in mitochondrial membrane potential and increase in lipid peroxidation and DNA damage was observed in cells exposed to tert-BOOH. Pretreatment of cells with quercetin, catechin and epicatechin significantly inhibited tert-BOOH induced cytotoxicity by inhibiting ROS generation. The flavonoids inhibited DNA damage induced by tert-BOOH and preserved the mitochondrial transmembrane potential significantly. Epicatechin and catechin were found to be more efficient than quercetin in inhibiting tert-BOOH induced cellular damage.     

Caffeic acid and quercetin protect erythrocytes against the oxidative stress and the genotoxic effects of lambda-cyhalothrin in vitro

Lambda-cyhalothrin (LTC) is a synthetic pyrethroid with a broad spectrum of insecticidal and acaricidal activities used to control wide range of insect pests in a variety of applications. The aim of this study was to examine (i) the potency of LTC to induce oxidative stress response in rat erythrocytes in vitro and (ii) the role of caffeic acid (20 μM) and/or quercetin (10 μM) in preventing the cytotoxic effects. Erythrocytes were divided into four portions. The erythrocytes of the first portion were incubated for 4 h at 37°C with different concentrations (0, 50 and 100 μM) of LTC. The others portions were pretreated with caffeic acid and/or quercetin for 30 min prior to LTC incubation. Lipid peroxidation, protein oxidation, antioxidant enzyme activities and DNA damage were examined. LTC at different concentrations causes increased levels of lipid peroxidation, protein oxidation, DNA damage and decreased antioxidant enzyme activities. Combined caffeic acid and quercetin pretreatments significantly reduced the levels of lipid peroxidation markers, that is thiobarbituric acid reactive substance (TBARS), protein carbonyls (PCO) and decreased DNA damage in LTC portion. Further, combined caffeic acid and quercetin pretreatment maintain antioxidant enzyme activities and glutathione content near to normal values. These results suggest that LTC exerts its toxic effect by increasing lipid peroxidation, altering the antioxidant enzyme activities and DNA damage. Caffeic acid and quercetin pretreatments prevent the toxic effects of LTC, suggesting their role as a potential antioxidant.     

Antioxidant and free radical-scavenging activity of Choto-san and its related constituents

The antioxidant properties of Choto-san and its related constituents such as Chotoko and Choto-san without Chotoko, and phenolic compounds contained in Chotoko such as epicatechin, caffeic, acid and quercetin were evaluated. In the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging assay, the scavenging activity of Chotoko (IC(50) 14.3 microg/ml) was found to be higher than that of Choto-san (IC(50) 206.2 microg/ml) and Choto-san without Chotoko (IC(50) 244.3 microg/ml). Epicatechin (IC(50) 10.4 microM), caffeic acid (IC(50) 13.8 microM), and quercetin (IC(50) 7.1 microM) also revealed scavenging activity against DPPH radicals. Choto-san (IC(50) 67.7 microg/ml) exhibited stronger inhibitory activity against superoxide anion formation than Choto-san without Chotoko (IC(50) 92.4 microg/ml) but weaker activity than Chotoko (IC(50) 18.3 microg/ml). The generation of superoxide anion was also inhibited by epicatechin (IC(50) 175.2 microM), caffeic acid (IC(50) 141.7 microM), and quercetin (IC(50) 18.7 microM). In a hydroxyl radical-scavenging experiment, Choto-san (IC(50) 2.4 mg/ml), Chotoko (IC(50) 2.2 mg/ml), Choto-san without Chotoko (IC(50) 2.8 mg/ml), epicatechin (IC(50) 3.9 mM), caffeic acid (IC(50) 3.6 mM), and quercetin (IC(50) 1.9 mM) exhibited activity. In NG108-15 cells, when added simultaneously with H(2)O(2) (500 microM), Choto-san (250 microg/ml), Chotoko (250 microg/ml), Choto-san without Chotoko (500 microg/ml), epicatechin (200 microM), caffeic acid (200 microM), and quercetin (200 microM) effectively protected cells from oxidative damage. In conclusion, the present results provide evidence that Choto-san acts as an antioxidant and cytoprotective agent against oxidative damage, which is due at least partly to the phenolic compounds contained in Chotoko.     

Oxidative and non-oxidative metabolism of ethanol by the rabbit lung.

Pulmonary ethanol metabolism in the rabbit was investigated in vitro by examining oxidative (alcohol dehydrogenase) and non-oxidative (fatty acid ethyl ester synthase) pathways. Pulmonary alcohol dehydrogenase (ADH) activity was very low when measured at close to normal physiological conditions but increased at higher pH, although it was lower than that in the liver. Significant esterification of oleic acid was detected using rabbit lung homogenates, with the rate being intermediate to that of the other tissues examined. The results indicate that oxidative metabolism of ethanol (via ADH) by the rabbit lung is minimal compared to other tissues, but non-oxidative metabolism of ethanol leading to FAEE formation occurs at significant rates that may alter lipid metabolism or membrane composition.     

The protective effect of caffeic acid phenethyl ester against cyclosporine A-induced cardiotoxicity in rats

Cyclosporine A (CsA) is the immunosuppressor, which is most frequently used in transplant surgery and in the treatment of autoimmune diseases. Oxidative stress has been considered as one of the possible mechanisms of CsA-induced cardiotoxicity. The present investigation examines the ability of caffeic acid phenethyl ester (CAPE), which is an active component of propolis extracts, as a natural antioxidant to protect against CsA-induced oxidative stress and cardiotoxicity. CsA cardiotoxicity was induced by subcutaneous injection of CsA at a dose of 15 mg/kg/body weight daily for 21 days in rats. Cardiotoxicity was evaluated by morphological and biochemical studies. CsA treated rats showed degenerative changes with cardiac fibrosis localized around the fibers. These latters were disorganised and the network was disappeared. The ROS production was increased whereas cytochrome-c-oxidase decreased. The expression and levels of matrix metalloproteinase 2 (MMP2) were increased whereas those of its inhibitor were downregulated. CAPE subcutaneous administration (15 micromol/kg/day) improved cardiac cytoarchitecture, decreased the levels and the expression of MMP2, and increased those of TIMP2 proteins. Moreover, it increased cytochrome-c-oxidase activity and decreased ROS production. These results suggest that CAPE could have protective effect against CsA-induced cardiotoxicity.     

Seasonal variation of phenolic constituents and medicinal activities of Northern Labrador tea, Rhododendron tomentosum ssp. subarcticum, an Inuit and cree First Nations traditional medicine

Northern Labrador tea, Rhododendron tomentosum ssp. subarcticum, is one of the most commonly used medicinal plants by Inuit and other First Nations peoples of Canada. The phenolic profile and seasonal variation of this commonly used medicinal plant remains largely unknown. To assess optimal harvesting time, R. tomentosum was collected in accordance with traditional knowledge practices bimonthly throughout the snow-free summer in Iqaluit, Nunavut. The antioxidant potency was measured in a DPPH radical scavenging assay, and the anti-inflammatory activity was determined with a TNF-α production assay. The seasonal variation of phenolic content was assessed with HPLC-DAD for fifteen of the most abundant phenolic compounds; (+)-catechin, chlorogenic acid, PARA-coumaric acid, quercetin 3-O-galactoside (hyperoside), quercetin 3-O-glucoside (isoquercitrin), quercetin 3-O-rhamnoside (quercitrin), quercetin pentoside, myricetin, quercetin, 3 procyanidins, and 3 caffeic acid derivatives. The most abundant constituent was (+)-catechin, which made up 19?% of the total weight of characterized phenolics. There was significant seasonal variation in the quantity of all fifteen constituents assessed, whereas there was no seasonal variation of their total sum. The antioxidant activity was positively correlated with phenolic content and negatively correlated with daylight hours. The anti-inflammatory activity was negatively correlated with caffeic acid derivative 1 and daylight hours. Together these results demonstrate that the timing of harvest of R. tomentosum impacts the plant's phenolic content and its antioxidant and anti-inflammatory activities.     

Protective effect of KR-31378 on oxidative stress in cardiac myocytes

In this study, we investigated whether a novel anti-ischemic KATP opener KR-31378 [(2S,3S,4R)-N"-cyano-N-(6-amino-3,4-dihydro-3-hydroxy-2-methly-2-dimethoxymethly-2H-benzopyran-4-yl)-N'-benzylguanidine] has protective effect against oxidative stress-induced death in heart-derived H9c2 cells. Cell death was induced by BSO, butionine sulfoximine, which inhibits GSH synthesis and subsequently increases reactive oxygen species (ROS) level. Cell death was quantitatively determined by measuring lactate dehydrogenase (LDH) activity and stained by Hoechst 33258. BSO-induced ROS production and mitochondrial membrane potential (MMP) were measured using 2',7'-dichlorofluorescein diacetate oxidation and rhodamine 123, respectively. Both the LDH release and the ROS elevation induced by treatment of H9c2 cells with 10 mM BSO, were significantly decreased by KR-31378. These protective effect and antioxidant effect of KR-31378 appeared to be independent on KATP channel opening. Cells exposed to BSO showed an early reduction in MMP, and this reduction in MMP was significantly reversed by treatment with KR-31378. Caspase-3 activity in BSO treated H9c2 cells was remarkably increased, and this increased caspase-3 activity was significantly reversed by KR-31378. In conclusion, our results suggest that KR-31378 can produce cardioprotective effect against oxidative stress-induced cell death through antioxidant mechanism.     

Olive oil hydroxytyrosol reduces toxicity evoked by acrylamide in human Caco-2 cells by preventing oxidative stress

Humans are exposed to dietary acrylamide (AA) during their lifetime, it is therefore necessary to investigate the mechanisms associated with AA-induced toxic effects. Accumulating evidence indicates that oxidative stress contributes to AA cytotoxicity, thus, dietary antioxidants might have a protective role in colonic cells against AA toxicity. We have recently reported that hydroxytyrosol (HTy), a natural antioxidant abundant in olive oil, is able to enhance the cellular antioxidant defence capacity, thereby protecting cells from oxidative stress. In this study, we evaluate the protective role of HTy on alterations of the redox balance induced by AA in Caco-2 intestinal cells. AA cytotoxicity was counteracted by HTy by powerfully reducing ROS generation, recovering the excited enzyme antioxidant defences and decreasing phospho-Jun kinase concentration and caspase-3 activity induced by AA. Therefore, AA-induced cytotoxicity and apoptosis are closely related to oxidative stress in Caco-2 cells and the olive oil natural dietary antioxidant HTy was able to contain AA toxicity by improving the redox status of Caco-2 cells and by partly restraining the apoptotic pathway activated by AA.     

Effects of Choto-san and its related constituents on endogenous antioxidant systems

We previously reported that Choto-san acts as an antioxidant and cytoprotective agents against H2O2-induced oxidative damage in NG108-15 cells, and the effect is due at least partly to the phenolic compounds. To further investigate the detail mechanisms of this cytoprotection effects of Choto-san and related compounds on enzyme activities of antioxidant systems were examined. Choto-san (5-100 microg/ml) and Chotoko (5-100 mg/ml) stimulated the activity of superoxide dismutase (SOD), catalase and glutathione peroxidase (GPX). These also increased the level of glutathione. Although Choto-san without Chotoko (w/o CKO) did not show the effects on SOD and catalase, GPX activity and glutathion content also, but weakly, stimulated by w/o CKO. The effects of phenolic compounds, epicatechin, caffeic acid and quercetin were also investigated. Epicatechin stimulated catalase, GPX and glutathion content, but not SOD. On the other hand, caffeic acid stimulated SOD activity but had no effects on others. Quercetin stimulated all, although intensities were different among. These results suggest that simultaneous induction of cellular antioxidant defense systems by Choto-sam and its related constituents may be an important mechanisms underlying the protective effects of Choto-san on ischemia-induced neuronal cells injury, and the characteristics of the stimulative effects of phenolic compounds were depend on enzymes.     

Silencing of cytosolic NADP<Superscript>+</Superscript>-dependent isocitrate dehydrogenase gene enhances ethanol-induced toxicity in HepG2 cells

It has been shown that acute and chronic alcohol administrations increase the production of reactive oxygen species, lower cellular antioxidant levels and enhance oxidative stress in many tissues. We recently reported that cytosolic NADP⁺-dependent isocitrate dehydrogenase (IDPc) functions as an antioxidant enzyme by supplying NADPH to the cytosol. Upon exposure to ethanol, IDPc was susceptible to the loss of its enzyme activity in HepG2 cells. Transfection of HepG2 cells with an IDPc small interfering RNA noticeably downregulated IDPc and enhanced the cells’ vulnerability to ethanol-induced cytotoxicity. Our results suggest that suppressing the expression of IDPc enhances ethanol-induced toxicity in HepG2 cells by further disruption of the cellular redox status.     

何首乌炮制前后及其主要成分体外抗氧化活性研究

目的：研究何首乌生品、炮制品及其主要成分的体外抗氧化活性,探讨何首乌的炮制机理及其抗氧化作用物质基础。方法：采用2,2-二苯基-1-苦肼基（DPPH）自由基清除实验及铁离子还原（FRAP）法抗氧化能力检测方法评价何首乌炮制前后及其主要化学成分的体外抗氧化活性。结果：DPPH自由基清除实验中,何首乌生品（S）,随着乙醇浓度的增加,对DPPH自由基的清除率也增加,制品（Q/H/Z）中,以90%乙醇提取所得样品的平均清除率为最高,其次是水、70%乙醇和50%乙醇提取物;FRAP法检测结果显示何首乌生品（S）、黑豆汁制品（H）、清蒸制品（Q）及市售制品（Z）的FRAP值范围分别为2.23~9.27 mmol·L^-1,5.94~9.66 mmol·L^-1,5.74~10.49 mmol·L^-1,3.99~10.40 mmol·L^-1 FeSO₄,不同溶剂提取物的FRAP值由大到小为70%醇 > 50%醇 > 水 > 90%醇;何首乌的组成成分中虎杖苷、白藜芦醇、槲皮苷、槲皮素、芦丁、儿茶素、金丝桃苷、二苯乙烯苷和没食子酸有明显的清除DPPH自由基的作用,没食子酸、槲皮素、槲皮苷、儿茶素、芦丁和白藜芦醇的FRAP值较高,具有较强的抗氧化作用。结论：何首乌炮制前后均有较强的抗氧化作用,虎杖苷、白藜芦醇、槲皮苷、槲皮素、芦丁、儿茶素、金丝桃苷、二苯乙烯苷和没食子酸对何首乌的抗氧化活性有一定的贡献作用。     

Protective effect of hopeahainol A, a novel acetylcholinesterase inhibitor, on hydrogen peroxide-induced injury in PC12 cells

In this study, we evaluated the effects of hopeahainol A, a novel acetylcholinesterase inhibitor (AChEI) from Hopea hainanensis, on H(2)O(2)-induced cytotoxicity in PC12 cells and the possible mechanism. Exposure of PC12 cells to 200μM H(2)O(2) caused cell apoptosis, reduction in cell viability and antioxidant enzyme activities, increment in malondialdehyde (MDA) level, and leakage of lactate dehydrogenase (LDH). Pretreatment of the cells with hopeahainol A at 0.1-10μM before H(2)O(2) exposure significantly attenuated those changes in a dose-dependent manner. Moreover, hopeahainol A could mitigate intracellular accumulation of reactive oxygen species (ROS) and Ca(2+), the loss of mitochondrial membrane potential (MMP), and the increase of caspase-3, -8 and -9 activities induced by H(2)O(2). These results show that hopeahainol A protects PC12 cells from H(2)O(2) injury by modulating endogenous antioxidant enzymes, scavenging ROS and prevention of apoptosis. There was potential for hopeahainol A to be used in treating Alzheimer's disease (AD) that involved acetylcholinesterase, free radical, oxidative damage and cell apoptosis.     

The antioxidative and antihistaminic properties of quercetin in ethanol-induced gastric lesions

The role of reactive oxygen species in the pathogenesis of acute ethanol-induced gastric mucosal lesions and the effects of quercetin were evaluated in an experimental model. In addition, the effects of quercetin on gastric damage were evaluated histopathologically. Rats were divided into three groups as control rats, ethanol treated rats and ethanol+quercetin treated rats. Ethanol group was given a gastric gavage containing 1 ml of 80% ethanol (v/v) prepared in distilled water. Quercetin (200 mg/kg body wt.) was given by intragastric gavage 120 min before the administration of ethanol. Gastric tissue thiobarbituric acid reactive substance levels, carbonyl compounds, histamine levels and myeloperoxidase activities were found to be increased in ethanol treated rats and quercetin treatment reversed these increases. No statistically significant changes were found between all groups in catalase activity. The superoxide dismutase activity dropped significantly after ethanol treatment and quercetin treatment increased this enzyme activity. Gastric damage was confirmed histomorphometrically by significant increases in the number of mast cells and gastric erosions in ethanol treated rats. It was also confirmed that quercetin treatment significantly decreased the number of mast cells and reduced the area of gastric erosions. The results suggest that the gastroprotective effect of quercetin in this experimental model could be due to its antiperoxidative, antioxidant and antihistaminic effects.     

Non-oxidative ethanol metabolism in human hepatic cells in vitro: Involvement of uridine diphospho-glucuronosyltransferase 1A9 in ethylglucuronide production

Ethanol is the most frequently psychoactive substance used in the world, leading to major public health problems with several millions of deaths attributed to alcohol consumption each year. Metabolism of ethanol occurs mainly in the liver via the predominant oxidative metabolism pathway involving phase I enzymes including alcohol dehydrogenases (ADH), cytochrome P450 (CYP) 2E1 and catalase. In a lesser extent, an alternative non-oxidative pathway also contributes to the metabolism of ethanol, which involves the uridine diphospho-glucuronosyltransferase (UGT) and sulfotransferase (SULT) phase II enzymes. Using liquid chromatography-high resolution mass spectrometry, ethylglucuronide (EtG) and ethylsulfate (EtS) produced respectively by UGT and SULT conjugation and detected in various biological samples are direct markers of alcohol consumption. We report herein the efficient non-oxidative metabolic pathway of ethanol in human differentiated HepaRG cells compared to primary human hepatocytes (HH). We showed dose- and time-dependent production of EtS and EtG after ethanol (25 or 50 mM) treatment in culture media of differentiated HepaRG cells and HH and a significant induction of CYP2E1 mRNA expression upon acute ethanol exposure in HepaRG cells. These differentiated hepatoma cells thus represent a suitable in vitro human liver cell model to explore ethanol metabolism and more particularly EtG and EtS production. In addition, using recombinant HepG2 cells expressing different UGT1A genes, we found that UGT1A9 was the major UGT involved in ethanol glucuronidation.     

Protective effect of quercetin in gentamicin-induced oxidative stress in vitro and in vivo in blood cells. Effect on gentamicin antimicrobial activity

We have evaluated the effect of gentamicin and gentamicin plus quercetin on ROS production, endogenous antioxidant defenses (SOD and CAT) and lipid peroxidation in vitro on human leukocytes and in vivo on whole rat blood. Gentamicin generated ROS production in human leukocytes, produced a dual effect on both enzymes dosage-dependent and generated an increase in lipid peroxidation. Quercetin, in leukocytes stimulated by gentamicin, showed more inhibitory capacity in ROS production than the reference inhibitor (vitamin C) in mononuclear cells and a similar protective behavior at this inhibitor in polymorphonuclear cells. Quercetin, in both cellular systems, tend to level SOD and CAT activities, reaching basal values and could prevent lipidic peroxidation induced by gentamicin. The results in Wistar rats confirmed that therapeutic doses of gentamicin can induce oxidative stress in whole blood and that the gentamicin treatment plus quercetin can suppress ROS generation, collaborate with SOD and CAT and diminish lipid peroxidation. Finally, flavonoid and antibiotic association was evaluated on the antimicrobial activity in S. aureus and E. coli, showing that changes were not generated in the antibacterial activity of gentamicin against E. coli strains, while for strains of S. aureus a beneficial effect observes. Therefore, we have demonstrated that gentamicin could induce oxidative stress in human leukocytes and in whole blood of Wistar rats at therapeutic doses and that quercetin may to produce a protective effect on this oxidative stress generated without substantially modifying the antibacterial activity of gentamicin against E. coli strains, and it contributes to this activity against S. aureus strains.     

Eckol inhibits ultraviolet B-induced cell damage in human keratinocytes via a decrease in oxidative stress

In previous reports, the antioxidant effects of eckol were shown to protect cells against hydrogen peroxide- and gamma ray-induced oxidative stress. In this study, the role of eckol in protecting human skin keratinocytes (HaCaT) against UVB-induced oxidative cell damage was investigated. Also, triphlorethol-A, one of the chemical components in Ecklonia cava, and quercetin a well known antioxidant, were compared with eckol in terms of antioxidant activity based on chemical structure. Eckol decreased UVB-induced intracellular reactive oxygen species (ROS), decreased injury to cellular components resulting from UVB-induced oxidative stress, and restored cell viability. In addition, eckol reduced UVB-induced apoptosis by inhibiting the disruption of mitochondrial membranes. These results suggest that eckol protects human keratinocytes against UVB-induced oxidative stress by scavenging ROS, thereby lessening injury to cellular components.     

Inhibitory activity of liposomal flavonoids during oxidative metabolism of human neutrophils upon stimulation with immune complexes and phorbol ester

Context and objective: The massive production of reactive oxygen species by neutrophils during inflammation may cause damage to tissues. Flavonoids act as antioxidants and have anti-inflammatory effects. In this study, liposomes loaded with these compounds were evaluated as potential antioxidant carriers, in attempt to overcome their poor solubility and stability. Materials and methods: Liposomes containing quercetin, myricetin, kaempferol or galangin were prepared by the ethanol injection method and analyzed as inhibitors of immune complex (IC) and phorbol ester-stimulated neutrophil oxidative metabolism by luminol (CLlum) and lucigenin-enhanced (CLluc) chemiluminescence (CL) assays. The mechanisms involved this activity of liposomal flavonoids, such as cytotoxicity and superoxide anion scavenging capacity, and their effect on phagocytosis of ICs were also investigated. Results and discussion: The results showed that the inhibitory effect of liposomal flavonoids on CLlum and CLluc is inversely related to the number of hydroxyl groups in the flavonoid B ring. Moreover, phagocytosis of liposomes by neutrophils does not seem to necessarily promote such activity, as the liposomal flavonoids are also able to reduce CL when the cells are pretreated with cytochalasin B. Under assessed conditions, the antioxidant liposomes are not toxic to the human neutrophils and do not interfere with IC-induced phagocytosis. Conclusion: The studied liposomes can be suitable carriers of flavonoids and be an alternative for the treatment of diseases in which a massive oxidative metabolism of neutrophils is involved.