Antioxidant activity of Hypericum androsaemum infusion: scavenging activity against superoxide radical,hydroxyl radical and hypochlorous acid

Hypericum androsaemum is a medicinal plant species containing many polyphenolic compounds, namely flavonoids and phenolic acids. Since polyphenolic compounds have high antioxidant potential, the ability of H. androsaemum infusion to act as a scavenger of reactive oxygen species (superoxide radical, hydroxyl radical and hypochlorous acid) was investigated. Superoxide radical was generated by the xanthine/xanthine oxidase and phenazine methosulphate/NADH systems. The infusion-mediated prevention of nitroblue tetrazolium reduction by the superoxide radical was used as the measured endpoint. Hydroxyl radical was generated by the Fe3+-EDTA/ascorbate Fenton system, and assayed by evaluating deoxyribose degradation using the thiobarbituric acid method. Hypochlorous acid scavenging activity was tested by measuring the inhibition of hypochlorous acid-induced 5-thio-2-nitrobenzoic acid oxidation to 5,5'-dithiobis(2-nitrobenzoic acid). The tested infusion mainly exhibited a potent scavenging effect on superoxide radicals (although a noncompetitive inhibitory effect on xanthine oxidase was also observed). The infusion also acted as a moderate scavenger of hydroxyl radicals and hypochlorous acid. A phytochemical study of the infusion was also undertaken, and nine phenolic compounds were identified.     

肉桂酸衍生物清除羟基自由基活性研究

目的:比较研究咖啡酸、氯原酸、连翘酯苷、阿魏酸、对-羟基香豆酸、邻-羟基香豆酸、间-羟基香豆酸、肉桂酸等中草药有效成分对HO·的清除能力.方法:利用邻菲罗啉化学发光体系.结果:咖啡酸、氯原酸、连翘酯苷、阿魏酸对HO·具有较强的清除能力,对-羟基香豆酸、邻-羟基香豆酸、间-羟基香豆酸对HO·的清除能力较弱,而肉桂酸对HO·的清除作用很小.结论:肉桂酸衍生物的酚羟基邻位有含孤对电子的杂原子取代基(-OH,-OCH3)时,清除HO·的能力可被提高;在-CHCHCOOH的对位含有酚羟基的化合物比在邻位或间位含有酚羟基化合物的活性强.     

On the antioxidant properties of kynurenic acid: Free radical scavenging activity and inhibition of oxidative stress

Kynurenic acid (KYNA) is an endogenous metabolite of the kynurenine pathway for tryptophan degradation and an antagonist of both N-methyl-d-aspartate (NMDA) and alpha-7 nicotinic acetylcholine (α7nACh) receptors. KYNA has also been shown to scavenge hydroxyl radicals (OH) under controlled conditions of free radical production. In this work we evaluated the ability of KYNA to scavenge superoxide anion (O₂⁻) and peroxynitrite (ONOO⁻). The scavenging ability of KYNA (expressed as IC₅₀ values) was as follows: OH = O₂⁻ > ONOO⁻. In parallel, the antiperoxidative and scavenging capacities of KYNA (0-150 μM) were tested in cerebellum and forebrain homogenates exposed to 5 μM FeSO₄ and 2.5 mM 3-nitropropionic acid (3-NPA). Both FeSO₄ and 3-NPA increased lipid peroxidation (LP) and ROS formation in a significant manner in these preparations, whereas KYNA significantly reduced these markers. Reactive oxygen species (ROS) formation were determined in the presence of FeSO₄ and/or KYNA (0-100 μM), both at intra and extracellular levels. An increase in ROS formation was induced by FeSO₄ in forebrain and cerebellum in a time-dependent manner, and KYNA reduced this effect in a concentration-dependent manner. To further know whether the effect of KYNA on oxidative stress is independent of NMDA and nicotinic receptors, we also tested KYNA (0-100 μM) in a biological preparation free of these receptors - defolliculated Xenopus laevis oocytes - incubated with FeSO₄ for 1 h. A 3-fold increase in LP and a 2-fold increase in ROS formation were seen after exposure to FeSO₄, whereas KYNA attenuated these effects in a concentration-dependent manner. In addition, the in vivo formation of OH evoked by an acute infusion of FeSO₄ (100 μM) in the rat striatum was estimated by microdialysis and challenged by a topic infusion of KYNA (1 μM). FeSO₄ increased the striatal OH production, while KYNA mitigated this effect. Altogether, these data strongly suggest that KYNA, in addition to be a well-known antagonist acting on nicotinic and NMDA receptors, can be considered as a potential endogenous antioxidant.     

珠芽蓼清除羟自由基作用研究

目的为了解珠芽蓼根茎作为蝙蝠蛾幼虫食物的生理作用,研究它的提取物对羟自由基的清除能力。方法采用邻二氮菲—Fe2+氧化法检测珠芽蓼根茎的提取物对H2O2/Fe2+体系产生的羟自由基的清除作用。结果珠芽蓼根茎蒸馏水提取物和丙酮提取物均对羟自由基有清除作用,其中丙酮提取物的作用最强。结论珠芽蓼根茎具有明显的清除羟自由基作用,其活性主要来自丙酮提取物部分。     

Effect of the natural antioxidant 2"-O-glycosylisovitexin on superoxide and hydroxyl radical generation.

The inhibitory effect of 2"-O-glycosyl isovitexin (2"-O-GIV) isolated from green barley leaves on superoxide and hydroxyl radical generation was investigated using electron spin resonance spectroscopy. 2"-O-GIV exhibited dose-related inhibitory activity both towards superoxide generation and hydroxyl radical generation. 2"-O-GIV inhibited superoxide formation by 97% at a level of 25 mM and inhibited hydroxyl radical formation by 91% at a level of 500 microM. The results suggest that the inhibitory activity of 2"-O-GIV is due to trapping radicals.     

Investigations concerning the correlation of COX-1 inhibitory and hydroxyl radical scavenging activity

The aim was to study the COX-1 inhibiting efficacy in context with hydroxyl radical scavenging properties of compounds bearing a carboxylic acid and ester function, respectively. In general, the acids are more potent radical scavengers than the corresponding esters but there is no clear correlation with their COX-1 inhibiting potencies. A feasible scavenging mechanism of carboxylic acids is discussed.     

Primaquine-NSAID twin drugs: Synthesis, radical scavenging, antioxidant and Fe2+ chelating activity

Novel primaquine conjugates with non-steroidal anti-inlammatory drugs (PQ-NSAIDs, 4a-h) were prepared, fully chemically characterized and screened for radical scavenging and antioxidant activities. The synthetic procedure leading to twin drugs 4a-h involved two steps: i) preparation of NSAID benzotriazolides 3a-h from the corresponding NSAID (ibuprofen, ketoprofen, fenoprofen, ketoprofen hydroxy and methylene analogues, diclofenac or indomethacin) and benzotriazole carboxylic acid chloride (BtCOCl, 1), ii) reaction of intermediates 3a-h with PQ. The prepared PQ-NSAIDs exerted moderate activities in the DPPH free radical test and β-carotene-linoleic acid assay. Moreover, ketoprofen derivatives 4d and 4b demonstrated a notable Fe2+ chelating ability as well. On the other hand, negligible antiproliferative and antituberculotic effects of conjugates 4a-h were observed.     

Study on the hydroxyl radical scavenging activity changes of ginseng and ginsenoside-Rb2 by heat processing

The free radical scavenging activities of Panax ginseng C.A. MEYER are known to increase by heat processing. Phenolic acids and Maillard reaction products (MRPs) have been suggested as active free radical scavenging components from our previous research, but heat processing-induced chemical and activity changes of ginsenosides considering the Maillard reaction have not yet been fully elucidated. In this study, we investigated the hydroxyl radical (.OH) scavenging activity changes of ginsengs and ginsenoside-Rb2 (Rb2)) by heat processing using an electron spin resonance spectrometer. Especially, Rb2 was heat processed with the same amount of glycine, a frequently used amino acid in the Maillard reaction model system. As a result, the .OH scavenging activities and brown compound levels of ginseng and glycine-Rb2 mixture were increased by heat processing. However, the increase in .OH scavenging activities were not in accordance with the extents of browning. On the other hand, less-polar ginsenosides such as Rg3, Rg5, and Rk1 were generated from the glycine-Rb2 mixture by heat processing. The sugar moieties at carbon-20 of Rb2 were separated by the steaming process, less-polar ginsenosides were produced, and then the separated sugar moieties were thought to form MRPs with glycine. From the .OH scavenging activity tests of Rb2, glycine, less-polar ginsenosides, and maltol, the increase in .OH scavenging activity was thought to be more closely related to the generation of .OH scavenging ginsenosides such as 20(S)-Rg3 and Rg5 by heat processing than MRPs.     

The contribution of the pyrogallol moiety to the superoxide radical scavenging activity of flavonoids

Sixteen flavonoids including flavonols, flavones, flavanonol and catechins, and five aromatic compounds were examined for their ability to scavenge superoxide radical (O2-*) generated enzymatically in a xanthin-xanthinoxidase system and non-enzymatically in a phenazine methosulfate-NADH system. Pyrogallol, gallic acid and its ester, were much more efficient in scavenging O2-* than catechol. The superiority of pyrogallol over catechol in the flavonoidal nucleus is apparent from the much higher O2-* scavenging activity of myricetin and epigallocatechin, which contain 3',4',5'-trihydroxyl substitution in the B-ring, compared to quercetin and epicatechin, which contain 3',4'-dihydroxyl substitution, respectively. The strong O2-* scavenging ability of pyrogallol appears to function even in the A-ring, as in baicalein, and also in the form of a pyrogalloyl ester at the C-3 position in the C-ring, as in epicatechin gallate and epigallocatechin gallate. It can be concluded that the pyrogallol moiety is an active component of flavonoids for displaying high O2-* scavenging activity. Flavonoids and aromatics were also examined to correlate their O2-* scavenging activity with their oxidizability, which was measured on the basis of electrochemical redox potential and the reducing ability of the Cu2+ ion. Aromatics such as pyrogallol, gallic acid and its ester, and flavonoids such as baicalein, epicatechin gallate and epigallocatechin gallate, in which the O2-* scavenging activity is enhanced by the presence of a pyrogallol moiety which does not belong to the B-ring, reduced the correlation between the higher O2-* scavenging activity and the lower redox potential. The O2-* scavenging activity was well correlated with the Cu2+ reducing ability of flavonoids and aromatics.     

Interactions of a series of coumarins with reactive oxygen species. Scavenging of superoxide, hypochlorous acid and hydroxyl radicals.

Sixteen plant-derived or synthetic coumarins with various hydroxyl and other substitutions were tested for their ability to inhibit lipid peroxidation and to scavenge hydroxyl radicals, superoxide radicals and hypochlorous acid. Seven unsubstituted or monosubstituted coumarins were essentially inactive in all tests except for ability to scavenge OH with rate constants approximately greater than 1 x 10(9) M-1. sec-1. Of the remaining nine, six containing dihydroxy substitutions were effective inhibitors of Fe3+-ascorbate-dependent microsomal lipid peroxidation (IC50 less than 20 microM), with ortho-dihydroxy + one additional substitution optimal (IC50 less than 10 microM). ortho-Dihydroxylated coumarins were pro-oxidant (enhanced OH generation) in the Fe3+-EDTA-H2O2 deoxyribose system but decreased OH' generation in the Fe3+-ascorbate-H2O2 deoxyribose system, indicating that these compounds can both chelate iron ions and also readily donate electrons for redox cycling of Fe3+. The meta-dihydroxycoumarin did not show this behaviour, but was an effective scavenger of hypochlorous acid, a property shared by only one other compound. Several other coumarins with one or more hydroxyl substituents were also capable of effectively removing superoxide anions (IC50 3.7-72 microM), although some could not be quantified due to direct rapid reduction of cytochrome c. We conclude that several compounds, notably 5,7-dihydroxy-4-methylcoumarin, possess beneficial biochemical profiles of interest in relation to pathophysiological processes dependent upon reactive oxygen species.     

Aqueous extracts of Lippia turbinata and Aloysia citriodora (Verbenaceae): assessment of antioxidant capacity and DNA damage

The aim of the present work was to make a contribution to the knowledge of aqueous extracts of Lippia turbinata and Aloysia citriodora (Verbenaceae; infusion and decoction) in relation with the establishment of its antioxidant activity and lack of DNA damage, for its potential use in therapeutics. The cytogenotoxic profile was evaluated through genotoxic biomarkers such as mitotic index, cellular proliferation kinetics, sister chromatid exchanges, single-cell gel electrophoresis assay, and micronucleus test in human peripheral blood lymphocyte cultures. No statistical differences were found (P > .05) between control and exposed cultures, even between both aqueous extracts. The total antioxidant capacity was shown to be higher in the decoction than in the infusion and both aqueous extracts protected against protein carbonylation and lipid peroxidation, the decoction being more efficient than the infusion (P < .005). These results suggest the safe use of these medicinal plants as chemoecologic agents in therapeutics.     

The iron-binding and hydroxyl radical scavenging action of anti-inflammatory drugs

1. Hydroxyl radicals (.OH) are thought to be generated at sites of inflammation and to contribute to tissue damage. All anti-inflammatory drugs tested were able to scavenge .OH generated in free solution at almost diffusion-controlled rates (rate constants about 10(10)M-1s-1). 2. Much .OH generation in vivo occurs at specific sites, where bound metal ions (such as Fe2+) react with H2O2 to produce .OH that immediately attacks the site. Only .OH scavengers that have sufficient metal-binding ability to withdraw metal ions from this site can protect against site-specific damage. 3. All anti-inflammatory drugs tested were able to protect against site-specific damage by .OH in a simple model system in vitro. Penicillamine, diclofenac sodium, piroxicam, azathioprine, primaquine, chloroquine and hydroxychloroquine were especially effective. 4. The ability of an anti-inflammatory drug to protect against .OH formation in vivo depends not only on its rate constant for reaction with .OH, but also on its metal-binding ability and on the geometry and redox potential of any metal complex formed.     

Characterization of the radical scavenging and antioxidant activities of danshensu and salvianolic acid B.

Danshensu (3-(3,4-dihydroxyphenyl) lactic acid) and salvianolic acid B, two natural phenolic acids of caffeic acid derivatives isolated from Salvia miltiorrhiza root of the most widely used traditional Chinese medicine for the treatment of various cardiovascular diseases, have been reported to have potential protective effects from oxidative injury. To better understand their biological functions, the in vitro radical scavenging and antioxidant activities of danshensu and salvianolic acid B were evaluated along with vitamin C. Both danshensu and salvianolic acid B exhibited higher scavenging activities against free hydroxyl radicals (HO()), superoxide anion radicals (O(2)(-)), 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radicals and 2-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radicals than vitamin C. In contrary, danshensu and salvianolic acid B showed weaker iron chelating and hydrogen peroxide (H(2)O(2)) scavenging activities than vitamin C. As expressed as vitamin C equivalent capacity (VCEAC), the relative VCEAC values (mg/100ml) were in the order of salvianolic acid B (18.59) > danshensu (12.89) > vitamin C (10.00) by ABTS radical assay. The protective efficiencies against hydrogen peroxide induced human vein vascular endothelial cell damage were correlated with their antioxidant activities. Analysis of structure-activity relationship of these two compounds showed that the condensation and conjugation of danshensu and caffeic acid appears important for antioxidant activity. These results indicated that danshensu and salvianolic acid B are efficient radical scavengers and antioxidants, and salvianolic acid B is superior to danshensu. Their radical scavenging and antioxidant properties might have potential applications in food and healthcare industry.     

羊栖菜中清除自由基活性物质的研究

目的 通过不同溶剂对羊栖菜的有效成分进行提取和体外活性检测,以期筛选出具有一定开发潜力的自由基清除荆.方法 采用DPPH(二苯代苦味酰自由基)体系、超氧阴离子体系对不同提取物进行清除自由基活性测定,并与海带、紫菜的提取物及抗坏血酸(Vit C)和荼多酚作活性对照.结果 不同溶剂对羊栖菜的提取率不同,各种提取物之间的清除自由基活性存在一定差异.结论 0.5%Na2CO3溶液提取率最高,且提取物清除自由基的能力最强,以DPPH体系作为检测系统时测量结果较为稳定,效果也较明显.     

Superoxide, hydroxyl and fatty acid radical scavenging by aminosalicylates. Direct evaluation with electron spin resonance spectroscopy.

Reactive oxygen radicals such as superoxide and hydroxyl radicals, as well as intermediate unsaturated fatty acid radicals, have been proposed as playing an important role in various diseases including inflammatory bowel disease (IBD). In this study we evaluated radical scavenger properties of aminosalicylates used in the therapy of IBD using spin trapped electron spin resonance spectroscopy. 5-Aminosalicylic acid (5-ASA), 4-aminosalicylic acid and olsalazine had superoxide radical scavenger properties (IC50 = 0.4, 0.4 and 1.0 mM, respectively). 5-ASA and benzalazine also inhibited hydroxyl radicals (IC50 = 6.5 mM). Fatty acid radicals were not inhibited by aminosalicylates. Our results support the hypothesis that therapeutically active compounds may be oxygen radical scavengers and that fatty acid radical scavenging has to be performed by drugs other than aminosalicylates.     

Effects of hydroxyl radical scavenging on cisplatin-induced p53 activation, tubular cell apoptosis and nephrotoxicity

Nephrotoxicity is a major side effect of cisplatin, a widely used cancer therapy drug. Recent work has suggested a role of p53 in renal cell injury by cisplatin. However, the mechanism of p53 activation by cisplatin is unclear. This study determined the possible involvement of oxidative stress in p53 activation under the pathological condition using in vitro and in vivo models. In cultured renal proximal tubular cells, cisplatin at 20 microM induced an early p53 phosphorylation followed by protein accumulation. Cisplatin also induced reactive oxygen species (ROS), among which hydroxyl radicals showed a rapid and drastic accumulation. Dimethylthiourea (DMTU) and N-acetyl-cysteine (NAC) attenuated hydroxyl radical accumulation, and importantly, diminished p53 activation during cisplatin treatment. This was accompanied by the suppression of PUMA-alpha, a p53-regulated apoptotic gene. Concomitantly, mitochondrial cytochrome c release and apoptosis were ameliorated. Notably, DMTU and NAC, when added post-cisplatin treatment, were also inhibitory to p53 activation and apoptosis. In C57BL/6 mice, cisplatin at 30 mg/kg induced p53 phosphorylation and protein accumulation, which was also abrogated by DMTU. DMTU also ameliorated tissue damage, tubular cell apoptosis and cisplatin-induced renal failure. Collectively, this study has suggested a role of oxidative stress, particularly hydroxyl radicals, in cisplatin-induced p53 activation, tubular cell apoptosis and nephrotoxicity.     

Biologically-significant scavenging of the myeloperoxidase-derived oxidant hypochlorous acid by some anti-inflammatory drugs

Neutrophils contain the enzyme myeloperoxidase, which oxidizes Cl- ions into the powerful oxidant hypochlorous acid (HOCl). HOCl inactivates alpha 1-antiprotease, permitting uncontrolled protease activities. Most anti-inflammatory drugs tested are capable of reacting with HOCl, but the reactions seem insufficiently rapid under physiological conditions to protect alpha 1-antiprotease against inactivation by HOCl. However, rapid scavenging of HOCl might contribute to the anti-inflammatory effects of penicillamine, gold sodium thiomalate, phenylbutazone and primaquine.     

Streptonigrin-induced deoxyribose degradation: inhibition by superoxide dismutase, hydroxyl radical scavengers and iron chelators

The aminoquinone antitumour antibiotic streptonigrin stimulates deoxyribose degradation in cell-free systems. This degradation is dependent both on reduction of the drug to a semiquinone and on traces of molecular oxygen in the reaction. Inhibition by a variety of hydroxyl radical scavengers and by catalase implicates a radical species with properties similar to the hydroxyl radical. Iron salts appear to play some part in radical formation as DETAPAC and desferrioxamine partly inhibit. Deoxyribose degradation under conditions of low oxygen concentration is strongly inhibited by superoxide dismutase.     

Antimutagenic, antioxidant and free radical scavenging activity of ethyl acetate extracts from white, yellow and red onions.

The beneficial effects of red, yellow and white onion extracts have been assessed by antioxidant activity and antimutagenic activity. And the effects compared to BHT and ascorbic acid. Total phenolic compounds and flavonoids in onion extracts were determined. Yellow onion extract had more organic acid and free sugar than those detected in the white and red onion extract. The scavenging activity of DPPH radical and H(2)O(2) were increased depending on the concentration. The antioxidant activities using beta-carotene-linoleate system and reducing power were increased but the effect was small to that of BHT and ascorbic acid. After digested, extracts showed antimutagenic activities, and it seems that they inhibit the mutagenicity for digesting. This study demonstrated that the antimutagenicities and antioxidant properties of ethyl acetate extract against mutagens were related to their phenols and flavonoids, which are heat stable and losses digestive juices are relatively low.     

淫羊藿的自由基清除与抗氧化活性

目的研究淫羊藿不同部位的体外抗自由基与抗氧化活性。方法自由基清除活性采用2,2-二苯基-1-苦味基肼自由基清除（DPPH）实验法;抗氧化活性采用β-胡萝卜素漂白体系实验法。结果乙酸乙酯部位自由基清除活性与抗氧化活性较好。结论淫羊藿乙酸乙酯措取部位含有较多抗氧化成分,可为日后淫羊藿抗氧化提取物在药品领域的开发提供依据。