比色法测定Fenton反应产生的羟自由基

Fenton反应产生的羟自由基能与水杨酸生成羟基化产物2,3－二羟基苯甲酸,用比色法测定其含量能间接测定羟自由基的生成量．通过对测定条件的研究,得到最佳的测定方案．可作为一种简便的筛选羟自由基清除剂的方法     

丝裂霉素促进活性氧自由基生成的研究

本文探讨了抗癌药丝裂霉素对活性氧自由基,超氧负离子自由基(O_2~-),羟自由基(OH)生成反应的作用,实验表明,丝裂霉素对O_2~-及OH的生成有明显的促进作用,其作用随着丝裂霉素浓度的增加而加强。分别测定了它对两种自由基生成反应的促进率。     

维生素C对视网膜色素上皮蓝光损伤保护作用的研究（简报）

维生素C为6碳多羟化合物,在化学反应中易失去电子,依次生成半脱氧抗坏血酸和脱氧抗坏血酸。因此,维生素C可作为自由基清除剂,能迅速与超氧阴离子、氢化氧基、过氧化氢、羟自由基反应,生成抗坏血酸自由基。蓝光作为一种短波长,靠近紫外线频段的光,具有能量高的特点,是自然界中导     

维生素C对视网膜色素上皮蓝光损伤保护作用的研究(简报)

维生素C为6碳多羟化合物,在化学反应中易失去电子,依次生成半脱氧抗坏血酸和脱氧抗坏血酸。因此,维生素C可作为自由基清除剂,能迅速与超氧阴离子、氢化氧基、过氧化氢、羟自由基反应,生成抗坏血酸自由基。蓝光作为一种短波长,靠近紫外线频段的光,具有能量高的特点,是自然界中导致视网膜色素上皮细胞损伤甚至凋亡的主要光线。本实验通过观察蓝光照射视网膜色素上皮,对其DNA的损伤产生光损伤作用,并比较加入维生素C后对这种光损伤的保护作用,以期探讨维生素C在     

迷迭香酸对羟自由基所致小鼠肝线粒体损伤的保护作用

探索迷迭香酸对羟自由基致小鼠肝脏线粒体氧化损伤的保护作用。采用羟自由基(.OH),诱导小鼠肝线粒体损伤后,通过测定线粒体肿胀度、膜流动性、丙二醛(MDA)含量及琥珀酸脱氢酶(SDH)活性等指标以确定迷迭香酸对小鼠肝线粒体羟自由基损伤的保护作用。结果迷迭香酸剂量依赖地抑制线粒体肿胀,提高膜流动性,降低MDA的生成,增强SDH活性,差异显著。本实验证明迷迭香酸可以抑制.OH所致的线粒体损伤。     

Fenton反应及其可能的活性产物

活性氧对许多生物分子,如脂质、蛋白质和DNA等均可引起损伤,它与许多疾病过程相联系.由超氧阴离子自由基和过氧化氢所引起的许多损伤被认为与它们转变为反应活性更强的组分有关,这些组分包括羟自由基及可能的高价铁组分.实验材料及理论结果表明,当铁盐与过氧化氢混合时,除羟自由基产生以外,高价铁组分也被认为同时产生.Fenton试剂的活性中间体是一亲核加合物,其反应活性及其产物不同于游离态羟自由基的反应活性及产物.Fenton试剂的产物分布依赖于不同的过渡金属离子、不同的配位体、不同的反应底物以及不同的溶剂基体效应.     

葡聚糖糖基化处理对绿豆蛋白抗氧化性影响研究

以绿豆分离蛋白（MBPI）和葡聚糖为原料,通过湿热法制备MBPI-Dextran共价复合物。本研究通过测定产物的还原力、DPPH自由基清除能力、超氧阴离子自由基、羟自由基清除能力共同评价糖基化反应对MBPI抗氧化能力的影响。结果表明：MBPI-Dextran共价复合物具有一定的抗氧化能力,特别是90℃条件下反应得到的产物,抗氧化能力较80℃产物显著提升,这与美拉德反应程度有关,较高温度下美拉德反应进程加快,促使更多具有抗氧化能力的物质生成。因此,糖基化改性后的绿豆蛋白在抗氧化能力研究上具有一定的应用价值。     

北虫草抗氧自由基和羟自由基作用的研究

基于很多疾病与脂质过氧化有关 ,探讨了利用人工培育的北虫草的抗脂质过氧化作用。结果显示 :人工培育的北虫草子座对 Fenton反应生成的羟自由基具有较强的清除作用 ,且作用明显强于相同剂量的羟自由基特异清除剂甘露醇 (P<0 .0 1 ) ;北虫草对邻苯三酚自氧化体系产生的氧自由基亦具有清除作用 ,与对照组比较 P<0 .0 1 ,但作用弱于相同剂量的抗坏血酸。结果提示 :北虫草具有抗脂质过氧化作用。     

葎草多糖含量测定及其抗氧化性研究

对8月和10月采集的葎草全草、嫩头、叶和茎中多糖含量进行测定,并对各部位提取纯化的多糖进行体外抗氧化作用研究。结果表明,8月份嫩头中多糖含量最高,达到(42.897±2.996)mg/g;浓度为5mg/mL的各部位葎草多糖溶液,对Fenton反应生成的羟自由基具有较强的清除能力,达85.15%～98.52%;对邻苯三酚自氧化法产生的超氧阴离子自由基也有较好的抑制能力,达57.15%～67.54%。葎草多糖对羟自由基和超氧阴离子自由基具有较高的清除能力。     

库克诺你果汁提取物体外清除自由基及抗氧化活性研究

本文对诺你果汁多糖、乙醇溶出物和乙酸乙酯萃取物体外对超氧阴离子(O2·)、羟自由基(·OH)、DPPH和脂质过氧化(LPO)的抑制作用进行了研究。超氧阴离子(O2·)由邻苯三酚自氧化产生;羟自由基(·OH)由Fenton反应产生;利用Fe2 诱发卵黄脂蛋白产生丙二醛(MDA),TBA法测定。所有测定均为分光光度法。结果表明,与已知抗氧化剂L抗坏血酸相比,乙醇溶出物和乙酸乙酯萃取物均有明显的捕捉自由基和抗氧化能力,而多糖捕捉自由基和抗氧化能力很低,且对O2·没有抑制作用,反而会增加其生成速度。     

Antioxidant properties of EPC-K1: a study on mechanisms.

Scavenging effects of L-ascorbic acid 2-[3,4-dihydro-2,5,7,8- tetramethyl-2-(4,8,12-trimethytridecyl)-2H-1-benzopyran- 6-yl-hydrogen phosphate] potassium salt (EPC-K1) on hydroxyl radicals, alkyl radicals and lipid radicals were studied with ESR spin trapping techniques. The inhibition effects of EPC-K1 on lipid peroxidation were assessed by TBA assay. The kinetics of EPC-K1 reacting with hydroxyl radicals and linoleic acid radicals were studied by pulse radiolysis. The active site of EPC-K1 and the structure-antioxidative activity relationships were discussed. The superoxide radicals scavenging capacity of the brain homogenate of EPC-K1-treated rats was measured. The results revealed that in comparison with Trolox and vitamin C, EPC-K1 showed better overall antioxidative capacity in vitro and in vivo. EPC-K1 was a moderate scavenger on hydroxyl radicals and alkyl radicals, a potent scavenger on lipid radicals, and an effective inhibitor on lipid peroxidation. EPC-K1 could react with hydroxyl radicals with a rate constant of 7.1 x 10(8) dm3 mol-1 s-1 and react with linoleic acid radicals with a rate constant of 2.8 x 10(6) dm3 mol-1 s-1. The active site of EPC-K1 was the enolic hydroxyl group. After administration of EPC-K1, the ability of rat brain to scavenge superoxide radicals was significantly increased. The potent scavenging effects of EPC-K1 on both hydrophilic and hydrophobic radicals were relevant with its molecular structure, which consisted of both hydrophilic and hydrophobic groups.     

A new approach for EPR detection of hydroxyl radicals by reaction with sterically hindered cyclic amines and oxygen

Sterically hindered cyclic amines react with hydroxyl radicals in the presence of oxygen to yield stable nitroxide radicals which can be detected by EPR. This reaction provides a nonconventional spin-trapping tool for detection of hydroxyl radicals.     

Effect of oxygen free radicals on ubiquinone in aqueous solution and phospholipid vesicles

The purpose of this study was to evaluate the direct effect of oxygen free radicals produced by ultrasonic irradiation on ubiquinone and to compare the efficiency with which the antioxidant can compete with these radicals when it is both in aqueous solution and within the lipid bilayer. The main product obtained after insonation of aqueous solutions of ubiquinone-0 was ubiquinol, moreover some degradation occurred. The direct electron donor responsible for most of the ubiquinol generated by ultrasonic irradiation appeared to be superoxide radical. Addition reactions of hydroxyl radicals with aromatic ring structure led probably to degradation products of ubiquinone, which were not identified. Experiments were also performed to evaluate the efficiency with which ubiquinone-3 could react with oxygen radicals when it was within the lipid bilayer. The effect of presence or absence of a net surface charge was studied selecting a suitable bilayer including dimyristylphosphatidic acid or stearylamine in uncharged dimyristylphosphatidylcholine vesicles. In these systems hydroxyl radicals did not represent a potential danger for the antioxidant, the reaction between superoxide and ubiquinone-3 instead was significant only in positively charged membranes and gave rise to ubiquinol. It is suggested that ubiquinone acts as an antioxidant by stopping the propagation reaction.     

DNA damage during glycation of lysine by methylglyoxal: assessment of vitamins in preventing damage

Summary. Amino acids react with methylglyoxal to form advanced glycation end products. This reaction is known to produce free radicals. In this study, cleavage to plasmid DNA was induced by the glycation of lysine with methylglyoxal in the presence of iron(III). This system was found to produce superoxide as well as hydroxyl radicals. The abilities of various vitamins to prevent damage to plasmid DNA were evaluated. Pyridoxal-5-phosphate showed maximum protection, while pyridoxamine showed no protection. The protective abilities could be directly correlated to inhibition of production of hydroxyl and superoxide radicals. Pyridoxal-5-phosphate exhibited low radical scavenging ability as evaluated by its TEAC, but showed maximum protection probably by interfering in free radical production. Pyridoxamine did not inhibit free radical production. Thiamine and thiamine pyrophosphate, both showed protective effects albeit to different extents. Tetrahydrofolic acid showed better antioxidant activity than folic acid but was found to damage DNA by itself probably by superoxide generation.     

Electrons initiate efficient formation of hydroperoxides from cysteine

Amino acid and protein hydroperoxides can constitute a significant hazard if formed in vivo. It has been suggested that cysteine can form hydroperoxides after intramolecular hydrogen transfer to the commonly produced cysteine sulfur-centered radical. The resultant cysteine-derived carbon-centered radicals can react with oxygen at almost diffusion-controlled rate, forming peroxyl radicals which can oxidize other molecules and be reduced to hydroperoxides in the process. No cysteine hydroperoxides have been found so far. In this study, dilute air-saturated cysteine solutions were exposed to radicals generated by ionizing radiation and the hydroperoxides measured by an iodide assay. Of the three primary radicals present, the hydroxyl, hydrogen atoms and hydrated electrons, the first two were ineffective. However, electrons did initiate the generation of hydroperoxides by removing the –SH group and forming cysteine-derived carbon radicals. Under optimal conditions, 100% of the electrons reacting with cysteine produced the hydroperoxides with a 1:1 stoichiometry. Maximum hydroperoxide yields were at pH 5.5, with fairly rapid decline under more acid or alkaline conditions. The hydroperoxides were stable between pH 3 and 7.5, and decomposed in alkaline solutions. The results suggest that formation of cysteine hydroperoxides initiated by electrons is an unlikely event under physiological conditions.     

Hydrogen peroxide-induced base damage in deoxyribonucleic acid

Aqueous solutions of calf thymus deoxyribonucleic acid (DNA) were exposed to hydrogen peroxide in the presence of air. Base products formed in DNA were identified and quantitated following acid hydrolysis and trimethylsilylation using gas chromatography-mass spectrometry. The yields of these products were dependent upon the hydrogen peroxide concentration, and increased in the following order: 8-hydroxyadenine, cytosine glycol, 2,6-diamino-4-hydroxy-5-formamidopyrimidine, 8-hydroxyguanine, thymine glycol, and 4,6-diamino-5-formamidopyrimidine. Previous studies have shown that these compounds are typically formed in DNA in aqueous solution by hydroxyl radicals generated by ionizing radiation. Hydrogen peroxide is thought to participate in a Fenton-like reaction with transition metals, which are readily bound to DNA in trace quantities, resulting in the production of hydroxyl radicals close to the DNA. This proposed mechanism was examined by exposing DNA to hydrogen peroxide either in the presence of a hydroxyl radical scavenger or following pretreatment of DNA with metal-ion chelators. The results indicate that trace quantities of transition metal ions can react readily with hydrogen peroxide to produce radical species. The production of radical species was monitored by determining the altered bases that resulted from the reaction between radicals and DNA. The yields of the base products were reduced by 40 to 60% with 10 mmol dm-3 of dimethyl sulfoxide. A 100-fold increase in the concentration of dimethyl sulfoxide did not result in a further reduction in hydrogen peroxide-induced base damage. DNA which was freed from bound metal ions by pretreatment with metal ion chelators followed by exhaustive dialysis was found to be an ineffective substrate for hydrogen peroxide. The yields of base products measured in this DNA were at background levels. These results support the role of metal ions bound to DNA in the site-specific formation of highly reactive radical species, most likely hydroxyl radicals, in hydrogen peroxide-induced damage to the bases in DNA.     

Hydroxylation of p-coumaric acid by horseradish peroxidase. The role of superoxide and hydroxyl radicals.

1. In the presence of dihydroxyfumarate, horseradish peroxidase catalyses the conversion of p-coumaric acid into caffeic acid at pH 6. This hydroxylation is completely inhibited by superoxide dismutase. 2. Dihydroxyfumarate cannot be replaced by ascorbate H2O2, NADH, cysteine or sulphite. Peroxidase can be replaced by high (10 mM) concentrations of FeSO4, but this reaction is almost unaffected by superoxide dismutase. 3. Hydroxylation by the peroxidase/dihydroxyfumarate system is completely inhibited by low concentrations of Mn2+ or Cu2+. It is proposed that this is due to the ability of these metal ions to react with the superoxide radical O2--. 4. Hydroxylation is partially inhibited by mannitol, Tris or ethanol and completely inhibited by formate. This seems to be due to the ability of these reagents to react with the hydroxyl radical -OH. 5. It is concluded that O2-- is generated during the oxidation of dihydroxyfumarate by peroxidase and reacts with H2O2 to produce hydroxyl radicals, which then convert p-coumaric acid into caffeic acid.     

Chemiluminescence studies on the generation of oxygen radicals from the interaction of NADPH-cytochrome P-450 reductase with iron

The ability of NADPH-cytochrome P-450 reductase to interact with iron and generate oxygen radicals was evaluated by assaying for low level chemiluminescence. The basic reaction system which contained the reductase, an NADPH-generating system, ferric-EDTA as an electron acceptor, and t-butyl hydroperoxide as the oxidant acceptor, resulted in the production of chemiluminescence. Omission of any of these components resulted in a complete loss of chemiluminescence. The light emission was completely sensitive to inhibition by glutathione and butylated hydroxytoluene, partially sensitive (about 60% decrease) to catalase and hydroxyl radical scavengers, and relatively insensitive (about 20% decrease) to superoxide dismutase. The ability of other ferric chelates to replace ferric-EDTA in catalyzing the reductase-dependent chemiluminescence was evaluated. Ferric-citrate, -ADP, -ATP, and ferric-ammonium sulfate were ineffective in promoting chemiluminescence, whereas ferric-diethylenetriaminepentaacetic acid was even more effective than ferric-EDTA. Thus, the ferric chelates, which catalyze reductase-dependent chemiluminescence, are those which are efficient electron acceptors from the reductase and were previously shown to be those capable of catalyzing hydroxyl radical production by microsomes and the reductase. It is suggested that chemiluminescence results from (a) the direct interaction of the reduced iron chelate with the hydroperoxide (Fenton-type of reaction) to generate alkoxyl and peroxyl radicals, and (b) the generation of hydroxyl radicals, which subsequently react with the hydroperoxide to generate secondary radicals. The latter, but not the former, would be sensitive to inhibition by catalase and competitive hydroxyl radical scavengers, whereas both would be sensitive to antioxidants such as butylated hydroxytoluene. Chemiluminescence appears to be a versatile tool for studying the reductase-dependent generation of oxygen radicals and for the interaction of reductase with iron.     

Hydroxyl radical scavenging activity of beta-N-oxalyl-L-alpha, beta-diaminopropionic acid.

Exogenous hydroxyl radical treatments increased the levels of beta-N-oxalyl-L-alpha, beta-diaminopropionic acid (ODAP) in grass pea seedlings. Lipid peroxidation initiated by hydroxyl radicals can be alleviated when grass pea seedlings were pretreated with exogenous ODAP, suggesting that ODAP might act as an hydroxyl radical scavenger in vivo. Competing with salicylate for hydroxyl radicals in the hydroxyl radical generating/detecting system, ODAP's in vitro hydroxyl radicals scavenging activity was assessed to confirm this result, and the role of ODAP as an effective hydroxyl radical scavenger is discussed.