Inactivation of alpha1-antiproteinase induced by phenylbutazone: participation of peroxyl radicals and hydroperoxide

To clarify the action of a side-effect of phenylbutazone, we investigated the inactivation of alpha(1)-antiproteinase induced by phenylbutazone in the presence of horseradish peroxidase (HRP) and H(2)O(2) (HRP-H(2)O(2)). The activity of alpha(1)-antiproteinase was rapidly lost during the interaction of phenylbutazone with HRP-H(2)O(2) under aerobic conditions. Phenylbutazone showed a marked spectral change under aerobic conditions but not under anaerobic conditions. Spin trap agents were very effective in inhibiting alpha(1)-antiproteinase inactivation induced by phenylbutazone. Oxidation of phenylbutazone was stopped by catalase, but the inactivation reaction of alpha(1)-antiproteinase proceeded even after removal of H(2)O(2) in the reaction mixture. Formation of the peroxidative product from phenylbutazone was detected by iodometric assay. These results indicate that both peroxyl radicals and the peroxidative product of phenylbutazone participated in the inactivation of alpha(1)-antiproteinase. Other anti-inflammatory drugs did not inactivate alpha(1)-antiproteinase during interaction with HRP-H(2)O(2). Inactivation of alpha(1)-antiproteinase may contribute to serious side effects of phenylbutazone.     

Ambroxol inhibits peroxynitrite-induced damage of alpha1-antiproteinase and free radical production in activated phagocytic cells

The present study examined the effect of ambroxol on toxic action of peroxynitrite and the respiratory burst in activated phagocytic cells. Ambroxol decreased the inactivation or destruction of alpha1-antiproteinase induced by peroxynitrite (ONOO-) or hypochlorous acid (HOCl), which was similar to penicillamine and glutathione and was greater than diclofenac sodium and naproxen sodium. Ambroxol significantly decreased ONOO--mediated tyrosine nitration and iron plus EDTA-mediated degradation of 2-deoxy-D-ribose. Ambroxol significantly attenuated the production of superoxide, hydrogen peroxide, HOCl, and nitric oxide in fMLP- or IL-1-activated phagocytic cells, while the inhibitory effects of antiinflammatory and thiol compounds were only observed in HOCl production. Ambroxol and antiinflammatory drugs did not show a cytotoxic effect on macrophages. The results suggest that ambroxol protects tissue components against oxidative damage by an action different from antiinflammatory drugs. Ambroxol may interfere with oxidative damage of alpha1-antiproteinase through a scavenging action on ONOO- and HOCl and inhibition of the respiratory burst of phagocytic cells.     

Modulation of peroxynitrite- and hypochlorous acid-induced inactivation of alpha1-antiproteinase by mercaptoethylguanidine.

1. Peroxynitrite is a cytotoxic species that can be formed, among other mechanisms, by the rapid reaction of superoxide with nitric oxide. Peroxynitrite formation has been implicated in a wide range of neurodegenerative and chronic inflammatory diseases, as has the formation of hypochlorous acid by myeloperoxidase. 2. There is considerable interest in the development of peroxynitrite scavengers as therapeutic agents. The thiol compound mercaptoethylguanidine has been suggested to fulfil this role since it has recently been shown to be not only a potent inhibitor of inducible nitric oxide synthase but also a scavenger of peroxynitrite. Indeed, it has been shown to be protective in some experimental models of circulatory shock and inflammation at plasma levels in the approximate range 100-300 microM. 3. One protein inactivated by peroxynitrite is the major inhibitor of serine proteinases in human body fluids, alpha1-antiproteinase. At high (250-1000 microM) concentrations, mercaptoethylguanidine was found to be effective in preventing peroxynitrite-mediated tyrosine nitration and alpha1-AP inactivation. 4. By contrast, lower concentrations of mercaptoethylguanidine (1-60 microM) enhanced the inactivation of alpha1-antiproteinase by peroxynitrite. 5. At all concentrations tested (1-1000 microM), mercaptoethylguanidine decreased the inactivation of alpha1-antiproteinase by hypochlorous acid. 6. We suggest that products of reaction of mercaptoethylguanidine with peroxynitrite or peroxynitrite-derived products could cause damage to alpha1-antiproteinase, and possibly other proteins in vivo, whereas scavenging of hypochlorous acid by mercaptoethylguanidine could contribute to its anti-inflammatory action in vivo.     

Antioxidant and pro-oxidant properties of active rosemary constituents: carnosol and carnosic acid.

1. Carnosol and carnosic acid have been suggested to account for over 90% of the antioxidant properties of rosemary extract. 2. Purified carnosol and carnosic acid are powerful inhibitors of lipid peroxidation in microsomal and liposomal systems, more effective than propyl gallate. 3. Carnosol and carnosic acid are good scavengers of peroxyl radicals (CCl3O2.) generated by pulse radiolysis, with calculated rate constants of 1-3 x 10(6) M-1 s-1 and 2.7 x 10(7) M-1 s-1 respectively. 4. Carnosic acid reacted with HOCl in such a way as to protect the protein alpha 1-antiproteinase against inactivation. 5. Both carnosol and carnosic acid stimulated DNA damage in the bleomycin assay but they scavenged hydroxyl radicals in the deoxyribose assay. The calculated rate constants for reaction with .OH in the deoxyribose system for carnosol and carnosic acid were 8.7 x 10(10) M-1 s-1 and 5.9 x 10(10) M-1 s-1 respectively. 6. Carnosic acid appears to scavenge H2O2, but it could also act as a substrate for the peroxidase system. 7. Carnosic acid and carnosol reduce cytochrome c but with a rate constant significantly lower than that of O2(-.).     

The inhibitory effect of ambroxol on hypochlorous acid-induced tissue damage and respiratory burst of phagocytic cells

Ambroxol (100 microM and 1 mM) and the thiols (all 1 mM), glutathione, tiopronin and cysteine, significantly attenuated the myeloperoxidase, H(2)O(2) and Cl(-) system-caused destruction of alpha(1)-antiproteinase and the HOCl-induced destruction of collagen, whereas they did not affect the elastase-induced destruction of collagen. Glutathione, tiopronin and cysteine almost completely decomposed both HOCl and H(2)O(2), while ambroxol up to 1 mM did not show a scavenging action on H(2)O(2). Ambroxol (1 to 100 microM) and 1 mM thiol compounds markedly inhibited the HOCl-induced alteration of elastase activity. Thiol compounds significantly attenuated the HOCl production caused by degraded immunoglobulin G-activated neutrophils. Ambroxol depressed superoxide and H(2)O(2) production induced by degraded immunoglobulin G-activated neutrophils and by lipopolysaccharide-activated alveolar macrophages in a dose-dependent manner. The results show that ambroxol may interfere with oxidative tissue damage and decrease proteolytic tissue destruction by attenuation of oxidative stress-induced inactivation of alpha(1)-antiproteinase through both decomposition of HOCl and inhibition of the respiratory burst in phagocytic cells.     

Does the anaerobic formation of hydroxyl radicals by paraquat monocation radicals and hydrogen peroxide require the presence of transition metals?

This in vitro study investigated the formation of hydroxyl radicals (*OH) under anaerobic conditions through the direct reaction between paraquat radicals (PQ(+)*) and hydrogen peroxide (H(2)O(2)) by quantitative UV-VIS and electron spin resonance (ESR) spectroscopy. PQ(+)* was formed by paraquat reduction using either sodium dithionite or the xanthine/xanthine oxidase reaction as electron donors. The anaerobic formation of PQ(+)* was quantified both by measuring light absorption at 605 nm or by ESR techniques respectively, using either the absorption coefficient or ultramarine as a stable spin standard. Detection of *OH took place with aid of the spin trap 5-diethoxyphosphoryl-5-methyl-1-pyrroline- N-oxide (DEPMPO). Generation or addition of H(2)O(2) to PQ(+)* eliminates the 35-line ESR signal of PQ(+)* and subsequently generates the 8-line ESR signal of the DEPMPO-OH adduct. The elimination of PQ(+)* as well as the formation of OH-DEPMPO adduct was not influenced by 1.0 mM deferoxamine, indicating that iron or other transition metals are, at least under anoxic conditions, not necessarily involved in the generation of the most aggressive reactive oxygen species *OH.     

Plant phenolic antioxidant and prooxidant activities: phenolics-induced oxidative damage mediated by metals in plants

Plant phenolic compounds such as flavonoids and lignin precursors are important constituents of the human diet. These dietary phytophenolics have been recognized largely as beneficial antioxidants that can scavenge harmful active oxygen species including O(2)(.-), H(2)O(2), .OH, and (1)O(2). Here we review our current understanding of the antioxidant and prooxidant actions of phenolics in plant cells. In plant systems, phytophenolics can act as antioxidants by donating electrons to guaiacol-type peroxidases (GuPXs) for the detoxification of H(2)O(2) produced under stress conditions. As a result of such enzymatic as well as non-enzymatic antioxidant reactions, phenoxyl radicals are formed as the primary oxidized products. Until recently, phenoxyl radicals had been difficult to detect by static electron spin resonance (ESR) because they rapidly change to non-radical products. Application of Zn exerts spin-stabilizing effects on phenoxyl radicals that enables us to analyze the formation and decay kinetics of the radicals. The ESR signals of phenoxyl radicals are eliminated by monodehydroascorbate radical (MDA) reductase, suggesting that phenoxyl radicals, like the ascorbate radical, are enzymatically recycled to parent phenolics. Thus, phenolics in plant cells can form an antioxidant system equivalent to that of ascorbate. In contrast to their antioxidant activity, phytophenolics also have the potential to act as prooxidants under certain conditions. For example, flavonoids and dihydroxycinnamic acids can nick DNA via the production of radicals in the presence of Cu and O(2). Phenoxyl radicals can also initiate lipid peroxidation. Recently, Al, Zn, Ca, Mg and Cd have been found to stimulate phenoxyl radical-induced lipid peroxidation. We discuss the mechanism of phenoxyl radical prooxidant activity in terms of lifetime prolongation by spin-stabilizing agents.     

On the reaction mechanism of tirapazamine reduction chemistry: unimolecular N-OH homolysis, stepwise dehydration, or triazene ring-opening

The initial steps of the activation of tirapazamine (TPZ, 1, 3-amino-1,2,4-benzotriazine 1,4-N,N-dioxide) under hypoxic conditions consist of the one-electron reduction of 1 to radical anion 2 and the protonation of 2 at O(N4) or O(N1) to form neutral radicals 3 and 4, respectively. There are some questions, however, as to whether radicals 3 and/or 4 will then undergo N-OH homolyses 3 → 5 + ·OH and 4 → 6 + ·OH or, alternatively, whether 3 and/or 4 may react by dehydration and form aminyl radicals via 3 → 11 + H(2)O and 4 → 12 + H(2)O or phenyl radicals via 3 → 17 + H(2)O. These outcomes might depend on the chemistry after the homolysis of 3 and/or 4, that is, dehydration may be the result of a two-step sequence that involves N-OH homolysis and formation of ·OH aggregates of 5 and 6 followed by H-abstraction within the ·OH aggregates to form hydrates of aminyls 11 and 12 or of phenyl 17. We studied these processes with configuration interaction theory, perturbation theory, and density functional theory. All stationary structures of OH aggregates of 5 and 6, of H(2)O aggregates of 11, 12, and 17, and of the transition state structures for H-abstraction were located and characterized by vibrational analysis and with methods of electron and spin-density analysis. The doublet radical 17 is a normal spin-polarized radical, whereas the doublet radicals 11 and 12 feature quartet instabilities. The computed reaction energies and activation barriers allow for dehydration in principle, but the productivity of all of these channels should be low for kinetic and dynamic reasons. With a view to plausible scenarios for the generation of latent aryl radical species without dehydration, we scanned the potential energy surfaces of 2-4 as a function of the (O)N1-Y (Y = C5a, N2) and (O)N4-Z (Z = C4a, C3) bond lengths. The elongation of any one of these bonds by 0.5 ? requires less than 25 kcal/mol, and this finding strongly suggests the possibility of bimolecular reactions of the spin-trap molecules with 2-4 concomitant with triazene ring-opening.     

Oxygen dependence of oxidative stress. Rate of NADPH supply for maintaining the GSH pool during hypoxia

NADPH supply for oxidized glutathione (GSSG) reduction was studied in hepatocytes under different steady-state O2 concentrations with controlled infusions of diamide, a thiol oxidant. When bis-chloro-nitrosourea (BCNU) was used to inhibit GSSG reductase, the rate of GSH depletion approximated the rate of diamide infusion, showing that diamide reacted preferentially with GSH under these experimental conditions. Under aerobic conditions without BCNU treatment, the GSH and NADPH pools were largely unaffected and little diamide accumulation or protein thiol oxidation occurred with diamide infusion rates up to 5.3 nmol/10(6) cells per min. However, at greater infusion rates, GSH and NADPH decreased, diamide and GSSG concentrations increased, and protein thiols were oxidized. This critical infusion rate was easily discernible and provided a convenient means to assess the capacity of cells to reduce GSSG as a function of O2 concentration. As the O2 concentration was decreased below 15 microM, the critical infusion rate decreased from the aerobic value of 5.3 to less than 2 nmol/10(6) cells per min in anoxic cells; half-maximal change occurred at 5 microM O2. Although cells could not maintain normal thiol and NADPH pools at infusion rates above the critical value, analysis of the rates of thiol depletion showed that the maximal NADPH supply rate for GSSG reduction under aerobic conditions was 7-8 nmol/10(6) cells per min and was affected by hypoxia to the same degree as the critical value. Thus, hypoxia and anoxia impair the capability of cells to supply NADPH for the reduction of thiol oxidants. This could be an important factor in the sensitivity of hypoxic and ischemic tissues to oxidative injury.     

Comparative study of the antioxidant activity of some thiol-containing substances

Background  

Therapeutic thiol administration has been shown to have great potential in a variety of pathological conditions associated with oxidative stress. In the present study, the free radical scavenging effects against superoxide anion (O_2^- {\rm O}_{2^ - } ) and hydroxyl (·OH) radicals of captopril were compared with those of cysteamine and mercaptoethanol. Methods: The O_2^- {\rm O}_{2^ - } and ·OH were generated in vitro. Deoxyribose (DR) was used as a detector of ·OH radicals. The degradation of DR was measured in terms of the formation of thiobarbituric acid reactive substances, which were quantified spectrophotometrically. Superoxide anion radicals were generated photochemically and O_2^- {\rm O}_{2^ - } -produced nitro-blue tetrazolium (NBT) reduction was measured.     

Inhibitory effect of reactive oxygen species on angiotensin I-converting enzyme (kininase II)

1. Somatic angiotensin I-converting enzyme (ACE) is a protein that contains two similar domains (N- and C-terminal), each possessing an active site. We have examined the effects of a generator of hydroxyl radicals (g*OH: 2,2'-azo-bis(2-amidinopropane)) and hydrogen peroxide (H2O2) on ACE using an in vitro approach. 2. The generator of hydroxyl radicals inactivated ACE in a time (2-6 h)- and concentration (0.3-3 mmol/L)-dependent manner at 37 degrees C. When ACE was coincubated for 4 h with g*OH (3 mmol/L), its activity decreased by 70%. Addition of dimethylthiourea or mannitol + methionine, two *OH scavengers, resulted in a significant protection of ACE activity. Mercaptoethanol and dithiotreitol, two thiol-reducing agents, also efficiently protected ACE activity. 3. The hydrolysis of two natural and domain-specific substrates was explored. The hydrolysis of angiotensin I, preferentially cleaved by the C-domain, was significantly inhibited (57-58%) after 4 h exposure to g*OH (0.3-1 mmol/L). Under the same conditions of exposure, the hydrolysis of N-acetyl-Ser-Asp-Lys-Pro, a specific substrate for the N-domain, was only slightly inhibited by 1 mmol/L g*OH. 4. Hydrogen peroxide, another source of *OH, was used. After exposure to H2O2 (3 mmol/L; 4 h), an 89% decrease in ACE activity was observed. Pretreatment with the iron chelator deferoxamine (1 mmol/L) attenuated H2O2-mediated ACE inactivation, demonstrating that the effect of H2O2 was partly due to its conversion into *OH (Fenton reaction). 5. In summary, our findings demonstrate that g*OH and H2O2 inhibit ACE activity and suggest a preferential action of g*OH on the C-domain of the enzyme.     

Antioxidant and pro-oxidant actions of the plant phenolics quercetin, gossypol and myricetin. Effects on lipid peroxidation, hydroxyl radical generation and bleomycin-dependent damage to DNA

The plant-derived phenolic compounds gossypol, quercetin and myricetin are powerful inhibitors of iron-induced lipid peroxidation in rat liver microsomes, under all five experimental conditions tested and at low micromolar concentrations (IC50 less than or equal to 1.5 microM). However, they greatly accelerate the generation of hydroxyl radicals (.OH) from H2O2 in the presence of Fe3+-EDTA at pH 7.4, as measured by the deoxyribose assay. At 100 microM, the three phenolic compounds enhanced .OH formation up to eight-fold. The hydroxyl radical generation was inhibited by catalase and superoxide dismutase, suggesting a mechanism in which the phenols oxidize to produce superoxide radical, which then assists .OH generation from H2O2 in the presence of Fe3+-EDTA. At concentrations up to 75 microM, quercetin and myricetin also accelerate bleomycin-dependent DNA damage in the presence of Fe3+, possibly by reducing the Fe3+-bleomycin-DNA complex to the Fe2+ form. Hence these naturally-occurring substances can have pro-oxidant effects under some reaction conditions and cannot be classified simplistically as "antioxidants".     

Radical degradation of high molecular weight hyaluronan: inhibition of the reaction by ibuprofen enantiomers

The antioxidative and/or free-radical-scavenging activities of R-(-)- and S-(+)-ibuprofen enantiomers, as well as of the drug racemate, were studied in vitro on measuring the kinetics of (uninhibited or drug-inhibited) degradation of high molecular weight hyaluronan by hydroxyl radicals. The continual flux of OH radicals at aerobic conditions was maintained by the H2O2 + Cu2+ system. The kinetics of hyaluronan degradation was monitored indirectly by capillary viscometry. Under experimental conditions, with no drug addition, the relative viscosity ([eta]rel) decreased continuously, reaching 13% of the initial [eta]rel. value in 4 h. Each drug tested exhibited a dose-dependent protective effect against hyaluronan degradation, however R-(-)-ibuprofen demonstrated a slightly greater activity than the drug S-(+)-enantiomer.     

The effects of reactive oxygen species on calcium- and carbachol- induced contractile responses in β-escin permeabilized rat bladder

The effect of reactive oxygen species on contractions in beta-escin permeabilized rat detrusor was investigated. Cumulative calcium contractions were inhibited by hydrogen peroxide (H(2)O(2)) and hydroxyl (*OH) but not by superoxide (O(2) *). The sarcoplasmic reticulum calcium-ATPase inhibitor cyclopiazonic acid (CPA) and the mitochondrial blocker carbonyl cyanide p-trifluromethoxyphenylhydrazone (FCCP) decreased the calcium contractions, however in their presence, H(2)O(2) and *OH did not have further effect. Carbachol contractions were inhibited by either H(2)O(2)/*OH/O(2) * or CPA/FCCP. In the presence of CPA, carbachol contractions were not affected by H(2)O(2) and *OH but further decreased by O(2) *. On the other hand, only H(2)O(2) and *OH elicited additional inhibition in carbachol responses in the presence of FCCP. Inositol triphosphate contraction was inhibited by *OH whereas none of the radicals affect carbachol induced calcium sensitization. These results show that H(2)O(2) and *OH affects sarcoplasmic reticulum where O(2) * acts on mitochondria to change contractions in rat detrusor smooth muscle.     

Antioxidant activity of nasunin, an anthocyanin in eggplant peels

The free radical scavenging activities and inhibitory effect of lipid peroxidation of a delphinidin derivative in eggplant were investigated. Delphinidin-3-(p-coumaroylrutinoside)-5-glucoside (nasunin), an anthocyanin, was isolated as purple colored crystals from eggplant peels. Using electron spin resonance spectrometry and 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), hydroxyl radicals (OH) or superoxide anion radicals (O(2)(-)) generated by the Fenton reaction or the hypoxanthine-xanthine oxidase system were measured as DMPO-OH or DMPO-OOH spin adducts. L-Ascorbic acid 2-[3, 4-dihydro-2,5,7,8-tetramethyl-2-(4,8, 12-trimethyltridecyl)-2H-1-benzopyran-6yl-hydrogen phosphate] potassium salt (EPC-K1) and bovine erythrocyte superoxide dismutase (SOD) were used as standards for OH and O(2)(-) scavengers, respectively. Nasunin showed potent O(2)(-) scavenging (143+/-8 SOD-equivalent U/mg) and OH scavenging (0. 65+/-0.07 EPC-K1-equivalent micromol/mg) activities. Then, by changing the concentration of DMPO to vary the trapping rate of OH, the presence of a competitive reaction between nasunin and OH was studied. The 50% inhibition dose (ID(50)) obtained from the inhibition curve did not change, indicating OH scavenging of nasunin is not due to direct scavenging but inhibition of OH generating system by chelating ferrous ion. Nasunin protection against H(2)O(2)-induced lipid peroxidation in rat brain homogenate was measured at 586 nm using the indicator of malonaldehyde and 4-hydroxyalkenals. Nasunin (<50 microM) protected against lipid peroxidation of brain homogenates. The findings suggest that nasunin is a potent O(2)(-) scavenger and has protective activity against lipid peroxidation.     

Hydroxyl radical participation in the in vitro effects of gram-negative endotoxin on cardiac sarcolemmal Na,K-ATPase activity.

The effect of in vitro exposure of sarcolemmal membrane (SL) vesicles to Gram-negative endotoxin lipopolysaccharides (LPS) was studied. LPS decreased the Na,K-ATPase activity of SL vesicles; this effect was inhibited by hydroxyl radical (.OH) scavengers such as dimethylthiourea and dimethyl sulfoxide, but not by superoxide dismutase, a scavenger of superoxide anion radicals or by the hydrogen peroxide scavenger catalase. ESR spin-trapping with 5,5-dimethyl-1-pyrroline N-oxide verified the generation of .OH from LPS itself under the conditions used; .OH generated from LPS was not affected by deferoxamine, a powerful iron chelator. The Na,K-ATPase activity was reduced by an .OH radical generating system consisting of dihydroxyfumarate and Fe3(+)-ADP. Furthermore, exposure of SL vesicles to LPS caused an increase in malondialdehyde formation. It can be concluded that LPS damages cardiac SL by an oxygen free radical mechanism by the generation of .OH, due to inhibition of Na,K-ATPase activity and peroxidation of lipids, and that the effect of LPS is not dependent on the presence of contaminating iron.     

Effect of thiol drugs on the oxidative hemolysis in human erythrocytes

The effect of different thiol drugs and 2-methyl-thiazolidine-2,4-dicarboxylic acid on the oxidative stress, induced by hydrogen peroxide, was examined in human erythrocytes. The results indicated that captopril (CA), methimazole, N-acetylcysteine (NAC), penicillamine and precursor of L-cysteine 2-methyl-thiazolidine-2,4-dicarboxylic acid (CP) might protect the erythrocyte membrane against lipid peroxidation in the experimental conditions. Captopril, methimazole and penicillamine had the strongest antioxidative properties at the concentration level of 0.5 mM. The protective effects gradually decreased at higher and lower concentrations of these drugs. Contrary, the antioxidative properties of N-acetylcysteine increased with its levels growing in the reaction mixture, and only N-acetylpenicillamine did not protect erythrocytes against oxidative damages. The effect of 2-methyl-thiazolidine-2,4-dicarboxylic acid showed in these in vitro experimental conditions that it could act as an antioxidant at the concentration as high as 5 mM and higher.     

Studies on the activation of molecular oxygen and the biological defence mechanism against active oxygen species

One of the active oxygen species, superoxide (O2-), was generated by the electrolytic reduction of molecular oxygen in acetonitrile. O2- was determined by the ultraviolet (UV) (lambda max/nm = 255, epsilon = 1.48 x 10(3) M-1 cm-3) and the electron spin resonance (ESR) (g parallel = 2.083, g perpendicular = 2.008) spectrum. O2- could easily react with tocopherols (vitamin E and its derivatives) to give the corresponding chromanoxyl radicals of which structures were determined by ESR. ESR studies of the reactions of O2- with tocopherols or their model compounds indicate that the radical concentrations from tocopherol models correlate with the physiological activities of the tocopherols. O2- could also react with some biologically active quinones such as vitamin K3 and vitamin E quinone to give the corresponding semiquinone radicals. The fact that vitamin E quinone, an irreversible metabolite of vitamin E, was reduced by O2- to the semiquinone radical suggests that, like vitamin E, vitamin E quinone may also scavenge O2- and protect living cells from the effects of O2- in a hydrophobic environment. Further, O2- could react with some metalloporphyrins. In this case, non-redox metalloporphyrins such as Zn(II)TPP (TPP: tetraphenylporphine), Cd(II)TPP, Mg(II)TPP generated the superoxide adduct by the reaction with O2-. On the other hand, redox-active metalloporphyrins such as Cr(III)TPP.Cl, Mn(III)TPP.Cl, Co(II)TTP TTP: tetra-p-tolylporphine) and Co(III)TTP.Cl underwent the addition and/or redox reactions with O2-. Another active oxygen species, hydroxyl radical (OH.), was first detected from some copper (II) coplexes such as Cu(en)2 (en: ethylenediamine) with hydrogen peroxide (H2O2) by ESR spin trapping and thiobarbituric acid (TBA) methods. Further, by using Cu(en)2-H2O2 system the most active OH. scavenger was determined. This Cu(en)2-H2O2 system will be useful for determing the antioxidant ability against OH..     

棉花皮苷的抗氧化活性及对肝癌细胞生长的抑制作用研究

目的探讨棉花皮苷的抗氧化性及对肝癌细胞生长的抑制作用。方法本研究首次应用化学发光法和分光光度法系统评价了棉花皮苷对超氧阴离子自由基(O2.)、羟基自由基(.OH)、H2O2等活性氧和二苯代苦味酰基自由基(DPPH.)自由基的清除作用。采用MTT实验评价了棉花皮苷对人肝癌HepG2细胞生长的影响。结果棉花皮苷对O2.、.OH、H2O2和DPPH.等自由基均有较强的清除能力。与常用的抗氧化剂Vit C相比,其抗氧化效果明显强于VitC。棉花皮苷可以以剂量及时间依赖方式抑制肝癌细胞生长、增殖。结论棉花皮苷具有较强的抗氧化和抗肝癌生物活性。     

甘草类黄酮对脂质过氧化和活性氧自由基的作用

本文用比色法测定过氧化脂质研究甘草类黄酮(FG)的抗脂质过氧化作用,并用化学发光法和自旋捕集技术检测该药对O₂^-和OH^·的清除作用。体外实验表明:FG 2.8～25μg/ml可明显抑制小鼠肝匀浆在振荡温育条件下引起的丙二醛升高;FG 0.265～26.5μg/ml或2.58～258 μg/ml分别对碱性二甲基亚砜或黄嘌呤/黄嘌呤氧化酶体系生成的O₂^-有显著清除作用;以上作用呈浓度依赖性变化。FG 144 μg/ml或258 μg/ml分别对PMA刺激多形核白细胞释放的O₂^-及OH^·或Fenton's反应生成的OH^·有明显清除作用。实验证明FG有抗氧化作用。