Free radical research communications最新文献

Electron paramagnetic resonance/spin trapping studies were applied, to verify the superoxide radical scavenging activity of two non-toxic, water soluble dihydroquinoline type antioxidants, CH402 (Na-2,2-dimethyl-1,2-dihydroquinoline-4-yl methane sulphonate and MTDQ-DA (6,6-methylene bis 2,2-dimethyl-4-methane sulphonic acid: Na-1,2-dihydroquinoline). Results were compared with other indirect methods such as the amperometric, spectrophotometric and luminometric methods, respectively. Both dihydroquinoline type antioxidants scavenged superoxide in vitro specifically. MTDQ-DA scavenged superoxide an order of magnitude faster than CH-402. Neither CH402 nor MTDQ-DA affected the hypoxanthine/xanthine oxidase superoxide generating system, nor did they inhibit xanthine oxidase directly.

In HeLa cells evidence is provided that active oxygen species such as hydrogen peroxide and superoxide at low levels are important growth regulatory signals. They may constitute a novel regulatory redox system of control superimposed upon the established cell growth signal transduction pathways. Whilst for example hydrogen peroxide can be added exogenously to elicit growth responses in these cells, it is clear that cellularly generated superoxide and hydrogen peroxide are important. Experiments with superoxide dismutase, superoxide dismutase mimics and inhibitors of both superoxide dismutase and xanthine oxidase suggest that superoxide generated intracellularly and superoxide released extracellularly are both relevant to growth control in HeLa cells.

The production of nitrate (NO3-) and nitrite (NO2-) from macrophage-derived NO was studied using EPR and spin trapping. The formation of NO3- was determined via EPR in reactions involving the iron-binding protein, lactoferrin. The formation of NO2- was determined via EPR/spin trapping in the reaction between NO2- and H2O2. Dissolved nitric oxide (NO.) was reacted with lactoferrin yielding an EPR spectrum (77 degrees K) different from the normal EPR spectrum obtained for lactoferrin, suggesting that NO. interacts with the ferric ions bound to lactoferrin forming a ferric-nitrosyl type complex. The EPR spectrum (77 degrees K) of this ferric-nitrosyl type complex was also observed in the supernatant fluid of macrophage cell suspensions following their stimulation with lipopolysaccharide (LPS). During LPS stimulation of macrophages, these cells generate NO. which in turn produces NO3- and NO2-. The ferric-nitrosyl type complex is formed in a reaction mixture containing apolactoferrin and bicarbonate following the reaction of Fe+2 with NO3-, generated from macrophage-derived NO(.), to produce Fe+3 and NO(.). Furthermore, in an acidic medium, NO2- reacts with H2O2 forming peroxynitrous acid (HOONO) which rapidly decomposes into hydroxyl radicals (.OH) and the nitrogen dioxide (NO2.) radical. In the supernatant fluid of LPS-stimulated macrophage suspensions, the production of .OH was verified by spin trapping using 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) as the spin trap and ethanol as the .OH scavenger. The EPR spectra corresponding to the DMPO-OH and the DMPO-hydroxyethyl adducts were identified.(ABSTRACT TRUNCATED AT 250 WORDS)

Spin-labeled oligonucleotides (S-probes) were synthesized and examined as DNA probes to monitor hybrid formation. TEMPO was introduced either at the internucleotide linkage of 5'-terminus (Type 1) or at the 5'-terminal hydroxyl group (Type 2) and both types of S-probes were used in this study. The presence of target DNA was detected in solution by EPR spectroscopy for both types of S-probes. Hybridization of the S-probes resulted in notable broadening of EPR line width, accompanied by a decrease in the EPR signal height ratio for I(-1)/I(0).I(-1)/I(0) of S-probes having no spacer between oligonucleotide and TEMPO decreased more markedly than that of S-probes with a spacer, indicating that TEMPO should be introduced to an oligonucleotide directly to monitor hybrid formation. When M13mp8 single-stranded DNA with or without an EcoRI recognition site was selected as a target DNA, hybrid formation was detected only for DNA containing EcoRI site in solution using spin-labeled oligonucleotides.

Allopurinol, a potent inhibitor of xanthine oxidase, is known to effectively protect the heart against damage in patients undergoing cardiac bypass surgery. There is still an ambiguity concerning the presence of xanthine oxidase in the human heart. Thus, the mechanism underlying the protective effect of allopurinol is unclear. Transition metal ions, such as iron and copper, can participate in single-electron reactions and mediate the formation of oxygen-derived free radicals. In this study the interaction between allopurinol and Cu(II) was investigated. Spectrophotometric investigation shows that allopurinol (0-0.8 mM) form a 1:1 complex with Cu(II) ions (0-0.8 mM) with a specific absorbance peak at 364 nm. Also, the rate constant (k) for the copper-catalyzed aerobic oxidation of ascorbate was markedly decreased in the presence of allopurinol (from 0.068 min-1 to 0.014 min-1). Allopurinol substantially reduced the copper-mediated and ascorbate-driven DNA breakage. Spectrophotometric measurements did not indicate a specific interaction between iron ions and allopurinol. It is suggested that the beneficial effects of allopurinol during reperfusion of the heart could stem from its chelation of copper, yielding a complex with low redox activity.

The relationships between structure and antioxidant activity of dihydrolipoic acid (DHLA) were studied using homologues of DHLA: bisonor-DHLA (a derivative which lacks two carbons in the hydrophobic tail), tetranor-DHLA (which lacks four carbons) and a methyl ester derivative. It was observed that: i) DHLA homologues with shorter hydrocarbon tails (i.e., bisnor- and tetranor-DHLA) had greater ability to quench superoxide radicals (O2-); ii) no differences among homologues with different chain lengths were found for peroxyl radical (ROO.) scavenging in aqueous solution, and iii) DHLA was the best membrane antioxidant in terms of ROO. scavenging and lipid peroxidation inhibition. Differences among the DHLA homologues in their antioxidant properties in polar and apolar environments generally agreed with differences in their partition coefficients. The methyl ester was the least effective antioxidant both in aqueous phase and in membranes. Tetranor-DHLA was found not only to be less effective in preventing ROO.-induced lipid peroxidation, but also to induce lipid peroxidation in the presence of residual iron. Thus, the complexity of biological systems seems to complicate generalizations on the correlation of molecular structure with antioxidant activity of DHLA.

Administration of daily doses of 0.1 mg of 3,5,3'-triiodothyronine (T3)/kg body weight for 3 consecutive days to fed rats elicited a calorigenic response in the animals, in concomitance with a 36% increase in the rate of 0(2) consumption by the liver. In these conditions, liver submitochondrial particles (SMP) from T3-treated rats exhibited marked increases in the rate of superoxide radical generation, both in the presence of NADH (142%) or succinate (152%). Furthermore, liver SMP from hyperthyroid animals released hydrogen peroxide at higher rates than those of euthyroid rats, either under basal conditions or in the succinate-supported process, both in the absence and presence of antimycin-A. It is concluded that the hyperthyroid state in the rat leads to a drastic enhancement in the capacity of liver mitochondria to produce active oxygen species, which correlates with the elevated respiratory rate observed in the intact organ.

The toxicity of the antineoplastic agent doxorubicin (DOX) has been shown to be moderated by the antioxidant enzyme glutathione peroxidase. It has been reported that acute doses of DOX can cause an inhibition of glutathione peroxidase in cardiac tissue, that may render this tissue especially susceptible to further prooxidant damage. In this study, multiple DOX treatments at a therapeutic dose were assessed for their effect on the antioxidant enzyme status of cardiac and kidney tissue. DOX was administered i.p. (5 mg/kg) once a week for two weeks to male balb/c mice. The activities of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPOX) and glutathione reductase (GR) were measured 1, 2 and 7 days following the second DOX treatment in both heart and kidney. Levels of reduced glutathione (GSH) were also measured in cardiac tissue at these same times. Cardiac levels of GPOX and GR showed a time-dependent decrease in activity, with 10% and 12% inhibition for GPOX and GR, respectively, at 7 days post second treatment. Cardiac levels of GSH also showed a significant decrease, approximately 15%, at 7 days post second treatment. Cardiac levels of SOD and CAT as well as kidney levels of all four antioxidant enzymes were not affected by DOX treatment. These data suggest that DOX given in a therapeutic regimen, at a therapeutic dose, can cause decreases in cardiac levels of GPOX, GR and GSH that could render the heart especially susceptible to further oxidative challenge.

The alpha-tocopheroxyl radical (alpha TR.)generated in the reaction with 1,1-diphenyl-2-picrylhydrazyl in n-butanol decayed according to second-order kinetics with a rate constant k alpha = 3 x 10(3) M-1s-1 as determined by EPR spectroscopy. Various biologically and pharmacologically active substances like isoprenaline (ISO), epinephrine (EPI), histamine (HIS), stobadine (STO), nafazatrom (NAF) and Kampo C medicine (KMC) accelerated the decay rate of alpha TR(.). The whole process is formally a third-order reaction with the rate constants (in 10(9) M-2s-1): ks(ISO) = 1.28, ks(NAF) = 1.25, ks(EPI) = 0.6, ks(HIS) = 0.4, and ks(STO) = 0.1. In the kinetics of the reaction mechanism, bimolecular intermediates are assumed and the rate constants of their formation were determined.

The effect of lipopolysaccharide (LPS) and/or bile acids on rat erythrocyte membranes was studied in vitro. Addition of LPS isolated from E. coli (J5 mutant) into the erythrocyte resulted in the decrease of membrane fluidity as determined by spin labelling using electron paramagnetic resonance (EPR). This was accompanied by membrane fragility. It was found that hydroxyl radicals were generated from erythrocytes treated with LPS by using DMPO spin trapping. However, pretreatment of erythrocytes with taurine-conjugated bile acids was found to modify the membrane response induced by LPS. Taurocholic acid (TCA) and tauroursodeoxycholic acid (TUDCA) prevented the decrease of membrane fluidity induced by LPS, and, as a result, the membrane integrity was maintained although no significant changes were observed in the amount of hydroxyl radicals produced by LPS addition. However, taurochenodeoxycholic acid (TCDCA) exhibited little beneficial effect on the dynamic properties and the function of the erythrocyte membranes, although the hydroxyl radical declined markedly in the erythrocytes. Therefore, it is suggested that TCA and TUDCA have a protective effect against LPS-induced membrane fragility by modulating membrane fluidity.