Chem. Res. Toxicol. 1996, 9, 3-7
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Communications An Adduct between Peroxynitrite and 2′-Deoxyguanosine: 4,5-Dihydro-5-hydroxy-4-(nitrosooxy)-2′-deoxyguanosine Thierry Douki,† Jean Cadet,† and Bruce N. Ames*,‡ CEA, De´ partement de Recherche Fondamentale sur la Matie` re Condense´ e, SESAM/LAN, F-38054 Grenoble Cedex 9, France, and Division of Biochemistry and Molecular Biology, Barker Hall, University of California, Berkeley, California 94720-3202 Received July 12, 1995X
The reactivity of peroxynitrite (OONO-) with DNA is of interest because it is released during chronic inflammation, a major contributor to cancer. Peroxynitrite was found to undergo homolytic addition to the C4-C5 double bond of 2′-deoxyguanosine with formation of 4,5dihydro-5-hydroxy-4-(nitrosooxy)-2′-deoxyguanosine (nox-dG). This adduct may be a useful marker in the study of the chemical processes associated with the mutagenicity of peroxynitrite.
Introduction Peroxynitrite (ONOO-) is produced in vivo by reaction of nitric oxide (NO) with superoxide anion (O2•-) (1, 2). Concomitant release of NO + O2•- by activated macrophages and neutrophils results in the formation of peroxynitrite by the phagocytic cells of the immune system (3, 4). Recent studies suggest its relevance in several degenerative processes associated with inflammation, neurodegeneration, and sclerosis (5-9). A major exogenous source of peroxynitrite is cigarette smoke (10). Under physiological conditions, the major reactive form of peroxynitrite (pKa about 7 (11)) is the protonated form, peroxynitrous acid (HOONO), which decomposes with a half-life of about 1 s, yielding products with the reactivity of •NO2 and •OH (1). Oxidation of small molecules (12, 13), proteins (14, 15), and lipids (16) and nitration and hydroxylation of the aromatic rings of amino acids have been described (17, 18). The cytotoxicity of peroxynitrite has been shown in Escherichia coli (19), where it is more lethal than nitric oxide (20). In isolated DNA, nucleotide strand breaks have been observed upon peroxynitrite treatment (21). We describe here a major adduct of peroxynitrite with 2′-deoxyguanosine, 4,5-dihydro-5-hydroxy-4-(nitrosooxy)-2′-deoxyguanosine (nox-dG), a possible contributor to the mutagenicity and carcinogenicity of inflammation.
Experimental Procedures Chemicals. 2′-Deoxyguanosine (dG) was purchased from Pharma-Waldorf (Geneva, Switzerland). Sodium nitrite and hydrogen peroxide (30% w/w) were obtained from Merck (Darmstadt, Germany) and Sigma (St Louis, MO), respectively. tertButyl nitrite and 99.9% iron powder were products from Aldrich (Milwaukee, WI). Deuterium oxide (99.98% and 99.8%) and DMSO-d6 (99.96%) were purchased from Eurisotop (St. Aubin, France). † ‡ X
CEA. University of California, Berkeley. Abstract published in Advance ACS Abstracts, December 15, 1995.
0893-228x/96/2709-0003$12.00/0
HPLC Systems. The high performance liquid chromatography system consisted of a L6000 Hitachi pump (Tokyo, Japan) equipped with a Rheodyne Model 7125 loop injector (Cotati, CA) and a self-packed semipreparative Nucleosil 100-10 C18 octadecylsilyl silica gel column (250 × 7 mm i.d., particle size: 10 µm) (Macherey Nagel, Du¨ren, Germany). The isocratic eluent was a 95:5 (v/v) mixture of 25 mM ammonium formate aqueous solution and methanol. The flow rate was 3 mL‚min-1. A Hypersil NH2 amino silica gel column (250 × 4.6 mm i.d., particle size: 5 µm) (Interchim, Montluc¸ on, France) was used in the final purification. The isocratic eluent was a 95:5 (v/v) mixture of acetonitrile and 25 mM ammonium formate, at a flow rate of 1.5 mL‚min-1. The elution was monitored with a 2151 variable wavelength UV spectrophotometer (LKB-Bromma, Uppsala, Sweden) at either 230 or 380 nm. Spectroscopic Analysis. NMR analysis was carried out either in 99.98% D2O (after evaporation of the sample in 99.8% D2O to remove exchangeable protons) or in 99.96% DMSO-d6 solutions on a AC 200 Bruker spectrometer (Wissenbourg, France) at 200.13 MHz and 50.32 for 1H and 13C, respectively. Chemical shifts of nonexchangeable protons and secondary and tertiary carbons were inferred from first order spectrum analysis and are expressed with respect to potassium 3-([2,2,3,3-2H]trimethylsilyl)propionate (TSP) in water and tetramethylsilane (TMS) in DMSO-d6. Chemical shifts of carbons C4 and C5 were determined by 1H/13C long distance coupling analysis in 99.98% D2O on a Unity 400 spectrometer (Varian). The UV absorption spectra were determined in water with a DU-88 spectrophotometer (Beckman, Irvine, CA). GC/MS analysis was performed after trimethylsilylation in 500 µL of a 1:1 (v/v) mixture of pyridine and bis(trimethylsilyl)fluoroacetamide (Pierce, Rockford, IL). The sample was analyzed on a HP 5890 GC interfaced to a 5989 MS engine (Hewlett-Packard, Palo Alto, CA). A DB5 column (12 m long, 0.25 mm internal diameter, 0.2 µm film thickness, J&W Scientifics, Folsom, CA) was used with helium as a carrier gas at a linear velocity of 35 cm‚s-1. The column was maintained at 100 °C for 1 min, raised to 280 °C with a 10 °C‚min-1 rate, and maintained at this temperature for 5 min. Fast atom bombardment mass spectrometry in the positive mode (FAB+) was carried out on a VG ZAB 2-EQ apparatus (Manchester, U.K.). Isolation of the Peroxynitrite-2′-Deoxyguanosine Addition Product. Sodium peroxynitrite was synthesized by oxidation of nitrous acid with hydrogen peroxide (22). The concentration of the sodium peroxynitrite solution was deter-
© 1996 American Chemical Society
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mined to be 30 mM by measuring the UV absorption at 302 nm (302 ) 1670 M-1‚cm-1 in 1 M sodium hydroxide). 2′-Deoxyguanosine (500 mg) was added under stirring to the solution of sodium peroxynitrite in an ice bath. The pH was monitored and adjusted from 11 to 7.5, after which it spontaneously decreased to 5 within 2 min. The solution was neutralized with sodium hydroxide, the volume was reduced to 20 mL in vacuo, and the resulting solution was freeze-dried. The residue was extracted with 2 mL of hot water and centrifuged. The supernatant fraction was prepurified by reversed phase HPLC on a semipreparative column. The fraction containing the major UV-absorbing peak (from 4.5 to 8 min) was collected. After concentration in vacuo, the sample was purified further on the same HPLC system. The fraction corresponding to the peak observed at 5.1 min on the chromatogram was collected, concentrated in vacuo, and freeze-dried. The residue was dissolved in acetonitrile and water (95:5 v/v) and injected on the HPLC system equipped with an amino silica gel column. The adduct eluted at 17 min and was collected, providing 3 mg of pure material (overall yield: 3%). The purity was inferred from analysis on both HPLC systems with the UV spectrophotometer set at 230 nm. An aliquot (500 µg) of the sample was analyzed by GC/MS. The tri-trimethylsilylated (tR ) 18.8 min) and di-trimethylsilylated (tR ) 17.6 min) products were observed. One milligram of adduct was dried in vacuo and acetylated overnight in a 3:1 (v/v) mixture of pyridine and acetic anhydride. After removal of the excess of reagent in vacuo, the residue was suspended in water and injected on the RP-HPLC column, using a mixture of water and methanol (50:50) as isocratic eluent. The 380 nm absorbing compound eluted after 5 min and was collected and analyzed by FAB+ mass spectrometry. 4,5-Dihydro-5-hydroxy-4-(nitrosooxy)-2′-deoxyguanosine: UV (H2O, λmax): 230 and 380 nm. 1H NMR (D2O, TSP, 200.13 MHz): δ (ppm) 7.82 (1 H, H8), 6.19 (1 H, H1′), 4.63 (1 H, H3′), 4.17 (1 H, H4′), 3.89 (1 H, H5′), 3.79 (1 H, H5′′), 2.72 (1 H, H2′), 2.56 (1 H, H-2′′); 13C NMR (D2O, TSP, 50.13 MHz): δ (ppm) 41.4 (C2′) 62.0 (C5′), 73.4 (C3′), 86.6 (C4′), 89.7 (C1′), 134.8 (C8), 135.5 (C5), 145.8 (C4); GC/MS, m/z (rel intensity) 546 ([M: dG + HOONO + 3TMS], 10), 535 ([M - 1CH3], 7), 516 ([M NO], 100), 501 ([M - NO - 1CH3], 78); FAB+-MS, m/z (rel intensity) 413 ([M+: dG + HOONO + 2OAc + H+], 80) 383 ([M+ - NO], 42) 341 ([M+ - NO - CH3CO + H], 100) 299 ([M+ NO - 2CH3CO + 2H], 33). Reduction and Acidic Hydrolysis Experiments. Reduction of the adduct was carried out by addition of 50 mg of iron powder and 500 µL of 12 M HCl. After 5 min at room temperature, the sample was neutralized by addition of 600 µL of 10 M sodium hydroxide and centrifuged. The UV absorption spectra of the supernatant fraction were determined after dilution with 1 mL of water. A similar procedure was used for tert-butyl nitrite. Acidic treatment of both the adduct and tertbutyl nitrite were carried out by dissolving the products in 500 µL of concentrated HCl followed by neutralization.
Results HPLC analysis of the reaction mixture (Figure 1) revealed a polar yellow adduct, nox-dG. The adduct was further purified by a combination of reverse phase and normal phase HPLC and then characterized by UV, GC/ MS, FAB+-MS, and NMR. An additional compound eluted at 9 min and was isolated. It was identified as the 8-nitroguanine previously characterized (23) on the basis of its 400 nm absorption, a typical feature of aromatic rings carrying a nitro group (24), and its GC/ MS mass spectrum. A recent paper has also reported the isolation and the characterization of 8-nitroguanine after treatment of guanine with peroxynitrite (25). In our experiment 8-nitroguanine could have been produced either from guanine (depurinated dG) by peroxynitrite or by depurination of 8-nitro-2′-deoxyguanosine generated from dG during the decomposition of HOONO.
Communications
Figure 1. HPLC elution profile of the crude reaction mixture of peroxynitrite and dG. HPLC conditions: semipreparative C18 column; eluent: 25 mM ammonium formate in water and methanol (95:5); flow rate: 3 mL‚min-1; detection wavelength: 380 nm.
The nox-dG adduct was analyzed by GC/MS following trimethylsilylation (Figure 2a). A FAB+-MS analysis was done after acetylation (Figure 2b) in order to decrease its polarity and therefore to facilitate its desorption from the glycerol matrix. The results are consistent with a molecular weight of 330, i.e., consistent with the addition of HOONO to 2′-deoxyguanosine. Further support for a 2′-deoxyribonucleoside structure for the adduct was provided by the 1H and 13C NMR spectra (Figure S1). Moreover, the adduct was converted into a characteristic gray compound when heated on a thin layer chromatography plate after spraying a solution of cysteine in 3 M sulfuric acid, a specific reagent for 2-D-erythro-pentose (e.g., deoxyribose) (26). Evidence that the adduct contains a nitrosooxy function (sONdO) is as follows. The adduct exhibited absorption at 380 nm, which was lost after HCl/iron reduction, likely to be due to the conversion of -ONO into -OH. tert-Butyl nitrite, used as a model compound of an organic nitrous ester, was found to have a 380 nm absorption which was lost on reduction. This reaction is not definitive proof of structure as it also reduces -NO2 to -NH2 with the loss of absorption. Acidic treatment did not induce the loss of the 380 nm absorption, either in tert-butyl nitrite or in the adduct. Unambiguous
Communications
Chem. Res. Toxicol., Vol. 9, No. 1, 1996 5 Table 1. 1H NMR Features of Nox-dG and dG in D2Oa δ (ppm) protons
nox-dG
dG
H1′ H2′ H2′′ H3′ H4′ H5′ H5′′ H8
6.19 2.72 2.56 4.63 4.17 3.89 3.79 7.82
6.38 2.86 2.58 4.70 4.21 3.91 3.83 8.05 J (Hz)
a
proton-proton
nox-dG
dG
1′-2′ 1′-2′′ 2′-2′′ 2′-3′ 2′′-3′ 3′-4′ 5′-4′ 5′-5′′ 5′′-4′
6.6 6.7 -14.0 6.4 4.5 3.9 3.8 -12.4 5.1
6.4 6.3 -14.0 7.3 3.6 2.7 3.7 -12.0 4.6
Chemical shifts are expressed in ppm with respect to TSP.
Table 2. Chemical Shifts of the Exchangeable Protons of Nox-dG and dG in DMSO-d6 Expressed in ppm with Respect to TMS δ (ppm)
Figure 2. (a) GC/MS mass spectrum of the trisilylated derivative of the adduct. The spectrum is the average of the peak eluting at 18.6 min. (b) FAB+ mass spectrum of the diacetylated derivative of the adduct.
characterization of the nitrosooxy function was provided by the observation of the loss of a NO group (M - 30) on both GC/MS and FAB+-MS mass spectra of the derivatized adduct. This ruled out the possibility of nox-dG as a nitration product, since loss of a NO2 fragment should be observed at a higher intensity than the M - 30 ion. Extensive 1H and 13C NMR analysis has made possible the determination of the structure of the base moiety of the adduct, mainly on the basis of a comparison with dG (Table 1). Heteronuclear long distance coupling was observed between H8 on the one hand, and C4 and C5 on the other hand. These results unambiguously showed that no cleavage of the C5-N7 bond had taken place. Moreover, the observation of a pseudo triplet in both D2O and DMSO-d6 for the signal of H1′ indicates the absence of a proton on N9. This provided evidence that the C8N9 bond had not been hydrolyzed. Together, these results show that the imidazole ring had not opened. This ruled out the possibility that the adduct had a structure similar to that of formamidopyrimidine (27) or oxazolone (28), two products formed from the reaction of •OH with dG. The signals corresponding to the protons of a NH2 and a NH group were observed at 6.11 and 8.80 ppm, respectively. This observation suggests that the pyrimidine ring of dG is not modified. This result was confirmed by the lack of detectable long distance coupling of the NH2 group. This is likely due to the fact that, under our NMR experimental conditions, mainly coupling through three bonds was observed. In the pyrimidine ring on dG, the protons of the amino group on position 2 are only three bonds from two nitrogen atoms. Altogether, it can thus be concluded that the purine structure is still intact.
protons
nox-dG
dG
NH2 NH 5′-OH 3′-OH
6.11 8.80 5.25 4.97
6.45 10.61 5.27 4.95
Table 3.
13C
NMR Features of Nox-dG and dG in D2Oa δ (ppm)
a
carbons
nox-dG
dG
1′ 2′ 3′ 4′ 5′ 4 5 8
89.7 41.4 73.4 86.6 64.1 145.8 135.5 134.8
89.0 38.9 73.4 85.2 84.6 154.0 119.6 138.7
Chemical shifts are expressed in ppm with respect to TSP.
The positions of the hydroxyl and nitrosooxy groups on the purine ring were determined by analysis of the UV and NMR spectra. The lack of an absorption maximum around 260-270 nm in the UV spectrum is strongly indicative of loss of aromaticity on both the imidazole and the pyrimidine ring of the guanine moiety. This is confirmed by a low upfield shift of the signal of the 1′, 2′, and 3′ protons of the deoxyribose ring (0.19, 0.14, and 0.07 ppm, respectively), due to the modification of the electronic structure of the base and the observation of a downfield shift of the signal of the NH group of nox-dG compared to dG (∆δ ) -1.81 ppm). The similarity of the 1H NMR spectra of the glycosidic moieties of dG and that of the adduct suggests that both compounds are in the same anti conformation around the C1′-N9 bond. A dramatic upfield shift of H2′ due to a preferential syn conformation is expected for 8-substituted derivatives, as reported for 7,8-dihydro-8-oxo-2′-deoxyguanosine (29). This, together with the observation that the H8 proton is still present, provides evidence that the C8 position is not modified in the adduct. Moreover, the high similarity between the NMR feature of the deoxyribose moieties of
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dG and nox-dG shows that no sugar modification occurred in the formation of the latter compound. In particular, the observation of the protons of the 3′ and 5′ OH rules out the possibility of an esterification by nitrous acid of either one of these positions. Altogether, these results make the C4-C5 double bond very likely to be the reaction site, as shown by the observation of marked changes in the chemical shift of the C4 and C5 carbons with respect to dG (∆δ ) -8.2 and +15.9 ppm, respectively). The high chemical shifts of H8 (δ ) 7.82 ppm) and C8 (δ ) 134.8 ppm) show that the N7-C8 double bond has not been saturated during the addition of HOONO. The small upfield shift of C8 (∆δ ) 3.9 ppm) with respect to dG is likely to be due to the loss of the aromaticity. Finally, observation of a resonance at higher frequency for C4 (δ ) 145.8 ppm) than C5 (δ ) 135.5 ppm) makes the C4 position more likely to carry the ONO group which is more electronegative than a hydroxyl group. We therefore conclude that the most likely structure for the adduct is 4,5-dihydro-5-hydroxy-4(nitrosooxy)-2′-deoxyguanosine. No trace of nox-dG was detected in dG solution acidified in the presence of either H2O2, NaNO2, or decomposed NaOONO. This indicates that the products used for the synthesis of the peroxynitrite solution are not involved in the reaction. Similarly, the decomposition products of peroxynitrite do not react with dG to yield nox-dG. These observations show that peroxynitrite is the actual reactive species. This is confirmed by the formation of nox-dG upon acidification of a solution of dG containing 100 mM H2O2 and 100 mM NaNO2 (data not shown). Under the latter conditions, peroxynitrous acid is produced in situ and leads to the formation of the same adduct as is produced from NaOONO. Involvement of peroxynitrite in the formation of nox-dG was unambiguously established by the observation of a 7-fold increase in the yield of nox-dG when the reaction was carried out at 0 °C instead of room temperature. This ratio is very close to that of the lifetime of HOONO (7 and 1 s at 0 and 37 °C, respectively) (11), which strongly suggests that the limiting factor in the yield of nox-dG is the concentration of peroxynitrous acid, in agreement with a bimolecular mechanism.
Discussion The formation of an adduct with a nitrosooxy function in the addition reaction between peroxynitrite and 2′deoxyguanosine is described. This addition reaction represents a new type of reaction of HOONO with biomolecules in addition to oxidation (1, 16) and nitration (17, 18, 24). Though nitrosooxy derivatives are not very common compounds, they are produced upon HOONOinduced lipid peroxidation (30) or by hydroperoxides reacting with NO (31) or NO2 (32). The isolation of a product exhibiting both nitrosooxy and hydroxyl functions on the C4 and C5 positions is strongly indicative that the addition of HOONO occurs with homolytic cleavage of the peroxide bond (-O-O-). This is the weakest bond within the peroxynitrous acid molecule (11). The reaction is likely to occur via a concerted mechanism, since if a cleavage of peroxynitrite into HO• and •NO2 had taken place before the reaction, a nitro group would have been expected because the latter radical is centered on the nitrogen atom (33). The reaction appears to be stereoselective because of the 2-deoxyribose ring. Even in the case of a syn addition, two diastereoisomers should be
Communications
produced. We did not observe any evidence for such a result since all NMR spectra exhibit only one signal per proton or carbon. In addition, no double peak was observed on the HPLC elution profiles, either on the reverse phase or on the amino column. Isomerization of HOONO from the cis form to a more reactive trans form (18, 34) has been postulated. The trans form may be the species that yields the dG adduct, and the adduct may be a unique probe of HOONO. This work describes the first identification of a specific modification of a DNA component with peroxynitrite. The detection of this compound in cellular DNA could be used as a marker of the action of peroxynitrite in vivo and also could provide insights on nitric oxide mediated mutagenesis (35, 36). The generation of peroxynitrite by phagocytic cells is likely to be a key link in the relation between chronic inflammation and cancer (37, 38).
Acknowledgment. We are indebted to J. Beckman, S. Christen, W. Pryor, and M. Shigenaga for suggestions. This work was supported by National Cancer Institute Outstanding Investigator Grant CA39910 and National Institute of Environmental Health Sciences Center Grant ESO1896 to B.N.A. Supporting Information Available: The supporting information consists of Figure S1 which is the 1H and 13C NMR spectra for the 2′-deoxyribonucleoside structure of the nox-dG adduct (1 page). Ordering information is given on any current masthead page.
References (1) Beckman, J. S., Beckman, T., W., Chen, J., Marshall, P. A., and Freeman, B. (1990) Apparent hydroxyl radical production by peroxynitrite: implications for endothelial injury from nitric oxide and superoxide. Proc. Natl. Acad. Sci. U.S.A. 87, 1620-1624. (2) Blough, N. V., and Zafiriou, O. C. (1985) Reaction of superoxide with nitric oxide to form peroxonitrite in alkaline aqueous solution. Inorg. Chem. 24, 3502-3504. (3) Ischiropoulos, H., Zhu, L., and Beckman, J. S. (1992) Peroxynitrite formation from macrophage-derived nitric oxide. Arch. Biochem. Biophys. 288, 446-451. (4) Carreras, M. C., Pargament, G. A., Catz, S. D., Poderoso, J. J., and Boveris, A. (1994) Kinetics of nitric oxide and hydrogen peroxide production and formation of peroxynitrite. FEBS Lett. 341, 65-68. (5) Lipton, S. A., Chol, Y.-B., Pan, Z.-H., Lei, S. Z., Chen, H.-S. V., Sucher, N. J., Loscalzo, J., Singel, D. J., and Stamler, J. S. (1993) A redox-based mechanism for the neuroprotective and neurodestructive effects of nitric oxide and related nitroso-compounds. Nature 364, 626-631. (6) Olanow, C. W. (1993) A radical hypothesis for neurodegeneration. Trends Neurol. Sci. 11, 439-444. (7) Beckman, J. S., Carson, M., Smith, C. D., and Koppenol, W. H. (1993) ALS, SOD and peroxynitrite. Nature 364, 584. (8) Hogg, N., Darley-Usmar, V. M., Graham, A., and Moncada, S. (1993) Peroxynitrite and atherosclerosis. Biochem. Soc. Trans. 21, 358-362. (9) White, C. R., Brock, T. A., Chang L.-Y., Crapo, J., Briscoe, P., Ku, D., Bradley, W. A., Gianturco, S. H., Gore, J., Freeman, B. A., and Tarpey, M. M. (1994) Superoxide and peroxynitrite in atherosclerosis. Proc. Natl. Acad. Sci. U.S.A. 91, 1044-1048. (10) Pryor, W. A., and Stone, K. (1993) Oxidants in cigarette smoke. Tabacco smoking and nutrition. Ann. N.Y. Acad. Sci. 686, 1228. (11) Edwards, J. O., and Plumb, R. C. (1994) The chemistry of peroxynitrites. Prog. Inorg. Chem. 41, 599-634. (12) Pryor, W. A., Jin, X., and Squadrito, G. L. (1994) One- and twoelectron oxidations of methionine by peroxynitrite. Proc. Natl. Acad. Sci. U.S.A. 91, 11173-11177. (13) Hogg, N., Joseph, J., and Kalyaraman, B. (1994) The oxidation of R-tocopherol and Trolox by peroxynitrite. Arch. Biochem. Biophys. 314, 153-158. (14) Moreno, J. J., and Pryor, W. A. (1992) Inactivation of R1proteinase inhibitor by peroxynitrite. Chem. Res. Toxicol. 5, 425431.
Communications (15) Schmidt, K., Klatt, P., and Mayer, B. (1994) Reaction of peroxynitrite with oxyhaemoglobin: interference with photometrical determination of nitric oxide. Biochem. J. 301, 645-647. (16) Radi, R., Beckman, J. S., Bush, K. M., and Freeman, B. A. (1991) Peroxynitrite-induced membrane lipid peroxidation: the cytotoxic potential of superoxide and nitric oxide. Arch. Biochem. Biophys. 288, 481-487. (17) Ischiropoulos, H., Zhu, L., Chen, J., Tsai, M., Martin, J. C., Smith, C. D., and Beckman, J. S. (1992) Peroxynitrite-mediated tyrosine nitration catalyzed by superoxide dismutase. Arch. Biochem. Biophys. 298, 431-437. (18) Beckman, J. S., Ischiropoulos, H., Zhu, L., van der Woerd, M., Smith, C., Chen, J., Harrison, J., Martin, J. C., and Tsai, M. (1992) Kinetics of superoxide dismutase- and iron-catalyzed nitration of phenolics by peroxynitrite. Arch. Biochem. Biophys. 298, 438445. (19) Zhu, L., Gunn, C., and Beckman, J. (1992) Bactericidal acitivity of peroxynitrite. Arch. Biophys. Biochem. 298, 452-457. (20) Brunelli, L., Crow, J. P., Beckman, J. (1995) The comparative toxicity of nitric oxide and peroxynitrite to Escherichia coli. Arch. Biochem. Biophys. 316, 327-334. (21) King, P. A., Anderson, V., Edwards, J. O., Gustafson, G., Plumb, R. C., and Suggs, J. W. (1992) A stable solid that generates hydroxyl radical upon dissolution in aqueous solutions: reaction with proteins and nucleic acids. J. Am. Chem. Soc. 114, 54305432. (22) Hughes, B. N., and Nicklin, H. G. (1968) The chemistry of pernitrites. Part I. Kinetics of decomposition of pernitrous acid. J. Chem. Soc. (A), 450-452. (23) Shigenaga, M. K., Christen, S., Douki, T., Yeo, H. C., Lee, H. H., and Ames, B. N. (1994) Nitration of guanine and its nucleoside derivatives at the C-8 position by nitronium ion, nitrogen dioxide, and peroxynitrite. Abstract. International Society for Free Radical Research, 7th Biennial Scientific Meeting, Sydney, Australia. (24) van der Vliet, A., O’Neil, C. A., Helliwell, B., Cross, C. E., and Kaur, H. (1994) Aromatic hydroxylation and nitration of phenylalanine and tyrosine by peroxynitrite. Evidence for hydroxyl radical production from peroxynitrite. FEBS Lett. 339, 89-92. (25) Yermilov, V., Rubio, J., Becchi, M., Friesen, M. D., Pignatelli, B., and Ohshima, H. (1995) Formation of 8-nitoguanine by the reaction of guanine with peroxynitrite in vitro. Carcinogenesis 16, 2045-2050.
Chem. Res. Toxicol., Vol. 9, No. 1, 1996 7 (26) Buchanan, J. G. (1951) Detection of deoxyribonucleotides on paper chromatograms. Nature 168, 1091. (27) Cadet, J., and Berger, M. (1985) Radiation-induced decomposition of the purine bases within DNA and related model compounds. Int. J. Radiat. Biol. 47, 127-143. (28) Cadet, J., Berger, M., Buchko, G. W., Joshi, P. C., Raoul, S., and Ravanat, J.-L. (1994) 2,2-Diamino-4-[(3,5-di-O-acetyl-2-deoxy-βD-erythro-pento-furanosyl)amino]-5-(2H)-oxazolone: a novel and predominant radical oxidation product of 3′-5′-di-O-acetyl-2′deoxyguanosine. J. Am. Chem. Soc. 116, 7403-7404. (29) Culp, S. J., Cho, B. P., Kadlubar, F. F., and Evans, F. E. (1989) Structural and conformational analyses of 8-hydroxy-2′-deoxyguanosine. Chem. Res. Toxicol. 2, 416-422. (30) Rubbo, H., Radi, R., Trujillo, M., Telleri, R., Kalyanaraman, B., Barnes, S., Kirk, M., and Freeman, B. A. (1994) Nitric oxide regulation of superoxide and peroxynitrite-dependent lipid peroxidation. J. Biol. Chem. 269, 26066-26075. (31) Shelton, J. R., and Kopczewski, R. F. (1967) Nitric oxide induced free-radical reactions. J. Org. Chem. 32, 2908-2910. (32) Pryor, W. A., Castle, L., and Church, D. F. (1985) Nitrosation of organic hydroperoxides by nitrogen dioxide/dinitrogen tetraoxide. J. Am. Chem. Soc. 107, 211-217. (33) Cotton, F. A., and Wilkinson, F. R. S. (1972) Nitrogen. In Advanced Inorganic Chemistry, pp 357, John Wiley and Sons, New York. (34) Koppenol, W. H., Moreno, J. J., Pryor, W. A., Ischiropoulos, H., and Beckman, J. S. (1992) Peroxynitrite, a cloaked oxidant formed by nitric oxide and superoxide. Chem. Res. Toxicol. 5, 834-842. (35) Arroyo, P. L., Hatch-Pigott, V., Mower, H. F., and Cooney, R. V. (1992) Mutagenicity of nitric oxide and its inhibition by antioxidants. Mutat. Res. 281, 190-202. (36) Stamler, J. S., Singel, D. J., and Loscalzo, J. (1992) Biochemistry of nitric oxide and its redox-activated forms. Science 258, 18981902. (37) Ames, B. N., Shigenaga, M. K., and Hagen, T. M. (1993) Oxidants, antioxidants, and the degenerative diseases of aging. Proc. Natl. Acad. Sci. U.S.A. 90, 7915-7922. (38) Ames, B. N., Gold, L. S., and Willett, W. C. (1995) The causes and prevention of cancer. Proc. Natl. Acad. Sci. U.S.A. 92, 5258-5265.
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