- Hydantoin derivative formation from oxidation of 7,8-dihydro-8-oxo- 2′-deoxyguanosine (8-oxodG) and incorporation of 14C-labeled 8-oxodG into the DNA of human breast cancer cells
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One-electron oxidation of 7,8-dihydro-8-oxo-2′-deoxyguanosine (8-oxodG) yielded a guanidinohydantoin derivative (dGh) and a spiroiminodihydantoin derivative (dSp), both putatively mutagenic products that may be formed in vivo. The nucleoside dGh was the major product at room temperature, regardless of pH. The results are contrary to previously published model studies using 2′,3′,5′-triacetoxy-8-oxo-7,8- dihydroguanosine (Luo, W.; Miller, J. G.; Rachlin, E. M.; Burrows, C. J. Org. Lett. 2000, 2, 613; Luo, W.; Miller, J.G.; Rachlin, E.M.; Burrows, C.J. Chem. Res. Toxicol. 2001, 14, 927), who observed a spiroiminodihydantoin derivative as the major product at neutral pH. Clearly, the functional groups attached to the ribose moiety of 8-oxodG influence the oxidation chemistry of the nucleobase derivative. To explore this chemistry in vivo, 14C-labeled 8-oxodG was synthesized and incubated with growing MCF-7 human breast cancer cells, resulting in the incorporation of the compound into cellular DNA as measured by a novel accelerator mass spectrometry assay.
- Sang, Soo Hah,Kim, Hyung M.,Sumbad, Rhoda A.,Henderson, Paul T.
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- Time and concentration dependence of fenton-induced oxidation of dG
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The influence of incubation time and Fenton reagent concentrations was investigated on the oxidation of 2′-deoxyguanosine. The compounds identified and quantified, through use of an LC-MS/MS system, were 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) and 8,5′-cyclo- 2′-deoxyguanosine (8,5′cyclodG) and the secondary oxidation products guanidinohydantoin and dehydro-guanidinohydantoin. 8-oxodG and 8,5′cyclodG formed very quickly, reaching a maximum rapidly, but with 8-oxodG a rapid decline occurred thereafter due to its further oxidation into the secondary products, which formed more slowly. Due to the better stability, 8,5′cyclodG correlated better with the general level of oxidation than 8-oxodG. The results emphasize the advantages of measuring other oxidation adducts than 8-oxodG alone. Copyright Taylor & Francis Group, LLC.
- Grey, Carl,Adlercreutz, Patrick
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- Susceptibility of channel catfish (Ictalurus punctatus) and brown bullhead (Ameriurus nebulosus) to oxidative stress: a comparative study
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Many pollutants in aquatic systems have been shown to exert toxic effects related to oxidative stress. Biochemical parameters of xenobiotic metabolism and oxidative stress were examined in two benthic fish species, channel catfish and brown bullhead, which differ in their apparent susceptibility to pollutant-mediated neoplasia. These parameters included enzyme activities and production of reactive oxygen species by hepatic subcellular fractions. Hepatic ethoxyresorufin O-deethylase (EROD) activity was markedly higher in catfish than in bullhead. This interspecific difference was also observed following induction of EROD activities by β-naphthoflavone (βNF). Conversely, hepatic microsomal cytochrome P-450 reductase (P-450R) activity was higher in bullhead and was unaffected in both species by βNF treatment. Hepatic catalase, DT diaphorase, glutathione reductase, glutathione S-transferase and total glutathione content were significantly higher in catfish as well. Hepatic glutathione peroxidase activity was significantly higher in bullhead. No significant interspecific difference was observed in hepatic superoxide dismutase activity. Cytosolic rates of menadione-mediated superoxide anion and hydrogen peroxide production were higher in pooled catfish liver samples as compared to bullhead. Conversely, microsomal rates of superoxide and hydrogen peroxide production were higher in bullhead, consistent with relative P-450R activities in the two species. Catfish liver homogenate was more effective than bullhead homogenate at inhibiting iron/ascorbate-mediated production of 8-hydroxydeoxyguanosine (8-OHdG) in vitro. - Keywords: Oxidative stress; Channel catfish; Brown bullhead; Menadione; Antioxidant
- Hasspieler, Bruce M.,Behar, Jeffrey V.,Carlson, David B.,Watson, David E.,Giulio, Richard T. Di
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- Hydroxylation of Deoxyguanosine at the C-8 Position in the Thiosulfate-Hydrogen Peroxide Reaction System. Evidence of Hydroxyl Radical Generation in the System
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The thiosulfate-hydrogen peroxide reaction system generates thiosulfate radical anion and hydroxyl radical.When 2'-deoxyguanosine (dG) was included in the system, it was hydroxylated at the C-8 position, and the hydroxylation was markedly enhanced by addition of a trace amount of iron ion.
- Seto, Hiroshi,Koike, Hiroshi,Sasano, Hideo
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- Measurement of oxidative DNA damage by catechol estrogens and analogues in vitro
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The growth-promoting effects of estrogens in hormone-dependent tumor tissues involve receptor-mediated pathways that are well-recognized; however, the role of estrogens in tumor initiation remains controversial. Estrogen metabolites, primarily the catechol estrogens (CE's), have been implicated in tumor initiation via a redox cycling mechanism. We have developed metabolically stable CE analogues for the study of receptor versus redox cycling effects on DNA damage. Comparisons between hydroxy estradiols (HE2's), methoxy estradiols (ME2's), and hydroxymethyl estradiols (HME2) in potentiometric and DNA damaging studies were made. DNA damage was assessed in calf thymus DNA using 8-oxo-2'-deoxyguanosine (8-oxo-dG) as a genotoxic marker for oxidative stress. Increases in the number of 8-oxo-dG/105 dG were significant for each 2-HE2 and 4-HE2. Cu(II)SO4, a transition metal known to catalyze the redox cycling of o-quinones, substantially increased the amount of DNA damage caused by both CE's. However, DNA damage was only observed at concentrations of 10 μM or higher, much greater than what is found under physiologic conditions. Furthermore, the presence of endogenous antioxidants such as glutathione, SOD, and catalase drastically reduced the amount of DNA damage induced by high concentrations of 2-HE2. There was no DNA damage observed for the non-redox cycling HME2's, making these compounds useful probes in the study of receptor-mediated carcinogenesis. Thus, both 2- HE2 and 4-HE2 are capable of producing oxidative DNA damage at micromolar concentrations in vitro. However, since the amount of CE's has not been shown to surpass nanomolar levels in vivo, it is unlikely that free radical production via redox cycling of CE's is a causative factor in human tumorigenesis.
- Mobley, James A.,Bhat, Abhijit S.,Brueggemeier, Robert W.
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- Optimization of the workup procedure for the analysis of 8-oxo-7,8-dihydro-2′-deoxyguanosine with electrochemical detection
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The artifactual generation of the biomarker for oxidative stress, 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG), during the workup procedure for its analysis is a difficult problem to solve, and the responsible factors are unclear. Here, peroxide removal and other antioxidant procedures during workup were compared using a limited amount of rat liver (50 mg) as starting material, with subsequent hydrolysis of 50 μg of DNA. A cold (0 °C) high salt GTC (4 M guanidine thiocyanate) nonphenol DNA extraction method was developed where DNA is quickly isolated. GSH (reduced glutathione) generated artifactual formation of 8-oxodG during the workup procedure, whereas H2O2 removal using catalase, Fe3+ removal and passivation using desferal, peroxide removal using glutathione peroxidase, ebselen and a peroxidase mimic lowered the 8-oxodG levels, all identifying peroxides as the responsible oxidants. Desferal was more protective when excluding Mg2+ and Ca2+ from buffers but was found to disturb the electrochemical detector when repeatedly injected five to six times, even at 100 μM. Addition of the OH-scavenger ethanol in all steps at 2% v/v had no protective effect. Zn2+ was found necessary for efficient DNA hydrolysis using nuclease P1, which was poor below 37 °C. Use of water substitutes was tested but inhibited DNA hydrolysis completely. H218O could, however, work for mass spectrometry methods. Long-term (38 days) storage of 0.5% v/v Triton X-100 generated more 8-oxodG than Tween 20 when incubated with free dG. The cold GTC DNA extraction method was used for analysis of freshly isolated human lymphocytes/monocytes from 60 healthy men using catalase and TEMPO as antioxidants, giving a background level of 0.074 ± 0.027 8-oxodG/105 dG (or 16 8-oxodG/108 nucleotides or 1943 8-oxodG/nuclei) which is probably the lowest value obtained yet. No increase with age was seen. Oxidation of dG to 8-oxodG during workup was found to fit a mathematically defined curve, and a calculated background level of 0.047 8-oxodG/105 dG was obtained. To obtain more reliable results it is recommended that control samples are included during the workup procedure, having an equal amount of cells (or DNA) as the exposed samples.
- Hofer, Tim,Moeller, Lennart
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- Dietary Antioxidants Fail in Protection against Oxidative Genetic Damage in In Vitro Evaluation
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Carcinogenesis is believed to be induced through the oxidative damage of DNA, and antioxidants are expected to suppress it. So, the polyphenolic antioxidants in daily foods were investigated to see whether they protect against genetic damage by active oxygen. In the evaluation, we used a bioassay and a chemical determination, a Salmonella mutagenicity test for mutation by a N-hydroxyl radical from one of the dietary carcinogens 3-amino-1-methyl-5H-pyrido[4,3-b]indole and the formation of 8-hydroxyl (8-OHdG) from 2′-deoxyguanosine (2′-dG) in a Fenton OH-radical generating system. Thirty-one antioxidants including flavonoids were compared in terms of radical-trapping activity with bacterial DNA and 2′-dG. Antioxidants inhibited the mutation but the IC50 values were in the mM order. Against 8-OHdG formation, only α-tocopherol had a suppressive effect with an IC50 of 1.5 μM. Thus, except α-tocopherol, the dietary antioxidants did not scavenge the biological radicals faster than bacterial DNA and intact 2′-dG, indicating that they failed to prevent oxidative gene damage and probably carcinogenesis.
- Sun, Mingzhou,Sakakibara, Hiroyuki,Ashida, Hitoshi,Danno, Gen-Ichi,Kanazawa, Kazuki
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- 8-Hydroxy-2′-deoxyguanosine formation and DNA damage induced by a dinuclear manganese(IV) complex and hydrogen peroxide
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The dinuclear manganese(IV) complex [Mn2O3L2][PF6]2 exhibits high activity for 8-hydroxy-2′-deoxyguanosine formation and relaxation of pBR322 form I DNA in the presence of H2O2, where L = 1,4,7-trimethyl-1,4,7-triazacyclononane; the active species is postulated to be a unidentate peroxide adduct of the manganese(IV) complex.
- Kobayashi, Teruyuki,Tsuchiya, Kazuyoshi,Nishida, Yuzo
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- Antioxidant Properties of Novel Tetraoxygenated Phenylindan Isomers Formed during Thermal Decomposition of Caffeic Acid
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Mild pyrolysis (228 °C, 15 min) of rosmarinic, chlorogenic, and caffeic acids increased their antioxidative efficacy in a biological rat liver membrane assay by 4-, 11-, and 460-fold, respectively. The active components in the caffeic acid pyrolysates were identified as the recently isolated novel tetraoxygenated 1,3-cis- and 1,3-trans-phenylindan isomers, which showed comparable IC50 values (0.041 and 0.04 μM, respectively) and were ?8-fold more active than butylated hydroxytoluene (BHT). Comparison of nonroasted, light-roasted, and dark-roasted coffee extracts showed that the degree of roasting is positively correlated to the inhibition of lipid peroxidation in rat liver membranes. The potent reducing properties of the phenylindan isomers resulted in (a) prooxidative effects at relatively higher concentrations in an ethyl linoleate peroxidation assay, and (b) promotion of hydroxylation of 2′-deoxyguanosine to afford 8-oxo-2′-deoxyguanosine. However, the results of the rat liver homogenate model system show that pyrolysis of caffeic acid and its esters chlorogenic acid and rosmarinic acid can procure potent antioxidants and underlines the potential use of heat processing to generate novel bioactive molecules.
- Guillot, Francois L.,Malnoe, Armand,Stadler, Richard H.
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- Spiroiminodihydantoin nucleoside formation from 2′-deoxyguanosine oxidation by [18O-labeled] singlet molecular oxygen in aqueous solution
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The main singlet molecular oxygen (1O2) oxidation products of free 2′-deoxyguanosine (dGuo) in aqueous solution were identified as a pair of diastereomeric spiroiminodihydantoin 2′-deoxyribonucleosides (dSp) together with 8-oxo-7,8-dihydro-2′- deoxyguanosine (8-oxodGuo). In the present work, evidence is provided from 18[1O2] and H218O labeling experiments, using HPLC-ESI-MS/MS, that the formation of dSp is explained by the addition of water to a reactive quinonoid intermediate, and a second reaction pathway leading to dSp involves 1O2 oxidation of initially generated 8-oxodGuo. Copyright
- Martinez, Glaucia Regina,Ravanat, Jean-Luc,Cadet, Jean,De Medeiros, Marisa Helena Gennari,Di Mascio, Paolo
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- High-intensity UV laser photolysis of DNA and purine 2'- deoxyribonucleosides: Formation of 8-oxopurine damage and oligonucleotide strand cleavage as revealed by HPLC and Gel electrophoresis studies
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Emphasis was placed in this work on the measurement of purine oxidation products generated upon nano- and picosecond UV laser biphotonic photolysis of 2'-deoxyadenosine, 2'-deoxyguanosine, calf thymus DNA, and a synthetic duplex oligonucleotide (37-mer) in aerated aqueous solutions. The overall formation of 8-oxo-7,8-d hydro- 2'-deoxyguanosine and 8-oxo-7,8-dihydro-2' - deoxyadenosme was determined using a HPLC-electrochemical assay. Denaturing gel electrophoresis analysis in association with a formamidopyrimidineNA glycosylase treatment was applied to reveal the sites recognized by this DNA repair enzyme. Both 8-oxo-7,8-dihydro-2'-deoxyguanosine and 8-oxo-7,8 dihydro-2'-deoxyadenosine were shown to be minor decomposition products of the related nucleoside purine radical cations in agreement with earlier observations. Interestingly, a dramatic increase in the yield of both photoproducts, this applying particularly to 8-oxo-7,8-dihydro-2'- deoxyguanosine, was observe in DNA. It should be noted that the yield of 8- oxo-7,8-dihydro-2'-deoxyguanosine was about 3-fold lower in heat-denatured DNA than in double-stranded DNA. These observations provide strong support to the significant involvement of base stacking and probably DNA solvatation in the chemical reactions of the purine radical cations. Other interesting information dealt with the similarity in the level of 8-oxo-7,8-dihydro-2'- deoxyguanosine and the number of formanidopyrimidine- DNA glycosytase sensitive guanine lesions. This strongly suggests that the latter formidopyrimidine-DNA glycosyiase pine nucleoside is the major DNA photodamaged product recognized by the DNA repair glycosylase. Another striking feature is the almost 10-fold decrease in the saturation dose E(s) for the two-quantum ionization of the guanine base in double-stranded DNA as compared to that observed for free 2'-deoxyguanosine. This can be explained by either an enhancement of the quantum yield of photoionization from the intermediate excited state in DNA (φ2) and/or hole migration with preferential trapping by guanine residues.
- Angelov,Spassky,Berger,Cadet
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- Synthesis of 8-oxo-dGTP and its β,γ-CH2-, β,γ-CHF-, and β,γ-CF2- analogues
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Three novel bisphosphonate analogues of 8-oxo-dGTP 3 in which the bridging β,γ-oxygen is replaced by a methylene, fluoromethylene or difluoromethylene group (4–6, respectively) have been synthesized from 8-oxo-dGMP 2 by reaction of its morpholine 5′-phosphoramidate 14 or preferably, its N-methylimidazole 5′-phosphoramidate 15 with tri-n-butylammonium salts of the appropriate bisphosphonic acids, 11–13. The latter method also provides a convenient new route to 3. Analogues 4–6 may be useful as mechanistic probes for the role of 3 in abnormal DNA replication and repair.
- Zheng, Yiying,Haratipour, Pouya,Kashemirov, Boris A.,McKenna, Charles E.
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- Sources of 2,5-diaminoimidazolone lesions in DNA damage initiated by hydroxyl radical attack
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The present study reports radiation-chemical yields of 2.5-diaminoimidazolone (Iz) derivatives in X-irradiated phosphate-buffered solutions of guanosine and double-stranded DNA. Various gassing conditions (air, N20/O2 (4:1), N2O, vacuum) were employed to elucidate the contribution of several alternative pathways leading to Iz in reactions initiated by hydroxyl radical attack on guanine. In all systems, Iz was identified as the second by abundance guanine degradation product after 8-oxoguanine, formed in 1:5 (guanosine) and 1:3.3 (DNA) ratio to the latter in air-saturated solutions. Experimental data strongly suggest that the addition of molecular oxygen to the neutral guanine radical G(-H)? plays a major in Iz production in oxygenated solutions of double-stranded DNA while in other systems it may compete with recombination of G(-H)? with superoxide and/or alkyl peroxyl radicals. The production of Iz through hydroxyl radical attack on 8-oxoguanine was also shown to take place although the chemical yield of Iz (ca 6%) in this process is too low to compete with the other pathways. The linearity of Iz accumulation with dose also indicates a negligible contribution of this channel to its yield in all systems.
- Thomas, Caroline Suzanne,Pollard, Hannah Catherine,Razskazovskiy, Yuriy,Roginskaya, Marina
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p. 517 - 524
(2020/09/07)
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- Isolation of new photoadducts from UVA-irradiated N-nitrosoproline with 2'-deoxyadenosine and characterization of photoadducts from DNA irradiated with N-nitrosoproline
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N-nitrosoproline (NPRO) is formed from nitrosation of proline and has been reported to be non-carcinogenic and non-mutagenic. However, earlier studies in our laboratory showed that pre-irradiated NPRO can be converted to a mutagenic form. We previously investigated the reaction of NPRO with dA or dG under UVA irradiation and identified the formation of 2-pyrrolidyl-dA adducts (P1 & P2) and 8-pyrrolidyl-dG adducts (G1 & G2) as well as four known modified nucleosides, although several peaks found in the HPLC profiles of UVA-irradiated mixtures of dA and NPRO remain unidentified. In the present study we isolated new photoproducts from irradiated mixtures of dA and NPRO and identified (R)- and (S)-8-(2-pyrrolidyl)-2′-deoxyadenosine (A1 and A2) as products by MS and NMR. We also investigated the photoadducts formed in DNA treated with NPRO under UVA irradiation, and detected A1 and/or A2 (probably both), P1, P2, G1 and/or G2, and 8-oxodG as products. Under anaerobic conditions, formation of A1 and A2 was greater than that under aerobic conditions, suggesting that photo-reactions comprising pyrrolidyl radical with dA may increase under anaerobic conditions given reduced competition with oxidative photo-reactions which may decompose pyrrolidyl-dA adducts.
- Aoyama, Shuhei,Arimoto-Kobayashi, Sakae,Asahi, Chiharu,Hatano, Tsutomu,Kimura, Sachiko,Suzuki, Toshinori
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- 5-Carboxamido-5-formamido-2-iminohydantoin, in Addition to 8-oxo-7,8-Dihydroguanine, Is the Major Product of the Iron-Fenton or X-ray Radiation-Induced Oxidation of Guanine under Aerobic Reducing Conditions in Nucleoside and DNA Contexts
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Exogenously and endogenously produced reactive oxygen species attack the base and sugar moieties of DNA showing a preference for reaction at 2′-deoxyguanosine (dG) sites. In the present work, dG was oxidized by HO? via the Fe(II)-Fenton reaction or by X-ray radiolysis of water. The oxidized lesions observed include the 2′-deoxynucleosides of 8-oxo-7,8-dihydroguanine (dOG), spiroiminodihydantoin (dSp), 5-guanidinohydantoin (dGh), oxazolone (dZ), 5-carboxamido-5-formamido-2-iminohydantoin (d2Ih), 5′,8-cyclo-2′-deoxyguanosine (cyclo-dG), and the free base guanine (Gua). Reactions conducted with ascorbate or N-acetylcysteine as a reductant under aerobic conditions identified d2Ih as the major lesion formed. Studies were conducted to identify the role of O2 and the reductant in product formation. From these studies, mechanisms are proposed to support d2Ih as a major oxidation product detected under aerobic conditions in the presence of the reductant. These nucleoside observations were then validated in oxidations of oligodeoxynucleotide and λ-DNA contexts that demonstrated high yields of d2Ih in tandem with dOG, dSp, and dGh. These results identify dG oxidation to d2Ih to occur in high yields leading to a hypothesis that d2Ih could be found from in cells stressed with HO?. Further, the distorted ring structure of d2Ih likely causes this lesion to be highly mutagenic. (Chemical Equation Presented)
- Alshykhly, Omar R.,Fleming, Aaron M.,Burrows, Cynthia J.
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p. 6996 - 7007
(2015/07/27)
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- Mapping structurally defined guanine oxidation products along DNA duplexes: Influence of local sequence context and endogenous cytosine methylation
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DNA oxidation by reactive oxygen species is nonrandom, potentially leading to accumulation of nucleobase damage and mutations at specific sites within the genome. We now present the first quantitative data for sequence-dependent formation of structurally defined oxidative nucleobase adducts along p53 gene-derived DNA duplexes using a novel isotope labeling-based approach. Our results reveal that local nucleobase sequence context differentially alters the yields of 2,2,4-triamino-2H-oxal-5-one (Z) and 8-oxo-7,8-dihydro-2′- deoxyguanosine (OG) in double stranded DNA. While both lesions are overproduced within endogenously methylated MeCG dinucleotides and at 5′ Gs in runs of several guanines, the formation of Z (but not OG) is strongly preferred at solvent-exposed guanine nucleobases at duplex ends. Targeted oxidation of MeCG sequences may be caused by a lowered ionization potential of guanine bases paired with MeC and the preferential intercalation of riboflavin photosensitizer adjacent to MeC:G base pairs. Importantly, some of the most frequently oxidized positions coincide with the known p53 lung cancer mutational hotspots at codons 245 (GGC), 248 (CGG), and 158 (CGC) respectively, supporting a possible role of oxidative degradation of DNA in the initiation of lung cancer.
- Ming, Xun,Matter, Brock,Song, Matthew,Veliath, Elizabeth,Shanley, Ryan,Jones, Roger,Tretyakova, Natalia
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p. 4223 - 4235
(2014/04/03)
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- Near ultraviolet radiation-mediated reaction between N-nitrosoproline and DNA: Isolation and identification of two new adducts, (R)- and (S)-8-(2-pyrrolidyl)-2′-deoxyguanosine
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To explore possible genotoxicity of N-nitrosoproline (NPRO), we investigated near-ultraviolet radiation (UVA)-mediated chemical reaction of NPRO with 2′-deoxyguanosine (dG). An acidic solution containing NPRO and dG was irradiated with UVA and products we
- Aoyama, Shuhei,Sano, Kayoko,Asahi, Chiharu,Machida, Masaki,Okamoto, Keinosuke,Negishi, Tomoe,Kimura, Sachiko,Suzuki, Toshinori,Hatano, Tsutomu,Arimoto-Kobayashi, Sakae
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- An ammonium bicarbonate-enhanced stable isotope dilution UHPLC-MS/MS method for sensitive and accurate quantification of acrolein-DNA adducts in human leukocytes
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Acrolein (Acr), a ubiquitous environmental pollutant, can react directly with genomic DNA to form mutagenic adducts without undergoing metabolic activation. To sensitively and accurately quantify Acr-DNA adducts (including structural isomers and stereoiso
- Yin, Ruichuan,Liu, Shengquan,Zhao, Chao,Lu, Meiling,Tang, Moon-Shong,Wang, Hailin
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p. 3190 - 3197
(2013/06/04)
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- Synthesis of N 2-Alkyl-8-oxo-7,8-dihydro-2′-deoxyguanosine derivatives and effects of these modifications on RNA duplex stability
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N2-Alkyl analogues of 8-oxo-7,8-dihydro-2′-deoxyguanosine (OG) were synthesized (alkyl = propyl, benzyl) via reductive amination of the protected OG nucleoside and incorporated into various positions of an RNA strand. Thermal stability studies of duplexes containing A or C opposite a single modified base revealed only moderate destabilization. Both OG as well as its N2-alkyl analogues can pair opposite A or C with nearly equal stability, potentially offering a new means of modulating RNA-protein interactions in the minor vs major grooves.
- Kannan, Arunkumar,Burrows, Cynthia J.
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p. 720 - 723
(2011/03/20)
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- Radiation-induced formation of purine 5′,8-cyclonucleosides in isolated and cellular DNA: High stereospecificity and modulating effect of oxygen
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The present work is aimed at gaining conclusive mechanistic insights into the radiation-induced formation of the 5′R and 5′S diastereomers of both adenine and guanine 5′,8-cyclo-2′-deoxyribonucleosides, with emphasis on the delineation of the inhibitory effect of O2 in isolated and cellular DNA. The levels of purine 5′,8-cyclo-2′- deoxyribonucleosides as assessed by HPLC-MS/MS were found to decrease steadily with the increase of O2 concentration, the 5′,8-cyclo-2′- deoxyguanosine being produced more efficiently than the 5′,8-cyclo- 2′-deoxyadenosine for low O2 concentrations. A high stereoselectivity was observed in the intramolecular addition of the C5′ radical to the C8 of the purine leading, after the creation of the C5′-C8 bond and a subsequent oxidation step, to the predominant formation of the 5′R diastereomer for both purine 5′,8-cyclonucleosides. The reduced formation yield of the 4 tandem lesions in the presence of O2 explains, at least partly, the low efficiency of radiation-induced yields of the purine 5′,8-cyclo-2′-deoxyribonucleosides in cellular DNA, which are about two orders of magnitude lower than the previously reported data obtained from HPLC-MS analysis. The Royal Society of Chemistry 2010.
- Belmadoui, Nourreddine,Boussicault, Fabien,Guerra, Maurizio,Ravanat, Jean-Luc,Chatgilialoglu, Chryssostomos,Cadet, Jean
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experimental part
p. 3211 - 3219
(2010/08/21)
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- Fe(II) phthalocyanine catalyzed oxidation of dGMP by molecular oxygen
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We show that iron(II)-phthalocyanines are able to catalyze guanosine oxidation by molecular oxygen in the presence of reducing agents such as ascorbic acid and 2-mercaptoethanol. The products of 5′-monophosphate-2′-deoxyguanosine (dGMP) oxidation were dir
- Kuznetsova, Aleksandra A.,Solovyeva, Lyudmila I.,Kaliya, Oleg L.,Lukyanets, Evgeniy A.,Knorre, Dmitri G.,Fedorova, Olga S.
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experimental part
p. 4335 - 4338
(2010/04/24)
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- Identification of the α and β anomers of 1-(2-deoxy-D-erythro- pentofuranosyl)-oxaluric acid at the site of riboflavin-mediated photooxidation of guanine in 2′-deoxyguanosine and thymidylyl-3′-5′- deoxyguanosine
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Products of riboflavin-mediated photosensitization of 2′- deoxyguanosine (dG) and thymidylyl-(3′-5′)-2′-deoxyguanosine (TpdG) by 350-nm light in oxygen-saturated aqueous solution have been isolated and identified as 1-(2-deoxy-β-erythro-pentofuranosyl) oxaluric acid (β-dOx) and thymidylyl-(3′-5′)-1-(2-deoxy-β-D-erythro- pentofuranosyl) oxaluric acid (Tpβ-dOx), respectively. In aqueous solution the modified β-deoxyribonucleoside is slowly converted to the α-anomer, generating α-dOx and Tpα-dOx. These modified nucleosides and dinucleoside monophosphates have been isolated by HPLC and characterized by proton and carbon NMR spectroscopy, fast atom bombardment mass spectrometry, and enzymatic analyses. Both α-dOx and Tpα-dOx slowly convert back into the modified β-deoxyribonucleoside, indicating that the furanosidic anomers are in dynamic equilibrium. Relative to TpdG, the rate of hydrolysis of Tpβ-dOx and Tpα-dOx by spleen phosphodiesterase is greatly reduced. Hot piperidine (1.0 M, 90°C, 30 min) destroys Tpβ-dOx and Tpα-dOx. Riboflavin-mediated photosensitization of TpdG in D 2O instead of H2O has no detectable effect on the yield of Tpβ-dOx, suggesting that oxaluric acid is generated through a Type-I reaction mechanism, likely through the intermediary on initially generated 8-oxo-7,8-dihydro-2′-deoxyguanosine.
- Buchko, Garry W.,Cadet, Jean
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p. 191 - 199
(2008/02/05)
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- Singlet oxygen oxidation of 2′-deoxyguanosine. Formation and mechanistic insights
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Emphasis was placed in this work on the delineation of mechanistic aspects of the singlet oxygen-mediated oxidation reactions of 2′-deoxyguanosine 1 used as a DNA model compound in aerated aqueous solution. For this purpose a thermolabile naphthalene endoperoxide derivative was used allowing the generation of [18O]-labeled singlet oxygen for dedicated mechanistic studies. The analysis and characterization of the oxidized nucleosides of the 1O2 reactions were achieved on the basis of accurate HPLC-tandem mass spectrometry measurements. Thus it was found that primary oxidation products include, in addition to the previously identified 8-oxo-7,8-dihydro-2′-deoxyguanosine 5 and the two diastereomers of spiroiminodihydantoin 8, two relatively minor nucleosides, namely the two diastereomers of 4-hydroxy-8-oxo-4,8-dihydro-2′-deoxyguanosine 9.
- Ravanat, Jean-Luc,Martinez, Glaucia R.,Medeiros, Marisa H.G.,Di Mascio, Paolo,Cadet, Jean
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p. 10709 - 10715
(2007/10/03)
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- Formation of a diimino-imidazole nucleoside from 2′-deoxyguanosine by singlet oxygen generated by methylene blue photooxidation
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Singlet oxygen (1O2) is capable of inducing genotoxic, carcinogenic and mutagenic effects. It has previously been reported that the reaction of 1O2 with 2′-deoxyguanosine, which is a major target of 1O2 among the DNA constituents, leads to formation of various oxidized products including 8-oxo-7,8-dihydro-2′-deoxyguanosine and spiroiminodihydantoin, amino-imidazolone and diamino-oxazolone nucleosides. In addition to these products, we report that a novel diimino-imidazole nucleoside, 2,5-diimino-4-[(2-deoxy-β-D-erythro-pentofuranosyl)amino]-2H,5H- imidazole (dD), is formed by reaction of 2′-deoxyguanosine with 1O2 generated by irradiation with visible light in the presence of methylene blue under aerobic conditions. Its identification is based on identical chromatographic and spectroscopic data with an authentic compound, which we recently isolated and characterised from the reaction mixture of 2′-deoxyguanosine with reagent HOCl and a myeloperoxidase-H2O2-Cl- system. The yield of dD was increased by D2O and decreased by azide. dD was not generated from 8-oxo-7,8-dihydro-2′-deoxyguanosine. These results indicate that dD is generated by 1O2 directly from 2′-deoxyguanosine, but not via 8-oxo-7,8-dihydro-2′-deoxyguanosine. dD may play a role in the genotoxicity of singlet oxygen in cells.
- Suzuki, Toshinori,Friesen, Marlin D.,Ohshima, Hiroshi
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p. 2157 - 2162
(2007/10/03)
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- Spiroiminodihydantoin is a major product in the photooxidation of 2′-deoxyguanosine by the triplet states and oxyl radicals generated from hydroxyacetophenone photolysis and dioxetane thermolysis
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(formula presented) Photolysis of hydroxyacetophenone and thermolysis of the corresponding dioxetane afford spiroiminodihydantoin rather than 4,8-dihydro-4-hydroxy-8-oxo-2′-deoxyguanosine (4-HO-8-oxodG) through the oxidation of 2′-deoxyguanosine (dG) by triplet-excited hydroxyacetophenone and the peroxyl radicals derived thereof by α cleavage and subsequent oxygen trapping. The structure of the spiroiminodihydantoin is assigned by the SELINQUATE NMR technique, which unequivocally establishes the spirocyclic connectivity.
- Adam, Waldemar,Arnold, Markus A.,Gruene, Matthias,Nau, Werner M.,Pischel, Uwe,Saha-Moeller, Chantu R.
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p. 537 - 540
(2007/10/03)
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- Type II guanine oxidation photoinduced by the antibacterial fluoroquinolone rufloxacin in isolated DNA and in 2′-deoxyguanosine
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The role played by type I (radical) and type II (singlet oxygen) mechanisms in the Rufloxacin (RFX)-photoinduced production of 8-hydroxy-2′-deoxyguanosine in DNA has been evaluated. This fluoroquinolone drug has been shown to be able to photoinduce increased levels of some DNA base oxidation products, such as 8-OH-dGuo, that are indicative of mutagenic and carcinogenic events, with probable implications in aging processes. The relative weight of the two photosensitization mechanisms was obtained via determination of two different photoproducts of 2′-deoxyguanosine (dGuo), which are diagnostic of the two different pathways, namely, (4R*)- and (4S*)-4,8-dihydro-4-hydroxy-8-oxo-2′-deoxyguanosine and 2,2-diamino-4-[(2-deoxy-β-D-erythro-pentofuranosyl)amino]-2, 5-dihydrooxazol-5-one. The observed predominance of type II reaction is in agreement with the fact that the triplet state of RFX is able to transfer with high efficiency its energy to molecular oxygen, giving rise to singlet oxygen. Photophysical measurements suggest that hydrated electrons produced by Rufloxacin photoionization react with dGuo, Thd, and DNA, whereas these biomolecules quench the RFX triplet state with low efficiency. Static quenching of Rufloxacin fluorescence indicates an interaction of this drug both with DNA and with dGuo. On the basis of these experimental data, Rufloxacin photosensitization of DNA is proposed to occur by a type II mechanism.
- Belvedere, Alessandra,Bosca, Francisco,Catalfo, Alfio,Cuquerella, Maria C.,De Guidi, Guido,Miranda, Miguel A.
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p. 1142 - 1149
(2007/10/03)
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- Low-energy, low-yield photoionization, and production of 8-oxo-2a?2-deoxyguanosine and guanine from 2a?2-deoxyguanosine
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Experiments employing electron scavenging methods and high performance liquid chromatography with mass spectrometry detection indicate that electrons, formed via one-photon ionization, guanine (G) and small amounts of 8-oxo-2a?2-deoxyguanosine (8-oxo-dG)
- Papadantonakis, George A.,Tranter, Robert,Brezinsky, Kenneth,Yang, Yanan,Van Breemen, Richard B.,LeBreton, Pierre R.
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p. 7704 - 7712
(2007/10/03)
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- Analysis of peroxynitrite reactions with guanine, xanthine, and adenine nucleosides by high-pressure liquid chromatography with electrochemical detection: C8-nitration and -oxidation
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Peroxynitrite, the reaction product of nitric oxide and superoxide anion, and a powerful oxidant, was found to nitrate as well as oxidize adenine, guanine, and xanthine nucleosides. A highly sensitive reverse-phase HPLC method with a dual-mode electrochemical detector, which reduces the nitro product at the first electrode and detects the reduced product by oxidation at the second electrode, was applied to detect femtomole levels of 8-nitroguanine and 8-nitroxanthine. This method was used to separate and identify the products of nitration and oxidation from the reactions of nucleosides with peroxynitrite. Peroxynitrite nitrates deoxyguanosine at neutral pH to give the very unstable 8-nitrodeoxyguanosine, in addition to 8-nitroguanine. 8-Nitrodeoxyguanosine, with a half-life of ～10 min at room temperature and ≤3 min at 37 °C, hydrolyzes at pH 7 to 8-nitroguanine. A decrease in the reaction pH resulted in a decrease in the level of C8-nitration. Peroxynitrite also oxidizes deoxyguanosine in a pH-dependent manner, to give 8-oxodeoxyguanosine with a maximum yield (0.5-0.7%) at pH 5. Guanosine and xanthosine exhibit reactivity similar to that of deoxyguanosine toward peroxynitrite at neutral pH, producing only the corresponding 8-nitronucleosides as well as 8-nitroguanine and 8-nitroxanthine, respectively. 8-Nitroguanosine at pH 7, with a half-life of several weeks at 5 °C and 5 h at 37 °C, was much more stable than 8-nitrodeoxyguanosine. C8-nitration was confirmed by dithionite reduction to the corresponding amino nucleosides, which cochromatographed with synthesized 8-amino nucleoside standards. In contrast to guanine nucleosides, adenine nucleosides undergo peroxynitrite-mediated C8 oxidation even at neutral pH to give the corresponding 8-oxoadenine nucleosides in ～0.3% yield. Adenine nitration, though minor compared to C8-oxidation, appears to occur at both C2 and C8 positions of the adenine ring. Lowering the reaction pH from 7 to 5 results in 2.4- and 2.2-fold increases in the yields of 8-oxo-dA and 8-oxo-Ado, respectively, but the level of nitration is not altered.
- Sodum, Rama S.,Fiala, Emerich S.
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p. 438 - 450
(2007/10/03)
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- Mechanistic and kinetic aspects of photosensitization in the presence of oxygen
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Determining whether the first step of photooxygenation is Type I or Type II is a necessary prerequisite in order to establish the mechanism of photodynamic action. But this distinction is not sufficient, because other processes, both consecutive and competitive, commonly participate in the overall mechanism. Thus, in both Type I and Type II reactions, the initial products are often peroxides that can break down and induce free radical reactions. These aspects of photosensitization are discussed and illustrated by sensitizer/substrate systems involving (1) only radical reactions (decatungstate/alkane) and (2) reactions of singlet oxygen occurring in competitive and consecutive processes and possibly followed by radical reactions (methylene blue/2′-deoxyguanosine). Two other previously investigated systems involving, respectively, a Type II interaction followed by radical processes (methylene blue/alkene) and Type II reactions, some of which being competitive or consecutive (rose bengal/alkene), are briefly reconsidered.
- Tanielian, Charles,Mechin, Robert,Seghrouchni, Rachid,Schweitzer, Claude
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- N-hydroxy-4-(4-chlorophenyl)thiazole-2(3H)-thione as a photochemical hydroxyl-radical source: Photochemistry and oxidative damage of DNA (strand breaks) and 2′-deoxyguanosine (8-oxodG formation)
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On irradiation of N-hydroxythiazole-2(3H)-thione 3 at 300 nm, the photoproducts disulfide 4, bisthiazole 5 and thiazole 6 are formed. During this photolysis, hydroxyl radicals are released, which have been detected by spin trapping with 5,5-dimethyl-1-pyrroline N-oxide (DMPO), coupled with electron paramagnetic resonance spectroscopy. In the presence of supercoiled pBR322 DNA, irradiation of thiazolethione 3 induces strand breaks through the photogenerated hydroxyl-radicals, as confirmed by control experiment with the hydroxyl-radical scavenger isopropanol. Singlet oxygen appears not to be involved, as attested by the lack of a D2O isotope effect. During the photoreaction of thiazolethione 3 in the presence of 2′-deoxyguanosine (dG), the latter is photooxidized (ca 10% conversion after 2 h of irradiation) to the 7,8-dihydro-8-oxo-2′-deoxyguanosine as the main oxidation product. The dG conversion levels off after complete consumption of thiazolethione 3 and is suppressed by the addition of the hydroxyl-radical scavenger 2,6-di-tert-butylcresol or DMPO. Since the photoproducts 4-6 are ineffective as sensitizers for the photooxidation of dG and DNA, the hydroxyl radicals released in the photolysis of thiazolethione 3 are the oxidizing species of DNA and dG. These results suggest that the thiazolethione 3 may serve as a novel and effective photochemical hydroxyl-radical source for photobiological studies.
- Adam, Waldemar,Hartung, Jens,Okamoto, Hideki,Saha-Moeller, Chantu R.,Spehar, Kristina
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p. 619 - 624
(2007/10/03)
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- Efficient synthesis of 8-oxo-dGTP: A mutagenic nucleotide
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An efficient synthesis of mutagenic and oxidative DNA damage product, 8-oxo-dGTP (4) has been achieved in high yield, along with a serendipitous generation of 8-dimsyl-dG (2). In combination with dPTP (5), 8-oxo-dGTP (4) can be formulated into a kit for investigating DNA random mutagenesis. (C) 2000 Elsevier Science Ltd. All rights reserved.
- Nampalli, Satyam,Kumar, Shiv
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p. 1677 - 1679
(2007/10/03)
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- Formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine under anaerobic conditions by reductively activated nitro 5-deazaflavin derivatives
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Electrolytically reduced 6- and 8-nitro-5-deazaflavin derivatives have been found to interact to react specifically with guanine base by means of cyclic voltammetry. Electrolytic reductions of 6- and 8-nitro-5-deazaflavin derivatives in the presence of the 2'-deoxyguanosine under anaerobic conditions resulted in prominent formation of 8-oxo-7,8-dihydro-2'- deoxyguanosine.
- Mikata, Yuji,Kishigami, Maki,Nishida, Mamiko,Yano, Shigenobu,Kawamoto, Tetsuji,Ikeuchi, Yoshihiro,Yoneda, Fumio
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p. 2141 - 2144
(2007/10/03)
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- Hydroxyl radical-induced degradation of 2′-deoxyguanosine under reducing conditions
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Addition of hydroxyl radical to the base moiety of 2′-deoxyguanosine (dGuo) leads to the formation of two main radicals exhibiting oxidising and reducing properties, respectively. The oxidising radical reacts with oxygen to yield 2,2-diamino-5-[2-deoxy-β-D-erythro-pentofuranosyl)amino]oxazol-5(2H)-one (oxazolone) as the final product. The reducing radical is either preferentially oxidised into 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodGuo) or reduced into a 2,6-diamino-4-hydroxy-5-formamidopyrimidine derivative (FapydGuo) depending on conditions. We report here that the presence of reducing compounds (ascorbate or cysteine) strongly modifies the distribution of modified nucleosides upon γ irradiation of an aerated aqueous solution of dGuo. The yield of oxazolone decreases while that of 8-oxodGuo and FapydGuo increases. This was explained by the reduction of the oxidising radical which prevents the occurrence of the restitution of dGuo through a reaction between the oxidising and the reducing purine radicals. The study was extended to the decomposition of dGuo upon photochemical release of .OH by N-hydroxypyrimidine-2-thione (HPT). The analysis of the base modification products of dGuo induced by the latter system showed that HPT exhibits reducing properties and cannot be used as a pure photochemical source of ?OH radical.
- Douki, Thierry,Spinelli, Sandrine,Ravanat, Jean-Luc,Cadet, Jean
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p. 1875 - 1880
(2007/10/03)
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- Oxidative DNA damage by radicals generated in the thermolysis of hydroxymethyl-substituted 1,2-dioxetanes through the α cleavage of chemiexcited ketones
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The 3-(hydroxymethyl)-3,4,4-trimethyl-1,2-dioxetane (HTMD) highly efficiently damages DNA compared to the merely alkyl-substituted derivative 3,3,4,4-tetramethyl-1,2-dioxetane (TMD). To elucidate this difference in oxidative reactivity, two additional hydroxymethyl-substituted 1,2- dioxetanes, namely cis/trans-3-(hydroxymethyl)-3,4-dimethyl-4-(phenylmethyl)- (lα/1β) and 3-(hydroxymethyl)-4,4-dimethyl-3(phenylmethyl)-1,2-dioxetane (2), were investigated in regard to their photochemical and photobiological properties. The high genotoxic effects of the hydroxymethyl-substituted 1,2- dioxetanes, which are reflected in the significant formation of single- strand breaks in plasmid pBR 322 DNA and the efficient oxidation of guanine in calf thymus DNA and the nucleoside 2'-deoxyguanosine (dGuo), are for the first time understood in terms of radical chemistry. The reactivity order of the dioxetanes 1α/1β > HTMD > 2 >> TMD to damage DNA parallels the propensity of these dioxetanes to generate radicals. These reactive species are formed in the thermolysis of the dioxetanes through α cleavage of the intermediary triplet-excited α-hydroxy- and α-phenyl-substituted carbonyl products. The presence of radicals was confirmed by spin-trapping experiments with 5,5-dimethyl-1-pyrroline N-oxide and by laser-flash photolysis. These carbon-centered radicals are efficiently scavenged by molecular oxygen to produce peroxyl radicals, which are proposed as the active DNA damaging species in the thermal decomposition of the hydroxymethyl-substituted 1,2- dioxetanes HTMD, 1α/1β, and 2.
- Adam, Waldemar,Andler, Simone,Nau, Werner M.,Saha-M?ller, Chantu R.
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p. 3549 - 3559
(2007/10/03)
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- The equine estrogen metabolite 4-hydroxyequilenin causes DNA single- strand breaks and oxidation of DNA bases in vitro
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Premarin (Wyeth-Ayerst) is the estrogen replacement treatment of choice and continues to be one of the most widely dispensed prescriptions in North America. In addition to endogenous estrogens, Premarin contains unsaturated equine estrogens, including equilenin [1,3,5(10),6,8-estrapentaen-3-ol-17- one]. In previous work, we showed that the equilenin metabolite 4- hydroxyequilenin (4-OHEN) can be autoxidized to 4-OHEN-o-quinone which readily entered into a redox couple with the semiquinone radical catalyzed by NAD(P)H, P450 reductase, or quinone reductase, resulting in generation of reactive oxygen species [Shen, L., Pisha, E., Huang, Z., Pezzuto, J. M., Krol, E., Alam, Z., van Breemen, R. B., and Bolton, J. L. (1997) Carcinogenesis 18, 1093-1101]. As oxidative damage to DNA by reactive oxygen species generated by redox active compounds has been proposed to lead to tumor formation, we investigated whether 4-OHEN could cause DNA damage. We treated λ phage DNA with 4-OHEN and found that extensive single-strand breaks could be obtained with increasing concentrations of 4-OHEN as well as increasing incubation times. If scavengers of reactive oxygen species are included in the incubations, DNA could be completely protected from 4-OHEN- mediated damage. In contrast, NADH and CuCl2 enhanced the ability of 4-OHEN to cause DNA single-strand breaks presumably due to redox cycling between 4- OHEN and the semiquinone radical generating hydrogen peroxide and ultimately copper peroxide complexes. We also confirmed that 4-OHEN could oxidize DNA bases since hydrolysis of 4-OHEN-treated calf thymus DNA and HPLC separation with electrospray MS detection revealed oxidized deoxynucleosides, including 8-oxodeoxyguanosine and 8-oxodeoxyadenosine. Our data suggest that DNA single-strand breaks and oxidation of DNA bases by 4-OHEN could contribute to the carcinogenic mechanism(s) of equine estrogens.
- Chen, Yumei,Shen, Li,Zhang, Fagen,Lau, Serrine S.,Van Breemen, Richard B.,Nikolic, Dejan,Bolton, Judy L.
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p. 1105 - 1111
(2007/10/03)
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- Type I and type II photosensitized oxidative modification of 2'-deoxyguanosine (dGuo) by triplet-exicted ketones generated thermally from the 1,2-dioxetane HTMD
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The nucleoside 2'-deoxyguanosine (dGuo) was treated with 3-(hydroxymethyl)-3,4,4-trimethyl-1,2-dioxetane (HTMD), the latter generates efficiently triplet-excited carbonyl products on thermal decomposition in the dark. The type I photooxidation products, 2,2-diamino-[(2-deoxy-β-D-erythro-pentofuranosyl)-4-amino]-5(2H)-oxaz olone (oxazolone) and the cyclic nucleoside 2-(S)-2,5'-anhydro-1-(2-deoxy-β-D-erythro-pentofuranosyl)-5-guanidiny lidene-2-hydroxy-4-oxoimidazolidine (oxoimidazolidine), as well as the type II photooxidation products 4-(R)*- and 4-(S)*-4-hydroxy-8-oxo-4,8-dihydro-2'-deoxyguanosine (4-HO-8-oxodGuo) and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo), were quantitatively determined by appropriate selective and sensitive HPLC assays. The concentration and time profiles revealed that about 40% of the triplet ketones derived from the thermal decomposition of HTMD led to photooxidation of dGuo. Essentially equal amounts of type I and type II photooxidation products were found, as could be established by comparison with predominant type I (benzophenone, riboflavin) and type II (Rose Bengal, methylene blue) photosensitizers. The participation of singlet oxygen (type II activity) was confirmed by the substantial D2O effect in the formation of 8-oxodGuo. The results demonstrate that dioxetanes, particularly HTMD, are efficient photooxidants of dGuo on thermal activation in the dark and constitute excellent chemical tools to study photobiological processes without the use of light, in the present case, photogenotoxicity.
- Adam, Waldemar,Saha-M?ller, Chantu R.,Sch?nberger, André
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p. 719 - 723
(2007/10/03)
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- Photochemistry and photobiology of furocoumarin hydroperoxides derived from imperatorin: Novel intercalating photo-fenton reagents for oxidative DNA modification by hydroxyl radicals
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Photochemical and photobiological properties of the imperatorin-derived furocoumarin hydroperoxides 1a, 1a′, 2a and 2a′ have been investigated. Irradiation (350 nm) of the hydroperoxide 2a′ afforded the alcohol 2b (2%), a diastereomeric mixture of the hydroxy epoxide 2c (40%; diastereomeric ratio = 80:20) and the epoxide 2d (8%). The formation of these products was rationalized in terms of homolysis of the hydroperoxide bond initiated by intramolecular energy transfer from the photoexcited furocoumarin chromophore. The quantum yields for the photolytic decomposition of hydroperoxides were estimated to be in the range of 0.03-0.85 and decreased in the order 2a ? 2a′ ? 1a′ ≥ 1a. The involvement of hydroxyl radicals in these reactions was established by trapping experiments with benzene and spectroscopic evidence was obtained by EPR spin trapping with 5,5-dimethylpyrroline-N-oxide. Fluorescence titration, DNA melting and linear dichroism studies of furocoumarins indicated that these compounds undergo efficient complexation and also intercalation into the DNA. The binding parameters K (intrinsic binding constant) and 1/n (frequency of binding sites) of complexes between furocoumarin derivatives and DNA were determined to be in the range of 3900-23900 M-1 and 0.017-0.045. The photoreaction of 1a′ and 1b′ with 2′-deoxyguanosine (dGuo) afforded exclusively 7,8-dihydro-8-oxo-2′-deoxy-guanosine (8-oxodGuo), presumably through singlet oxygen, which was formed in a type II photooxidation process. In contrast, the hydroperoxide 2a oxidized dGuo to oxazolone as major and 8-oxodGuo as minor products through hydroxyl radicals, which were generated from 2a under photolytic conditions. Interestingly, the photoreactions of furocoumarins with salmon testes DNA showed that the highly reactive (φ = 0.85) hydroperoxide 2a is also most efficient in inducing the mutagenic DNA oxidation product 8-oxodGuo. Hence, the novel furocoumarin hydroperoxide 2a constitutes the first intercalating photo-Fenton reagent and serves as convenient hydroxyl radical source for genotoxicity studies.
- Adam, Waldemar,Berger, Maurice,Cadet, Jean,Dall'Acqua, Francesco,Epe, Bernd,Frank, Silvia,Ramaiah, Danaboyina,Raoul, Sebastian,Saha-Moeller, Chantu R.,Vedaldi, Daniela
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p. 768 - 778
(2007/10/03)
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- Reaction of Singlet Oxygen with 2'-Deoxyguanosine and DNA. Isolation and Characterization of the Main Oxidation Products
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The reaction of singlet molecular oxygen with 2'-deoxyguanosine and DNA was studied. Emphasis was placed on the identification and characterization of the main methylene blue mediated type II (singlet oxygen) oxidation products of 2'-deoxyguanosine and its corresponding 3',5'-di-O-acetylated derivative. Two major oxidation products of 2'-deoxyguanosine were isolated and characterized by mass spectrometry analysis and extensive 1H and 13C NMR measurements as the two 4R* and 4S* diastereomers of 4,8-dihydro-4-hydroxy-8-oxo-2'-deoxyguanosine. The addition of 1O2 was also found to occur to the base moiety of the corresponding 3',5'-di-O-acetylated derivative. Methylene blue mediated photosensitization of 2'-deoxygunosine led also to the production of 7,8-dihydro-8-oxo-2'-deoxyguanosine, but in a relatively lower yield with respect to the two above diastereomers. The participation of singlet oxygen in the mechanism of formation of these oxidation products was confirmed. A reasonable mechanism involving the transient formation of an unstable endoperoxide produced through a Diels-Alder 1,4-cycloaddition of singlet oxygen to the purine ring is suggested. Quantitative analysis allowed us to demonstrate that the two diastereomers of 4,8-dihydro-4-hydroxy-8-oxo-2'-deoxyguanosine are the main singlet oxygen oxidation products of the guanine moiety within nucleosides, whereas 7,8-dihydro-8-oxoguanine was found to be the major 1O2 oxidation product of guanine in double-stranded DNA.
- Ravanat, Jean-Luc,Cadet, Jean
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p. 379 - 388
(2007/10/03)
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- Photosensibilisierte Bildung von 8-Hydroxy-2'-desoxyguanosin in Lachs-Testes-DNA durch Furocumarinhydroperoxide: ein neuartiges, intercalierendes "Photo-Fenton"-Reagens fuer oxidative DNA-Schaedigung
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Stichworte: DNA-Oxidation * Furocumarine * Hydroperoxide * Hydroxylradikale * 8-Hydroxy-2'-desoxyguanosin
- Adam, Waldemar,Cadet, Jean,Dall'Acqua, Francesco,Epe, Bernd,Ramaiah, Danaboyina,Saha-Moeller, Chantu R.
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- High Activity of an Fe-tfda (tfda = 2-aminomethyltetrahydrofuran-N,N-diacetic acid) Complex for Hydroxylation at the Aromatic and Alkane Rings of 2'-Deoxyguanosine in the Presence of Hydrogen Peroxide
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An iron(III) complex with tfda (2-aminomethyltetrahydrofuran-N,N-diacetic acid) exhibits high activity for hydroxylation at the 8 and 2' positions of deoxyguanosine in the presence of hydrogen peroxide; this demonstrates that a facile transformation from deoxyribonucleotide to ribonucleotide in vivo may be possible.
- Nishida, Yuzo,Ito, Sayo
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p. 1211 - 1212
(2007/10/02)
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- Significant formation of 8-hydroxydeoxyguanosine through interaction of diesel particulate matter with deoxyguanosine
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The role of diesel particulate matter (DPM) in generating oxidative DNA damage was investigated. Specifically, the effect of DPM on hydroxylation of guanine base at the C-8 position was investigated. We demonstrated significant formation of 8-hydroxydeoxyguanosine through interaction of DPM with 2-deoxyguanosine in aqueous solution. Notably, the most effective part of DPM is the residue obtained after organic solvent extraction, and the next most effective part is the polar material extracted with acetone. The hydroxylating activity of organic pollutants in DPM is discussed.
- Seto,Ohkubo,Koike,Saito,Sasano
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p. 789 - 795
(2007/10/03)
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- Photooxydation sensibilisee de la desoxy-2' guanosine par des phtalocyanines et naphtalocyanines. Determination de l'importance des mecanismes de type I et de type II
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The photodynamic properties of sulfonated derivatives of phthalocyanine and naphthalocyanine were investigated using 2'-deoxyguanosine as a model compound for DNA.The major degradation photoproducts of this nucleoside were characterized and classified into two categories.The degradation involving the radical mechanism (type I) led to the formation of 2,2-diamino-5-oxazolone.The two main photooxidation products involving type II mechanism were characterized as the 4R* and 4S* diastereoisomers of 9-(2-β-D-deoxyerythropentofuranosyl)-4,8-dihydro-4-hydroxy-8-oxoguanine.In addition, 8-oxo-7,8-dihydro-2'-deoxyguanosine was shown to be generated (probably through type II mechanism).By identifying and quantifying these photoproducts, the ratio of type I/type II photoprocess was established for three (aluminum, gallium and zinc) disulfonated phthalocyanines and a tetrasulfonated aluminum naphthalocyanine.It was found that the type II mechanism is predominant.
- Ravanat, J. L.,Berger, M.,Buchko, G. W.,Benard, J. F.,Lier, J. E. van,Cadet, J.
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p. 1069 - 1076
(2007/10/02)
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- Etude comparative de l'oxydation radicalaire de l'ADN et de ses nucleosides par les radicaux hydroxyles et les ions ferryles issus de la reaction de Fenton
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A comparative study of the reaction of hydroxyl radicals and Fenton type oxidative species with DNA and 2'-deoxyribonucleosides was investigated.This study was based on the characterization of the diamagnetic products resulting from the chemical transformation of the transient radicals.Emphasis was placed on the radical oxidative reactions of the purine nucleosides.It is interesting to note that oxidative purine radicals can be reduced by reagents such as ascorbic acid or N,N,N',N'-tetramethyl-1,4-p-phenylenediamine.The observed differences in the nature of the decomposition products resulting from the Fenton reaction are not consistent with the nature of the oxidative species (hydroxyl radicals or ferryl ions) involved, but due to the presence of ferrous sulfate.
- Mouret, J. F.,Berger, M.,Anselmino, C.,Polverelli, M.,Cadet, J.
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p. 1053 - 1060
(2007/10/02)
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- Phototherapy using methylene blue
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The present invention is a method for using thiazin dyes, especially methylene blue, in combination with light to hydroxylate guanosine or deoxyguanosine at the C8 of the purine ring. The number of guanosines in a nucleic acid strand converted to 8-OH-deoxyguanosine (8-OH-dG) or 8-OH-guanosine (8-OH-G) can be controlled through manipulation of the concentration of methylene blue, light intensity and length of exposure, pH, and buffer strength. The method can be used for the selective mutation or modification of either a DNA or a RNA sequence, or the protein expressed therefrom. The method can also be used in the treatment of viral infectons and in cancer. Methylene blue is FDA approved for topical, i.v., and oral administration. Viruses, bacteria, and cells undergoing rapid DNA synthesis are all inactivated by methylene blue in the presence of light or when irradiated.
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- 8-Substituted guanosine and 2'-deoxyguanosine derivatives as potential inducers of the differentiation of Friend erythroleukemia cells
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A variety of 8-substituted guanosine and 2'-deoxyguanosine derivatives were synthesized and tested as inducers of the differentiation of Friend murine erythroleukemia cells in culture. The most active agents in the guanosine series were 8-substituted -N(CH3)2, -NHCH3, -NH2, -OH, and -SO2CH3, which caused 68, 42, 34, 33, and 30% of erythroleukemia cells to attain benzidine positivity, a functional measure of maturation, at concentrations of 5, 1, 0.4, 5, and 5 mM, respectively. The 8-OH derivative of the 2'-deoxyguanosine series produced comparable activity, causing 62% benzidine-positive cells at a level of 0.2 mM. These findings indicate that 8-substituted analogues of guanosine and 2'-deoxyguanosine have the potential to terminate leukemia cell proliferation through conversion to end-stage differentiated cells.
- Lin,Cheng,Ishiguro,Sartorelli
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p. 1194 - 1198
(2007/10/02)
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