Oxidation Of Pyrimidines Research Articles

Expanding the π -conjugate structures: Synthesis of high-energy and insensitive Tricyclic fused energetic material

In order to meet the growing demand for different applications, great efforts have been made to develop new energetic materials with high energy and good safety. Fused cyclic energetic material, a special category of π-conjugate structures containing two or more rings, typically exhibits excellent performance and is currently a key focus of research. In this paper, we have designed and synthesized a new tricyclic, fused energetic material: 5-amino-[1,2,5]oxadiazolo[3,4-e]tetrazolo[1,5-c]pyrimidine 1-oxide (3). The reaction mechanism of the one-step reaction, including nucleophilic substitution and cyclization, has been theoretically investigated. Notably, compound 3 has a high density (1.864 g cm−3) at room temperature and good detonation properties (Dv: 8391 m s−1 and P: 28.2 GPa). Compound 3 has a relatively low impact and friction sensitivity (IS > 40 J, FS > 300 N), and acceptable thermal stability (Td: 175 °C), demonstrating the great potential of compound 3 as a high-energy, insensitive tricyclic fused energy-containing material.

Nanoemulsions for the Encapsulation of Hydrophobic Actives

This work analyzes the dispersion of two highly hydrophobic actives, (9Z)-N-(1,3-dihydroxyoctadecan-2-yl)octadec-9-enamide (ceramidelike molecule) and 2,6-diamino-4-(piperidin-1-yl)pyrimidine 1-oxide (minoxidil), using oil-in-water nanoemulsions with the aim of preparing stable and safe aqueous-based formulations that can be exploited for enhancing the penetration of active compounds through cosmetic substrates. Stable nanoemulsions with a droplet size in the nanometric range (around 200 nm) and a negative surface charge were prepared. It was possible to prepare formulations containing up to 2 w/w% of ceramide-like molecules and more than 10 w/w% of minoxidil incorporated within the oil droplets. This emulsions evidenced a good long-term stability, without any apparent modification for several weeks. Despite the fact that this work is limited to optimize the incorporation of the actives within the nanoemulsion-like formulations, it demonstrated that nanoemulsions should be considered as a very promising tool for enhancing the distribution and availability of hydrophobic molecules with technological interest.

Oxidative Stress and DNA Damage of Zebrafish Sperm at Different Stages of the Cryopreservation Process

Although gamete cryopreservation has facilitated advancement of reproduction research by allowing the storage of cells over prolonged periods of time, during freezing-thawing cycles, cells inevitably suffer from cryoinjuries. Here, we evaluate oxidative stress and DNA damage of zebrafish sperm at different stages of the cryopreservation process. It was generally observed that the freezing and thawing of the samples led to an increase in the generation of reactive oxygen species and the activity of the catalase enzyme and a reduction in the generation of sulfhydryl groups and superoxide dismutase activity. The alkaline comet assay demonstrated that DNA damage increased after equilibration time, with an even greater increase after freezing and thawing. The comet assay modified with the enzyme formamidopyrimidine glycosylase, and Endonuclease III demonstrated greater DNA damage than the standard comet assay, demonstrating a high degree of oxidation of purines and pyrimidines at all stages of cryopreservation. Our results show that the freeze and thaw processes cause greater oxidative stress and DNA damage than cryoprotectant toxicity during exposure at the equilibrium stage.

Genotoxicity and oxidative stress in Caiman latirostris hatchlings exposed to pesticide formulations and their mixtures during incubation period.

Agricultural expansion and the consequent use of pesticides lead to the loss and fragmentation of natural habitats of several wild species. Then, many species are inevitably exposed to a wide amount of pesticide formulations. Glyphosate (GLY)-based formulations are the most used herbicide, whereas two of the most employed insecticides are chlorpyrifos (CPF) and cypermethrin (CYP). The aim of this study was to evaluate genotoxicity, oxidative damage, and the modulation of antioxidants defenses in peripheral blood of Caiman latirostris after embryonic exposure to pesticide formulations and their mixtures. Pesticides concentrations employed were equivalent to those recommended in agricultural practices for application in soybean crops and a half of them: GLY: 2% and 1%; CYP: 0.12% and 0.06%; CPF: 0.8% and 0.4%. Two similar experiments (E1 and E2) were carried out in consecutive years, where C. latirostris eggs were exposed to pesticide formulations separately and in different mixtures through application on the incubation material. After hatching, blood samples were taken and genotoxicity and oxidative stress was evaluated through the micronucleus (MN) test, the modified comet assay, the lipid peroxidation (LPO) levels and the activities of catalase (CAT) and superoxide dismutase (SOD) antioxidant enzymes. The results indicated the presence of DNA damage, oxidation of purines and pyrimidines, and increased frequency of micronucleus (FMN) in the case of GLY, CYP, and CPF formulations exposure, as well as in all the mixtures tested, with respect to the control groups. Specifically, the results observed for the mixtures would indicate independent action or antagonism of the components for DNA damage and base oxidation (purines and pyrimidines) and a possible potentiation interaction for the FMN in two binary mixtures. However, there were not differences regarding lipid peroxidation, the activity of antioxidant enzymes and growth parameters. This study proved that the use of pesticide formulations at concentrations used in the field generate deleterious genetic effects on this species, then, exposure to them could threaten its survival and health status.

Methamphetamine (“crystal meth”) causes induction of DNA damage and chromosomal aberrations in human derived cells

Methamphetamine (METH) is a widely consumed psychostimulant drug; its acute toxic effects in brain and liver are well known, furthermore, there is some evidence in regard to its DNA damaging properties in humans. Therefore, we studied the impact of the drug on genomic stability in human derived hepatoma (HepG2) cells, which reflect the activation/detoxification of drugs better than other cell lines. Furthermore, experiments with human buccal derived cells (TR146) were conducted as the drug is consumed orally. Induction of DNA damage in both cell types with doses reflecting the exposure in abusers was found in single cell gel electrophoresis (SCGE) assays (which detect single and double strand breaks as well as apurinic sites). Furthermore, induction of micronuclei (formed as a consequence of structural and numerical chromosomal aberrations) and formation of nuclear buds resulting from gene amplifications was detected. Additional experiments with lesion-specific enzymes showed that the drug causes oxidation of purines and pyrimidines, indicating that its genotoxic effects may be due to oxidation of the DNA. Our findings support the assumption that the drug may cause adverse health effects (such as cancer and infertility) in long-term users which are causally related to DNA damage.

Simultaneous Electrochemical Determination of Purine and Pyrimidine Bases Using Cu Doped CeO2 Nanoparticles Modified Glassy Carbon Electrode

Deoxyribonucleic acid (DNA) is an important hereditary substance in humans and in almost all other living organisms. It plays a vital role in the storage of genetic information and instructs the biological synthesis of proteins and enzymes through the process of replication and transcription of heritage information Thus, the identification and quantification of either individual and/or group of purine and pyrimidine bases are important for diagnosis of certain diseases and genetic disorders. Though there exits quite a few methods for such sensing, there is a great demand for a cheap, sensitive and interference free electrode for simultaneous determination of all the four neuclobases due to their important role in medical diagnosis, crime detection and biocomputing. In the present work, an efficient electrochemical sensor has been fabricated for simultaneous determination of purine bases adenine (AD), guanine (GU) and pyrimidines bases thymine (TY), cytosine (CY) using Cu doped CeO2 nanoparticles modified glassy carbon electrode (Cu-CeO2/GCE). Direct electrocatalytic activities of DNA bases have been studied in phosphate buffer solution (PBS, pH 7.0) without any enzyme using voltammetric techniques. 3 wt% Cu doped CeO2 modified GCE showed two well defined anodic peaks each corresponding to the oxidation of purine (AD, GU) and pyrimidine (TY, CY) bases with significant peak to peak potential separation of 312 mV (AD-GU) and 200 mV (TY-CY) which were large enough for the selective and simultaneous determination of these bases in their mixture. Under optimum conditions, calibration plots for the simultaneous detection of the purine and pyrimidine bases were linear in the concentration range of 0.1– 500 μM for AD, 1–650 μM for GU, 1-300 μM for TY and 1-250 for CY with detection limit values of 0.021, 0.031, 0.024 and 0.038 μM respectively. Additionally the developed sensor exhibited good repeatability, reproducibility, sufficient stability and good anti-interference ability and was successfully applied for simultaneous detection of AD, GU, TY and CY in denatured DNA sample with satisfactory results. References Y. Deng, Y.L. Xia, C.H. Xiao, Biosens. Bioelectron. 31 (2012) 469–474. N. Lavanya, C. Sekar, R. Murugan, G. Ravi, Mater. Sci. Engg. C 65 (2016) 278–286.

Highly Chemoselective and Enantioselective Catalytic Oxidation of Heteroaromatic Sulfides via High-Valent Manganese(IV)–Oxo Cation Radical Oxidizing Intermediates

A manganese complex with a porphyrin-like ligand that catalyzes the highly chemoselective and enantioselective oxidation of heteroaromatic sulfides, including imidazole, benzimidazole, indole, pyridine, pyrimidine, pyrazine, sym-triazine, thiophene, thiazole, benzothiazole, and benzoxazole, with hydrogen peroxide is described, furnishing the corresponding sulfoxides in good to excellent yields and enantioselectivities (up to 90% yield and up to >99% ee) within a short reaction time (0.5 h). The practical utility of the method has been demonstrated in the gram-scale synthesis of chiral sulfoxide. Mechanistic studies, performed with 18O-labeled water (H218O), hydrogen peroxide (H218O2), and cumyl hydroperoxide, reveal that a high-valent manganese–oxo species is generated as the oxygen atom delivering agent via carboxylic acid assisted heterolysis of O–O bonds. Density functional theory (DFT) calculations were also carried out to give further insight into the mechanism of manganese-catalyzed sulfoxidation. O...

Thymine and Other Prebiotic Molecules Produced from the Ultraviolet Photo-Irradiation of Pyrimidine in Simple Astrophysical Ice Analogs

The informational subunits of RNA or DNA consist of substituted N-heterocyclic compounds that fall into two groups: those based on purine (C₅H₄N₄) (adenine and guanine) and those based on pyrimidine (C₄H₄N₂) (uracil, cytosine, and thymine). Although not yet detected in the interstellar medium, N-heterocycles, including the nucleobase uracil, have been reported in carbonaceous chondrites. Recent laboratory experiments and ab initio calculations have shown that the irradiation of pyrimidine in ices containing H₂O, NH₃, or both leads to the abiotic production of substituted pyrimidines, including the nucleobases uracil and cytosine. In this work, we studied the methylation and oxidation of pyrimidine in CH₃OH:pyrimidine, H₂O:CH₃OH:pyrimidine, CH₄:pyrimidine, and H₂O:CH₄:pyrimidine ices irradiated with UV photons under astrophysically relevant conditions. The nucleobase thymine was detected in the residues from some of the mixtures. Our results suggest that the abundance of abiotic thymine produced by ice photolysis and delivered to the early Earth may have been significantly lower than that of uracil. Insofar as the delivery of extraterrestrial molecules was important for early biological chemistry on early Earth, these results suggest that there was more uracil than thymine available for emergent life, a scenario consistent with the RNA world hypothesis.

Visible light-sensitive properties of 1,2-dimethyl-2-(2-nitrophenyl)-2,3-dihydro-1H-perimidine

Four o- and p-nitro substituted 2-methyl-2-phenyl-2,3-dihydro-1H-perimidine derivatives were synthesized via a condensation–cyclization of naphthalene-1,8-diamine with nitro substituted acetophenones to study their photochemical properties. Only the o-nitrophenyl substituted perimidine 1 was found to be sensitive to light. When exposed to visible light under either aerobic conditions or argon atmosphere, it undergoes pyrimidine ring-opening/deoxygenation and pyrimidine oxidation to give the 2H-indazole 10 and perimidinone 11, respectively. A new visible light-sensitive o-nitrobenzylamino scaffold is introduced.

The Human Base Excision Repair Enzyme SMUG1 Directly Interacts with DKC1 and Contributes to RNA Quality Control

Single-strand-selective monofunctional uracil-DNA glycosylase 1 (SMUG1) is a base excision repair enzyme that removes uracil and oxidised pyrimidines from DNA. We show that SMUG1 interacts with the pseudouridine synthase Dyskerin (DKC1) and colocalizes with DKC1 in nucleoli and Cajal bodies. As DKC1 functions in RNA processing, we tested whether SMUG1 excised modified bases in RNA and demonstrated that SMUG1 has activity on single-stranded RNA containing 5-hydroxymethyldeoxyuridine, but not pseudouridine, the nucleoside resulting from isomerization of uridine by DKC1. Moreover, SMUG1 associates with the 47S rRNA precursor processed by DKC1, and depletion of SMUG1 leads to a reduction in the levels of mature rRNA accompanied by an increase in polyadenylated rRNA. Depletion of SMUG1, and, in particular, the combined loss of SMUG1 and DKC1, leads to accumulation of 5-hydroxymethyluridine in rRNA. In conclusion, SMUG1 is a DKC1 interaction partner that contributes to rRNA quality control, partly by regulating 5-hydroxymethyluridine levels.

Poly(alizarin red)/Graphene modified glassy carbon electrode for simultaneous determination of purine and pyrimidine

In this work, a poly(alizarin red)/Graphene composite film modified glassy carbon electrode (PAR/Graphene/GCE) was prepared for simultaneous determination of four DNA bases (guanine, adenine, thymine and cytosine) without any pretreatment. The morphology and interface property of PAR/Graphene films were examined by scanning electron microscopy and electrochemical impedance spectroscopy. The PAR/Graphene/GCE exhibited excellent electrocatalytic activity toward purine (guanine and adenine) and pyrimidine (thymine and cytosine) in 0.1M phosphate buffer solution (pH 7.4). Under optimum conditions, differential pulse voltammetry was used to detect the oxidation of purine and pyrimidine. The results showed that PAR/Graphene/GCE exhibited well-separated peaks, low detection limit, high sensitivity and wide linear range for simultaneous detection of purine and pyrimidine. The proposed sensor also has good stability and reproducibility. Furthermore, the modified electrode was applied for the detection of DNA bases in a fish sperm DNA sample with satisfactory results.

Mechanism for the abiotic synthesis of uracil via UV-induced oxidation of pyrimidine in pure H2O ices under astrophysical conditions

The UV photoirradiation of pyrimidine in pure H(2)O ices has been explored using second-order Moller-Plesset perturbation theory and density functional theory methods, and compared with experimental results. Mechanisms studied include those starting with neutral pyrimidine or cationic pyrimidine radicals, and reacting with OH radical. The ab initio calculations reveal that the formation of some key species, including the nucleobase uracil, is energetically favored over others. The presence of one or several water molecules is necessary in order to abstract a proton which leads to the final products. Formation of many of the photoproducts in UV-irradiated H(2)O:pyrimidine=20:1 ice mixtures was established in a previous experimental study. Among all the products, uracil is predicted by quantum chemical calculations to be the most favored, and has been identified in experimental samples by two independent chromatography techniques. The results of the present study strongly support the scenario in which prebiotic molecules, such as the nucleobase uracil, can be formed under abiotic processes in astrophysically relevant environments, namely in condensed phase on the surface of icy, cold grains before being delivered to the telluric planets, like Earth.

Development of Tester Strains Deficient in Nth/Nei DNA Glycosylases to Selectively Detect the Mutagenicity of Oxidized DNA Pyrimidines

Oxidative DNA damage is a major cause of mutation and cell death in aerobic organisms. In addition to 8-hydroxyguanine, oxidized DNA pyrimidines play important roles in mutagenesis. Salmonella typhimurium TA1535, widely used in mutagenicity assays, carries a hisG46 missense mutation and efficiently detects mutations at G:C base pairs. To detect oxidative mutagens that selectively modify pyrimidines, we constructed a derivative of strain TA1535, termed YG3206, which lacks the Nei and Nth DNA glycosylases that excise oxidized pyrimidines from DNA. This novel strain easily detected the mutagenicity of L-cysteine, L-penicillamine, dopamine-HCl, and phenazine methosulfate, which are non-mutagenic or only weakly mutagenic in the TA1535 parent strain. A second strain that is equivalent to YG3206 but harbors the plasmid pKM101 which carries mucAB encoding DNA polymerase R1, termed YG3216, was significantly sensitive to phenazine ethosulfate. The compound was not mutagenic in either YG3206 or the Fapy-glycosylase-defective strain YG3001. Potassium bromate and methylene blue plus visible light with metabolic activation induced mutations in YG3001 but not YG3206 or YG3216. The number of spontaneous His+ revertants per plate was 82 ± 16 (YG3206, ΔnthΔnei), 19 ± 4 (YG3001, Δfpg), and 6 ± 2 (TA1535), suggesting a significant contribution to spontaneous mutagenesis by endogenous pyrimidine oxidation. In the absence of exogenous chemical treatment, exposure to fluorescent light enhanced the spontaneous mutation frequency by approximately two-fold (YG3206), 13-fold (YG3001), and 10-fold (TA1535). These results suggest that certain environmental chemicals may selectively introduce mutagenic damage at DNA pyrimidines, and that these changes can be monitored by the use of YG3206.

Accurate micro- and macro- gas phase basicities of hydroxyl-radical-modified pyrimidines estimated by advanced quantum chemistry methods

The gas phase basicities (GPB) of 16 hydroxyl-radical-derived analogues of model pyrimidine nucleosides were analysed at B3LYP/aug-cc-pVDZ and G3MP2B3 levels of theory. All possible tautomeric equilibriums of neutral and protonated forms of pyrimidine analogues were taken into account. The oxidation of pyrimidines usually reduces the GPB values with respect to canonical values. The only exception was observed in the case of 5,6-dihydroxycytidine, which is more basic than model cytidine if both macro- and micro-scopic measures of basicities are used for comparison. In addition, 6-hydroxycytidine is characterised more by the basic character of the O2 atom compared to the more basic centre of cytidine, despite the fact that the GPB values of both these compounds are almost identical. Although the B3LYP/aug-cc-pVDZ approach seems to be an accurate and robust method for GPB estimation, the microscopic protonation properties are much more sensitive to this method since the difference in energy between some tautomers is often less than 1 kcal/mol with method-dependent succession. In such cases, G3MP2B3 is recommended.

Genotoxicity of chloroquine in rat liver cells: Protective role of free radical scavengers

The genotoxic effect of chloroquine (CQ), a 4-aminoquinoline antimalarial drug was investigated in rat liver cells using the alkaline comet assay. Chloroquine (0-1000 micromol/L) significantly increased DNA strand breaks of rat liver cells dose-dependently. Rat liver cells exposed to CQ (100-500 micromol/L) and treated with endonuclease III and formamidopyrimidine-DNA glycosylase, the bacterial DNA repair enzymes that recognize oxidized pyrimidine and purine, respectively, showed greater DNA damage than those not treated with the enzymes, providing evidence that CQ induced oxidation of purines and pyrimidines. Treatment of cells with 5 mmol/L N-acetylcysteine, an intracellular reactive oxygen species (ROS) scavenger, and 100 micromol/L and 250 micromol/L deferoxamine, an established iron chelator, significantly decreased the CQ-induced strand breaks and base oxidation, respectively. Similarly, the formation of DNA strand breaks and oxidized bases was prevented by vitamin C (10 micromol/L) (a water-soluble antioxidant), quercetin (50 micromol/L) (an antioxidant flavonoid), and kolaviron (30 micromol/L and 90 micromol/L) (an antioxidant and a liver hepatoprotective phytochemical). The results indicate that the genotoxicity of CQ in rat liver cells might involve ROS and that free radical scavengers may elicit protective effects in these cells.

Red wine polyphenols influence carcinogenesis, intestinal microflora, oxidative damage and gene expression profiles of colonic mucosa in F344 rats

Polyphenols from tea and other beverages such as red wine have been regarded with interest as possible chemopreventive agents against cancer. Here we report that red wine polyphenols (50 mg/kg) administered with the diet to F344 rats for 16 weeks inhibited colon carcinogenesis induced by azoxymethane (AOM, 7.4 mg/kg, total dose 74 mg/kg) or dimethylhydrazine (DMH, 30 mg/kg, total dose, 300 mg/kg). Polyphenol-treated animals had a consistently lower tumour yield compared to controls. In polyphenol-treated rats, the main bacterial strains in the faeces at sacrifice were Bacteroides, Lactobacillus and Bifidobacterium spp., whereas microorganisms predominantly identified in control-fed rats were Bacteroides, Clostridium and Propionibacterium spp. Wine polyphenols (57 mg/kg for 10 days, by gavage), administered to rats not treated with carcinogens, produced a significant decrease in the basal level of DNA oxidative damage of the colon mucosa as measured with the comet assay (average pyrimidine oxidation was reduced by 62% and purine oxidation by 57%, p < 0.05). To further explore the molecular effects of wine polyphenols we used the microarray technology to study gene expression profiles: rats were treated with 50 mg/kg wine polyphenols for 14 days, mixed in the diet. Global expression analysis of 5707 genes revealed an extensive down-regulation of genes involved in a wide range of physiological functions, such as metabolism, transport, signal transduction and intercellular signalling. By analysing metabolic pathways with the GenMAPP software program we observed that two major regulatory pathways were down-regulated in the colon mucosa of polyphenols-treated rats: inflammatory response and steroid metabolism. We also found a down-regulation of many genes regulating cell surface antigens, metabolic enzymes and cellular response to oxidative stress. In conclusion, reduction of oxidative damage, modulation of colonic flora and variation in gene expression may all concur in the modulation of intestinal function and carcinogenesis by wine polyphenols.

Different roles of Fpg and Endo III on catechol-induced DNA damage in extended-term cultures of human lymphocytes and L5178Y mouse lymphoma cells

Catechol is a genotoxic agent assumed to induce DNA damage via the oxidative pathway. Using the comet assay and the repair-specific enzymes formamido pyrimidine glycosylase (Fpg) and endonuclease III (Endo III), we examined the ability of catechol to induce DNA damage in extended-term cultures of human lymphocytes and mouse lymphoma cells. Our results suggest that mouse lymphoma cells are somewhat more sensitive towards catechol-induced DNA damage than the extended-term cultures of human lymphocytes. At high concentrations, the catechol-induced damage seemed to be independent of both Fpg and Endo III, possibly indicating a non-oxidative pathway for the DNA damage (involving, for example, a bulky adduct). The fact that Endo III, but not Fpg, enhanced the DNA damaging effect of catechol, suggests that this metabolite of benzene either mediates oxidation of pyrimidines rather than purines, or that oxidised purines are repaired more efficiently, at least in human lymphocytes. In the latter cells, low concentrations of catechol were found to reduce the DNA migration. Considering the role of Fpg and it’s adduct specific detection of 8-oxoguanine, this suggests that a low concentration of catechol has an antioxidative effect reducing the background levels of oxidized purines.

Heterogeneous oxidation of pyrimidine and alkyl thioethers in ionic liquids over mesoporous Ti or Ti/Ge catalysts.

Heterogeneous catalytic oxidation of a series of thioethers (2-thiomethylpyrimidine, 2-thiomethyl-4,6-dimethyl-pyrimidine, 2-thiobenzylpyrimidine, 2-thiobenzyl-4,6-dimethylpyrimidine, thioanisole, and n-heptyl methyl sulfide) was performed in ionic liquids by using MCM-41 and UVM-type mesoporous catalysts containing Ti, or Ti and Ge. A range of triflate, tetrafluoroborate, trifluoroacetate, lactate and bis(trifluoromethanesulfonyl)imide-based ionic liquids were used. The oxidations were carried out by using anhydrous hydrogen peroxide or the urea-hydrogen peroxide adduct and showed that ionic liquids are very effective solvents, achieving greater reactivity and selectivity than reactions performed in dioxane. The effects of halide and acid impurities on the reactions were also investigated. Recycling experiments on catalysts were carried out in order to evaluate Ti leaching and its effect on activity and selectivity.

Photooxidation of substituted pyrimidines in presence of peroxydisulphate in aqueous solution

Photooxidation of pyrimidine bases viz. thymine, uracil, cytosine, 5-bromouracil and 6-methyluracil in presence of peroxydisulphate (PDS) in aqueous solution at natural pH (7.5) has been carried out in a quantum yield reactor using a high-pressure mercury lamp. The reaction between pyrimidine and SO . - 4 is followed by measuring the absorbance of pyrimidine at its λ m a x . The rate of oxidation of pyrimidine is calculated from the plot of absorbance vs time using microcal origin computer program on a personal computer. On the basis of these experimental results and product analysis a probable mechanism has been suggested in which peroxydisulphate ion on photolysis gives sulphate radical anions which initiates the reactions by adding to C(5) or C(6) position of pyrimidine base leading to the formation of pyrimidine base radical via radical cation or hydrolysis. The pyrimidine radical further reacts with PDS to give the final products.

Stereoselective excision of thymine glycol lesions by mammalian cell extracts

Thymine glycols (Tg) are major pyrimidine oxidation products produced by chemical agents and ionizing radiation. Recent improvements in purification procedures gave us the opportunity to examine the incision of DNA duplexes containing a single (5 S,6 R)- or (5 R,6 S)-Tg lesion by mouse NTH1 DNA glycosylase and mammalian cell nuclear extracts. Time course experiments and steady state enzyme kinetics indicated that mNTH1 discriminates between the cis-Tg isomers. In addition, a variety of mammalian cell nuclear extracts showed a similar discrimination between the cis-Tg isomers. Trapping of Schiff base intermediates with sodium borohydride demonstrated that a single protein–DNA complex was formed in the presence of the nuclear extracts. The electrophoretic mobility of trapped complexes formed with both Tg isomers was identical to one another and similar to that of the complex formed with recombinant mNTH1. These results suggest that among all Tg-active DNA glycosylases, NTH1 is the major enzyme in mammalian cell nuclear extracts responsible for incision of duplexes containing cis-Tg isomers.