Nuclear and mitochondrial genome instability induced by fractions of ethanolic extract from Hovenia dulcis Thunberg in Saccharomyces cerevisiae strains.

Hovenia dulcis is a plant commonly used as a pharmaceutical supplement, having displayed important pharmacological properties such antigiardic, antineoplastic and hepatoprotective. The purpose of this work was investigate the cytotoxic, genotoxic and mutagenic potential from fractions of Hovenia dulcis ethanolic extract on Saccharomyces cerevisiae strains FF18733 (wild type) and CD138 (ogg1). Ethanolic extract from Hovenia dulcis leaves was fractioned using organic solvents according to increasing polarity: Hexane (1:1), dichlorometane (1:1), ethyl acetate (1:1) and butanol (1:1). Three experimental assays were performed, such as (i) inactivation of cultures; (ii) mutagenesis (canavanine resistance system) and (iii) loss of mitochondrial function (petites colonies). The findings shown a decrease in cell viability in FF18733 and CD138 strains; all fractions of the extract were mutagenic in CD138 strain; only ethyl acetate and butanol fractions increased the rate of petites colonies for CD138 strains. Ethyl acetate and n-butanol fractions induces mutagenicity, at the evaluated concentrations, in mitochondrial and genomic DNA in CD138 strain, mediated by oxidative lesions. In conclusion, it is possible to infer that the lesions caused by the extract fractions could be mediated by reactive oxygen species and might reach multiple molecular targets to cause cellular damage.

Insights and controversies on sunscreen safety.

Although sunlight provides several benefits, ultraviolet (UV) radiation plays an important role in the development of various skin damages such as erythema, photoaging, and photocarcinogenesis. Despite cells having endogenous defense systems, damaged DNA may not be efficiently repaired at chronic exposure. In this sense, it is necessary to use artificial defense strategies such as sunscreen formulations. UV filters should scatter, reflect, or absorb solar UV radiation in order to prevent direct or indirect DNA lesions. However, the safety of UV filters is a matter of concern due to several controversies reported in literature, such as endocrine alterations, allergies, increased oxidative stress, phototoxic events, among others. Despite these controversies, the way in which sunscreens are tested is essential to ensure safety. Sunscreen regulation includes mandatory test for phototoxicity, but photogenotoxicity testing is not recommended as a part of the standard photosafety testing program. Although available photobiological tests are still the first approach to assess photosafety, they are limited. Some existing tests do not always provide reliable results, mainly due to limitations regarding the nature of the assessed phototoxic effect, cell UV sensitivity, and the irradiation protocols. These aspects bring queries regarding the safety of sunscreen wide use and suggest the demand for the development of robust and efficient in vitro screening tests to overcome the existing limitations. In this way, Saccharomyces cerevisiae has stood out as a promising model to fill the gaps in photobiology and to complete the mandatory tests enabling a more extensive and robust photosafety assessment.

Saccharomyces cerevisiae strains as bioindicators for titanium dioxide sunscreen photoprotective and photomutagenic assessment.

Although several short-term assays are available for cosmetic photosafety assessment, cell models are usually highly sensitive to UV radiation, tending to overestimate both phototoxic and photomutagenic risks. In addition, these assays are performed with UV doses/fluences that do not correspond to actual environmental conditions. In this sense, Saccharomyces cerevisiae has already proved to be an interesting tool to predict photomutagenic potential of several compounds, including sunscreens. Yeast can support environmental UVB doses compatible with human daily sunlight exposure, allowing the use of irradiation sources to faithfully mimic the external conditions of ambient sunlight. Herein, we used a set of S. cerevisiae mutant strains sensitive to UVA, UVB and Solar Simulated Light sources in order to evaluate their potential as bioindicators for sunscreen development. The bioindicator potential of the strains was tested with the widely-used titanium dioxide inorganic sunscreen. The AWP001 (yno1) and LPW002 (ogg1yno1) strains obtained in this study stood out as promising experimental tools for the validation of this assay. Overall, our results evidenced a set of S. cerevisiae strains particularly useful for evaluating both photoprotective (efficacy) and photo/antiphotomutagenic (safety) potential of UV filters, meeting the industries and regulatory agencies demand for robust and efficient in vitro screening tests.

Photoprotection assessment of olive (Olea europaea L.) leaves extract standardized to oleuropein: In vitro and in silico approach for improved sunscreens.

Olive leaves contain higher amount of polyphenols than olive oil and represent a waste product from olive harvest and pruning of olive trees. The most abundant compound in olive leaves is oleuropein. Benefits of the topical application of olive leaves extract were previously reported, but little information is available on its photoprotective potential and the result of the association of this extract with organic UV filters in topical sunscreen formulations. The olive leaves extract photoprotective potential is less explored for both oral and topical photoprotection in comparison with other plants extracts and polyphenols, such as Polypodium leucotomos extract and resveratrol. There are increasing efforts towards developing more efficient sunscreens and a photoprotection assessement along with a better understanding of the photochemistry of naturally occurring sunscreens could aid the design of new and improved commercial sunscreen formulations. This study was designed to investigate the photoprotective potential of olive leaves extract standardized for oleuropein performing a set of in vitro and in silico tools as an innovative approach, highlighting yeast assays, in vitro Sun Protection Factor (SPF) and molecular modelling studies of UV absorption. This study supports the use of olive leaves extract for photoprotection, as an effective photoprotective, anti-mutagenic and antioxidant active, also showing a synergistic effect in association with UV filters with an improvement on in vitro SPF of sunscreen formulations.

Microbiological Decontamination of Water: Improving the Solar Disinfection Technique (SODIS) with the Use of Nontoxic Vital Dye Methylene Blue.

Rational use of water is a major challenge for governments and global organizations, with easy and inexpensive interventions being sought by communities that are not supplied with drinking water. In this context, solar disinfection (SODIS) has shown great efficiency for water disinfection. To speed up the process and improve inactivation, we studied the effects of methylene blue (MB) as a photodynamic agent because of its ability to absorb visible light (red wavelength) and generate singlet oxygen as a reactive species, thereby inactivating bacteria and viruses present in water. In this study, samples of clean mineral water were artificially contaminated with Gram-positive (Staphylococcus epidermidis or Deinococcus radiodurans) or with Gram-negative strains (Escherichia coli or Salmonella typhimurium) and exposed to traditional SODIS or to MB-SODIS. A lethal synergistic effect was observed when cultures were illuminated in the presence of MB. The obtained results indicate that bacterial inactivation can be achieved in a much shorter time when using MB associated with SODIS treatment. Therefore, this technique was able to provide safe water for consumption through the inactivation of microorganisms in general, including pathogens and some strains resistant to the traditional SODIS procedure, thus allowing its use in areas usually less exposed to sunlight.

Effect of the FE2+ chelation by 2,2'-dipyridyl in the doxorubicin-induced lethality in breast tumor cell lines.

Breast cancer cells may exhibit changes in iron homeostasis, which results in increased labile iron pool (LIP) levels. Several studies highlight the crucial role of high LIP levels in the maintenance of tumor cell physiology. Iron chelators have been tested in anticancer therapy in combination with chemotherapeutic agents, to improve drug efficacy. Thus, the aim of this study was to evaluate the effect of 2,2'-dipyridyl (DIP), a Fe2+ chelator, in combination with doxorubicin (DOX) in breast tumor cells. The maximum concentration of DIP that did not significantly reduce the viability of MDA-MB-231 cells was 10µM and for MCF-7 cells was 50µM. We observed that MCF-7 had higher LIP levels than MDA-MB-231 cells. DIP alone increased ROS generation in MCF-7 cells, and DIP pretreatment reduced ROS generation induced by DOX treatment. In conclusion, the increase in MCF-7 cell viability induced by DIP pretreatment in DOX-treated cells seems to be related to an increase in the cellular antioxidant capacity and the iron chelator did not improve drug efficacy in the two breast tumor cell lines analyzed.

Influence of Ogg1 repair on the genetic stability of ccc2 mutant of Saccharomyces cerevisiae chemically challenged with 4-nitroquinoline-1-oxide (4-NQO).

In Saccharomyces cerevisiae, disruption of genes by deletion allowed elucidation of the molecular mechanisms of a series of human diseases, such as in Wilson disease (WD). WD is a disorder of copper metabolism, due to inherited mutations in human copper-transporting ATPase (ATP7B). An orthologous gene is present in S. cerevisiae, CCC2 gene. Copper is required as a cofactor for a number of enzymes. In excess, however, it is toxic, potentially carcinogenic, leading to many pathological conditions via oxidatively generated DNA damage. Deficiency in ATP7B (human) or Ccc2 (yeast) causes accumulation of intracellular copper, favouring the generation of reactive oxygen species. Thus, it becomes important to study the relative importance of proteins involved in the repair of these lesions, such as Ogg1. Herein, we addressed the influence Ogg1 repair in a ccc2 deficient strain of S. cerevisiae. We constructed ccc2-disrupted strains from S. cerevisiae (ogg1ccc2 and ccc2), which were analysed in terms of viability and spontaneous mutator phenotype. We also investigated the impact of 4-nitroquinoline-1-oxide (4-NQO) on nuclear DNA damage and on the stability of mitochondrial DNA. The results indicated a synergistic effect on spontaneous mutagenesis upon OGG1 and CCC2 double inactivation, placing 8-oxoguanine as a strong lesion-candidate at the origin of spontaneous mutations. The ccc2 mutant was more sensitive to cell killing and to mutagenesis upon 4-NQO challenge than the other studied strains. However, Ogg1 repair of exogenous-induced DNA damage revealed to be toxic and mutagenic to ccc2 deficient cells, which can be due to a detrimental action of Ogg1 on DNA lesions induced in ccc2 cells. Altogether, our results point to a critical and ambivalent role of BER mediated by Ogg1 in the maintenance of genomic stability in eukaryotes deficient in CCC2 gene.

The anabolic androgenic steroid nandrolone decanoate disrupts redox homeostasis in liver, heart and kidney of male Wistar rats.

The abuse of anabolic androgenic steroids (AAS) may cause side effects in several tissues. Oxidative stress is linked to the pathophysiology of most of these alterations, being involved in fibrosis, cellular proliferation, tumorigenesis, amongst others. Thus, the aim of this study was to determine the impact of supraphysiological doses of nandrolone decanoate (DECA) on the redox balance of liver, heart and kidney. Wistar male rats were treated with intramuscular injections of vehicle or DECA (1 mg.100 g(-1) body weight) once a week for 8 weeks. The activity and mRNA levels of NADPH Oxidase (NOX), and the activity of catalase, glutathione peroxidase (GPx) and total superoxide dismutase (SOD), as well as the reduced thiol and carbonyl residue proteins, were measured in liver, heart and kidney. DECA treatment increased NOX activity in heart and liver, but NOX2 mRNA levels were only increased in heart. Liver catalase and SOD activities were decreased in the DECA-treated group, but only catalase activity was decreased in the kidney. No differences were detected in GPx activity. Thiol residues were decreased in the liver and kidney of treated animals in comparison to the control group, while carbonyl residues were increased in the kidney after the treatment. Taken together, our results show that chronically administered DECA is able to disrupt the cellular redox balance, leading to an oxidative stress state.

Titanium dioxide-montmorillonite nanocomposite as photoprotective agent against ultraviolet B radiation-induced mutagenesis in Saccharomyces cerevisiae: a potential candidate for safer sunscreens.

Photoprotective potential and biological consequences (mutagenic potential) of octyl-dimethyl-PABA (ODP), titanium dioxide (TiO2 ), and montmorillonite (MMT) upon ultraviolet B (UVB) irradiation, alone and in different associations [physical mixtures (PMs)], were evaluated using a Saccharomyces cerevisiae ogg1 mutant (deficient) strain. In addition, we developed and characterized a delaminated TiO2-pillared MMT, called the TiO2 -MMT nanocomposite (NC), which was also investigated in terms of its photoprotective and mutagenic potential. Overall, our results revealed an interesting TiO2 -MMT NC endowed with antimutagenic activity that can be associated to organic sunscreen molecule (ODP) and still maintain its positive effect, whereas its respective PM is unable to grant antimutagenic protection against UVB.

Insight from the draft genome of Dietzia cinnamea P4 reveals mechanisms of survival in complex tropical soil habitats and biotechnology potential.

The draft genome of Dietzia cinnamea strain P4 was determined using pyrosequencing. In total, 428 supercontigs were obtained and analyzed. We here describe and interpret the main features of the draft genome. The genome contained a total of 3,555,295 bp, arranged in a single replicon with an average G+C percentage of 70.9%. It revealed the presence of complete pathways for basically all central metabolic routes. Also present were complete sets of genes for the glyoxalate and reductive carboxylate cycles. Autotrophic growth was suggested to occur by the presence of genes for aerobic CO oxidation, formate/formaldehyde oxidation, the reverse tricarboxylic acid cycle and the 3-hydropropionate cycle for CO(2) fixation. Secondary metabolism was evidenced by the presence of genes for the biosynthesis of terpene compounds, frenolicin, nanaomycin and avilamycin A antibiotics. Furthermore, a probable role in azinomycin B synthesis, an important product with antitumor activity, was indicated. The complete alk operon for the degradation of n-alkanes was found to be present, as were clusters of genes for biphenyl ring dihydroxylation. This study brings new insights in the genetics and physiology of D. cinnamea P4, which is useful in biotechnology and bioremediation.

Genotoxic and Cytotoxic Safety Evaluation of Papain (Carica papaya L.) Using In Vitro Assays.

Papain, a phytotherapeutic agent, has been used in the treatment of eschars and as a debriding chemical agent to remove damaged or necrotic tissue of pressure ulcers and gangrene. Its benefits in these treatments are deemed effective, since more than 5000 patients, at the public university hospital at Rio de Janeiro, Brazil, have undergone papain treatment and presented satisfactory results. Despite its extensive use, there is little information about toxic and mutagenic properties of papain. This work evaluated the toxic and mutagenic potential of papain and its potential antioxidant activity against induced-H(2)O(2) oxidative stress in Escherichia coli strains. Cytotoxicity assay, Growth inhibition test, WP2-Mutoxitest and Plasmid-DNA treatment, and agarose gel electrophoresis were used to investigate if papain would present any toxic or mutagenic potential as well as if papain would display antioxidant properties. Papain exhibited negative results for all tests. This agent presented an activity protecting cells against H(2)O(2)-induced mutagenesis.

Enzymatic recognition of DNA damage induced by UVB-photosensitized titanium dioxide and biological consequences in Saccharomyces cerevisiae: evidence for oxidatively DNA damage generation.

Although titanium dioxide (TiO(2)) has been considered to be biologically inert, finding use in cosmetics, paints and food colorants, recent reports have demonstrated that when TiO(2) is attained by UVA radiation oxidative genotoxic and cytotoxic effects are observed in living cells. However, data concerning TiO(2)-UVB association is poor, even if UVB radiation represents a major environmental carcinogen. Herein, we investigated DNA damage, repair and mutagenesis induced by TiO(2) associated with UVB irradiation in vitro and in vivo using Saccharomyces cerevisiae model. It was found that TiO(2) plus UVB treatment in plasmid pUC18 generated, in addition to cyclobutane pyrimidine dimers (CPDs), specific damage to guanine residues, such as 8-oxo-7,8-dihydroguanine (8-oxoG) and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG), which are characteristic oxidatively generated lesions. In vivo experiments showed that, although the presence of TiO(2) protects yeast cells from UVB cytotoxicity, high mutation frequencies are observed in the wild-type (WT) and in an ogg1 strain (deficient in 8-oxoG and FapyG repair). Indeed, after TiO(2) plus UVB treatment, induced mutagenesis was drastically enhanced in ogg1 cells, indicating that mutagenic DNA lesions are repaired by the Ogg1 protein. This effect could be attenuated by the presence of metallic ion chelators: neocuproine or dipyridyl, which partially block oxidatively generated damage occurring via Fenton reactions. Altogether, the results indicate that TiO(2) plus UVB potentates UVB oxidatively generated damage to DNA, possibly via Fenton reactions involving the production of DNA base damage, such as 8-oxo-7,8-dihydroguanine.

Genotoxicity and mutagenicity of Echinodorus macrophyllus (chapéu-de-couro) extracts.

Echinodorus macrophyllus, commonly known as chapéu-de-couro, is a medicinal plant used in folk medicine to treat inflammation and rheumatic diseases. In this work, we used short-term bacterial assays based on the induction of SOS functions to examine the genotoxicity and mutagenicity of an aqueous extract of E. macrophyllus leaves. Whole extract and an ethyl acetate fraction showed similar genotoxicity and caused an ~70-fold increase in lysogenic induction. The extract also gave a positive result in the SOS chromotest with an increase of 12-fold in ß-Galactosidase enzymatic units. There was a strong trend towards base substitutions and frameshifts at purine sites in the mutations induced by the extract in Escherichia coli (CC103 and CC104 strains) and Salmonella typhimurium test strains (22-fold increase in histidine revertants in TA98 strain). Since reactive oxygen species may be implicated in aging process and in degenerative diseases, we used antioxidant compounds as catalase, thiourea and dipyridyl in the lysogenic induction test. All this compounds were able to reduce the induction factor observed in the treatment with chapéu-de-couro, thus suggesting that the genotoxicity and mutagenicity were attributable to the production of reactive oxygen species that targeted DNA purines.

Differential survival of Escherichia coli uvrA, uvrB, and uvrC mutants to psoralen plus UV-A (PUVA): Evidence for uncoupled action of nucleotide excision repair to process DNA adducts.

The nucleotide excision repair mechanism (NER) of Escherichia coli is responsible for the recognition and elimination of more than twenty different DNA lesions. Herein, we evaluated the in vivo role of NER in the repair of DNA adducts generated by psoralens (mono- or bi-functional) and UV-A light (PUVA) in E. coli. Cultures of wild-type E. coli K12 and mutants for uvrA, uvrB, uvrC or uvrAC genes were treated with PUVA and cell survival was determined. In parallel, kinetics of DNA repair was also evaluated by the comparison of DNA sedimentation profiles in all the strains after PUVA treatment. The uvrB mutant was more sensitive to PUVA treatment than all the other uvr mutant strains. Wild-type strain, and uvrA and uvrC mutants were able to repair PUVA-induced lesions, as seen by DNA sedimentation profiles, while the uvrB mutant was unable to repair the lesions. In addition, a quadruple fpg nth xth nfo mutant was unable to nick PUVA-treated DNA when the crude cell-free extract was used to perform plasmid nicking. These data suggest that DNA repair of PUVA-induced lesions may require base excision repair functions, despite proficient UvrABC activity. These results point to a specific role for UvrB protein in the repair of psoralen adducts, which appear to be independent of UvrA or UvrC proteins, as described for the classical UvrABC endonuclease mechanism.

Genotoxicity and mutagenicity of Echinodorus macrophyllus (chapéu-de-couro) extracts

Echinodorus macrophyllus, commonly known as chapéu-de-couro, is a medicinal plant used in folk medicine to treat inflammation and rheumatic diseases. In this work, we used short-term bacterial assays based on the induction of SOS functions to examine the genotoxicity and mutagenicity of an aqueous extract of E. macrophyllus leaves. Whole extract and an ethyl acetate fraction showed similar genotoxicity and caused an ~70-fold increase in lysogenic induction. The extract also gave a positive result in the SOS chromotest with an increase of 12-fold in beta-Galactosidase enzymatic units. There was a strong trend towards base substitutions and frameshifts at purine sites in the mutations induced by the extract in Escherichia coli (CC103 and CC104 strains) and Salmonella typhimurium test strains (22-fold increase in histidine revertants in TA98 strain). Since reactive oxygen species may be implicated in aging process and in degenerative diseases, we used antioxidant compounds as catalase, thiourea and dipyridyl in the lysogenic induction test. All this compounds were able to reduce the induction factor observed in the treatment with chapéu-de-couro, thus suggesting that the genotoxicity and mutagenicity were attributable to the production of reactive oxygen species that targeted DNA purines.

Comparative studies of phenotypic and genetic characteristics between two desulfurizing isolates of Rhodococcus erythropolis and the well-characterized R. erythropolis strain IGTS8.

Two Rhodococcus erythropolis isolates, named A66 and A69, together with the well-characterized R. erythropolis strain IGTS8 were compared biochemically and genetically. Both isolates, like strain IGTS8, desulfurized DBT to 2-hydroxybiphenyl (2-HBP), following the 4S pathway of desulfurization. Strain IGTS8 showed the highest (81.5%) desulfurization activity in a medium containing DBT at 30 degrees C. Strain A66 showed approximately the same desulfurization activity either when incubated at 30 degrees C or at 37 degrees C, while strain A69 showed an increase of desulfurization efficiency (up to 79%) when incubated at 37 degrees C. Strains A66 and A69 were also able to grow using various organosulfur or organonitrogen-compounds as the sole sulfur or nitrogen sources. The biological responses of A66, A69 and IGTS8 strains to a series of mutagens and environmental agents were evaluated, trying to mimic actual circumstances involved in exposure/handling of microorganisms during petroleum biorefining. The results showed that strains A69 and IGTS8 were much more resistant to UVC treatment than A66. The three desulfurization genes (dszA, dszB and dszC) present in strains A66 and A69 were partially characterized. They seem to be located on a plasmid, not only in the strain IGTS8, but also in A66 and A69. PCR amplification was observed using specific primers for dsz genes in all the strains tested; however, no amplification product was observed using primers for carbazole (car) or quinoline (qor) metabolisms. All this information contributes to broaden our knowledge concerning both the desulfurization of DBT and the degradation of organonitrogen compounds within the R. erythropolis species.

Enhanced sensitivity of Escherichia coli uvrB mutants to mitomycin C points to a UV-C distinct repair for DNA adducts.

Nucleotide excision repair (NER) in Escherichia coli repairs DNA by incising the damaged strand on the 3' and 5' sides of the lesion within pyrimidine dimers and DNA cross-linking adducts. Cross-linking adducts belong to a class of chemical damage to DNA that prevent strand separation, and thus, replication and transcription. For this reason, cross-linking agents such as mitomycin C (MC) have been used in cancer chemotherapy. The mechanisms involved in MC binding to DNA have already been defined; however, the repair of these lesions is not fully understood. Our goal was to study the repair of MC DNA lesions in E. coli cells. Several bacterial strains with specific mutations were tested for cellular inactivation and kinetics of DNA repair through analysis of DNA sedimentation profiles in alkaline sucrose gradients. The results obtained show that uvrB mutants are extremely sensitive to MC in contrast to the other isogenic uvrA and uvrC mutant strains. uvrB mutant strains are unable to repair DNA strand breaks produced by MC. Thus, UvrB might play a NER-uncoupled role in the repair of lesions induced by MC in vivo, different from its role on the repair of lesions produced by UV-C. Also it is suggested that a modified NER system is taking place in the repair of MC-adducts.

Interaction of stannous chloride leads to alteration in DNA, triphosphate nucleotides and isolated bases.

Stannous chloride (SnCl2) is a reducing chemical agent used in several man-made products. SnCl2 can generate reactive oxygen species (ROS); therefore, studies have been carried out in order to better understand its damaging action in biological systems. In this work, calf thymus DNA, triphosphate nucleotides and isolated bases were incubated with SnCl2 and the results were analyzed through UV spectrophotometry. The presence of stannous ions altered the absorption spectra of all three isolates. The amount of stannous ions associated to DNA was measured by atomic absorption spectrophotometry. Data showed that more than 40% of the initial SnCl2 concentration was present in the samples. Our results are in accordance with the damaging potential of this salt and present evidence that stannous ions can complex with DNA, inducing ROS in its vicinity, which may be responsible for the observed lesions.

Reevaluating the role of 1,10-phenanthroline in oxidative reactions involving ferrous ions and DNA damage.

It is widely believed that the iron chelator 1,10-phenanthroline (phen) is able to fully block the Fenton reaction by forming a complex (Fe(phen)3(2+), also known as ferroin) that cannot react with H2O2. We observed that phen cannot fully prevent 2-deoxyribose (5 mM) degradation induced by Fenton reagents (30 microM Fe(II) plus 100-500 microM H2O2); protection varied from 55% to 66% when the phen/Fe(II) ratio was 3:1 to 20:1. Inhibition of 2-deoxyribose damage was nearly unchanged if phen was pre-incubated with Fe(II). Moreover, preformed Fe(phen)3(2+) complex added to the solution containing H2O2 was able to induce 2-deoxyribose degradation and methane sulfinic acid formation from the oxidation of 5% DMSO. The partially protective effect of phen was unchanged with the use of either phosphate or HEPES as buffers (5 mM, pH 7.2), or in unbuffered media (pH 5.1). Both DMSO oxidation and 2-deoxyribose degradation correlated with the increase in Fe(phen)3(2+) concentration. Strand breaks in plasmid pTARGETtrade mark DNA induced by Fenton reagents (1 microM Fe(II) plus 25 microM H2O2) in HEPES buffer could only be partially prevented by phen, even when the chelator was 16 times more concentrated than Fe(II). In these experiments, Fe(phen)3(2+) and DNA were pre-incubated from 1 to 10 min before addition of H2O2. Moreover, a high level of DNA strand breakage was observed when iron and phen are added to the reaction immediately before H2O2. On the other hand, phen fully prevented 2-deoxyribose degradation induced by the autoxidation of 30 microM Fe(II) in phosphate-buffered (3 to 30 mM) media. Our data provide evidence that the Fe(phen)3(2+) complex induces in vitro oxidative damage in the presence of H2O2 (possibly by means of Fe(phen)3(2+) dissociation into Fe(phen)2(2+)), but they show that the complex cannot undergo autoxidation.