João P. Telo

DNA cleavage activity of VIVO(acac)2 and derivatives

The DNA cleavage activity of several beta-diketonate vanadyl complexes is examined. Vanadyl acetylacetonate, V(IV)O(acac)(2), 1, shows a remarkable activity in degrading plasmid DNA in the absence of any activating agents, air and photoirradiation. The cleaving activity of several related complexes V(IV)O(hd)(2) (2, Hhd=3,5-heptanedione), V(IV)O(acac-NH(2))(2) (3, Hacac-NH(2)=acetoacetamide) and V(IV)O(acac-NMe(2))(2) (4, Hacac-NMe(2)=N,N-dimethylacetoacetamide) is also evaluated. It is shown that 2 exhibits an activity similar to 1, while 3 and 4 are much less efficient cleaving agents. The different activity of the complexes is related to their stability towards hydrolysis in aqueous solution, which follows the order 1 approximately 2>>3 approximately 4. The nature of the pH buffer was also found to be determinant in the nuclease activity of 1 and 2. In a phosphate buffered medium DNA cleavage by these agents is much more efficient than in tris, hepes, mes or mops buffers. The reaction seems to take place through a mixed mechanism, involving the formation of reactive oxygen species (ROS), namely OH radicals, and possibly also direct cleavage at phosphodiester linkages induced by the vanadium complexes.

Insights into the Mechanisms of Toxicity and Tolerance to the Agricultural Fungicide Mancozeb in Yeast, as Suggested by a Chemogenomic Approach

Abstract Saccharomyces cerevisiae was used to uncover the mechanisms underlying tolerance and toxicity of the agricultural fungicide mancozeb, linked to cancer and Parkinsons disease development. Chemogenomics screening of a yeast deletion mutant collection revealed 286 genes that provide protection against mancozeb toxicity. The most significant Gene Ontology (GO) terms enriched in this dataset are associated to transcriptional machinery, vacuolar organization and biogenesis, intracellular trafficking, and cellular pH regulation. Clustering based on physical and genetic interactions further highlighted the role of oxidative stress response, protein degradation and carbohydrate/energy metabolism in mancozeb stress tolerance. Mancozeb was found to act in yeast as a thiol-reactive compound, but not as a free radical or reative oxygen species (ROS) inducer, leading to massive oxidation of protein cysteins, consistent with the requirement of genes involved in glutathione biosynthesis and reduction and in protein degradation to provide mancozeb resistance. The identification of Botrytis cinerea homologues of yeast mancozeb tolerance determinants is expected to guide studies on mancozeb mechanisms of action and tolerance in phytopathogenic fungi. The generated networks of protein-protein associations of yeast mancozeb tolerance determinants and their human orthologues share a high degree of similarity. This toxicogenomics analysis may, thus, increase the understanding of mancozeb toxicity and adaptation mechanisms in humans.

Mechanism of free radical oxidation of caffeine in aqueous solution

The oxidation of caffeine with persulfate and hydroxyl radicals in aqueous solution has been studied by EPR spectroscopy and HPLC analysis. In both cases the formation of 1,3,7-trimethyluric acid is observed as the main final product. A C8–OH radical adduct is postulated as the intermediate after reaction with OH˙, and leads to the final product after further oxidation. This radical is too short-lived to be observed by EPR. After oxidation of caffeine with SO4˙- its radical cation is detected by EPR. This radical reacts with water to produce the above mentioned C8–OH radical adduct after deprotonation. In the presence of dibasic phosphate a spectrum attributed to the C8-phosphate radical adduct is observed. This radical results from the nucleophilic attack of the buffer on the radical cation of caffeine. Hydrolysis and oxidation of the phosphate radical intermediate results in the formation of 1,3,7-trimethyluric acid.

Radicals derived from uric acid and its methyl derivatives in aqueous solution: an EPR spectroscopy and theoretical study

The oxidation of uric and of four N-methyluric acids in aqueous solution was studied by EPR spectroscopy. The primary oxidising radicals react with uric acid and its methyl derivatives by formal hydrogen abstraction from an NH group to yield radical-anions in neutral or moderately basic solutions and the respective radical-dianions in basic media. In the case of uric acid, the radical-trianion was detected at very high pH. The pKa values of the radical-anions were determined to be in the range 9.5-11.2. The pKa of uric acid radical-dianion was estimated to be 13.0. DFT calculations were performed to assign the hyperfine coupling constants and to determine the predominant tautomeric structure of the radicals. The uric acid radical-anion exists as the N1H, N9H tautomer, while in the radical-dianion the N1H structure is the most stable one. The intrinsic acidity of the NH protons both in uric acid and in its radicals seems to follow the order N1H < N9H < N3H.

Intramolecular electron exchange in the 2,7-dinitronaphthalene radical anion : electron paramagnetic resonance and kinetic data

The rates of intramolecular electron exchange in the radical anion of 2,7-dinitronaphthalene in protic and aprotic solvents have been determined from alternating line-broadening effects in EPR spectra. Rate constants in alcohols range from 9.0 × 105 s–1(methanol, 297 K) to 1.7 × 108 s–1(propan-2-ol, 361 K), while in aprotic solvents they range from 1.2 × 108 s–1(acetonitrile, 232 K) to 3.7 × 109 s–1(dimethylformamide, 342 K). The reactions were found to be adiabatic and uniform.Applying Marcus theory and the Rips–Jortner approach to the solvent dependence of the rates, the transferred charge ze0 was determined. It was found that z decreases with increasing temperature, changing from 1.0 at 260 K to 0.98 at 320 K in protic solvents and from 0.58 at 280 K to 0.47 at 340 K in aprotic solvents. This result is explained on the basis of the effect of temperature on the asymmetric solvation of the anions.The outer-sphere reorganization energy, λ0, was obtained by applying the ellipsoidal cavity model and was found to be strongly dependent on the solvent polarity, changing at 293 K from 57.7 kJ mol–1 in hexamethylphosphoric triamide to 235.8 kJ mol–1 in methanol. Activation energies were found to be higher in protic than in aprotic solvents.Results for the 2,7-dinitronaphthalene radical anion are compared with previous data reported for the 1,3-dinitrobenzene radical anion. Rate constants are smaller in the former while outer-sphere reorganization energies are larger. Activation parameters were found of the same order of magnitude for both radical anions.

Production of poly (3-hydroxybutyrate-co-4-hydroxybutyrate) by Burkholderia sacchari using wheat straw hydrolysates and gamma-butyrolactone

Burkholderia sacchari DSM 17165 is able to grow and produce poly(3-hydroxybutyrate) both on hexoses and pentoses. In a previous study, wheat straw lignocellulosic hydrolysates (WSH) containing high C6 and C5 sugar concentrations were shown to be excellent carbon sources for P(3HB) production. Using a similar feeding strategy developed for P(3HB) production based on WSH, fed-batch cultures were developed aiming at the production of the copolymer P(3HB-co-4HB) (poly(3-hydroxybutyrate-co-4-hydroxybutyrate)) by B. sacchari. The ability of this strain to synthesize P(3HB-co-4HB) was first shown in shake flasks using gamma-butyrolactone (GBL) as precursor of the 4HB units. Fed-batch cultures using glucose as carbon source (control) and GBL were developed to achieve high copolymer productivities and 4HB incorporations. The attained P(3HB-co-4HB) productivity and 4HB molar% were 0.7g/(Lh) and 4.7molar%, respectively. The 4HB incorporation was improved to 6.3 and 11.8molar% by addition of 2g/L propionic and acetic acid, respectively. When WSH were used as carbon source under the same feeding conditions, the values achieved were 0.5g/(Lh) and 5.0molar%, respectively. Burkholderia sacchari, a strain able to produce biopolymers based on xylose-rich lignocellulosic hydrolysates, is for the first time reported to produce P(3HB-co-4HB) using gamma butyrolactone as precursor.

Electronic communication in linear oligo(azobenzene) radical anions.

The radical anions of five bis(azobenzene) and one tris(azobenzene) compounds were studied by optical and electron paramagnetic resonance (EPR) spectroscopies in polar aprotic solvents. The radicals with planar or almost-planar bridges are charge-delocalized mixed-valence species. Localization of charge occurs only on radicals with highly twisted biphenyl bridges. The electronic coupling between the azobenzene charge-bearing units, calculated as half the energy of the intervalence band for the charge-delocalized and by the Hush equation for the charge-localized radicals, decreases with the distance and torsion angle between azobenzene units. These radicals have smaller electronic couplings between charge-bearing units than other mixed-valence organic radicals with similar bridges. However, the application of a three-stage model to the tris(azobenzene) radical anion intervalence band yields an electronic coupling between consecutive azobenzenes that is higher than in any of the bis(azobenzene) radicals studied.

EPR spectroscopic study of the radical oxidation of hydroxypurines in aqueous solution: acid–base properties of the derived radicals

EPR spectroscopy has been employed to study the reactions of SO4˙– and OH˙ radicals with xanthine, 8-methylxanthine, hypoxanthine and 2-hydroxypurine.Radicals obtained on the reaction of SO4˙– with these hydroxypurines resulted, depending on the pH, from one-electron oxidation of either the parent compound or its anion, followed by deprotonation. For xanthine and 8-methylxanthine the respective radical anion and radical dianion were observed, for hypoxanthine the species detected correspond to the neutral radical and radical anion, whereas for 2-hydroxypurine only its radical anion was observed.The pKa values of the radical anion of xanthine and 8-methylxanthine were estimated as 12.0 ± 0.1 and 12.1 ± 0.5, respectively, whereas the pKa value of the hypoxanthine neutral radical was 7.7 ± 0.1.In the reaction with OH˙, only the radical anion of xanthine was detected.

Gold and nickel alkyl substituted bis-thiophenedithiolene complexes: anionic and neutral forms

The Au and Ni monoanionic complexes of tert-butyl and diisopropyl substituted thiophenedithiolate ligands, [M(α-tb-tpdt)2] and [M(α-dp-tpdt)2], were synthesized and characterized namely by single crystal X-ray diffraction and magnetic susceptibility measurements. These complexes, prepared in a first step as monoanionic species, are easier to oxidize than the related non-substituted thiophenedithiolates and could be obtained also as stable neutral species. As expected, the peripheral alkyl groups in the ligands also confer a high solubility to the complexes in common organic solvents. The neutral gold complex [Au(α-tb-tpdt)2] presents a significant ligand asymmetry indicative of unpaired electron localization in one ligand at variance with [Au(α-dp-tpdt)2] that is within experimental uncertainty fully symmetric illustrating the role of intermolecular interactions in the stabilization of SOMO⋯SOMO interactions. While in [Au(α-tb-tpdt)2] a significant intermolecular interaction between paramagnetic molecules is possible leading to diamagnetic dimers of molecules, in [Au(α-dp-tpdt)2] the bulkier substituents prevent the intermolecular interactions, leading to a regular stacking of molecules in symmetrical configuration. The regular stacks of paramagnetic [Au(α-dp-tpdt)2] units behave, at high temperatures, as antiferromagnetic chains undergoing an AFM transition at ca. 25 K.

Structure and redox properties of radicals derived from one-electron oxidised methylxanthines.

Abstract The repair of the one electron oxidised form of a xanthine derivative by another xanthine derivative was studied by reacting aqueous binary mixtures of xanthine derivatives with sulphate radical anion (SO4˙−) and following the concentration of both compounds as a function of time. The relative behaviour observed enabled the establishment of a qualitative order of antioxidant capacities for the several xanthines studied in acidic and neutral media. The same order was confirmed quantitatively by measuring the reduction potentials of the xanthines by cyclic voltammetry. Theoretical DFT calculations were used to calculate the relative stabilities of the tautomers of each xanthine neutral radical. It was also demonstrated that the deprotonation of a xanthine radical cation never occurs from N1, unless no other possibility is available. At high pH values, it was possible to obtain the ESR spectra of the radical anions derived from 1-methylxanthine, 3-methylxanthine and xanthosine. The theoretical calculations also enabled the assignment of the ESR hyperfine coupling constants of the spectra of these radical anions. The coupling constants calculated are in good agreement with the experimental values.