Characterization of O(2) ((1)Delta(g))-Derived Oxidation Products of Tryptophan: A Combination of Tandem Mass Spectrometry Analyses and Isotopic Labeling Studies

The fragmentation mechanisms of singlet oxygen [O(2) ((1)Delta(g))]-derived oxidation products of tryptophan (W) were analyzed using collision-induced dissociation coupled with (18)O-isotopic labeling experiments and accurate mass measurements. The five identified oxidized products, namely two isomeric alcohols (trans and cis WOH), two isomeric hydroperoxides (trans and cis WOOH), and N-formylkynurenine (FMK), were shown to share some common fragment ions and losses of small neutral molecules. Conversely, each oxidation product has its own fragmentation mechanism and intermediates, which were confirmed by (18)O-labeling studies. Isomeric WOH lost mainly H(2)O + CO, while WOOH showed preferential elimination of C(2)H(5)NO(3) by two distinct mechanisms. Differences in the spatial arrangement of the two isomeric WOHs led to differences in the intensities of the fragment ions. The same behavior was also found for trans and cis WOOH. FMK was shown to dissociate by a diverse range of mechanisms, with the loss of ammonia the most favored route. MS/MS analyses, (18)O-labeling, and H(2)(18)O experiments demonstrated the ability of FMK to exchange its oxygen atoms with water. Moreover, this approach also revealed that the carbonyl group has more pronounced oxygen exchange ability compared with the formyl group. The understanding of fragmentation mechanisms involved in O(2) ((1)Delta(g))-mediated oxidation of W provides a useful step toward the structural characterization of oxidized peptides and proteins. (J Am Soc Mass Spectrom 2009, 20, 188-197) (C) 2009 Published by Elsevier Inc. on behalf of American Society for Mass Spectrometry

DNA strand breaks and base modifications induced by cholesterol hydroperoxides

Cholesterol (Ch) can be oxidized by reactive oxygen species, forming oxidized products such as Ch hydroperoxides (ChOOH). These hydroperoxides can disseminate the peroxidative stress to other cell compartments. In this work, the ability of ChOOH to induce strand breaks and/or base modifications in a plasmid DNA model was evaluated. In addition, HPLC/MS/MS analyses were performed to investigate the formation of 8-oxo-7,8-dihydro-2`-deoxyguanosine (8-oxodGuo) after the incubation of 2`-deoxyguanosine (dGuo) with ChOOH and Cu(2+). In the presence of copper ions, ChOOH induced DNA strand breaks in time and concentration-dependent manners. Purine and pyrimidine base modifications were also observed, as assessed respectively by the treatment with Fpg and Endo III repair enzymes. The detection of 8-oxodGuo by HPLC/MS/MS is in agreement with the dGuo oxidation in plasmid DNA. ChOOH-derived DNA damage adds further support to the role of lipid peroxidation in inducing DNA modifications and mutation.

Detection and Characterization of Cholesterol-Oxidized Products Using HPLC Coupled to Dopant Assisted Atmospheric Pressure Photoionization Tandem Mass Spectrometry

Oxidation of cholesterol (Ch) by a variety of reactive oxygen species gives rise mainly to hydroperoxides and aldehydes. Despite the growing interest in Ch-oxidized products, the detection and characterization of these products is still a matter of concern. In this work, the main Ch-oxidized products, namely, 3 beta-hydroxycholest-5-ene-7 alpha-hydroperoxide (7 alpha-OOH), 3 beta-5 alpha-cholest-6-ene-5-hydroperoxide (5 alpha-OOH), 3 beta-hydroxycholest-4-ene-6 alpha-hydroperoxide (6 alpha-OOH), 3 beta-hydroxycholest-4-ene-6 beta-hydroperoxide (6 beta-OOH), and 3 beta-hydroxy-5 beta-hydroxy-B-norcholestane-6 beta-carboxaldehyde (ChAld), were detected in the same analysis using high-performance liquid chromatography (HPLC) coupled to dopant assisted atmospheric pressure photoionization tandem mass spectrometry. The use of selected reaction monitoring mode (SRM) allowed a sensitive detection of each oxidized product, while the enhanced product ion mode (EPI) helped to improve the confidence of the analyses. Isotopic labeling experiments enabled one to elucidate mechanistic features during fragmentation processes. The characteristic fragmentation pattern of Ch-oxidized products is the consecutive loss of 1120 molecules, yielding cationic fragments at m/z 401, 383, and 365. Homolytic scissions of the peroxide bond are also seen. With (18)O-labeling approach, it was possible to establish a fragmentation order for each isomer. The SRM transitions ratio along with EPI and (18)O-labeled experiments give detailed information about differences for water elimination, allowing a proper discrimination between the isomers:Phis is of special interest considering the emerging role of Ch-oxidized products in the development of diseases.

The antioxidative effect of de novo generated vitamin B-6 in Plasmodium falciparum validated by protein interference

The malaria parasite Plasmodium falciparum is able to synthesize de novo PLP (pyridoxal 5'-phosphate), the active form of vitamin B-6. In the present study, we have shown that the de novo synthesized PLP is used by the parasite to detoxify O-1(2) (singlet molecular oxygen), a highly destructive reactive oxygen species arising from haemoglobin digestion. The formation of O-1(2) and the response of the parasite were monitored by live-cell fluorescence microscopy, by transcription analysis and by determination of PLP levels in the parasite. Pull-down experiments of transgenic parasites overexpressing the vitamin B-6-biosynthetic enzymes PfPdx1 and PfPdx2 clearly demonstrated an interaction of the two proteins in vivo which results in an elevated PLP level from 12.5 mu M in wild-type parasites to 36.6 mu M in the PfPdx1/PfPdx2-overexpressing cells and thus to a higher tolerance towards O-1(2). In contrast, by applying the dominant-negative effect on the cellular level using inactive mutants of PfPdx1 and PfPdx2, P. falciparum becomes susceptible to O-1(2). Our results demonstrate clearly the crucial role of vitamin B-6 biosynthesis in the detoxification of O-1(2) in P falciparum. Besides the known role of PLP as a cofactor of many essential enzymes, this second important task of the vitamin B-6 de novo synthesis as antioxidant emphasizes the high potential of this pathway as a target of new anti-malarial drugs.

Oxidation of thiones by singlet and triplet oxygen

Molecular oxygen (012) i8 eatabliehed to be a good electrophile' and haabean Pound to yield many interesting moleculae upon reaction with olefinic, aromatic and other mu1 tipla bonded compounda. Although, oxidation of carbon ulphur double bond (thiones) by air her bean know for a longtime, nai the r the aechaniam nor the reactive species involved in theae oxidationa have bean etabliahodo Although there is no clear experimental verification, involvement of malecular oxygen in such types of oxidationa oP activated thiocarbonyl coc pounds has been recently auggeetad.4.

Singlet oxygen reacts with 2 `,7 `-dichlorodihydrofluorescein and contributes to the formation of 2 `,7 `-dichlorofluorescein

There are controversial reports in the literature concerning the reactivity of singlet oxygen ((1)O(2)) with the redox probe 2`,7`-dichlorodihydrofluorescein (DCFH). By carefully preparing solutions in which (1)O(2) is quantitatively generated in the presence of DCFH, we were able to show that the formation rate of the fluorescent molecule derived from DCFH oxidation, which is 2`,7`-dichlorofluorescein (DCF), increases in D(2)O and decreases in sodium azide, proving the direct role of (1)O(2) in this process. We have also prepared solutions in which either (1)O(2) or dication (MB(center dot 2+)) and semi-reduced (MB(center dot)) radicals of the sensitizer and subsequently super-oxide radical (O(2)(center dot-)) are generated. The absence of any effect of SOD and catalase ruled out the DCFH oxidation by O(2)(center dot-), indicating that both (1)O(2) and MB(center dot 2+) react with DCFH. Although the formation of DCF was 1 order of magnitude larger in the presence of MB(center dot 2+) than in the presence of (1)O(2), considering the rate of spontaneous decays of these species in aqueous solution, we were able to conclude that the reactivity of (1)O(2) with DCFH is actually larger than that of MB(center dot 2+). We conclude that DCFH can continue to be used as a probe to monitor general redox misbalance induced in biologic systems by oxidizing radicals and (1)O(2).

Singlet oxygen generation in the reaction centers of Rhodobacter sphaeroides

Singlet oxygen ((1)O(2)) generation in the reaction centers (RCs) of Rhodobacter sphaeroides wild type was characterized by luminescent emission in the near infrared region (time resolved transients and emission spectra) and quantified to have quantum yield of 0.03 +/- 0.005. (1)O(2) emission was measured as a function of temperature, ascorbate, urea and potassium ferricyanide concentrations and as a function of incubation time in H(2)O: D(2)O mixtures. (1)O(2) was shown to be affected by the RC dynamics and to originate from the reaction of molecular oxygen with two sources of triplets: photoactive dimer formed by singlet-triplet mixing and bacteriopheophytin formed by direct photoexcitation and intersystem crossing.

Generation and suppression of singlet oxygen in hair by photosensitization of melanin

We have studied the spectroscopic properties of hair (white, blond, red, brown, and black) under illumination with visible light, giving special emphasis to the photoinduced generation of singlet oxygen ((1)O(2)). Irradiation of hair shafts (lambda(ex)>400 nm) changed their properties by degrading the melanin. Formation of C3 hydroperoxides in the melanin indol groups was proven by (1)H NMR. After 532-nm excitation, all hair shafts presented the characteristic (1)O(2) emission (lambda(em) = 1270 nm), whose intensity varied inversely with the melanin content. (1)O(2) lifetime was also shown to vary with hair type, being five times shorter in black hair than in blond hair, indicating the role of melanin as a (1)O(2) suppressor. Lifetime ranged from tenths of a nanosecond to a few microseconds, which is much shorter than the lifetime expected for (1)O(2) in the solvents in which the hair shafts were suspended, indicating that (1)O(2) is generated and suppressed inside the hair structure. Both eumelanin and pheomelanin were shown to produce and to suppress (1)O(2), with similar efficiencies. The higher amount of (1)O(2) generated in blond hair and its longer lifetime is compatible with the stronger damage that light exposure causes in blond hair. We propose a model to explain the formation and suppression of (1)O(2) in hair by photosensitization of melanin with visible light and the deleterious effects that an excess of visible light may cause in hair and skin. 2011 Published by Elsevier Inc.

Implication of the oep16-1 mutation in a flu-independent, singlet oxygen-regulated cell death pathway in Arabidopsis thaliana

Singlet oxygen is a prominent form of reactive oxygen species in higher plants. It is easily formed from molecular oxygen by triplet–triplet interchange with excited porphyrin species. Evidence has been obtained from studies on the flu mutant of Arabidopsis thaliana of a genetically determined cell death pathway that involves differential changes at the transcriptome level. Here we report on a different cell death pathway that can be deduced from the analysis of oep16 mutants of A. thaliana. Pure lines of four independent OEP16-deficient mutants with different cell death properties were isolated. Two of the mutants overproduced free protochlorophyllide (Pchlide) in the dark because of defects in import of NADPH:Pchlide oxidoreductase A (pPORA) and died after illumination. The other two mutants avoided excess Pchlide accumulation. Using pulse labeling and polysome profiling studies we show that translation is a major site of cell death regulation in flu and oep16 plants. flu plants respond to photooxidative stress triggered by singlet oxygen by reprogramming their translation toward synthesis of key enzymes involved in jasmonic acid synthesis and stress proteins. In contrast, those oep16 mutants that were prone to photooxidative damage were unable to respond in this way. Together, our results show that translation is differentially affected in the flu and oep16 mutants in response to singlet oxygen.

Synthesis of FeCoMnAPO-5 molecular sieve and catalytic activity in cyclohexane oxidation by oxygen

A new FeCoMnAPO-5 with AFI structure was synthesized under hydrothermal conditions and characterized by XRD, FT-IR, X-ray fluorescence, nitrogen adsorption and SEM. The oxidation of cyclohexane with molecular oxygen was studied over the catalyst at 403 K. It show d higher activity compared to FeAPO-5, CoAPO-5 and MnAPO-5. The FeCoMnAPO-5 catalyst was recycled twice without loss of activity or selectivity.

Interaction Of Oxygen With Zn(0001) Surface

EELS and XPS studies show the presence of both adsorbed atomic and molecular oxygen at low temperatures. The nature of the oxide layer formed on the surface has been characterized by angular dependent and variable temperature EELS. A loss peak around 550 cm−1 is assigned to an electronic transition.

Position preference and diffusion path of an oxygen ion in apatite-type lanthanum silicate La9.33Si6O26: A density functional study

Using density functional theory, we investigated the position preference and diffusion mechanisms of interstitial oxygen ions in lanthanum silicate La9.33Si6O26, which is an apatite-structured oxide and a promising candidate electrolyte material for solid oxide fuel cells. The reported lanthanum vacancies were explicitly taken into account by theoretically determining their arrangement with a supercell model. The most stable structures and the formation energies of oxygen interstitials were determined for each charged state. It was found that the double-negatively charged state is stable over a wide range of the Fermi level, and that the excess oxygen ions form split interstitials with the original oxygen ions, while the neutral and the single-negatively charged states preferably form molecular oxygen. These species were found near the lanthanum vacancy site. The theoretically determined migration pathway along the c-axis essentially follows an interstitialcy mechanism. The obtained migration barrier is sensitive to the charge state, and is also affected by the lanthanum vacancy. The barrier height of the double-negatively charged state was calculated to be 0.58 eV for the model structure, which is consistent with the measured activation energy.

Electroreduction of oxygen on some novel cobalt phthalocyanine complexes

The catalytic activity of cobalt phthalocyanine monomer and some of its polymeric derivatives towards the electroreduction of molecular oxygen in salt and alkaline solutions is examined. It is found that most of these complexes exhibit a higher catalytic activity than the cobalt phthalocyanine monomer.

Effect of HCl on the adsorption of oxygen on Cu(110) surface: An eels study

Oxygen is shown to adsorb molecularly on a clean Cu(110) surface at 80 K and dissociate around 150 K forming atomic oxygen. Adsorption of oxygen on an HCl covered surface at low temperatures results in the formation of adsorbed hydroxyl groups and water in addition to adsorbed molecular oxygen. The molecular oxygen species is stable up to 190 K on the HCl covered surface.

Nafion®-bound carbon electrodes containing transitionmetal phthalocyanines for oxygen reduction in solid-polymer-electrolyte fuel cells

Catalytic activities of some transition metal-phthalocyanine complexes towards electroreduction of molecular oxygen are examined on Nafion®-bound and bare porous carbon electrodes in 2.5 M H2SO4 electrolyte. It is found that these metal complexes exhibit better catalytic activities towards oxygen reduction with the Nafion®-bound electrodes.