Further evidence for double-strand breaks originating from a paired radical precursor from studies of oxygen fixation processes

By using a fast reaction technique which employs H2S gas as a fast-reacting chemical repair agent, it is possible to measure the competition kinetics between chemical repair reactions and oxygen fixation reactions in model DNA and cellular systems. In plasmid pBR322 DNA irradiated with electrons, we have compared the oxygen fixation reactions of the free radical precursors that lead to the production of single-strand (SSBs) and double-strand breaks (DSBs). For the oxygen-dependent fixation of radical damage leading to SSBs, a second-order rate constant of 2.3 x 10(8) dm(3) mol(-1) s(-1) was obtained compared to 8.9 x 10(7) dm(3) mol(-1) s(-1) for DSBs. The difference is in general agreement with predictions from a multiple-radical model where the precursor of a DSB originates from two radicals. The fixation of this precursor by oxygen will require both radicals to be fixed for the DSB to be formed, which will have slower kinetics than that of single free-radical precursors of SSBs. (C) 1999 by Radiation Research Society.

Plasma parameter characterization of a reflex plasma source operating in oxygen

A reflex discharge plasma, obtained as a hybrid between a Penning discharge plasma (PDP) and a hollow-cathode discharge (HCD) plasma, is analysed as a possible direction-current, high-density plasma source. The experiment is run in oxygen at pressures of 10 mTorr and 1 mTorr, and for discharge currents of 100 to 200 mA. Although the gas pressure is considerably lower than those used in HCDs, the hollow-cathode effect (HCE) occurs for current levels higher than 100 mA and leads to plasma densities comparable with those obtained using inductive plasma sources. The presence of a constant magnetic field leads to the enhancement of electron emission from cathodes under ion bombardment, and to the decreasing of the ion loss by diffusion to the wall.

THE ROLE OF GAS-PHASE OXIDATION AND COMBINATION DURING LASER DEPOSITION OF YBA2CU3O7-X IN AMBIENT OXYGEN

Spectroscopic absorption and emission measurements have been used to study laser deposition of YBCO films. They show that >95% of the monatomic Y and Ba initially ablated from the target undergo gas-phase chemical combination before film deposition. In contrast, considerable monatomic Cu persists into the deposition region. in this region, equilibrated gas temperatures are of the order of 2700 K. It is suggested that this high temperature facilitates film crystallization and epitaxial growth. The survival of monatomic Cu in the plume to the site of deposition is a manifestation of its endothermic reaction with O-2.

In vitro resistance of Burkholderia cepacia complex isolates to reactive oxygen species in relation to catalase and superoxide dismutase production

The Burkholderia cepacia complex comprises groups of genomovars (genotypically distinct strains with very similar phenotypes) that have emerged as important opportunistic pathogens in cystic fibrosis (CF) patients. The inflammatory response against bacteria in the airways of CF individuals is dominated by polymorphonuclear cells and involves the generation of oxidative stress, which leads to further inflammation and tissue damage. Bacterial catalase, catalase-peroxidase and superoxide dismutase activities may contribute to the survival of B. cepacia following exposure to reactive oxygen metabolites generated by host cells in response to infection. In the present study the authors investigated the production of catalase, peroxidase and SOD by isolates belonging to various genomovars of the B. cepacia complex. Production of both catalase and SOD was maximal during late stationary phase in almost all isolates examined. Native PAGE identified 13 catalase electrophoretotypes and two SOD electrophoretotypes (corresponding to an Fe-SOD class) in strains belonging to the six genomovars of the B. cepacia complex. Seven out of 11 strains displaying high-level survival after H(2)O(2) treatment in vitro had a bifunctional catalase/peroxidase, and included all the genomovar III strains examined. These isolates represent most of the epidemic isolates that are often associated with the cepacia syndrome. The majority of the isolates from all the genomovars were resistant to extracellular O(-)(2), while resistance to intracellularly generated O(-)(2)was highly variable and could not be correlated with the detected levels of SOD activity. Altogether the results suggest that resistance to toxic oxygen metabolites from extracellular sources may be a factor involved in the persistence of B. cepacia in the airways of CF individuals.

Solubility of carbon dioxide, ethane, methane, oxygen, nitrogen, hydrogen, argon, and carbon monoxide in 1-butyl-3-methylimidazolium tetrafluoroborate between temperatures 283 K and 343 K and at pressures close to atmospheric

Experimental values for the solubility of carbon dioxide, ethane, methane, oxygen, nitrogen, hydrogen, argon and carbon monoxide in 1-butyl-3- methylimidazolium tetrafluoroborate, [bmim][BF4] - a room temperature ionic liquid - are reported as a function of temperature between 283 K and 343 K and at pressures close to atmospheric. Carbon dioxide is the most soluble gas with mole fraction solubilities of the order of 10-2. Ethane and methane are one order of magnitude more soluble than the other five gases that have mole fraction solubilities of the order of 10-4. Hydrogen is the less soluble of the gaseous solutes studied. From the variation of solubility, expressed as Henry's law constants, with temperature, the partial molar thermodynamic functions of solvation such as the standard Gibbs energy, the enthalpy, and the entropy are calculated. The precision of the experimental data, considered as the average absolute deviation of the Henry's law constants from appropriate smoothing equations is of 1%. © 2005 Elsevier Ltd. All rights reserved.

Regulation of erythropoietin gene expression depends on two different oxygen-sensing mechanisms

Erythropoietin (Epo), a glycoprotein hormone produced principally in the fetal kidney and in the adult liver in response to hypoxia, is the prime regulator of growth and differentiation in erythroid progenitor cells. The regulation of Epo gene expression is not fully understood, but two mechanisms have been proposed. One involves the participation of a heme protein capable of reversible oxygenation and the other depends on the intracellular concentration of reactive oxygen species (ROS), assumed to be a function of pO2. We have investigated the production of Epo in response to three stimuli, hypoxia, cobalt chloride, and the iron chelator desferrioxamine, in Hep3B cells. As expected, hypoxia caused a marked rise in Epo production. When the cells were exposed to the paired stimuli of hypoxia and cobalt no further increase was found. In contrast, chelation of iron under hypoxic conditions markedly enhanced Epo production, suggesting that the two stimuli act by separate pathways. The addition of carbon monoxide inhibited hypoxia-induced Epo production, independent of desferrioxamine concentration. Taken together these data support the concept that pO2 and ROS are sensed independently.

Erythropoietin production:evidence for multiple oxygen sensing pathways

The production of erythropoietin (Epo), the glycoprotein hormone which controls red blood cell formation, is regulated by feedback mechanisms sensing tissue oxygenation. The mechanism of the putative oxygen sensor has yet to be elucidated. There is evidence that at least two pathways participate in hypoxia signal transduction. One appears to involve a specific haem protein, and a second implicates reactive oxygen species (ROS). Iron catalyses the generation of intracellular ROS and therefore alters the cellular redox state. We have investigated the effect of modulating intracellular iron content on Epo production in Hep 3B cells. Iron chelation stimulates Epo production at 20% O2 and enhances Epo production at 1% O2, but it has no additive effect on cobalt-induced Epo production. Excess molar iron inhibited Epo production in response to hypoxia, desferrioxamine (DFO) and cobalt chloride and inhibited the DFO-enhancing effect of hypoxia-induced Epo production. We found that sulphydryl oxidising agents exert a differential inhibitory effect on hypoxia-induced versus DFO-induced Epo production, providing further evidence that multiple pathways of oxygen sensing exist.

Can we trust mass spectrometry for determination of arsenic peptides in plants:comparison of LC-ICP-MS and LC-ES-MS/ICP-MS with XANES/EXAFS in analysis of Thunbergia alata

The weakest step in the analytical procedure for speciation analysis is extraction from a biological material into an aqueous solution which undergoes HPLC separation and then simultaneous online detection by elemental and molecular mass spectrometry (ICP-MS/ES-MS). This paper describes a study to determine the speciation of arsenic and, in particular, the arsenite phytochelatin complexes in the root from an ornamental garden plant Thunbergia alata exposed to 1 mg As L(-1) as arsenate. The approach of formic acid extraction followed by HPLC-ES-MS/ICP-MS identified different As(III)-PC complexes in the extract of this plant and made their quantification via sulfur (m/z 32) and arsenic (m/z 75) possible. Although sulfur sensitivity could be significantly increased when xenon was used as collision gas in ICP-qMS, or when HR-ICP-MS was used in medium resolution, the As:S ratio gave misleading results in the identification of As(III)-PC complexes due to the relatively low resolution of the chromatography system in relation to the variety of As-peptides in plants. Hence only the parallel use of ES-MS/ICP-MS was able to prove the occurrence of such arsenite phytochelatin complexes. Between 55 and 64% of the arsenic was bound to the sulfur of peptides mainly as As(III)(PC(2))(2), As(III)(PC(3)) and As(III)(PC(4)). XANES (X-ray absorption near-edge spectroscopy) measurement, using the freshly exposed plant root directly, confirmed that most of the arsenic is trivalent and binds to S of peptides (53% As-S) while 38% occurred as arsenite and only 9% unchanged as arsenate. EXAFS data confirmed that As-S and As-O bonds occur in the plants. This study confirms, for the first time, that As-peptides can be extracted by formic acid and chromatographically separated on a reversed-phase column without significant decomposition or de-novo synthesis during the extraction step.

Age-related changes in non-receptor dependent generation of reactive oxygen species from phagocytes of healthy adults

Several authors have shown that neutrophil generation of reactive oxygen species (ROS) declines with advancing age. Similar changes have also been suggested in monocytes. In both cases alterations in second messenger activity have been implicated as the most likely explanation for these observations. The aim of this study was to investigate the effect of age on phagocyte ROS generation, stimulated by the direct activation of protein kinase C (PKC). Venous blood was drawn from normal healthy subjects, cells were separated on a double density gradient into mononuclear and polymorphonuclear (pmn) cells. Phorbol myristate acetate (PMA) was employed as a cell stimulus. Superoxide generation was measured by cytochrome c reduction and myeloperoxidase (MPO) products by measurement of peak luminol chemiluminescence (CL). Fifty-eight subjects, 25 males and 33 females, were studied, median age 49 years (range 26-88 years). Polymorphonuclear cell superoxide generation was significantly higher in males and there was a trend towards higher pmn MPO product generation in males. Using Spearman's ranked correlation coefficient, monocyte superoxide generation was negatively correlated with age (r = -0.473, P <0.001). No changes in the generation of MPO products was found. There were also trends towards a negative correlation of pmn cytochrome c reduction and peak luminol CL with age in males but not females. Since PMA directly activates protein kinase C, reduced monocyte superoxide generation with increasing age appears to be related to alterations in the ROS generating system downstream of the cell receptor. Impaired monocyte superoxide generation may have implications for non-specific defence against certain infections and early tumour growth in the elderly. Factors underlying these changes in monocyte function therefore require further study.