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PubMed1975

Hemoglobin function in the vertebrates: an evolutionary model.

Comparative data on quaternary structure, cooperativity, Bohr effect and regulation by organic phosphates are reviewed for vertebrate hemoglobins. A phylogeny of hemoglobin function in the vertebrates is deduced. It is proposed that from the monomeric hemoglobin of the common ancestor of vertebrates, a deoxy dimer, as seen in the lamprey, could have originated with a single amino acid substitution. The deoxy dimer has a Bohr effect, cooperativity and a reduced oxygen affinity compared to the monomer. One, or two, additional amino acid substitutions could have resulted in the origin of a tetrameric deoxy hemoglobin which dissociated to dimers on oxygenation. Gene duplication, giving incipient alpha and beta genes, probably preceded the origin of a tetrameric oxyhemoglobin. The origin of an organic phosphate binding site on the tetrameric hemoglobin of an early fish required only one, or two, amino acid substitutions. ATP was the first organic phosphate regulator of hemoglobin function. The binding of ATP by hemoglobin may have caused the original elevation in the concentration of ATP in the red blood cells by relieving end product inhibition of ATP synthesis. The switch from regulation of hemoglobin function by ATP to regulation by DPG may have been a consequence of the curtailment of oxidative phosphorylation in the red blood cell. The basic mechanisms by which ATP and DPG concentrations can respond to strss on the oxygen transport system were present before the origin of an organic phosphate binding site on hemoglobin. A switch from ATP regulation to IP5 regulation occurred in the common ancestor of birds.

Adenosine Triphosphate↗
PubMed1987

Spin trapping of oxygen and carbon-centered free radicals in ischemic canine myocardium.

Oxygen free radical injury has been postulated to occur during myocardial ischemia. We have used Electron Spin Resonance and Spin Trapping techniques to directly demonstrate the production of carbon-centered (R.) and oxygen-centered lipid radical (RO.) in ischemic canine heart. In addition, venous effluent from the ischemic region showed that conjugated dienes (lipid peroxidation products) increased with ischemic duration. Our results suggest that the formation of the oxygen-centered and carbon-centered lipid radical species during ischemia are a consequence of oxy-radical peroxidation of myocardial membrane lipids.

Animals↗
PubMed1998

Electron and light microscopy studies on particulate EPR spin probes lithium phthalocyanine, fusinite and synthetic chars.

This is the first detailed study on the morphology of the widely used particulate electron paramagnetic resonance (EPR) probes lithium phthalocyanine (LiPc), methyl-LiPc, methoxy-LiPc, fusinite and synthetic carbon-based chars, by means of both light and electronmicroscopy (LM and EM). The importance of these EPR probes for the measurement of O2/NO has been reported previously. Under LM, LiPc with its distinct crystalline structure differs significantly from the noncrystalline black flakes of both LiPc derivatives, methyl-LiPc and methoxy-LiPc. Unlike the shiny carbon-based synthetic chars, which have no characteristic morphology, fusinite, a fraction of fossilized coal maceral, displays distinctive fine, parallel channels. SEM studies reveal a striated surface and interlocking multilayered structure of LiPc that is markedly different from either the multilayered stacked methyl-LiPc or- botryoidal methoxy-LiPc. The regularly spaced pores and channels of fusinite, a reflection of its plant origin, contrast sharply with the randomly distributed pores of all sizes of the various synthetic chars. Furthermore, the combined results of both LM and EM studies strongly suggest the nonperturbing nature of fusinite and LiPc when they were used as EPR probes to measure oxygen in cells or tissues. We hope that this study, in conjunction with the numerous EPR functional studies of probes reported earlier, provides valuable information for the future development of new particulate EPR spin probes.

Animals↗
PubMed2004

Bioremediation of chlorobenzene-contaminated ground water in an in situ reactor mediated by hydrogen peroxide.

New in situ reactive barrier technologies were tested nearby a local aquifer in Bitterfeld, Saxonia-Anhalt, Germany, which is polluted mainly by chlorobenzene (CB), in concentrations up to 450 microM. A reactor filled with original aquifer sediment was designed for the microbiological remediation of the ground water by indigenous bacterial communities. Two remediation variants were examined: (a) the degradation of CB under anoxic conditions in the presence of nitrate; (b) the degradation of CB under mixed electron acceptor conditions (oxygen+nitrate) using hydrogen peroxide as the oxygen-releasing compound. Under anoxic conditions, no definite degradation of CB was observed. Adding hydrogen peroxide (2.94 mM) and nitrate (2 mM) led to the disappearance of CB (ca. 150 microM) in the lower part of the reactor, accompanied by a strong increase of the number of cultivable aerobic CB degrading bacteria in reactor water and sediment samples, indicating that CB was degraded mainly by productive bacterial metabolism. Several aerobic CB degrading bacteria, mostly belonging to the genera Pseudomonas and Rhodococcus, were isolated from reactor water and sediments. In laboratory experiments with reactor water, oxygen was rapidly released by hydrogen peroxide, whereas biotic-induced decomposition reactions of hydrogen peroxide were almost four times faster than abiotic-induced decomposition reactions. A clear chemical degradation of CB mediated by hydrogen peroxide was not observed. CB was also completely degraded in the reactor after reducing the hydrogen peroxide concentration to 880 microM. The CB degradation completely collapsed after reducing the hydrogen peroxide concentration to 440 microM. In the following, the hydrogen peroxide concentrations were increased again (to 880 microM, 2.94 mM, and 880 microM, respectively), but the oxygen demand for CB degradation was higher than observed before, indicating a shift in the bacterial population. During the whole experiment, nitrate was uniformly reduced during the flow path in the reactor.

Biodegradation, Environmental↗
PubMed2006

Reduction in air bubble size using perfluorocarbons during cardiopulmonary bypass in the rat.

BACKGROUND: Perfluorocarbon (PFC) emulsions are artificial oxygen-carrying compounds with a high solubility for gases that have experimentally been shown to ameliorate cerebral air embolism. Cerebral air embolism has been associated with adverse cerebral outcomes after cardiac surgery using cardiopulmonary bypass (CPB). We designed this study to test whether PFC emulsions could reduce the volume of bubbles within the CPB circuit. METHODS: Male Sprague-Dawley rats undergoing 60 min of normothermic nonpulsatile CPB were randomized to one of the three groups. The PFC group (n = 10) received 60% O(2)/36% N(2)/4% CO(2) via the membrane oxygenator and 2.7 g/kg (4.5 mL/kg) of PFC into the venous reservoir; the control group (n = 10) received the same gas mixture and 4.5 mL/kg of saline; the N(2)O group (n = 6) was exposed to 36% N(2)O/60% O(2)/4% CO(2) and received 4.5 mL/kg of saline. After 10 min and 35 min of CPB, 400 microL of air was injected into a bubble chamber in the CPB circuit. After 20 min, the bubble was removed for volumetric analysis. RESULTS: Compared with baseline, the bubble decreased 13% +/- 5% in size in the PFC group and increased 46% +/- 9% in the nitrous oxide group, both of these changes significantly different from the control group (P < 0.0001). CONCLUSION: The results suggest that PFC administration may be useful in reducing the volume of gaseous bubbles present during CPB.

Animals↗
PubMedDate not supplied

Mechanism of inhibiting effect of biologically active compounds on the active forms of oxygen reduction.

The impulse voltammetry methods with recording short-living radical-natured products of the reduction (in particular, hydroxyl radicals) are used to study the process of oxygen reduction steps. A possibility to study mechanisms of antioxidative action of biologically active compounds (BAC) through studying their influence on the separate steps of oxygen reduction (initial molecular oxygen, radical particles and peroxides) is shown. It provides a possibility for the purposeful search of BAC capable to affect certain steps of chain oxidation in biosystems.

Electrochemistry↗
PubMed2006

PET imaging of tumour hypoxia.

Tumour hypoxia represents a significant challenge to the curability of human tumours leading to treatment resistance and enhanced tumour progression. Tumour hypoxia can be detected by non-invasive and invasive techniques but the inter-relationships between these remains largely undefined. [18F]Fluoromisonidazole-3-fluoro-1-(2'-nitro-1'-imidazolyl)-2-propanol ([18F]MISO) and Cu-diacetyl-bis(N4-methylthiosemicarbazone (Cu-ATSM)-positron emission tomography (PET), and blood oxygen level-dependent (BOLD)-magnetic resonance imaging (MRI) are the lead contenders for human application based on their non-invasive nature, ease of use and robustness, measurement of hypoxia status, validity, ability to demonstrate heterogeneity and general availability; PET techniques are the primary focus of this review.

Cell Hypoxia↗
PubMed2006

Linking denitrifier community structure and prevalent biogeochemical parameters in the pelagial of the central Baltic Proper (Baltic Sea).

The oxic-anoxic interface of the water column of the Gotland Basin (central Baltic Sea) is characterised by defined biogeochemical gradients and is hypothesised to be a zone of pronounced denitrification. Our aim was to analyse the composition and distribution of pelagic denitrifying microorganisms in relation to the physico-chemical gradients in the water column. PCR-amplified nirS genes--coding for dissimilatory nitrite reductase--were analysed as functional markers by terminal restriction fragment length polymorphism and cloning. The overall nirS diversity was low, with the lowest levels found at the oxic-anoxic interface. Only a few terminal restriction fragments dominated the denitrifier communities throughout the water column, and these could be assigned to several new Baltic Sea clusters that were revealed by phylogenetic analysis. The novel clusters were separated in two groups corresponding to the oxygen concentrations within specific layers of the water column. Gradients of prevalent biogeochemical parameters (H(2)S, NH(4) (+), NO(3) (-) and O(2)) largely determined the composition of the nirS-type denitrifier communities within the water column of the Gotland Basin.

Bacteria↗
PubMed1988

Effect of hypoxia and acidosis on the cytotoxicity of four platinum complexes at normal and hyperthermic temperatures.

The cytotoxicities of cis-diamminedichloroplatinum(II) (CDDP) and of three recently developed dichloro complexes of bivalent platinum with radiosensitizing ligands [(1,2-diamino-4-nitrobenzene)dichloroplatinum(II) (Plato), trans-bis(2-amino-5-nitrothiazole)dichloroplatinum(II) (Plant), and trans-bis(2-nitroimidazole)dichloroplatinum(II) (NIPt)] were evaluated at 37 degrees C, 42 degrees C, and 43 degrees C at normal pH, at pH 6.45, and under normally oxygenated and hypoxic conditions in EMT6 cells in vitro. For CDDP, marked hyperthermic sensitization to the drug was evident in normally oxygenated cells, but hypoxic cells showed essentially no sensitization to the cytotoxicity of CDDP at elevated temperature at normal pH. Low pH further increased the cytotoxicity of CDDP toward normally oxygenated but not hypoxic cells at 37 degrees C and 42 degrees C. At 43 degrees C, however, low pH increased the cytotoxicity of CDDP toward both normally oxygenated and hypoxic cells, restoring nearly the full sensitizing effect of hyperthermia on CDDP cytotoxicity in the hypoxic cells. Plato was much more cytotoxic toward hypoxic than normally oxygenated cells under all culture conditions. At normal pH, hyperthermia increased the cytotoxicity of Plato in both hypoxic and normally oxygenated cells. At low pH, however, the cytotoxicity of Plato was inhibited at all temperatures and in both normally oxygenated and hypoxic cells. Plant was also more toxic to both normally oxygenated and hypoxic cells at elevated temperatures at normal pH. In contrast to Plato, however, Plant became much more cytotoxic toward hypoxic cells and showed increased cytotoxicity in normally oxygenated cells at low pH. Hyperthermia, however, did not further increase the rate of cell killing by Plant at low pH. NIPt, at the concentrations tested, was essentially nontoxic to cells at normal pH at 37 degrees C. Hyperthermia significantly increased the killing of hypoxic cells by NIPt under both normal and low pH conditions, but little cytotoxicity was noted for NIPt in normally oxygenated cells under any culture conditions. These results demonstrate that pH and the level of oxygenation of cells significantly affect the cytotoxicity of drugs at both normal and elevated temperatures. This sort of investigation may help delineate optimum drugs for use against environmentally determined subpopulations of cells within tumors.

Animals↗
PubMed1993

[Isopropoxyhermatran--a stimulant of hepatocyte regeneration].

It was shown that isoproxihermatran (IPH) (40 mg/kg) administered intraperitoneally in rats with lobectomy of heparin stimulates development of hepatocytes' dimensions and decrease of oxygen transmission velocity through the mitochondrial membranes in this cells. Antioxidant properties of IPH have been detected. Stimulating activity of IPH at heparin secretion of bile.

Animals↗
PubMed1996

Effects of pteridines on chloramine-T-induced growth inhibition in E. coli strains: correlations with molecular structure.

Little is known about the biological significance of most pteridines, despite their ubiquitous occurrence in living cells. Seventeen different pteridines were tested for their ability to modulate the growth inhibitory effect of the disinfectant chloramine-T on three different strains of Escherichia coli bacteria. We found striking differences between the pteridine derivatives: whereas aromatic pterins with a hydroxy function at side chain atom C2' increased the growth inhibition, those with a 7,8-dihydro structure exerted a suppressive effect. These results are in excellent agreement with previously observed effects of pteridine derivatives on chloramine-T-induced luminol-dependent chemiluminescence, and together, are highly suggestive of a general interaction of these compounds with oxygen or chlorine free radicals. This interaction is likely to have biological significance and might offer an explanation for the widespread occurrence of pteridines.

Anti-Infective Agents, Local↗
PubMed2004

Mechanisms of coupling between DNA recognition specificity and catalysis in EcoRI endonuclease.

Proteins that bind to specific sites on DNA often do so in order to carry out catalysis or specific protein-protein interaction while bound to the recognition site. Functional specificity is enhanced if this second function is coupled to correct DNA site recognition. To analyze the structural and energetic basis of coupling between recognition and catalysis in EcoRI endonuclease, we have studied stereospecific phosphorothioate (PS) or methylphosphonate (PMe) substitutions at the scissile phosphate GpAATTC or at the adjacent phosphate GApATTC in combination with molecular-dynamics simulations of the catalytic center with bound Mg2+. The results show the roles in catalysis of individual phosphoryl oxygens and of DNA distortion and suggest that a "crosstalk ring" in the complex couples recognition to catalysis and couples the two catalytic sites to each other.

Adenine Nucleotides↗
PubMed2004

Mechanistic studies on the formation and reactivity of dioxygen adducts of diiron complexes supported by sterically hindered carboxylates.

Dioxygen activation by enzymes such as methane monooxygenase, ribonucleotide reductase, and fatty acid desaturases occurs at a nonheme diiron active site supported by two histidines and four carboxylates, typically involving a (peroxo)diiron(III,III) intermediate in an early step of the catalytic cycle. Biomimetic tetracarboxylatodiiron(II,II) complexes with the familiar "paddlewheel" topology comprising sterically bulky o-dixylylbenzoate ligands with pyridine, 1-methylimidazole, or THF at apical sites readily react with O(2) to afford thermally labile peroxo intermediates that can be trapped and characterized spectroscopically at low temperatures (193 K). Cryogenic stopped-flow kinetic analysis of O(2) adduct formation carried out for the three complexes reveals that dioxygen binds to the diiron(II,II) center with concentration dependences and activation parameters indicative of a direct associative pathway. The pyridine and 1-methylimidazole intermediates decay by self-decomposition. However, the THF intermediate decays much faster by oxygen transfer to added PPh(3), the kinetics of which has been studied with double mixing experiments in a cryogenic stopped-flow apparatus. The results show that the decay of the THF intermediate is kinetically controlled by the dissociation of a THF ligand, a conclusion supported by the observation of saturation kinetic behavior with respect to PPh(3), inhibition by added THF, and invariant saturation rate constants for the oxidation of various phosphines. It is proposed that the proximity of the reducing substrate to the peroxide ligand on the diiron coordination sphere facilitates the oxygen-atom transfer. This unique investigation of the reaction of an O(2) adduct of a biomimetic tetracarboxylatodiiron(II,II) complex provides a synthetic precedent for understanding the electrophilic reactivity of like adducts in the active sties of nonheme diiron enzymes.

Carboxylic Acids↗
PubMed2006

Effect of nitric oxide release from NOR-3 on urea synthesis, viability and oxygen consumption of rat hepatocyte cultures.

As nitric oxide is considered a mediator of liver oxidative metabolism during sepsis, we studied the effects of exogenous nitric oxide, produced by NO-donor, (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (NOR-3), on cell viability, urea biosynthesis and oxygen consumption in rat hepatocyte cultures. Nitric oxide release from NOR-3 was studied using 4,5-diaminofluorescein diacetate. Urea levels were measured by the spectrophotometric method. Cell viability was determined by the MTT test and trypan blue exclusion test, whereas oxygen consumption was measured by a polarographic technique. After 2 h treatment, NOR-3 induced an increase in the levels of nitric oxide. After 2 h of treatment and 24 h after the end of the treatment with NOR-3, both cell viability and urea synthesis were significantly reduced in comparison to the controls for NOR-3 concentrations equal to or greater than 50 microM. A reduction in oxygen consumption was observed in hepatocytes after 40 min treatment with 100 microM NOR-3, even if the cell viability was unchanged. Reduction of oxygen consumption is an early indicator of the metabolic alterations in hepatocytes exposed to nitric oxide. These findings suggest that nitric oxide accumulation acts on hepatocyte cultures inducing cell death and reduction of urea synthesis after 2 hours.

Animals↗
PubMed2006

A novel luminescent lifetime-based optrode for the detection of gaseous and dissolved oxygen utilising a mixed ormosil matrix containing ruthenium (4,7-diphenyl-1,10-phenanthroline)3Cl2 (Ru.dpp).

A novel luminescent lifetime optrode is presented for the detection of gaseous and dissolved oxygen. The optrode utilises ruthenium (4,7-diphenyl-1,10-phenanthroline)(3)Cl(2) as the sensing fluorophore immobilised in a hydrophobic ormosil matrix. The ormosil matrix is synthesised at room temperature from octyltriethoxysilane and methyltriethoxysilane precursors. Investigations of different ormosils were conducted and the most effective one was selected for optrode production. Optrodes were tested for responses to gaseous and dissolved oxygen. Their responses were modelled using traditional two-site or two-exponential methods and feed-forward artificial neural networks. Comparison of the two modelling methodologies is presented and further improvements in modelling and ormosil design are suggested.

Chlorides↗
PubMed1990

Photodynamic action of sulphonated aluminium phthalocyanine (SALPC) on isolated rat pancreatic acini.

The photodynamic action of SALPC has been investigated on dispersed, perifused, acini isolated from the rat pancreas. Stimulation of secretion was assessed by measuring amylase release and membrane permeabilization determined by the leakage of cytoplasmic lactate dehydrogenase (LDH) and by the efflux of 86Rb from preloaded acini. Light alone (greater than 570 nm, less than or equal to 18,400 lux), or SALPC (less than or equal to 1 microM) in the absence of light, had no effect on pancreatic acini but cellularly bound SALPC when illuminated caused a dose-dependent, light intensity-dependent and temperature-dependent release of amylase. Singlet oxygen generated by photon-activation of SALPC was measured by the formation of an imidazole adduct and bleaching of the secondary substrate, RNO. Whereas illumination caused a rapid increase in photodynamically-evoked pancreatic amylase release, the efflux of 86Rb and loss of cytosolic LDH were markedly delayed in onset: similar results were obtained with monochromatic laser light (633 nm). In contrast, the muscarinic agonist bethanechol evoked a rapid increase in amylase release but with an almost immediate efflux of 86Rb. Finally, electron microscopy confirmed that the structural integrity of the pancreatic acinar cells was maintained after the photodynamic action of SALPC. It is concluded that the stimulation of amylase secretion and membrane permeabilization by SALPC is due to the generation of singlet oxygen. However, the consistent difference between the time course of amylase secretion and membrane permeabilization makes it likely that an initial stage in photodynamic drug action involves oxidation of plasma membrane protein and activation of secretagogue receptors or the G-proteins and their effector systems.

Amylases↗
PubMed1981

Carbon monoxide- and oxygen-reacting haemoproteins in the mitochondrial fraction from the soil amoeba Acanthamoeba castellanii. Studies at subzero temperatures.

1. Mitochondria-enriched fractions of the soil amoeba Acanthamoeba castellanii contained four haemoproteins that in their reduced forms reacted with CO to give photodissociable CO complexes; these were cytochromes a 3, a 614, b- and c-type cytochromes. 2. Non-photodissociable oxygen-containing compounds were formed at temperatures between -130 and -150 degrees C after photodissociation of CO in the presence of 200 microM-O2, 3. Electron transport, indicated by the oxidation of cytochromes a + a3 and cytochrome c, did not occur until the temperature was raised to -80 degrees C.

Amoeba↗
PubMedDate not supplied

Polyelectrolyte microshells as carriers for fluorescent sensors: loading and sensing properties of a ruthenium-based oxygen indicator.

A strategy for the design and fabrication of microcapsule-based fluorescent biosensors containing indicators and internal references is described. The rationale for this work is the physical immobilization and chemical separation of assay chemistry for use in biological environments. Using the general approach of depositing oppositely charged species on colloidal micro/nanotemplates, a sensor system employing polyelectrolyte microshells for uptake of functional molecules is proposed, and experiments to demonstrate the feasibility of nanoengineering the sensor properties are described in the context of an oxygen sensor. Methods for immobilization and entrapment of fluorescent indicator and reference dyes are shown, along with the pH dependence of this process. Embedded dyes are shown to be stable and retain their function, as demonstrated with oxygen-sensitivity experiments of loaded microcapsules. Although oxygen sensitivity is presented as an example of a specific application, the overall strategy is likely more generally useful. The work suggests that polyelectrolyte microshells may be used as a platform to develop novel sensors by entrapment of functional materials.

Biosensing Techniques↗