Litter-decay fungi have recently been shown to secrete heme-thiolate peroxygenases that oxidize various organic chemicals, but little is known about the physiological role or the mechanism of these enzymes. The aromatic peroxygenase of Agrocybe aegerita (AaeAPO) was purified and catalytically characterized. An overall reaction mechanism was proposed. The results show that AaeAPO catalyzed diverse H2O2-dependent monooxygenations (two-electron oxidations) including (a) the cleavage of aliphatic and aromatic ethers, (b) the regio- and enantioselective hydroxylation of aromatic compounds, (c) the stepwise oxygenation of benzylic compounds, (d) the N-dealkylation of secondary amines and (e) the dehalogenation of halogenated aliphatic compounds as well as typical peroxidase reactions (suggested to involve one-electron oxidation) such as (f) oxidation and polymerization of phenols and (g) halogenations. The enzyme failed to oxidize polymers such as polyethylene glycol (PEG). Mechanistic studies with several model substrates provided information about the reaction cycle of AaeAPO: (1) stoichiometry of tetrahydrofuran cleavage showed that the reaction was a two-electron oxidation that generated one aldehyde group and one alcohol group, yielding the ring-opened product 4-hydroxybutanal, (2) steady-state kinetics results with methyl 3,4-dimethoxybenzyl ether, which was oxidized to 3,4-dimethoxybenzaldehyde, gave parallel double reciprocal plots suggestive of a ping-pong mechanism, (3) the cleavage of methyl 4-nitrobenzyl ether, the hydroxylation of aromatics such as diclofenac and nitrophenol and the oxygenation of benzylic compounds, resulted in incorporation of 18O into the reaction product in the presence of H218O2, and (4) the demethylation of 1-methoxy-4-trideuteromethoxybenzene showed an distinct observed intramolecular deuterium isotope effect. These results support a mechanism similar to that envisaged for the peroxygenase activity of P450s in which the enzyme heme is oxidized by H2O2 to give an iron species that carries one of the peroxide oxygen. This intermediate then abstracts a hydrogen from the substrate, which is followed by rebound of an •OH equivalent to produce the monooxygenated reaction product (hydrogen abstraction and oxygen rebound mechanism). AaeAPO may accordingly have a role in the biodegradation of natural and anthropogenic low molecular weight compounds in soils and plant litter. Moreover, the results raise the possibility that fungal peroxygenases may be useful for versatile, cost-effective, and scalable syntheses of drug metabolites and herbicide precursors. / Die Peroxygenase des Südlichen Ackerling (Agrocybe aegerita, AaeAPO) wurde gereinigt, ihr Katalysepotential ermittelt und ein allgemeiner Reaktionsmechanismus postuliert. Die AaeAPO katalysiert sowohl H2O2-abhängige Monooxygenierungen (Zwei-Elektron Oxidationen) wie (a) die Spaltung aliphatischer und aromatischer Ether, (b) die regio- und enantioselektive Hydroxylierung von Aromaten, (c) die schrittweise Monooxygenierung von Toluolderivaten, (d) die N-Dealkylierung sekundärer Amine und (e) die Dehalogenierung chlorierter Aliphaten als auch typische Reaktionen bekannter Peroxidasen (vermutlich Ein-Elektron-Oxidation) unter anderem (f) die Oxidation/ Polymerisierung von Phenolen und (g) die Halogenierung von Aromaten. Polymere Verbindungen wie Polyethylenglycol (PEG) werden nicht oxidiert. Mechanistische Untersuchungen zur Etherspaltung am Beispiel der AaeAPO haben Einblick in den generellen Reaktionsmechanismus dieses neuen Enzymtyps ermöglicht: (1) die Stöchiometrie der Spaltung von Tetrahydrofuran entspricht der einer zwei-Elektron-Oxidation, (2) die Spaltung von Methyl-3,4-Dimethoxybenzylether zu 4-Dimethoxybenzaldehyd und Methanol ergaben parallele Verläufe für die ermittelten Ausgleichsgeraden in der doppelt reziproken Darstellung, was einem „Ping-Pong“-Reaktionsmechanismus entspricht (3) die Monooxygenierungen haben stets den Einbau eines aus dem Peroxid (H2O2) stammenden Sauerstoffatoms in das Produkt zur Folge, (4) die O-Dealkylierung von 1-Methoxy-4-Trideuterummethoxybenzol zeigt einen ausgeprägten Deuterium Isotopen Effekt, was auf die primäre Abspaltung eines Wasserstoffatoms vom Substratmolekül hindeutet. Demnach verläuft die Peroxygenase-katalysierte Monooxygenierung über Wasserstoffabstraktion und eine unmittelbar anschließende Sauerstoffrückbindung (hydrogen abstraction - oxygen rebound mechanism). Diese Reaktionsabfolge ähnelt dem sogenannten peroxide "shunt" pathway, der von einer Reihe Cytochrom-P450-abhängiger Monooxygenasen her bekannt ist. Die physiologische Funktion der AaeAPO besteht möglicherweise in der extrazellulären Transformation und Detoxifikation niedermolekularer Pflanzeninhaltsstoffe, mikrobieller Metabolite und anthropogener Xenobiotika. Aufgrund der Stabilität und Unabhängigkeit der AaeAPO von teuren Kofaktoren ergeben sich vielversprechende biotechnologische Möglichkeiten zum Einsatz isolierter Biokatalysatoren in selektiven (bio)chemischen Synthesen monooxygenierter Metabolite.

This thesis investigates how benthic fluxes of phosphorus (P), nitrogen (N), and silicon (Si) change upon oxygenation of anoxic soft bottoms in the brackish, eutrophicated Baltic Sea. Direct measurements in situ by benthic landers demonstrated that fluxes of dissolved inorganic P (DIP) from anoxic bottom sediments in the Eastern Gotland Basin are higher than previously thought (Paper I). It is argued that the benthic DIP flux has a much larger influence on the DIP inventory in the Baltic proper than the external sources. Similarly, benthic fluxes of DIP and dissolved inorganic N (DIN) from anoxic sediment in the coastal Kanholmsfjärden Basin, Stockholm archipelago, were sufficiently high to renew the pools of these nutrients below the upper mixed layer in roughly one year (Paper II). A natural inflow of oxygen rich water into the deep, and previously long-term anoxic part of Kanholmsfjärden Basin, increased the P content in the sediment by 65% and lowered DIP and dissolved silica (DSi) concentrations in the pore water. These changes, as well as the large increases in benthic effluxes of these solutes following de-oxygenation of the bottom water, suggest that they are influenced similarly by changing oxygen conditions. Experimental results in papers III and IV show that common benthic macrofauna species in the Baltic Sea can stimulate benthic release of DIN and DSi, as well as dissolved organic and particulate bound nutrients. Thus, if benthic oxygen conditions would improve in the Baltic, initial effects on benthic–pelagic nutrient coupling will change due to animal colonisation of currently azoic soft bottoms. A new box corer was designed (Paper V) which can be used to obtain highly needed virtually undisturbed samples from soft bottom sediments – if lowered slowly and straight into the bottom strata – as demonstrated by in situ videography and turbidimetry. The commonly used USNEL box corer caused severe biasing during sediment collection. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: Manuscript. Paper 4: Manuscript. Paper 5: Manuscript.</p> / Baltic oxygenation project

Hollow fiber membranes are widely used in blood oxygenators to remove carbon dioxide and add oxygen during cardiopulmonary bypass operations. These devices are now widely used off-label by physicians to perform extracorporeal blood oxygenation for patients with lung failure. Unfortunately, the hollow fiber membranes used in these devices fail prematurely due to blood plasma leakage and gas emboli formation. This project formed ultrathin (~100nm) polymer coatings on polymer hollow fiber membranes. The coatings were intended to “block” existing pores on the exterior surfaces while permitting high gas fluxes. This coating is synthesized using surface imitated control radical polymerization. The coating was durable and did not peel or degrade. Fibers modified using this coating technique did not substantially degrade the mechanical properties of the membrane. This coating technique prevented blood plasma leakage and gas emboli formation. The coating permitted blood oxygenation and carbon dioxide removal from in a mock circulation module. Coating formation on polymeric hollow fiber membranes using surface initiated controlled radical polymerization allows for the formation of membranes that have the potential for long term blood oxygenation. This coating technique would allow these long term blood membranes to be produced more inexpensively than currently existing membranes used for long term use.

Relative changes are detectable in the blood of end-stage renal disease (ESRD) patients during hemodialysis (HD) treatment using optical spectroscopy. However, the potential impacts of several confounding factors that could affect the detection of these changes have not been evaluated. The objectives of this thesis were to: 1) investigate how the variations and/or changes in acid-base and oxygen parameters during HD treatment can affect the optical signature of whole blood of ESRD patients, 2) to investigate the effect of heparin on the optical properties of whole blood and its impact on our method. Blood samples were drawn from 23 ESRD patients at 5 time points during a 4 hour HD treatment and sent for blood gas and blood spectroscopy analyses. No significant correlations were found between the changes in the blood transmittance spectra and acid-base and oxygen parameters. This indicates that the perturbations in these parameters due to HD procedures do not confound the detection of changes in the blood transmittance spectra of ESRD patients during HD treatment. Additionally, the effect of heparin in modifying the optical properties of whole blood does not confound the detection of changes in the blood of ESRD patients due to HD treatment using whole blood-based optical spectroscopy. ANOVA revealed significant (P<0.05) measurable changes in the blood transmittance spectra of ESRD patients during HD treatment. Significant spectral differences (P<0.05) were found between ESRD patients. The lack of uniform spectral characteristics across patients is

Doctor of Philosophy / In this thesis, three experiments were conducted to monitor: (i) muscle oxygenation and electromyographic activity of the biceps brachii after exercise-induced muscle damage (ii) muscle oxygenation after downhill walking-induced muscle damage, and, (iii) muscle oxygenation following a bout of vigorous concentric exercise. Maximal eccentric exercise (EE) of biceps brachii resulted in significantly increased mean resting oxygen saturation and decreased deoxyhaemoglobin. During isometric contractions at 50% and 80% of subjects’ maximum voluntary torque (MVT), oxygen desaturation and resaturation kinetics and volume were significantly decreased after EE, and these declines were significantly prevalent over the following 6 days. Additionally, a significant shift in median frequency intercept (measured by electromyography; EMG) towards lower frequencies was observed during isometric contractions at both 50% and 80% MVT after EE in the exercised arm. After an exhaustive session of downhill walking, another form of EE, resting total haemoglobin and oxyhaemoglobin decreased. Furthermore, during isometric contractions at 30%, 50% and 80% of MVT, prolonged and significant increases were observed in oxygen desaturation and resaturation kinetics and volumes after ambulatory EE. In contrast to the two EE experiments, concentric contractions did not evoke any prolonged changes in muscle oxygenation. Collectively, the findings of this thesis revealed significant and prolonged changes in muscle oxygenation at rest and during exercise, following sessions of strenuous eccentric exercise. Although not clear, the possible mechanism responsible for the changes in muscle oxygenation after EE could be increased resting muscle oxygen utilization due to probable muscle damage and a subsequent requirement of energy demanding repair processes. Concentric exercise resulted in fatigue, but it did not affect muscle oxygenation. Although a prolonged reduction in EMG median frequency intercept was observed after EE, this was not closely time-associated with the biochemical, anthropometric or functional markers of muscle damage.

The purpose of this study was to compare the exercise and recovery muscle oxygenation response of Women and Girls during two 30s Wingate anaerobic tests separated by two minutes of active cycling recovery (resistance ≈ 2.5% body weight, 60-80rpm). Oxygenated hemoglobin (HbO2), deoxygenated hemoglobin (HHb), total hemoglobin (tHb), and tissue saturation index (TSI) were monitored at the right vastus lateralis muscle using near infrared spectroscopy (NIRS) throughout exercise, recovery, and a post-exercise femoral artery occlusion to TSI plateau. Pressure was preset at 250mmHg for Women and 210mmHg for Girls, achieved by rapid inflation in 0.3 seconds, and maintained until a 2 minute TSI plateau occurred or 10 minutes had passed. Twenty Women (23.8[2.12] years) and 13 Girls (9[1] years, combined Tanner stage <4) completed all tasks excepting 1 girl who did not complete occlusion. Significant group, time, and group by time interaction effects were observed for peak and mean power (Watts.kgFFM-1). Women had significantly greater power output compared to Girls for both Wingates. While both groups had reduced power output in Wingate 2, the reduction was significantly greater in Women compared to Girls. No significant group differences were found for resting TSI, recovery TSI, minimum TSI during either Wingate test, or for minimum TSI during occlusion, however a time main effect for Women was observed with minimum TSI being significantly lower in Wingate 1 compared to Wingate 2. Girls had similar minimum TSI for both Wingate tests. Women also demonstrated a significantly greater difference between Wingate minimum TSI and occlusion minimum TSI in Wingate 2 compared to Wingate 1. During Wingate 1, HHb increase was greater in Girls compared to Women and remained elevated during recovery compared to women. Changes in HbO2, HHb, and tHb were reduced in Wingate 2 for both groups, more so in Women for tHb and in Girls for HHb. Recovery was not different between groups with the exception of a faster TSI time constant of recovery in Women (τ =20.25 [13.01]s) compared to Girls (τ =36.77 [13.38]s) which is attributed to a faster HHb time constant in Women (τ =13.6 [0.44]s) compared to Girls (τ =30.77[19.47]s). Both groups demonstrated similar power output results and TSI response across the two Wingate tests but Girls were better able to repeat the anaerobic performance with a consistent TSI minimum between the two tests despite a faster recovery of HHb and TSI in women. These findings, in the context of observed Hb variable differences between groups, provide evidence of greater oxidative metabolism in Girls during a high intensity exercise. / Graduate / December 18, 2016

Ce travail méthodologique a porté sur la mise en place et l'évaluation d'une technique de cartographie de l'oxygénation cérébrale en imagerie par résonance magnétique (IRM) à 3 T. La technique repose sur la quantification de signaux IRM issus du contraste BOLD (pour « blood level oxygen dependent ») à l'aide d'une approche multiparamétrique (approche mqBOLD pour multiparametric quantitative BOLD) et permet d'obtenir une estimation de la saturation tissulaire en oxygène (StO2). Un premier travail de simulation numérique a porté sur la prise en compte du biais induit par la susceptibilité magnétique de la myéline dans la mesure de StO2. Ce travail a permis de montrer l'influence de l'orientation des fibres de substance blanche par rapport au champ magnétique principal sur la mesure de StO2. La méthode mqBOLD a tout d'abord été appliquée chez l'animal, sur le primate non-humain. L'approche semble suffisamment précise pour observer des différences inter-espèces. Puis, cette approche a été adaptée pour une utilisation en clinique et appliquée à différentes pathologies. Chez des patients souffrant d'un accident vasculaire cérébral à la phase aiguë, nous avons observé une différence spatiale entre les régions pour lesquelles le débit sanguin et la StO2 sont abaissés, différence qui répondrait aux critères définissant la pénombre ischémique. Chez des patients atteints d'une sténose intracrânienne artérielle sévère, nous avons étudié la relation entre la mesure de la vasoréactivité cérébrale avec celle de l'oxygénation. Les résultats montrent une corrélation entre l'altération de la vasoréactivité et celle de l'oxygénation dans le territoire de l'artère cérébrale moyenne. Enfin, nous montrons au travers d'une analyse par cluster que l'étude de l'oxygénation apporte des informations complémentaires aux paramètres de perfusion dans la détermination du grade tumoral. L'ensemble des résultats obtenus au cours de ce travail démontrent la faisabilité de la cartographie de StO2 par IRM et la complémentarité de cette approche avec les techniques de caractérisation de perfusion tissulaire. / This work focuses on the implementation and the evaluation of a technique to map cerebral oxygenation by MRI at 3T. The technique is based on the quantification of MRI signals from the BOLD contrast (for "blood oxygen level dependent") using a multiparametric (mqBOLD multiparametric approach for quantitative BOLD) and provides an estimate of the tissue oxygen saturation (StO2). A study based on a numerical simulation focused on taking into account the bias induced by the magnetic susceptibility of myelin in the measurement of StO2. This work demonstrated the influence of the orientation of white matter fibers with respect to the main magnetic field on the measurement of StO2. The mqBOLD method was applied in animals, the non-human primate. The approach seemed accurate enough to observe differences between species. Then, this approach was adapted for clinical practice and applied to different pathologies. In acute stroke patients, we observed a spatial difference between the regions with lowered cerebral blood flow and StO2, a difference that could meet the criteria of the ischemic penumbra. In patients with severe intracranial arterial stenosis, we studied the relationship between an estimate of cerebral vasoreactivity and tissue oxygen saturation. The results showed a correlation between impaired vascular reactivity and the oxygenation in the territory of the middle cerebral artery. Finally, we showed, through a cluster analysis, that the study of oxygenation provides additional information to the perfusion parameters and could be useful to determine the tumor grade. The overall results obtained in this thesis demonstrate the feasibility of mapping StO2 by MRI and the complementarity of this approach with techniques for characterizing tissue perfusion.

No description available.

L’apport d’oxygène joue un rôle prépondérant dans le développement d’un incendie en milieu confiné. La quantité d’oxygène disponible pour la combustion va dépendre de son appauvrissement dû au feu mais aussi de l’apport d’air par le système de ventilation et / ou les ouvertures. Une baisse du niveau d’oxygène du mélange oxydant va conduire à une diminution du flux de chaleur provenant de la flamme vers la surface du combustible, ce qui, en retour, va entraîner une diminution du débit de pyrolyse. La présente étude expérimentale a un double objectif : comprendre les effets d’une atmosphère appauvrie en oxygène sur la combustion de matériaux solides et liquides et fournir des données nécessaires à la validation des outils de simulation numérique d’un incendie. Les essais expérimentaux ont été menés dans le calorimètre à atmosphère contrôlée CADUCEE de l’IRSN. Le poly méthacrylate de méthyle (PMMA) et l’heptane ont été utilisés comme combustibles, et pour chacun d’eux, différentes tailles de foyer ont été considérées. Les données collectées dans cette étude ont été comparées avec celles de la littérature pour divers matériaux et différentes échelles, révélant un bon accord. En supposant l’équilibre chimique, le calcul de la richesse du milieu réactionnel, à partir de la concentration de CO2 dans les fumées extraites, donne une richesse proche de l’unité, ce qui révèle une combustion incomplète faiblement réductrice. En fait, la flamme, et donc le débit de pyrolyse, s’adaptent à la concentration d’oxygène dans le mélange oxydant pour rester proche de la stœchiométrie. / Oxygen supply has a leading role in fire growth in confined spaces. The oxygen quantity available for combustion depends on the oxygen consumption by the fire and on the air supply from the mechanical ventilation system or openings. A decrease of oxygen concentration of the oxidant flow will lead to a decrease of the heat flux feedback from the flame to the fuel surface, which in turn will lead to a decrease in mass loss rate. The present experimental study has a dual objective: understand the effects of an under-oxygenated atmosphere on the combustion of solids and liquids and collect data for model validation. The Controlled Atmosphere Device for Unburnt and Carbon Emission Evaluation (CADUCEE) of IRSN has been used to conduct experiments at oxygen concentrations ranging from the limiting oxygen concentration for extinction to 21%vol. Fuels used are polymethylmethacrylate (PMMA) and heptane at different scales. A good agreement with literature data for various fuels and scales is found. Assuming chemical equilibrium, it is also found that the global equivalence ratio, deduced from the concentration of CO2 in the extracted gases, is close to unity, which reveals a weakly reductive incomplete combustion. This suggests that the flame, and thus the mass loss rate, adapt themselves to the available concentration of oxygen in the oxidant flow, to stay close to stoichiometry.

Made available in DSpace on 2014-06-11T19:35:12Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-07-12Bitstream added on 2014-06-13T20:26:18Z : No. of bitstreams: 1 barros_jmp_dr_botfm.pdf: 645820 bytes, checksum: e0369b9c2f84abbf1ff69b061c6def53 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / As variáveis hemodinâmicas e de oxigenação sistêmica não refletem com precisão a hipoperfusão esplâncnica durante o choque hipovolêmico, dificultando o tratamento adequado. A expansão volêmica após a reposição fluídica é fundamental para promover a oxigenação sistêmica e regional após o choque hemorrágico. Em contraste com as soluções convencionais de reposição volêmica, a menor expansão plasmática proporcionada pela administração de hidroxietilamido em solução hipertônica de cloreto de sódio, durante o choque hemorrágico, poderia determinar uma menor oferta de oxigênio sistêmico, com prejuízo à oxigenação gástrica. O estudo teve como objetivo comparar a expansão volêmica e os efeitos imediatos na oxigenação sistêmica e da mucosa gástrica após a administração de hidroxietilamido a 6% (peso molecular de 130 kDa, grau de substituição de 0,4) em solução hipertônica de cloreto de sódio a 7,5% (HHEA), Ringer lactato (RL) e hidroxietilamido a 6% (130/0,4) em cloreto de sódio a 0,9% (HEA), em cães submetidos à choque hemorrágico. Trinta cães, sem raça definida, sob anestesia e esplenectomizados, foram submetidos a sangramento (30 ml/kg) visando manter a pressão arterial média de 40 a 50 mm Hg durante 45 9 Introdução e Literatura minutos, sendo feita a reposição volêmica após este período com RL (n=10), na razão de 3:1 para o sangue removido; HEA (n=10), na razão de 1:1 para o sangue removido; e HHEA (n=10), 4 ml/kg. A expansão do volume intravascular (através da diluição do azul de Evans e da hemoglobina), e os atributos hemodinâmicos, e de oxigenação sistêmica e gástrica (através da tonometria gástrica), foram determinados no momento basal, após 45 minutos de hemorragia, e aos 5, 45 e 90 minutos após a reposição volêmica. A solução de HHEA aumentou o volume sanguíneo, devido à alta eficiência na... / Hemodynamic and global oxygen transport variables have failed to reflect splanchnic hypoperfusion, resulting in a failure to recognize inadequately treated hemorrhagic shock. Volemic expansion after fluid resuscitation is essential to improve global and regional oxygen in hemorrhagic shock. We hypothesized that, in contrast with conventional plasma expanders, the smaller volemic expansion from hypertonic hydroxyethyl starch solution administration in hemorrhagic shock may determine lesser systemic oxygen delivery and gastric oxygenation. We used hemorrhaged dogs to compare the early intravascular volume expansion and systemic and gastric oxygenation effects of 7.5% NaCl 6% hydroxyethyl starch 130/0.4 (HHES), lactated Ringer’s (LR), and 0.9% NaCl 6% hydroxyethyl starch (HES) solutions. Thirty mongrel dogs anesthetized and submitted to splenectomy, were bled (30mL/kg) to hold mean arterial pressure at 40-50 mm Hg over 45 minutes and were randomly resuscitated in three groups: LR (n=10) at 3:1 ratio to shed blood; HES (n=10) at 1:1 to shed blood; and HHES (n=10), 4mL/kg. Intravascular volume expansion (Evans blue and hemoglobin dilution), hemodynamic, systemic oxygenation and gastric intramucosal- arterial PCO2 gradient (PCO2 gap) variables were measured at baseline, after 45 min of hemorrhage, and 5, 45, and 90 min after fluid resuscitation. HHES increased blood volume, due to the 11 Introdução e Literatura high volume expansion efficiency, but intravascular volume expansion with this solution was the smallest of the solutions. All three solutions induced a similar hemodynamic performance but HHES showed lower mixed venous oxygen saturation and higher systemic oxygenation extraction and PCO2 gap than LR and HES. In conclusion, the smaller volume state from HHES after resuscitation provides worse systemic and gastric oxygenation recovery compared to LR and HES in dogs submitted... (Complete abstract click electronic access below)