Dissertations / Theses on the topic 'Active oxidants'

Selective oxidations are important for the functionalization of compounds in organic synthesis and chemical industry. Using O2 as a terminal e- acceptor would be ideal because it is cheap and environmentally friendly, but aerobic oxidations are often prone to unselective free radical autoxidation. Recently developed palladium catalysts use O2 as a selective multi-electron oxidant for various organic transformations. Although these methods are powerful and sophisticated, the lower cost of base metals makes them attractive as potential alternatives. The challenge is to develop methods for effecting multi-electron transformations at metals that typically prefer one electron changes. To this end, the development of manganese(III) complexes containing redox-active ligands as catalysts for selective oxidase-type oxidation of organic substrates was pursued. Bis(tetrabromocatecholato) manganese(III) complexes were shown to aerobically oxidize catechols to form quinones and H2O2. This reactivity was extended to other alcohol and amine substrates. In these reactions, the non-innocent tetrabromocatecholate ligands may impart a multi-electron character to the metal. To directly probe the intermediacy of ligand-centered radicals in catalytic turnover, a series of structurally similar manganese(III) complexes with aminophenol-derived ligands were prepared and characterized. The capacity of these ligands to undergo low-energy redox changes, allowed for isolation of an electron transfer series spanning two redox states without a change in oxidation state at the metal center. The ligand-centered redox events were a key feature in aerobic homocoupling of Grignard reagents.

Abstract Glutaredoxins (Grx) are small thiol disulphide oxidoreductases with a conserved active site sequence -CXXC/S- and a glutathione (GSH) binding site. They catalyze the reduction of protein disulphides, preferring protein-GSH mixed disulphides as substrates. The accumulation of protein-GSH mixed disulphides has been observed during oxidative stress, where they may serve both a regulatory and an antioxidant function by protecting the enzymes from irreversible oxidation. Once oxidative stress has been removed the GSH-protein mixed disulphides are reduced by GSH or, more efficiently, by Grx. The present study showed for the first time that Grx1 and Grx2 can be detected in healthy human lung. Highly specific expression of Grx1 was observed in alveolar macrophages, but it could also be detected from sputum supernatant. Grx1 levels in alveolar macrophages were lower in selected inflammatory diseases than in control lung samples. Grx1 was also mainly negative in the fibrotic areas in usual interstitial pneumonia, an aggressive fibrotic lung disease. Overall, the present study suggests that Grx1 is a potential redox modulatory protein regulating the intracellular as well as extracellular homeostasis of glutathionylated proteins and GSH not only in healthy lung, but also in inflammatory and fibrotic lung diseases. In order to study the mechanism of action of glutaredoxins in vitro, a new real-time fluorescence-based method for measuring the deglutathionylation activity of glutaredoxins using a glutathionylated peptide as a substrate was developed. The first reaction intermediate in the deglutathionylation reaction was shown to be exclusively Grx-GSH mixed disulphide and this specificity was solely dependent on the unusual γ-linkage present in glutathione. The study also demonstrated the role of conserved residues in the proximity of proposed GSH binding site to the GSH binding specificity of E. coli Grx1. Opening the binding groove and removing charged residues enabled Grx to form more readily mixed disulfides with other molecules besides GSH. Different members of the PDI family showed considerably lower activity levels compared to glutaredoxins and, in contrast to the glutaredoxin-GSH mixed disulphide, the only intermediate in the PDI catalysed reaction was PDI-peptide mixed disulphide.

Master of Science
Department of Grain Science and Industry
J.M. Faubion
The market for frozen goods is expanding and the frozen dough goods sector still has potential to expand its market. It is well known that deterioration in bread quality occurs during frozen dough/bread production. In addition, it is known that dough rheology influences bread quality. To prevent deterioration of bread quality, many additives have been used and researched. Combinations of oxidants (potassium bromate and ascorbic acid) are widely used worldwide. However, potassium bromate may be carcinogenic to humans, and it has been detected in bread after baking. Since it has been prohibited or strictly limited in many countries, many researchers have tried to find a replacement. Ascorbic acid is safe for human intake, and does not persist in bread. However, it is not as effective as potassium bromate. Possible replacements in frozen doughs include oxidant (ascorbic acid)-enzyme combinations. This study evaluated the effects of ascorbic acid-specific enzyme combinations as a replacement for the potassium bromate in frozen dough and related the effects to dough behavior (gluten network strength) as evaluated by dynamic oscillation rheometry. Bread quality was evaluated by test baking. Based on the results from fresh baking studies, potassium bromate can be replaced by an optimum level combination of ascorbic acid and hemicellulase/endo-xylanase. This combination clearly improved loaf volume, and crumb grain over both control and potassium bromate containing doughs. For frozen dough/bread production, the addition of all additives improved bread quality, but ascorbic acid and endo-xylanase containing dough resulted in higher volume, and better crumb structure than did dough containing potassium bromate. Dough rheology experiments show that rheology was affected by both the process and additives. Strain sweeps gave the information about dough stability. Both the additives and proofing improved dough stability. Dough behavior (gluten network strength) was assessed by frequency sweeps. Dough containing ascorbic acid and endoxylanase was most stable during frozen dough processing.

[ES] Las estaciones depuradoras de aguas (EDAR) tienen un papel fundamental en la protección del medio ambiente, evitan la llegada de nutrientes (nitrógeno y fósforo) y otros contaminantes a los ecosistemas acuáticos. El sistema más utilizado para la eliminación de nitrógeno en las EDAR es el proceso biológico de nitrificación-desnitrificación vía nitrato. El rendimiento de los sistemas biológicos está directamente relacionado con la estructura de la comunidad bacteriana y su metabolismo. En este trabajo se monitorizaron las variaciones temporales de las características fisicoquímicas del afluente, los parámetros operacionales y los rendimientos de eliminación del amonio en 6 reactores biológicos con sistemas de fangos activos. Para la caracterización de comunidades involucradas en el proceso de nitrificación se utilizaron diferentes técnicas de biología molecular: hibridación in situ con sondas marcadas con fluoróforos (FISH), secuenciación de segunda generación Illumina y secuenciación de tercera generación SMRT de PacBio, la cual no había sido utilizada hasta la fecha para el análisis de la microbiota de sistemas convencionales de eliminación de nutrientes. Para valorar la actividad de la biomasa nitrificante y de la biomasa heterótrofa se utilizaron técnicas respirométricas y técnicas para la cuantificación del ATP de última generación. Para analizar el gran volumen de datos generado tras la aplicación de las diferentes técnicas, se utilizaron técnicas estadísticas de análisis multivariante, como los modelos de regresión lineal multivariante basados en la distancia (DISTLM). Estas técnicas estadísticas permitieron valorar la contribución de las variables ambientales a la variabilidad observada en la estructura de las comunidades de bacterias nitrificantes, los rendimientos de eliminación del nitrógeno y su actividad. Las técnicas moleculares empleadas permitieron determinar que Nitrosomonas oligotropha, Nitrospira spp. y Nitrotoga sp. fueron las especies responsable de los procesos de nitrificación en las EDAR analizadas. Los resultados alcanzados con las técnicas FISH e Illumina sobre la estructura de la población de bacterias nitrificantes fueron similares, y permitieron detectar los sesgos de la secuenciación SMRT de PacBio. Los modelos de regresión permitieron valorar la contribución de las bacterias nitrificantes a la eliminación del amonio y cuáles fueron los factores que influían en su abundancia en cada una de las EDAR. La carga orgánica, la concentración de sólidos volátiles, la concentración de oxígeno y la temperatura fueron las variables de mayor influencia en la abundancia de estas especies. Mientras que la carga de fósforo influenció significativamente en su actividad. Este estudio reveló que la aplicación de ozono disminuye significativamente a los rendimientos de eliminación del amonio. Los resultados obtenidos ayudaron a mejorar la comprensión sobre el proceso de nitrificación en cada una de las EDAR y manifiestan la importancia de la dinámica poblacional de las bacterias nitrificantes en los rendimientos de eliminación del amonio en las EDAR. Estos resultados establecen que las técnicas moleculares combinadas con respirometría y los modelos de ordenación multivariante empleados en esta tesis, son una herramienta fiable para la monitorización y el control del proceso de nitrificación en sistemas de eliminación biológica de nitrógeno. Los resultados de esta tesis sugieren que la medida de los sólidos suspendidos volátiles activos mediante técnicas de determinación de ATP de segunda generación puede mejorar el cálculo de las variables de diseño y control más habituales de las EDAR.
[CAT] Les estacions depuradores d'aigües residuals (EDAR) tenen un paper fonamental en la protecció del medi ambient, eviten l'arribada de nutrients (nitrogen i fòsfor) i altres substàncies contaminants als ecosistemes aquàtics. El sistema més utilitzat per a l'eliminació de nitrogen en les EDAR és el procés biològic de nitrificació-desnitrificació via nitrat. El rendiment dels sistemes biològics està directament relacionat amb l'estructura de la comunitat bacteriana i el seu metabolisme. En aquest treball es van monitorar les variacions temporals de les característiques fisicoquímiques de l'afluent, els paràmetres operacionals i els rendiments d'eliminació de l'amoni en 6 reactors biològics amb sistemes de fangs actius. Per a caracteritzar les comunitats involucrades en el procés de nitrificació s'han utilitzat diferents tècniques de biologia molecular: hibridació in situ amb sondes marcades amb fluoròfors (FISH), seqüenciació de segona generació Illumina i seqüenciació de tercera generació SMRT de PacBio, la qual no havia estat utilitzada fins avui per a l'anàlisi de la microbiota dels sistemes convencionals d'eliminació de nutrients. Per a valorar l'activitat de la biomassa nitrificant i de la biomassa heteròtrofa es van utilitzar tècniques respiromètriques i tècniques per a la quantificació de l'ATP d'última generació. Per a analitzar el gran volum de dades generat després de l'aplicació de les diferents tècniques, es van utilitzar tècniques estadístiques d'anàlisi multivariant, com els models de regressió lineal multivariant basats en la distància (DISTLM). Aquestes tècniques estadístiques van permetre valorar la contribució de les variables ambientals a la variabilitat observada en l'estructura de les comunitats de bacteris nitrificants, els rendiments d'eliminació del nitrogen i la seua activitat. Les tècniques moleculars emprades van permetre determinar que Nitrosomonas oligotropha, Nitrospira spp. i Nitrotoga sp resultaren les espècies responsables del procés de nitrificació en les EDAR analitzades. Les tècniques FISH i Illumnina van mostrar resultats molt similars sobre l'estructura de la població de bacteris nitrificants i van permetre detectar els biaixos de la seqüenciació SMRT de PacBio. Els models de regressió van permetre valorar la contribució dels bacteris nitrificants a l'eliminació de l'amoni i quins van ser els factors d'influència en la seua abundància en cadascuna de les EDAR. La concentració de matèria orgànica, sòlids volàtils en suspensió, la concentració d 'oxigen i la temperatura foren les variables amb més influència en l'abundància d'aquestes especies. Així mateix la càrrega de fòsfor va influir en la seua activitat. Aquest estudi va determinar que l 'aplicació ozó va causar una disminució significativa dels rendiments d 'eliminació del amoni. Aquests models van ajudar a millorar la comprensió sobre el procés de nitrificació en cadascuna de les EDAR i ressalten la importància de la dinàmica poblacional dels bacteris nitrificants en el rendiments de l 'eliminació de l 'amoni. Els resultats estableixen que les tècniques moleculars combinades amb respirometría i els models d'ordenació multivariant emprats en aquesta tesi, són una eina fiable per al monitoratge i el control del procés de nitrificació en sistemes d'eliminació biològica de nitrogen. Els resultats d'aquesta tesi suggereixen que la mesura dels sòlids suspesos volàtils actius mitjançant tècniques de determinació d'ATP de segona generació pot millorar el càlcul de les variables de disseny i control més habituals de les EDAR.
[EN] Wastewater treatment plants play an important role in environmental protection. These facilities protect aquatic ecosystems from excessive inputs of nutrients (nitrogen and phosphorous) and other pollutants. The most widespread system of nitrogen removal in wastewater treatment plants is a conventional method involving a biological nitrification-denitrification via nitrate. The efficiency of biological systems is directly related to bacterial community structure and its metabolism. In this work, the temporal variations of influent characteristics, operational parameters and ammonium removal efficiency in 6 bioreactors with activated sludge systems were monitored. To characterize the microbial communities involved in the nitrification process different molecular biology techniques were used: fluorescence in situ hybridization (FISH); second generation sequencing (Illumina), and third generation sequencing (SMRT PacBio). To assess the activity of nitrifying and heterotrophic bacteria, respirometric tests and second-generation ATP determination techniques were used. To analyse the large volume of data generated, statistical multivariate analysis techniques were used, including distance-based multivariate linear regression models (DISTLM). These statistical techniques allowed us to assess the contribution of environmental variables to the variability observed in the nitrifying community structure, in nitrogen removal performance and nitrifying activity. The molecular techniques employed determined that Nitrosomonas oligotropha, Nitrospira spp. and Nitrotoga sp. where the dominant nitrifying bacteria in the monitored WWTP. FISH and Illumnina technique showed very similar results and allowed for the detection of biases in PacBio SMRT sequencing. The regression models determined the contribution of nitrifying bacteria to ammonium oxidation and the factors influencing their abundance and activity. The main factors influencing the nitrifying bacteria abundance were organic load, volatile suspended solids concentration, dissolved oxygen and temperature. The activity of nitrifying bacteria also was influenced by phosphorous loading rate. This study revealed that ozone concentration was the main factor determining the low ammonium removal performance. These models helped to improve our understanding of the nitrification process in each WWTP and highlight the importance of nitrifying bacterial community structure in nitrogen removal performance. The results establish that the molecular techniques combined with respirometry and the multivariate ordering models used in this thesis, are a reliable tool for the monitoring and control of the nitrification process. The results of this thesis suggest that the measurement of active volatile suspended solids by means of second- generation ATP determination techniques can improve calculation of the most common design and control parameters of WWTPs.
A la Entidad de Saneamiento y Depuración de la Región de Murcia (ESAMUR) por la financiación del proyecto “Influencia de las variables operacionales y fisicoquímicas en la dinámica y estructura de la población de bacterias nitrificantes”, al Grupo de Química y Microbiología del Agua del IIAMA. A la Entidad Pública de Saneamiento de Aguas Residuales de la Comunidad Valenciana (EPSAR), por la financiación del proyecto “Estudio integrado del proceso biológico en plantas de tratamiento por fangos activos, análisis de interrelación entre los distintos componentes y optimización de métodos moleculares para la identificación de bacterias formadoras de espumas” al Grupo de Química y Microbiología del Agua del IIAMA
Barbarroja Ortiz, P. (2019). Estudio de la dinámica poblacional y actividad de los organismos nitrificantes en sistemas de depuración de aguas residuales [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/124063
TESIS

This investigation focused on Rhoicissus tomentosa, belonging to the family, Vitaceae in an attempt to assess the phytochemistry of this plant which is widely used by traditional healers in South Africa to ensure the safe delivery during pregnancy and childbirth (Hutchings et al., 1996).

Nicotine exposure to the fetus through tobacco smoking or nicotine replacement therapy during the whole period of gestation and lactation causes diverse effects on fetal and neonatal lung development, integrity and maturation which compromise the gas exchange function of the lungs and renders this vital organ susceptible to gradual damage and different diseases in latter life. Maternal nicotine exposure during gestation and lactation results in gradual destruction of the lung parenchyma, and this leads to the combination of many small air sacs in one bigger alveoli which is a sign of emphysema. Many researchers speculated that the way in which, nicotine causes emphysema and other damage, is by inducing the formation of many reactive oxygen species (ROS), and creating an imbalance between the oxidants and the antioxidants of the body, which is termed oxidative stress. The aim of this study was to assess the effects of nicotine exposure on the lung of the fetal and neonate rat during gestation and lactation as gas exchanger, and also to see whether the supplementation of tomato juice containing lycopene, a powerful carotenoid antioxidant could protect the lungs against these effects of maternal nicotine exposure.

Differents facteurs tels les rayonnements uv ou ionisants, les agents oxydants, les radicaux libres, peuvent engendrer des dommages au niveau des bases de l'adn. Ces modifications peuvent avoir une influence notable sur la structure et les proprietes du biopolymere. Afin de mieux comprendre la nature de ces lesions, leur mode de formation et les effets qu'elles peuvent avoir sur l'integrite de l'adn, l'etude de certains degradations de la thymine a ete realisee au moyen des outils de la chimie quantique. La premiere partie de ce travail est consacree a l'etude comparative des proprietes conformationnelles et electroniques de produits d'oxydation du groupement methyle de la thymidine par l'intermediaire d'une methode locale de la dft. Le chapitre suivant traite des intermediaires radicalaires formes par l'addition d'un atome d'hydrogene ou du radical hydroxyle sur la double liaison 5,6 de la thymine. Un protocole general d'etude de la structure et des constantes de couplage hyperfin isotrope rpe, incluant une methode hybride de la dft (b3lyp), les effets du solvant, des vibrations et de la temperature a ete mis en place. Ce protocole a ete applique a l'analyse des produits radicalaires formes par addition de l'atome d'hydrogene. D'autre part, les parametres thermodynamiques et cinetiques des differentes reactions d'addition ont ete determines. Un modele de reactivite en solution est finalement propose pour la comprehension des mecanismes d'addition du radical hydroxyle. La derniere partie concerne l'etude des produits finaux non radicalaires formes par l'addition initiale du radical hydroxyle sur la double liaison 5,6 de la thymine. La comparaison des conformations et des proprietes electroniques des deux isomeres cis d'hydroxyhydroperoxydes de thymine et des deux isomeres cis de la 5,6-dihydrothymidine a ete effectuee.

The cytochrome P450 heme-thiolate enzymes catalyse a multitude of oxidation reactions and, in humans, carry out drug metabolism. P450s perform hydroxylation, epoxidation, N-, O- and Sdealkylation, sulfoxidation, alkyne oxidation and aldehyde oxidation of organic molecules (and many other reactions). These reactions are predominantly performed by the reactive intermediate Compound I, but other intermediates in the catalytic cycle may mediate some types of reactions. It would be appealing to exploit these enzymes as environmentally benign catalysts in the synthesis of fine chemicals. Their widespread use in industrial synthesis is, however, impractical given the high cost of the required cofactor NAD(P)H, but engineering P450s to instead use cheap H₂O₂ would overcome this problem. CYP199A4 is a soluble bacterial P450 enzyme from Rhodopseudomonas palustris HaA2 that favours 4-methoxybenzoic acid and other para-substituted benzoic acids as substrates. It tightly binds these substrates and the para-substituent is rapidly oxidised. This enzyme has been used as a model system to study the mechanism of P450-catalysed reactions. While 4-methoxybenzoic acid is oxidatively demethylated at a rate of 1220 μM (μM-P450)⁻¹ min⁻¹, CYP199A4 displays no detectable activity towards the meta isomer, 3-methoxybenzoic acid. In vitro reactions were performed with a range of other meta-substituted benzoic acids (3-methylthio-, 3-methylamino-, 3- formyl-, 3-methyl-, 3-isopropyl-, 3-tert-butyl- and 3-ethoxy-benzoic acid) to assess whether CYP199A4 had activity towards these substrates. These meta-substituted substrates, except for 3- tert-butylbenzoic acid, were all metabolised by CYP199A4, but with low activity compared to the corresponding para isomers. Compared to the para isomers, the meta isomers had lower binding affinity and induced smaller type I spin-state shifts to high-spin. To rationalise CYP199A4’s preference for para- over meta-substituted benzoic acid substrates and to investigate the requirements for efficient monooxygenase activity, crystal structures were solved of CYP199A4 in complex with 3-methoxy-, 3-methylamino-, 3-methylthio-, and 3- and 4-methyl-benzoic acid. These structures revealed that the heme-bound water ligand to the heme is retained when these substrates bind (water occupancy 21-90%) and is hydrogen-bonded to the heteroatom (N, S, O) of the substrate. The corresponding para isomers displace the iron-bound water. 3-Ethoxybenzoic acid, which has a bulkier meta-substituent, shifted the spin-state to 85% high-spin and in the crystal structure the iron-bound water was removed. The meta-substituent of each substrate is held in close proximity to the iron. 3-Methoxybenzoic acid is positioned near the iron but is not oxidised. This was attributed to the fact that the C-H bonds are oriented away from the heme, whereas those of 4-methoxybenzoic acid are ideally oriented for H-atom abstraction by Cpd I. These results emphasise that close proximity of the methyl carbon to the heme iron does not guarantee that hydroxylation will occur if the C-H bonds are not oriented appropriately for abstraction, and subtle modification of the substrate’s position relative to the heme can abolish catalytic activity. X-ray crystallography, CW and HYSCORE EPR and other experiments were performed to elucidate the binding modes of 4-pyridin-2-yl-, 4-pyridin-3-yl- and 4-imidazol-1-yl-benzoic acid in the CYP199A4 active site. These heterocyclic aromatic compounds are not metabolised and induce substantially different type II UV-Vis spectra. 4-Pyridin-3-yl- and 4-imidazol-1-yl-benzoic acid redshifted the Soret band from 419 to 424 nm. They induced ‘normal’ type II spectra, characterised by a less intense α-band than β-band and an increase in δ-band intensity. The UV-Vis spectra of ferrous CYP199A4 in complex with these ligands indicated that 4-pyridin-3-ylbenzoic acid was directly ligated to the heme iron via the pyridine nitrogen, but the Fe-N bond between the iron and 4-imidazol-1-ylbenzoic acid was ruptured upon heme reduction. 4-Pyridin-2-ylbenzoic acid induced a smaller Soret band red-shift (to 422 nm) when added to the ferric enzyme. It produced an ‘abnormal’ type II spectrum, with no decrease in the α-band intensity. 4-Pyridin-2-ylbenzoic acid also induced a smaller Soret band trough in the difference spectrum than 4-pyridin-3-ylbenzoic acid. HYSCORE EPR and X-ray crystallography revealed that 4-pyridin-3-yl- and 4-imidazol-1-ylbenzoic acid were directly ligated to the ferric heme iron, but 4-pyridin-2-ylbenzoic acid was hydrogen-bonded to the heme-bound water. This study revealed that optical spectroscopy can distinguish between water-bridged and directly bound nitrogen donor ligands. 4-Pyridin-3-yl- and 4-imidazol-1-yl-benzoic acid-bound CYP199A4 were both reduced to the ferrous form by ferredoxin, ferredoxin reductase and NADH. On the other hand, binding of 4-pyridin-2-ylbenzoic acid to CYP199A4 lowered the reduction potential and prevented heme reduction by even the powerful reductant dithionite. This implies that water-bridged nitrogen ligands may in some instances be more effective P450 inhibitors than those that bind directly to the iron. The T252E mutant of CYP199A4 was produced and characterised. This variant was no longer able to operate using NADH but was a more efficient peroxygenase (H₂O₂-utilising enzyme) than the wild-type (WT) enzyme. EPR indicated that the sixth axial ligand to the heme was a mixture of hydroxide and water. Crystal structures showed that this aqua/hydroxo ligand was tightly bound due to strong interactions with the carboxylate of E252. The axial aqua/hydroxo ligand was not displaced by substrates, even sterically bulky substrates, explaining the lack of substrate-induced spin-state shifts and the exceedingly slow rate of electron transfer from the ferredoxin to the P450. Because this ligand is not displaced by substrates, the active species could potentially be generated via light-driven oxidation of the water-bound ferric resting state to Compound I. Type II nitrogen ligands were also unable to displace the aqua/hydroxo ligand. 4-Pyridin-3-ylbenzoic acid, when added to the T252E mutant, induced an ‘abnormal’ type II spectrum, confirming that optical spectroscopy can distinguish between water-bridged and directly bound type II ligands. X-ray crystallography revealed that the T252 → E mutation only subtly altered the orientation of substrates in the CYP199A4 active site. In the absence of substrate, the heme signal of the T252E mutant was rapidly bleached by 50 mM H₂O₂. When substrate was present, the T252E variant remained catalytically active for several hours. The T252E variant was able to perform a range of reactions using H₂O₂ (O-dealkylation, hydroxylation/desaturation, epoxidation, sulfoxidation, alkyne oxidation and aldehyde oxidation). When the surrogate oxygen donor tert-butyl hydroperoxide was substituted for H₂O₂, the T252E mutant had negligible activity. t-BuOOH is presumably too bulky to access the heme iron in this P450. WT CYP199A4 and the T252A and D251N mutants also catalysed these reactions using H₂O₂ but afforded less product over a 4-hour period than the T252E mutant. H₂O₂- and NADH-driven epoxidation of 4-vinylbenzoic acid catalysed by WT and mutant CYP199A4 proceeded with high enantioselectivity, yielding almost exclusively the (S)-enantiomer. In NADH-supported reactions, WT CYP199A4 catalysed O-demethylation of 4- methoxybenzoic acid more efficiently than sulfoxidation of 4-methylthiobenzoic acid. In H₂O₂- driven reactions, the T252E variant had higher activity towards sulfoxidation compared to Odemethylation, hydroxylation, epoxidation, aldehyde oxidation and alkyne oxidation. This may indicate the involvement of a second oxidant in sulfoxidation (e.g. the FeIII–H₂O₂ species), allowing sulfoxidation to occur in the absence of Compound I as proposed by Shaik.
Thesis (MPhil) -- University of Adelaide, School of Physical Sciences, 2020

As espécies reativas de oxigénio e azoto desempenham um papel importante em certas funções fisiológicas, como na inflamação, mas em situações de sobreprodução ou desregulação podem ter vários efeitos lesivos para as biomoléculas. O objetivo foi construir um modelo matemático de equações diferenciais ordinárias (ODEs) que descrevesse a rede de reações químicas entre as diferentes espécies reativas e as biomoléculas. Para tal: a) foram recolhidas da literatura e utilizadas as constantes cinéticas das várias reações para escrever as equações cinéticas de todas as reações que constituíram o sistema estudado; b) com base nas várias equações cinéticas foram construídas as equações diferenciais que descreviam a evolução temporal de cada espécie química no sistema, recorrendo à abordagem do tipo Continuously Stirred Batch Reactor (CSBR); c) otimizou-se os parâmetros do modelo de modo a obter um sistema em estado estacionário que mimetizou as condições observadas na circulação sistémica; d) foi feita a análise do modelo com recurso à análise de fluxos e de coeficientes de controlo, que identificaram quais os principais pontos de controlo do sistema. Com estes passos obteve-se um modelo matemático robusto que permitiu a simulação de situações fisiológicas como a desregulação dos sistemas de produção de espécies reativas ou o efeito de antioxidantes sobre o nível global de espécies reativas e sobre o nível de lesões induzidas às biomoléculas: ▪ As concentrações de estado estacionário das espécies tamponantes do sangue bem como da água, do hidrónio e do anião hidróxido mantiveram-se constantes em todas as variações; ▪ As variações do óxido nítrico e do superóxido ocorreram exatamente de acordo com o que se esperava; ▪ A variação da concentração do anião nitrato correspondeu à variação da concentração do peroxinitrito, indicando a composição do mesmo; ▪ Todas as espécies reativas tiveram a variação das suas concentrações como esperado em relação da variação das concentrações de estado estacionário dos antioxidantes. Contudo, não foi possível obter um modelo que descreva adequadamente o sistema de tamponamento do sangue, tendo-se identificado a origem desse erro no excessivo detalhe utilizado para descrever essas reações.
Reactive oxygen and nitrogen species play an important role in certain physiological functions, such as in inflammation, but in situations of overproduction or deregulation, they may have several detrimental effects on biomolecules. The goal was to construct a mathematical model of ordinary differential equations (ODEs) that describes the network of chemical reactions between different reactive species and biomolecules. For such: a) they were collected from literature and kinetic constants of the various reactions were used to write the kinetic equations of all the reactions that constituted the studied system; b) based on the various kinetic equations the differential equations that described the temporal evolution of each chemical specie in the system were constructed, using the Continuously Stirred Batch Reactor (CSBR); c) the model parameters were optimized to obtain a steady-state system that mimicked the conditions observed in the systemic circulation; d) the analysis of the model using flow and control coefficients analysis was performed, which identified the main control points of the system. These steps yielded a robust mathematical model that allowed the simulation of physiological situations such as the deregulation of reactive species production systems or the effect of antioxidants on the global level of reactive species and on the level of biomolecule induced lesions: ▪ Steady-state concentrations of the buffering species in the blood as well as water, hydronium and hydroxide anion remained constant in all variations; ▪ Changes of nitric oxide and superoxide went exactly according to what was expected; ▪ The variation of the concentration of the nitrate anion corresponded to the variation of the peroxynitrite concentration, indicating its composition; ▪ All reactive species had the variation of their concentrations as expected in relation to the variation of the steady-state concentrations of the antioxidants. However, it was not possible to obtain a model that adequately describes the blood-buffering system, and the origin of this error was identified in the excessive detail used to describe these reactions.

Theses (MSc. (Biochemistry)) -- University of Limpopo, 2015
Lithium is an FDA-approved psychiatric drug that has been used for more than half a century as a preferred, gold standard treatment for bipolar disorders (Cade, 1949; Freland and Beaulieu, 2012). Numerous studies reported that lithium can play a key role in regulation of inflammation and oxidative stress beyond bipolar disorders since it’s known to target GSK3-β and NF-κB key inflammation molecules (Yestevelasco et al., 2008; Zhang et al., 2008). The choice of this drug was derived from its easy accessibility, less cost, minor side effects and efficacy as opposed to other anti-depressant drugs used currently. This study is based on a body of experimental evidence that has outlined the link between uncontrolled inflammation and some chronic ailments. Thus, this study was aimed at investigating the molecular mode of action of lithium chloride on amelioration of oxidative stress and inflammation in activated Raw 264.7 since, preliminary results of this work suggested the anti-inflammatory properties of lithium. Lithium chloride (LiCl) is not cytotoxic to Raw 264.7 and NIH 3T3 cell lines up to 20 mM and no change in cell proliferation, viability, growth, and cell adhesion were observed as demonstrated using xCELLigence Real Time Cell Analyser (RTCA) and MTT assays. In addition, this drug was shown to be unable to induce programmed cell death in Raw 264.7 and NIH 3T3 cell after 10 mM LiCl treatment as demonstrated using Annexin-V/ PI apoptosis detection assay. Using the Griess and DAF2-DA assays pre-treatment with low doses of lithium (LiCl) was shown to reduce Nitric Oxide production in LPS-induced macrophages. A reduced internal H2DCF-DA fluorescence intensity is indicative of reduced ROS production, observed in lithium-treated Raw 264.7 macrophages stimulated with LPS, FMLP and PMA. Real Time PCR analysis revealed that lithium modulates expression of inflammation inhibitory genes such as IκB-α, TRAF3, Tollip and NF-κB1/p50. This inhibitors are known to play a vital role up stream (Tollip and TRAF3) and downstream (IκB-α and NF-κB1/p50) of NF-κB inflammation signalling pathway. Thus, these molecules are thought to be anti-inflammatory molecular targets of lithium. Moreover, immunocytochemistry suggest that lithium blocks nuclear translocation of NF-κB. This study associates lithium to reduced oxidative stress in LPS, FMLP and PMA-activated Raw 264.7 macrophages in a non-neuronal setting and further suggests lithium as a potential candidate for regulation of oxidative stress conditions beyond bipolar disorders.