The processing of bitumen in the Athabasca oil sands region (AOSR) produces extensive volumes of oil sands process-affected water (OSPW) and tailings, which are stored within tailings ponds and settling basins to promote the consolidation of solids and the recycling of water. Oil sands operators are actively investigating dry and wet reclamation strategies in order to reduce their inventory of tailings and return disturbed land back to its original state. An important component of the reclamation of tailings is understanding the environmental fate of naphthenic acids (NAs), which are considered the most toxic constituents of OSPW and tailings. However, since NAs exist as a complex mixture comprised of thousands of compounds from dozens of chemical classes, the characterization of NAs within environmental samples poses significant challenges to analytical chemists. This dissertation is focused on the characterization of naphthenic acids by comprehensive two-dimensional gas chromatography coupled to mass spectrometry (GC×GC/MS). GC×GC/MS offers unparalleled chromatographic separation and peak capacity and has been used in recent years to resolve individual constituents within complex mixtures, including structural isomers. Since the biodegradation and toxicity of NAs is structure-specific and can vary between structural isomers, the profiling of individual NAs by GC×GC/MS is expected to enhance the monitoring of NAs within environmental samples impacted by oil sands activity. In this thesis, GC×GC coupled with time-of-flight mass spectrometry (TOFMS) was used to structurally elucidate a number of ‘unknown’ classical and sulfur-containing naphthenic acids by interpretation of their electron ionization (EI) mass spectra and, if available, confirmed by comparison with the spectra of references standards. GC×GC/TOFMS was also utilized as a fingerprinting tool to assess the temporal and spatial variability at two reclamation sites in the AOSR: Syncrude’s Sandhill Fen reclamation site and Base Mine Lake. Lastly, a methodology was developed which coupled GC×GC with a high resolution quadrupole time-of-flight mass spectrometer (QTOFMS) for the improved profiling of NAs. GC×GC/QTOFMS is advantageous for the monitoring of NAs since it can provide useful fingerprints via isomer distributions, differentiate NAs from several chemical classes, and provide a global overview of the elemental compositions (assigned by mass accuracy) within NA mixtures. / Thesis / Doctor of Philosophy (PhD)

[pt] O objetivo deste estudo foi determinar a concentração de metais, compostos orgânicos e inorgânicos por suspensão aquosa e análise dos metais por abertura ácida presentes nas amostras de PM10 coletadas em áreas urbanas, industrial e rural da Região Metropolitana do Rio de Janeiro. A concentração de PM10 foi determinada por análise gravimétrica. Espécies inorgânicas e orgânicas presentes nos extratos aquosos foram determinadas por cromatografia de íons e carbono orgânico total (TOC). Os extratos ácidos, e também os aquosos, foram analisados por ICP-MS (espectrometria de massa com plasma indutivamente acoplado) e por ICP OES (espectrometria de emissão óptica com plasma indutivamente acoplado) para determinar a concentração de metais. As correlações entre as concentrações de PM10 e de metais foram avaliadas, considerando os dados meteorológicos para cada ponto de coleta e origem das massas de ar. Os resultados mostraram que as concentrações médias de PM10 foram de 36 micrograma m-3 na área rural, 50 micrograma m-3 na área urbana e 74 micrograma m-3 na área industrial. A concentração de PM10 medida na área industrial ultrapassou, em geral, o limite estabelecido pela resolução CONAMA, de 50 micrograma m-3. O teor de metais apresentou maior concentração na área industrial, sendo predominantes ferro, zinco, alumínio, titânio, manganês, cromo, níquel, cádmio e chumbo. Na área de maior tráfego, foi detectada a maior concentração de cobre e vanádio. As espécies iônicas foram maiores na área urbana, devido à proximidade do oceano. Na área rural, menores concentrações de espécies antropogênicas, como sulfato, nitrato e alguns metais (Pb, Cr, V e Ni) foram medidas. Assim, as concentrações das espécies estudadas são influenciadas pelas características locais. Porém, também foi observado a influencia do transporte de poluentes de longa distância, como queimadas e poeiras dos desertos do continente africano. Os percentuais dos extratos ácido/aquoso demonstraram que os metais extraídos de forma mais eficiente foram V e Ni (maior que 45 porcento), enquanto que os menos extraídos foram Al e Fe (menor que 3 porcento), conforme o esperado. A fração aquosa é importante para avaliar a biodisponibilidade de metais associados a danos à saúde. / [en] The scope of this study was to measure the concentration of metals and organic and inorganic anions in aqueous and acid extracts of PM10 samples collected in urban, industrial and rural zones of Rio de Janeiro. PM10 concentration was determined by gravimetric analysis and the organic and inorganic species in the aqueous extracts were determined by ion chromatography and total organic carbon (TOC). The aqueous and acid extracts were analyzed by inductively coupled plasma mass spectrometry (ICP-MS) and by ICP OES (inductively coupled plasma optical emission spectrometry), in order to determine metal content. Correlations between PM10 and metal concentrations were evaluated, considering meteorological data for each site. Results showed that the average PM10 concentrations were 36 microgram m-3; 50 microgram m-3 and 74 microgram m-3 at the rural, downtown and industrial sites, respectively. The measured PM10 concentration in the industrial site exceeded the limit allowed by the Brazilian law, which is 50 microgram m-3. While industrial area had the highest concentrations of metals, such as Fe, Zn, Al, Ti, Mn, Cr, Ni, Cd and Pb, the highest concentrations of Cu and V were found in areas with higher traffic densities. Ionic species had higher concentrations in the urban site due to the proximity to the ocean. The lowest concentrations of species from anthropogenic sources, such as sulfate, nitrate and some metals (Pb, Cr, V and Ni) were measured in the rural area. Hence, studied species concentrations are influenced by site locations, but the influence by the transportation of long distance pollutants, such as biomass burning and dust from African deserts was also observed. The acid/aqueous percentiles showed that the most efficiently extracted metals in the aqueous phase were V and Ni (greater than 45 percent) while the less efficiently extracted were Al and Fe (less than 3 percent), as expected. The aqueous fraction is important to evaluate the bioavailability of metals that are associated to damage to the human health.

Monitoring the impact of anthropogenic activities, particularly in industrial regions, requires ecological screening tools and frameworks that provide a comprehensive understanding of ecosystem responses to environmental changes. Biological indicators, organisms like algae, insects, fish, and sentinel mammals, are critical for assessing ecosystem health, particularly in areas of high industrial activity. The aim of this thesis was to identify a cross-species biomarker that can assess organismal health and environmental stress across various species, organs, and biological matrices. A range of biological systems and signaling pathways related to xenobiotic metabolism, energy homeostasis, immune responses, and stress adaptation were explored, leading to the identification of the Tryptophan-Kynurenine Pathway, which consumes 60-90% of tryptophan in vertebrates. Tryptophan and its metabolites play key roles in diverse physiological processes, including cell growth and maintenance, immunity, disease states, and the coordination of adaptive responses to environmental and dietary cues. This adaptive response suggests that kynurenine-tryptophan ratio (KTR) may serve as a marker for exposure to a variety of environmental stress conditions, including toxicants, nutrient scarcity, predatory stress, and habitat loss—stressors that are prevalent in areas of high industrial activity. In recent years, the KTR is increasingly recognized as a sensitive biomarker in human diseases induced or exacerbated by stress; however, its role in environmental exposure and wildlife health remains unexplored. This thesis explores the question of whether KTR can be utilized as a cross-species biomarker for environmental stress or environmental exposure to toxicants, particularly focusing on the Athabasca Oil Sands Region (AOSR). In vitro studies with mammalian hepatocytes exposed to polycyclic aromatic compounds (PACs): benzo[a]pyrene (BaP), and a Bitumen Water Accommodated Fraction (BitWAF) demonstrated that KTR increases were driven by elevated kynurenine levels, indicating disruption of tryptophan metabolism via the aryl hydrocarbon receptor (AhR). Further studies using acid extractable organics from Oil Sands Process-Affected Water (OSPW), Naphthenic Acid Fraction Components (NAFCs) showed metabolic reprogramming, including altered glucose and fatty acid uptake and mitochondrial dysfunction, mediated through PPARα activation and upregulation of Tdo2, the enzyme responsible for kynurenine production. In vivo studies of longnose and white suckers from the AOSR were conducted to assess the relationship between KTR and CYP1 enzyme activity (EROD). These studies revealed species-specific responses, with an inverse correlation between KTR and EROD in longnose suckers and a direct correlation in white suckers. These findings validate KTR as a biomarker for environmental exposure in wildlife, with significant implications for monitoring ecosystem health. Collectively, this work demonstrates the potential of KTR as a novel biomarker for environmental toxicology, offering a valuable tool for assessing organismal stress across species in response to environmental contaminants. / Thesis / Doctor of Philosophy (PhD) / Human activities, especially industrial operations, can significantly impact the environment. To monitor these effects, scientists use various tools and organisms to assess ecosystem health. This research introduces a new approach to measuring environmental stress in wildlife by focusing on two key molecules: tryptophan and kynurenine. These molecules are part of a conserved biological pathway that helps all organisms manage stress, repair cells, adapt to their environment, and maintain overall health. Tryptophan, an essential amino acid, is broken down into kynurenine, and the balance between them— known as the kynurenine-tryptophan ratio (KTR)—can indicate the level of stress an organism is experiencing. This thesis investigates whether KTR can detect environmental stress caused by industrial activity, particularly from petroleum-derived chemicals in the Athabasca Oil Sands Region (AOSR). In laboratory experiments, mammalian liver cells were exposed to oil sands compounds and complex mixtures from oil sands wastewater. These compounds changed KTR, showing that the liver’s stress response was activated, and tryptophan metabolism was disrupted. The study also found that these chemicals affected cellular energy use and the way cells process fats and sugars. Furthermore, we examined fish species in the AOSR: longnose and white suckers. Results showed that KTR varied depending on the species and the location of exposure. In white suckers, KTR increased in response to stress, while in longnose suckers, it decreased, indicating species-specific responses to environmental changes. Overall, our findings suggest that KTR could serve as a useful tool for measuring environmental stress in different species and ecosystems, especially in areas affected by anthropogenic or industrial activity. Understanding how KTR changes in response to pollution can help scientists better monitor and protect wildlife and ecosystem health.