Thesis (Ph. D.)--Oregon State University, 2007. / Printout. Includes bibliographical references. Also available on the World Wide Web.

Polycyclic aromatic hydrocarbons (PAHs) are environmentally significant compounds due to the toxicity of some members. They are ubiquitous and are persistent bioaccumulative toxins(PBTs). The toxicity of PAHs represents a risk to human health, and there are varied risk assessment approaches to quantifying the risk posed by PAHs based on exposure routes and scenarios. PAHs are not carcinogenic until they are metabolically activated as the body attempts to break them down and forms reactive metabolites that bind to the DNA causing subsequent replication in the cells. Fundamental to assessing the risk posed by PAHs is understanding the metabolism of PAHs. Since exposure to PAHs is never to single PAHs, understanding what differences may occur in mixtures of PAHs gives accurate assessment of the dangers of PAHs. Understanding the dynamics of complex metabolism vis-a-vis single metabolism of PAHs and possible effects on the toxicity expression of PAHs is a necessary advancement to accurately impact and guide remediation strategies. Studies were carried out comparing the metabolism of the PAHs Phenanthrene (PHE), Flouranthene (FLA) and Benzo[a]pyrene (BAP) in single, binary and ternary mixtures by monitoring the disappearance of the parent compound. It was observed that PAH metabolism in the single PAH experiment differed from metabolism in both binary and ternary mixtures. Enzyme competition was evident in the metabolism of mixtures, changing significantly the metabolism patterns of individual PAHs. PAH structure was also seen to affect metabolism in mixtures and the possible creation of toxicity effects during mixture metabolism. PAH concentration changed over time with faster change during single PAH metabolism followed by ternary mixture metabolism and finally binary metabolism. These results affirm that substrate interactions must be considered in the risk assessment approaches to the dangers posed by exposure to PAHs.

Polycyclic aromatic hydrocarbons (PAH) have been identified as a serious environmental problem. In past research it has been proven that naphthalene, the simplest PAH, could be biodegraded using roller bioreactors and Pseudomonas putida. In this previous work it became apparent that the mass transfer rate of the hydrophobic naphthalene was the rate limiting factor in biodegradation, as the bacteria could degrade the naphthalene as fast as it entered solution. The challenge for the present research was to find a simple, inexpensive method for increasing the mass transfer rates within the framework of the previously successful reactor. <p>After some deliberation, the addition of inert particles (glass beads) was determined to be the preferred option to increase mass transfer. The inert particles visibly increased the turbulence in the reactor and significant increases in both mass transfer and bioremediation rates were achieved. The augmentation of mass transfer rates was found to be dependent on the type, size and relative loading of the particles. Two types of inert particles were investigated to increase mass transfer rates, spherical glass beads and Raschig rings. Glass beads were found to be far superior to Raschig rings for the intended purpose. Three sizes of spherical glass beads were then compared experimentally (1, 3, and 5mm). It was discovered that the 3mm beads were vastly superior to 1mm beads and 5 mm beads were slightly superior to 3mm beads. Different bead loadings (volume of particles / total working volume) were then explored with 10%, 25% and 50% bead loading investigated. Although slight increases in mass transfer were observed at higher bead loadings, the reduction in working volume for biodegradation meant that 50% was accepted as the optimum loading parameter. <p>The optimum conditions for maximum mass transfer occurred using 5 mm spherical glass beads at 50% loading. The increase in mass transfer and biodegradation rates compared to a traditional roller bioreactor were found to be 10 fold and 11 fold, respectively. The optimum mass transfer conditions were then applied to 2-methylnaphthalene with increases in mass transfer and biodegradation equal to 6 fold and 8 fold, respectively. The candidate bacteria used in this study was found incapable of degrading 1,5 dimethylnaphthalene although the mass transfer results demonstrate promise for the developed technology. To determine the effects of scale on the process, two larger reactors were finally studied. They were eight times and twenty-one times the size of the initial bioreactor. The process was shown to speed up at larger scale which shows great promise for future applications. The maximum degradation rate achieved in the larger reactor was 148 mgL-1h-1. This compares very well with the best result found in literature, 119 mgL-1h-1, which was achieved in a much more complex system. Clearly, the bead mill bioreactor designed during the present work is a simple concept that shows superior performance for the bioremediation of PAHs.

The shikimate pathway is a primary metabolic pathway involved in the synthesis of aromatic compounds in plants, fungi, apicomplexan parasites and microbes. The absence of this pathway in animals makes it ideal for the synthesis of antimicrobial compounds and herbicides. Additionally, its branching into indole hormone synthesis and phenylpropanoid secondary metabolism makes this pathway attractive for metabolic engineering. Here, the focus is on the first step of the shikimate pathway catalyzed by DAHP synthase. This step consists of the condensation of phosphoenol pyruvate and erythrose-4-phosphate to make DAHP, which undergoes another six catalytic steps to synthesize chorismate, the precursor of the aromatic amino acids. Arabidopsis thaliana contains three DAHP synthase isozymes, which are known to indirectly regulate downstream pathways in response to wounding and pathogen stress. The model presented here proposes that DAHP synthase isozymes are regulated by the end products tyrosine, tryptophan and phenylalanine.

The photodebromination of selected bromoarenes has been studied at 300 nm to determine the the possible mechanistic pathways leading to product. Irradiation of 4-bromobiphenyl (BpBr) at this wavelength leads to the product biphenyl. The quantum yield of intersystem crossing (0.98) and quenching studies with cis-1 ,3-pentadiene suggest that the reaction occurs only via the triplet state. The observed increase of quantum yield of reaction with increasing concentration of BpBr suggests formation of a triplet excimer between the triplet state and ground state of BpBr as the key reactive intermediate. The log of the rate constant for excimer formation showed a linear increase with solvent polarity. The correlation of rate constants for excimer formation with linear solvation energy parameters indicates only a weak polarization of the excimer species. In order to understand the extent of radical anion character in the excimer, the regiochemistry of the photo-debromination of 3,4-dibromobiphenyl (3,4-BpBr) was studied. 3,4-BpBr was irradiated with and without an electron donor (triethylamine). 3,4-BpBr was also chemically reduced with lithium biphenylide (LiDBB). The difference in the regiochemistries under these conditions has been interpreted in terms of reaction via a free radical anion in the case of reactions with triethylamine and LiDBB and a weakly polarized excimer in the case of direct irradiation. In order to understand the extent of polarization in the excimers and their conformation, it was decided to study systems where the structure of the molecule would fix the geometry of potential intramolecular charge-transfer complexes. Towards this goal, brominated [2.2]paracyclophanes, 4-bromo[2.2]para-cylophane (CpBr), pseudo-para-dibromo[2.2]cyclophane (ps-p-CpBr) and pseudo-ortho-dibromo[2.2]cyclophane (ps-o-CpBr), were studied. The effect of substitution pattern of the bromines on the efficiency of excimer formation was also investigated. The brominated [2.2]para-cyclophanes showed varying efficiencies of formation of intermolecular excimer and intramolecular charge-transfer. A probable conformation for the excimer from BpBr has been proposed based on these results. / Graduation date: 1991

The shikimate pathway is a primary metabolic pathway involved in the synthesis of aromatic compounds in plants, fungi, apicomplexan parasites and microbes. The absence of this pathway in animals makes it ideal for the synthesis of antimicrobial compounds and herbicides. Additionally, its branching into indole hormone synthesis and phenylpropanoid secondary metabolism makes this pathway attractive for metabolic engineering. Here, the focus is on the first step of the shikimate pathway catalyzed by DAHP synthase. This step consists of the condensation of phosphoenol pyruvate and erythrose-4-phosphate to make DAHP, which undergoes another six catalytic steps to synthesize chorismate, the precursor of the aromatic amino acids. Arabidopsis thaliana contains three DAHP synthase isozymes, which are known to indirectly regulate downstream pathways in response to wounding and pathogen stress. The model presented here proposes that DAHP synthase isozymes are regulated by the end products tyrosine, tryptophan and phenylalanine.

Polycyclic aromatic hydrocarbons (PAHs) and metals are co-contaminants of aquatic environments near industrial and urbanized areas. Mixtures could result in synergistic toxicity impairing macrophyte growth and potentially causing bioaccumulation and biomagnification throughout the ecosystem. In this study, combinations of 1,2-dihydroxyanthraquinone (1,2-dhATQ) and copper (Cu) at low concentrations synergistically inhibited Lemna gibba (duckweed) growth. Further analysis using fluorescence techniques showed an increase in reactive oxygen species (ROS) levels upon Cu exposures possibly through redox cycling in the chloroplasts. Pulse amplitude modulated (PAM) and fast repetition rate fluorometry (FRRF) indicated that plants exposed to 1,2-dhATQ had impaired photosynthetic electron transport that manifested as a decrease in the yield of photosynthesis and change in the redox status of the plastoquinone (PQ) pool. At the gene expression level acetyl coA carboxylase (ACC), a key enzyme in membrane repair, and serine decarboxylase (SDC), another enzyme needed for membrane repair were up-regulated in response to copper and 1,2-dhATQ, respectively. The mechanism for mixtures toxicity is thought to involve the reduced PQ pool which could serve as a source of electrons for copper redox cycling thereby increasing ROS production and causing synergistic growth inhibition. When the antioxidant glutathione (GSH) was added, copper toxicity was ameliorated but 1,2-dhATQ toxicity increased possibly through formation of reactive conjugates or suppression of the native antioxidant system. This study emphasizes that mixtures of toxicants at low concentrations can cause more biological damage than individual toxicants via alterations of the redox status and increases in ROS production.

Polycyclic aromatic hydrocarbons (PAHs) and metals are co-contaminants of aquatic environments near industrial and urbanized areas. Mixtures could result in synergistic toxicity impairing macrophyte growth and potentially causing bioaccumulation and biomagnification throughout the ecosystem. In this study, combinations of 1,2-dihydroxyanthraquinone (1,2-dhATQ) and copper (Cu) at low concentrations synergistically inhibited Lemna gibba (duckweed) growth. Further analysis using fluorescence techniques showed an increase in reactive oxygen species (ROS) levels upon Cu exposures possibly through redox cycling in the chloroplasts. Pulse amplitude modulated (PAM) and fast repetition rate fluorometry (FRRF) indicated that plants exposed to 1,2-dhATQ had impaired photosynthetic electron transport that manifested as a decrease in the yield of photosynthesis and change in the redox status of the plastoquinone (PQ) pool. At the gene expression level acetyl coA carboxylase (ACC), a key enzyme in membrane repair, and serine decarboxylase (SDC), another enzyme needed for membrane repair were up-regulated in response to copper and 1,2-dhATQ, respectively. The mechanism for mixtures toxicity is thought to involve the reduced PQ pool which could serve as a source of electrons for copper redox cycling thereby increasing ROS production and causing synergistic growth inhibition. When the antioxidant glutathione (GSH) was added, copper toxicity was ameliorated but 1,2-dhATQ toxicity increased possibly through formation of reactive conjugates or suppression of the native antioxidant system. This study emphasizes that mixtures of toxicants at low concentrations can cause more biological damage than individual toxicants via alterations of the redox status and increases in ROS production.

Polycyclic aromatic hydrocarbons (PAH) have been identified as a serious environmental problem. In past research it has been proven that naphthalene, the simplest PAH, could be biodegraded using roller bioreactors and Pseudomonas putida. In this previous work it became apparent that the mass transfer rate of the hydrophobic naphthalene was the rate limiting factor in biodegradation, as the bacteria could degrade the naphthalene as fast as it entered solution. The challenge for the present research was to find a simple, inexpensive method for increasing the mass transfer rates within the framework of the previously successful reactor. <p>After some deliberation, the addition of inert particles (glass beads) was determined to be the preferred option to increase mass transfer. The inert particles visibly increased the turbulence in the reactor and significant increases in both mass transfer and bioremediation rates were achieved. The augmentation of mass transfer rates was found to be dependent on the type, size and relative loading of the particles. Two types of inert particles were investigated to increase mass transfer rates, spherical glass beads and Raschig rings. Glass beads were found to be far superior to Raschig rings for the intended purpose. Three sizes of spherical glass beads were then compared experimentally (1, 3, and 5mm). It was discovered that the 3mm beads were vastly superior to 1mm beads and 5 mm beads were slightly superior to 3mm beads. Different bead loadings (volume of particles / total working volume) were then explored with 10%, 25% and 50% bead loading investigated. Although slight increases in mass transfer were observed at higher bead loadings, the reduction in working volume for biodegradation meant that 50% was accepted as the optimum loading parameter. <p>The optimum conditions for maximum mass transfer occurred using 5 mm spherical glass beads at 50% loading. The increase in mass transfer and biodegradation rates compared to a traditional roller bioreactor were found to be 10 fold and 11 fold, respectively. The optimum mass transfer conditions were then applied to 2-methylnaphthalene with increases in mass transfer and biodegradation equal to 6 fold and 8 fold, respectively. The candidate bacteria used in this study was found incapable of degrading 1,5 dimethylnaphthalene although the mass transfer results demonstrate promise for the developed technology. To determine the effects of scale on the process, two larger reactors were finally studied. They were eight times and twenty-one times the size of the initial bioreactor. The process was shown to speed up at larger scale which shows great promise for future applications. The maximum degradation rate achieved in the larger reactor was 148 mgL-1h-1. This compares very well with the best result found in literature, 119 mgL-1h-1, which was achieved in a much more complex system. Clearly, the bead mill bioreactor designed during the present work is a simple concept that shows superior performance for the bioremediation of PAHs.

<p>Dibenzothiophene (DBT) is a constituent of creosote and petroleum waste contamination, it is a model compound for more complex thiophenes, and its degradation by mixed microbial communities has received little attention. The chemical characteristics, environmental fate and ecotoxicology of DBT degradation products are not well understood. This research investigated DBT degradation in an enrichment culture derived from creosote-contaminated estuarian sediment using a suite of assays to monitor bacterial populations, bacterial growth, degradation products, DBT loss, and toxicity. Ultraviolet (UV) irradiation was evaluated as a sequential treatment following biodegradation. Additionally, to advance SYBR-Green qPCR methodology for characterizing mixed microbial communities, an alternative approach for evaluating qPCR data using a sigmoidal model to fit the amplification curve was compared to the conventional approach in artificial mixed communities. The overall objective of this research was to gain a comprehensive understanding of the degradation of a model heterocyclic PAH, DBT, by a mixed microbial community, particularly within the context of remediation goals.</p><p>DBT biodegradation was evaluated in laboratory scale cultures with and without pH control. The microbial community was monitored with 10 primer sets using SYBR-Green quantitative polymerase chain reaction (qPCR). Twenty-seven degradation products were identified by gas chromatography and mass spectrometry (GC/MS). The diversity of these products indicated that multiple pathways functioned in the community. DBT degradation appeared inhibited under acidic conditions. Toxicity to bioluminescent bacteria <italic>Vibrio fischeri</italic> more than doubled in the first few days of degradation, was never reduced below initial levels, and was attributed in part to one or more degradation products. UV treatment following biodegradation was explored using a monochromatic (254 nm) low-pressure UV lamp. While DBT was not extensively photooxidized, several biodegradation products were susceptible to UV treatment. At higher doses, UV treatment following DBT biodegradation exacerbated cardiac defects in <italic>Fundulus heteroclitus</italic> embryos, but slightly reduced toxicity to <italic>V. fischeri</italic>.</p><p>This research provides a uniquely comprehensive view of the DBT degradation process, identifying bacterial populations previously unassociated with PAH biodegradation, as well as potentially hazardous products that may form during biodegradation. Additionally, this research contributes to development of unconventional remediation strategies combining microbial degradation with subsequent UV treatment.</p> / Dissertation