Graduate Theses & Dissertations

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effects of environmental variables and dissolved organic matter characteristics on the diffusion coefficient of dissolved organic matter using diffusive gradients in thin films
The efficacy of the diffusive gradients in thin films (DGT) passive samplers to provide accurate measurements of free metal ions and those complexed with dissolved organic matter (DOM) was investigated. DOM controls the diffusive properties of DOM-complexed metal species in natural systems. Knowing the diffusion coeiffiecent (D) for DOM of different molecular weights (MW) and the major environmental variables influencing D is critical in developing the use of DGT passive samplers and understanding labile species. D and MW were determined for natural and standard DOM. No noticeable changes in DOM MW were observed during the diffusion process, suggesting that DOM remains intact following diffusion across the diffusive gel. Data analysis revealed that MW had the greatest influence on D, with a negative relationship between D and MW, except in tidal areas where ionic strength influence on D was significant. This study provides further characterization of the variables influencing D using the DGT technique. Author Keywords: Diffusion coefficient, Diffusive gradients in thin films, Dissolved organic matter, Flow field-flow fractionation, Principal Component Analysis, UV-Vis Spectroscopy
effects of Dissolved Organic Matter (DOM) sources on Pb2+, Zn2+ and Cd2+ binding
Metal binding to dissolved organic matter (DOM) determines metal speciation and strongly influences potential toxicity. The understanding of this process, however, is challenged by DOM source variation, which is not always considered by most existing metal speciation models. Source determines the molecular structure of DOM, including metal binding functional groups. This study has experimentally showed that the allochthonous-dominant DOM (i.e. more aromatic and humic) consistently has higher level of Pb binding than the autochthonous-dominant DOM (i.e. more aliphatic and proteinaceous) by more than two orders of magnitude. This source-discrimination, however, is less noticeable for Zn and Cd, although variation still exceeds a factor of four for both metals. The results indicate that metal binding is source-dependent, but the dependency is metal-specific. Accordingly, metal speciation models, such as the Windermere Humic Aqueous Model (WHAM), needs to consider DOM source variations. The WHAM input of active fraction of DOM participating in metal binding (f) is sensitive to DOM source. The commonly-used f = 0.65 substantially overestimated the Pb and Zn binding to autochthonous-dominant DOM, indicating f needs to be adjusted specifically. The optimal f value (fopt) linearly correlates with optical indexes, showing a potential to estimate fopt using simple absorbance and/or fluorescence measurements. Other DOM properties not optically-characterized may be also important to determine fopt, such as thiol, which shows strong affinity to most toxic metals and whose concentrations are appreciably high in natural waters (< 0.1 to 400 nmol L-1). Other analytical techniques rather than Cathodic Stripping Voltammetry (CSV) are required to accurately quantify thiol concentration for DOM with concentration > 1 mg L-1. To better explain the DOM-source effects, the conditional affinity spectrum (CAS) was calculated using a Fully Optimized ContinUous Spectrum (FOCUS) method. This method not only provides satisfactory goodness-of-fit, but also unique CAS solution. The allochthonous-dominant DOM consistently shows higher Pb affinity than autochthonous-dominant DOM. This source-discrimination is not clearly observed for Zn and Cd. Neither the variability of affinity nor capacity can be fully explained by the variability of individual DOM properties, indicating multiple properties may involve simultaneously. Together, the results help improve WHAM prediction of metal speciation, and consequently, benefit geochemical modelling of metal speciation, such as Biotic Ligand Model for predicting metal toxicity. Author Keywords: Dissolved organic matter, Metal binding, Source, Windermere Humic Aqueous Model
Using ultra high-resolution mass spectrometry to characterize the biosorbent Euglena gracilis and its application to dysprosium biosorption
Euglena gracilis is an enigmatic and adaptable organism that has great bioremediationpotential and is best known for its metabolic flexibility. The research done in this dissertation addresses (1) how growth conditions impact cellular composition, and (2) how chemometric approaches (such as statistical design of experiments and artificial neural networks) are viable alternatives to the conventional biosorption models for process optimization. Using high-resolution mass spectrometry for biosorbent characterization is a powerful way to assess the chemical characteristics of lyophilized and fractionated cells with high precision, especially to screen for compound classes that may have potentiality for rare earth element removal. Growth conditions impacted cellular composition and separated size fractions of cells yielded different molecular/chemical properties as described by compositional abundances, thus different biosorptive potential. Untargeted analysis demonstrated that exponential dark-grown cells with glucose supplementation were abundant in polyphenolic- and carbohydrate-like compounds, molecular species highly involved in rare earth element binding. Light grown cells had more heterogeneity and the highest molecular weighted fractions from light grown cells (fraction D) had the most abundances of polyphenolic- and protein-like structures. Chemometric modeling used identified the best and worst conditions for iii dysprosium sorption and showed that pH had the most significant influence on bioremoval. Bioremoval ranged from 37% at pH 8 to 91% at pH 3 at Dy concentration ranging from 1 to 100 μg L-1. The work presented in the PhD dissertation will aid in understanding the chemical characteristics of biosorbents by using a Van krevelen analysis of elemental ratios whether algal cells are grown in different environmental growth conditions, or when algal cell are size fractionated. This is especially applied for the screening for metal binding potentiality to Dysprosium. Chemometric methods provide an alternative method for the investigating factors for bioremoval, and applications for process optimization and for real-world applications. This dissertation will aid in understanding chemical characteristics when a biosorbent is grown in a given condition and which factors are important for rare earth element (REE) bioremoval. The significance of this work aims to look for alternate ways to screen biosorbents and using a more efficient experimental design for REE bioremoval. Author Keywords: bioremoval, biosorption, chemometrics, dysprosium, euglena, mass spectrometry
Using Fluorescent Carbon Dots for Biosensing Applications of Amino Acids
Amino acids make up proteins, which are the building blocks of life. A balance of amino acids is needed to maintain a healthy state. Tyrosine (Tyr) is synthesized from the metabolism of phenylalanine, which is an essential amino acid, meaning it can only be obtained from the diet. It is related to many metabolic and neurodegenerative diseases. Tyr can undergo post-translational modifications such as phosphorylation and nitration, which are implicated in cancer and nitrative stress, respectively. Although there are many methods to detect Tyr and its analogues, phosphotyrosine (pTyr) and nitrotyrosine (nTyr), these methods are time-consuming, involve expensive instruments and involve tedious process. This research proposes a new type of nanomaterials, carbon dots (CDs), to detect these amnio acids. Data indicate that CDs can be used to detect nTyr with a limit of detection of 34 μM in the linear range of 20 - 105 μM. The amenability of CD-nTyr assay was also tested in various biological matrices and biological molecules and was shown to be sensitive to nTyr. Nitration of Tyr was carried out in the presence of sodium nitrite and hydrogen peroxide catalyzed by either Cu(II) or Fe(III) to mimic biological reactions and CDs were tested as both inhibitors and indicators of Tyr nitration. Although CDs did not inhibit the nitration reaction of Tyr, they did not serve as indicators of Tyr nitration due to the quenching of CDs by the nitrating agents. This shows the importance of using CDs to detect nTyr and further use it for biological applications to detect diseased states. Author Keywords: amino acids, carbon dots, nanomaterials, sensor, spectroscopy, tyrosine
Speciation of Aluminum and Zinc in Three Streams of a Forested Catchment of the Boreal Zone
This study presents a detailed assessment of the chemical speciation of aluminum and zinc in three streams of a small, acid-sensitive forested catchment on the southern edge of the Precambrian Shield. Speciation analysis was achieved using an in-situ analytical technique known as Diffusive Gradient in Thin film (DGT) which measures labile metals, and a predictive computer algorithm (WHAM VI) which calculates metal species concentrations. Three types of DGT with different metal scavenging capabilities were used and a total of 11 deployments performed across four seasons. WHAM VI predictions showed that the organic fraction of aluminum was the main contributor to the dissolved concentrations in the main inflow stream (PC1) (~ 80 %) and the lake's outflow (PCO) (~ 75%); in the upland stream (PC1-08) the inorganic fraction contributed ~ 75%. For zinc the free ion was the single most important contributor to the dissolved concentration (< 90%) in all three streams. A comparative study of the DGT and WHAM methods showed an agreement between their inorganic concentrations during the spring season. Both methods indicate the greatest environmental impact for Al takes place during snow melt period in PCO and PC1-08 and in the summer for PC1. The greatest environmental impact for Zn predicted with WHAM VI, occurs during the spring in all three streams. Author Keywords: Aluminum, DGT, Metal speciation, WHAM, Zinc
Size and fluorescence properties of allochthonous dissolved organic matter
Dissolved organic matter (DOM) is a mixture of molecules with dynamic structure and composition that are ubiquitous in aquatic systems. DOM has several important functions in both natural and engineered systems, such as supporting microorganisms, governing the toxicity of metals and other pollutants, and controlling the fate of dissolved carbon. The structure and composition of DOM determine its reactivity, and hence its effectiveness in these ecosystem functions. While the structure, composition, and reactivity of riverine and marine DOM have been previously investigated, those of allochthonous DOM collected prior to exposure to microbes and sunlight have received scant attention. The following dissertation constitutes the first in-depth study of the structure, composition, and reactivity of allochthonous DOM at its point of origin (i.e. leaf leachates, LLDOM), as detected by measuring its size and optical properties. Concomitantly, novel chemometric methods were developed to interpret size-resolved data obtained using asymmetrical flow field-flow fractionation, including spectral deconvolution and the application of machine learning algorithms such as self-organizing maps to fluorescence data using a dataset of more than 1000 fluorescence excitation-emission matrices. The size and fluorescence properties of LLDOM are highly distinct. Indeed, LLDOM was correctly classified as one of 13 species/sources with 92.5% accuracy based on its fluorescence composition, and LLDOM was distinguished from riverine DOM sampled from eight different rivers with 98.3% accuracy. Additionally, both fluorescence and size properties were effective conservative tracers of DOC contribution in pH-controlled mixtures of leaf leachates and riverine DOM over two weeks. However, the structure of LLDOM responded differently to pH changes for leaves/needles from different tree species, and for older needles. Structural changes were non-reversible. Copper-binding strength (log K) differed for the different fluorescent components of DOM in a single allochthonous source by more than an order of magnitude (4.73 compared to 6.11). Biotransformation preferentially removed protein/polyphenol-like fluorescence and altered copper-binding parameters: log K increased from 4.7 to 5.5 for one fluorescent component measured by fluorescence quenching, but decreased from 7.2 to 5.8 for the overall DOM, as measured using voltammetry. The complexing capacity of DOM increased in response to biotransformation for both fluorescent and total DOM. The relationship between fluorescence and size properties was consistent for fresh allochthonous DOM, but differed in aged material. Since the size and fluorescence properties of LLDOM are strikingly different from those of riverine DOM, deeper investigation into transformative pathways and mixing processes is required to elucidate the contribution of riparian plant species to DOM signatures in rivers. Author Keywords: Analytical chemistry, Chemometrics, Dissolved organic matter (DOM), Field-flow fractionation, Fluorescence spectroscopy, Parallel factor analysis (PARAFAC)
SARS-CoV-2 Protein-based Detection Using Localized Surface Plasmon Resonance
During the COVID-19 pandemic, nucleic acid and antibody-based testing methods were heavily relied upon, but can be costly, time-consuming and exhibit high false -negative and -positive rates. Thus, alternative strategies are needed. Viral antigens such as the SARS-CoV-2 spike (S) glycoprotein are critical in the function of the virus and useful as diagnostic biomarkers for viral infections. For biosensing applications, aptamers are suitable high-affinity and cost-effective binding partners for their specific targets. Using localized surface plasmon resonance (LSPR), real-time, rapid acquisition of results can be achieved, essential for improving the efficacy of a sensor. Herein, LSPR aptamer sensors were fabricated for the detection of the SARS-CoV-2 protein. Data indicate that the best performing aptasensor was the streptavidin-biotin sensor, while the current gold aptasensor exhibited lower sensitivity and the fabrication of the carboxyl aptasensor was unsuccessful. The S1 aptamer selectively bound the S1 protein with high binding affinity. Excellent shelf-life stability, reusability, and high recovery in complex matrices was also maintained. Additionally, a receptor binding domain (RBD) functionalized sensor was fabricated to examine the interactions with angiotensin converting enzyme 2 (ACE2), for future assessment of inhibitors used in drug therapies. Overall, LSPR has been demonstrated as a viable tool for measuring SARS-CoV-2 related aptamer-protein and protein-protein interactions, and this strategy may be applied to other viral or non-viral antigen targets. Author Keywords: Antigen-based Detection, Coronavirus, COVID-19, Inhibition, Localized Surface Plasmon Resonance, SARS-CoV-2
Purification and Identification of Selenium-containing C-phycocyanin from Spirulina
Selenium is an essential trace nutrient to many organisms, yet in high concentrations it is toxic. Organic selenium is more bioavailable to aquatic biota than inorganic selenium, but is usually found in much lower concentrations. Algae are known to biotransform inorganic selenium into several organo-selenium compounds, but it is unknown whether any of these bioaccumulate in the food chain. In this study, selenium was incorporated into the methionine residues of an algal photosynthetic protein, c-phycocyanin from Spirulina spp. The extent of selenium incorporation was quantified by inductively coupled plasma-mass spectrometry (ICP-MS), and the protein was identified using electrospray mass spectrometry (ES-MS). C-phycocyanin was isolated and purified from Spirulina with a final recovery of 20-30 % of the total c-phycocyanin present. Selenomethionine replaced 92.8% ± 1.22 of the methionine residues in c-phycocyanin when grown in 2.5 ppm sodium selenite. ES-MS was used to obtain protein spectra, and pure c-phycocyanin was identified. Data of full scans provided estimated masses of both protein subunits--α-chain measured at 18,036 Da; β-chain measured at 19,250 Da--close to the theoretical masses. Protein fragmentation by collision-induced dissociation and electron capture dissociation provided approximately 52 % amino acid sequence match with c-phycocyanin from Spirulina platensis. This study demonstrates the incorporation of selenium into an algal protein, and the identification of c-phycocyanin using electrospray ionization-mass spectrometry. Author Keywords:
Novel Silica Sol-Gel Passive Sampler for Mercury Monitoring in Aqueous Systems
A novel passive sampler for mercury monitoring was prepared using organosilica sol-gel materials. It comprises a binding layer with thiol groups for mercury complexation and a porous diffusive layer through which mercury can diffuse and arrive at the binding layer. Our study demonstrated that this new sampler follows the principle of passive sampling. The mass of mercury accumulated in the binding layer depends linearly on the mercury concentration in solution, the sampling rate and the exposure time. A typical sol-gel sampler is characterized by a diffusive layer of 1.2 &mum, in which mercury ions diffuse with a coefficient of D = 0.09~10-6 cm2/s. The capacity for mercury uptake is approximately 0.64 &mug/cm2. Mercury diffusion and binding in the passive sampler are independent of the type of mercury-chloride complex. Its sampling rate increases with increasing water turbulence and decreases with increasing DOM amount. The field trial of sol-gel sampler in Miller Creek shows the concentration gained from the sol-gel passive sampler is slightly lower than that from the spot sampling. Author Keywords:
Molecular Composition of Dissolved Organic Matter Controls Metal Speciation and Microbial Uptake
Aquatic contaminant mobility and biological availability is strongly governed by the complexation of organic and inorganic ligands. Dissolved organic matter (DOM) is a complex, heterogeneous mixture of organic acids, amino acids, lipids, carbohydrates and polyphenols that vary in composition and can complex to dissolved metals thereby altering their fate in aquatic systems. The research conducted in this doctoral dissertation addresses 1) how DOM composition differs between phytoplankton taxa and 2) how DOM composition affects metal speciation and its subsequent microbial bioavailability in laboratory and field conditions. To accomplish this, a series of analytical methods were developed and applied to quantify thiols, sulphur containing DOM moieties, and the molecular composition of DOM. The works presented in this thesis represents one of the first comprehensive and multipronged analyses of the impact of phytoplankton metabolite exudates on microbial metal bioavailability. This dissertation demonstrated the analytical versatility of high-resolution mass spectrometry as a tool for compound specific information, as well as having the capabilities to obtain speciation information of organometallic complexes. The work presented in this PhD strengthens the understanding compositional differences of both autochthonous and allochthonous DOM and their effects on metal biogeochemistry. Author Keywords: Dissolved Organic Matter, Mercury, Metal Accumulation, Phytoplankton, Spring Melts, Thiol
Investigating the sources and fate of monomethylmercury and dimethylmercury in the Arctic marine boundary layer and waters
Monomethylmercury (MMHg), the most bioavailable form of mercury (Hg) and a potent neurotoxin, is present at elevated concentrations in Arctic marine mammals posing serious health threats to the local populations relying on marine food for their subsistence living. The sources of MMHg in the Arctic Ocean surface water and the role of dimethylmercury (DMHg) as a source of MMHg remain unclear. The objective of this research was to determine the sources and fate of methylated Hg species (MMHg and DMHg) in the marine ecosystem by investigating processes controlling the presence of methylated Hg species in the Arctic Ocean marine boundary layer (MBL) and surface waters. A method based on solid phase adsorption on Bond Elut ENV was developed and successfully used for unprecedented measurement of methylated Hg species in the MBL in Hudson Bay (HB) and the Canadian Arctic Archipelago (CAA). MMHg and DMHg concentrations averaged 2.9 ± 3.6 (mean ± SD) and 3.8 ± 3.1 pg m-3, respectively, and varied significantly among sampling sites. MMHg in the MBL is suspected to be the product of marine DMHg degradation in the atmosphere. MMHg summer (June to September) atmospheric wet deposition rates were estimated to be 188 ± 117.5 ng m-2 and 37 ± 21.7 ng m-2 for HB and CAA, respectively, sustaining MMHg concentrations available for bio-magnification in the pelagic food web. The production and loss of methylated Hg species in surface waters was assessed using enriched stable isotope tracers. MMHg production in surface water was observed from methylation of inorganic Hg (Hg(II)) and, for the first time, from DMHg demethylation with experimentally derived rate constants of 0.92 ± 0.82 x 10-3 d-1 and 0.04 ± 0.02 d-1 respectively. DMHg demethyation rate constant (0.98 ± 0.51 d-1) was higher than that of MMHg (0.35 ± 0.25 d-1). Furthermore, relationships with environmental parameters suggest that methylated Hg species transformations in surface water are mainly biologically driven. We propose that in addition to Hg(II) methylation, the main processes controlling MMHg production in the Arctic Ocean surface waters are DMHg demethylation and deposition of atmospheric MMHg. These results are valuable for a better understanding of the cycle of methylated Hg in the Arctic marine environment. Author Keywords: Arctic Ocean, Atmosphere, Demethylation, Dimethylmercury, Methylation, Monomethylmercury
Identification and Quantification of Organic Selenium Species Produced by Microbiological Activity in Freshwater Environments
Despite being an essential nutrient at trace levels, selenium can be devastating to aquatic environments when present in excess. There is no apparent correlation between total aqueous selenium concentrations and observed toxic effects because bioaccumulation varies over several orders of magnitude depending on the chemical species of selenium and the biological species present in the lowest trophic level of the aquatic food chain. Despite being used in toxicity models due to its high bioavailability, free selenomethionine had not been found previously in the environment outside of a biological entity. Here, it is confirmed that selenomethionine is produced during the biological treatment of selenium-contaminated wastewater, and released in the effluent along with other discrete organic selenium species, including selenomethionine oxide. This identification followed the development of a rigorous preconcentration and cleanup procedure, allowing for the analysis of these organic selenium species in high-ionic strength matrices. A newly optimized anion-exchange chromatographic separation was coupled to inductively-coupled plasma mass spectrometry for the simultaneous quantification of these organic selenium species along with the more ubiquitous selenium oxyanions, selenite and selenate. This separation method was also coupled to electrospray tandem mass spectrometry for structural confirmation of selenomethionine and selenomethionine oxide. High resolution orbitrap mass spectrometry was used to identify another oxidation product of selenomethionine – a cyclic species which was tentatively identified, by coelution, in a selenium-contaminated river water sample. The production and release of selenomethionine, selenomethionine oxide, Se-(methyl) selenocysteine, and methyl selenic acid were observed for various laboratory algal cultures. Once the presence of free selenomethionine in a water system was confirmed, factors affecting its uptake into algal cultures were examined. The uptake of selenomethionine into Scenedesmus obliquus was noted to be significantly higher under low nitrate conditions, where it was incorporated into selenium-containing proteins more readily than at higher nitrate conditions where other metabolites were produced. With the increasing popularity of biological treatment systems for the remediation of selenium-contaminated waters, these observations, combined with existing knowledge, could be used to make predictions regarding the potential toxicity of selenium in various environmental scenarios. Author Keywords: bioremediation, electrospray mass spectrometry, inductively-coupled plasma mass spectrometry, selenium, selenoamino acids, selenomethionine

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