Chemistry

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Open Access
Protein Structure Dynamics In Gram Negative Bacterial Secretion Systems
(2025-04-10) Nicholas Joseph Bragagnolo; Audette, Gerald F.
Bacteria use a variety of secretion systems that enhance their virulence. The conjugative type IV secretion system attributes transmission of mobile DNA elements in bacteria, whereas type IV pili are akin to the type II secretion system and are responsible for host adhesion and twitching motility. Understanding the structure and function of components in these systems has the potential to elucidate the mechanisms of bacterial commensalism and antagonism, and would aid in the development of inhibitors to bacterial pathogenesis by disrupting the tools virulent species use for stable, long-term infections. The entry exclusion protein of the F-like type IV secretion system, TraG, consists of a membrane-bound N-terminal domain and a periplasmic C-terminal domain, denoted TraG*. TraG* is essential in preventing redundant DNA transfer through interaction with a cognate TraS in the inner membrane of the recipient cell to prevent conjugation when the recipient cell carries the same plasmid. Using a multitude of biophysical methods including thermofluor, circular dichroism, collision induced unfolding mass spectrometry, and small angle X-ray scattering, the structural dynamics of TraG* was investigated. This allowed for the characterisation of the 50 N-terminal residues, the 77 C-terminal residues, and the TraS-interacting region of the protein as highly dynamic, providing evidence for the mechanism of the long-distance interactions made by TraG. This dissertation also describes the 1.3 Å crystal structure of the N-terminally truncated Type IV pilin of Pseudomonas aeruginosa from strain P1 (ΔP1), the first structure of its phylogenetically linked group (group Ia) to be reported. The structure was solved from X-ray diffraction data collected 20 years ago with a molecular replacement search model generated using AlphaFold. Comparisons of the ΔP1 structure to other Type IV pilins using ProSMART indicated that there are cases of higher structural homology between different phylogenetic groups of P. aeruginosa than there are between pilins from the same phylogenetic group, indicating that the classification of pilins based on structural homology may be necessary in developing anti-virulence drugs and vaccines.
Open Access
A Passive Air Sampling Technique For Ultra-Trace Quantitation Of Gas-Phase Perfluoroalkyl Carboxylic Acids
(2024-11-07) Vanhauwaert, Eric; VandenBoer, Trevor ; Young, Cora
Perfluoroalkyl carboxylic acids (PFCAs) are persistent chemicals distributed ubiquitously in the environmental despite their anthropogenic origin. Recent studies have shown that atmospheric transport is an important transport mechanism for PFCAs. Despite this, measuring mixing ratios of gaseous PFCAs in the atmosphere has been an analytical challenge, limiting the availability of atmospheric measurements. A new nylon-based passive air sampling (PAS) technique addresses many of these challenges by offering a cost effective, low-labour solution. These samplers were calibrated for trifluoroacetic acid (TFA) in an atmospheric chamber and field measurements were compared to measurements from an ambient ion monitoring ion chromatography mass spectrometer, where good agreement was found. They were also deployed in various locations in Canada for gas-phase PFCA measurements which revealed many different trends, including the wastewater treatment plant as a point source of gas-phase TFA. These measurements have enhanced our understanding of PFCAs with calibrated, cost-effective samplers capable of ultra-trace measurements.
Open Access
The Utility Of Hydrogen Deuterium Exchange Mass Spectrometry For Characterization Of Proteins And Their Interactions.
(2024-11-07) Wolf, Esther Zvia; Wilson, Derek
Mass spectrometry (MS) offers immense versatility to the structural biology field. Particularly, Hydrogen Deuterium Exchange MS (HDX-MS) can overcome obstacles encountered by more traditional, higher resolution structural characterization methods. What happens when your protein system is just too big, dynamic, dilute, or poorly soluble? Recent advancements in structural prediction and HDX-MS automation have enabled some of the fastest data turnaround to date. This work follows many different scenarios where HDX-MS and MS have been employed, including protein-peptide interactions of the Bcl-2 family, affinity ranking and prediction of a focused small molecule library targeting WDR5, USP3 binding characterization of a small molecule screen hit, the detection of proteolysis targeting chimera (PROTAC)-induced ternary complex formation, and epitope mapping of antibodies targeting the SARS-CoV-2 Spike protein, all made possible by the collaboration of the Dickinson Lab (University of Chicago), Icosagen (Estonia), the Ingerman-James Lab (University of North Carolina at Chapel Hill), and the Structural Genomics Consortium.
Open Access
Exploring the heterogeneous uptake of gas-phase PFCAs to the condensed phase
(2024-07-18) Indos, Dylan James; Young, Cora
Perfluoroalkyl carboxylic acids (PFCAs) are a class of per- and polyfluoroalkyl substances (PFAS) which are completely fluorinated, accompanied by a carboxylic head functional group. With varying toxic effects on animals, plants, and humans, it has become imperative to understand the fate of these compounds in the atmosphere. The most important aspect in helping define a chemical’s environmental fate is its physicochemical properties and partition tendencies. Partitioning of a gas phase chemical from air to water begins with the heterogeneous uptake of the gas-phase molecule onto the liquid’s surface. Heterogeneous reactions are important in identifying sinks and the fate of atmospheric gas-phase chemicals. Currently there exists no experimental gas-phase uptake data for gas-phase PFCAs onto liquid surfaces which limits our understanding on their fate, movement, and sinks in the atmosphere. This work will aim to fill in gaps of knowledge for the heterogeneous uptake of a short chain PFCA, TFA, onto various liquid surfaces.
Open Access
Crystalline and amorphous cobalt-based metal oxides for water oxidation reaction – structure, composition, morphology and electrochemical
(2024-07-18) Thekkoot, Sreena Raju; Morin, Sylvie
Cu, Ni, and Fe-substituted cobalt-based amorphous materials (CoOx, CuCo2Ox, Ni0.5Cu0.5Co2Ox, Fe0.1Cu0.9Co2Ox, and Fe0.1Ni0.9Co2Ox) were prepared by thermal decomposition method and employed as electrocatalysts for oxygen evolution reaction (OER). These materials were characterized by scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), X-ray diffraction (XRD), transmission electron microscopy (TEM), high-resolution transmission electron microscopy (HR-TEM) and X-ray photoelectron spectroscopy (XPS). CuxCo3-xO4 (0 ≤ x ≤ 1), NixCu1-xCo2O4 (0 ≤ x ≤ 0.75) and FeyCux-yCoxO4 (x=1, 1.5 and y = 0.1, 0.15) were also by thermal decomposition method. Surface area measurements were performed using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). The catalytic activity of spinel oxides and amorphous metal oxides for OER was analyzed by CV, while kinetic studies were carried out using a rotating disc electrode (RDE). TEM and HR-TEM analysis were performed on polarized and nonpolarized CuCo2O4. The study clearly indicates that the as-prepared CuCo2O4 possesses a higher degree of crystallization compared to the polarized sample. The crystallite size of the samples measured by TEM and XRD analysis is very similar. Surface area measurements indicate that the incorporation of Cu and Ni increased the surface area of amorphous and crystalline samples. The incorporation of Fe increased the surface area of the amorphous samples, while an opposite trend was observed in the case of spinel oxides. XPS analysis indicates that the surface of both amorphous and crystalline samples contains different species and the metal ions exist in different oxidation states. Catalytic activity was measured as a function of geometric and real surface area. Both amorphous and spinel oxides were found to be active for OER. However, when corrected for real surface area, spinel oxides provided much higher current density compared to amorphous samples. Our study indicates that spinel oxides outperform amorphous samples for OER.
Open Access
Structural Modifications of Dithienophospholes for Applications as Functional Materials
(2024-03-16) Asok, Nayanthara; Baumgartner, Thomas
Recent breakthroughs in synthetic chemistry have revolutionized main-group molecules, elevating them from mere laboratory curiosities to powerful materials with broad applications. A primary focus has been on electron-accepting or -deficient materials, driven by their historical limitations in availability and stability, which have hindered practical applications. The incorporation of heavier main-group elements, including Si, Ge, P, As, Sb, Bi, S, Se, and Te, has proven advantageous for electron-accepting materials due to their polarizable molecular orbitals (MOs) readily accessible to electrons and nucleophiles. This foundation has spurred research in materials chemistry across various applications, encompassing optoelectronic devices (OLEDs, OPVs), energy storage (batteries, capacitors), fluorescent sensors (biological, physiological), catalysis, and synthesis. Among main-group-element-based materials, organophosphorus compounds hold a privileged status, with their frontier orbitals easily modifiable through chemical, structural, or electronic means at the phosphorus center itself, without necessitating kinetic stabilization. The five-membered phosphorus-based heterocycle, phosphole, is particularly captivating in this context. Extensive studies have unveiled the intricate σ*-π* interaction within phospholes, endowing them with intriguing electron-accepting properties, while preserving morphological and physiological stability for practical utilization. Furthermore, phosphorus introduces easily accessible, low-lying antibonding orbitals, leading to Lewis acidic phosphorus species, a departure from the conventional perception of phosphorus as an electron-rich element. These species exhibit unconventional chemical reactivity through hypervalency. This thesis advances conjugated materials by employing the unique structures and electronics of organophosphorus compounds. It discusses how these materials can be harnessed to design functional materials with exceptional electronic, chemical, and structural properties, contributing to the realm of functional materials.
Open Access
Machine Learning Estimation of Reaction Energy Barriers and its Applications in Astrochemistry
(2024-03-16) Ji, Hongchen; Fournier, Rene Andre
We developed a machine learning model for fast estimating reaction energy barriers. The model was trained on data for 11,730 elementary reactions and barriers computed with an estimated accuracy of 2.3 kcal/mol by Grambow and coworkers. Although it was trained, and then applied, only for reactions involving atoms of H, C, N and O, our model can readily be generalized to molecules and reactions involving other elements. We designed and tested many molecular representations. Our best model has 363 features calculated from the chemical composition, structure, and energy of products and reactants that can all be obtained at a small computational cost. A Kernel Ridge Regression with Laplacian kernel was found to give the best fit to the data. It makes predictions with a mean absolute error of 4.1 kcal/mol for barriers smaller than 40 kcal/mol. We used this machine learning model to estimate the barriers of 136,081 hypothetical association reactions between molecules known to exist in the circumstellar envelopes and interstellar medium, where temperatures range between 10 and 150 K. A screening procedure identified reactions likely to occur (those with near zero barriers) that could play a role in the formation of relatively complex organic molecules (those with at least one double bond and containing at least one atom each of the elements H, C, N and O). Reactions identified as the most promising were investigated with density functional theory and coupled cluster quantum chemical methods to obtain reaction pathways and energies of reactants, intermediates, transition states and products with an accuracy of roughly 1 kcal/mol. We found no barrierless reaction but found two reactions with low barriers leading to the formation of N-methyleneformamide (CH2NCHO) and imine acetaldehyde (NHCHCHO). These molecules have not yet been observed in space. The low barriers that we calculated, and the possibility that these reactions would be facilitated by adsorption on ice covered dust grains in the interstellar medium, suggest that these two molecules may exist in space and could be detected by determinations of rotational constants from spectroscopic lines in the microwave region.
Open Access
Structural and Functional Insights into the F Plasmid Type IV Secretion System proteins TrbI and TrbB
(2024-03-16) Apostol, Arnold Jan; Audette, Gerald F.
Bacteria have evolved elaborate mechanisms to thrive in stressful environments. One mechanism that bacteria utilize are secretion systems that can traverse protective lipid cell membranes and serve as mediators for a diverse set of goals, including the secretion of toxins implicated with target host pathogenesis. F-like plasmids in gram-negative bacteria encode for the multi-protein Type IV Secretion System (T4SSF) that is functional for bacterial proliferation and adaptation through the process of conjugation. The periplasmic protein TrbB is believed to have a stabilizing chaperone role in the T4SSF assembly, with TrbB exhibiting disulfide isomerase (DI) activity. In the current report, we demonstrate that residues W57-K181, which include the active thioredoxin motif, are sufficient for DI activity. Moreover, a structural model of GST-TrbBWT based on ColabFold-AlphaFold2 and Small Angle X-Ray Scattering data indicate that TrbBWT’s N-terminus is disordered, and this disordered nature likely contributes to the protein’s dynamicity and recalcitrance to crystallization. A truncation construct, TrbB57-181, was designed and found to exhibit higher physicochemical stability using 1H-15N Heteronuclear Single Quantum Correlation spectroscopy and Circular Dichroism spectroscopy. Binding studies of TrbB and other T4SSF proteins TrbI and TraW were performed, and results do not support the inference of a stable complex forming in vitro. Comparative studies of TrbB, TraF, and TrbI also provide insights into the structure of these T4SSF component proteins. Lastly, crystallization trials of GST-TrbBWT and GST-TrbI provide leads for future crystallization campaigns.
Open Access
Alkylidene Dihydropyridines as Useful Intermediates for Functionalization of 4-Alkylpyridines
(2024-03-16) Shi, Jiaqi; Orellana, Arturo
Pyridines are widely used and extensively studied heterocyclic compounds in industry and academia. Developing mild and selective methods for functionalizing pyridines would facilitate the synthesis of more structurally diverse pyridine derivatives, which could aid the development of potential drug candidates and streamline the drug discovery process. Through a “soft-enolization” logic, 4-alkylpyridines can be readily converted to corresponding alkylidene dihydropyridines (ADHPs), enabling mild and selective palladium-catalyzed allylation and dehydrogenation, as previously shown by our group. During the mechanistic study of the allylation reaction, prompted by the low enantioselectivity of pyridine allylation using optically active ligands, our group demonstrated that ADHPs are “soft” nucleophiles towards the allylpalladium(II) complex. We thus began employing this class of intermediates as soft nucleophiles in other reactions. Here I will first discuss the work towards enantioselective allylation of 4-alkylpyridines. Then I will describe the conjugate addition of ADHPs to a,b-unsaturated ketones that are activated by triethylsilyl triflate. Finally, I will introduce the approach to unite piperidine and pyridine with a carbon atom through addition of ADHPs to protonated pyridines with subsequent transfer hydrogenation reaction.
Open Access
Advancing Kinetic Capillary Electrophoresis for High-Efficiency Screening of Oligonucleotide Libraries in Drug Discovery
(2024-03-16) Le, An Thi Hoai; Krylov, Sergey N.
Identifying protein binders is the first step in drug discovery. The combinatorial approach, in which a library of compounds is subjected to affinity screening against a target protein, is a major way for identifying protein binders. Oligonucleotide libraries constitute the largest source of material for such affinity screening. Selecting protein binders from such libraries requires a highly efficient method for separation of protein−oligonucleotide complexes from the excess of unbound oligonucleotides. Kinetic Capillary Electrophoresis (KCE) is a rapidly advancing technique in affinity applications. It reportedly has superior partitioning efficiency, but screening oligonucleotide libraries by KCE has many challenges which must be addressed before KCE can compete with conventional surface-based screening. The goal of my research is to transform KCE into a versatile technology for screening protein binders from oligonucleotide libraries. To overcome the nonbinder background issue in KCE-based partitioning, I introduce Ideal-Filter Capillary Electrophoresis (IFCE), where binders and nonbinders travel in opposite directions. While successfully implementing IFCE conditions to be compatible with physiological buffers, a remarkable partitioning efficiency of 109 is achieved, the highest recorded so far. Further, I develop the first quantitative characterization of all KCE-based partitioning modes for a diverse range of protein target sizes. This systematic analysis provides guidance for CE users on selecting appropriate KCE-based partitioning conditions for a given protein target. Next, I conduct the first experimental investigation into the influence of target concentration on binder selection, aiding researchers in identifying an appropriate range of target concentrations to prevent selection failures. Finally, I gather insights from all these works to demonstrate the first highly efficient KCE-based selection of protein binders from DNA-encoded library of small molecules (DEL). This pioneering achievement showcases the capabilities of KCE-based partitioning within the context of DEL-based drug discovery, marking a significant advancement in the field.
Open Access
Robust MircroRNA detection by Capillary Electrophoresis
(2023-12-08) Hu, Liang; Krylov, Sergey N.
MicroRNA (miRNA) are short single-stranded RNA molecules that function as post-transcriptional regulators in gene expression, The abnormal expression of small subsets of miRNAs, termed “miRNA fingerprints”, have been found as potential biomarkers in diagnosis of various diseases including cancers. However, none of miRNA-based biomarkers has been approved yet for clinical use due to the lack of a robust quantitative method for miRNA analysis. To address this issue, there have been significant efforts towards developing robust quantitative miRNA detection methods. Among a variety of initiatives, DQAMmiR is a promising approach developed by our lab which is capable of direct, quantitative analysis of multiple miRNAs in capillary electrophoresis (CE). In my PhD project, I have been focusing on investigating and improving the robustness of DQAMmiR to increase its applicability for practical use. Firstly, I demonstrated that DQAMmiR is capable of directly quantitating miRNA in crude cell lysates, indicating its robustness to the change of sample matrices, revealing its great potential to be used to analyzing miRNAs in biological samples directly without the need for complex sample processing. Secondly, I developed a second-generation DQAMmiR by replacing the ssDNA probes with the electrically neutral peptide nucleic acid (PNA) probes, which further improves the robustness of the assay by removing the need for single-stranded DNA binding protein (SSB) in the assay. Thirdly, to increase the applicability of the assay, I introduced a new sample preconcentration technique based on the integration of field-amplified sample stacking (FASS) and isotachophoresis (ITP) prior to CE separation in DQAMmiR, which successfully decreases its limit of quantification (LOQ) by 140 times. Lastly, I investigated the applicability of DQAMmiR for analyzing urinary miRNAs. I demonstrated that the concentration ratio of two miRNAs, miR-16 and miR-21, could be relative stable from different urine samples collected from the same person, although the individual concentrations of these miRNAs varied significantly in these samples. These findings suggests that the ratiometric measurements between multiple miRNAs in urine would serve as better biomarkers than individual miRNA concentrations for clinical diagnostic tests. And DQAMmiR would be an ideal analytical methodology for the development and use of such tests.
Open Access
Instrumental Analysis of Pasylated Asparaginase, JZP-341, A Pre-Clinical Drug Candidate
(2023-12-08) Ghaffari, Shakiba; Krylov, Sergey N.
Bio-betters are second-generation biopharmaceutical drugs that aim to improve the original drug’s pharmacokinetic or pharmacodynamic properties through minor physical modifications. Bio-betters require extensive characterization. Here, we investigate: JZP-341, a long-acting asparaginase bio-better used to treat leukemia. JZP-341 has a disordered proline-alanine-serine (PAS) tail that increases the drug’s size and thereby its serum half-life. A long serum half-life decreases the dosage frequency, providing more freedom to the patient. We assess the structural heterogeneity, charge heterogeneity, and enzyme kinetics of JZP-341 to better understand the effects of the PAS tail on the drug via the methods of capillary electrophoresis (CE), mass spectrometry (MS), and chromatography. We observe size heterogeneity and charge heterogeneity. We also developed a native capillary gel electrophoresis assay and an automated label-free CE-MS enzyme activity assay to study JZP-341. A detailed understanding of the role of PAS tail on JZP-341 requires further assay development and sophisticated equipment.
Open Access
Sustainable Synthesis of 4-acylpyridines through air oxidation of dearomatized 4-alkylpyridines
(2023-12-08) Puzhitsky, Matthew; Orellana, Arturo
Nitrogen containing heterocycles such as pyridines are a frequently encountered motif found in small molecule drugs approved by the FDA. Due to their prevalence in the pharmaceutical industry, green and effective synthetic routes towards these molecules are appealing. We detail a novel synthetic route towards 4-acypyridines via the ambient oxidation of alkylidene dihydropyridines (ADHPs). We then establish the robust and chemoselective nature of this oxidation by exploring substrates with various functional groups and multiple pyridylic sites. Finally, a series of time-course NMR studies reveal how to manipulate the stability of ADHPs by varying the nature of the chloroformate used to make these conjugated intermediates.
Open Access
Mild Electrocyclization of Heptatrienyl Anions
(2023-12-08) Guo, You Xuan Bill; Orellana, Arturo
Carbocycles are prevalent in natural products and drug-like molecules. Despite their importance in medicinal chemistry, functionalized cycloheptanes remain a challenging scaffold to access. We have taken a keen interest in developing a strategy for cycloheptane formation through electrocyclization of heptatrienyl anions. Historically, this strategy has faced many challenges, including the use of strong bases to form the heptatrienyl anion, the lack of product selectivity, and low yields. We aim to overcome these deficiencies by the strategic use of electron withdrawing groups to activate both the heptatriene towards deprotonation and stabilize the resultant heptadienyl anion. In this report we detail the mild catalytic electrocyclization of functionalized heptatrienyl anions.
Open Access
Exploring selected modern Mass Spectrometry techniques in applied sciences
(2023-12-08) Taemeh, Maryam Yousefi; Ifa, Demian R.
Mass Spectrometry (MS) is a powerful analytical technique that has revolutionized our ability to analyze complex mixtures and has diverse applications in various scientific disciplines. It is widely used in chemistry and biology to determine the molecular composition, structure, and quantification of samples. In this dissertation, we aimed to explore novel applications of mass spectrometry techniques, including electrospray ionization (ESI) and tandem mass spectrometry (MS/MS), as well as mass spectrometry imaging (MSI) techniques such as matrix-assisted laser desorption/ionization (MALDI) and desorption electrospray ionization (DESI). The objective was to strengthen our knowledge and skills in various MS methodologies and gain practical experience in using these techniques, for developing methods to chemically profile, separate, detect, map, and quantify a wide range of targeted and untargeted analytes from complex matrices. Given the versatility of MS-based techniques, we focused on specific applications within this field. First and foremost, forensic and pharmaceutical cannabis-related applications require accurate and sensitive analytical methods. Hence, we developed and validated a DESI-MS method for detecting cannabinoids in cannabis-infused chocolate, utilizing TLC-DESI-MS and QuEChERS extraction for THC quantification. Additionally, we investigated metabolic differences associated with soybean alleles (QTLs E and M) conferring resistance to leaf-chewing insects. Non-targeted mass spectrometry reveals distinct metabolite sets influenced by the QTLs, highlighting daidzein as a significant marker. This suggests a novel mechanism impeding the insects' ability to evolve tolerance. Moreover, we explored the sublimation technique for the application of a matrix coating in MALDI-MSI. Specifically, we investigated the optimal conditions for the sublimation of the 5-chloro-2-mercaptobenzothiazole (CMBT) matrix on mouse kidney samples. We successfully obtained high-quality MALDI-MSI images of phospholipids (PC, PG) and phosphatidylinositol (PI) in mouse kidney sections, providing valuable spatially resolved information. Lastly, we discussed the importance of phosphatidylinositols (PI) and phosphoinositides and their involvement in various diseases, emphasizing the need to optimize the MALDI-matrix type and thickness for enhanced sensitivity in detecting these molecules. The label-free data obtained through this optimized approach holds the potential for investigating the relative m/z factors associated with PI-related pathogenesis. Overall, this dissertation demonstrated the versatility and potential of mass spectrometry techniques. The findings lay the groundwork for future research in the field of MS analysis, particularly in the detection, quantification, and understanding of phosphatidylinositols and phosphoinositides in biological systems and disease pathologies.
Open Access
Investigating the Structural Dynamics of Therapeutic Proteins using Time resolved Hydrogen deuterium Exchange Mass Spectrometry
(2023-12-08) ElSabaawy, Marwa Abdalla Abdelmaksoud Mohamed; Wilson, Derek
Biopharmaceutical products have drastically grown as essential treatments in clinical settings. Hence, the drug industry is moving towards protein-based therapeutics with increasing demands for parallel advancement in bioanalytical technologies. This thesis centers around displaying the versatility of coupling front-end analytical methods to mass spectrometers. We implement time-resolved electrospray ionization hydrogen-deuterium exchange (TRESI-HDX) and ion mobility spectroscopy (IMS) MS for uncovering new and exciting insights into therapeutic protein dynamics and protein-substrate interactions on millisecond timescale. Using automated continuous flow injection (CFI) has enhanced the analytical throughput of our results. Collectively, the implications for these results widen the fundamental understanding of the studied proteins. Understanding the crosstalk between dynamics and function may help scientists to design new and improved therapeutics.
Open Access
Quantification of Methane Emissions by Surface Mass Balance Method
(2023-08-04) Kim, Yeuhyun; McLaren, Robert
This thesis presents a surface mass balance method as a cost-effective top-down technique to conveniently validate the bottom-up inventories. Mobile methane measurements were performed for two large landfills, Keele Valley Landfill and Greenlane Landfill and the city of Sarnia which included petrochemical industries and residential areas by employing a Cavity Ringdown Spectrometer (CRDS) mounted in a vehicle to capture downwind enhancements of methane. Methane emission from the Greenlane landfill was estimated to be 3300 ± 730 kg h-1 by a mass balance approach. An estimation by a gaussian dispersion model provided a similar emission rate of 3320 ± 250 kg h-1. The regression analysis of the mixing ratios of CO2 and CH4 showed positive correlation with an average molar ratio of 0.99 ± 0.04 mole mole-1 which was used to estimate CO2 emission to be 7600 ± 1700 kg h-1. The city of Sarnia including its industrial complex and residential areas showed a total methane emission rate of 2450 ± 560 kg h-1. It is estimated the city emits 21.5 ± 4.9 kt CH4 annually accounting for 45% of Ontario’s oil and gas methane emission. These estimated source rates from facilities were consistently 9-10 times greater than the GHGRP estimates. The discrepancies confirmed in the study emphasizes that it is significant to reconcile top-down measurements with the bottom-up inventories to provide a more accurate understanding of methane sources and sinks in Canada.
Open Access
Nanocrystals for Photocatalysis and Imaging Applications
(2023-08-04) Malile, Brian; Chen, Jennifer I-Ling
Nanocrystals, contrary to the bulk counterparts, can exhibit size-dependent optical, electronic, magnetic, and catalytic properties. These materials can be tailored for specific applications including sensing, bioimaging, drug delivery, optoelectronics, and catalysis. This dissertation explores two types of nanocrystals, namely CdS-based quantum dots (QDs) and DNA-conjugated gold nanoparticles (DNA-AuNPs), as photocatalysts for reductive organic transformations and high mass probes for the emerging Imaging Mass Cytometry bioanalytical platform, respectively. QDs are zero-dimensional semiconductor nanocrystals with attractive properties arising from quantum confinement effects. The tunable absorption and emission profiles make QDs desirable candidates in display technologies, lasing, and solar energy applications. On the other hand, non-radiative photophysical processes enhanced by quantum confinement in QDs are underutilised and often perceived as undesirable. One such process – Auger relaxation – can produce hot electrons with high reducing power and can be amplified further by doping the nanocrystal with Manganese (II). Part one of this thesis examines the photoreduction capabilities of Mn2+-doped CdS/ZnS core/shell QDs (Mn:CdS/ZnS QDs). The doped QDs were implemented as photocatalytic coatings on reaction vessels, and several model organic reactions were evaluated including the 6-electron reduction of nitrobenzene to aniline that reached an overall internal quantum efficiency of ~3%. The findings demonstrate several-fold increase in the photoreduction efficiency of Mn:CdS/ZnS over undoped CdS/ZnS QDs, and the film set up allows for facile post-reaction workup and a range of solvent compatibility. Additionally, surface characterizations were performed to probe the changes and address the reusability of the QDs. Lastly, the initial implementation of QD coatings in flow reactors showed success. This work presents new opportunities and diversifies the toolbox of heterogeneous photocatalysts for prospective use in organic reactions. Imaging Mass Cytometry (IMCTM) is a multiparametric imaging technique that utilizes metal-tagged antibodies as probes for investigating subcellular components via mass spectrometry. However, low-abundant cellular components can generate weak or no signals due to the small number of antibodies that bind to them. In the second part of this thesis, DNA-functionalized gold nanoparticles, each comprising >10 000 Au atoms, were examined as high mass probes for targeting low abundant microRNA. The interaction between the DNA strands on the AuNPs and microRNA-210, a biomarker for preeclampsia and hypertensive diseases, leads to the accumulation of DNA-AuNPs in cells as readily imaged with IMC. The results from IMC corroborated with traditional fluorescence-based methods, but with an enhanced sensitivity of a thousand-fold. This work is the first demonstration that DNA-AuNP can serve as high mass probes in IMC for detecting low-abundant nucleic acids.
Open Access
An Evaluation of the Contributions of Short and Long Range Pollutant Transport on Southern Ontario's Atmospheric Sulfur Content
(2023-08-04) Pritchard-Oh, Austin Michael; McLaren, Robert
Sulfur dioxide (SO2), particle sulfate (p-SO4-2), and total sulfur (SOx) were evaluated at York University, in Toronto, Canada, using a ThermoFisher SO2 analyzer and an Aerodyne Aerosol Mass Spectrometer. These measurements were compared with a dataset from 2002-2020, which demonstrated a downward trend in atmospheric sulfur. Toronto’s atmospheric sulfur decline has plateaued in recent years. Current mean SO2 was 0.37 ppb, p-SO4-2 was 0.60 μg/m3, and SOx was 1.6 μg/m3. There were no significant seasonal or daytime/nighttime differences. The effects of pollutant transport and major contributors to Toronto’s sulfur were determined. Major contributors to SO2 included Hamilton metal refineries, Greater Toronto Area chemical production facilities, the Nanticoke petroleum refinery, and vehicle emissions. Major contributors to p-SO4-2 are Sarnia petroleum and chemical production, Sudbury metal refineries, and long-range transport from the Ohio River Valley. A field study mass-balance determination of Sarnia’s SO2 emissions was estimated at 0.18 tonnes/hr (4,300 kg/day).
Open Access
Selective Chemical Reactions for Nucleic Acid Sequencing and DNA-Encoded Library Synthesis
(2023-03-28) Mahdavi Amiri, Yasaman; Hili, Ryan
Part One: The ability to map methylation sites in the human genome and epitranscriptome has transformed our understanding of how these modifications govern and influence a host of cellular processes and human diseases. Amongst the most widely studied methylations is N6-methyladenine, known as 6mA in DNA and m6A in RNA. While traditional methods to sequence these modifications have depended on antibody pulldowns, chemistry-based approaches are often less sequence dependent, can work on either DNA or RNA, and thus can provide a robust, inexpensive, and universal sequencing approach. In part one of this thesis, the first chemistry-based single-nucleotide resolution sequencing method for the detection of N6-adenine methylation sites in DNA and RNA is presented. This method takes advantage of the chemoselective deamination of unmodified adenines under acidic nitrite conditions, resulting in a (d)A to (d)G transition, while leaving methylated adenine sites unaffected. As changes in N6-adenine methylation of RNA and DNA have been implicated in a range of human diseases, especially cancers, the method has been rapidly adopted by researchers globally as an affordable and straightforward sequencing approach to assist in understanding the role and impact of the epigenome and epitranscriptome on human health. The ability of this method to detect other nucleotide modifications was also evaluated and described. Part Two: DNA-encoded libraries (DELs) comprise millions to billions of small-molecules covalently linked to a unique DNA barcode that can be read using standard next-generation sequencing (NGS). This technology has revolutionized the field of drug discovery as a method to rapidly identify small molecules that can serve as novel leads for drug development. The success of a drug discovery campaign involving a DEL depends on the chemical diversity presented within the DEL; methods that can generate DELs with new molecular architectures and with greater chemical diversity are critically needed to advance drug discovery efforts both within industry and academia. To this end, the use of photoredox chemistry as a facile method to generate DELs with drug-like properties is presented as part two of this thesis. An efficient approach for the photoredox-catalysed hydroaminoalkylation between on-DNA secondary N-substituted (hetero)arylamines and vinylarenes is explored as a method to generate DELs with known bioactive architectures. The developed reaction proceeds efficiently with a broad and well-explored substrate scope, working best with electron poor to neutral vinylarenes. This method is well suited for the construction of DELs enabling an expansion of drug-like chemical space.