Chemistry - Theses, Dissertations, and other Required Graduate Degree Essays

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Development and validation of a LC-MS/MS method for the pharmacokinetic study of thiamet-G and its analogues in rat

Date created: 
2010-12-09
Abstract: 

Thiamet-G inhibits the activity of N-acetyl-β-glucosaminidase, a glycoside hydrolase known as OGA. A validated bioanalytical method has been developed to enable pharmacokinetic studies of Thiamet-G and its related analogues. The bioanalysis was carried out using high performance liquid chromatography (HPLC) coupled to a tandem mass spectrometer (MS/MS). In the MS/MS, multiple reaction monitoring (MRM) was used to monitor the transition of analyte parent ions to diagnostic daughter ions. The validated method utilized the Hypercarb SPE cartridge as the cleanup tool and the ZIC-HILIC column as the suitable stationary phase. The method was validated for linearity, specificity, accuracy, precision, recovery, matrix effect, stability, and sensitivity. Pharmacokinetic samples obtained from rats treated by oral gavage with Thiamet-G were subjected to analysis using the validated method. Thiamet-G was found to be absorbed with a C max of 370 ± 20 ng / mL and showed a t max of 2 h.

Document type: 
Thesis
File(s): 
Supervisor(s): 
David J. Vocadlo
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) M.Sc.

Development of methods for mass spectrometry analysis of proteins in supernatants harvested from in vitro human lung cells (A549)

Date created: 
2010-12-09
Abstract: 

Proteins are primarily responsible for functionalizing cells and, with respect to an initiative in soft ionization mass spectrometry (MS), the relative abundances of selected proteins have been used as indicators of normal or stressed physiological states. Methods developed for this thesis describe the use of matrix assisted laser desorption ionization time of flight (MALDI-ToF) MS to monitor the ion signals of C-X-C motif chemokine 5 and ubiquitin protein. Identification of these ion signals were carried out by performing additional experiments using liquid chromatography interfaced to an electrospray ionization (LC-ESI) equipped MS capable of tandem MS. A total of 78 proteins were identified by LC-ESI tandem MS, but not all corresponded to MALDI-MS data. In addition to the use of commercially available instrumentation, a separate study was performed to investigate the potential for an AC trap to generate molecular ions from a single levitated droplet having undergone Coulomb explosion.

Document type: 
Thesis
File(s): 
Supervisor(s): 
George Agnes
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) M.Sc.

Novel inhibitors of human intestinal N-terminal maltase-glucoamylase and influenza virus A neuraminidase

Author: 
Date created: 
2010-11-26
Abstract: 

This thesis work deals with the syntheses and biological evaluation of novel inhibitors of two clinically relevant glycosidases, namely, human intestinal N-terminal maltase-glucoamylase (ntMGAM) and influenza virus neuraminidase (NA, also known as sialidase). Kotalanol, a naturally-occurring sulfonium-ion glucosidase inhibitor isolated from Salacia reticulata, with potent inhibitory activities against rat intestinal glucosidases, has potential to become a lead candidate in the treatment of type 2 diabetes. However, the lack of information about its stereostructure posed a major challenge for its total synthesis and structure-activity relationship studies. In this thesis, we successfully established the absolute stereostructure of kotalanol and also completed its total synthesis and that of its de-O-sulfonated derivative. The synthetic route was also extended to obtain heteroatom analogues and the six-membered ring nitrogen analogues of kotalanol and their de-O-sulfonated derivatives, and the inhibitory activities of these compounds were studied against ntMGAM. Neuraminidase, a glycoprotein located on the surface of the influenza virus, plays a crucial role in the viral replication process and hence, has proven to be a useful target enzyme for the treatment of influenza infections. The X-ray crystal structures of certain subtypes of influenza group-1 neuraminidases (N1, N4 and N8 subtypes) revealed that there exists a cavity (known as the 150-cavity) near the active-site resulting from a loop of amino acid residues (known as the 150-loop) that adopts an unusual open-loop conformation. Accordingly, we report here the exploitation of the 150-cavity in the active-sites of group-1 neuraminidases for the design of new triazole-containing carbocycles related to oseltamivir. Inhibition studies with virus-like particles (VLPs) containing the influenza virus neuraminidase-1 (N1) activity indicate that several candidates are inhibitors, with Ki values in the 10-5 - 10-8 M range. The proposed binding model for the triazole series of compounds was confirmed using the X-ray crystallographic analysis of the N8 subtype complexed with two compounds from the triazole series. We have also assessed the crossreactivity of these novel inhibitors against human neuraminidase isoform-3 (NEU3) as well as their ability to inhibit the replication of two different strains of influenza virus A in vitro using a cell-based assay.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Dr. B. Mario Pinto
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) Ph.D.

Glycoproteins: biosynthesis, structure and biological functions

Date created: 
2010-11-25
Abstract: 

Glycoproteins are formed by the covalent attachment of monosaccharides or their oligomers to proteins. The intermediates through which glycoproteins are biosynthesized are nucleotide sugars. The research described within this thesis focuses on the use of chemical methods by which the structures of glycans (the carbohydrate portion of a glycoconjugate) can be experimentally manipulated. Most of the glycan-altering compounds discussed herein prevent the normal biosynthesis of glycoproteins either because of their metabolism into non-natural nucleotide sugars or by preventing the normal utilization of nucleotide sugars. The perturbation of glycan structures is one of the main ways in which the biological functions of glycans can be assessed. Three different classes of glycoproteins were investigated, namely, glycoproteins containing asparagine- or serine/threonine-linked glycans, and those containing only serine/threonine-linked N-acetylglucosamine residues. Two thiosugar-containing disaccharide analogues were discovered, in a cell-based screening assay, to significantly impair the biosynthesis of asparagine-linked glycans within treated cells. These compounds, generally called 5-thiomannosides, were extensively characterized in the context of their effects on proprotein convertases, a class of proteases that are responsible for the production of a wide variety of bioactive peptides. The 5-thiomannosides investigated inhibited the production of endorphin in pituitary cells and were used to demonstrate the importance of correct glycosylation on the regulated peptide-hormone-targeting inside cells. Furthermore, this class of compounds was found to inhibit another key proprotein convertase involved in cholesterol biosynthesis. Several different techniques were used to investigate the structures of these glycans isolated from treated cells, the most significant of which was capillary electrophoresis (CE). This technique was used to deduce the structures of glycans obtained from cells by comparison with known standards or on the basis of their sensitivity in various enzyme assays. Furthermore, CE methods for the analysis of nucleotide sugars or nucleotides, and carbohydrate-like natural products were developed. These methods were used to develop an assay for an important drug target in the bacterium Mycobacterium tuberculosis and to investigate the biosynthesis of unusual nucleotide sugars within cultured cells. Of particular interest was the biosynthesis of nucleotide sugars containing thiosugar moieties, that is, sugars that contain an endocyclic sulfur atom.

Document type: 
Thesis
File(s): 
Supervisor(s): 
B. Mario Pinto
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) Ph.D.

On the microstructure of PEM fuel cell catalyst layers

Author: 
Date created: 
2010-11-02
Abstract: 

In this work, the microstructure of conventional catalyst layers of PEM fuel cells (PEMFCs) is investigated and correlated to the catalyst layer water sorption and retention properties, electrochemical properties and fuel cell performance. Two types of conventional carbon support were chosen for investigation: Ketjen Black and Vulcan XC-72. The microstructure of carbon supports, Pt/carbon catalyst powders, and 3-component, Pt/carbon/ionomer catalyst layers is studied in order to evaluate the effects of carbon support microstructure and Nafion ionomer loadings on the resultant CL microstructure. The microstructure of the carbon support is found to be a significant factor in the formation of the structure in the 3-component CLs serving as a rigid porous framework for distribution of Pt and ionomer. It is found that deposition of Pt particles on Ketjen Black significantly changes its porosity possibly by forming at the mouth of the support’s micropores, thus affecting its effective microporosity, whereas Pt deposited on Vulcan XC-72 does not significantly affect the support’s microporosity. The co-deposition of ionomer in the CL strongly influences porosity of the catalyst layer, especially for pore sizes < 20 nm, which are ascribed to the pores within primary carbon particles (pore sizes < 2 nm) and to the pores within agglomerates of the particles (pore sizes of 2-20 nm). Ketjen Black and Ketjen Black-based CLs were found to posses higher water sorption and retention capacity than Vulcan-XC-72 based catalyst layers due to carbon microstructure. Electrochemical properties and overall fuel cell performance were found to be strongly dependent on carbon support microstructure, ionomer loading and water sorption and retention capacity of the catalyst layers. It is postulated that carbons with a pore size distribution in the mesoporous range are beneficial for improved Pt utilization, especially, for fuel cell operation in dry conditions. It was also found that in these CLs the ionomer loading can be significantly decreased without significant reduction in performance.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Steven Holdcroft
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) Ph.D.

Selective DNA dehybridization induced by laser irradiation on gold nanostructures

Author: 
Date created: 
2010-08-11
Abstract: 

The objective of this thesis is to explore the feasibility of using light to control the surface reactions of DNA-Au nanoconjugates. Two types of gold nanostructures (Au nanoparticles vs. Au/SiO2 nanoshells) were modified with DNA oligonucleotides with different sequences. Due to the variation of their surface plasmon resonance peaks (520 nm for nanoparticles, 1100 nm for nanoshells), laser beams with corresponding wavelengths were used to irradiate the mix samples. It has been demonstrated that the laser-induced (photo-thermal effect) dehybridization of the dsDNAs on these nanostructures is selective, i.e., we can either release the single strands from the nanoparticles or from the nanoshells in the same solution independently. The experimental conditions have been optimized based on the selection of irradiation time and laser power. The selective, “remote” control of DNA reactions on the surface of nanostructures has potential applications in various clinical areas such as drug delivery and in vivo diagnostics.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Hua-Zhong Yu
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) M.Sc.

Effect of the local environment on the structure and reactivity of metalloporphyrins

Author: 
Date created: 
2010-08-09
Abstract: 

Phosphonium ionic liquids were utilized as novel reaction media in order to detect new chemistry for catalytic metalloporphyrins. This research involves two general areas: (1) EPR and ENDOR spectroscopic studies of metalloporphyrins in different hosts: ionic liquids, porous materials, and conventional molecular solvents; (2) catalytic reactivity and mechanism studies of metalloporphyrins in different hosts. Spectroscopic results indicate a distortion in geometry of copper porphyrins in ionic liquids, which shifts the spin density distribution within the molecule. Similar studies of iron porphyrins indicate that component ions from ionic liquids can bind to the metal centres of these catalytic molecules. Some ionic liquids were found to improve the reactivity of porphyrin catalysts in oxidation reactions due to their ability to solubilize all reaction components and to prevent aggregation of porphyrins. A porphyrin radical cation intermediate was detected in the oxidation reaction of 4-methoxythioanisole by iron meso-tetra(4-sulfonatophenyl)porphyrin.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Charles J. Walsby
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) M.Sc.

Thermal expansion of d10 dicyanometallate-based coordination polymers

Date created: 
2010-08-11
Abstract: 

The thermal expansion of several new d10, dicyanometallate-based coordination polymers was examined by powder and single-crystal X-ray diffraction. In isostructural KNi[Au(CN)2]3, KCd[M(CN)2]3, and In[M(CN)2]3 (M=Ag(I), Au(I)), extremely large positive and negative thermal expansion (PTE/NTE) coefficients were discovered. Weaker metallophilic interactions promote larger PTE and NTE. Similarly, the M[Au(CN)2]2 series (M=Mn, Fe, Co, Zn, Hg) showed sizeable PTE and NTE effects. HgCN(NO3) exhibited similar behaviour, but significantly smaller PTE/NTE coefficients than the Ag and Au systems. Hg(CN)2 demonstrated entirely (anisotropic) PTE, partly attributed to a new structural mechanism resembling rigid unit modes in metal oxides. Hg(CN)2 is one of the first cyanide-containing frameworks not showing NTE. This work has demonstrated that Ag(I) and Au(I) in coordination polymers drive large expansivity, and when coupled with flexible cyanide frameworks, can also yield large NTE.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Daniel Leznoff
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) M.Sc.

Rational design and synthesis of chemical probes for O-GlcNAcase and OGT

Date created: 
2010-07-21
Abstract: 

Attachment of GlcNAc via a β-O-linkage to serine or threonine residues of various nucleocytoplasmic proteins is a posttranslational modification in multicellular eukaryotes. A glycosyl transferase, UDP-N-acetyl-D-glucosamine: polypeptide-N-acetylglucosaminyl transferase (OGT) installs O-GlcNAc while a β-N-acetylglucosaminidase, O-GlcNAcase, removes this modification from proteins. A dynamic relationship between O-GlcNAc and phosphorylation has been proposed and is implicated in cellular signalling and disorders such as Alzheimer disease. In this thesis, the facile syntheses of two series of inhibitors of O-GlcNAcase are described. The potency of these compounds toward O-GlcNAcase is also detailed. Several of these compounds are highly potent and selective inhibitors of O-GlcNAcase. Additionally, 2-acetamido-2-deoxy-5-thio-D-glucopyranosose was synthesized as an analogue of GlcNAc. This compound was used for cell studies and to synthesize para-methoxyphenyl 2-acetamido-2-deoxy-5-thio-D-glucopyranoside as a potential substrate for O-GlcNAcase. Together, these compounds should serve as useful chemical probes to study the function of O-GlcNAc in cells and in vivo.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Dr. David J. Vocadlo
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) M.Sc.

The effect of molecular shape and symmetry on discotic mesogens

Author: 
Date created: 
2010-04-01
Abstract: 

In the field of material science, structure/property relationships can allow a more efficient design of materials exhibiting the desired properties. Self-assembled super-structures possess several advantages that can be exploited. The formation of complex architecture is dictated by the properties of the small sub-units. The formation of liquid crystalline phases requires molecular anisotropy, which is often achieved by using flat rigid rod or disc-shaped cores decorated with aliphatic chains. Other structural morphologies have also proven to allow the formation of these ordered fluid phases: plate, bent-core, bowl etc. Our research group is interested in disc-shaped molecules that have the ability to form liquid crystalline phases constituted of columns distributed in a two-dimensional lattice. The discs within a column are stacked on top of each other that could be used to conduct either energy or charges. The research projects presented in this thesis were concerned in establishing the effect of molecular symmetry and shape on the mesogenic properties of compounds based on a dibenz[a,c]phenazine core decorated with alkoxy chains (four and six). Series of structural isomers have been synthesized and their mesogenic behaviour analyzed. Reducing the molecular symmetry of mesogens lowers the melting temperature and, to a smaller extent, the clearing temperature leading to broader liquid temperature ranges of mesogenic behaviour. To study the effect of molecular shape, compounds with chains of different length and different motifs were prepared. This type of substitution leads to different mesogenic behaviour. Comparisons between structural isomers showed that this new substitution pattern lowered the clearing temperatures for these mesophases. The molecular shape of these compounds does have an effect on the mesophase and the results could allow more efficient engineering of systems used in various devices.

Document type: 
Thesis
File(s): 
Supervisor(s): 
Vance E. Williams
Department: 
Science: Department of Chemistry
Thesis type: 
(Thesis) Ph.D.