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Improvement of Gastroparesis Management By Addressing Challenges in Drug Metabolism: Studies with Metabolite Identification, Reaction Phenotyping and In Vitro Drug-Drug InteractionsYoussef, Amir Samaan Bishara January 2013 (has links)
Gastroparesis is a disorder characterized by delayed gastric emptying due to chronic abnormal gastric motility. Prokinetic agents such as domperidone and metoclopramide are the cornerstone in treatment of gastroparesis. Although these medications have been used for decades, essential information about their metabolism is not available. Lack of knowledge about the metabolites formed in the body upon administration of the aforementioned medications as well as the enzymes involved in their metabolism limits key information needed to make sound medical decisions. Accurate and comprehensive identification of the metabolites along with reaction phenotyping of prokinetic agents will ensure safe and effective use of these drugs and hence enhance the clinical outcome. The thesis starts with an introductory chapter which comprises a comprehensive literature review on gastroparesis and the available pharmacological treatment options. The chapter also emphasizes the importance of metabolic profiling of prokinetic agents (domperidone and metoclopramide) and its impact on enhancing the safety and efficacy of these medications. Chapter 2 of this project was aimed to determine phase oxidative and conjugative metabolites of domperidone in the plasma and urine of gastroparesis patients using tandem mass spectrometry. First, the metabolites were identified in in-vitro human subcellular fractions. The knowledge gained in this experiment helped identifying the metabolites in the biological fluids of patients. In total, 12 metabolites including 7 new metabolites were identified, 5 of which were not reported previously. Chapter 3 aimed to identify the cytochrome P450 (CYP) enzymes responsible for the metabolism of metoclopramide. The parent depletion approach was used and a novel LC-MS/MS method was developed and validated to enable metoclopramide quantification. CYP2D6 was showed to the predominant isoform in metoclopramide metabolism; other isoforms also contribute to a minor extent. Chapter 4 discusses the possibility of potential drug-drug interaction (DDI) in the current management practice of gastroparesis. We identified and investigated some frequently used drug combinations that are known to share common metabolic pathways. Domperidone in combination with pioglitazone and ondansetron was evaluated. Results showed that pioglitazone inhibited domperidone metabolism in-vitro. Our experiments did not predict a DDI for the domperidone - ondansetron combination. In summary, the ultimate goal of this thesis was to improve the management of gastroparesis by increasing information about the metabolic disposition of prokinetic agents and to investigate the magnitude of putative drug combinations. The knowledge provided by this work will help in making more effective and less hazardous clinical decisions which will ultimately lead to more successful gastroparesis management. / Pharmaceutical Sciences
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Quantitative Bioanalysis : Liquid separations coupled to targeted mass spectrometric measurements of bioactive compoundsArvidsson, Björn January 2008 (has links)
Performing quantitative analysis of targeted bioactive compounds in biological samples, such as blood plasma, cerebrospinal fluid or extracts from pig liver, put high demands on the ruggedness of the method acquiring the results. In addition to the complexity of the sample matrix, the bioactive compounds targeted for analysis usually have low levels of natural abundance, further increasing the demand on the analytical method sensitivity. Furthermore, quantitation of analytes at trace levels in the presence of large amounts of interfering species in biofluids must aim for repeatable precision, high accuracy ensuring the closeness to the true values, a linear response spanning over several orders of magnitude and low limits of quantitation to be valid for monitoring disease states in clinical analysis. An analytical method most commonly follow a certain order of events, called the analytical chain, which includes; experimental planning, sampling, sample pre-treatment, separation of species, detection, evaluation, interpretation and validation, all equally important for the outcome of the results. In this thesis, the scope has been to create novel methods, or to refine already existing methods, in order to achieve even better performances of the different events in the analytical chain. One of the aspects has been to sample and enrich analytes in vivo by the use of solid supported microdialysis, giving the advantage of almost real-time monitoring of analyte levels within a living host with targeted selectivity due to the analyte affinity for solid particles. Another aspect to selectively clean and enrich analytes in a complex matrix has been developed and automated on-line by the use of a column-switching technique before the analytical separation. By using several steps of extraction and separation coupled on-line to selected detection by the use of a triple quadrupole mass spectrometer facilitates great selectivity of species. The mass spectrometer also gives the ability to distinguish between isotopically labelled analogues coeluting with the analytes yielding the necessary accuracy for quantitative evaluation. Both development of analytical methods and clinical applications has been performed, as well as improvements of existing techniques, all to improve the quantitation of trace levels of targeted analytes in biofluids.
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The Gas-Phase Ligand Exchange of Calcium β-diketonate ComplexesGatte, Brandi J. 02 May 2022 (has links)
No description available.
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Charakterizace glykoforem haptoglobinu v lidském séru / Characterization of hapt oglobin glycoforms in human serumDarebná, Petra January 2015 (has links)
Characterization of haptoglobin glycoforms in human serum Petra Darebná (Katedra biochemie, Přírodovědecká fakulta, Univerzita Karlova v Praze, Česká republika) Changes in glycosylation of proteins are associated with several types of cancer, including hepatocelular carcinoma and colorectal carcinoma. This project deals with data independent analysis using ion cyclotron resonance with Fourier transform and tandem mass spectrometry with liquid chromatogramy and multiple reaction monitoring to quantify these changes in hepatocelular cancinoma and colorectal carcinoma with liver metastases. In the first part of the project the haptoglobin was enriched from pooled serum samples of pacients with hepatocellular carcinoma, colorectal cancer and colorectal carcinoma with metastases using hemoglobin immobilized on CNBr-activated Sepharose 4B and then purified by high pressure liquid chromatography. Individual haptoglobin glycopeptides were analyzed using ion cyclotron resonance with Fourier transform. In the second part of the project we analyzed changes in glycosylation depending on diferent tumor diseases in partially depleted serum of individual patients using ethanol precipitation. Individual fucosylated glycoforms of N-glycopeptides of serum proteins were compared with their nonfucosylated forms. In...
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Analysis of Glycerophospholipids and Sphingolipids in Murine Brain Using Liquid Chromatography – Electrospray Ionization - Tandem Mass Spectrometry and Matrix-Assisted Laser Desorption Ionization – Imaging Mass SpectrometryNguyen, Thao January 2017 (has links)
Mass spectrometry is an indispensable tool in lipidomics research. Current advances and progress in the technology of mass spectrometry have allowed for the identification, quantification and characterization of lipid molecular species to further our understanding of their biological roles. In this thesis, I assessed the influence post-mortem times have on quantitative lipidomics. Using liquid chromatography - electrospray ionization tandem mass spectrometry (LC-ESIMS/MS) on a triple-quadrupole mass spectrometer and multiple-reaction-monitoring (MRM) mode, the glycerophosphocholine (GPC) metabolites and second messengers in the hippocampus of N3 & N4 C57BL/6 x 129/SV were profiled at various post-mortem interval (PMI). I found that disruption to the GPC metabolite and second messengers lipidome occured as early as 1 hour postmortem and fluctuate up till at least 12 hours post-mortem. Therefore, PMI is a variable in lipidomic studies that must be controlled for, and brain samples which are collected with PMI variations must be matched to avoid misinterpretation. Subsequently, I developed a working protocol to visualize the location and distribution of different classes of glycerophospholipids, ceramides, and sphingomyelin in whole mouse brain sections. This visualization technique is novel because it does not require tissue staining or immunohistochemistry; instead, it was performed using an atmospheric-pressure matrix-assisted laser desorption/ionization (AP-MALDI) source coupled to an orbitrap mass spectrometer. As part of this lipid visualization technique, I also developed a protocol for sublimation as a simple, effective and reproducible matrix application
method for brain tissue. The lipid-compatible matrix, 2,5-dihydroxybenzoic acid (DHB), was assessed and optimized for imaging lipid targets. The high mass-resolution and accuracy characteristics of the orbitrap mass spectrometer and its capability to perform tandem mass spectrometry via high-collision dissociation allowed for the identification of approximately 200 different lipid species directly from brain tissue using the visualization technique I developed.
Altogether, the work in this thesis has showed that post-mortem changes in the lipidome are quantifiable and has provided a novel avenue to further assess these changes by means of imaging mass spectrometry.
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The Investigation of Reactions of Atomic Metal Anions with Small Hydrocarbons and Alcohols in the Gas PhaseHalvachizadeh, Jaleh 21 February 2014 (has links)
Hydrocarbons are an abundant resource of carbon and hydrogen. For example, fossil can be used to produce useful organic compounds. However hydrocarbons seem to be inert. Thus, the activation of the C-H bond is a popular research area. Metals play the main role in most catalysts that convert hydrocarbons to starting materials in industry. The study of metals is important because the properties of the metal core greatly influences the reactivity of a catalyst.1
The study of the chemistry of metals in the gas phase provides valuable information about the properties of metals. This information can be expanded to the chemistry of metals in the condensed phase. Furthermore, it is often both more accurate and more manageable to study the profile of a reaction in the gas phase than in the condensed phase.2,3
There are many studies about metal cations in the gas phase due to ease of their production. However metals have low electronegativity, limiting the study of gas phase metal anions. Recently, a simple and efficient method to generate atomic metal anions was developed at the University of Ottawa in Dr. Mayer's research laboratory.4-6 Atomic metal anions of Fe-, Co-, Cu-, Ag-, Cs- and K- were generated in an electrospray ionization (ESI) source of a mass spectrometer (MS).
In this thesis study generated metal anions were reacted with small hydrocarbons of pentane, 1-pentene, 2-pentene and 1-pentyne to investigate the role of different metal anions in the activation of the C-H bond. Also metal anions were reacted with small alcohols of 1-butanol, 2-butanol and 2-methyl-2-propanol to compare the results.
Metal anions showed a variety of reactions with these hydrocarbons and alcohols. Fe- was the only metal anion to show the electron transfer reaction, indicating that alcohols are more electronegative than Fe- and less electronegative than other metal anions. Fe-, Co- and Ag- showed the complex formation reaction. All metal anions showed the deprotonation reaction. A deprotonation reaction follows the harpoon mechanism, the long range proton abstraction7, and depends on the gas phase acidity of fragments. The most informative reaction observed was the dehydrogenation reaction because a metal-containing fragment is observed as a product in the spectrum of this reaction. The observation of a metal-containing fragment in the spectrum is significant because it emphasizes the important role that metal anions play in this reaction. This suggests that a dehydrogenation reaction involves metal insertion into a C-H bond. Among the transition metal anions, it was observed that Fe- and Cu- are more reactive than Co- and Ag- with regards to the dehydrogenation reaction, probably because Fe- and Cu- have a greater hydrogen affinity than Co- and Ag- that facilitates the hydrogen abstraction reaction. Another reason could be that Fe- and Cu- have a greater gas phase acidity that leads to a more stable intermediate in the course of the reaction. The results of this thesis study revealed that Cs- and K- could not abstract H from these substrates, probably due to the absence of occupied d orbitals that would facilitate insertion into a C-H bond.
Some metal anions not only can insert into a C-H bond of alcohols but also can insert into a C-O bond of alcohols to form metal hydroxide anions. Alcohols are more reactive than hydrocarbons with regards to reactions with metal anions because they contain a functional group.
This thesis study shows that some atomic metal anions are able to activate the C-H bond and abstract two hydrogens to form a double bond in hydrocarbons. It is probable that the electronic configuration, gas phase acidity and hydrogen affinity of the metal anions governs their reactivity.
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The Investigation of Reactions of Atomic Metal Anions with Small Hydrocarbons and Alcohols in the Gas PhaseHalvachizadeh, Jaleh January 2014 (has links)
Hydrocarbons are an abundant resource of carbon and hydrogen. For example, fossil can be used to produce useful organic compounds. However hydrocarbons seem to be inert. Thus, the activation of the C-H bond is a popular research area. Metals play the main role in most catalysts that convert hydrocarbons to starting materials in industry. The study of metals is important because the properties of the metal core greatly influences the reactivity of a catalyst.1
The study of the chemistry of metals in the gas phase provides valuable information about the properties of metals. This information can be expanded to the chemistry of metals in the condensed phase. Furthermore, it is often both more accurate and more manageable to study the profile of a reaction in the gas phase than in the condensed phase.2,3
There are many studies about metal cations in the gas phase due to ease of their production. However metals have low electronegativity, limiting the study of gas phase metal anions. Recently, a simple and efficient method to generate atomic metal anions was developed at the University of Ottawa in Dr. Mayer's research laboratory.4-6 Atomic metal anions of Fe-, Co-, Cu-, Ag-, Cs- and K- were generated in an electrospray ionization (ESI) source of a mass spectrometer (MS).
In this thesis study generated metal anions were reacted with small hydrocarbons of pentane, 1-pentene, 2-pentene and 1-pentyne to investigate the role of different metal anions in the activation of the C-H bond. Also metal anions were reacted with small alcohols of 1-butanol, 2-butanol and 2-methyl-2-propanol to compare the results.
Metal anions showed a variety of reactions with these hydrocarbons and alcohols. Fe- was the only metal anion to show the electron transfer reaction, indicating that alcohols are more electronegative than Fe- and less electronegative than other metal anions. Fe-, Co- and Ag- showed the complex formation reaction. All metal anions showed the deprotonation reaction. A deprotonation reaction follows the harpoon mechanism, the long range proton abstraction7, and depends on the gas phase acidity of fragments. The most informative reaction observed was the dehydrogenation reaction because a metal-containing fragment is observed as a product in the spectrum of this reaction. The observation of a metal-containing fragment in the spectrum is significant because it emphasizes the important role that metal anions play in this reaction. This suggests that a dehydrogenation reaction involves metal insertion into a C-H bond. Among the transition metal anions, it was observed that Fe- and Cu- are more reactive than Co- and Ag- with regards to the dehydrogenation reaction, probably because Fe- and Cu- have a greater hydrogen affinity than Co- and Ag- that facilitates the hydrogen abstraction reaction. Another reason could be that Fe- and Cu- have a greater gas phase acidity that leads to a more stable intermediate in the course of the reaction. The results of this thesis study revealed that Cs- and K- could not abstract H from these substrates, probably due to the absence of occupied d orbitals that would facilitate insertion into a C-H bond.
Some metal anions not only can insert into a C-H bond of alcohols but also can insert into a C-O bond of alcohols to form metal hydroxide anions. Alcohols are more reactive than hydrocarbons with regards to reactions with metal anions because they contain a functional group.
This thesis study shows that some atomic metal anions are able to activate the C-H bond and abstract two hydrogens to form a double bond in hydrocarbons. It is probable that the electronic configuration, gas phase acidity and hydrogen affinity of the metal anions governs their reactivity.
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Electron Transfer and Other Reactions Using Atomic Metal AnionsButson, Jeffery M. 04 February 2014 (has links)
The atomic metal anions Rb-, Cs-, Cu-, Ag- and Fe- have been generated in the gas phase and reacted with various neutral reactants in a triple quadrupole mass spectrometer. The metal anions were formed via electrospray ionization of the metal-oxalate solutions and form in gas phase between the capillary and the first quadrupole. Neutral gas phase reactants investigated include NO, NO2, SO2, C6F5OH, C6F5NH2, C6F6, E-octafluoro-butene and 1,2,3/1,2,4/1,3,5 trifluoro-benzene. When possible, CBS-4M methods were used to suggest the lowest energy products based on relative energy. Observed reactions of atomic metal anions with the aforementioned neutral species include electron transfer and dissociative electron transfer to the neutral gas phase reactants. In addition, hydrogen abstraction and fluorine abstraction forming a neutral metal hydride or fluoride as well as the formation of multiply substituted metal-oxide/fluoride anions was also observed. Metal-complex anions observed from the gas phase reactions include CuF-,CuF2-,CuO-,CuO2-, FeO-, FeO2-, FeO3-, FeF-, FeF2-, FeF3-, CsF- and CsF2-.
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Electron Transfer and Other Reactions Using Atomic Metal AnionsButson, Jeffery M. January 2014 (has links)
The atomic metal anions Rb-, Cs-, Cu-, Ag- and Fe- have been generated in the gas phase and reacted with various neutral reactants in a triple quadrupole mass spectrometer. The metal anions were formed via electrospray ionization of the metal-oxalate solutions and form in gas phase between the capillary and the first quadrupole. Neutral gas phase reactants investigated include NO, NO2, SO2, C6F5OH, C6F5NH2, C6F6, E-octafluoro-butene and 1,2,3/1,2,4/1,3,5 trifluoro-benzene. When possible, CBS-4M methods were used to suggest the lowest energy products based on relative energy. Observed reactions of atomic metal anions with the aforementioned neutral species include electron transfer and dissociative electron transfer to the neutral gas phase reactants. In addition, hydrogen abstraction and fluorine abstraction forming a neutral metal hydride or fluoride as well as the formation of multiply substituted metal-oxide/fluoride anions was also observed. Metal-complex anions observed from the gas phase reactions include CuF-,CuF2-,CuO-,CuO2-, FeO-, FeO2-, FeO3-, FeF-, FeF2-, FeF3-, CsF- and CsF2-.
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