Thesis (Ph.D.)--Duquesne University, 2004. / Title from document title page. Abstract included in electronic submission form. Includes bibliographical references and abstracts.

Neurotransmitters are endogenous compounds located in the brain and can occur in other parts of the body at low concentrations. They are a challenging group of compounds when it comes to analysis. Compounds with amine functionality such as dopamine and serotonin are sensitive to high pH and light and can spontaneously oxidize and degrade. Since neurotransmitters exist at low levels, it can be difficult to achieve sufficient spectrometry data and clean chromatography. In order to improve data acquisition, derivatization and the use of multidimensional liquid chromatography (2D-LC) was evaluated. Dopamine, L-3,4-dihydroxyphenylalanine (L-DOPA), serotonin, tyrosine, tryptophan, and noradrenaline were the compounds of interest with tyramine included as an internal standard. Data was collected before and after derivatization to compare the resulting chromatography. Chromatography was completed using a 6x6 grid of methods with variables of organic solvent, elution pH, loading pH, and trapping column chemistry utilizing different elution column chemistries. Derivatization was examined with Dabsyl-Cl and Dansyl-Cl at pH 8.5, 9.5, 10.5, and 14. A final method was chosen with Dabsyl at pH 8.5 for the Multiple Reaction Monitoring (MRM) scans. Before derivatization, the chromatography had poor peak shapes such as tailing, fronting, or shouldering or too much distortion to be able to distinguish a peak for all of the examined methods. Resulting chromatography after derivatization showed overall improvement in peak shape and intensity for a majority of the methods. Derivatization aided in increasing the mass and stability of the compounds which allowed for more sensitive detection. Multidimensional liquid chromatography improved the separation of structurally similar compounds and increased sensitivity. By combining the two, better analysis of neurotransmitters was possible.

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There continue to be great strides in the field of proteomics but as samples become more complex, the ability to increase sequence coverage and confidence in the identification becomes more important. Several methods of derivatization have been developed that can be used in combination with tandem mass spectrometry to identify and characterize proteins. Three types of activation, including infrared multiphoton dissociation, ultraviolet photodissociation, and electron transfer dissociation, are enhanced in this dissertation and compared to the conventional method of collisional induced dissociation (CID) to demonstrate the improved characterization of proteins. A free amine reactive phosphate group was synthesized and used to modify the N-terminus of digested peptides. This phosphate group absorbs at the IR wavelength of 10.6 µm as well as the Vacuum-ultraviolet (VUV) due to an aromatic group allowing modified peptides to be dissociated by infrared multi-photon dissociation (IRMPD) or ultraviolet photodissociation (UVPD) whereas peptides without this chromophore are less responsive to IR or UV irradiation. The PD spectra for these modified peptides yield simplified MS/MS spectra due to the neutralization of all N-terminal product ions from the incorporation the negatively charged phosphate moiety. This is especially advantageous for UVPD due to the great number of product ions produced due to the higher energy deposition of the UV photons. The MS/MS spectra also produce higher sequence coverage in comparison to CID of the modified or unmodified peptides due to more informative fragmentation pathways generated upon PD from secondary dissociation and an increased ion trapping mass range. IRMPD is also implemented for the first time on an orbitrap mass spectrometer to achieve high resolution analysis of IR chromophore-derivatized samples as well as top-down analysis of unmodified proteins. High resolution/high mass accuracy analysis is extremely beneficial for characterization of complex samples due to the likelihood of false positives at lower resolutions/accuracies. For electron transfer dissociation, precursor ions in higher charge states undergo more exothermic electron transfer and thus minimize non-dissociative charge reduction. In this dissertation, cysteine side chains are alkylated with a fixed charge to deliberately increase the charge states of peptides and improve electron transfer dissociation. ETD can also be used to study protein structure by derivatizing the intact structure with a hydrazone reagent. A hydrazone bond will be preferentially cleaved during ETD facilitating the recognition of any modified residues through a distinguishing ETD fragmentation spectrum. / text

Pharmaceuticals are a group of emerging organic compounds of environmental concern used extensively in human and veterinary medicine. They are continually released into the environment as a result of manufacturing operations and excretion from humans and animals. These compounds enter directly into the municipal sewage systems and into wastewater treatment plants. A large number of important and potentially harmful organic contaminants, such as these pharmaceuticals, are not regulated in drinking and other waters. As a result, conventional technologies at most waste water treatment plants (WWTPs) discharge water that meet regulatory standards, yet are not specifically designed to remove these organic contaminants. Therefore, pharmaceutical compounds and their metabolites remain in discharged effluent and enter into the natural aquatic environment. Concentrations of pharmaceutical residues measured in water are typically reported in the ranges of ug/L to ng/L, which are at least three to four orders of magnitude lower than that required to produce a pharmacological effect. The probability of risks to humans arising from such an acute exposure is unlikely, but the possible effects resulting from life-long exposures and synergistic effects from exposure to many chemicals have yet to be determined. It has been widely reported that pharmaceuticals and their metabolites that enter into the aquatic environment can have a potential harmful effect on the aquatic ecosystem and can reach drinking water sources. This research focuses on non-steroid anti-inflammatory drugs (NSAIDs), a group of pharmaceuticals which are widely used as analgesic, antipyretic and anti-inflammatory agents. NSAIDs are frequently used because they are easily accessible as over the counter medication and are a group of drugs that do not produce addiction, respiratory depression, or drowsiness. There is an incentive for removing NSAIDs and other pharmaceuticals from the aquatic environment. Thus, quantitative evaluation of the fate of pharmaceuticals, proper risk assessment and improvement of the efficiency of WWTPs need sensitive and reliable analytical methods. The purpose of this project was to provide a method for detecting three common NSAIDs, IBF, KTF, and NAP, in purified water with LLE-GC-FID. And, an investigation of UV photolysis, UV/H2O2, and UV/TiO2 AOPs was performed to determine their effectiveness in treating IBF, KTF, and NAP in purified water. All treatment methods were successful in degrading target compounds with a total degradation of 86% or greater after 45 minutes. A liquid-liquid extraction technique using methylene chloride and BSTFA + 1%TMCS derivatizing agent was determined for detecting low concentrations of IBF, KTF, and NAP with calibration curves showing good linearity with all R2 values greater than 0.9880.

La recherche de vie extraterrestre et en particulier de vie sur Mars est devenue un des enjeux majeurs des prochaines explorations spatiales. Dans ce cadre, la recherche de molécules organiques et l’étude de leur énantiomérie pourrait permettre d'obtenir les premiers indices en faveur d'une présence passée ou présente de vie sur cette planète. En effet la vie telle que nous la connaissons est uniquement basée sur des acides aminés L et sucres D. Ces molécules organiques et leur éventuel excès énantiomérique pourraient avoir été conservés dans les conditions martiennes. La chromatographie en phase gazeuse couplée à la spectrométrie de masse (CPG-SM) est actuellement la technique analytique spatialisable la plus pertinente pour la détection de composés organiques volatils. Afin de rendre volatils les composés réfractaires tels que les acides aminés, nous proposons une fonctionnalisation chimique au diméthylformamide diméthyl-acétal (DMF-DMA). Cette fonctionnalisation couplée à une séparation chromatographique sur une colonne chirale est une technique parfaitement adaptée à la séparation énantiomérique et à la spatialisation. Nous avons donc mis au point une méthode de fonctionnalisation des molécules organiques chirales par l’agent de méthylation DMF-DMA afin de séparer les énantiomères et de les étudier in situ en CPG-SM lors des futures missions martiennes. Cette technique d'analyse des molécules réfractaires chirales a ensuite été adaptée à une utilisation spatiale. Ainsi, la méthode d’analyse est constituée d’un four réactionnel dans lequel l’échantillon solide estplacé, les molécules d’intérêt exobiologique sont ensuite successivement extraites et fonctionnalisées au DMFDMA pour être analysées en CPG-SM. Les résultats ont permis de montrer qu’ à partir d’une faible quantité de sols d'analogues martiens (50 mg de sol du désert d’Atacama, Chili) et après optimisation de chacune des étapes, plusieurs types de molécules organiques présentes à l’état de traces ont pu être détectées, telles que des acides aminés, des acides carboxyliques et des bases nucléiques. Parmi ces molécules chirales, plusieurs ont pu être séparées selon leur forme énantiomérique. Le protocole est élaboré dans les limites technologiques et opératoires d’une analyse spatiale ; chaque étape est pensée pour un usage robotisé. Nous avons ainsi développé une expérience globale, sensible et robuste, totalement compatible avec les exigences d’une analyse in situ. C’est la raison pour laquelle cette expérience sera intégrée sur la prochaine mission martienne Exomars 2016/2018 afin de rechercher des traces de matière organique à la surface et en subsurface de Mars. La découverte d’un excès énantiomérique, voire d’une homochiralité, apporterait des indices forts quant à la présence d’une vie passée ou présente sur la planète rouge. / Looking for extraterrestrial life, particularly on Mars, has become a challenging aim for future space missions. In this framework, looking for organic molecules and studying their enantiomeric properties could lead to the first clues of an extinct or extant life on this planet. Indeed, life as we know it consists in one of the two specular forms of organic molecules only, and it has been shown that conditions at Mars could preserve, at least partially, this enantiomeric excess. Gas Chromatograph coupled to a Mass Spectrometer (GC-MS) is nowadays the most pertinent space compatible analytical tool for the detection of volatile organics. In order to transform refractory molecules suchas amino acids into volatile ones, we are putting forward a chemical derivatization with dimethylformamidedimethyl-acetal (DMF-DMA). This reaction, when coupled with an analytical step using a chiral chromatographic column, is perfectly adapted to enantiomeric separation within the space constraints. This thesis develops a method of derivatization of the chiral organic molecules with methylation reagent DMFDMA in order to separate and analyze in situ the enantiomers by GC-MS during the future Martian missions. This analytical tool consists in a reaction chamber into which the solid sample is dropped. From this sample, the molecules of interest are successively extracted and derivatized with DMF-DMA to be detected byGC-MS. From a low quantity of Martian analogue soil (50 mg of Atacama soil) and after a complete optimization of each step, numerous molecules of different kinds present at traces levels have been successfully detected, such as amino, carboxylic and nucleic acids. Among these molecules, some of the chiralones have been resolved regarding their enantiomeric forms. The whole procedure is elaborated within the technological and operational limits imposed by a space experiment, each step having been designed to becarried out robotically. Thus, we have developed a complete experiment, sensitive, robust and entirely meeting the requirements of an in situ analysis. That is the reason why this experiment has been selected to be integrated onboard the next Martian mission Exomars 2016/2018 in order to search traces of organic matter at Mars surface and subsurface. Determining an enantiomeric excess, or even homochirality, would be a remarkable breakthrough regarding the presence of an extinct or extant life on the red planet.

Orientador: Marco Aurelio Zezzi Arruda / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica / Made available in DSpace on 2018-08-08T22:29:56Z (GMT). No. of bitstreams: 1 Matos_GeraldoDomingues_D.pdf: 2963776 bytes, checksum: 63761ac61fa005bd6133cb5e9831941d (MD5) Previous issue date: 2007 / Resumo: O objetivo desta Tese foi a ampliação das potencialidades do método TS-FF-AAS, no que diz respeito à melhora da sensibilidade para Cd e Co. Com relação ao Cd, foi desenvolvido um procedimento de pré-concentração em linha em uma minicoluna contendo bagaço de uva. Inicialmente, foi realizada uma caracterização físico-química do material adsorvente empregando espectrometria na região do infravermelho, ressonância magnética nuclear de C, análise termogravimétrica, difratometria de raios-X e microscopia eletrônica de varredura. As variáveis otimizadas incluem o pH de adsorção, tipo, concentração e volume do eluente e vazão de pré-concentração. O procedimento apresentou faixa linear de 0,11-5 mg/L, limite de detecção (LD) de 0,03 mg/L e precisão de 5%, além da boa estabilidade do material. O ganho no LD foi de 16 vezes em relação a TS-FF-AAS sem a etapa de pré-concentração. A exatidão foi confirmada com o uso de materiais de referência certificados, ao nível de 95% de confiança. Para o Co, foi desenvolvido um procedimento com derivatização em linha com posterior quantificação por TS-FF-AAS. Assim, foram estudados alguns complexantes, sendo o dietilditiocarbamato de sódio o mais adequado. Os parâmetros otimizados foram: pH de complexação, volume e concentração do complexante, volume de amostra, composição dos gases da chama e área total de furos do tubo de níquel. Estas duas últimas apresentaram-se como os mais importantes. O uso do complexante proporcionou uma maior eficiência de vaporização da amostra, comprovado por meio de filmagens de formação do spray térmico. O procedimento apresentou faixa linear de 23-400 mg/L e LD de 7 mg/L, o que representa um ganho de 17 vezes em relação a FAAS. A exatidão foi verificada empregando materiais de referência certificados com concordância com valores certificados ao nível de 95% de confiança. / Abstract: The main objective of this Thesis was the improvement of the TS-FF-AAS technique concerning sensitivity for Cd and Co. For Cd, an on-line preconcentration procedure in a mini-column containing grape bagasse was developed. Initially, physical-chemical characterization of this material was made using infrared spectroscopy, nuclear magnetic resonance C, thermogravimetric analysis, X-ray difractometry and scanning electron microscope. The optimized variables include the pH of sorption, type, concentration and volume of the eluent and preconcentration flow rate. The procedure provided linear range from 0.11-5.0 mg/L, limit of detection (LD) of 0.03 mg/L and precision of 5% as well as good stability of the material. The gain in the power of detection was of 16-fold when compared to TS-FF-AAS without preconcentration step. The accuracy was confirmed by using certified reference materials. For Co, a procedure using on-line derivatization and quantification by TS-FF-AAS was developed. In this way, some complexing agents were evaluated and the most appropriate was sodium diethyldithiocarbamate. The optimized parameters were complexation pH, complexing volume and concentration, sample volume, flame composition and nickel tube with total hole area. These two last were the most important. The use of complexing agents provided a larger vaporization efficiency of the sample, checked by means of filming of formation of the thermospray. This procedure provided a linear range from 23-400 mg/L and LD of 7 mg/L, what represents a gain of 17-fold in relation to FAAS. The accuracy was evaluated using certified reference materials with agreement with certificate values at the 95% confidence level. / Doutorado / Quimica Analitica / Doutor em Quimica

β-Aminocarbonyl motifs are a privileged substructures in medicinal chemistry and peptidomimetics. As part of our efforts toward metal free aminations, we developed a method for intermolecular amino-carbonylation of alkenes using hydrazones. This method provides access to cyclic azomethine imines containing a β-aminocarbonyl motif. Conceptually, these dipoles can be derivatized into many bioactive compounds, such as 1,3-diamines, β-amino amides and β-amino acids. The first part of this thesis will present the results on the derivatization of our aminocarbonylation products into various nitrogen-containing molecules, such as β-amino amides, β-amino acids and pyrazolones. More specifically, a short, chromatography-free derivatization of azomethine imines into N-Boc-β-amino amides will be presented. Following these results, the next chapter will focus on attempts at develop novel aminocarbonylation reactivity between 1,2-diacylhydrazines and alkenes followed by results from our reductive N-N bond cleavage experiments on our cyclic hydrazides.

Alkylphenol ethoxylates (APEOs), alcohol ethoxylates (AEOs), and alcohol propoxylates (APOs) are non-ionic surfactants used in daily care products and detergents. They are formed as an oligomeric series with a varying distribution, which determines their commercial application. The goal of the research performed was the development of sample characterization methods for non-ionic surfactants utilizing supercritical fluid chromatography (SFC) under mild instrument operating conditions. The aryl group present in APEOs allowed ultraviolet (UV) detection, with an equal molar response for oligomers, allowing average molar oligomer values to be calculated. APEOs were separated by ethoxylate unit via SFC-UV as well as normal phase HPLC-UV employing packed columns. Stationary phase and column length were varied in the SFC setup to produce the most favorable separation conditions. Fractions from SFC runs of APEOs were collected and analyzed by flow injection analysis electrospray ionization mass spectrometry (FIA-ESI-MS) to identify fraction composition. SFC provided shorter retention times with similar resolution as HPLC for separation of APEOs and consumed a smaller amount of organic solvent. AEOs and APOs lack functionality capable of absorbing UV light outside the UV cut-off of normal organic solvents. SFC was able to separate AEOs and APOs derivatized as trimethylsilyl ethers (TMS) with pure CO2 with detection at 195 nm. The instrumental conditions, however, needed for separation necessitated high temperature and high CO2 pressure. Derivatization of alcohol polyether samples with an UV absorbing agent was achieved with phenylated disilazane-chlorosilane mixtures forming phenylsilylethers detected at 215 nm. Use of an organic solvent-modified CO2 mobile phase afforded lower pressure and temperature conditions for oligomer separation. The use of polar embedded alkyl phases combined with use of organic modified CO2 produced good resolution between oligomers. Better peak shape and shorter retention times were realized with methanol-modified CO2 than acetonitrile-modified CO2. Peak assignments were made via SFC coupled with ESI-MS detection in the positive ion mode. SFC-UV and SFC-ESI-MS data were jointly used for calculation of average molar oligomer values. Proton nuclear magnetic resonance (1H-NMR) analysis of non-derivatized samples was performed to determine average molar oligomer values and was used for comparison with values calculated from SFC-UV data. / Ph. D.

Alginates are (1and4) linked linear copolysaccharides composed of B-D-mannuronic acid (M) and its C-5 epimer, a-L-guluronic acid (G). Several strategies to synthesize organically modified alginate derivatives have been reported, but almost all chemistries are performed in either aqueous or aqueous-organic media. The ability to react alginates homogeneously in organic solvents would open up access to a wide range of new chemistries and derivatives. However, past attempts have been restricted by the absence of methods for alginate dissolution in organic media. We therefore report a strategy to solubilize tetrabutylammonium (TBA) salts of alginic acid in polar aprotic solvents containing tetrabutylammonium fluoride (TBAF). Acylation of TBA-alginate was performed in DMSO/TBAF to get products with DSacetyl up to ~ 1.0. We further report that by using appropriate solvent conditions, placement of acyl groups can be controlled to achieve either random or M-selective substitution. Alginate acetates synthesized in an M-selective fashion were used to study the ability of these derivatives to form Ca-crosslinked hydrogels. Detailed structure-property analyses were performed to identify acetylation reaction conditions and product properties that may be ideal for hydrogel formation. Furthermore, alginate esters were synthesized via modification of carboxylate groups on the backbone. These derivatives dissolved in polar aprotic solvents without the need to add TBAF. A proof of concept study showed their utility in the solubility enhancement of the poorly water soluble flavonoid naringenin. / Ph. D.