Un des enjeux majeurs dans la gestion de la plaie de pied diabétique est l’obtention d’informations permettant d’anticiper l’évolution de ces infections. Actuellement, il n’existe pas d’outils suffisamment efficaces qui permettent de distinguer une plaie colonisée d’une plaie infectée. L’approche proposée est basée sur discrimination de plusieurs bactéries fréquemment retrouvées dans les plaies chroniques de pied diabétique à partir de leur profil métabolique, et plus particulièrement des métabolites volatils qu’elles produisent. En effet, le dynamisme du métabolisme bactérien serait à même de mettre en évidence les changements qui s’opèrent dans la plaie. Dans un premier temps, une nouvelle méthodologie de concentration des métabolites volatils par Stir Bar Sorptive Extraction (SBSE) a été développée. Elle est basée sur l’utilisation de barreaux qui sont placés à la fois dans le milieu de culture et en espace de tête, suivie d’une analyse par GC-MS. La méthode a ensuite été comparée avec une autre méthode de concentration utilisant des fibres (la SPME) et a montré une meilleure capacité de concentration, permettant ainsi une détection plus sensible. Cette méthodologie a ensuite été utilisée pour suivre la production métabolique de six souches bactériennes cultivées dans des conditions mimant la plaie chronique. Grâce à leur profil métabolique, il a été possible de distinguer des espèces bactériennes. De plus, de manière plus surprenante, il a été possible de distinguer deux souches de Staphylococcus aureus présentant des profils de virulence différents. Enfin, une étude en co-culture a mis en évidence que 83% des métabolites produit en culture simple étaient retrouvés, prouvant l’intérêt de la méthodologie pour distinguer des souches bactériennes d’une même espèce au sein d’une plaie. / One of the major challenges in the management of diabetic foot wounds is to obtain information to anticipate the evolution of these infections. Currently, there are no sufficiently effective tools to distinguish a colonized wound to an infected wound. The proposed approach is based on the discrimination of several bacteria frequently found in chronic diabetic foot wounds from their metabolic profile, and more specifically the volatile metabolites they produce. Indeed, the dynamism of bacterial metabolism would be able to highlight the changes that are occurring in the wound. First, a new methodology for the concentration of volatile metabolites by Stir Bar Sorptive Extraction (SBSE) was developed. It is based on the use of stir bars that are placed both in the culture medium and in headspace, followed by GC-MS analysis. The method was then compared with another concentration method using the fibres (SPME) and we highlighted a better concentration capacity with a more sensitive detection. This methodology was then used to monitor the metabolic production of six bacterial strains grown under conditions mimicking the chronic wound. Their metabolic profile allowed us to distinguish bacterial species. Moreover, more surprisingly, it was possible to distinguish two strains of Staphylococcus aureus with different virulence profiles. Finally, a co-culture was performed and we showed that 83% of the metabolites produced in simple culture were found, proving the interest of the methodology to distinguish bacterial strains of the same species within a wound.

Identifying the best control method for problematic wildlife is an ever present issue in wildlife management. Popular control methods have ranged from lethal techniques, extirpating the animal, to multiple non-lethal methods focused on deterring undesired behavior. In the past, lethal methods were the preferred choice. However, with increased awareness of the need for biodiversity conservation, new management methods focus on non-lethal control, with emphasis on exploiting aspects of naturally occurring organismal behaviors and ecology. Over the past decade, technological advances in extraction method’s and equipment have also developed new techniques providing a broader range of information about species biology for management use. One of the most well documented conflicts between wildlife and humans is that of the wolf. Using advanced technology and new techniques, we investigated the implication of using chemosensory signals in canid urine to modify behavior as a possible non-lethal alternative in large predator management. Here we used the SBSE method coupled with improved GC/MS equipment to analyze the volatile organic compounds in the urine of four canid species, gray wolf (Canis lupus), red wolf (Canis rufus), wolf-dog hybrids (Canis familiaris) and the domestic dog (Canis familiaris) in order to create working urinary profiles. The extraction method identified several compounds also seen in the urinary profiles of other large predators. In addition, similarities and differences were also noted between taxa and the sexes, and these can be further explored in future studies. Two identified urinary compounds, acetophenone and methyl propyl sulfide, were selected for further behavioral evaluation. We focused on these compounds and their influence as chemosensory signals triggering urine marking events in both the gray wolf and red wolf. Behavioral observations of the effects of these two chemicals indicated they elicited responses from captive wolves. At each of the three study sites, the combination of these chemicals produced urine-marking events along the territory boundary by dominant animals. As a result, the investigation focused on what triggered the urine-marking events, the chemicals themselves, their combination, or the breakdown of the chemicals producing other odorants. It was found that there was no significant degradation of the chemicals over time and environmental conditions produced no significant breakdown of the acetophenone prior to the addition of methyl propyl sulfide. This posed a number of new questions and illustrated the need for additional behavioral studies. The results of this study analyzing chemosensory signals in canid urine, provides biologists with new information to aid in the development of new non-lethal management strategies for handling problematic wildlife as well as providing useful information for future research involving reproduction, predator/prey dynamics, territory maintenance, and a host of other studies focusing on animal ecology in association with chemosensory signaling.

Neste trabalho desenvolveram-se diferentes tipos de barras para extração por sorção em barras de agitação (SBSE). Esta técnica foi desenvolvida para extrair analitos orgânicos de matrizes aquosas por meio da sorção destes em uma barra de agitação. As barras c omerciais consistem em um magneto inserido no interior de um tubo de vidro, o qual é recoberto por uma camada de poli(dimetilsiloxano) (PDMS). A SBSE apresenta várias vantagens em relação às técnicas de extração convencionais, tais como: não utiliza solvente orgânico, não requer instrumentação analítica sofisticada, as barras de extração são reutilizáveis e, principalmente, possui um volume maior de recobrimento quando comparada com a microextração em fase sólida. As barras desenvolvidas para este estudo contêm fases mais seletivas, já que, comercialmente disponível, existe apenas um único tipo de recobrimento - o PDMS. Estas barras \"home-made\" foram preparadas com PDMS modificado com diferentes tipos de fases, as quais são usualmente empregadas no preparo de colunas de cromatografia gasosa. O objetivo desta modificação foi aumentar a polaridade das mesmas e, consequentemente, melhorar a eficiência e a seletividade da extração.para analitos com características mais polares. As barras desenvolvidas foram aplicadas para a análise de anticonvulsivantes e antidepressivos em plasma humano e também para a análise de anti-helmínticos em plasma bovino. Em todas as aplicações realizadas, as barras \"home-made\" demonstraram um aumento de seletividade quando comparadas à barra comercial. Além disso, as barras foram empregadas em estudos visando o acoplamento entre as técnicas SBSE e HPLC, através de uma interface desenvolvida no Laboratório de Cromatografia. O acoplamento apresentou bons resultados para a análise de antidepressivos em plasma humano, sendo observada uma maior dessorção em temperaturas mais altas, devido ao aumento da transferência de massa dos analitos, do recobrimento da barra \"home-made\" para o solvente de dessorção. / The development of different kinds of bars for stir bar sorptive extraction (SBSE) was described in this work. This technique has been applied for the extraction of organic compounds from aqueous matrices due to their sorption on the stir bar. Stir bars are composed of a magneto inserted into a glass tube and coated with a polydimethylsiloxane (PDMS) layer. SBSE presents many advantages over the conventional extraction techniques such as, solventless sample preparation, simple analytical instrumentation, reuse of stir bars and mainly the higher volume of coating when compared to solid phase microextraction. The bars have been developed with more selective phases, once the unique commercially available bar presents the disadvantage of being coated solely with PDMS polymer. The home-made stir bars were prepared using PDMS modified with different materials, often used in gas chromatography columns. The goal of this modification was to increase coating polarity and to improve efficiency and selectivity of the extraction for polar compounds. The developed bars were applied to the analysis of anticonvulsivant and antidepressant in human plasma and anthelmintic in bovine plasma. In all applications, the home-made bars showed better selectivity when compared to commercial bars. An interface for coupling SBSE to HPLC developed at Laboratory of Chromatography was also evaluated for antidepressant analysis in human plasma. This coupling presented satisfactory results, being desorption more efficient when high temperatures was used. Temperature increases affinity between analytes and desorption solvent.

Neste trabalho desenvolveram-se diferentes tipos de barras para extração por sorção em barras de agitação (SBSE). Esta técnica foi desenvolvida para extrair analitos orgânicos de matrizes aquosas por meio da sorção destes em uma barra de agitação. As barras c omerciais consistem em um magneto inserido no interior de um tubo de vidro, o qual é recoberto por uma camada de poli(dimetilsiloxano) (PDMS). A SBSE apresenta várias vantagens em relação às técnicas de extração convencionais, tais como: não utiliza solvente orgânico, não requer instrumentação analítica sofisticada, as barras de extração são reutilizáveis e, principalmente, possui um volume maior de recobrimento quando comparada com a microextração em fase sólida. As barras desenvolvidas para este estudo contêm fases mais seletivas, já que, comercialmente disponível, existe apenas um único tipo de recobrimento - o PDMS. Estas barras \"home-made\" foram preparadas com PDMS modificado com diferentes tipos de fases, as quais são usualmente empregadas no preparo de colunas de cromatografia gasosa. O objetivo desta modificação foi aumentar a polaridade das mesmas e, consequentemente, melhorar a eficiência e a seletividade da extração.para analitos com características mais polares. As barras desenvolvidas foram aplicadas para a análise de anticonvulsivantes e antidepressivos em plasma humano e também para a análise de anti-helmínticos em plasma bovino. Em todas as aplicações realizadas, as barras \"home-made\" demonstraram um aumento de seletividade quando comparadas à barra comercial. Além disso, as barras foram empregadas em estudos visando o acoplamento entre as técnicas SBSE e HPLC, através de uma interface desenvolvida no Laboratório de Cromatografia. O acoplamento apresentou bons resultados para a análise de antidepressivos em plasma humano, sendo observada uma maior dessorção em temperaturas mais altas, devido ao aumento da transferência de massa dos analitos, do recobrimento da barra \"home-made\" para o solvente de dessorção. / The development of different kinds of bars for stir bar sorptive extraction (SBSE) was described in this work. This technique has been applied for the extraction of organic compounds from aqueous matrices due to their sorption on the stir bar. Stir bars are composed of a magneto inserted into a glass tube and coated with a polydimethylsiloxane (PDMS) layer. SBSE presents many advantages over the conventional extraction techniques such as, solventless sample preparation, simple analytical instrumentation, reuse of stir bars and mainly the higher volume of coating when compared to solid phase microextraction. The bars have been developed with more selective phases, once the unique commercially available bar presents the disadvantage of being coated solely with PDMS polymer. The home-made stir bars were prepared using PDMS modified with different materials, often used in gas chromatography columns. The goal of this modification was to increase coating polarity and to improve efficiency and selectivity of the extraction for polar compounds. The developed bars were applied to the analysis of anticonvulsivant and antidepressant in human plasma and anthelmintic in bovine plasma. In all applications, the home-made bars showed better selectivity when compared to commercial bars. An interface for coupling SBSE to HPLC developed at Laboratory of Chromatography was also evaluated for antidepressant analysis in human plasma. This coupling presented satisfactory results, being desorption more efficient when high temperatures was used. Temperature increases affinity between analytes and desorption solvent.

Thesis (PhD (Chemistry and Polymer Science))--University of Stellenbosch, 2008. / The main goal of this study was the development of new technologies based on modern analytical techniques for analysis of volatiles in wines. Due to the exponential growth of the wine industry and consumer demands for an enjoyable, safe-to-consume, and high quality product, the need for arose for methodologies aiding the understanding of wine better arose. Chemical analysis is a valuable way of studying the composition of wine in depth. Very sophisticated instrumentation is available nowadays but almost always the sample needs to be cleaned up or concentrated before such analysis. This study investigates the use of stir bar sorptive extraction (SBSE) as such a technique. It is shown that SBSE combined with gas chromatography-mass spectrometry (GC-MS) is extremely suited for a wide number of analyses and during the course of the study the technique was applied for troublesome analytical challenges in various beverages and foodstuffs. The study focuses on the development of a screening technique for volatiles in wine using SBSE and the application of the data to various chemometrical techniques for classification purposes. A second part of the study shows the applicability of SBSE for extraction of pesticides, contaminants and preservatives from wine, water, lemon flavoured beverages and yoghurt. The method is also elaborated upon by development of faster analysis methods for wine and beer and the investigation of using SBSE for headspace sampling of wine. In all the applications, SBSE technology was shown to be sensitive, repeatable, robust and very simple to use.

Este trabalho apresenta o desenvolvimento de diferentes métodos para a determinação de fluoxetina, norfluoxetina e bisfosfonatos, utilizando técnicas modernas de preparo de amostras. Dentre eles, um método simples e sensível, empregando microextração em fase sólida (SPME) e cromatografia líquida foi desenvolvido para a análise de fluoxetina e norfluoxetina em plasma. As condições de SPME foram otimizadas empregando planejamento fatorial. A extração foi realizada utilizando fibras com PDMS-DVB, e a etapa de dessorção foi realizada em uma nova interface homemade com sistema de aquecimento. Extração com sorção em barra de agitação (SBSE) com derivatização in-situ, combinada com dessorção com solventes ou dessorção térmica, acoplada à cromatografia gasosa e espectrometria de massas (GC-MS), foi também empregada para a determinação de fluoxetina em plasma. Avaliou-se a derivatização dos analitos in situ com cloroformato de etila, bem como os parâmetros tempo de extração, solvente e tempo de dessorção. A combinação de SBSE e LC-MS foi também avaliada. As amostras de plasma foram submetidas à precipitação de proteínas com ácido tricloroacético, extraídas com SBSE, e analizadas por LC-MS. Os métodos SPME-LC, SBSE-GC e SBSE-LC desenvolvidos foram completamente validados, demonstrando serem adequados para analisar fluoxetina em amostras de plasma. Métodos rápidos para a determinação de etidronato, clodronato, pamidronato e alendronato, baseados em cromatografia de troca iônica com detecção indireta no ultravioleta, foram também desenvolvidos. Os métodos são simples e demonstraram precisão, exatidão e especificidade. Além disso, os métodos validados empregam colunas de sílica, mais baratas e de mais disponibilidade do que as colunas poliméricas, frequentemente utilizadas em métodos descritos na literatura para o mesmo propósito. / This study describes different methods to analyze fluoxetine, norfluoxetine and bisphosphonates, employing modern sample preparation techniques. A simple and sensitive procedure using solid-phase microextraction (SPME) coupled with liquid chromatography was developed for the analysis of fluoxetine and norfluoxetine in plasma samples. SPME conditions were optimized employing a factorial design. The sampling step was performed using a PDMS-DVB fiber and desorption was carried out in a novel homemade heated interface. Stir bar sorptive extraction (SBSE) with in situ derivatization, in combination with either thermal or liquid desorption on-line coupled to gas chromatography-mass spectrometry (GC-MS), was employed for the analysis of fluoxetine. In situ derivatization using ethylchloroformate was evaluated as well as parameters such as solvent polarity, time for analytes desorption, and extraction time. SBSE combined with liquid chromatography and mass spectrometry was also used to analyze fluoxetine in plasma. Plasma samples were first submitted to protein precipitation with trichloroacetic acid, subjected to SBSE, and thereafter analyzed by LC-MS. SPME-LC, SBSE-GC, and SBSE-LC methods were completely validated showing to be adequate to assess fluoxetine in plasma samples. Rapid methods for etidronate, clodronate, pamidronate and alendronate assay were also developed. The methods were based on ion chromatography with indirect ultraviolet detection, which avoids the need for chemical derivatization procedures. The methods were simple, rapid, and demonstrated precision, accuracy, and specificity. Furthermore, they employed silica-based columns, cheaper and more readily available than polymeric columns, frequently used in previous reported methods.

Este trabalho apresenta o desenvolvimento de diferentes métodos para a determinação de fluoxetina, norfluoxetina e bisfosfonatos, utilizando técnicas modernas de preparo de amostras. Dentre eles, um método simples e sensível, empregando microextração em fase sólida (SPME) e cromatografia líquida foi desenvolvido para a análise de fluoxetina e norfluoxetina em plasma. As condições de SPME foram otimizadas empregando planejamento fatorial. A extração foi realizada utilizando fibras com PDMS-DVB, e a etapa de dessorção foi realizada em uma nova interface homemade com sistema de aquecimento. Extração com sorção em barra de agitação (SBSE) com derivatização in-situ, combinada com dessorção com solventes ou dessorção térmica, acoplada à cromatografia gasosa e espectrometria de massas (GC-MS), foi também empregada para a determinação de fluoxetina em plasma. Avaliou-se a derivatização dos analitos in situ com cloroformato de etila, bem como os parâmetros tempo de extração, solvente e tempo de dessorção. A combinação de SBSE e LC-MS foi também avaliada. As amostras de plasma foram submetidas à precipitação de proteínas com ácido tricloroacético, extraídas com SBSE, e analizadas por LC-MS. Os métodos SPME-LC, SBSE-GC e SBSE-LC desenvolvidos foram completamente validados, demonstrando serem adequados para analisar fluoxetina em amostras de plasma. Métodos rápidos para a determinação de etidronato, clodronato, pamidronato e alendronato, baseados em cromatografia de troca iônica com detecção indireta no ultravioleta, foram também desenvolvidos. Os métodos são simples e demonstraram precisão, exatidão e especificidade. Além disso, os métodos validados empregam colunas de sílica, mais baratas e de mais disponibilidade do que as colunas poliméricas, frequentemente utilizadas em métodos descritos na literatura para o mesmo propósito. / This study describes different methods to analyze fluoxetine, norfluoxetine and bisphosphonates, employing modern sample preparation techniques. A simple and sensitive procedure using solid-phase microextraction (SPME) coupled with liquid chromatography was developed for the analysis of fluoxetine and norfluoxetine in plasma samples. SPME conditions were optimized employing a factorial design. The sampling step was performed using a PDMS-DVB fiber and desorption was carried out in a novel homemade heated interface. Stir bar sorptive extraction (SBSE) with in situ derivatization, in combination with either thermal or liquid desorption on-line coupled to gas chromatography-mass spectrometry (GC-MS), was employed for the analysis of fluoxetine. In situ derivatization using ethylchloroformate was evaluated as well as parameters such as solvent polarity, time for analytes desorption, and extraction time. SBSE combined with liquid chromatography and mass spectrometry was also used to analyze fluoxetine in plasma. Plasma samples were first submitted to protein precipitation with trichloroacetic acid, subjected to SBSE, and thereafter analyzed by LC-MS. SPME-LC, SBSE-GC, and SBSE-LC methods were completely validated showing to be adequate to assess fluoxetine in plasma samples. Rapid methods for etidronate, clodronate, pamidronate and alendronate assay were also developed. The methods were based on ion chromatography with indirect ultraviolet detection, which avoids the need for chemical derivatization procedures. The methods were simple, rapid, and demonstrated precision, accuracy, and specificity. Furthermore, they employed silica-based columns, cheaper and more readily available than polymeric columns, frequently used in previous reported methods.

Extraction is required to separate and concentrate trace level analytes from the sample matrix prior to gas chromatography (GC). Classical extraction procedures utilise large amounts of hazardous solvents, generate waste, and sensitivity limitations are associated with the injection of microlitre amounts of the final solvent extract. In response to real world challenges, and to overcome the problems associated with solvent extraction, novel silicone rubber (polydimethylsiloxane (PDMS)) samplers were developed for solvent free enrichment of trace level analytes from indoor air, contaminated soil, desert soil, ultra high temperature (UHT) milk and Pinotage wine. Versatile PDMS samplers as a loop, a multichannel trap, or a denuder for trace environmental forensics, geochemical and aroma investigations are presented. A unique off-line multidimensional GC approach involving heart-cut gas chromatographic fraction collection is described, as is off-line olfactory assessment of recombined heart-cuts for synergistic odour effects. PDMS loop samplers were used for the extraction of DDT (1,1,1- trichloro-2,2-bis(p-chlorophenyl)ethane) and its associated environmental pollutants from soil samples. Miniature denuder samplers accomplished separate concentration of vapour phase and of particulate phase fractions of DDT and its associated environmental pollutants from indoor air, in a single step. Ratios of airborne p,p’-DDD/p,p’-DDT and of o,p’-DDT/p,p’-DDT are unusual and do not match the ideal certified ingredient composition required of commercial DDT. Results suggest that commercial DDT used for indoor residual spraying may have been compromised with regards to insecticidal efficacy, demonstrating the power of this new environmental forensics tool. Multichannel PDMS trap samplers were used in a unique heart-cut multidimensional GC approach for off-line enantiomeric separation of o,p’-DDT and o,p’-DDD in air and soil. This alternative multidimensional method is compared to the complementary technique of comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry detection (GCxGC-TOFMS). PDMS loop samplers were also employed for the solvent free extraction of hydrocarbons from desert soil to investigate, for the first time, a possible geochemical origin of the enigmatic fairy circles of Namibia. It is proposed that microseepages of natural gas and low volatility hydrocarbons are expressed at the surface as a geobotanical anomaly of barren circles and circles of altered vegetation. Multichannel PDMS trap samplers were utilised for sampling of the headspace of UHT milk and of Pinotage wine, and to study off-line, using a portable olfactometer, synergistic effects between recombined heart-cut aroma compounds. Olfactory results show that a synergistic combination of 2- heptanone and 2-nonanone was responsible for a pungent cheese-like odour in UHT milk, while a synergistic combination of furfural and 2-furanmethanol was responsible for a roast coffee bean-like odour in coffee style Pinotage wine. The small, low cost samplers are quick and easy to assemble and they fit commercial thermal desorber systems. The PDMS samplers are reusable. Solvent extraction of the sampling materials, extract clean-up and pre-concentration are not required. Thus, potential loss of analyte, introduction of contaminants and waste disposal are minimised. Solvent free thermal desorption permits transfer of the entire sample mass to the cooled injection system (CIS) inlet of a GC resulting in greater sensitivity when compared to injection of microlitre amounts of a solvent extract. This allowed for sampling of smaller sized soil samples, shorter air sampling times and lower air sampling flow rates when compared to solvent based methods. / Thesis (PhD)--University of Pretoria, 2013. / Chemistry / unrestricted

A depressão em idosos é uma desordem persistente e recorrente, resultado do stress psicossocial ou efeito de doenças fisiológicas, que podem acarretar a desabilidade do indivíduo, aumento dos sintomas das doenças clínicas, na maior utilização dos serviços de saúde e altas taxas de suicídios.A monitorização terapêutica permite a ndividualização do regime de dosagem, assegurando a eficácia clínica e minimizando os efeitos adversos dos fármacos, prescritos na clínica. Os antidepressivos têm sido monitorados, pois, apresentam intervalos terapêuticos bem estabelecidos, ou seja, a maioria dos pacientes, que apresentam concentrações plasmáticas dentro deste intervalo fixo, tem as desordens psiquiátricas mantidas sob controle e efeitos adversos aceitáveis. Os antidepressivos tricíclicos (ADTs): imipramina, amitriptilina, nortriptilina e desipramina, embora eficazes e ainda muito utilizados, apresentam efeitos adversos, não desejáveis. Os antidepressivos, inibidores seletivos da recaptação de serotonina (SSRIs): citalopram, fluoxetina, paroxetina e sertralina, apresentam eficácia clínica comparável aos clássicos ADTs, mas destituídos dos efeitos adversos associados aos mesmos. Os métodos convencionais, empregados no tratamento de amostras biológicas, para análises de antidepressivos por técnicas cromatográficas, têm sido a extração líquido-líquido e extração em fase sólida. A microextração em fase sólida tem sido empregada em diferentes análises em fluidos biológicos, porém essa técnica apresenta certas limitações como, o número limitado de fases extratoras disponíveis no comércio que sejam adequadas para a análise de compostos não iônicos. A avaliação de novas fases extratoras, mais seletivas, estáveis e de baixo custo tem sido requerida para a análise de fármacos. O interesse no uso de poli(pirrol) (PPY), como fase extratora para SPME, está relacionado às diferentes interações dos fármacos (hidrofóbicas, -, com o grupo funcional polar, troca iônica, ácido-base) aos grupos multifuncionais deste polímero. Sua polimerização pode ser alcançada tanto por oxidação química, quanto por eletrodeposição em meio aquoso ou orgânico. O método eletroquímico apresenta algumas vantagens, tais como: o polímero ou mistura de polimeros podem ser revestidos diretamente em um metal, incorporação de diferentes grupos funcionais, entre outras, a polimerização pode ser eletroquimicamente controlada através da voltametria cíclica. Neste trabalho o poli(pirrol) (PPY) foi eletrodepositado em eletrodo de aço inox e empregado para a microextração em fase sólida dos antidepressivos: paroxetina, fluoxetina, mirtazapina, duloxetina, sertralina e citalopram. As variáveis da eletrodeposição, número de ciclos e contra-íon empregado no processo de eletrodeposição foram otimizadas. Assim como as variáveis SPME: tempo, temperatura, pH e volume de amostra, e tempo e solvente de dessorção; almejando maior sensibilidade para o método SPME-PPY/LC UV proposto. A extração sortiva em barra de agitação (SBSE), técnica recente de preparo de amostras, para a pré-concentração de compostos orgânicos presentes em amostras biológicas, baseia-se na extração estática, através do polímero polidimetilsiloxano (PDMS), no qual ocorre a dissolução (sorção) do analito. Neste trabalho, as técnicas SBSE e cromatografia líquida de alta eficiência com detecção UV (SBSE/LC UV) foram avaliadas para a análise simultânea de antidepressivos em amostras de plasma para fins de monitorização terapêutica. As condições cromatográficas de análise, assim como as variáveis SBSE de extração (tempo, temperatura, força iônica, pH da matriz) e tempo de dessorção, foram otimizadas, visando adequada sensibilidade analítica. A validação analítica foi realizada segundo normas da ANVISA, para ambos os métodos propostos, em diferentes concentrações plasmáticas, as quais contemplam o intervalo terapêutico. Segundo os parâmetros de validação avaliados, os métodos SBSE/LCUV e SPMEPPY/LCUV padronizados poderão ser empregados nas análises dos antidepressivos, para fins de monitorização terapêutica. / Depression in the elderly is a persistent and recurrent disorder resulting from psychosocial stress or physiological effect or disease. This condition can lead to disability, cognitive impairment, enhanced symptoms of medical illnesses, increased use of health care services and, increased of suicide rates. Therapeutic monitoring allows individualization of the dose regimen, ensuring clinical effectiveness and minimizing the adverse effects of drugs, prescribed at the clinic. Antidepressants have been monitored because they present well - established therapeutic intervals; in other words, most of the patients present plasmatic concentrations within this fixed range, so that their psychiatric disorders are kept under control and the adverse effects are acceptable. The tricyclic antidepressants (ADTs) imipramine, amitriptyline, nortryptiline, and desipramine, have adverse effects. The selective serotonin reuptake inhibitors (SSRIs) antidepressants citalopram, fluoxetine, paroxetine, and sertraline, are clinically effectiveness as the classic ADTs, but they do not lead to the adverse effects associated to the latter. The conventional methods employed in the treatment of biological samples for analysis of antidepressants by chromatographic techniques have been the liquid-liquid extraction (LLE) and solid phase extraction (SPE) techniques. Solid-phase microextraction (SPME) has been used in various analyses of biological fluids. However, this technique has limitations such as the small number of comemercially available extracting phases that are appropriate for the analysis of non-ionic compounds. Investigation of new extraction phases that are more selective and stable, as well as inexpensive, has been requested for drug analysis. The interest in the use of poly(pirrole) (PPY) as an extraction phase for SPME is related to the different interactions of the drugs (hydrophobic, - , with the polar functional group, ionic exchange and acid-base) with the multifunctional groups on this polymer. The PPY polymerization can be achieved by chemical oxidation or electropolymerization in aqueous solution or organic solvents. The electrochemical method has advantages such as, the polymer or a mixes of polymers can be directly deposited on a metal wire, different functional groups can be incorporated, polymerization can be electrochemically controlled by cyclic voltammetry. In this work poly(pyrrole) was electropolymerized on stainless steel electrodes and employed for the solid phase microextraction of the antidepressants paroxetine, fluoxetine, mirtazapine, duloxetine, sertraline, and citalopram. The electropolymerization variables, the number of cycles and counterion employed in the process were optimized, as well as the SPME variables, time, temperature, pH, sample volume, and desorption solvent; aiming at a better sensibility for the proposed method SPME-PPY/LC-UV. The stir bar sorptive extraction (SBSE), recently established technique for samples preparation, that targets the pre concentration of organic compounds present in biological samples. It is based on static extraction, through the polymeric polydimethylsiloxane (PDMS), where there is analyte dissolution (sorption). In this work, the SBSE technique and liquid chromatography with UV detector (SBSE/LC-UV) were evaluated for the simultaneous determination of antidepressants in plasma samples for therapeutic monitoring purposes. The cromatographic conditions, the SBSE extraction variables (time, temperature, ionic strength and matrix pH), and the dessorption time were optimized for appropriate analytical sensibility. The analytical validation was accomplished according to the norms of ANVISA for both of the proposed methods, in different plasmatic concentrations, which contemplate the therapeutic interval. According to the evaluated validation parameters, the standardized methods SBSE/LC-UV and SPME-PPY/LC-UV can be used in the analyses of antidepressants for therapeutic drug monitoring purposes.

Uma fase polimérica inédita de polidimetilsiloxano/carvão ativado (PDMS-ACB) é proposta como fase extratora para extração por sorção em barras de agitação (SBSE). A barra de PDMS-ACB foi desenvolvida no laboratório usando um molde de teflon simples, demonstrando estabilidade e resistência aos solventes orgânicos. Utilizando uma única barra de PDMS-ACB mais de 150 extrações foram realizadas sem qualquer dano. A barra de SBSE contém aproximadamente 100 µL de revestimento polimérico, 2,36 mm de diâmetro e 2,2 cm de comprimento. A barra PDMS-ACB foi aplicada na extração de seis pesticidas (ametrina, atrazina, bifentrina, carbofurano, metribuzim e tebutiurom), com propriedades predominantemente polares, comumente aplicados em plantações de cana-de-açúcar. A barra PDMS-ACB foi utilizada na determinação de pesticidas em garapa através de cromatografia gasosa hifenada à espectrometria de massa (GC-MS). O planejamento experimental foi empregado na etapa de otimização da extração SBSE, um planejamento fatorial fracionado avaliou a influência dos principais parâmetros envolvidos. Posteriormente, o planejamento composto central (CCD) com conformação estrela foi explorado para otimizar os fatores significativos na extração. Apenas 200 µL de acetato de etila foram utilizados como solvente na dessorção líquida (LD). Para os compostos avaliados, o método apresentou limite de quantificação (LOQ) no intervalo de 0,5 - 40 µg L-1, as recuperações variaram entre 0,18 - 49,50 % e a precisão intra-dia variou de 0,072 - 8,40 %. Concluída a etapa de validação, o método foi aplicado em amostras reais de garapa disponíveis comercialmente em São Carlos-SP. / A novel polydimethylsiloxane/activated carbon (PDMS-ACB) is proposed as new polymeric phases for stir bar sorptive extraction (SBSE). The PDMS-ACB was developed in lab using simple teflon\'s mold, demonstrating remarkable stability and resistance to organic solvents, over 150 extractions without any damage. SBSE bar contained 100 µL of polymeric coating, a diameter of 2.36 mm and a length of 2.2 cm. It was applied to the determination of pesticides (ametryn, atrazine, bifenthrin, carbofuran, metribuzine and tebuthiuron) having predominantly polar properties, applied in sugarcane crops. PDMS-ACB was employed in the determination of pesticides in sugarcane juice using gas chromatography coupled to mass spectrometry (GC-MS). Experimental design was employed in the optimization step, a fractional factorial evaluated the main parameters involved in the extraction procedure. Afterwards, central composite design (CCD) was exploited to optimize the significant factors on the extraction. About 200 µL of ethyl acetate were employed as solvent in the liquid desorption (LD). The method presented limit of quantification (LOQ) from 0.5 to 40 µg L-1, recoveries varied 0.18 - 49.50% and precision intra-day 0.072 - 8.40%. Hence, the method was applied to the analysis of real sugarcane juice samples commercially available in São Carlos-SP.