Neste trabalho estudou-se o desenvolvimento de metodologia analitica para a determinação e quantificação de sete anti-inflamatórios não esteroides, amplamente consumidos para tratamento humano, além da cafeína, determinada como marcador antropogênico, em amostras aquosas ambientais. Os analitos estudados foram acetaminofeno (AC), ácido salicílico (AS), cafeína (CA), cetoprofeno (CE), diclofenaco (DC), dipirona (DP), ibuprofeno (IB) e naproxeno (NA) . Como padrão internos, foi utilizado o lindano (LI). A determinação destes analitos foi realizada utilizando-se cromatografia a gás acoplada à espectrometria de massas (GC-MS) utilizando MSTFA/Imidazol como derivatizante. As condições para a reação de derivatização foram otimizadas utilizando o planejamento Doehlert. O procedimento de pré-concentração utilizado foi a extração em fase sólida (SPE), tendo como sorvente um sistema de fase polimérica reversa. O procedimento de SPE foi baseado em trabalhos publicados na literatura, com metanol e água (ajustada a pH 3,0) para o condicionamento e eluição com acetonitrila. Os parâmetros de desempenho do método, como recuperação, linearidade, limite de detecção, limite de quantificação, repetitividade e reprodutibilidade foram avaliados de acordo com as normas estabelecidas pela ANVISA e INMETRO. Os limites de detecção ficaram entre 1,09 e 1079,14 ng L-1 e os de quantificaçõ entre 3,63 e 3597,12 ng L-1. Após as definições em relação ao método, amostras de água do Arroio Dilúvio (em Porto Alegre – RS) e de uma Estação Piloto de tratamento de efluente pertencente ao Instituto de Pesquisas Hidráulicas (IPH/UFRGS) , foram analisadas para avaliar a presença destes analitos. As amostras da ETE-piloto do IPH, foram coletadas nos meses de janeiro e fevereiro e as amostras do Arroio Dilúvio também foram coletadas no mesmo período. Considerando as amostras reais analisadas em algumas foram detectados e em outras quantificados os antiinflamatórios estudados. Nas amostras que estes compostos foram quantificados, a faixa de concentração variou entre 0,09 e 23,16 μg L-1. / In this work the development of an analytic methodology was studied to determine and quantify seven non-steroidal anti-inflammatory drugs, widely used in human treatment, and caffeine considered an anthropogenic marker. The compounds studied were acetaminophen (AC), salicylic acid (AS), caffeine (CA), ketoprofen (CE), diclofenac (DC), dipyrone (DP), ibuprofen (IB), naproxen (NA) and lindane (LI) as internal standard. The determination of these compounds was performed using gas chromatography coupled to mass spectrometry (GC-MS) with derivatization of the target analytes using MSTFA/Imidazole. The derivatization conditions were optimized with Doehlert design. Solid phase extraction (SPE) with a polymeric reversed solid phase was used as a pre-concentration system. The SPE procedure conditions were based previously established data in the literature, using methanol and water (adjusted to pH 3.0) for the conditioning step and acetonitrile for the elution step The performance parameters, such recovery, linearity, detection limit, quantification limit, repeatability and reproducibility were evaluated according to ANVISA and INMETRO determinations. The detection limits was in the range of 1,09 and 1079,14 ng L -1 and the quantification limits 3,63 and 3597,12 gn L-1. After method establishment, samples from a piloto sewage treatment plant and river water, both in Porto Alegre-RS, were analysed. The samples were collected in January and February 2012, both in the pilot plant and in the river water. In some real samples the antiinflammatories were detected and in others was possible quantify. In the samples were the analytes were quantified, the concentration range was 0,09 to 23,16 μg L-1.

Neste trabalho estudou-se o desenvolvimento de metodologia analitica para a determinação e quantificação de sete anti-inflamatórios não esteroides, amplamente consumidos para tratamento humano, além da cafeína, determinada como marcador antropogênico, em amostras aquosas ambientais. Os analitos estudados foram acetaminofeno (AC), ácido salicílico (AS), cafeína (CA), cetoprofeno (CE), diclofenaco (DC), dipirona (DP), ibuprofeno (IB) e naproxeno (NA) . Como padrão internos, foi utilizado o lindano (LI). A determinação destes analitos foi realizada utilizando-se cromatografia a gás acoplada à espectrometria de massas (GC-MS) utilizando MSTFA/Imidazol como derivatizante. As condições para a reação de derivatização foram otimizadas utilizando o planejamento Doehlert. O procedimento de pré-concentração utilizado foi a extração em fase sólida (SPE), tendo como sorvente um sistema de fase polimérica reversa. O procedimento de SPE foi baseado em trabalhos publicados na literatura, com metanol e água (ajustada a pH 3,0) para o condicionamento e eluição com acetonitrila. Os parâmetros de desempenho do método, como recuperação, linearidade, limite de detecção, limite de quantificação, repetitividade e reprodutibilidade foram avaliados de acordo com as normas estabelecidas pela ANVISA e INMETRO. Os limites de detecção ficaram entre 1,09 e 1079,14 ng L-1 e os de quantificaçõ entre 3,63 e 3597,12 ng L-1. Após as definições em relação ao método, amostras de água do Arroio Dilúvio (em Porto Alegre – RS) e de uma Estação Piloto de tratamento de efluente pertencente ao Instituto de Pesquisas Hidráulicas (IPH/UFRGS) , foram analisadas para avaliar a presença destes analitos. As amostras da ETE-piloto do IPH, foram coletadas nos meses de janeiro e fevereiro e as amostras do Arroio Dilúvio também foram coletadas no mesmo período. Considerando as amostras reais analisadas em algumas foram detectados e em outras quantificados os antiinflamatórios estudados. Nas amostras que estes compostos foram quantificados, a faixa de concentração variou entre 0,09 e 23,16 μg L-1. / In this work the development of an analytic methodology was studied to determine and quantify seven non-steroidal anti-inflammatory drugs, widely used in human treatment, and caffeine considered an anthropogenic marker. The compounds studied were acetaminophen (AC), salicylic acid (AS), caffeine (CA), ketoprofen (CE), diclofenac (DC), dipyrone (DP), ibuprofen (IB), naproxen (NA) and lindane (LI) as internal standard. The determination of these compounds was performed using gas chromatography coupled to mass spectrometry (GC-MS) with derivatization of the target analytes using MSTFA/Imidazole. The derivatization conditions were optimized with Doehlert design. Solid phase extraction (SPE) with a polymeric reversed solid phase was used as a pre-concentration system. The SPE procedure conditions were based previously established data in the literature, using methanol and water (adjusted to pH 3.0) for the conditioning step and acetonitrile for the elution step The performance parameters, such recovery, linearity, detection limit, quantification limit, repeatability and reproducibility were evaluated according to ANVISA and INMETRO determinations. The detection limits was in the range of 1,09 and 1079,14 ng L -1 and the quantification limits 3,63 and 3597,12 gn L-1. After method establishment, samples from a piloto sewage treatment plant and river water, both in Porto Alegre-RS, were analysed. The samples were collected in January and February 2012, both in the pilot plant and in the river water. In some real samples the antiinflammatories were detected and in others was possible quantify. In the samples were the analytes were quantified, the concentration range was 0,09 to 23,16 μg L-1.

A monitorização terapêutica permite a individualização do regime de dosagem, assegurando a eficácia clínica e minimizando os efeitos adversos dos fármacos prescritos. Atualmente, as estatinas têm sido monitoradas, pois, embora eficazes e muito utilizadas, apresentam alguns efeitos adversos não desejáveis. Neste trabalho, as técnicas SPME e cromatografia gasosa acoplada à espectrometria de massas (GC-MS) foram avaliadas para a análise de estatinas (fluvastatina, simvastatina e atorvastatina) em plasma humano para fins de monitorização terapêutica. As condições de extração e derivatização foram otimizadas empregando planejamento experimental e avaliando a influência dos principais parâmetros envolvidos. Para otimização das condições de extração avaliou-se parâmetros como tempo e temperatura de extração, pH, volume de solução tampão e força iônica; e para otimização das condições de derivatização avaliou-se parâmetros como volume de reagente derivatizante, volume de agente pareador iônico e pH. A extração foi realizada utilizando fibras de PDMS-DVB, e a dessorção feita termicamente no injetor do cromatógrafo a gás. O método desenvolvido foi validado segundo normas da ANVISA, apresentando linearidade na faixa de 20 a 500 ng mL-1, precisão com coeficientes de variação menor que 14% e recuperação relativa de 20 a 40%. Concluída a validação analítica, a metodologia proposta foi aplicada em amostras reais de plasma de pacientes em terapia com simvastatina concedidas pelo Laboratório Médico Dr. Maricondi e pela Casa de Saúde de São Carlos. / Therapeutic drug monitoring allows individualization of dosage regimen, ensuring the clinical efficacy and minimizing the adverse effects of prescribed drugs. Currently, statins have been monitored; despite they are effective, some statins have undesirable adverse effects. In this work, SPME technique and gas chromatography / mass spectrometry (GC-MS) were evaluated for analysis of statins (fluvastatin, simvastatin and atorvastatin) in human plasma for therapeutic drug monitoring. The extraction and derivatization conditions were optimized using experimental design and evaluating the influence of the main parameters involved in the SPME procedure. To optimize the extraction conditions were evaluated parameters such as time and extraction temperature, pH, volume of buffer and ionic strength, and to optimize the derivatization conditions were evaluated parameters as derivatization reagent volume, ionic agent pareador volume and pH. The extraction was performed using PDMS-DVB fibers, and thermal desorption performed in the injector of the gas chromatograph. The method was validated according to ANVISA, showing linearity in the range 20 to 500 ng mL-1, the precision with coefficients of variation less than 14% and relative recovery from 20 to 40%. The proposed methodology was applied to real samples of plasma from patients on therapy with simvastatin provided by the Laboratório Médico Dr. Maricondi and Casa de Saúde of São Carlos.

Esta tese tem como objetivo a derivatização da celulose sob condições homogêneas de reação, através de um método simples, reprodutível e eficiente. As três etapas de derivatização da celulose (ativação, dissolução, e reação) foram estudadas com ênfase na relação entre a estrutura do biopolímero, as condições da reação e as propriedades dos derivados obtidos. Foi realizado estudo do intumescimento de celuloses (microcristalina, algodão, eucalipto, algodão mercerizado e eucalipto mercerizado) em solventes próticos e apróticos. O intumescimento foi correlacionado com as propriedades dos solventes. Nos próticos, as celuloses nativas e mercerizadas foram sensíveis aos mesmos quatro descritores (acidez, basicidade, dipolaridade/polarizabilidade e volume molar), porém, a contribuição relativa dessas propriedades foi diferente, sendo que no caso da celulose MC e das nativas a dipolaridade/polarizabilidade foi mais significativo e para as celuloses mercerizadas o volume molar foi o descritor mais importante. Para o intumescimento em solventes apróticos, para as celuloses mercerizadas, boas correlações foram obtidas empregando apenas um descritor (dipolaridade/polarizabilidade). No intumescimento intracristalino, investigamos as mudanças nas propriedades físico-químicas após a mercerização. Este tratamento resultou em: ocorreu aumento no teor de α-celulose; diminuição da cristalinidade; redução da espessura da fibra; remoção de fragmentos da superfície; aumento na área superficial. Salientamos que não ocorre uma diminuição significativa no grau de polimerização das celuloses. Empregamos uma nova classe de solvente (líquidos iônicos) para a dissolução e subsequente funcionalização da celulose. Tanto na dissolução por aquecimento térmico ou por micro-ondas a integridade física da celulose foi mantida. Alem disto, observamos que a dissolução facilita a acessibilidade/reatividade da celulose, pois ocorre a diminuição do índice de cristalinidade e o aumento da área superficial. O uso de aquecimento por micro-ondas mostrou-se mais rápido e eficiente do que aquecimento térmico, sendo que tempo necessário para a dissolução e acilação da celulose foi de 25 a 30% menor daquele com aquecimento térmico. O líquido iônico (AlBuImCl) mostrouse um excelente solvente em termos de controle do GS e reprodutibilidade do mesmo. Realizou-se um estudo da dependência das propriedades dos acetatos de celulose e seus filmes com o grau de substituição do biopolímero. Foram empregadas sondas solvatocrômicas e medidas térmicas para investigar as propriedades de filmes de acetatos de celulose. Os resultados obtidos mostraram que a dependência de tais propriedades sobre o grau de substituição dos acetatos de celulose pode ser descrita por equações simples. Por fim, desenvolveu-se um método simples, rápido, e preciso para a determinação do GS de ésteres de celulose (acetato, butirato e hexanoato), através do uso de sondas solvatocrômicas / The objective of this thesis is to study some aspects of the derivatization of cellulose under homogeneous reaction conditions. The three steps of this reaction scheme (activation, dissolution and derivatization) were studied, with emphasis on the relationship between the biopolymer structure, the conditions of the reaction and the properties of the products. Swelling of different celluloses (microcrystalline (MC), cotton linter, eucalyptus, cotton linter mercerized, eucalyptus mercerized) by protic and aprotic solvents was determined. The results were correlated with a series of solvent descriptors. For protics solvents, although native and mercerized celluloses are sensitive to the same set of four solvent properties (acidity, basicity, dipolarity/polarizability, and molar volume), the relative contributions are different; π* is most important for MC and native celluloses, VS is most important for mercerized celluloses. Because of absence of hydrogen-bon donation by aprotics solvents, a smaller number of solvent descriptors is required in order to describe cellulose swelling. For example, the swelling of mercerized cellulose can be reasonably described by a single descriptor (dipolarity/polarizability). The effect of mercerization on cellulose properties was investigated; it leads to: increase of α-cellulose content; decrease of Ic; decrease of fiber cross section, smoothing of its surface, and increase of its surface area. The ionic liquid 1-allyl-3-(1-butyl)imidazolium chloride showed to be a convenient reaction medium for the dissolution and derivatization of cellulose, with reproducible easily controlled degree of substitution, DS. The physical integrity of the cellulose is maintained during dissolution by using either \"conventional\" or convection-, or microwave heating. Compared with the former, MW irradiation has resulted in considerable decrease in dissolution- and reaction times. The value DS was found to be DSethanoate > DSpropanoate > DSbutanoate. The values of DSpentanoate and DShexanoate were found to be slightly higher than DSethanoate. The dependence of the properties of cellulose acetates and their films on the degree of biopolymer substitution was investigated. Solvatochromic probes and thermal analysis have been employed. The results showed that the dependence of the properties (solvatochromic parameters and thermal properties) of CAs on their DS can be described by simple equations; a consequence of the substitution of the OH by the ester group. Finally, an expedient, accurate method for the determination of the degree of substitution of cellulose carboxylic esters (ethanoate, butanoate and hexanoate) was developed, using Uv-vis spectroscopy (dye perichromism)

Aim of the master thesis was the development and the optimization of the HPLC method with fluorescence detection for the determination of the bile acids in the bile of mice. On the basis of literature, the HPLC-FLD method has been proposed and optimized for the quantification of eight bile acids (cholic acid, glycocholic acid, deoxycholic acid, glycodeoxycholic acid, chenodeoxycholic acid, glycochenodeoxycholic acid, α-muricholic acid, and λ-muricholic acid) in the biological material. 1,2-benzo-3,4-dihydrocarbazole-9-ethyl-p-toluenesulfonate is suitable derivatization reagent for labeling of the bile acids. The calibration dependences, limits of detection and quantification (in the range of 0.88-1.78 μmol dm -3 ) for all bile acids are measured. In the biological sample five bile acids have been successfully identified and then determined (cholic acid, glycocholic acid, glycodeoxycholic acid, glycochenodeoxycholic acid, and α-muricholic acid). Key words: bile acids, HPLC, fluorescence detector, derivatization reaction, bile

Short-chain fatty acids (SCFAs) and the tricarboxylic acid (TCA) cycle metabolites aresmall hydrophilic compounds that play crucial roles in biological species ranging fromenergy metabolism, immune homeostasis to cellular signalling. There is a need for reliableand precise quantification of these metabolites in biological matrices as they can providecrucial information of metabolic status and potentially be used as diagnostic biomarkersfor different pathological and physiological conditions. However, their retention andseparation in traditional reversed-phase system, without chemical derivatization, is oftenproblematic due to their volatile and hydrophilic characteristics. The aim of this studywas to implement a facile and effective derivatization method for the simultaneousquantitation of SCFAs and TCA cycle metabolites by LC-qToF-MS in negative ion mode. Inthis work, 3-nitrophenylhydrazine (3-NPH) was employed for preanalyticalderivatization to convert the compounds to their respective 3-nitrophenylhydrazones.Analytical standards and faecal samples were used to assess the linearity, matrix effect,accuracy, extraction efficiency, precision, retention-time shift and short-term stability.The compounds were successfully separated within 6 minutes on a reverse-phase C18column. All the compounds showed good linearity (R2≥ 0.97) in both solvent-only andfaecal samples. The matrix effect was minimal and did not affect the compoundsquantitation. The extraction efficiency ranged from 80% to 110% (CV≤9.7%, n = 6). Theaccuracy of quantitation was determined to be between 82.8% to 113.8% (CV≤9.0%, n =6). The intra-day (CV%) demonstrated good precision for all analytes, the inter-day (%)were more variable due to the derivatives’ chemical instability. However, most of thederivatives were chemical stable up to 5 days in the autosampler (10°C). The method wasalso applied to explore the levels of these metabolites in human faecal samples and mousebrain samples.

This diploma thesis deals with the development and validation of an analytical method using high performance liquid chromatography method with fluorescence detection for the simultaneous determination of aflatoxins and ochratoxin A in herbal and fruit tea. The theoretical part describes the most common groups of mycotoxins and the most important methods for their determination in food. The great attention is devoted to HPLC method and the overview of the derivatization techniques for aflatoxins B1 and G1 fluorescence response enhancement. The practical part of this study focuses on the optimization of sample extraction and purification, the settings of the instrumental analysis and the photochemical reactor assembly. The thesis also involves the determination of the basic performance characteristics for the successful method validation.

This thesis deals with the passive sampling of polar compounds from water using passive samplers POCIS (Polar Organic Chemical Integrative Sampler). These devices were exposed for 4 weeks in the waste water at the inflow of municipal waste water treatment plant in Brno – Modřice, and also at the outflow of the treated water. After exposition sequestered compounds were released by extraction with a mixture of methanol, toluene and dichlormethane. Extracts were analysed by liquid chromatography/mass spectrometry with electrospray ionization. Further, methylated and trimethylsilylated extracts were analysed by orthogonal comprehensive two-dimensional gas chromatography with mass spectrometric detection. Separated compounds were identified on the base of their retention and mass spectra.

This thesis focused on optimization of separation methods for underivatized theanine enantiomers on two teicoplanin-based chiral stationary phases and separation methods for enantiomers derivatized by 9-fluorenylmethyloxycarbonyl chloride and dansylchloride. Using these optimized conditions the limits of detection and quantification were calculated. Under optimized conditions were also investigated the contents of theanine enantiomers in green tea and in selected dietary supplement containing L-theanine.

The diploma thesis focuses on analysis of fatty alcohols with mass spectrometry. Theoretical part describes distribution, main properties, usage and analytical methods for analysis of fatty alcohols. Experimental work focuses on the search for a proper derivatisation procedure for fatty alcohols, which would enable their detection with soft ionisation techniques of mass spectrometry, as well as their structural analysis. The main aim of the structural analysis is to find the methyl branching in the fragmentation spectra of derivatives of the fatty alcohols.