Neste trabalho foi desenvolvida, otimizada, e validada, uma metodologia para análise de 15 pesticidas organoclorados em sedimento através de HS/SPME-GC/MS. Esta metodologia foi aplicada para análise de amostras reais visando à realização de um diagnóstico da contaminação por pesticidas organoclorados do Rio Atibaia, principalmente em sua região próxima a contaminação oriunda da empresa Shell Brasil S.A. unidade de Paulínia. Para realização deste diagnóstico, também foi desenvolvida uma metodologia padronizada para diagnóstico ambiental de áreas contaminadas, embasada principalmente em conceitos de Gestão de Qualidade e Meio Ambiente abordadas nas normas ISO 17025, ISO 14001 e GLP. Finalmente, aplicando a metodologia analítica validada, e a metodologia para realização de um diagnóstico ambiental, foi realizado o diagnóstico da presença de pesticidas organoclorados no Rio Atibaia, indicando a existência de diferentes pesticidas organoclorados em sedimento, com destaque para os compostos DDE e BHC que se encontraram em valores acima dos recomendados pela legislação canadense. Os resultados encontrados também demonstraram que as metodologias desenvolvidas foram adequadas para análise de uma contaminação real. / In this work a methodology was developed, optimized and validated for the analysis of 15 organochlorine pesticides in sediment through HS/SPME-GC/MS. This methodology was applied to the analysis of real samples for diagnosis of the organochlorine pesticides contamination of the Atibaia River, mainly in its region close the contamination from company Shell Brasil S.A. site of Paulínia. For accomplishment of this diagnosis, also a standard methodology for environmental diagnosis of contaminated areas was developed, mainly based in concepts of Quality Management and Environmental Management find in standards like ISO 17025, ISO 14001 and GLP. Finally, applying the validated analytical methodology and the methodology proposed for environmental diagnosis, the diagnosis of the organoclhorine pesticide presence in the Atibaia River was performed, indicating different organoclorine pesticides in sediment, mainly DDE and yBHC, which were found in values above of those recommended by the Canadian legislation. The results also demonstrated that the developed methodologies are adequate for analysis of a real contamination.

A poluição dos corpos d\'água é de grande preocupação mundial, pois a maioria da população utiliza a água doce de reservatórios, represas ou rios como principal fonte de água potável. A presença no Brasil de florações de cianobactérias capazes de produzir anatoxina-a, revela a necessidade de métodos simples e rápidos que permitam sua detecção e monitoramento. Neste trabalho foram desenvolvidos, otimizados e validados dois métodos analíticos por GC/MS para identificação e quantificação de anatoxina-a em amostras de água e florações, respectivamente. A norcocaína foi usada como padrão interno em ambos os métodos. Os íons escolhidos para serem monitorados foram (íons quantificadores sublinhados): anatoxina-a: 191,164, 293 e norcocaína: 195, 136, 168. As curvas de calibração dos métodos mostraram-se lineares nas faixas de 2.5-200 ng.mL-1 e 13-250 ng.mg-1. Os limites de detecção obtidos foram 2 ng.mL-1 e 10 ng.mg-1. Os métodos demonstraram sensibilidade e especificidade adequada para seu uso no monitoramento ambiental da anatoxina-a. / The water pollution is a big concern around the world, since the most of cities use freshwater reservoirs, dams or rivers as the main drinking water suppliers. Cyanobacterial blooms capable to produce anatoxin-a are regularly present in Brazilian waters. Therefore, there is a necessity of simple and rapid analytical methods to monitor this cyanotoxin. In the present work, two analytical methods by GC/MS for identification and quantification of anatoxin-a in water and algae bloom samples were developed, optimized and validated. Norcocaine was used as internal standard in both methods. The ions chosen to be monitorated were (quantification ions underlined): anatoxin-a 191, 164, 293 and norcocaine: 195, 136, 168. Both method calibration curves showed linearity in the ranges of: 2.5-200 ng.mL-1 and 13-250 ng.mg-1. The obtained limit of detection were: 2 ng.mL-1 and 10 ng.mg-1. The methods showed sensitivity and specificity enough to be used routinely as a tool for anatoxin-a monitoring.

Esta tese de doutoramento tem o objetivo de identificar os padrões de organização espacial dos principais componentes da paisagem na Baixa Nhecolândia e analisar suas relações com as estruturas regionais e as dinâmicas hídricas. Para tanto, foram utilizados produtos de sensoriamento remoto e técnicas do geoprocessamento de imagens de satélites com bancos de dados georreferenciados dos elementos da área de estudo, além de procedimentos de controle de campo das informações levantadas. A Baixa Nhecolândia sofre influência da dinâmica das estruturas lineares aparentes nas serranias de seu entorno e, a partir de um estudo mais acurado, pode-se evidenciar no sutil relevo do Pantanal registros morfoestruturais correlacionados a esses lineamentos. Por outro lado, feições do relevo associadas a hidrodinâmica do Leque Aluvial do Rio Taquari distribuem-se na região, apresentando padrões de organização diferenciados das formas ligadas às morfoestruturas. Desta forma, os elementos da paisagem da Baixa Nhecolândia apresentam distribuição, padrão de organização espacial e formas resultantes, dentre outros, de 2 processos dinâmicos que são objetos desta tese: 1) processos estruturais regionais, 2) processos hidrodinâmicos do leque aluvial do rio Taquari. / The goal of this thesis is to identify the spatial organization patterns of the main landscape elements in the Baixa Nhecolândia and to analyze its relations with the regional structures and the hydrological dynamic. The products of remote sensing and geoprocessing techniques applied on the elements of the study area had been used, beyond procedures of information field control. The Baixa Nhecolândia suffers influences from the linear structures dynamics from its around and can be evidenced in the subtle relief of the Pantanal correlated to morphostructures evidences. Otherwise, reliefs forms associates to the hydrodynamics of the Aluvial Fan of the River Taquari, distributed in the region, presenting differentiated patterns of organization of the morfostructures forms. Them, the elements of the landscape of the Baixa Nhecolândia presents distribution resultant patterns of spatial organization and forms, amongst others, of 2 dynamic processes that are objects of this thesis: 1) regional structural processes, 2) hydrodynamic processes of the aluvial fan of the Taquari river.

Synthetic piperazines are chemically-produced compounds that contain a six-member ring with two opposing nitrogen atoms. Several piperazine derivatives, namely 1- benzylpiperazine (BZP), 1-(3-trifluoromethylphenyl)-piperazine (TFMPP), and 1-(3- chlorophenyl)-piperazine (mCPP), have fallen into the “designer drugs” category due to their increasing recreational use as a “legal” alternative to ecstasy (3,4-methylenedioxymethamphetamine). These compounds share similar stimulant and physiological effects with amphetamines which make them desirable to young adults in party-type atmospheres. BZP, a Schedule I drug for its high abuse potential and no accepted medical use, is the only recreationally-abused synthetic piperazine currently federally controlled in the United States. The purpose of this research was to develop and validate a reliable method to identify and quantify eight forensically significant synthetic piperazines in blood and urine using ultra-fast liquid chromatography-electrospray ionization-tandem mass spectrometry (UFLC-ESI-MS/MS). The method was validated according to the Scientific Working Group for Forensic Toxicologists (SWGTOX) guidelines for quantitative analysis for both matrices and includes the following analytes: 1-benzylpiperazine (BZP), 1-(4-fluorobenzyl)-piperazine (FBZP), 4-methyl-1-benzylpiperazine (MBZP), 1-(4-methoxyphenyl)-piperazine (MeOPP), 1-(para-fluorophenyl)-piperazine (pFPP), 1-(3-chlorophenyl)-piperazine (mCPP), 2,3-dichlorophenylpiperazine (DCPP), and 1-(3-trifluoromethylphenyl)-piperazine (TFMPP). All samples were prepared by fortifying 100 µL of certified drug-free whole blood and urine (UTAK Laboratories, Inc., Valencia, CA, U.S.A.) with certified reference standards (Cayman Chemical, Ann Arbor, MI, U.S.A.) of each analyte at desired concentrations and standard additions of 1-benzylpiperazine-d7, 1-(3-chlorophenyl)-piperazine-d8, and 1(-3-trifluoromethylphenyl)-piperazine-d4 internal standards (Cerilliant, Round Rock, TX, U.S.A). After pretreatment with 1 mL phosphate buffer, samples underwent solid phase extraction (SPE) on mixed-mode copolymeric columns (Clean Screen®, UCT Inc., Levittown, PA, U.S.A.). Eluents were evaporated to dryness with low heat (65°C) and nitrogen gas. Samples were reconstituted with a 50:50 mixture of methanol and 2mM ammonium formate buffer with 0.2% formic acid before being analyzed by a UFLC (Shimadzu Corporation, Kyoto, Japan) and 4000 QTRAP ESIMS/MS (SCIEX, Framingham, MA, U.S.A.) system. Analyses were performed with multiple reaction monitoring scans in positive ionization mode using ions and voltages obtained from a manual compound optimization. Analytes were separated on a reversed phase column (Kinetex® F5, Phenomenex®, Torrance, CA, U.S.A.) with a binary gradient consisting of a 2mM ammonium formate buffer with 0.2% formic acid and methanol with 0.1% formic acid. The flow rate was 0.400 mL/min. Analyst™ (SCIEX) software was used for data collection and MultiQuant™ (SCIEX) software was used for quantitation. The total run time was 11.5 minutes with equilibrations. All calibration curves in both matrices exhibited R2 values > 0.99 using a weighting factor of 1/x. A linear dynamic range of 20-2000 ng/mL was used for all analytes in both matrices, except for BZP in urine which ranged from 50-2000 ng/mL. In blood, the limit of quantitation was 10 ng/mL for mCPP and TFMPP and 20 ng/mL for BZP, FBZP, MBZP, MeOPP, pFPP and DCPP. In urine, the limit of quantitation was 10 ng/mL for MeOPP, mCPP, TFMPP and DCPP, 20 ng/mL for FBZP, MBZP and pFPP and 50 ng/mL for BZP. When a 200 ng/mL concentration was evaluated, the SPE procedure showed percent recoveries ranging from 80-95% for blood; except for BZP, FBZP, and MeOPP which had recoveries of 60%, 60%, and 105%, respectively. Percent recoveries ranged from 82-94% for urine; except for BZP and FBZP which had recoveries of 66% and 68%, respectively. Bias and precision were assessed at concentrations of 50, 200, and 700 ng/mL. All samples were calculated within ±20% bias and within ±20% coefficient of variation. The highest concentration evaluated that did not produce carryover in subsequent matrix blanks was 5000 ng/mL. Ionization was suppressed for all analytes in both matrices by 45-95%. Matrix effects were present but were determined to be insignificant. Of the drugs evaluated, caffeine, dibenzylpiperazine, and 1-(4-chlorophenyl)-piperazine (pCPP) produced chromatographic peaks in the method; however, pCPP was the only substance that affected quantitation of an analyte. It increased the peak area of mCPP by almost 50% when present at the same concentration which suggests this method is unable to differentiate between isomeric pairs. This is a sensitive, reliable, and robust method with a wide linear dynamic range to account for the presence of these analytes in both blood and urine. This research will provide for the identification and quantitation of these substances in forensic casework.

Objetivo: Avaliar a farmacocinética da vildagliptina em animais sadios e diabéticos, através da análise dos níveis plasmáticos totais e livres teciduais, empregando-se a técnica de microdiálise. Metodologia: A doença foi induzida nos animais através da administração de 42mg/kg de aloxano através da via intravenosa (i.v.). A vildagliptina foi administrada nas doses de 50 mg/kg (n = 6) e 75 mg/kg (n = 6) via i.v. nos animais diabéticos e na dose de 50 mg/kg (n = 6) nos animais sadios. As concentrações plasmáticas foram quantificadas por CLAE-EM-EM em método desenvolvido e validado. A ligação às proteínas plasmáticas foi determinada por microdiálise, assim como a avaliação tecidual. As sondas de microdiálise foram calibradas in vitro através de diálise e retrodiálise e in vivo utilizando retrodiálise. Para determinação das concentrações teciduais, uma segunda metodologia foi desenvolvida e validada em CLAE-EM-EM. Avaliações compartimentais (software Scientist ®) e não compartimentais (software Excel ®) foram realizadas. Resultados e Discussão: A ligação as proteínas plasmáticas apresentou um valor médio de 9,44 % ± 3,23, condizente com valores encontrados na literatura. Os valores de Ke, clearance, tempo de meia vida, MRT e VDss não apresentaram diferença estatística significativa entre as diferentes doses administradas nos animais diabéticos e entre os animais sadios. As calibrações in vitro por diálise e retrodiálise apresentaram uma recuperação média de 30%, sem diferença estatística entre as duas metodologias empregadas (α = 0,05). A recuperação in vivo também apresentou o mesmo valor médio de recuperação. A penetração tecidual do fármaco em animais diabéticos para as diferentes doses estudadas apresentou mesmo valor nos tecidos estudados, uma média de 0,20. A penetração tecidual semelhante no animal diabético pode ser devido ao dano similar entre os órgãos sofrido durante a indução da doença. Já os animais sadios apresentaram penetração tecidual similar no músculo sem diferença estatística significativa em relação aos diabéticos, entretanto no fígado foi observada uma penetração quarenta e quatro vezes inferior a observada no músculo. Essa disparidade pode ser atribuída a diferença de expressão de proteínas transportadoras no fígado do animal diabetico quando comparado ao sadio. O perfil farmacocinético plasmático foi semelhante entre os dois grupos avaliados, sendo que os parâmetros não diferiram estatisticamente (α = 0,05). Foi empregado o modelo de dois compartimentos para prever as concentrações teciduais. A previsão supõe concentrações superiores as encontradas experimentalmente, contradizendo dados de literatura. Esses dados são inéditos na literatura e demostram a importância da determinação do fármaco em tecidos alvo, uma vez que nem sempre modelos matemáticos conseguem prever a realidade fisiológica. Conclusões: As metodologias analíticas para quantificação da vildagliptina em microdialisado e plasma foram desenvolvidas e validadas, seguindo os requisitos do FDA. O perfil farmacocinético plasmático foi adequadamente descrito pelo modelo de 2 compartimentos. Os perfis teciduais obtidos nesse trabalho podem contribuir para o melhor entendimento dos mecanismos farmacológicos envolvidos e contribuir para futura otimização de terapias. / Objective: To evaluate the pharmacokinetics of vildagliptin in healthy and diabetic animals using a microdialysis technique. Methodology: Diabetes was induced in animals by administration of 42 mg/kg of alloxan intravenously (iv). Vildagliptin was administered intravenously as 50 mg/kg (n = 6) and 75 mg/kg doses (n = 6) in the diabetic animals and as a 50 mg/kg dose (n = 6) in healthy animals. Plasma concentrations were quantified by a HPLC-MS-MS method developed and validated. The plasma protein binding was determined by microdialysis and tissue evaluation. Microdialysis probes were calibrated in vitro using dialysis and retrodialysis and in vivo using retrodialysis. A second method was developed and validated using HPLC-MS-MS to determine tissue concentrations. Results and Discussion: Mean plasma protein was 9.44% ± 3.23, consistent with values reported in the literature. The values of Ke, clearance, half-life, MRT and Vdss showed no statistical difference between the evaluated doses in diabetic animals and between healthy animals (α = 0.05). Calibrations in vitro by dialysis and retrodialysis showed mean recovery of 30%, with no statistical difference between the two methodologies. Mean recovery in vivo also showed the same value. The tissue penetration of the drug in diabetic animals for the different doses studied showed the same value in both tissues studied, an mean of 0.20. The tissue penetration similar in diabetic animals could be due to the similar damage suffered between organs during induction of the disease. The healthy animals showed similar muscle penetration, compared with diabetics animals, although the liver showed a penetration forty four times lower than muscle. This discrepancy can be attributed to differential expression of transporter proteins in the liver of diabetic animals, when compared to the healthy group. The plasma pharmacokinetic profile was similar between the investigated groups, and the parameters did not differ. The two-compartment model was employed to describe the data and used to predict the concentration in the tissues. This is the first study to present these tissue profiles, which presented concentrations below the estimated by the model. These data demonstrate the importance of determining the drug inside the target tissue, as the mathematical models sometimes cannot predict physiology. Conclusions: The analytical methods for the quantification of vildagliptin in microdialysate and plasma were developed and validated by following the requirements of the FDA. The plasma pharmacokinetic profile was correctly described by the model of two compartmental models. The novel tissue profiles obtained in this study may contribute to a better understanding of the pharmacological mechanisms involved and contribute to optimization of future therapies.

Metabólitos secundários produzidos por Streptomyces com atividade antibiótica apresentam relevante importância biotecnológica para as indústrias farmacêuticas e agroquímicas. Dentre estes metabólitos, podemos destacar a nigericina, um antibiótico poliéter usado como aditivo em ração animal atuando como promotor de crescimento e no tratamento de algumas doenças, como a malária, em carcinoma nasofaríngeo, a vaccínia, entre outras. Neste trabalho foram avaliadas duas cepas de actinobactérias potenciais produtoras de nigericina, a EUCAL 26 e a EUCAL 74. As duas actinobatérias foram fermentadas em cinco meios de cultivo diferentes (BD, Czapek, ISP2, M29 e TSB). A cepa EUCAL 26 foi a mais promissora na produção de nigericina em meio Czapeck. A partir da EUCAL 26, foi feito um estudo da máxima produção de nigericina em meio Czapek variando o pH do meio, temperatura de fermentação, e período de fermentação. As melhores condições encontradas foram em pH 7,0 a 25 °C por 27 dias. Foi realizado também um estudo de otimização de aumento de escala de fermentação, de um volume de meio Czapeck de 50 mL, para um volume de 4 L. Também foram avaliados dois resíduos agroindustriais (Farmal e Melaço de Soja) para a produção de nigericina. O meio de Melaço de Soja aumento a produção em aproximadamente 300x quando comparado com o meio Czapeck padrão. Os efeitos dos nutrientes do meio Czapeck também foram avaliados. A retida do K2HPO4 do meio produziu um aumento de 50x na produção de nigericina, quando comparado com o meio Czapeck controle. Também foi avaliado o efeito da adição de -butirolactonas sintéticas, moléculas de sinalização hormonal, para a produção de nigericina. Das 15 -butirolactonas testadas, a DP21A foi a mais eficiente, pois além de aumentar a produção de nigericina em 23x, também diminui o período máximo de sua produção. Todas as analises realizadas neste trabalho para o monitoramento da produção de nigericina, foram feitas empregando a espectrometria de massas sequencial acoplada à cromatografia liquida de ultra eficiência. / Secondary metabolites produced by Streptomyces with antibiotic activity have significant biotechnological importance for the pharmaceutical and agrochemical industries. Among them, nigericin stands out as an antibiotic polyether used as growth promoter in animal feed and for treatment of some diseases such as malaria, nasopharyngeal carcinoma, and vaccinia. In this study, two actinobacteria strains considered potential producers of nigericin named EUCAL 26 and 74 were tested. The two actinobacteria were fermented in five different culture media (BD, Czapek, ISP2, M29 and TSB). EUCAL 26 strain was the most promising in producing nigericin in amid Czapeck media. For EUCAL 26, a study of maximum production of nigericin in Czapek medium at varying the pH, fermentation temperature and fermentation period have been performed. As a result, the best conditions were pH 7.0, at 25 °C for 27 days. In addition, an optimization study for scale-up fermentation have been done, where a volume of 50 mL Czapeck medium have been expanded to 4 L, in order to obtain the highest production of nigericin. Two agroindustrial residues (FARMAL and Honey Soy) have also been evaluated for nigericin production. The honey soy medium increased nigericin production in the rate of 300 when compared with standard Czapeck medium. The effects of nutrients from Czapeck medium have also been evaluated. Removal of K2HPO4 from culture medium resulted in an increase of 50 times when compared with the control Czapeck medium. The effect of adding synthetics -butyrolactones (hormone signaling molecules) for the production of nigericin have also been evaluated. From 15 tested -butyrolactones, DP21A was the most efficient. In addition to increase nigericin yield in 23x it also reduced the period for its maximum production. All analyzes performed in this study to monitor the nigericin production were performed using tandem mass spectrometry coupled to ultra high performance liquid chromatography.

L’utilisation massive des antibiotiques contribue à leur accumulation dans les boues des stations d’épurations. L’application directe des boues est parmi les sources de dissémination des antibiotiques et des gènes de résistance aux antibiotiques. Le compostage et la méthanisation sont parmi les bioprocédés de traitement des boues qui permettent d’éliminer ou réduire les teneurs de certains antibiotiques. Dans ce travail, une boue primaire de la STEP de Marrakech a été contaminée par trois familles d’antibiotiques (macrolides, tétracyclines, fluoroquinolones) pour conduire 4 essais de compostage à différentes doses (dont un essai témoin) et un essai deméthanisation en mode semi-continu. Les résultats du compostage ont montré que l’augmentation des concentrations d’antibiotiques retarde la dégradation de la matière organique et affecte le ratioC/N. De même, la phase thermophile est perturbée, retardée et réduite dans le temps. Pour la méthanisation, une concentration unique et réaliste a été testée. Dans ces conditions, aucun effet sur la production du biogaz ou sur la dégradation de la matière organique n’a été observé. Afin de suivre la dissipation des trois familles d’antibiotiques utilisées au cours du compostage et de la méthanisation, une approche analytique basée sur l’extraction accélérée par solvant (ASE) suivie par l’application d’une méthode des ajouts dosés avant quantification par chromatographie liquide couplée à de la spectrométrie de masse en tandem (UPLC-MS/MS) a du être mise en point. Le compostage et la méthanisation permettent de réduire significativement les concentrations des molécules parents appartenant à la famille des macrolides et des tétracyclines. Par contre,l’élimination des fluoroquinolones est non-significative et ne dépasse pas 30%. Au cours du compostage, la dissipation des macrolides se fait en phase de stabilisation tandis que la phase de maturation est impliquée dans la dissipation des tétracyclines. Les concentrations encirprofloxacine (fluoroquinolone) semblent légèrement évoluer au cours du procédé probablement en raison d’une adsorption/désorption sur le co-substrat lignocellulosique utilisé. Concernant la méthanisation, l’élimination des macrolides et des tétracyclines est significative durant la stabilisation du procédé mais n’atteinds pas les rendements observés lors du compostage. Ladiminution des concentrations des molécules parents est probablement accompagnée par une biotransformation des antibiotiques sous forme de métabolites qui à ce stade ne sont pas connus.La question de la rémanence de certaines molécules comme les fluoroquinolones, interpelle quand au risque d’antibiorésistance. Ainsi, la valorisation des composts/digestats comme amendements organiques des sols dois à terme conduire à une réflexion concernant la réglementation qui inclus la présence de molécule de la classe des antibiotiques.

Mass Spectrometry (MS) and Ion Mobility - Mass Spectrometry (IM-MS) can be used to investigate protein structure and dynamics and are ideally positioned to study intrinsically disordered and amyloidogenic proteins, whose diverse conformational space and/or oligomeric state is hard to track accurately. This thesis uses hybrid MS approaches including IM-MS, Cross-linking IM-MS and ECD-FT-ICR MS to probe the structure of alpha-Synuclein and Amyloid-beta (Abeta). For alpha-Synuclein, the effect of solution pH and ionisation polarity on the species observed by MS and IM-MS is investigated. Conformational families observed by Cross-linking IM-MS provides a link between the solution and gas phase structures of alpha-Synuclein observed here and our data correlates with that reported by other groups. MS, IM-MS and HDX-MS are used to probe alpha-Synuclein during the early stages of aggregation. A specific aggregation competent conformer is not observed suggesting that the solution constituents remain conformationally dynamic. We observe shifts in the species observed by MS and IM-MS between samples and our data contributes to an array of conflicting structural studies indicating that alpha-Synuclein adopts a diverse range of species with significant variation. For Abeta(1-42) and Abeta(1-40) Collision Induced Unfolding and ETD/ETcaD demonstrate that Abeta(1-42) adopts a compact conformation bound by intramolecular interactions. Changes to the Abeta(1-42) and Abeta(1-40) ATDs following SID are correlated to known structure influencing intermolecular interactions and demonstrate the large structural difference between Abeta(1-42) and Abeta(1-40) despite differing by only two C-terminal amino acids. IM-MS is used to classify the mode of action of anti-aggregation drug candidates on Abeta(1-42). The anti-aggregation capacity of the retro-inverso peptide, RI-OR2 is shown to result from inducing the compaction or extension of Abeta(1-42), preventing the adoption of an aggregation competent structure. In contrast, the flavonoid Rutin is shown to act solely through inducing Abeta(1-42) compaction. This thesis demonstrates the power of MS based methods to investigate the diverse range of structures of intrinsically disordered aggregating proteins implicated in disease.

Neste estudo determinou-se o nonilfenol - um composto xenobiótico utilizados na fabricação de antioxidantes, aditivos para óleos lubrificantes e produção de surfactantes etoxilados - e o bisfenol A - um composto químico utilizado na produção de diversos produtos, como, plásticos policarbonatos, resinas epóxi e retardantes de chama. Ambos exibem atividade estrogênica em concentrações abaixo de 1ng L-1 - em cinco pontos de coleta na cidade de Americana, SP. Devido à ampla utilização, esses compostos estão presentes em esgoto industrial em quantidades substanciais. A primeira etapa deste estudo foi a validação dos método analítico seguindo o Método 8270D da United States Environmental Protection Agency (USEPA) que descreve o procedimento de validação para os compostos semivoláteis analisados por cromatografia gasosa acoplada a espectrometria de massas (GC/MS). Os limites de detecção e de quantificação do método para a determinação do nonilfenol foram, respectivamente, 1 e 5 µg L-1. O coeficiente de determinação (R2) e a recuperação média para o nonilfenol foram, respectivamente, 0,99 e 99,04%. Os limites de detecção e de quantificação do método para o bisfenol A foram, respetivamente, 1 e 5 µg L-1. O coeficiente de determinação (R2) e a recuperação média para o bisfenol A foram, respectivamente, 0,99 e 91,55%. Na segunda etapa desta pesquisa analisaram-se amostras de água coletadas na cidade de Americana para a determinação do nonilfenol e do bisfenol A. Como método de extração dos compostos, usou-se a extração em fase sólida (SPE) e como técnica analítica a cromatografia gasosa acoplada a espectrometria de massas (GC/MS). Os resultados indicaram a presença desses compostos em alguns pontos de amostragem sendo que a concentração do nonilfenol a variou de 5,028 a 10,524 µg L-1 e para o bisfenol A a concentração variou de 5,645 a 295,255 µg L-1. Foram feitos também testes ecotoxicólogicos utilizando como organismo teste o microcrustáceo Daphnia similis, porém o nonilfenol e o bisfenol A, na concentração testada que variou de 0,05 a 500 µg L-1 não apresentaram toxicidade aguda. / In this study it was determined nonylphenol - a xenobiotic compound used in the manufacture of antioxidants, additives for lubricating oils and production of ethoxylated surfactant - and bisphenol A - a chemical compound used in production of various products, such as polycarbonate plastic, epoxy resins and flame retardants. Both exhibit estrogenic activity at concentrations lower than 1 ng L-1 - in five sampling points in the city of Americana, SP. Due to the widespread use of these compounds, they are present in substantial quantities in industrial sewage. The first step of this study was to validate the analytical method following the Method 8270D of the United States Environmental Protection Agency (USEPA) that describes the validation procedure for semi-volatile compounds analized by gas chromatography-mass spectrometry (GC/MS). The limits of detection and quantification for determining the nonylphenol were, respectively, 1 and 5 µg L-1. The coefficient ofdetermination (R2) and recovery for nonylphenol were respectively 0.99 and 99.04%. The limits of detection and quantification for the bisphenol A were, respectively, 1 and 5 µg L-1. The coefficient of determination (R2) and recovery for bisphenol A were respectively 0.99 and 91.55%. In the second step of this study were analized in water samples of Americana city for the determination of nonylphenol and bisphenol A. As a method of extraction of the compounds it was used solid-phase extraction (SPE) and as analytical technique GC/MS. The results indicated the presence of such compounds in some sampling points and the concentration of nonylphenol ranged from 5.028 to 10.524 µg L-1 and bisphenol A concentration ranged from 5.645 to 295.255 µg L-1. Ecotoxicological tests were also performed using as test organism microcrustaceans Daphnia similis, but nonylphenol and bisphenol A, tested at concentration ranging from 0,05 to 500 µg L-1 showed no acute toxicity.

Malgré une sensible baisse des ventes depuis quelques années, la France reste le premier utilisateur européen de phytosanitaires. Ceci n’est pas sans conséquences sur l’environnement et les organismes non visés par ces produits. De nombreuses études ont permis d’une part de mieux comprendre les phénomènes de transfert des pesticides dans les compartiments eau-air sols et d’autre part de mettre en évidence leur contamination. Les données concernant l’eau sont plus nombreuses du fait de son statut de réceptacle ultime des pollutions.et des normes établies par les textes réglementaires. La Directive Cadre européenne sur l’Eau (DCE) adoptée en 2000 vise à atteindre le bon état chimique et écologique des eaux d’ici 2015 avec une obligation de résultats. Le bon état chimique dépend du respect des normes de qualité environnementales (NQE) établies notamment pour 24 pesticides. En 2012, 5% des points de mesure en cours d’eau et 19% des points de mesure en eaux souterraines ne respectent pas ces NQE.Le Languedoc Roussillon est depuis de nombreuses années la première région viticole de France et souffre d’une pression phytosanitaire importante. En 2014, un rapport de l’Agence Régionale de Santé (ARS) fait même état de l’utilisation encore récente de pesticides interdits (simazine, terbuthylazine…) depuis le début des années 2000. L’ARS a de plus affirmé que la plupart des cas de non-conformité avérés pouvaient être rattachés à la présence d’activité agricole dans le bassin versant du captage.Remédier à ces contaminations passe par le suivi de la qualité des eaux et la caractérisation de la pollution. Les outils actuels s’avèrent ne pas être toujours représentatifs de cette pollution ;de nouveaux outils sont donc mis au point. L’étude présentée ici porte sur l’utilisation de l’échantillonnage passif de type POCIS (Polar Organic Chemical Integrative Sampler) pour le suivi de la contamination en pesticides de trois bassins versants languedociens : Paulhan,Puissalicon, Roujan.Dans un premier temps, l’étude des limites possibles du dispositif POCIS a été faite pour évaluer son applicabilité dans les conditions d’exposition imposées par le terrain. Le comportement des molécules au sein de l’outil a été étudié pour une meilleure compréhension des phénomènes de diffusion qui s’y produisent. Ces études ont mené à la mise en place de calibrations en laboratoire en conditions contrôlées. [...] La mise en place des dispositifs POCIS sur les trois sites d’études a mis en évidence la prédominance de pesticides utilisés en viticulture et a montré tout l’intérêt de l’outil d’un point de vue qualitatif. La comparaison de l’outil avec l’échantillonnage automatisé montre des disparités entre concentration réelle et concentration estimée via le POCIS. Ce dernier reste à améliorer d’un point de vue quantitatif pour obtenir un échantillonneur fiable et robuste pour ce type de milieu d’exposition. / Despite a noticeable sales decrease since few years, France remains the first european user of phytosanitary products. This is not without consequences on the environment and the untargeted organisms. Several studies allowed on one hand to better understand phenomena of pesticides transfer in water-air and soils compartments and on the other hand to highlight their contamination. Data concerning the water are more numerous cause of its status of ultimate receptacle of pollution and standards established by regulatory texts. The european Water Framework Directive (WFD) adopted in 2000 aims to achieve good chemical and ecological status of water bodies by 2015 with an obligation of results. The good chemical status depends on respect for the Environmental Quality Standards (EQS) established, for instance, for 24 pesticides. In 2012, 5% of river measurement points and 19%of groundwater measurement points do not respect these EQS. For many years, Languedoc Roussillon is the first wine-producing region of France and suffers from a significant pesticide pressure. In 2014, a report from the Regional Health Agency (RHA) has mentioned a relatively recent use of banned pesticides since the early 2000’s like simazin and terbutylazin. Moreover, RHA stated that most of recognized cases of non-compliance could be linked to the presence of agricultural activity in the catchment basin.[...] The establishment of POCIS devices on the three study sites showed the predominance ofpesticides used in viticulture and showed all the interest of the tool on a qualitative point ofview. The comparison of the tool with the automated sampling shows disparities between real concentration and concentration estimated with the POCIS. However, it remains to improvethe tool on a quantitative way to obtain a reliable and robust sampler for this type of exposure medium.