[pt] Neste trabalho, polímeros de impressão molecular (MIP) foram desenvolvidos para aplicação em procedimentos de extração em fase sólida (SPE) visando separar e concentrar quercetina (flavonóide), piraclostrobina (pesticida) e kanamicina (antibiótico) em amostras contendo substâncias interferentes na determinação dos analitos-alvo. A seletividade em relação aos analitos de interesse foi conseguida pela interação específica dessas espécies químicas com os sítios de reconhecimento dos polímeros. A produção desses MIPs foi baseada na polimerização em bulk e, de modo a se comprovar a efetividade dos mesmos, o desempenho destes foram comparados com polímeros não-impressos (NIP) correspondentes, que foram produzidos sem o uso da molécula-molde. Os procedimentos para síntese são simples e o material produzido é quimicamente resistente nas condições de uso. A caracterização do material produzido foi feita por meio de microscopia de varredura eletrônica (MEV) e espectrometria de absorção na região do infravermelho (IV). O MIP preparado com quercetina foi empregado na extração seletiva deste flavonóide em amostras de urina e de suplemento alimentar e permitiu a obtenção de limite de detecção de 4,58 x 10-8 mol L-1 usando espectrofotometria de absorção no UV-vis com recuperações superiores a 90 porcento e separação efetiva de outros flavonóides como a flavona e naringenina. O MIP preparado com piraclostrobina foi usado na análise de amostra de água de rio e urina e permitiu o alcance de limite de detecção de 4,6 x 10-9 mol L-1, usando detecção por espectrofluorimetria, e recuperações maiores que 90 porcento na presença de outros pesticidas da classe das estrobilurinas. Para o MIP preparado com kanamicina, o limite de detecção alcançado usando detecção com sonda de nanopartículas fluorescentes foi 9 ng mL-1 (1,5 x 10-8 mol L-1) com aplicação na análise de urina e vacina contra febre amarela com recuperações maiores a 90 porcento. / [en] In this work, molecularly imprinted polymers (MIPs) have been developed for application procedures for solid phase extraction (SPE) in order to separate and concentrate quercetin (flavonoid), pyraclostrobin (pesticide) and kanamycin (antibiotic) in samples containing interfering substances in the determination of target analytes. The selectivity in respect of analytes has been achieved by specific binding of these chemical species with the recognition sites of the polymers. The production of MIP polymerization was based on "bulk" and in order to prove the effectiveness thereof, the performance of these polymers were compared with non-printed (NIP) thereof, that were produced without the use of template molecule. The procedures for synthesis are simple and relatively low cost and is chemically resistant material produced under the conditions of use. The characterization of the material produced was done by scanning electron microscopy (MEV), absorption spectroscopy in the infrared region and CHN elemental analysis. The MIP prepared with quercetin was used for the selective extraction of flavonoids in urine samples and dietary supplement and allowed to obtain a detection limit of 4,58 x 10-8 mol L-1 using absorption spectrophotometry in the UV-vis with recoveries exceeding 90 percent and effective separation of other flavonoids such as naringenin and flavone. The MIP prepared with pyraclostrobin was used to analyze samples of urine and river water and allowed to reach the detection limit of 4,6 x 10-9 mol L-1, detection using spectrofluorimetry and recoveries greater than 90 percent in presence of other pesticides from the class of strobilurins. For the MIP prepared with kanamycin, the detection limit using detection with fluorescent nanoparticles probe was 9 ng mL-1 (1,5 x 10-8 mol L-1) application to the analysis of urine and yellow fever vacine with recoveries greater than 90 percent.

Se han desarrollado métodos basados en análisis por inyección en flujo multijeringa (MSFIA) para determinar automáticamente sulfuro en muestras ambientales y aguas residuales, con detección espectrofotométrica previa transformación química del analito (acoplamiento oxidativo del sulfuro con N-N-dimetil-p-fenilendiamina en presencia de Fe(III) para formar azul de metileno).- Sistema MSFIA con difusión gaseosa para aislar el analito de matrices complejas sin ningún pretratamiento manual de la muestra.- Sistema MSFIA con pre-concentración en fase sólida (optrodo de reflectancia difusa sobre disco de membrana) para determinar niveles traza. - Método MSFIA que combina separación y pre-concentración en línea del sulfuro, para ofrecer elevadas selectividad y sensibilidad.- Sistema MSFIA inteligente totalmente automatizado para determinar sulfuro en un amplio rango de concentraciones, capaz de auto-ajustarse sin ninguna intervención del analista.- Estudio comparativo de metodologías en flujo para determinar sulfuro en matrices acuosas. Se discuten críticamente distintas técnicas y sistemas de detección empleados. / Multisyringe flow injection analysis (MSFIA) systems for automatic determination of sulfide in environmental and wastewater samples have been developed exploiting spectrophotometric detection prior derivatization of the analyte (oxidative coupling of sulfide with N-N-dimethyl-p-phenylenediamine in presence of Fe(III) to form methylene blue).- A MSFIA system was coupled to gas-diffusion separation to achieve the isolation from complex matrices without any pre-treatment batch sample.- An optrode using disk-based solid-phase extraction in a MSFIA set-up is proposed to determine trace levels.- A hyphenated method that combines the separation and pre-concentration is presented reaching high selectivity and sensitivity. - A fully automated smart MSFIA system for sulfide monitoring in a wide concentration range is proposed choosing by itself the best approach to quantify the analyte without the analyst intervention.- A review of flow-based methodologies for sulfide determination in aqueous matrices is presented. The characteristics of automated protocols using several detection systems are discussed.

Die Aufgabenstellung der vorliegenden Arbeit ergab sich aus der Notwendigkeit organische Schwefelverbindungen in stark belasteten wäßrigen Proben zu identifizieren, um das toxische Potential dieser Wässer abschätzen zu können. Bei der chromatographischen Trennung und Identifizierung der einzelnen Komponenten traten insbesondere dann Probleme auf, wenn die einzelnen Komponenten in Konzentrationsbereichen auftraten die um Größenordnungen differierten. Da auch durch selektive Detektion unbekannte Komponenten nicht direkt identifiziert werden können, wurde angestrebt durch geeignete Probenvorbereitungsschritte einerseits die gesuchten Zielsubstanzen anzureichern und andererseits störende Matrixbestandteile abzutrennen. Ziel der vorliegenden Arbeit war es effektive und möglichst selektive Verfahren zu entwickeln, um organische Schwefelverbindungen aus wäßrigen Proben anzureichern. Im Rahmen dieser Arbeit wurden zwei Möglichkeiten erarbeitet. Für die Anreicherung von leicht- bis mittelflüchtigen Substanzen erwies sich die Adsorption an Ag2S aus der Gasphase als geeignet. Zur Extraktion mittel- bis schwerflüchtiger Verbindungen wurde eine Festphasenextraktionstechnik an einem mit Blei(II)ionen modifizierten Kationenaustauschermaterial entwickelt. Ein Vergleich beider Techniken erfolgte mit dem Verfahren der Festphasenmikroextraktion (SPME). Die adsorptive Anreicherung an Ag2S wurde mit einem Membranextraktionsschritt (ME) , Thermodesorption (TD) und GC/MS gekoppelt. Wie die SPME kann sie für den Nachweis leicht- bis mittelflüchtiger Verbindungen aus flüssigen, festen und gasförmigen Proben eingesetzt werden. Gegenüber der Festphasenmikroextraktion ermöglicht sie den Einsatz größerer Probemengen, was in niedrigeren Nachweisgrenzen (oberer bis mittlerer ng/l-Bereich) resultiert. Nachteile der entwickelten Technik bilden der höhere experimentelle Aufwand und die längeren Analysenzeiten. Das Festphasenextraktionsverfahren an dem mit Pb(II)ionen beladenen Kationenaustauschermaterial erlaubt gegenüber der SPME ebenfalls die Anwendung größerer Probenmengen und höherer Konzentrationen. Beide Verfahren zeigen vergleichbare Nachweisgrenzen (unterer µg/l - bis oberer ng/l-Bereich) für die verwendeten Modellsubstanzen. Sowohl durch die adsorptive Anreicherung an Silbersulfid als auch durch die Festphasenextraktion an Pb(II)-modifizierten Ionenaustauschmaterialien wird die Identifizierung unbekannter organischer Schwefelverbindungen in stark belasteten Proben erheblich erleichtert. Beide Methoden bilden als einfache und leistungsfähige Techniken wirkungsvolle Ergänzungen zu bereits etablierten Anreicherungsverfahren. Neben der Identifizierung und Analyse können die Techniken ebenfalls zur Entfernung von schwefelhaltigen Substanzen aus verschiedenen Matrizes dienen. Anwendungsmöglichkeiten der entwickelten Methoden bestehen neben der Umweltanalytik auch in der Lebensmittelchemie.

Submitted by Ana Hilda Fonseca (anahilda@ufba.br) on 2014-09-17T17:51:04Z No. of bitstreams: 1 (Dissertação_mestrado__(03.11_)).pdf: 1756144 bytes, checksum: 6a5c8a2f486f154eba91b5aa9780dde0 (MD5) / Approved for entry into archive by Fatima Cleômenis Botelho Maria (botelho@ufba.br) on 2014-09-19T12:24:04Z (GMT) No. of bitstreams: 1 (Dissertação_mestrado__(03.11_)).pdf: 1756144 bytes, checksum: 6a5c8a2f486f154eba91b5aa9780dde0 (MD5) / Made available in DSpace on 2014-09-19T12:24:04Z (GMT). No. of bitstreams: 1 (Dissertação_mestrado__(03.11_)).pdf: 1756144 bytes, checksum: 6a5c8a2f486f154eba91b5aa9780dde0 (MD5) / CNPQ / A determinação de metais em águas naturais por Espectrometria de Emissão Óptica com Plasma Indutivamente Acoplado (ICP OES), em geral, deve envolver etapas de pré- concentração, devido os níveis de concentração dos metais em água serem muitas vezes abaixo dos limites de detecção da técnica. Dessa forma, o objetivo desse trabalho foi desenvolver um método de pré-concentração em fase sólida sensível e simples para a determinação de cobre, ferro, níquel, manganês, chumbo e zinco em amostras de água por ICP OES. As condições experimentais para pré-concentração foram otimizadas considerando os fatores: concentração do eluente, volume de eluente, volume da amostra, vazão e pH. As condições otimizadas corresponderam ao uso de 5 mL de solução de HCl 0,8 mol L-1 como eluente, volume da amostra de 50 mL, vazão de 1,9 mL min-1 e pH 9,0. Para a determinação dos metais, as amostras, após ajuste do pH, foram percoladas com o auxílio de uma bomba peristáltica em cartucho Sep-pak C18 com 1-(2-piridilazo)-2-naftol (PAN) imobilizado na sílica funcionalizada C18. A eluição dos analitos foi realizada com a passagem da solução de HCl pelo cartucho para posterior determinação por ICP OES. Nestas condições, os limites de detecção para Cu, Fe, Mn, Ni, Pb e Zn obtidos variaram entre 0,11 e 21 µg L-1 e os limites de quantificação entre 0,36-69 µg L-1, respectivamente. Os desvios padrão relativos (R.S.D.) para a determinação dos analitos não ultrapassaram 9 %. O efeito de possíveis espécies interferentes sobre a recuperação dos analitos também foram investigados. A exatidão do método foi avaliada mediante análise de material de referência certificado (SLEW-3 Estuariene Water Reference Material for Trace Metals) e o método proposto foi aplicado para análise de amostras de água subterrânea, água potável e água de rio coletadas nos municípios de Cachoeira e São Félix, Bahia, Brasil. As amostras foram analisadas por Espectrometria de Massas com Plasma Indutivamente Acoplado (ICP-MS) como método comparativo, não sendo observadas diferenças significativas entre estes resultados e os resultados obtidos pelo método proposto. / The determination of metals in natural waters by Inductively Coupled Plasma Optical Emission Spectrometry (ICP OES), in general, involves pre-concentration steps, due the concentration levels of metals in water are often below the limits of detection of this technique. In this way, the objective of this work was to develop a pre-concentration solid phase method for sensitive and simple determination of copper, iron, nickel, manganese, lead and zinc in water samples by ICP OES. The experimental conditions for pre- concentration were optimized considering the following factors: the concentration of the eluent, eluent volume, sample volume, flow-rate and pH. The optimized conditions corresponded to the use of 5 mL of HCl solution 0.8 mol L-1 as an eluent sample volume 50 ml, flow-rate 1.9 mL min-1 and pH 9.0. For the determination of metals by the procedure, the samples after pH adjustment were percolated with the aid of a peristaltic pump on Sep-Pak C18 cartridge with 1-(2-pyridylazo)-2-naphthol (PAN). The analyte elution was carried out with the passage of the HCl solution in the cartridge for subsequent determination by ICP OES. In these conditions, the detection limit for Cu, Fe, Mn, Ni, Pb and Zn varied between 0.11 to 21 μg L-1 and the limits of quantification from 0.36 to 69 μg L-1, respectively. The relative standard deviation (RSD) for the analytes of this study did not exceed 9%. The effect of possible interfering species on the recovery of the analytes was also investigated. The accuracy of the method was evaluated by analysis of certified reference material (SLEW-3 Estuariene Water Reference Material for Trace Metals) and the proposed method was applied to samples of ground water, drinking water and river water collected in Cachoeira and São Félix cities, Bahia, Brazil. The samples were analyzed by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) as a comparative method do not being found significant differences among these results and the results obtained by the proposed method.

Die Aufgabenstellung der vorliegenden Arbeit ergab sich aus der Notwendigkeit organische Schwefelverbindungen in stark belasteten wäßrigen Proben zu identifizieren, um das toxische Potential dieser Wässer abschätzen zu können. Bei der chromatographischen Trennung und Identifizierung der einzelnen Komponenten traten insbesondere dann Probleme auf, wenn die einzelnen Komponenten in Konzentrationsbereichen auftraten die um Größenordnungen differierten. Da auch durch selektive Detektion unbekannte Komponenten nicht direkt identifiziert werden können, wurde angestrebt durch geeignete Probenvorbereitungsschritte einerseits die gesuchten Zielsubstanzen anzureichern und andererseits störende Matrixbestandteile abzutrennen. Ziel der vorliegenden Arbeit war es effektive und möglichst selektive Verfahren zu entwickeln, um organische Schwefelverbindungen aus wäßrigen Proben anzureichern. Im Rahmen dieser Arbeit wurden zwei Möglichkeiten erarbeitet. Für die Anreicherung von leicht- bis mittelflüchtigen Substanzen erwies sich die Adsorption an Ag2S aus der Gasphase als geeignet. Zur Extraktion mittel- bis schwerflüchtiger Verbindungen wurde eine Festphasenextraktionstechnik an einem mit Blei(II)ionen modifizierten Kationenaustauschermaterial entwickelt. Ein Vergleich beider Techniken erfolgte mit dem Verfahren der Festphasenmikroextraktion (SPME). Die adsorptive Anreicherung an Ag2S wurde mit einem Membranextraktionsschritt (ME) , Thermodesorption (TD) und GC/MS gekoppelt. Wie die SPME kann sie für den Nachweis leicht- bis mittelflüchtiger Verbindungen aus flüssigen, festen und gasförmigen Proben eingesetzt werden. Gegenüber der Festphasenmikroextraktion ermöglicht sie den Einsatz größerer Probemengen, was in niedrigeren Nachweisgrenzen (oberer bis mittlerer ng/l-Bereich) resultiert. Nachteile der entwickelten Technik bilden der höhere experimentelle Aufwand und die längeren Analysenzeiten. Das Festphasenextraktionsverfahren an dem mit Pb(II)ionen beladenen Kationenaustauschermaterial erlaubt gegenüber der SPME ebenfalls die Anwendung größerer Probenmengen und höherer Konzentrationen. Beide Verfahren zeigen vergleichbare Nachweisgrenzen (unterer µg/l - bis oberer ng/l-Bereich) für die verwendeten Modellsubstanzen. Sowohl durch die adsorptive Anreicherung an Silbersulfid als auch durch die Festphasenextraktion an Pb(II)-modifizierten Ionenaustauschmaterialien wird die Identifizierung unbekannter organischer Schwefelverbindungen in stark belasteten Proben erheblich erleichtert. Beide Methoden bilden als einfache und leistungsfähige Techniken wirkungsvolle Ergänzungen zu bereits etablierten Anreicherungsverfahren. Neben der Identifizierung und Analyse können die Techniken ebenfalls zur Entfernung von schwefelhaltigen Substanzen aus verschiedenen Matrizes dienen. Anwendungsmöglichkeiten der entwickelten Methoden bestehen neben der Umweltanalytik auch in der Lebensmittelchemie.

Apresenta-se a comparação de duas metodologias para pré-concentração e determinação de Cd em amostras de águas estuarinas e água de mar. As metodologias comparadas foram a extração líquido-líquido no ponto nuvem e a extração por retenção dos analitos em fase sólida. Em ambos os métodos houve necessidade de formação de complexos de Cd previamente à extração. Dois agentes complexantes foram testados o O-O dietilditiofosfato (DDTP), e 1-(2- tiazolilazo)-2-naftol (TAN). A extração no ponto nuvem foi realizada utilizando o surfactante não iônico octil fenoxipolietoxietanol (Triton X-114) que à temperatura de \'40 graus\'C forma uma fase micelar viscosa de volume reduzido. Após separação das micelas, estas foram dissolvidas em solução de ácido nítrico facilitando a análise por ICP OES. A pré-concentração em fase sólida por retenção dos íons complexados foi realizada colocando-se uma coluna de Amberlite XAD 4 em sistema em fluxo. Foram ajustadas as vazões de carregamento da coluna e eluição de forma a aumentar os fatores de pré-concentração. Os volumes das soluções finais foram de 2 mL de forma a obter resultados em triplicata na análise por ICP OES. A quantificação foi efetuada pelo método das adições de padrão. Os dois procedimentos foram comparados considerando as eficiências relativas de pré-concentração, a facilidade para efetuar os procedimentos e a percentagem de recuperação das medidas / Two methodologies for determination of Cd in estuarine and seawater samples were studied. The methodologies for pre-concentration of analytes by surfactant-mediated cloud point extraction and solid phase retention in a resin column were compared. In both cases the prior formation of Cd ions complexes with O,O-diethyldithiophosphate (DDTP) and 1-(2-thiazolylazo)-2-naphtol (TAN) was carried out. The Cloud point extraction was attained with the non ionic surfactant ctylphenoxypolyethoxyethanol (Triton X-114) at \'40 graus\'C . The reduced volume of surfactant micelles was easily separated and dissolved using the solution containing nitric acid which facilitate the analysis by ICP OES. The solid phase pre-concentration was performed by retention of the complexed ions in the Amberlite XAD 4 resin column in a flow scheme. The flow-rates for sample loading and elution were adjusted to increase the preconcentration factor. The final volumes collected of 2 mL were further analyzed in triplicate by ICP OES. Quantification was performed by the standard addition method. Both pre-concentration procedures were compared considering the relative efficiencies, the complexity of procedures and recovery percent of results

Apresenta-se a comparação de duas metodologias para pré-concentração e determinação de Cd em amostras de águas estuarinas e água de mar. As metodologias comparadas foram a extração líquido-líquido no ponto nuvem e a extração por retenção dos analitos em fase sólida. Em ambos os métodos houve necessidade de formação de complexos de Cd previamente à extração. Dois agentes complexantes foram testados o O-O dietilditiofosfato (DDTP), e 1-(2- tiazolilazo)-2-naftol (TAN). A extração no ponto nuvem foi realizada utilizando o surfactante não iônico octil fenoxipolietoxietanol (Triton X-114) que à temperatura de \'40 graus\'C forma uma fase micelar viscosa de volume reduzido. Após separação das micelas, estas foram dissolvidas em solução de ácido nítrico facilitando a análise por ICP OES. A pré-concentração em fase sólida por retenção dos íons complexados foi realizada colocando-se uma coluna de Amberlite XAD 4 em sistema em fluxo. Foram ajustadas as vazões de carregamento da coluna e eluição de forma a aumentar os fatores de pré-concentração. Os volumes das soluções finais foram de 2 mL de forma a obter resultados em triplicata na análise por ICP OES. A quantificação foi efetuada pelo método das adições de padrão. Os dois procedimentos foram comparados considerando as eficiências relativas de pré-concentração, a facilidade para efetuar os procedimentos e a percentagem de recuperação das medidas / Two methodologies for determination of Cd in estuarine and seawater samples were studied. The methodologies for pre-concentration of analytes by surfactant-mediated cloud point extraction and solid phase retention in a resin column were compared. In both cases the prior formation of Cd ions complexes with O,O-diethyldithiophosphate (DDTP) and 1-(2-thiazolylazo)-2-naphtol (TAN) was carried out. The Cloud point extraction was attained with the non ionic surfactant ctylphenoxypolyethoxyethanol (Triton X-114) at \'40 graus\'C . The reduced volume of surfactant micelles was easily separated and dissolved using the solution containing nitric acid which facilitate the analysis by ICP OES. The solid phase pre-concentration was performed by retention of the complexed ions in the Amberlite XAD 4 resin column in a flow scheme. The flow-rates for sample loading and elution were adjusted to increase the preconcentration factor. The final volumes collected of 2 mL were further analyzed in triplicate by ICP OES. Quantification was performed by the standard addition method. Both pre-concentration procedures were compared considering the relative efficiencies, the complexity of procedures and recovery percent of results

In this thesis, improved techniques for biomolecule analysis using capillary electrophoresis (CE) and matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) and hyphenations between those have been presented.A pre-concentration method which is possible to apply in both techniques, has also been investigated. In this work the off-line MS mode has been used either in the form of fractionation (Paper I) or by incorporating the MALDI target in the CE separation system (Paper II).In Paper I, a protocol for CE-MALDI analysis of cyanogen bromide digested bacteriorhodopsin (BR) peptides as model integral membrane protein peptides were established. Also, an improved protocol for partially automated manufacturing of a concentration MALDI-target plate is presented. The design of the targets was suitable for the fractions from the CE. A novel technique for the integration of CE to MALDI-MS using a closed-open-closed system is presented in Paper II, where the open part is a micro canal functioning as a MALDI target window. A protein separation was obtained and detected with MALDI-MS analysis in the micro canal. A method has been developed for detection of monosaccharides originating from hydrolysis of a single wood fiber performed in a micro channel, with an incorporated electromigration pre-concentration step preceding CE analysis in Paper III. The pre-concentration showed to be highly complex due to the fact that several parameters are included that affecting each other. In Paper IV a protocol using enzymatic digestion, MALDI-TOF-MS and CE with laser induced fluorescence (LIF) detection for the investigation of the degree of substitution of fluorescein isothiocyanate (FITC) to bovine serum albumin (BSA), as a contact allergen model system for protein-hapten binding in the skin, is presented. The intention of a further CE-MALDI hyphenation has been considered during the work. In Paper V 2,6-dihydroxyacetophenone (DHAP) was investigated, showing promising MALDI-MS matrix properties for hydrophobic proteins and peptides. 2,5-dihydroxybenzoic acid (DHB) was undoubtedly the better matrix for the hydrophilic proteins, but its performance for the larger and hydrophobic peptides was not optimal. Consequently, DHAP can be used as a compliment matrix for improved analysis of hydrophobic analytes. / QC 20101214