Orientador: Marco Aurelio Zezzi Arruda / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Quimica / Made available in DSpace on 2018-08-12T19:20:38Z (GMT). No. of bitstreams: 1 Figueiredo_EduardoCostade_D.pdf: 6808404 bytes, checksum: 000ecbe23739fa5d1c64af83061c6da5 (MD5) Previous issue date: 2009 / Resumo: Essa Tese de Doutorado teve como objetivo promover a associação entre polímeros de impressão molecular (MIP) e espectrofotometria, sendo a seletividade conseguida pela ligação específica dos analitos com os sítios de reconhecimento molecular impresso no MIP. No capítulo 1 foi sintetizado e caracterizado um MIP seletivo a catecol, sendo o mesmo empregado na extração de catecol em amostras de guaraná (Paullinia cupana) e mate (Ilex paraguariensis), seguido de determinação espectrofotométrica (reação não específica de redução de Mn(VII) para Mn(II) pelo catecol). Obteve-se um limite de quantificação, um desvio padrão relativo (20 mmol L, n=10) e uma freqüência analítica de 2,7 mmol L, <5% e 15h, respectivamente. A exatidão foi comprovada por comparação dos resultados obtidos pelo método proposto e por HPLC. No Capítulo 2 um MIP foi empregado na extração de nicotina em amostras de urina de fumantes, seguido da quantificação por espectrofotometria (reação de redução do Mn(VII) a Mn(VI)). O limite de quantificação e a freqüência analítica foram de 1,1 mmol L e 11 h, respectivamente. As precisões intra-ensaio (10, 13 e 4%) e inter-ensaio (12, 10 e 5%) foram obtidas empregando-se padrões de 3, 10 e 30 mmol L, respectivamente e a exatidão foi comprovada por meio da técnica de HPLC. No capítulo 3 foi reportado o emprego do MIP na extração de fármacos fenotiazínicos (clorpromazina e perfenazina) em amostras de urina de pacientes. Após a extração, os fármacos foram eluidos e separados, à baixa pressão e alta velocidade, em uma coluna de C18 com tamanho de partícula entre 35 e 50 mm. Os limites de quantificação foram de 5 mmol L, para ambos os fármacos, e a exatidão foi comprovada por adição de analitos e pela técnica de HPLC / Abstract: The objective of this Thesis was the association between molecularly imprinted polymers and spectrophotometry, with the selectivity obtained through specific binding between analytes and imprinted binding sites of the polymer. In the Chapter 1, a selective MIP for catechol was synthesized, characterized, and employed for the extraction of catechol from guarana (Paullinia cupana) and mate (Ilex paraguariensis) samples, followed by indirect spectophotometric determination of catecol by reduction of Mn(VII) for Mn(II). A limit of quantification, a relative standard deviation (20 mmol L, n = 10) and an analytical frequency of 2.7 mmol L, <5% and 15h, were obtained, respectively. Accuracy was validated using HPLC. In the Chapter 2, a MIP for nicotine was used for its extraction in urine samples of smokers, followed by its indirect quantification by spectrophotometry (reduction of Mn(VII) to Mn(VI) by nicotine). The limit of quantification and analytic frequency were of 1.1 mmol L and 11 h, respectively. Intra (10, 13 and 4%) and inter-day (12, 10 and 5%) precisions were obtained using 3, 10 and 30 mmol L nicotine standard solutions, respectively, and accuracy was validated through HPLC. Finally, in the Chapter 3, a MIP was employed for phenothiazinics (chlorpromazine and perphenazine) extraction in urine samples. Soon after, the drugs were eluted and separated using a low pressure and high-speed system, comprising a C18 column with particle size between 35 and 50 mm. The quantification limits were 5 mmol L for chlorpromazine and perphenazine, and the accuracy was validated using HPLC / Doutorado / Quimica Analitica / Doutor em Ciências

Ces travaux de recherches concernent l'association de la technologie des polymères à empreintes moléculaires aux dispositifs acoustiques à onde de Love afin de réaliser un biocapteur dans le cadre d'un projet ANR Tecsan. La première partie de ces travaux de thèse a dressé plus spécifiquement la mise au point d'un protocole de dépôt localisé de polymères imprimés (MIP) et non imprimés (NIP) en films minces compatibles avec la propagation de l'onde élastique. La seconde aprtie des travaux a visé une caractérisation des films et des capteurs ainsi réalisés, par microscopie à balayage et apr mesures de détection sous gaz. Des éléments relatifs aux propriétés mécaniques (porosité, surfaces spécifiques) des MIPs et NIPs, avant extraction de la molécule cible, puis après extraction et après recapture, ont permis de valider le principe du capteur, ouvrant la voie à l'application en milieu liquide. / This work deals with the association of molecularly imprinted polymers technology and Love wave devices in order to develop a biosensor in the frame of ANR project, Tecsan. The first part of this work concerned the development and the validation of a protocol for localized deposition of imprinted (MIP) and non-imprinted (NIP) polymeric thin film compatible with the propagation of acoustic wave. The second part exhibited the cahracterization of the coated sensors, by scanning microscopy and deep gas characterization. The proposed characterization technique gives information on surface morphology and porosity of thin MIP films before extraction of the target molecule, then after extraction and after rebinding. It allowed hence, the validation of the sensor principle. These results constitute a god background for future application in medium liquid.

This dissertation evaluates how gold nanoparticle structure and local environment influence resulting sensor function when using these nanomaterials for complex sample analysis. Molecular imprinted polymers (MIPs), a class of plastic antibodies, are engineered and incorporated into these nanosensors thereby facilitating the quantitative detection of a variety of small molecules when Raman spectroscopy and surface enhanced Raman scattering (SERS) are used for detection. First, homogeneous seeded growth gold nanosphere synthesis is evaluated as a function of ionic double layer composition and thickness. Systematically increasing the citrate concentration during synthesis improves nanomaterial shape homogeneity; however, further elevations of citrate concentration increase the number of internal and/or external atomic defects in the nanomaterials which leads to decreasing solution-phase stability. Next, spherical gold nanoparticles are modified with self-assembled monolayer (SAM), modeled using interfacial energy calculations, and experimental characterized using transmission electron microscopy, NMR, extinction spectroscopy, zeta potential, X-ray photoelectron spectroscopy, and flocculation studies to assess the morphology, surface chemistry, optical properties, surface charge, SAM packing density, and nanoparticle stability, respectively. The number of molecules on the nanostructures increases with increasing ionic strength (by decreasing the electrostatic interfacial energy between assembled molecules) which subsequently promotes nanoparticle stability. Third, plastic antibodies that recognize three drugs commonly used to treat migraines are engineered. These methacrylate-based MIPs are synthesized, extracted, characterized, and used to quantitatively and directly detect over-the-counter drugs in complex samples using Raman microscopy. These results along with numerical approximation methods to estimate drug binding site densities and dissociation constants with the MIPs serve as a foundation for understanding how modest recognition selectivity of MIPs coupled with shifts in the vibrational energy modes from the drugs upon hydrogen binding to the polymer backbone promote sensitive and selective drug detection in complex samples. Finally, nanomaterial incorporation into MIPs for applications in SERS-based biosensors is evaluated. Importantly, gold nanorod concentration increases the detectability of the same drugs using MIPs as pre-concentration and recognition elements. This combination of materials, theory, and applications forms a solid foundation which should aid in the design and development of MIP nanobiosensors for specific and sensitive detection of small molecules in complex matrices.

京都大学 / 新制・課程博士 / 博士(工学) / 甲第23513号 / 工博第4925号 / 新制||工||1769(附属図書館) / 京都大学大学院工学研究科材料化学専攻 / (主査)教授 大塚 浩二, 教授 松原 誠二郎, 教授 秋吉 一成 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DFAM

Le vétérinaire aide le producteur laitier à garder son troupeau en santé. Lorsqu’une vache est malade, il peut prescrire des antibiotiques. Le cas échéant, le lait de la vache traitée aux antibiotiques est jeté. Il n’est donc pas vendu pour consommation. Tout le lait produit est analysé avant d’être pasteurisé et transformé afin de s’assurer que les produits laitiers ne contiennent pas de résidus d’antibiotiques. Si les analyses indiquent que le lait renferme des traces d’antibiotiques, il est jeté et le producteur en cause doit assumer la perte. Les épreuves de dépistage actuelles de ces résidus médicamenteux sont onéreuses et inapplicables sur le terrain. Pour résoudre cette problématique aux pieds de la vache, la solution proposée dans ce projet est la fabrication d’un kit de détection basé sur les polymères à empreinte moléculaire. Il s’agit de polymères dont la conformation moléculaire est complémentaire à celle des antibiotiques. Dans ce projet, il est question d’améliorer l’efficacité des épreuves de dépistages des résidus de tétracyclines en augmentant le nombre de sites d’interaction entre l’antibiotique et des polyesters. Trois polymères sont utilisés portant respectivement des groupements aromatiques, carboxyliques et hydroxyles. Une étude antérieure dans notre laboratoire avait déjà donné un pourcentage de rétention de tétracycline de 50% pour une composition de 1/3 PLAOH- 1/3 PLACOOH- 1/3 PLAOBn. Avec des ajustements, le pourcentage passe à 38.93 % pour une composition de 1/4 PLAOH- 1/2 PLACOOH- 1/4 PLAOPh, de 44.81 % pour une composition de 1/4 PLACOOH- 1/2 PLAOH- 1/4 PLAOPh, de 66.34 %pour une composition de 1/4 PLAOH- 1/4 PLACOOH- 1/2 PLAOPh et de 78.07 % pour une composition de 1/6 PLAOH- 1/6 PLACOOH- 2/3 PLAOPh. Notre hypothèse était que la présence accrue du groupement phényle augmenterait le nombre de sites d’interaction spécifique avec l’antibiotique augmentant ainsi le pourcentage de rétention de l’antibiotique à travers les MIP. Les résultats ont confirmé cette hypothèse. / The veterinarian is helping the dairy farmer to keep his flock healthy. When a cow is sick, he may prescribe antibiotics. But in that case, the cow milk treated with antibiotics is discarded, and not sold for consumption. All milk products are analyzed before being pasteurized and processed to ensure that dairy products do not contain residues of antibiotics. If the analyses show that milk contains traces of antibiotics, it is discarded and the producer must bear the loss. On place, the current screening tests for drug residues are expensive and inapplicable. To resolve this problem at the feet of the cow, the solution proposed in this project is the production of a detection kit based on molecular imprinted polymers. These are polymers whose molecular conformation is complementary to that of antibiotics. This project is about improving the effectiveness of screening tests for residues of tetracyclines by increasing the number of interaction sites between the antibiotic and polyester. Three polymers are used respectively, aromatic, carboxyl and hydroxyl groups. A previous study in our laboratory had already shown a tetracycline retention percentage of 50% for a composition of 1 / 3 PLAOH-1 / 3 PLACOOH-1 / 3 PLAOBn. After variation in the polymers ratio, the percentage obtained is, 38.93 % for a composition of 1 / 4 PLAOH-1 / 2 PLACOOH-1 / 4 PLAOPh, 44.81 % for a composition of 1 / 4 PLACOOH- 1 / 2 PLAOH- 1 / 4 PLAOPh, 66.34 % for a composition of 1/4 PLAOH- 1/4 PLACOOH- 1/4 PLAOPh and 78.07 % for a composition of 1/6 PLAOH- 1 / 6 PLACOOH- 2 / 3 PLAPh. Our hypothesis was that the increase of the amount of the phenyl group, increases the number of specific interaction sites with the antibiotic, and so increases the percentage of retention of the antibiotic through the MIP. The results confirmed this hypothesis.

Development and improvement of analytical techniques are vital in analytical chemistry research. This thesis describes the development and use of tailored surfaces for bioanalytical applications. In sample preparation, solid phase extraction is often used and the development of a protocol for extraction on a molecular imprinted polymer (MISPE) directly from plasma sample is presented. Molecular imprinted polymers (MIP) offer selective sorbents for the imprinted analyte. MISPE has mainly been used in organic phase but in this thesis the development of a protocol for direct extraction of the analyte form an aqueous phase is described. For analysis of complex samples a separation step is often needed. The growing interest in analysis of biological samples and analysis of the human proteome and potential biomarkers has increased the interest in developing new separation techniques. Capillary electrophoresis (CE) has evolved into an important technique for use in analysis of body fluids. In this thesis a novel polyamine coating named PolyE 323 tailored for minimizing the adsorption of basic proteins to the surface is introduced. A straightforward coating protocol, with four simple rinsing steps, was developed. The coating was highly reproducible and useable over a wide pH range. Successful protein separations on PolyE-323-coated capillaries coupled to electrospray ionization mass spectrometry (ESI-MS) were demonstrated. The coated capillaries were also used in studies of protein content of aqueous humor samples from cataract patients as a complement to capillary liquid chromatography. In the studies presented the protein content of aqueous humor samples from two clinical groups was compared. By using capillary liquid separation techniques coupled to matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and MS/MS in combination with isobaric tags for relative and absolute quantitation (iTRAQ) the identity and relative concentrations of proteins in the samples were evaluated. Earlier studies of the proteins in these kinds of samples have mainly been done with techniques using immunological detection where the proteins of interest were chosen in advance. In this thesis it was shown that liquid-based separation techniques are a good complement and by using the mass spectrometry approach presented the protein content of the samples could be evaluated without bias.

Le vétérinaire aide le producteur laitier à garder son troupeau en santé. Lorsqu’une vache est malade, il peut prescrire des antibiotiques. Le cas échéant, le lait de la vache traitée aux antibiotiques est jeté. Il n’est donc pas vendu pour consommation. Tout le lait produit est analysé avant d’être pasteurisé et transformé afin de s’assurer que les produits laitiers ne contiennent pas de résidus d’antibiotiques. Si les analyses indiquent que le lait renferme des traces d’antibiotiques, il est jeté et le producteur en cause doit assumer la perte. Les épreuves de dépistage actuelles de ces résidus médicamenteux sont onéreuses et inapplicables sur le terrain. Pour résoudre cette problématique aux pieds de la vache, la solution proposée dans ce projet est la fabrication d’un kit de détection basé sur les polymères à empreinte moléculaire. Il s’agit de polymères dont la conformation moléculaire est complémentaire à celle des antibiotiques. Dans ce projet, il est question d’améliorer l’efficacité des épreuves de dépistages des résidus de tétracyclines en augmentant le nombre de sites d’interaction entre l’antibiotique et des polyesters. Trois polymères sont utilisés portant respectivement des groupements aromatiques, carboxyliques et hydroxyles. Une étude antérieure dans notre laboratoire avait déjà donné un pourcentage de rétention de tétracycline de 50% pour une composition de 1/3 PLAOH- 1/3 PLACOOH- 1/3 PLAOBn. Avec des ajustements, le pourcentage passe à 38.93 % pour une composition de 1/4 PLAOH- 1/2 PLACOOH- 1/4 PLAOPh, de 44.81 % pour une composition de 1/4 PLACOOH- 1/2 PLAOH- 1/4 PLAOPh, de 66.34 %pour une composition de 1/4 PLAOH- 1/4 PLACOOH- 1/2 PLAOPh et de 78.07 % pour une composition de 1/6 PLAOH- 1/6 PLACOOH- 2/3 PLAOPh. Notre hypothèse était que la présence accrue du groupement phényle augmenterait le nombre de sites d’interaction spécifique avec l’antibiotique augmentant ainsi le pourcentage de rétention de l’antibiotique à travers les MIP. Les résultats ont confirmé cette hypothèse. / The veterinarian is helping the dairy farmer to keep his flock healthy. When a cow is sick, he may prescribe antibiotics. But in that case, the cow milk treated with antibiotics is discarded, and not sold for consumption. All milk products are analyzed before being pasteurized and processed to ensure that dairy products do not contain residues of antibiotics. If the analyses show that milk contains traces of antibiotics, it is discarded and the producer must bear the loss. On place, the current screening tests for drug residues are expensive and inapplicable. To resolve this problem at the feet of the cow, the solution proposed in this project is the production of a detection kit based on molecular imprinted polymers. These are polymers whose molecular conformation is complementary to that of antibiotics. This project is about improving the effectiveness of screening tests for residues of tetracyclines by increasing the number of interaction sites between the antibiotic and polyester. Three polymers are used respectively, aromatic, carboxyl and hydroxyl groups. A previous study in our laboratory had already shown a tetracycline retention percentage of 50% for a composition of 1 / 3 PLAOH-1 / 3 PLACOOH-1 / 3 PLAOBn. After variation in the polymers ratio, the percentage obtained is, 38.93 % for a composition of 1 / 4 PLAOH-1 / 2 PLACOOH-1 / 4 PLAOPh, 44.81 % for a composition of 1 / 4 PLACOOH- 1 / 2 PLAOH- 1 / 4 PLAOPh, 66.34 % for a composition of 1/4 PLAOH- 1/4 PLACOOH- 1/4 PLAOPh and 78.07 % for a composition of 1/6 PLAOH- 1 / 6 PLACOOH- 2 / 3 PLAPh. Our hypothesis was that the increase of the amount of the phenyl group, increases the number of specific interaction sites with the antibiotic, and so increases the percentage of retention of the antibiotic through the MIP. The results confirmed this hypothesis.

Advantages such as chemical, mechanical and thermal stability together with high selectivity for the templated analyte render molecularly imprinted polymers MIPs interesting alternatives to routinely applied separation materials or antibodies. Nevertheless, many factors such as the choice of functional monomer, cross-linker, and porogenic solvent, as well as the ratio between template, functional monomer, and cross-linker will affect the resulting imprinting efficiency and polymer particle size and morphology. The research described in this thesis contributes to the development of new synthetic strategies for the generation of imprinted micro- and nanospheres for 17beta-estradiol (E2) focusing on accurate control and optimization of the governing parameters for precipitation polymerization, including the polymerization temperature and the cross-linker, yielding a one-step synthetic approach with superior control on the bead diameter, shape, monodispersity and imprinting efficiency. Thus synthesized imprinting materials for E2 were successfully applied in HPLC separation, solid phase extraction and radioligand binding assays. As the optimization of imprinted materials is based on fundamental understanding of the binding site properties, the investigations is aimed at establishing a more rational basis for further tailoring imprinted materials to the desired analytical application. The relationships between the particle porosity and rebinding properties were detailed, providing useful guidelines for controlling the particle properties for the desired application including, SPE pre-concentration, HPLC separations, and biomimetic binding assays. Furthermore, analytical techniques (1H-NMR and IR, etc.) and molecular modeling were combined in this thesis to facilitate advanced understanding of the fundamental principles governing selective recognition of molecularly imprinted polymers at a molecular level. The molecular interactions involved in the templating process of molecularly imprinted polymers based on the self-assembly approach were simulated in molecular dynamic simulation model by building a modeling system include all the imprinting components with correct ratio, which has never been reported before. Molecular level interactions such as hydrogen bonding, π-π stacking interactions as well as the free energy governing complex formation of E2 with the functional monomers 4-vinylpyridine (4VP) and methacrylic acid (MAA), and the cross-linker divinylbenzene (DVB) were discussed.

No description available.

Capteur électrochimique basé sur des matériaux carbonés fonctionnalisés, pour la détection de deux micropolluants faisant partie des substances prioritaires de la Directive Cadre européenne sur l’Eau (DCE2000/60/CE) : le plomb et l’anthracène. Les capteurs électrochimiques permettent d’atteindre des limites de détection et une sélectivité adéquates pour l’analyse de micropolluants dont les concentrations sont de l’ordredu μg/L, et sont simples d’utilisation pour des analyses in situ à moindre coût comparés aux appareils d’analyses conventionnels. Leur robustesse est un paramètre important afin de permettre des mesures en continu ou semi-continu dans les eaux. Cette thèse propose tout d’abord le développement du capteur pour la détection du plomb. La formulation d’une encre conductrice de carbone est étudiée pour la sérigraphie de ’électrode réceptrice, permettant ainsi de contrôler la composition de l’encre et d’étudier l’influence de la phase carbonée sur les propriétés électrocatalytiques des électrodes. La fonctionnalisation des électrodes par greffage électrochimique d’un sel de diazonium est également étudiée afin de maîtriser la sensibilité et la reproductibilité des électrodes greffées, en contrôlant l’épaisseur et la qualité des couches. Avec cet objectif, la fonctionnalisation dans un liquide ionique protique qui permet le contrôle de la monocouche en modulant la viscosité de ce milieu a été étudiée. Les électrodes greffées montrent des performances analytiques améliorées notamment en termes de répétabilité et de reproductibilité. Enfin ce travail de thèse porte également sur le développement du capteur pour la détection électrochimique de l’anthracène, molécule sans fonctions chimiques. Les électrodes sont, dans ce cas, fonctionnalisées par un polymère à empreinte moléculaire, matériau connu pour sa très grande sélectivité. Les performances de ce capteur, dont la sélectivité est basée uniquement sur le facteur de forme de la molécule, pour la détection de l’anthracène sont alors mises en évidence. / Electrochemical sensor based on functionalized carbon materials, for the detection of two micropollutants, lead and anthracene, which are among of the priority substances of the European Framework Directive on Water(DCE 2000/60 / EC). Electrochemical sensors allow to achieve detection limits and selectivities for the analysis of micropollutants whose concentrations are of the order of μg/L. They are easy to use for in situ analyzes at lower costs compared to those of the conventional analysis equipment. Their robustness is an important parameter in order to allow continuous or semi-continuous measurements in water. First of all, this thesis proposes the development of a sensor for lead detection. The conductive carbon ink formulation is studied for the screen-printing of the receiving electrode, thus allowing to control the ink composition and to study the influence of the carbon phase on the electrocatalytic properties of electrodes. Functionalization of electrodes by electrochemical grafting of a diazonium salt is also studied in order to control the sensitivity and reproducibility of grafted electrodes, by controlling the thickness and the quality of the layers. With this aim it has been studied the functionalization in a protic ionic liquid in order to allow the control of the monolayer bymodulating the viscosity of this medium. The grafted electrodes show improved analytical performance especially in terms of repeatability and reproducibility. Finally, this work reports the development of a sensor for the electrochemical detection of anthracene, a molecule without chemical functions. In this case, a molecularly imprinted polymer, a material known for its very high selectivity, functionalizes the electrodes.Having a selectivity is only based on the form factor of the molecule, the performance of the sensor developed for the detection of anthracene is also highlighted.