Orientador: Maria Del Pilar Taboada Sotomayor / Banca: Flavio Santos Damos / Banca: Arnaldo Cesar Pereira / Resumo: Eletrodos quimicamente modificados à base de pasta de carbono foram construídos para determinação de captopril, empregando catalisadores biomiméticos em potencial da enzima P450. O complexo que permitiu obter os resultados mais satisfatórios foi o bis(piridil)ftalocianinaferro (II) [Fe(dipy)Pc]. O sensor proposto foi avaliado em batelada e em sistema de análise por injeção em fluxo (FIA), e as condições de análise foram devidamente otimizadas empregando a amperometria. Em batelada obtiveram-se os melhores resultados aplicando potencial de 0,175 V vs Ag|AgCl(KClsat) e usando solução tampão TRIS 0,1 mol L-1 como eletrólito em pH 8,0. Nestas condições observou-se uma faixa linear de resposta para o sensor entre 25 e 165 μmol L-1; com uma sensibilidade de 2.315(±61) μA L mol-1 e um limite de detecção e quantificação de 0,9 e 3,1 μmol L-1, respectivamente. Em sistema em fluxo, os melhores resultados foram obtidos aplicando potencial de 0,100 V vs Ag|AgCl (KClsat) em solução carregadora de tampão TRIS 0,1 mol L-1 e pH 8.0, com volume de amostra injetado de 75 μL e vazão de 1,4 mL min-1. A faixa linear de resposta obtida no sistema em fluxo (FIA) foi entre 5,0 x 10-5 e 2,5 x 10-2 mol L-1, com uma sensibilidade de 210(±1) μA L mol-1 e uma reprodutibilidade menor que 4%, avaliado pelo valor do desvio padrão médio relativo (R.S.D.) para sete injeções consecutivas de solução padrão de captopril 2,5 x 10-3 mol L-1. A biomimeticidade do sensor foi estudada, assim como a seletividade e seus possíveis interferentes. O sensor biomimético foi aplicado na análise de formulações comerciais, amostras biológicas e ambientais (águas de rios e esgoto). Os resultados obtidos quando comparados com o método oficial de análise baseado na cromatografia líquida de alta eficiência (HPLC), mostraram boa eficiência do sensor desenvolvido na quantificação do captopril / Abstract: Chemically modified electrodes based on carbon paste were constructed for determination of captopril, catalysts employing biomimetic of the P450 enzyme. The compound that allowed to obtain the most satisfying results was the iron (II) phthalocyanine bis(pyridine) complex [Fe(dipy)Pc]. The proposed sensor was evaluated in batch system and flow injection analysis (FIA) and the analysis conditions were properly optimized employing amperometry. In batch experiments were obtaining the best results by applying a potential of 0,175 V vs Ag | AgCl (KClsat) and using 0.1 mol L-1 TRIS buffer at pH 8.0, the electrolyte. These conditions was observed under the linear response range for the sensor between 25 and 165 μmol L-1, with a sensitivity of 2315 (± 61) μA L mol-1 and quantification and detection limits of 0.9 and 3.1 μmol L-1, respectively . In the flow system the best results were obtained at the potential of 0,1 V vs. Ag | AgCl (KClsat) in carrier solution of 0.1 mol L-1 TRIS buffer at pH 8.0, with the injected sample volume of 75 mL and flow rate of 1.4 mL min-1. The linear response range obtained in the FIA system was between 5.0 x 10-5 and 2.5 x 10-2 mol L-1, with a sensitivity of 210 (± 1) μA L mol L-1 and the reproducibility value lower than 4 %, calculated the relative standard deviation (RSD) for seven consecutive injections of standard solution of captopril 2.5 x 10-3 mol L-1. The biomimetic characteristics of the sensor was studied and the selectivity and possible interfering. The biomimetic sensor has been applied to analysis of commercial formulations, biological and environmental samples (river water and sewage). The results compared with the official method of analysis based on high performance liquid chromatography (HPLC) showed the good efficiency of this sensor is developed quantification of captopril / Mestre

Made available in DSpace on 2014-06-11T19:29:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-08-27Bitstream added on 2014-06-13T19:17:22Z : No. of bitstreams: 1 wong_a_me_araiq.pdf: 4048206 bytes, checksum: cdc95008c64d5a92757cafbd9ad5ad6c (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Eletrodos quimicamente modificados à base de pasta de carbono foram construídos para determinação de captopril, empregando catalisadores biomiméticos em potencial da enzima P450. O complexo que permitiu obter os resultados mais satisfatórios foi o bis(piridil)ftalocianinaferro (II) [Fe(dipy)Pc]. O sensor proposto foi avaliado em batelada e em sistema de análise por injeção em fluxo (FIA), e as condições de análise foram devidamente otimizadas empregando a amperometria. Em batelada obtiveram-se os melhores resultados aplicando potencial de 0,175 V vs Ag|AgCl(KClsat) e usando solução tampão TRIS 0,1 mol L-1 como eletrólito em pH 8,0. Nestas condições observou-se uma faixa linear de resposta para o sensor entre 25 e 165 μmol L-1; com uma sensibilidade de 2.315(±61) μA L mol-1 e um limite de detecção e quantificação de 0,9 e 3,1 μmol L-1, respectivamente. Em sistema em fluxo, os melhores resultados foram obtidos aplicando potencial de 0,100 V vs Ag|AgCl (KClsat) em solução carregadora de tampão TRIS 0,1 mol L-1 e pH 8.0, com volume de amostra injetado de 75 μL e vazão de 1,4 mL min-1. A faixa linear de resposta obtida no sistema em fluxo (FIA) foi entre 5,0 x 10-5 e 2,5 x 10-2 mol L-1, com uma sensibilidade de 210(±1) μA L mol-1 e uma reprodutibilidade menor que 4%, avaliado pelo valor do desvio padrão médio relativo (R.S.D.) para sete injeções consecutivas de solução padrão de captopril 2,5 x 10-3 mol L-1. A biomimeticidade do sensor foi estudada, assim como a seletividade e seus possíveis interferentes. O sensor biomimético foi aplicado na análise de formulações comerciais, amostras biológicas e ambientais (águas de rios e esgoto). Os resultados obtidos quando comparados com o método oficial de análise baseado na cromatografia líquida de alta eficiência (HPLC), mostraram boa eficiência do sensor desenvolvido na quantificação do captopril / Chemically modified electrodes based on carbon paste were constructed for determination of captopril, catalysts employing biomimetic of the P450 enzyme. The compound that allowed to obtain the most satisfying results was the iron (II) phthalocyanine bis(pyridine) complex [Fe(dipy)Pc]. The proposed sensor was evaluated in batch system and flow injection analysis (FIA) and the analysis conditions were properly optimized employing amperometry. In batch experiments were obtaining the best results by applying a potential of 0,175 V vs Ag | AgCl (KClsat) and using 0.1 mol L-1 TRIS buffer at pH 8.0, the electrolyte. These conditions was observed under the linear response range for the sensor between 25 and 165 μmol L-1, with a sensitivity of 2315 (± 61) μA L mol-1 and quantification and detection limits of 0.9 and 3.1 μmol L-1, respectively . In the flow system the best results were obtained at the potential of 0,1 V vs. Ag | AgCl (KClsat) in carrier solution of 0.1 mol L-1 TRIS buffer at pH 8.0, with the injected sample volume of 75 mL and flow rate of 1.4 mL min-1. The linear response range obtained in the FIA system was between 5.0 x 10-5 and 2.5 x 10-2 mol L-1, with a sensitivity of 210 (± 1) μA L mol L-1 and the reproducibility value lower than 4 %, calculated the relative standard deviation (RSD) for seven consecutive injections of standard solution of captopril 2.5 x 10-3 mol L-1. The biomimetic characteristics of the sensor was studied and the selectivity and possible interfering. The biomimetic sensor has been applied to analysis of commercial formulations, biological and environmental samples (river water and sewage). The results compared with the official method of analysis based on high performance liquid chromatography (HPLC) showed the good efficiency of this sensor is developed quantification of captopril

Orientadora: Maria del Pilar Taboada Sotomayor / Banca: Denis Ricardo Martins de Godoi / Banca: Éder Tadeu Gomes Cavalheiro / Resumo: Este trabalho propõe a síntese de MIP usando como molécula molde, ou template, o corante Verde Ácido 16 (AG16), um corante do grupo dos trifenilmetanos. A síntese foi feita pelo método em bulk; diretamente sobre a fibra óptica, usada como transdutor no desenvolvimento do sensor óptico; e também diretamente em placas de vidro, para fins de caracterização morfológica dos materiais impressos. O polímero sintetizado e o sensor construído foram caracterizados por Microscopia Eletrônica de Varredura (MEV), Espectroscopia vibracional da região no Infravermelho (IV), Reflectância Total Atenuada (RTA) e Microscopia Confocal. Os parâmetros otimizados para o desempenho do sensor foram temperatura, tempo de adsorção/pré-concentração e pH, obtendo-se os melhores resultados a 25 ºC, 60 minutos e pH 7,0; respectivamente. Sob estas condições os limites de detecção e quantificação do MIP-sensor foram 62,2 e 188 µmol L-1 respectivamente, com valores de repetibilidade e reprodutibilidade, em termos do desvio padrão relativo (RSD) abaixo de 4%. Nos experimentos realizados usando o MIP-optodo-AG16 na presença de outros quatro corantes diferentes, os valores de obtidos para a retenção/adsorção desses corantes mostraram a excelente seletividade do tip-sensor construído. Na aplicação em amostras de rio e de efluente industrial enriquecidas com AG16, os valores de recuperação obtidos foram próximos a 100% para o MIP-optodo, mostrando a eficiência do sensor em relação ao NIP-optodo. Desta forma, este ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: This work proposes the synthesis of a MIP using Acid Green 16 (AG16) dye as template molecule of the triphenylmethane group. The synthesis was made by bulk method directly on the optical fiber surface which is using as a transducer in the development of optical sensor and also synthesized directly on glass plates for morphological characterization of the printed materials. The synthesized polymer and the constructed sensor were characterized by Scanning Electron Microscopy (SEM), Infrared (IR), Attenuated Total Reflectance (ATR) and Confocal Microscopy. The optimized parameters were temperature, adsorption/pre-concentration time, pH, obtaining the best results at 25 ºC, 60 minutes and pH 7,0 respectively. Under these conditions for the MIP-sensor the limits of detection and quantification were 62.2 and 188 μmol L-1 respectively, with repeatability and reproducibility values in terms of relative standard deviation (RSD) below 4%. The experiments were performed using MIP-optode-AG16 in presence of four other different dyes and the values obtained for the retention/adsorption of those dyes showed the excellent selectivity of the constructed tip-sensor. For application the river samples and industrial effluent were enriched with AG16 and recovery values were obtained close to 100% for the MIP-optodo that showing the efficiency of the sensor in relation to the NIP-optodo. Thus this master's work shows the development of an exceptional optical sensor for the literature with excelle... (Complete abstract click electronic access below) / Mestre

The cochlea is known to exhibit a nonlinear, mechanical amplification which allows the ear to detect faint sounds, improves frequency discrimination, and broadens the range of sound pressure levels that can be detected. In this work, active artificial hair cells (AHC) are proposed and developed which mimic the nonlinear cochlear amplifier. Active AHCs can be used to transduce sound pressures, fluid flow, accelerations, or another form of dynamic input. These nonlinear sensors consist of piezoelectric cantilever beams which utilize various feedback control laws inspired by the living cochlea. A phenomenological control law is first examined which exhibits similar behavior as the living cochlea. Two sets of physiological models are also examined: one set based on outer hair cell somatic motility and the other set inspired by active hair bundle motility. Compared to passive AHCs, simulation and experimental results for active AHCs show an amplified response due to small stimuli, a sharpened resonance peak, and a compressive nonlinearity between response amplitude and input level. These bio-inspired devices could lead to new sensors with lower thresholds of sound or vibration detection, improved frequency sensitivities, and the ability to detect a wider range of input levels. These bio-inspired, active sensors lay the foundation for a new generation of sensors for acoustic, fluid flow, or vibration sensing. / Ph. D.

Cette thèse décrit le développement de capteurs à fibre optique hautement sélectifs, utilisant des polymères à empreintes moléculaires (MIPs, de l’anglais molecularly imprinted polymers) comme éléments de reconnaissance, et se basant sur la fluorescence pour la détection. Nous avons étendu l’étude à d’autres types de capteurs et de méthodes de détection optiques, toujours basés sur les MIPs. Les MIPs sont des récepteurs synthétiques biomimétiques possédant des cavités spécifiques pour une molécule cible. Produits par un processus de moulage à l’échelle moléculaire, les MIPs sont capables de reconnaître et de se lier à leurs molécules cibles, avec des spécificités et affinités comparables aux récepteurs naturels. De plus, comparé aux récepteurs biologiques, les MIPs sont plus stables, moins chers et plus faciles à intégrer dans les procédés standard industriels de fabrication. Ainsi, les MIPs apparaissent comme une alternative intéressante aux biomolécules entant qu’éléments de reconnaissance dans les biocapteurs. Dans la première partie de la thèse (Chapitre 2), les MIPs ont été synthétisés en tant que micropointe à l’extrémité d’une fibre optique, par polymérisation in-situ induite par un laser en seulement quelques secondes. Les paramètres photoniques et physico-chimiques ont été optimisés pour moduler les propriétés des micro-objets de polymères. Des nanoparticules d’or ont été incorporées dans la micropointe de MIP afin d’induire une exaltation du signal. Afin de prouver l’efficacité de notre capteur, les études initiales ont été réalisées avec un MIP synthétisé avec le Ncarbobenzyloxy- L-phenylalanine (Z-L-Phe) comme template et le dérivé d’acide aminé fluorescentdansyl-L-phenylalanine, comme analyte. La fluorescence a été collectée de l’extérieur au niveau de la micropointe par une fibre optique connectée à un spectrofluorimètre, ou par collection du signal fluorescent ré-émis dans l’un des bras d’une fibre bifurquée en Y. L’analyse fluorescent a pu être détecté à des concentrations de l’ordre du nM. Afin de quantifier les analytes non fluorescents, un monomère fluorescent, possédant un groupe naphthalimide, a été incorporé dans le MIP ; celui-çidéployant une augmentation de la fluorescence quand l’analyte se lie. Utilisant ce système avec un MIP spécifique pour l’herbicide 2,4-D (acide 2,4-dichlorophénoxyacétique), des concentrations aussi basses que 2,5 nM en 2,4-D ont pu être mesurées. Le capteur MIP a également été appliqué à des analytes d’intérêt pour la sécurité alimentaire et le domaine nanostructurées. Cette étude pose les bases pour le développement futur de nanocapteurs et de Dans la dernière partie (Chapitre 4), une méthode d’analyse novatrice, basée sur l’utilisation des MIPs et l’analyse par polarisation de fluorescence, a été développée en vue de permettre la quantification directe et rapide d’analytes dans des échantillons alimentaires et environnementaux. Cette technique a été appliquée avec succès pour détecter des antibiotiques fluoroquinolones dans l’eau du robinet et le lait, en-dessous de leur limite maximale de résidus. En conclusion, nous pouvons dire que ce travail ouvre la voie vers l'application d'une nouvelle génération de capteurs optiques portables, robustes et miniaturisables basés sur les MIPs, pour des mesures «sur-site » et la quantification en temps réel d’analytes biologiques et environnementaux dans des milieux complexes. / This thesis describes the development of highly selective fiber optic sensors using molecularly imprinted polymers (MIPs) as recognition elements associated with fluorescence for detection. Additionally, we extended the study to the development of other MIP-based optical sensors and sensing methods. MIPs are synthetic biomimetic receptors possessing specific cavities designed for a target molecule. Produced by a templating process at the molecular level, MIPs are capable of recognizingand binding target molecules with selectivities and affinities comparable to those of natural receptors. Compared to biological recognition elements, MIPs are more stable, cheaper and easier to integrate into standard industrial fabrication processes. Hence, MIPs have become interesting alternatives to biomolecules as recognition elements for biosensing. In the first part of this thesis (Chapter 2), MIPs were synthesized by in-situ laser-induced photopolymerization in only a few seconds, as a micrometer-sized tip at the extremity of a telecommunication optical fiber. Photonic and physico-chemical parameters were optimized to tailor the properties of the polymer micro-objects. Gold nanoparticles were incorporated into the MIP microtip for signal enhancement. To prove the efficiency of the sensor, initial studies were performed with a MIP templated with N-carbobenzyloxy-L-phenylalanine (Z-L-Phe) and the fluorescent amino acid derivative dansyl-L-phenylalanine as analyte. The fluorescence was collected either externally at the tip level by an optical fiber connected to a spectrofluorimeter or by collection of the fluorescent signal re-emitted into the fiber through the second arm of a Y-shaped bifurcated fiber. The fluorescent analyte could be detected in the low nM concentrations. In order to monitor nonfluorescent analytes, a naphthalimide-based fluorescent monomer was incorporated into the MIP during its synthesis; fluorescence enhancement was observed when analyte binding occurs. Using this system, the sensor containing a MIP specific for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), could detect and quantify this analyte at concentrations as low as 2.5 nM. The signaling MIP-based sensor was also applied to analytes of interest for food safety and biomedical applications, such as the mycotoxin citrinin and the sphingolipid, D-erythro-sphingosine-1-phosphate. In the second part of the thesis (Chapter 3), a different type of fiber optic sensor: cheap, fast and made for “single-use”, was developed by using 4-cm long disposable polystyrene evanescent wave optical fiber waveguides. The coating of the MIP was either performed ex-situ, by dip-coating the fiber in a suspension of MIP particles synthesized beforehand, or in-situ by evanescent-wave photopolymerization directly on the fiber. The resulting fiber optic sensor could detect 2,4-D in the low nM range and demonstrated specific and selective recognition of the herbicide over its structural analogues and other non-related carboxyl-containing analytes. Additionally, we demonstrated the versatility of the system by applying the evanescent wave fiber optic sensor to detect citrinin, a mycotoxin, by simply coating the waveguide with a MIP specific for citrinin. This type of technology could possibly be extended to detect other carboxyl-containing analytes, as long as a specific MIP for the concerned analyte is available. In parallel, the technique of evanescent-wave photopolymerization was used for the synthesis of signaling MIP microdots on continuous and nanostructured gold films. This study lays the foundations for future development of plasmonic MIP nanosensors and microchips. In the last part of the thesis (Chapter 4), an innovative sensing method, based on the use of MIPs and analysis by fluorescence polarization, was developed in order to allow the fast and directquantification of analytes in food and environmental samples.

This thesis describes the development of highly selective fiber optic sensors using molecularly imprinted polymers (MIPs) as recognition elements associated with fluorescence for detection. Additionally, we extended the study to the development of other MIP-based optical sensors and sensing methods. MIPs are synthetic biomimetic receptors possessing specific cavities designed for a target molecule. Produced by a templating process at the molecular level, MIPs are capable of recognizingand binding target molecules with selectivities and affinities comparable to those of natural receptors. Compared to biological recognition elements, MIPs are more stable, cheaper and easier to integrate into standard industrial fabrication processes. Hence, MIPs have become interesting alternatives to biomolecules as recognition elements for biosensing. In the first part of this thesis (Chapter 2), MIPs were synthesized by in-situ laser-induced photopolymerization in only a few seconds, as a micrometer-sized tip at the extremity of a telecommunication optical fiber. Photonic and physico-chemical parameters were optimized to tailor the properties of the polymer micro-objects. Gold nanoparticles were incorporated into the MIP microtip for signal enhancement. To prove the efficiency of the sensor, initial studies were performed with a MIP templated with N-carbobenzyloxy-L-phenylalanine (Z-L-Phe) and the fluorescent amino acid derivative dansyl-L-phenylalanine as analyte. The fluorescence was collected either externally at the tip level by an optical fiber connected to a spectrofluorimeter or by collection of the fluorescent signal re-emitted into the fiber through the second arm of a Y-shaped bifurcated fiber. The fluorescent analyte could be detected in the low nM concentrations. In order to monitor nonfluorescent analytes, a naphthalimide-based fluorescent monomer was incorporated into the MIP during its synthesis; fluorescence enhancement was observed when analyte binding occurs. Using this system, the sensor containing a MIP specific for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), could detect and quantify this analyte at concentrations as low as 2.5 nM. The signaling MIP-based sensor was also applied to analytes of interest for food safety and biomedical applications, such as the mycotoxin citrinin and the sphingolipid, D-erythro-sphingosine-1-phosphate. In the second part of the thesis (Chapter 3), a different type of fiber optic sensor: cheap, fast and made for "single-use", was developed by using 4-cm long disposable polystyrene evanescent wave optical fiber waveguides. The coating of the MIP was either performed ex-situ, by dip-coating the fiber in a suspension of MIP particles synthesized beforehand, or in-situ by evanescent-wave photopolymerization directly on the fiber. The resulting fiber optic sensor could detect 2,4-D in the low nM range and demonstrated specific and selective recognition of the herbicide over its structural analogues and other non-related carboxyl-containing analytes. Additionally, we demonstrated the versatility of the system by applying the evanescent wave fiber optic sensor to detect citrinin, a mycotoxin, by simply coating the waveguide with a MIP specific for citrinin. This type of technology could possibly be extended to detect other carboxyl-containing analytes, as long as a specific MIP for the concerned analyte is available. In parallel, the technique of evanescent-wave photopolymerization was used for the synthesis of signaling MIP microdots on continuous and nanostructured gold films. This study lays the foundations for future development of plasmonic MIP nanosensors and microchips. In the last part of the thesis (Chapter 4), an innovative sensing method, based on the use of MIPs and analysis by fluorescence polarization, was developed in order to allow the fast and directquantification of analytes in food and environmental samples.

This thesis describes the development of highly selective fiber optic sensors using molecularly imprinted polymers (MIPs) as recognition elements associated with fluorescence for detection. Additionally, we extended the study to the development of other MIP-based optical sensors and sensing methods. MIPs are synthetic biomimetic receptors possessing specific cavities designed for a target molecule. Produced by a templating process at the molecular level, MIPs are capable of recognizingand binding target molecules with selectivities and affinities comparable to those of natural receptors. Compared to biological recognition elements, MIPs are more stable, cheaper and easier to integrate into standard industrial fabrication processes. Hence, MIPs have become interesting alternatives to biomolecules as recognition elements for biosensing. In the first part of this thesis (Chapter 2), MIPs were synthesized by in-situ laser-induced photopolymerization in only a few seconds, as a micrometer-sized tip at the extremity of a telecommunication optical fiber. Photonic and physico-chemical parameters were optimized to tailor the properties of the polymer micro-objects. Gold nanoparticles were incorporated into the MIP microtip for signal enhancement. To prove the efficiency of the sensor, initial studies were performed with a MIP templated with N-carbobenzyloxy-L-phenylalanine (Z-L-Phe) and the fluorescent amino acid derivative dansyl-L-phenylalanine as analyte. The fluorescence was collected either externally at the tip level by an optical fiber connected to a spectrofluorimeter or by collection of the fluorescent signal re-emitted into the fiber through the second arm of a Y-shaped bifurcated fiber. The fluorescent analyte could be detected in the low nM concentrations. In order to monitor nonfluorescent analytes, a naphthalimide-based fluorescent monomer was incorporated into the MIP during its synthesis; fluorescence enhancement was observed when analyte binding occurs. Using this system, the sensor containing a MIP specific for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D), could detect and quantify this analyte at concentrations as low as 2.5 nM. The signaling MIP-based sensor was also applied to analytes of interest for food safety and biomedical applications, such as the mycotoxin citrinin and the sphingolipid, D-erythro-sphingosine-1-phosphate. In the second part of the thesis (Chapter 3), a different type of fiber optic sensor: cheap, fast and made for "single-use", was developed by using 4-cm long disposable polystyrene evanescent wave optical fiber waveguides. The coating of the MIP was either performed ex-situ, by dip-coating the fiber in a suspension of MIP particles synthesized beforehand, or in-situ by evanescent-wave photopolymerization directly on the fiber. The resulting fiber optic sensor could detect 2,4-D in the low nM range and demonstrated specific and selective recognition of the herbicide over its structural analogues and other non-related carboxyl-containing analytes. Additionally, we demonstrated the versatility of the system by applying the evanescent wave fiber optic sensor to detect citrinin, a mycotoxin, by simply coating the waveguide with a MIP specific for citrinin. This type of technology could possibly be extended to detect other carboxyl-containing analytes, as long as a specific MIP for the concerned analyte is available. In parallel, the technique of evanescent-wave photopolymerization was used for the synthesis of signaling MIP microdots on continuous and nanostructured gold films. This study lays the foundations for future development of plasmonic MIP nanosensors and microchips. In the last part of the thesis (Chapter 4), an innovative sensing method, based on the use of MIPs and analysis by fluorescence polarization, was developed in order to allow the fast and directquantification of analytes in food and environmental samples.

Submitted by Fabio Sobreira Campos da Costa (fabio.sobreira@ufpe.br) on 2016-08-31T14:13:50Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Tese doutorado Bárbara V M Silva.pdf: 8451335 bytes, checksum: 8f05c2e3a44caad45cfbe00e5b36c3eb (MD5) / Made available in DSpace on 2016-08-31T14:13:50Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Tese doutorado Bárbara V M Silva.pdf: 8451335 bytes, checksum: 8f05c2e3a44caad45cfbe00e5b36c3eb (MD5) Previous issue date: 2016-02-24 / CAPEs / A presente tese descreve o desenvolvimento de sensores eletroquímicos para imunoensaios empregando a tecnologia de eletrodos impressos com a finalidade de detectar a troponina T cardíaca, o marcador mais específico, atualmente, do infarto agudo do miocárdio. Um dos desafios na confecção de sensores eletroquímicos para imunoensaios é alcançar baixos limites de detecção. Nanomateriais de carbono são, recentemente, considerados excelentes estratégias no preparo de superfícies sensoras devido às suas excelentes propriedades, tais como rápida transferência elétrica e atividade catalítica, aumento da relação superfície/volume e, consequentemente, maior quantidade de biomoléculas imobilizadas. Nesta tese, nanotubos de carbono e grafeno foram utilizados sob diferentes abordagens para modificação de superfícies eletródicas. Um imunossensor baseado em eletrodos serigrafados obtidos pela impressão de filmes de nanotubos de carbono amino funcionalizados incorporados em tinta de carbono foi desenvolvido para detecção “livre de marcação”. Os grupos amino dos nanotubos expostos na interface sensora impressa foram utilizados para imobilização orientada dos anticorpos monoclonais anti-troponina T. Os nanofilmes impressos apresentaram uma excelente estabilidade e reprodutibilidade, exibindo um desvio padrão relativo (DP) menor que ~2% (n = 8), comparado com controle (DP ~9%, n = 8). A resposta analítica do sensor, obtida por voltametria de pulso diferencial, apresentou uma faixa linear entre 0,0025 e 0,5 ng/mL de troponina T (r = 0,995; p<0,0001; n=7), combinado a um baixo erro relativo (<<1%) e limite de detecção de 0,0035 ng/mL. Com o propósito de substituir os anticorpos anti-troponina T, visto que estes constituem parte onerosa do dispositivo, um sensor biomimético foi desenvolvido a partir de uma superfície nanoestruturada de grafeno e polipirrol. A técnica de impressão biomimética em superfície (“surface imprinting”) foi utilizada como estratégia para simplificar e reduzir em uma única etapa a produção das cavidades biomiméticas. Estas foram obtidas através da eletropolimerização do polipirrol e derivados copoliméricos orgânicos mimetizando grupos proteicos amino-reativos. As respostas analíticas do sensor foram geradas por voltametria de pulso diferencial, exibindo uma faixa linear de resposta variando de 0,01 a 0,1 ng/mL de troponina T (r = 0,9953; p<0,0001; n=5) e um limite de detecção de 0,006 ng/mL, mostrando um ótimo desempenho do sensor biomimético. As cavidades biomiméticas apresentaram uma constante de dissociação (KD) de 7,3 10- 13 mol/L, indicando boa afinidade à troponina quando comparadas com o sensor controle (sem troponina T), KD igual a 11,6 10-13 mol/L. Em conclusão, ambas as plataformas sensoras mostram potencial para detecção da troponina T em níveis de importância clínica no diagnóstico do infarto agudo, constituindo testes de pronto atendimento para emergências cardiológicas. / This thesis describes the development of electrochemical sensors for immunoassay by using a screen-printed electrodes technology in order to detect the human cardiac troponin T, the most important marker currently of the acute myocardial infarction. One of the challenges in the manufacturing of electrochemical sensors for immunoassays is to reach low limits of detection. Carbon nanomaterials are recently considered excellent strategies in preparing sensing surfaces due to theirs excellent properties, such as rapid electrical transfer and catalytic activity, increase surface / volume ratio and, consequently, offering higher amount of immobilized biomolecules. In this thesis, carbon nanotubes and graphene were used under different approaches in order to modify the sensors surfaces. An immunosensor based on screen printed electrode obtained by printing of amino functionalized carbon nanotubes films incorporated into carbon ink has been developed for "label-free" detection. The amino groups exposed on the imprinted sensor interface were utilized for oriented immobilization of the monoclonal antibody anti-troponin T. The imprinted nanofilms showed an excellent stability and reproducibility, exhibiting a relative standard deviation (RSD) less than ~2% (n = 8) compared to control (RSD ~9%, n = 8). The analytical response of the sensor, obtained by differential pulse voltammetry, showed a linear range between 0.0025 and 0.5 ng/mL (r = 0.995; p <0.0001, n = 7), combined with a low relative error (<< 1 %) and a calculated limit of detection of 0.0035 ng/mL. In order to replace the anti-troponin T antibody, since these are costly part of the device, a biomimetic sensor was developed from a nanostructured surface of graphene and polypyrrole. The biomimetic technique of surface imprinting was used as a strategy for simplify and reduce in a one-step production of the biomimetic cavities. These were obtained by electropolymerization of the pyrrole and its organic copolymers mimicking amino reactive protein groups. The analytical responses of the sensor were obtained by differential pulse voltammetry, exhibiting a linear range response in 0.01 and 0.1 ng/mL of troponin T (r = 0.9953; p <0.0001, n = 5) and a limit of detection of 0.006 ng/mL, showing a good performance of the biomimetic sensor. The biomimetic sites exhibited a dissociation constant (KD) of 7.3 10-13 mol/L, indicating a good affinity to troponin when compared to its control (without troponin T), KD equal to 11.6 10-13 mol/L. In conclusion, both sensor platform the sensor platforms showed a potential for troponin T detection in levels of clinical important for acute myocardial infarction diagnostic, constituting point-of-care testing for cardiac emergency departments.

Made available in DSpace on 2016-08-19T12:56:34Z (GMT). No. of bitstreams: 1 Jose de Ribamar Martins Neto.pdf: 683212 bytes, checksum: dd95cedfe01600e58259c96b38a5f571 (MD5) Previous issue date: 2010-01-15 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / This work presents an analytical study on determination of 4-aminophenol (4-APh) with a system based on molecurlaly imprinted polymer (MIP) and iron protoporphyrin IX as biomimetic sensor of the peroxidase enzyme. The MIP synthesis was performed by the precipitation method and the analytical determinations by amperometric measurements with a glassy carbon electrode modified with the biomimetic system. The best analytical conditions established for 4-APh determination were: applied potential of -0.1 V vs. Ag/Agl, 0.05 mol L-1 Tris buffer solution, pH 7 and hydrogen peroxide concentration of 100 μmol L-1. Under such optimized conditions, a good linear sensor response was observed in a concentration range from 10 to 90 μmol L-1 (r>0.998), with detection and quantification limits of 3 and 10 μmol L-1, respectively. Experiments performed with tap and river water samples spiked with 4-APh resulted in 93 to 112 % recovery values, suggesting the viability of using the MIP/iron protoporphyrin IX system as biomimetic sensor for determinations of 4-APh in real samples. / O presente trabalho apresenta um estudo analítico da determinação de 4-aminofenol com um sistema a base de um polímero de impressão molecular (MIP, do inglês Molecularly Imprinted Polymer) e protoporfirina IX de ferro como sensor biomimético da enzima peroxidase. A síntese do MIP foi realizada pelo método de precipitação e as determinações analíticas realizadas amperometricamente com um eletrodo de carbono vítreo modificado com o sistema biomimético. As melhores condições analíticas estabelecidas para a determinação de 4-aminofenol foram: potencial aplicado de - 0,1V vs Ag/AgCl, tampão Tris 0,05 mol L-1, pH 7,0 e concentração de peróxido de 100 μmol L-1. Sob tais condições, uma boa resposta linear do sensor foi observada numa faixa de concentração de 10 a 90 μmol L-1 (r>0,9978), com limites de detecção e de quantificação de 3 e de 10 μmol L-1, respectivamente Experimentos realizados com amostras de águas de torneira e de rio fortificadas com 4-aminofenol, resultaram em valores de recuperação entre 93 a 112 %, indicando a viabilidade da aplicação do sistema MIP/proptoporfirina IX de ferro como sensor amperométrico para a detecção de 4-aminofenol em amostras reais.