Une mauvaise étanchéité à l’air dans un bâtiment peut entraîner des surconsommations énergétiques et poser un certain nombre de problèmes tels que l’apparition de moisissures dans les murs ou encore une mauvaise qualité de l’air intérieur. Les constructions à ossature bois sont particulièrement sujettes aux infiltrations d’air, d’où la nécessité de mieux comprendre ces phénomènes et leurs conséquences afin que ces bâtiments puissent respecter les normes d’étanchéité de plus en plus strictes. Cette étude contribue par plusieurs aspects et à différentes échelles à l’évaluation de l’impact des infiltrations d’air sur les performances d’un bâtiment.Les infiltrations d’air à travers l’enveloppe peuvent perturber le bon fonctionnement de la ventilation mécanique et augmenter les pertes thermiques. Cette problématique est d’abord traitée numériquement à l’échelle du bâtiment, avec l’étude d’une grande variété de maisons et de conditions météorologiques. Des modèles simplifiés applicables à tout niveau d’étanchéité ont été établis pour la prise en compte des infiltrations naturelles dans les calculs de débit total de ventilation. Une plus petite échelle est ensuite considérée pour l’étude de l’étanchéité à l’air, avec la caractérisation expérimentale de parois ossature bois, de matériaux et de détails de construction, notamment grâce à la construction d’un banc d’essai adapté. Un certain nombre de tests de pressurisation ont permis de quantifier les fuites d’air induites par des défauts d’étanchéité spécifiques et peuvent être utilisés pour les simulations numériques à l’échelle du bâtiment.L’impact des infiltrations d’air sur les performances hygrothermiques d’une paroi est intimement lié à la dispersion de l’air à l’intérieur de celle-ci, mais il y a actuellement un manque d’études et de techniques expérimentales pour la déterminer. Une nouvelle méthode a donc été développée, à savoir l’utilisation de microparticules de fluorescéine comme traceur à l’intérieur des isolants. L’établissement de cartographies de la concentration en fluorescéine a permis d’étudier l’impact de certains paramètres tels que la vitesse d’air, le matériau isolant ou encore la géométrie sur les infiltrations d’air, et a mis en évidence des phénomènes tels que l’apparition de lames d’air entre les composants de la paroi. Par ailleurs un modèle du transport des particules de fluorescéine a été développé et couplé à un modèle CFD pour des analyses plus fines du chemin de l’air.Enfin, une étude de cas a été effectuée sur des parois simplifiées afin de comparer les différentes méthodes expérimentales, de vérifier leur applicabilité à l’étude du chemin de l’air, et d’obtenir des données pour la validation de modèles numériques. La dispersion de l’air en entrée/sortie de l’isolant a été étudiée par thermographie infrarouge et PIV. Le chemin de l’air à l’intérieur de l’isolant a lui été étudié par 3 techniques : des mesures de température avec des thermocouples ; d’humidité relative avec des capteurs capacitifs SHT 75 ; et l’utilisation de microparticules de fluorescéine. Les avantages et inconvénients de chaque méthode ont été identifiés pour aider à sélectionner la plus adaptée pour de futures études. / Poor airtightness in buildings can lead to an over-consumption of energy and to many issues such as moisture damage and poor indoor climate. The timber frame constructions are particularly subject to air leakage and further knowledge in this field is needed to meet the regulation requirements tightened by the development of low-energy and passive houses. This study focuses on the impact of air infiltration on the buildings’ performance, both at the building and the wall assembly scales.The air infiltration through the envelope can disrupt the proper functioning of mechanical ventilation and increase the global energy load. This issue was first investigated numerically at the building scale on a wide range of housing and weather conditions. Simplified models working across the whole airtightness spectrum were established for the inclusion of natural infiltration in buildings’ total ventilation rate calculations. The airtightness was then considered at a smaller scale with the experimental characterization of timber frame wall assemblies, components and construction details, in particular with an original test set-up built for this purpose. A number of pressurization tests enabled to quantify the additional leakage air flow induced by specific airtightness defects and may be of use for building scale numerical simulations.The impact of air infiltration on the hygro-thermal performance of a wall is closely linked to the air dispersion inside it, but there is a lack of experimental studies and methods for the air path investigation. A new technique has therefore been developed, consisting in an innovative use of fluorescein micro-particles as tracer inside the insulation material. It was first applied to specific configurations: straight/angled air channels in contact with porous media. A simple analysis of the fluorescein concentration mappings enabled to investigate the impact of parameters such as the flow velocity, the insulation material and the geometry on the air infiltration in the glass wool, and gave evidences of phenomena such as the appearance of thin air gaps between the components of the wall. A fluorescein transport model was developed and coupled to a CFD model for finer analysis.Finally a case study on simple wall assemblies was carried out to compare experimental techniques, to verify their applicability to the air path study and to provide data for possible numerical model validation. The air dispersion at the inlet/outlet of the insulation was studied with both infrared thermography and the PIV. The air path inside the insulation layer was investigated using three experimental approaches: a temperature monitoring with thermocouples; a relative humidity monitoring with capacitive sensors SHT 75; and the use of fluorescein tracer micro-particles. The respective benefits and limitations of the various methods were identified to help in the selection of the most appropriate one for further studies.

La formulation de médicaments regroupe des technologies innovantes où l'usage de matières premières naturelles émerge du fait de contraintes environnementales grandissantes. Les huiles végétales par exemple sont plébiscitées grâce à leurs propriétés remarquables en terme de solubilisation de principes actifs, de biocompatibilité et de biodégradabilité. Afin d'élargir leurs applications, notamment pour la libération prolongée de principes actifs, la solidification de ces huiles par réticulation a été envisagée à l'aide d'une chimie douce, c'est-à-dire respectueuse des principes de la chimie verte. La silice a été choisie pour réaliser la réticulation par réaction sol-gel d'huiles végétales et ainsi obtenir des matériaux hybrides inorganique/organique d'origine naturelle. L'étude a consisté à fonctionnaliser des huiles végétales à l'aide de précurseurs silylés en utilisant une chimie sans solvant ni catalyseur, pour obtenir des huiles réticulables par voie sol-gel. Deux voies chimiques de fonctionnalisation ont été employées. La première, difficilement contrôlable, été basée sur une réaction époxy-amine où des huiles de lin ou de soja époxydées ont été utilisées. La seconde reposait sur la valorisation d'huile de ricin porteuse d'un groupement hydroxyle via une réaction hydroxy-isocyanate. Les excellents résultats obtenus ont permis de poursuivre l'étude par la formulation de l'huile de ricin silylée grâce à un nouveau procédé d'émulsion huile/eau thermo-stabilisée. Simple et robuste, ce procédé a rendu possible de façon concomitante la mise en forme et la solidification de microparticules hybrides. Les microparticules présentent des distributions de taille homogènes (20 à 200 µm), sont sphériques et capables de piéger des molécules lipophiles. Ainsi une molécule modèle, a été encapsulé avec des rendements très satisfaisants et sa libération a été totale après 8h en milieu physiologiques simulé. En modifiant la composition des microparticules (ratio inorganique / organique), il a été possible de prolonger les cinétiques de libération et de réduire significativement « l’effet burst ». La biocompatibilité de ces microparticules a été démontrée in vitro. Enfin dans l’optique de correctement caractériser la réaction sol-gel, différentes techniques analytiques ont été explorées pour l’étude in situ de la réticulation et l’identification d’un catalyseur biocompatible. Ces travaux ont permis de détailler les mécanismes des réactions d’hydrolyse et de polycondensation et ont ouvert la voie à un meilleur contrôle des cinétiques de réticulation des microparticules. De plus, il est apparu que la maîtrise de cette réaction semble être indispensable afin d'obtenir des objets "stabilisés " aux propriétés identiques et aux cinétiques de libération reproductibles. En conclusion, il a été démontré que la chimie sol-gel appliquée aux huiles naturelles offre des possibilités d’innovation en formulation galénique tout en respectant les contraintes environnementales et de santé et a permis le développement de matériaux originaux aux propriétés modulables. / Drug formulation is gathering innovative technologies where the use of natural products for the preparation of drug delivery systems is getting more and more considerations because of environmental concerns. For instance, vegetable oils get increasingly used because of their outstanding properties in terms of drug solubilization, biocompatibility and biodegradability. For expanding their application, to drug sustained-release for instance, oils have to be hardened under mild conditions with respect to green chemistry principals. The silica condensation was therefore chosen as cross-linking reaction by the mean the sol-gel reaction which when applied to vegetable oils led to biosourced hybrid organic/inorganic materials. To do so, oils were functionalized with alkoxysilanes precursors without solvent nor catalyst in order to obtain cross-linkable systems. Two chemical paths were studied. The first one, based on an epoxy-amine reaction between epoxydized linseed or soybean oils and the alkoxysilane precursor ended up with an uncontrolled reaction and a triglyceride disruption. The second, used castor oil as an hydroxylated raw material and was based on the hydroxy-isocyanate reaction. Valuable results were obtained and this silylated oil was formulated thanks to a new oil/water thermo-stabilized emulsion process. Simple and robust, this process allowed to simultaneously shape and harden hybrid microparticles. Ranging between 20 and 200 µm in diameter, hybrid microparticles were spherical, homogeneously distributed and were capable of entrapping and releasing lipophilic molecules. As a model, ibuprofen was efficiently encapsulated and was fully released over 8 hours in a simulated buffer. Furthermore, by changing the composition of hybrid microparticles (inorganic/organic ratio), it was also possible to extend release kinetics and significantly reduce the burst effect. Then, biocompatibility of those hybrid microparticles was demonstrated in vitro and an innovative study of the cross-linking reaction was performed. This study aimed to properly understand hydrolysis and polycondensation mechanisms and took the form of an in situ the sol-gel reaction monitoring. It allowed to identify an alternative biocompatible catalysts and gave an insight on how those reactions can be controlled to reach “stabilized" hybrids with constant properties and exhibiting robust and reproducible drug sustained-releases. Finally, it has been demonstrated that the sol-gel chemistry applied to vegetable oils for the synthesis of original and tunable hybrid materials with concerns to environmental and health issues, opened the gate towards innovations in drug formulation.

Considerando a atividade antifúngica demonstrada, o óleo de Cymbopogon citratus apresenta-se como uma promissora matéria-prima no desenvolvimento de produtos farmacêuticos. Para a utilização em formulações, é necessário que se verifique a qualidade do óleo, determinada pelo teor de citral, seu componente majoritário. Além disso, cuidados relacionados à sua estabilidade devem ser considerados. Na busca por método alternativo à técnica de cromatografia a gás, comumente utilizada para óleos voláteis, um dos objetivos deste trabalho foi desenvolver e validar método analítico por cromatografia líquida de alta eficiência para quantificar o citral presente no óleo de C. citratus, na forma de seus isômeros geométricos, o neral e o geranial. Em virtude da volatilidade e suscetibilidade à degradação apresentadas pelo óleo de C. citratus, foram preparadas micropartículas contendo essa substância, a fim de melhorar sua estabilidade. Foram avaliadas duas técnicas de preparação, a secagem por aspersão e a precipitação, além de dois materiais encapsulantes, a beta-ciclodextrina e a hidroxipropil-beta-ciclodextrina. As micropartículas preparadas foram caracterizadas em relação ao teor de umidade, rendimento, porcentagem de inclusão, espectrofotometria na região do infravermelho e a morfologia foi avaliada por microscopia eletrônica de varreduta. Para determinar os teores de neral e geranial presentes nas micropartículas, método por CLAE foi validado. As micropartículas foram incorporadas em emulsões não-iônicas e géis hidrofílicos e estas formulações foram avaliadas quanto às suas propriedades físico-químicas e em relação a sua estabilidade. Os teores de neral e gernanial foram determinados utilizando-se método por CLAE adapatado do método descrito para as micropartículas. / Considering the antifungal activity demonstrated the oil of Cymbopogon citratus presents as a promising raw material for the development of pharmaceutical products. For use in pharmaceutical formulations, it is necessary to verify the quality of the oil, determined by the citral content, its major compound. Moreover, precautions related to its stability should be considered. In the search for alternative method to the technique of gas chromatography, commonly used for volatile oils, one of the purposes of this work was to develop and to validate analytical method by high performance liquid chromatography for quantitation of the citral contents in the C. citratus oil, as its geometric isomers, neral and geranial. Because of the volatility and susceptibility to degradation presented by C. citratus oil, microparticles containing this oil were prepared in order to improve its stability. Two techniques of preparations were evaluated; spraydrying and precipitation, and also two materials of encapsulation, betacyclodextrin and hydroxypropyl-beta-cyclodextrin were tested. The microparticles were characterized by their content of water, yielding, percentage of inclusion, infrared spectroscopy (IV), and the morphology was evaluated by scanning electronic microscopy (SEM). For the determination of neral and geranial contents present into microparticles an HPLC method was validated. The microparticles were incorporated in nonionic emulsions (NIE) and hydrophilic gels (HG), and physico-chemical properties as well as stability of these formulations were evaluated. The neral and geranial contents were determined using the HPLC method adapted from that described for the microparticles.

O presente trabalho visa desenvolver micropartículas na forma farmacêutica pó inalatório contendo levodopa, um fármaco empregado no tratamento da doença de Parkinson. As micropartículas foram preparadas pela técnica de secagem por aspersão utilizando os polímeros ácido hialurônico, quitosana e hidroxipropilmetilcelulose. Desenvolveu-se método analítico indicativo de estabilidade por cromatografia líquida de alta eficiência (CLAE) para o controle de qualidade da formulação, bem como, estudos preliminares de estabilidade e determinação da cinética de fotodegradação. Utilizou-se coluna analítica ACE® RP-18 com tampão fosfato monobásico 0,01 M, ajustado a pH 3,0 como fase móvel, com vazão de 1,0 mL/min e detecção em 280 nm. A linearidade foi obtida na faixa de concentração de 10-60 μg/mL (r2=0,9999) (α=5%). Os limites de quantificação e detecção foram 208 ng/mL e 46,8 ng/mL, respectivamente. Os excipientes e produtos de degradação não apresentaram interferência na eluição da levodopa. Resultados adequados foram encontrados para repetibilidade, precisão intermediária (<2% DPR), exatidão e robustez. Os resultados de recuperação estiveram na faixa de 100,01% a 100,93%. A cinética de fotodegradação em solução frente à luz UVC indicou reação de segunda ordem. A caracterização da formulação demonstrou resultados satisfatórios em relação ao teor, diâmetro aerodinâmico, densidade, teor de umidade e morfologia. A formulação apresentou tamanho de partícula inferior a 16,2 μm e formato arredondado com estrutura oca. A densidade de compactação mostrou valores entre 0,06-0,08 g/cm3 e diâmetro aerodinâmico abaixo de 5 μm, sugerindo que os pós são apropriados para a deposição nas regiões mais profundas do pulmão. Além disso, realizou-se estudo de citotoxicidade pulmonar in vivo, o qual demostrou que a administração intratraqueal das micropartículas não induziu aumentos significativos dos indicadores de toxicidade pulmonar, em comparação ao grupo controle-positivo. Portanto, a avaliação da toxicidade aguda sugere que a liberação pulmonar de levodopa pode ser uma nova e promissora via de administração para este fármaco. / The aim of this study was to develop microparticles containing levodopa for pulmonary delivery, a drug used in the treatment of Parkinson´s disease. The microparticles were prepared by spray-drying using the polymers hyaluronic acid, chitosan and hydroxypropyl methylcellulose. A stability-indicating method was developed and validated for quality control by high performance liquid chromatography (HPLC), as well as, stability studies and photodegradation kinetics. The analytical column ACE® RP-18 was operated with 0.01 M monobasic potassium phosphate, adjusted to a pH value 3.0 as mobile phase, at a flow rate of 1.0 mL/min with detection wavelength at 280 nm. Linearity was obtained over the concentration range of 10-60 μg/mL (r2=0.9999) (α=5%). The quantification limit and detection limit were 208 ng/mL and 46.8 ng/mL, respectively. Excipient ingredients and resulting degradation products had no interference in the levodopa elution. Adequate results were found for repeatability, inter-day precision (<2% RSD), accuracy and robustness. The recovery results were in the range of 100.01% to 100.93%. The photodegradation kinetics in solution front to UVC light indicated the second-order reaction. The formulation showed satisfactory results for drug content, aerodynamic diameter, density, water content and morphology. The formulation presented particle size below 16.2 μm and spherical shape presenting a hollow structure. The tapped density ranged from 0.06-0.08 g/cm3 and an aerodynamic diameter smaller than 5 μm, suggesting that the powders are appropriated for deep lung deposition. Besides that, a cytotoxicity study in vivo was performed which showed that microparticles intratracheal administration did not induce significant increases of lung toxicity indicators compared with the positive control. Therefore, the acute lung toxicity evaluation suggests that pulmonary levodopa delivery could be a new and promising administration route for this drug.

Made available in DSpace on 2014-12-17T14:16:37Z (GMT). No. of bitstreams: 1 EdileneGO_DISSERT.pdf: 1950686 bytes, checksum: 88f16924cd116b850ade306c274f71ac (MD5) Previous issue date: 2014-12-20 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / New drug delivery systems have been used to increase chemotherapy efficacy due the possible drug resistance of cancer cells. Poly (lactic acid) (PLA) microparticles are able to reduce toxicity and prolong methotrexate (MTX) release. In addition, the use of PLA/poloxamer polymer blends can improve drug release due to changes in the interaction of particles with biological surfaces. The aim of this study was developing spray dried biodegradable MTX-loaded microparticles and evaluate PLA interactions with different kinds of Pluronic? (PLUF127 and PLUF68) in order to modulate drug release. The variables included different drug:polymer (1:10, 1:4.5, 1:3) and polymer:copolymer ratios (25:75, 50:50, 75:25). The precision and accuracy of spray drying method was confirmed assessing drug loading into particles (75.0- 101.3%). The MTX/PLA microparticles showed spherical shape with an apparently smooth surface, which was dependent on the PLU ratio used into blends particles. XRD and thermal analysis demonstrated that the drug was homogeneously dispersed into polymer matrix, whereas the miscibility among components was dependent on the used polymer:copolymer ratio. No new drug- polymer bond was identified by FTIR analysis. The in vitro performance of MTX-loaded PLA microparticles demonstrated an extended-release profile fitted using Korsmeyer- Peppas kinetic model. The PLU accelerated drug release rate possible due PLU leached in the matrix. Nevertheless, drug release studies carried out in cell culture demonstrated the ability of PLU modulating drug release from blend microparticles. This effect was confirmed by cytotoxicity observed according to the amount of drug released as a function of time. Thus, studied PLU was able to improve the performance of spray dried MTX-loaded PLA microparticles, which can be successfully used as carries for modulated drug delivery with potential in vivo application / Novos sistemas de libera??o de f?rmacos v?m sendo utilizados para aumentar a efic?cia de quimioter?picos devido ? poss?vel resist?ncia de c?lulas cancer?genas. As micropart?culas de poli (?cido l?ctico) (PLA) constituem uma alternativa para diminuir a toxicidade e prolongar a libera??o do metotrexato (MTX). Al?m disso, o uso de blendas polim?ricas PLA-polox?meros pode melhorar o perfil de libera??o do f?rmaco devido a mudan?as nas intera??es das part?culas com superf?cies biol?gicas. O objetivo do estudo foi desenvolver micropart?culas biodegrad?veis de MTX produzidas por spray drying e avaliar intera??es PLA-Pluronic? (PLA-PLU) para modular a libera??o do f?rmaco, utilizando diferentes tipos de Pluronic? (PLUF127 e PLUF68). As vari?veis de composi??o inclu?ram raz?es f?rmaco:pol?mero (1:10; 1:4,5; 1:3) e pol?mero:copol?mero (25:75, 50:50, 75:25). A reprodutibilidade e a efic?cia do m?todo de produ??o foram confirmadas pela alta efici?ncia de incorpora??o dos sistemas (75,0-101,3%). As micropart?culas de MTX/PLA apresentaram-se esf?ricas com superf?cie aparentemente lisa. Este formato mostrou-se dependente da raz?o pol?mero:copol?mero nas part?culas contendo blendas. A an?lise t?rmica e a difra??o de raios-X sugerem que h? dispers?o do f?rmaco por toda a matriz, enquanto que a miscibilidade entre os componentes foi dependente da raz?o pol?mero:copol?mero. Nenhuma liga??o qu?mica entre o f?rmaco e o pol?mero foi identificada pela an?lise de FTIR. As micropart?culas de PLA contendo MTX apresentaram perfil de libera??o prolongada com um prevalente modelo cin?tico de Korsmeyer-Peppas. O PLU acelerou a taxa de libera??o do f?rmaco devido a sua poss?vel sa?da da matriz polim?rica. Por outro lado, estudos de libera??o do f?rmaco realizados em cultura de c?lulas demonstraram que o PLU modula a taxa de MTX liberado a partir de micropart?culas contendo blendas. Este efeito foi confirmado pela citotoxicidade dos sistemas estudados, de acordo com a quantidade de f?rmaco liberado em fun??o do tempo. Portanto, o uso de PLU foi capaz de melhorar o perfil de libera??o de micropart?culas de PLA contendo MTX, o qual pode ser utilizado como carreador para modular a libera??o do f?rmaco com potencial aplica??o in vivo

Considerando a atividade antifúngica demonstrada, o óleo de Cymbopogon citratus apresenta-se como uma promissora matéria-prima no desenvolvimento de produtos farmacêuticos. Para a utilização em formulações, é necessário que se verifique a qualidade do óleo, determinada pelo teor de citral, seu componente majoritário. Além disso, cuidados relacionados à sua estabilidade devem ser considerados. Na busca por método alternativo à técnica de cromatografia a gás, comumente utilizada para óleos voláteis, um dos objetivos deste trabalho foi desenvolver e validar método analítico por cromatografia líquida de alta eficiência para quantificar o citral presente no óleo de C. citratus, na forma de seus isômeros geométricos, o neral e o geranial. Em virtude da volatilidade e suscetibilidade à degradação apresentadas pelo óleo de C. citratus, foram preparadas micropartículas contendo essa substância, a fim de melhorar sua estabilidade. Foram avaliadas duas técnicas de preparação, a secagem por aspersão e a precipitação, além de dois materiais encapsulantes, a beta-ciclodextrina e a hidroxipropil-beta-ciclodextrina. As micropartículas preparadas foram caracterizadas em relação ao teor de umidade, rendimento, porcentagem de inclusão, espectrofotometria na região do infravermelho e a morfologia foi avaliada por microscopia eletrônica de varreduta. Para determinar os teores de neral e geranial presentes nas micropartículas, método por CLAE foi validado. As micropartículas foram incorporadas em emulsões não-iônicas e géis hidrofílicos e estas formulações foram avaliadas quanto às suas propriedades físico-químicas e em relação a sua estabilidade. Os teores de neral e gernanial foram determinados utilizando-se método por CLAE adapatado do método descrito para as micropartículas. / Considering the antifungal activity demonstrated the oil of Cymbopogon citratus presents as a promising raw material for the development of pharmaceutical products. For use in pharmaceutical formulations, it is necessary to verify the quality of the oil, determined by the citral content, its major compound. Moreover, precautions related to its stability should be considered. In the search for alternative method to the technique of gas chromatography, commonly used for volatile oils, one of the purposes of this work was to develop and to validate analytical method by high performance liquid chromatography for quantitation of the citral contents in the C. citratus oil, as its geometric isomers, neral and geranial. Because of the volatility and susceptibility to degradation presented by C. citratus oil, microparticles containing this oil were prepared in order to improve its stability. Two techniques of preparations were evaluated; spraydrying and precipitation, and also two materials of encapsulation, betacyclodextrin and hydroxypropyl-beta-cyclodextrin were tested. The microparticles were characterized by their content of water, yielding, percentage of inclusion, infrared spectroscopy (IV), and the morphology was evaluated by scanning electronic microscopy (SEM). For the determination of neral and geranial contents present into microparticles an HPLC method was validated. The microparticles were incorporated in nonionic emulsions (NIE) and hydrophilic gels (HG), and physico-chemical properties as well as stability of these formulations were evaluated. The neral and geranial contents were determined using the HPLC method adapted from that described for the microparticles.

O presente trabalho visa desenvolver micropartículas na forma farmacêutica pó inalatório contendo levodopa, um fármaco empregado no tratamento da doença de Parkinson. As micropartículas foram preparadas pela técnica de secagem por aspersão utilizando os polímeros ácido hialurônico, quitosana e hidroxipropilmetilcelulose. Desenvolveu-se método analítico indicativo de estabilidade por cromatografia líquida de alta eficiência (CLAE) para o controle de qualidade da formulação, bem como, estudos preliminares de estabilidade e determinação da cinética de fotodegradação. Utilizou-se coluna analítica ACE® RP-18 com tampão fosfato monobásico 0,01 M, ajustado a pH 3,0 como fase móvel, com vazão de 1,0 mL/min e detecção em 280 nm. A linearidade foi obtida na faixa de concentração de 10-60 μg/mL (r2=0,9999) (α=5%). Os limites de quantificação e detecção foram 208 ng/mL e 46,8 ng/mL, respectivamente. Os excipientes e produtos de degradação não apresentaram interferência na eluição da levodopa. Resultados adequados foram encontrados para repetibilidade, precisão intermediária (<2% DPR), exatidão e robustez. Os resultados de recuperação estiveram na faixa de 100,01% a 100,93%. A cinética de fotodegradação em solução frente à luz UVC indicou reação de segunda ordem. A caracterização da formulação demonstrou resultados satisfatórios em relação ao teor, diâmetro aerodinâmico, densidade, teor de umidade e morfologia. A formulação apresentou tamanho de partícula inferior a 16,2 μm e formato arredondado com estrutura oca. A densidade de compactação mostrou valores entre 0,06-0,08 g/cm3 e diâmetro aerodinâmico abaixo de 5 μm, sugerindo que os pós são apropriados para a deposição nas regiões mais profundas do pulmão. Além disso, realizou-se estudo de citotoxicidade pulmonar in vivo, o qual demostrou que a administração intratraqueal das micropartículas não induziu aumentos significativos dos indicadores de toxicidade pulmonar, em comparação ao grupo controle-positivo. Portanto, a avaliação da toxicidade aguda sugere que a liberação pulmonar de levodopa pode ser uma nova e promissora via de administração para este fármaco. / The aim of this study was to develop microparticles containing levodopa for pulmonary delivery, a drug used in the treatment of Parkinson´s disease. The microparticles were prepared by spray-drying using the polymers hyaluronic acid, chitosan and hydroxypropyl methylcellulose. A stability-indicating method was developed and validated for quality control by high performance liquid chromatography (HPLC), as well as, stability studies and photodegradation kinetics. The analytical column ACE® RP-18 was operated with 0.01 M monobasic potassium phosphate, adjusted to a pH value 3.0 as mobile phase, at a flow rate of 1.0 mL/min with detection wavelength at 280 nm. Linearity was obtained over the concentration range of 10-60 μg/mL (r2=0.9999) (α=5%). The quantification limit and detection limit were 208 ng/mL and 46.8 ng/mL, respectively. Excipient ingredients and resulting degradation products had no interference in the levodopa elution. Adequate results were found for repeatability, inter-day precision (<2% RSD), accuracy and robustness. The recovery results were in the range of 100.01% to 100.93%. The photodegradation kinetics in solution front to UVC light indicated the second-order reaction. The formulation showed satisfactory results for drug content, aerodynamic diameter, density, water content and morphology. The formulation presented particle size below 16.2 μm and spherical shape presenting a hollow structure. The tapped density ranged from 0.06-0.08 g/cm3 and an aerodynamic diameter smaller than 5 μm, suggesting that the powders are appropriated for deep lung deposition. Besides that, a cytotoxicity study in vivo was performed which showed that microparticles intratracheal administration did not induce significant increases of lung toxicity indicators compared with the positive control. Therefore, the acute lung toxicity evaluation suggests that pulmonary levodopa delivery could be a new and promising administration route for this drug.

Ce travail porte sur le rôle des microparticules procoagulantes (MPs) générées par les cellules vasculaires en réponse à un état septique. Après une introduction rappelant la structure et les propriétés des microparticules et la réponse del’hôte à un agent pathogène, en particulier en terme d’activation de la coagulation, nous rapportons nos travauxexpérimentaux et cliniques. Le premier travail a été réalisé sur un modèle cellulaire de vésiculation induite par le LPS. Il nous a permis de caractériser le transfert du complexe CD14/TLR4 à différents types cellulaires in-vitro dépourvus du récepteur au LPS. Ainsi, les MPs monocytaires pourraient avoir un rôle d’amplification de la réponse inflammatoire mais aussi dans la réponse anti-inflammatoire secondaire en participant à l’apoptose lymphocytaire. Le second travail aété réalisé chez l’animal. Après induction d’un choc septique, nous avons observé une amélioration hémodynamique enréponse à la perfusion de protéine C activée associée à une modulation du phénotype des MPs. Réinjectées à des ratsnaïfs, les MPs issues des rats septiques traités par protéine C activée développaient une moindre vasoplégie. Enfin, nous avons réalisé une étude prospective sur 100 patients en choc septique. Nous avons ainsi pu caractériser la présence d’une concentration élevée de microparticules procoagulantes, avec une variation phénotypique en présence de coagulation intravasculaire disséminée (CIVD) : réduction du contingent plaquettaire au profit des MPs d’origine leucocytaires qui deviennent prépondérantes et témoignent d’une activation leucocytaire accrue, et surtout une activation des cellules endothéliales avec génération de MPs porteuses d’endogline (CD105). En analyse multivariée,CD105+-MPs étaient fortement associée à la CIVD et pourraient constituer un marqueur précoce de l’atteinte endothéliale au cours du choc septique. / This work focused on procoagulant microparticles shed after vascular cells stress during sepsis. The first part gives an overview on MPs and host response during pathogen challenge. The first lab experimental work confirms direct and functional transfer of CD14/TLR4 LPS sensor by MPs shed to target cells after monocytic THP-1 challenge by LPS.CD14-MPs amplify LPS-induced apoptosis in monocytes but also prompted lymphocyte apoptosis and could play a role in secondary anti-inflammatory response. Then, septic shock was induced in rats after caecal ligature and puncture.Activated protein C (APC) infusion improved haemodynamic parameters and alter septic microparticular content. Infused in naïve rats, APC-treated MPs were associated with reduced hypotension and inflammatory response, confirming cytoprotective effect of both APC and APC-induced MPs. Finally, we performed a clinical prospective study in 3 medical ICU in France. Patients referred for septic shock had an increased level of circulating procoagulant MPs regardless disseminated intravascular coagulopathy (DIC) diagnosis. Nevertheless, DIC patients evidenced a specific pattern with lower platelet-MPs, increased leucocyte-MPs and specific endothelial cells activation with endoglin (CD105) shedding. In multiple logistic regression analysis, CD105-MPs were strongly associated with DIC and were evidenced before DIC diagnosis according to routine laboratory assays.

O presente trabalho visa desenvolver micropartículas na forma farmacêutica pó inalatório contendo levodopa, um fármaco empregado no tratamento da doença de Parkinson. As micropartículas foram preparadas pela técnica de secagem por aspersão utilizando os polímeros ácido hialurônico, quitosana e hidroxipropilmetilcelulose. Desenvolveu-se método analítico indicativo de estabilidade por cromatografia líquida de alta eficiência (CLAE) para o controle de qualidade da formulação, bem como, estudos preliminares de estabilidade e determinação da cinética de fotodegradação. Utilizou-se coluna analítica ACE® RP-18 com tampão fosfato monobásico 0,01 M, ajustado a pH 3,0 como fase móvel, com vazão de 1,0 mL/min e detecção em 280 nm. A linearidade foi obtida na faixa de concentração de 10-60 μg/mL (r2=0,9999) (α=5%). Os limites de quantificação e detecção foram 208 ng/mL e 46,8 ng/mL, respectivamente. Os excipientes e produtos de degradação não apresentaram interferência na eluição da levodopa. Resultados adequados foram encontrados para repetibilidade, precisão intermediária (<2% DPR), exatidão e robustez. Os resultados de recuperação estiveram na faixa de 100,01% a 100,93%. A cinética de fotodegradação em solução frente à luz UVC indicou reação de segunda ordem. A caracterização da formulação demonstrou resultados satisfatórios em relação ao teor, diâmetro aerodinâmico, densidade, teor de umidade e morfologia. A formulação apresentou tamanho de partícula inferior a 16,2 μm e formato arredondado com estrutura oca. A densidade de compactação mostrou valores entre 0,06-0,08 g/cm3 e diâmetro aerodinâmico abaixo de 5 μm, sugerindo que os pós são apropriados para a deposição nas regiões mais profundas do pulmão. Além disso, realizou-se estudo de citotoxicidade pulmonar in vivo, o qual demostrou que a administração intratraqueal das micropartículas não induziu aumentos significativos dos indicadores de toxicidade pulmonar, em comparação ao grupo controle-positivo. Portanto, a avaliação da toxicidade aguda sugere que a liberação pulmonar de levodopa pode ser uma nova e promissora via de administração para este fármaco. / The aim of this study was to develop microparticles containing levodopa for pulmonary delivery, a drug used in the treatment of Parkinson´s disease. The microparticles were prepared by spray-drying using the polymers hyaluronic acid, chitosan and hydroxypropyl methylcellulose. A stability-indicating method was developed and validated for quality control by high performance liquid chromatography (HPLC), as well as, stability studies and photodegradation kinetics. The analytical column ACE® RP-18 was operated with 0.01 M monobasic potassium phosphate, adjusted to a pH value 3.0 as mobile phase, at a flow rate of 1.0 mL/min with detection wavelength at 280 nm. Linearity was obtained over the concentration range of 10-60 μg/mL (r2=0.9999) (α=5%). The quantification limit and detection limit were 208 ng/mL and 46.8 ng/mL, respectively. Excipient ingredients and resulting degradation products had no interference in the levodopa elution. Adequate results were found for repeatability, inter-day precision (<2% RSD), accuracy and robustness. The recovery results were in the range of 100.01% to 100.93%. The photodegradation kinetics in solution front to UVC light indicated the second-order reaction. The formulation showed satisfactory results for drug content, aerodynamic diameter, density, water content and morphology. The formulation presented particle size below 16.2 μm and spherical shape presenting a hollow structure. The tapped density ranged from 0.06-0.08 g/cm3 and an aerodynamic diameter smaller than 5 μm, suggesting that the powders are appropriated for deep lung deposition. Besides that, a cytotoxicity study in vivo was performed which showed that microparticles intratracheal administration did not induce significant increases of lung toxicity indicators compared with the positive control. Therefore, the acute lung toxicity evaluation suggests that pulmonary levodopa delivery could be a new and promising administration route for this drug.

Considerando a atividade antifúngica demonstrada, o óleo de Cymbopogon citratus apresenta-se como uma promissora matéria-prima no desenvolvimento de produtos farmacêuticos. Para a utilização em formulações, é necessário que se verifique a qualidade do óleo, determinada pelo teor de citral, seu componente majoritário. Além disso, cuidados relacionados à sua estabilidade devem ser considerados. Na busca por método alternativo à técnica de cromatografia a gás, comumente utilizada para óleos voláteis, um dos objetivos deste trabalho foi desenvolver e validar método analítico por cromatografia líquida de alta eficiência para quantificar o citral presente no óleo de C. citratus, na forma de seus isômeros geométricos, o neral e o geranial. Em virtude da volatilidade e suscetibilidade à degradação apresentadas pelo óleo de C. citratus, foram preparadas micropartículas contendo essa substância, a fim de melhorar sua estabilidade. Foram avaliadas duas técnicas de preparação, a secagem por aspersão e a precipitação, além de dois materiais encapsulantes, a beta-ciclodextrina e a hidroxipropil-beta-ciclodextrina. As micropartículas preparadas foram caracterizadas em relação ao teor de umidade, rendimento, porcentagem de inclusão, espectrofotometria na região do infravermelho e a morfologia foi avaliada por microscopia eletrônica de varreduta. Para determinar os teores de neral e geranial presentes nas micropartículas, método por CLAE foi validado. As micropartículas foram incorporadas em emulsões não-iônicas e géis hidrofílicos e estas formulações foram avaliadas quanto às suas propriedades físico-químicas e em relação a sua estabilidade. Os teores de neral e gernanial foram determinados utilizando-se método por CLAE adapatado do método descrito para as micropartículas. / Considering the antifungal activity demonstrated the oil of Cymbopogon citratus presents as a promising raw material for the development of pharmaceutical products. For use in pharmaceutical formulations, it is necessary to verify the quality of the oil, determined by the citral content, its major compound. Moreover, precautions related to its stability should be considered. In the search for alternative method to the technique of gas chromatography, commonly used for volatile oils, one of the purposes of this work was to develop and to validate analytical method by high performance liquid chromatography for quantitation of the citral contents in the C. citratus oil, as its geometric isomers, neral and geranial. Because of the volatility and susceptibility to degradation presented by C. citratus oil, microparticles containing this oil were prepared in order to improve its stability. Two techniques of preparations were evaluated; spraydrying and precipitation, and also two materials of encapsulation, betacyclodextrin and hydroxypropyl-beta-cyclodextrin were tested. The microparticles were characterized by their content of water, yielding, percentage of inclusion, infrared spectroscopy (IV), and the morphology was evaluated by scanning electronic microscopy (SEM). For the determination of neral and geranial contents present into microparticles an HPLC method was validated. The microparticles were incorporated in nonionic emulsions (NIE) and hydrophilic gels (HG), and physico-chemical properties as well as stability of these formulations were evaluated. The neral and geranial contents were determined using the HPLC method adapted from that described for the microparticles.