Ausgangspunkt dieser vorliegenden Arbeit waren bisherige Untersuchungen unseres Arbeitskreises zum Memb-ranverankerungsverhalten (Phospholipidmembranen, LUV) von Nukleosiden und Oligonukleotiden, welche einen lipophilen Anker an der 5-Position der Pyrimidin- oder an der 8-Position der Purinbase tragen. Diese Nukleolipide ankern gut in der Membran, stehen aber nicht mehr für eine Watson-Crick-Basenpaarung an der Phasengrenzfläche zu Verfügung. Demnach wurde durch die Verwendung unterschiedlicher Reaktionen (Veresterung, Thioetherbildung, Carbamoylverknüpfung oder „Clickreaktion“ zu Triazolen) und verschiedener funktioneller Gruppen (Hydroxy, Thiohydroxy, Azid, Amin) an die 2´-Position der Nukleoside eine Reihe von lipophilen Resten (Alkylketten, Cholesterol, Pyren) eingeführt. Diese Konjugate verankerten ebenfalls gut in den Membranen und es zeigten sich erste Hinweise, dass durch die Einführung eines Spacers zwischen dem Nukleosid und dem lipophilen Anker, eine Basenpaarung an der Phasengrenzfläche möglich ist. Weiterhin zeigte es sich, dass Nukleolipide mit nur einem lipophilen Rest nicht stabil in Membranen verankern, vor allem, wenn dieser nicht verzweigt ist. Bei der Anwendung von Oligonukleotiden zum Ankern in Membranen ist es unbedeu-tend, an welcher Stelle der lipophile Rest am Nukleotid vorkommt, denn zum einen geht das entsprechende Nukleolipid selbst keine Basenpaarung ein und zum anderen erfolgt keine Basenpaarung über dieses hinweg. Für biotechnologische Anwendungen konnte mit Hilfe dieser synthetisierten lipophilen Oligonukleotide gezeigt werden, dass zwei vesikelmembranverankerte Oligonukleotide, welche komplementäre Enden tragen, eine Doppelhelix miteinander bilden und so diese beiden Vesikel auf einen definierten Abstand halten können. Da Nukleolipide einen amphiphilen Charakter aufweisen, sollte unter dem AFM untersucht werden, ob diese supramolekulare Strukturen zeigen. Dies wurde in der Tat auch beobachtet. Ebenso konnten mittels der LB-Technik LB-Schichten aus Nukleolipiden dargestellt werden. / The starting point of this work was found in our previous studies about anchoring behaviour of lipidated nucleo-sides and oligonucleotides in biocompatible phospholipid membranes (LUV). That nucleosides and oligonucleotides bear a lipophilic anchor at the 5-position of pyrimidine or at the 8-position of purinbases. This nucleolipi-des anchor well in such membranes, but were not longer available for a Watson-Crick base pairing at the interface to water. Therefore lipophilic groups (alkyl chain, cholesterol, Pyren etc.) were now connected to the 2''-position of nucleosides by several reactions (esterification, thioether binding, carbamoyl binding or "click reaction") and various functional groups (hydroxy, thiohydroxy, azide, amine) to the 2´-position of nucleosides. These nucleolipides also well anchored in the model membranes, and gave first evidence that by introducing a spacer between the nucleoside and the lipophilic anchor a base pairing at the interface to water is possible. However, only one anchor is not sufficient for a stable anchoring in the phospholipid membranes, especially if they are not branched. It was found out that it is insignifacant for the application of oligonucleotides in membrane anchoring, at which position of nucleotide the lipid is attached, because on the one hand, the corresponding nucleolipid can not form a pair with a corresponding nucleobase and secondly, there is no base pairing in the nucleotides situated between two lipidated positions. For biotechnology applications it might be interesting that two different vesicles each of it furnushed with a complementary lipidated oligonucleotide could be kept together in a defined distance by forming double strand DNA. Since nucleolipide possess amphiphilic character, there abillity to form supramolecular structures was investigated by atomic force microscope (AFM). In addition formation of LB-layers could be achieved by LB-technology.

O ibuprofeno é um antiinflamatório não esteróide, com baixa solubilidade em água, que apresenta diversos efeitos colaterais, incluindo lesão gástrica e intestinal. Esses efeitos podem depender da interação com a membrana celular, o que nos motivou a investigar, na primeira parte deste trabalho, a incorporação do ibuprofeno em monocamadas de Langmuir como modelos de membrana celular. Monocamadas de dipalmitoil fosfatidil glicerol (DPPG) e dipalmitoil fosfatidil colina (DPPC) co-espalhadas com o ibuprofeno ou depositadas sobre subfases contendo o fármaco foram estudadas por meio das isotermas de pressão e potencial de superfície. Foram observados efeitos significativos para monocamadas de DPPC, particularmente na transição de fase líquido-expandida para líquido-condensada, com modificações relevantes na elasticidade da monocamada. Esses efeitos aumentaram com a concentração do ibuprofeno. Para os dois tipos de fosfolipídios, o ibuprofeno pôde penetrar na região hidrofóbica, o que foi confirmado por espectroscopia de reflexão e absorção no infravermelho com modulação da polarização (PM-IRRAS), indicando assim a presença de interações hidrofóbicas. A análise por microscopia no ângulo de Brewster (BAM) mostrou que o ibuprofeno impede a formação de grandes domínios de DPPC, enquanto que não foram observadas alterações significativas para o DPPG. A interação entre o ibuprofeno e o DPPG também foi confirmada após a imobilização da monocamada mista em filmes LB com alterações na absorção no UV-Vis da molécula de ibuprofeno. No que diz respeito às implicações biológicas, a ação farmacológica que depende diretamente da interação com a membrana deve ocorrer primeiramente em regiões neutras via penetração do ibuprofeno na região hidrofóbica da membrana celular. A segunda parte deste trabalho foi dedicada à interação de capsaicinóides extraídos da pimenta malagueta com monocamadas de Langmuir constituídas de DPPG e DPPC. A capsaicina é um potente analgésico de uso tópico, que pode causar dessensibilização no local de aplicação dependendo da dose e, portanto há interesse na sua incorporação em sistemas de liberação controlada, como os lipossomos. A técnica de Langmuir foi empregada para verificar essa possibilidade. Os capsaicinóides expandiram as monocamadas de DPPG e aumentaram sua elasticidade. As isotermas de potencial de superfície indicaram que os capsaicinóides provocam aumento de 10% no momento de dipolo numa concentração de 30% em mol. Para os filmes mistos de DPPC e capsaicinóides, a área mínima diminuiu e a elasticidade da monocamada aumentou. De acordo com as isotermas de potencial, os momentos de dipolo diminuíram para as monocamadas de DPPC independentemente da concentração de capsaicinóides. Esses resultados sugerem que as moléculas de DPPC são solubilizadas para a subfase na presença do fármaco. A partir destes resultados, conclui-se que os capsaicinóides podem ser incorporados em estruturas lipídicas, constituídas principalmente de DPPG, o que é relevante para uso em sistemas de liberação de fármacos. / Ibuprofen is a nonsteroidal anti-inflammatory drug, with low solubility in water, which exhibits side effects including gastric and intestinal injury, often irreversible. Some of these effects may depend on the interaction with the cell membrane, which motivated us to investigate the incorporation of ibuprofen in Langmuir monolayers as cell membrane models, in the first part of this thesis. Dipalmitoyl phosphatidyl choline (DPPC) or dipalmitoyl phosphatidyl glycerol (DPPG) monolayers co-spread with ibuprofen or deposited on ibuprofen-containing aqueous subphases were studied using surface pressure and surface potential isotherms. Significant effects were observed for DPPC monolayers, particularly at the liquid-expanded to liquid-condensed phase transition, with relevant changes in the elasticity of the monolayer. These effects increased with the ibuprofen concentration. For both types of phospholipids, ibuprofen could penetrate into the hydrophobic part of the monolayer, which was confirmed with polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS), thus indicating the presence of hydrophobic interactions. BAM images showed that ibuprofen prevents the formation of large domains of DPPC, while no significant changes were observed for DPPG. The interaction between DPPG-ibuprofen was also confirmed for deposited layers in the form of LB films, with changes in the ibuprofen UV-Vis absorption. As for the biological implications, the pharmacological action depending directly on the membrane interaction should occur primarily with zwitterionic regions of the membrane via penetration of ibuprofen in the hydrophobic part of the monolayer. The second part of this thesis is dedicated to the interaction of capsaicinoids, extracted from malagueta pepper, with Langmuir monolayers of DPPC and DPPG. Capsaicin is a powerful analgesic of topical use, which can cause desensitization in the application site depending on the dose, and therefore there is interest in its incorporation in drug delivery systems, such as liposomes. The Langmuir technique was employed to verify this possibility. The capsaicinoids expanded the DPPG monolayer and increased its elasticity. Surface potential isotherms indicated that the capsaicinoids increased the average dipole moment by 10 % for 30 mol % of capsaicinoids. For the mixed films of DPPC and capsaicinoids, the minimum area decreased and the elasticity increased. According to the surface potential isotherms, the dipole moments decreased for DPPC monolayers regardless of the capsaicinoid concentrations. These results suggest that the DPPC molecules are solubilized into the subphase in the presence of the drug. From these results, it is concluded that the capsaicinoids can be incorporated into structures as the liposomes constituted mainly of DPPG, which is relevant for use in drug delivery systems.

Ces travaux de thèse portent sur les phénomènes d’amplification du signal de diffusion Raman par effet de surface et par effet de pointe. Des réseaux de motifs métalliques de taille nanométrique arrangés spatialement ont été fabriqués par la méthode de transfert Langmuir-Blodgett et par lithographie à faisceau d’électrons. De telles structures de géométries contrôlées déposées à la surface de lamelles de microscope ont été développées afin d’amplifier le signal Raman de molécules adsorbées par effet SERS (Surface Enhanced Raman Spectroscopy). Ces nanostructures triangulaires en or de taille proche de la longueur d’onde ont des bandes de résonance plasmon dans le domaine spectral visible. En utilisant une source de laser appropriée dans ce domaine spectral, les facteurs d’amplification Raman d’une couche mono-moléculaire d’un dérivé azobenzène sont de plusieurs ordres de grandeur, et ce pour les deux techniques de nano-lithographie employées. Afin de compléter ces premiers résultats, des réseaux de fils d’or avec de grands facteurs de forme ont été fabriqués. Ces derniers montrent des résonances plasmons multipolaires et des facteurs d’amplification de l’ordre de 105. Les techniques de microscopie en champ proche ont également été développées afin de localiser précisément l’exaltation Raman et d’accroitre la résolution spatiale de mesures Raman. Des pointes métalliques en or de taille nanométrique ont ainsi permis d’amplifier localement le signal de diffusion de molécules placées à leur proximité par effet TERS (Tip Enhanced Raman Spectroscopy). Les développements logiciels et mécaniques entre un microscope confocal Raman et un microscope à force atomique ont été implémentés afin de contrôler simultanément les deux instruments. Ce montage expérimental a été appliqué à l’étude de nanofils semi-conducteurs de nitrure de gallium permettant de suivre leur signal vibrationnel avec une résolution spatiale inférieure à 200 nm. / This thesis work focuses on Raman scattering enhancements by metallic nanostructures. In the first part of this work, arrays of metallic patterns with nanometer dimensions were fabricated by the Langmuir-Blodgett deposition technique and electron-beam lithography. Such structures made of gold were fabricated onto microscope slides with the goal to enhance the Raman signal through SERS effect (Surface Enhanced Raman Spectroscopy). These patterns formed by an assembly of triangular nanostructures with sizes of hundreds of nanometers, exhibit plasmon resonance bands in the visible spectral region. By using an appropriate excitation laser source with respect to the plasmon frequency, Raman enhancement factors of a monolayer were found to be of several order of magnitude for both Langmuir-Blodgett and electron-beam lithography platforms. To further complement these results, gold wires arrays with large aspect ratio made by electron-beam lithography showed multipolar plasmon resonances with enhancement factors up to 105. In the second part of this thesis, near-field Raman microscopy has been developed with the aim to localize precisely the Raman enhancement and improve spatial resolution of Raman measurements. Atomic force microscopy gold tips have been used to locally enhance scattering signal of molecules in close proximity to the tip opening new opportunities. This approach known as TERS (Tip Enhanced Raman Spectroscopy) is of significant interest to probe nanomaterials, nanostructures or monolayers. Software and mechanical developments have been made between a confocal Raman microscope and an atomic force microscope to control simultaneously both instruments. This experimental setup was used to characterize gallium nitride semi-conductors nanowires with spatial resolution better than 200 nm.

Une nouvelle méthode de détection sans marquage des interactions biomoléculaires a été développée. Elle est basée sur la conception d’une couche sensible supportée innovante réalisée par la technique de Langmuir‐Blodgett. Cette couche est composée d'un lipide à tête polaire immobilisant un cation métallique divalent capable (i) de chélater une molécule possédant une affinité pour ce type de cation et (ii) de catalyser la réaction de chimiluminescence du luminol. L'intensité du signal chimiluminescent est modulée par la présence de biomolécules fixées en surface de la couche sensible, qui modifient l'environnement immédiat du cation métallique. La variation du signal chimiluminescent (issu de la catalyse par le cation immobilisé) a pu être quantifiée et corrélée à une gamme de concentration d’histamine et d’anticorps. Les potentialités de cette approche ont finalement été exploitées pour développer une puce de PMMA de type macropuce immobilisant la monocouche de lipides. / A new label‐free detection method for biomolecular interactions was developed. It is based on the development of an original sensitive layer performed with the Langmuir‐ Blodgett technique. This layer is composed of a lipid which immobilized a bivalent metallic cation on his polar head, able (i) to chelate a molecule which has an affinity for this cation and (ii) to catalyze the luminol chemiluminescent reaction. The chemiluminescent signal intensity is modulated by the presence of immobilized biomolecules on the sensitive layer surface, which modifies the immediate environmentof the metallic cation. The chemiluminescent signal variation (from catalysis by the metallic cation) has been quantified and correlated to a histamine and antibody concentration range. The potentialities of this approach have finally been employed to achieve a PMMA chip (macroarray type), immobilizing the lipid monolayer.

Gegenstand der vorliegenden Arbeit ist die Entwicklung, Synthese und Charakterisierung neuartiger oligofunktionalisierter Amphiphile mit potentiellen Sammler- und LB-Filmbildungseigenschaften. Basierend auf dem entwickelten Konzept der supramolekularen Strukturbildung zum Aufbau präorganisierter und gebündelter Sammleraggregate wurden zwei Typen neuartiger oligofunktioneller Tenside synthetisiert, die sich insbesondere in ihrer konformativen Flexibilität und Hydrophilie-Lipophilie-Balance unterscheiden. Ausgeprägte oberflächenaktive Eigenschaften wurden bei den bi-, tri- und tetrafunktionellen Aminosäure-analogen Zielverbindungen (Typ II) mit linearer, trigonaler und tetragonaler Molekülstruktur gefunden. Diese Verbindungen bilden auch stabile monomolekulare Filme geringer Heterogenität aus. Im mineralischen Flotationsprozess ergeben diese präorganisierten Tenside mit geometrisch kontrollierter Positionierung der lateralen Funktionseinheiten im Vergleich zu konventionellen Sammlern bei spezifischem Verhalten eine bemerkenswerte Steigerung der flotativen Effizienz.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

El virus C (GBV-C) o virus de la hepatitis G (HGV) está relacionado con el virus de la hepatitis C (HCV) ya que tiene una organización genómica y una homología en la secuencia muy próxima pero, sin embargo, no parece causar hepatitis ni ningún otro tipo de patología. En los últimos años, se han publicado numerosos trabajos en los que se asocia la co-infección del GBV-C/HGV y el virus de la inmunodeficiencia humana (VIH) con una menor progresión de la enfermedad causada por el virus VIH así como con una mayor supervivencia de los pacientes una vez que el SIDA se ha desarrollado. Basándonos en un modelo de co-infección, existen diversas teorías de cómo el virus de la hepatitis G puede influir en la enfermedad del VIH, se cree que puede inducir las quemoquinas, regular negativamente los co-receptores del VIH, así como provocar otros efectos indefinidos en los linfocitos huésped. Pero el mecanismo responsable del efecto beneficioso que el GBV-C/HGV tiene sobre el curso de la infección causada por VIH no está todavía definido. Teniendo en cuenta el posible uso de los péptidos sintéticos, el objetivo de la actual tesis doctoral es llevar a cabo la selección de péptidos de la proteína de envoltura E1 del GBV-C/HGV con potencial capacidad de inhibición del péptido de fusión del VIH-1, mediante la síntesis múltiple en paralelo de secuencias peptídicas lineales que cubren la totalidad de la estructura primaria de esta proteína y su posterior evaluación mediante ensayos biofísicos. Con ello se pretende plantear la utilización de péptidos sintéticos del GBV-C/HGV en el diseño de inhibidores de la entrada del VIH en la célula huésped. Se pretende así tratar de comprender mejor el efecto de la presencia de anticuerpos anti-GBV-C/HGV en la progresión de la enfermedad causada por la infección del VIH y de profundizar en el conocimiento actual acerca de la interacción entre estos dos virus. Se realiza la síntesis en fase sólida de 58 péptidos correspondientes a la proteína de envoltura E1 del GBV-C/HGV y mediante el ensayo de liberación de contenidos vesiculares se seleccionan 5 péptidos capaces de inhibir la capacidad del péptido de fusión (PF) del VIH-1 de formar poros en las membranas. Se realizan estudios detallados de los péptidos y de la inhibición del proceso de fusión, inducido por el péptido de fusión del VIH-1, por parte de secuencias del GBV-C/HGV. Para ello se realizan estudios de citotoxicidad, ensayos de hemólisis, estudio de la estructura mediante dicroísmo circular, ensayos de agregación de liposomas inducida por los péptidos y ensayos de transferencia de energía por resonancia. Por otra parte se realizan ensayos utilizando monocapas de extensión como modelos de membrana mediante la técnica de Langmuir-Blodgett, y se calculan las concentraciones superficial de exceso y las de saturación, información que servirá para conocer la capacidad de los péptidos para formar monocapas estables en la interfase aire/agua. Los diferentes ensayos muestran una interacción de los péptidos de la proteína E1 con el péptido de fusión del VIH-1, dando lugar a una inhibición de la actividad que el péptido de fusión del VIH-1 tiene sobre las membranas lipídicas. Esto sugiere que estos péptidos pueden ser candidatos para ser utilizados en futuras terapias antivirales. / "BIOPHYSICAL STUDIES OF SYNTHETIC PEPTIDES OF THE ENVELOPE PROTEIN E1 OF GBV-C/HGV AND ITS RELATIONSHIP WITH HIV" The virus C (GBV-C) or hepatitis G virus (HGV) is related to hepatitis C virus (HCV) because it has similar genome organization and sequence homology; however, it does not seem to cause hepatitis or any other pathology. In recent years, there have been several studies in which the GBV-C/HGV and human immunodeficiency virus (HIV) co-infection is associated with less progression of the disease caused by HIV as well as improved survival of patients after AIDS has developed. Based on a model of co-infection, there are several theories of how the hepatitis G virus can affect HIV disease: it could induce chemokines, negatively regulate the HIV co-receptors and cause other undefined effects in the host cell. But the mechanism responsible for the beneficial effect that GBV-C/HGV has over the course of infection by HIV is not yet defined. Taking into account the possible use of synthetic peptides, the aim of the present thesis is to carry out the selection of peptides from the envelope protein E1 of GBV-C/HGV with potential ability to inhibit the HIV-1fusion peptide. The aim is to better understand the effect of the presence of antibodies anti-GBV-C/HGV in the progression of AIDS and to study the interaction between these two viruses. The solid phase synthesis of 58 peptides corresponding to the envelope protein E1 of GBV-C/HGV was made and 5 peptides capable of inhibiting the ability of the fusion peptide (FP) of HIV-1 to form pores in membranes were selected by means of the leakage of vesicular contents assay. Detailed studies of the E1 peptides and their ability to inhibit the fusion process induced by the fusion peptide of HIV-1 were done. Monolayers and liposomes were used as model membranes. The different tests show an interaction of the E1 peptides with the HIV-1fusion peptide, resulting in an inhibition of the activity that the fusion peptide has on lipid membranes. This suggests that these peptides could be good candidates for future antiviral therapies.

Este trabalho visa ao entendimento da interação de capsaicinóides com membranas celulares utilizando sistemas modelo. Dentre os alcalóides derivados de plantas do gênero Capsicum, a capsaicina e a dihidrocapsaicina respondem por 90% dos capsaicinóides, que são usados como analgésicos e antiinflamatórios, devido a sua interação específica com receptores. O mecanismo neurofarmacológico já foi bastante estudado, mas o modo de ação não neural ainda não foi elucidado. Usamos extratos brutos (EBs) de pimenta malagueta e de dedo-de-moça, que têm atividade superficial, e afetaram monocamadas de Langmuir de fosfatidil colina de dipalmitoíla (DPPC) e fosfatidil glicerol de dipalmitoíla (DPPG). Tais efeitos não tiveram dependência expressiva com a carga, pois o EB de dedo-de-moça interagiu mais fortemente com o DPPC do que com o DPPG, ao passo que o contrário se verificou para o EB de malagueta. Também não houve diferença significativa entre os EBs das duas pimentas. Nas monocamadas de Langmuir representativas para a bactéria S. aureus, ambos os EBs tiveram efeito, tanto nas isotermas de pressão quanto nos resultados de espectroscopia de absorção e reflexão no infravermelho com modulação de polarização (PM-IRRAS), sem distinção significativa entre malagueta e dedo-de-moça. No entanto, as medidas de vazamento com lipossomos mostraram maior interação com o EB de dedo-de-moça, o que é consistente com a atividade bactericida para S. aureus. De fato, a concentração inibitória mínima (MIC) foi 0,13 mg mL-1 para o EB da pimenta dedo-de moça e 4,0 mg mL-1 para o EB de malagueta. Para a E. coli, os EBs interagiram com as monocamadas de Langmuir sem diferenças dignas de nota para as duas pimentas, e nas medidas de vazamento o efeito maior foi para a dedo-de-moça. Não houve efeito bactericida para nenhum dos extratos. Isso se explica porque bactérias gram-negativas, como a E. coli, têm uma camada externa protetora de lipossacarídeos (LPS). Das medidas de monocamadas de Langmuir representativas da camada de LPS, observou-se pouca incorporação dos EBs. Conclui-se, assim, que os EBs não conseguem causar rompimento da camada de LPS. Do conjunto dos resultados, infere-se que o mecanismo de ação para a S. aureus envolve solubilização parcial da membrana, e não há relação entre pungência e atividade bactericida, pois a pimenta dedo-de-moça, que é menos pungente, teve maior efeito do que a malagueta. Depreende-se, também, que a ação de extratos de pimenta deve depender da interação com receptores na membrana, o que explica porque o uso de tais extratos tem sido principalmente em aplicações tópicas. / This study is aimed at understanding the interaction of capsaicinoids with cell membranes using model systems. Among the alkaloids derived from plants of the genus Capsicum, capsaicin and dihydrocapsaicin account for 90% of capsaicinoids, which are used as analgesic and anti-inflammatory due to their interaction with specific receptors. The neuropharmacological mechanism has been well studied, but the non-neural mode of action has not been elucidated. Here, we use crude extracts (EBs) of malagueta and dedo-de-moça chilli peppers, which are surface active, and affected Langmuir monolayers of dipalmitoyl phosphatidylcholine (DPPC) and dipalmitoyl phosphatidyl glycerol (DPPG). Such effects did not depend on the charge, since EB from dedo-de-moça interacted more strongly with DPPC than with DPPG, while the opposite applied for malagueta. In addition, there was no significant difference between the two EBs. For Langmuir monolayers representing the bacteria S. aureus, both EBs affected the surface pressure isotherms and the polarizationmodulated infrared reflection-absorption spectroscopy (PM-IRRAS) data, without significant distinction between dedo-de-moça and malagueta. However, in the leakage measurements with liposomes the EB from dedo-de-moça was more efficient in rupturing the liposome, which is consistent with the bactericidal activity for S. aureus. In fact, the minimum inhibitory concentration (MIC) was 0.13 mg mL-1 for dedo-de-moça and 4.0 mg mL-1 for malagueta. For the Langmuir monolayers mimicking the E. coli membrane, the EBs interacted much in the same way, while the EB from dedo-de-moça caused larger leakage in liposomes. There was no bactericidal effect of the EBs. This is explained by the fact that gram-negative bacteria, such as E. coli, have a protective outer layer of liposaccharides (LPS). In monolayers representing LPS, there was little incorporation of EBs, from which one infers that the EBs cannot cause disruption of the LPS layer. Taking all these results together, it appears that the mechanism of action for S. aureus involves partial solubilization of the membrane. Furthermore, there is no relationship between pungency and bactericidal activity because dedo-de-moça, which is less pungent, had greater effect than malagueta. It seems also that the action of pepper extracts must depend on the interaction with membrane receptors, which explains why the use of such extracts has been essentially in topical applications.

O ibuprofeno é um antiinflamatório não esteróide, com baixa solubilidade em água, que apresenta diversos efeitos colaterais, incluindo lesão gástrica e intestinal. Esses efeitos podem depender da interação com a membrana celular, o que nos motivou a investigar, na primeira parte deste trabalho, a incorporação do ibuprofeno em monocamadas de Langmuir como modelos de membrana celular. Monocamadas de dipalmitoil fosfatidil glicerol (DPPG) e dipalmitoil fosfatidil colina (DPPC) co-espalhadas com o ibuprofeno ou depositadas sobre subfases contendo o fármaco foram estudadas por meio das isotermas de pressão e potencial de superfície. Foram observados efeitos significativos para monocamadas de DPPC, particularmente na transição de fase líquido-expandida para líquido-condensada, com modificações relevantes na elasticidade da monocamada. Esses efeitos aumentaram com a concentração do ibuprofeno. Para os dois tipos de fosfolipídios, o ibuprofeno pôde penetrar na região hidrofóbica, o que foi confirmado por espectroscopia de reflexão e absorção no infravermelho com modulação da polarização (PM-IRRAS), indicando assim a presença de interações hidrofóbicas. A análise por microscopia no ângulo de Brewster (BAM) mostrou que o ibuprofeno impede a formação de grandes domínios de DPPC, enquanto que não foram observadas alterações significativas para o DPPG. A interação entre o ibuprofeno e o DPPG também foi confirmada após a imobilização da monocamada mista em filmes LB com alterações na absorção no UV-Vis da molécula de ibuprofeno. No que diz respeito às implicações biológicas, a ação farmacológica que depende diretamente da interação com a membrana deve ocorrer primeiramente em regiões neutras via penetração do ibuprofeno na região hidrofóbica da membrana celular. A segunda parte deste trabalho foi dedicada à interação de capsaicinóides extraídos da pimenta malagueta com monocamadas de Langmuir constituídas de DPPG e DPPC. A capsaicina é um potente analgésico de uso tópico, que pode causar dessensibilização no local de aplicação dependendo da dose e, portanto há interesse na sua incorporação em sistemas de liberação controlada, como os lipossomos. A técnica de Langmuir foi empregada para verificar essa possibilidade. Os capsaicinóides expandiram as monocamadas de DPPG e aumentaram sua elasticidade. As isotermas de potencial de superfície indicaram que os capsaicinóides provocam aumento de 10% no momento de dipolo numa concentração de 30% em mol. Para os filmes mistos de DPPC e capsaicinóides, a área mínima diminuiu e a elasticidade da monocamada aumentou. De acordo com as isotermas de potencial, os momentos de dipolo diminuíram para as monocamadas de DPPC independentemente da concentração de capsaicinóides. Esses resultados sugerem que as moléculas de DPPC são solubilizadas para a subfase na presença do fármaco. A partir destes resultados, conclui-se que os capsaicinóides podem ser incorporados em estruturas lipídicas, constituídas principalmente de DPPG, o que é relevante para uso em sistemas de liberação de fármacos. / Ibuprofen is a nonsteroidal anti-inflammatory drug, with low solubility in water, which exhibits side effects including gastric and intestinal injury, often irreversible. Some of these effects may depend on the interaction with the cell membrane, which motivated us to investigate the incorporation of ibuprofen in Langmuir monolayers as cell membrane models, in the first part of this thesis. Dipalmitoyl phosphatidyl choline (DPPC) or dipalmitoyl phosphatidyl glycerol (DPPG) monolayers co-spread with ibuprofen or deposited on ibuprofen-containing aqueous subphases were studied using surface pressure and surface potential isotherms. Significant effects were observed for DPPC monolayers, particularly at the liquid-expanded to liquid-condensed phase transition, with relevant changes in the elasticity of the monolayer. These effects increased with the ibuprofen concentration. For both types of phospholipids, ibuprofen could penetrate into the hydrophobic part of the monolayer, which was confirmed with polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS), thus indicating the presence of hydrophobic interactions. BAM images showed that ibuprofen prevents the formation of large domains of DPPC, while no significant changes were observed for DPPG. The interaction between DPPG-ibuprofen was also confirmed for deposited layers in the form of LB films, with changes in the ibuprofen UV-Vis absorption. As for the biological implications, the pharmacological action depending directly on the membrane interaction should occur primarily with zwitterionic regions of the membrane via penetration of ibuprofen in the hydrophobic part of the monolayer. The second part of this thesis is dedicated to the interaction of capsaicinoids, extracted from malagueta pepper, with Langmuir monolayers of DPPC and DPPG. Capsaicin is a powerful analgesic of topical use, which can cause desensitization in the application site depending on the dose, and therefore there is interest in its incorporation in drug delivery systems, such as liposomes. The Langmuir technique was employed to verify this possibility. The capsaicinoids expanded the DPPG monolayer and increased its elasticity. Surface potential isotherms indicated that the capsaicinoids increased the average dipole moment by 10 % for 30 mol % of capsaicinoids. For the mixed films of DPPC and capsaicinoids, the minimum area decreased and the elasticity increased. According to the surface potential isotherms, the dipole moments decreased for DPPC monolayers regardless of the capsaicinoid concentrations. These results suggest that the DPPC molecules are solubilized into the subphase in the presence of the drug. From these results, it is concluded that the capsaicinoids can be incorporated into structures as the liposomes constituted mainly of DPPG, which is relevant for use in drug delivery systems.

O estudo da interação de moléculas orgânicas com a água é de grande importância para diversas áreas das ciências, como por exemplo, na compreensão de sistemas biológicos. Uma elevada diversidade de estudos nesta área já foram realizados, no entanto, alguns aspectos do estudo de filmes finos de aminas anfifílicas não foram satisfatoriamente esclarecidos. Este trabalho aborda filmes de Langmuir de octadecilamina em função do pH e da força iônica de subfases aquosas, investigando a estrutura da monocamada orgânica e sua interação com a água, em diversos graus de ionização. Foi utilizada a Espectroscopia Vibracional por Geração de Soma de Frequências (Sum Frequency Generation - SFG), uma técnica que utiliza o princípio da óptica não-linear para estudar superfícies e interfaces, com o objetivo de investigar a conformação molecular dos filmes de Langmuir de aminas anfifílicas de cadeias longas, seu grau de ionização e a estrutura da água interagindo com a monocamada orgânica. Pôde-se acompanhar a ionização da monocamada em função do pH da subfase pela intensidade do modo de estiramento do grupo NH3+, e verificou-se que ela depende do pH local da interface, que por sua vez é alterado pela ionização da monocamada e é sensível à força iônica da subfase. A água interage com a monocamada via ligações de H com os grupos NH2/NH3+ e forma uma camada com alto grau de orientação perpendicular à superfície, mas relativamente desordenada do ponto de vista da rede de ligações de H (como na água líquida). A conformação das cadeias alquila depende da ionização do filme de Langmuir, sendo bastante compacta e ordenada a altos pHs (filme neutro) e mais expandida e desordenada a baixos pHs. Além disso, foram fabricados filmes de Langmuir-Blodgett (LB) de octadecilamina para comparar a estrutura dos filmes na superfície da água e em substratos sólidos. / The study of the interaction of organic molecules with water is of great importance for many areas of science, such as the understanding of biological systems. A large diversity of studies in this area have been performed, however some aspects of the study of thin films of amphiphilic amines have not been satisfactorily clarified. This dissertation focuses on Langmuir films of Octadecylamine as a function of pH and ionic strength of the aqueous subphase, investigating the structure of the organic monolayer and its interaction with water at various degrees of ionization. Sum-Frequency Vibrational Spectroscopy (SFG spectroscopy), a non-linear optical technique to study surfaces and interfaces, was used in order to investigate the molecular conformation of the Langmuir films, its degree of ionization and the structure of water interacting with the organic monolayer. The monolayer ionization could be monitored by the intensity of the NH3+ group stretch mode, and it was found that it depends on the local pH at the interface, which in turn is changed by the monolayer ionization and is sensitive to the subphase ionic strength. Water interacts with the monolayer by H bonds with NH2/NH3+ groups, forming a layer with a high degree of orientational order along the surface normal, but relatively disordered regarding the H-bonding network (like in the bulk liquid). The conformation of the alkyl chains depends on the monolayer ionization, being compact and all-trans at high pHs, but more expanded and disordered at low pHs. In addition, Langmuir-Blodgett (LB) films of Octadecylamine were fabricated to compare their structure on the solid substrate to those on the water surface