Made available in DSpace on 2014-07-29T15:12:48Z (GMT). No. of bitstreams: 1 Dissertacao - Wesley Renato Viali.pdf: 3929545 bytes, checksum: b4e900de56f7f751a05beaeb0b326f0b (MD5) Previous issue date: 2009-03-18 / In this work maghemite nanoparticles were synthesized by co-precipitating Fe2+ and Fe3+ in alkaline medium. The freshly prepared magnetite was oxidized by two different routes: by bubbling O2 through an acid suspension of magnetite nanoparticles or by hydrothermal treatment of magnetite with ferric nitrate/nitric acid solution. After oxidation, the nanoparticles were covered with oleic acid and the grafting coefficient was evaluated according to the oxidation method and pH employed in the adsorption experiments. The covered particles were dispersed in mineral and vegetable insulating oils, resulting in magnetic fluids whose colloidal stability was investigated in regard to the following parameters: dilution, heating, water content, chemical composition of insulating oils and addition of oleic acid. The nanoparticles were characterized by X-ray diffratometry, thermogravimetric and differential thermal analyses and infrared spectroscopy. The Fe3+/Fe2+ molar ratio in the solids was determined by UV-Vis spectrometry. The magnetic fluids were characterized regarding the volume fraction of nanoparticles, the water content, and their colloidal stability was monitored by dynamic light scattering measurements and visual observations. The results obtained from the analyses indicated that the oxidized magnetite nanoparticles (5 to 10 nm) have inverse spinel structure and Fe3+/Fe2+ molar ratio varied from 20 to 70, depending on the oxidation method and employed conditions. The grafting coefficient varied from 2,1 to 4,8, reflecting the oleic acid adsorption conditions, and the further treatments. It was also observed that by controlling the experimental conditions, a grafting coefficient very close to a Langmuir s monolayer can be achieved, together with a decrease in the amount of physisorbed species which contribute negatively to the colloidal stability. The study of colloidal stability showed that dilution, heating and XVII water content have a great influence in the sedimentation of diluted magnetic fluids, when they are diluted with minerals insulating oils. It was verified that the magnetic fluids diluted with vegetable insulating oil showed very high colloidal stability in the accelerated thermal aging experiment / Neste trabalho foram preparadas nanopartículas de maghemita pelo método da coprecipitação de íons Fe3+ e Fe2+ em meio básico. A magnetita inicialmente obtida foi oxidada utilizando-se gás oxigênio ou solução de nitrato férrico/ácido nítrico. Após a oxidação, as nanopartículas foram revestidas com ácido oleico e a eficiência deste processo foi avaliada em função do método de oxidação e do pH de adsorção. As partículas revestidas foram dispersas em óleos isolantes dos tipos naftênico, parafínico ou vegetal, resultando em fluidos magnéticos cujas estabilidades coloidais foram avaliadas em função dos seguintes parâmetros: diluição, aquecimento, teor de água, natureza do meio dispersante e adição de ácido oleico. As nanopartículas foram caracterizadas por difratometria de raios X, análise termogravimétrica e térmica diferencial e espectroscopia de infravermelho. A razão Fe3+/Fe2+ nos sólidos foi determinada por espectrofotometria no visível. Os fluidos magnéticos foram caracterizados quanto à fração volumétrica de nanopartículas e ao teor de água, e sua estabilidade coloidal foi acompanhada por espalhamento dinâmico de luz e por medidas visuais de turbidez. Os resultados das análises e experimentos indicaram que as nanopartículas de magnetita oxidada (5 a 10 nm) apresentaram estrutura espinélio inverso e razão Fe3+/Fe2+ desde 20 até 70, dependendo do método e condições de oxidação empregadas. O coeficiente de revestimento variou de 2,1 a 4,8, refletindo tanto as condições utilizadas nos procedimentos de adsorção do ácido oleico, como também os tratamentos subseqüentes. Demonstrou-se que através do controle das condições experimentais pode-se obter um revestimento muito próximo ao de uma monocamada de Langmuir, bem como reduzir a quantidade de XV espécies moleculares fisissorvidas que contribuem negativamente para a estabilidade coloidal. O estudo da estabilidade coloidal mostrou que a diluição, o aquecimento e o teor de água favorecem a sedimentação dos fluidos magnéticos quando estes são diluídos em óleos isolantes naftênico e parafínico. Verificou-se que os fluidos magnéticos em óleo vegetal quando em frações volumétricas baixas apresentaram alta estabilidade coloidal em experimentos de envelhecimento acelerado

The design and engineering of nanoscopic drug delivery vehicles that stably encapsulate lipophilic drug molecules, transport their loaded cargo to specific target sites, and release their payload in a controlled manner are of great interest in therapeutic applications, especially for cancer chemotherapy. This dissertation focuses on chemically cross-linked, water-soluble polymer nanoparticles, termed nanogels, which constitute a promising scaffold and offer the potential to circumvent encapsulation stability issues. A facile synthetic method for a new class of self-cross-linked polymer nanogels, synthesized by an intra/intermolecular disulfide cross-linking reaction in aqueous media, is described here. This simple emulsion-free method affords noncovalent lipophilic guest encapsulation and surface functionalization that may allow for targeted delivery. The encapsulation stability of lipophilic molecules sequestered within these nanoscopic containers is evaluated by a fluorescent resonance energy transfer (FRET) based method developed by our research group. We demonstrate that the encapsulation stability of noncovalently encapsulated guest molecules in disulfide cross-linked polymer nanogels can be tuned and that guest release can be achieved in response to a biologically relevant stimulus (GSH). In addition, varied hydrophobicity in the self-cross-linked nanogels affects the lipophilic loading capacity and encapsulation stability. We reveal that optimal loading capacity is limited by encapsulation stability, where over-loading of lipophilic molecules in the nanoscopic containers may cause undersirable leakage and severely compromise the viability of such systems for drug delivery and other biological applications.

Le développement de nanomédicaments capables de contrecarrer les propriétés biopharmaceutiques défavorables de l'amphotéricine B (AmB) représente un enjeu important. L'AmB est en effet une molécule très efficace pour le traitement des infections fongiques systémiques et aussi pour la leishmaniose, mais difficile à formuler efficacement, quelle que soit la voie d'administration. Cette molécule particulièrement hydrophobe souffre de limitations importantes en raison de sa tendance prononcée à l'agrégation dans les conditions physiologiques. La première partie de cette thèse a consisté à vérifier l'hypothèse selon laquelle le degré d'agrégation de l'AmB pourrait avoir un fort impact sur certaines de ses propriétés biopharmaceutiques et pharmacocinétiques. Dans cet objectif, l'albumine a été utilisée pour produire avec l'AmB des complexes de taille contrôlée. Les caractéristiques morphologiques des objets colloïdaux formés ont été déterminées par spectroscopie UV-Vis et par dichroïsme circulaire. Ainsi, l'impact de l'état de l'agrégation sur la perméabilité intestinale d'une part et la reconnaissance éventuellement des agrégats par le système immunitaire d'autre part a été étudié. La deuxième partie de ce travail a été axée sur le développement des micro et nanotransporteurs destinés à surmonter la barrière d'absorption élevée contre AmB après son administration orale. À cet effet, des micro- et nano-émulsions chargées en AmB ont été préparée afin d'estimer leur capacité à améliorer la perméabilité de l'AmB au travers de l'épithélium digestif de rat. Le modèle de la Chambre d’Ussing a été utilisé à cet effet. Aucun passage de l’AmB n'a pu être détecté dans chacune des conditions expérimentales testées. Toutefois, les données électrophysiologiques ont montré une diminution de la viabilité des tissus, attribuable à la grande toxicité de l'AmB, et dépendante de l'état d'agrégation de l'AmB lorsque ces objets sont au contact avec le tissu. Ces essais de perméation menés sur des tissus sains au niveau jéjunal suggèrent que le transport de l'AmB par les voies paracellulaire et/ou transcellulaire est sans doute marginal. Cependant, la littérature rapporte que qu'une absorption par voie orale de l'AmB, bien que peu importante, peut être observée in vivo. Ceci suggère donc que d'autres voies d'absorption pourraient être mises en oeuvre, parmi lesquelles la capture d'agrégats d'AmB au niveau des plaques de Peyer et l'accès à la voie lymphatique pourraient représenter des voies d'absorption alternatives. Enfin, l'emploi d'un autre système de transporteur conçu pour atteindre le colon et assurer la délivrance colonique grâce à l'action enzymatique bactérienne locale a été envisagé. Dans cet objectif, un biopolymère naturel et dégradable par des enzymes, le xylane, a été sélectionné. Le xylane est un polysaccharide présent dans les grains, de céréales et de plantes angiospermes qui est spécifiquement dégradé dans la région du côlon, grâce à l'action enzymatique du microbiote. Pour cela, un procédé original de préparation de particules a été mis en oeuvre consistant tout d'abord à produire une émulsion eau-dans-eau de xylane en présence de PEG, suivie d'une phase de réticulation du xylane au moyen du trimétaphosphate de trisodium. La méthode a permis la production de nano et de microparticules allant de 380 nm à 4,5 μm et les paramètres contrôlant le processus ont été identifiés. Ce processus, respectueux de l'environnement et ne nécessitant pas l'emploi de solvants organiques, pourrait être appliqué à la délivrance colonique de AmB. / This thesis is part of the development and evaluation of nanomedicines potentially able to overcome unfavorable biopharmaceutical properties of amphotericin B (AmB), a highly effective molecule used for the treatment of systemic fungal infections and leishmaniasis, but difficult to formulate efficiently, whatever the route of delivery. It is believed that this hydrophobic molecule suffers from severe limitations due to its prounounced tendency to aggregate under physiological conditions. The first part of the thesis was driven on the hypothesis that the degree of aggregation of AmB could have a strong impact on some of its pharmacokinetics properties. For this purpose albumin has been used to produce controlled complexes between albumin and AmB in order to control AmB aggregation states. The morphological characteristics of the resulting colloidal objects have been carefully characterized by UV-Vis spectroscopy and circular dichroism. Furthermore, the impact of aggregation state on both the intestinal permeability and a possibly expected recognition of the aggregates by the immunological system were investigated. The second part of this work was focused on the development of micro- and nanocarriers intended to overcome the absorption barrier raised against AmB after oral delivery. For this purpose, AmB was loaded into micro- and nanoemulsions to evaluate a possible permeability enhancement effect through the intestinal membrane, which was evaluated in ratas using the Ussing chamber model. No detectable permeation was seen in any of the experimental conditions. However, the electrophysiological data showed tissue viability losses due to the strong toxicity of AmB, that were dependent on the agregation state of AmB when in contact with the tissue. It was also concluded from detailed permeation experiments in healthy tissues that paracellular and transcellular routes were likely to be only marginal pathways when oral absorption are observed in vivo, as reported in the literature. The likeness of other possible absorption pathways, including Peyer's patches capture and lymphatic pathway implication for agregated particles has been discussed. Finally, another particulate system intended for colonic delivery and based on xylan, a natural and enzymatically degradable biopolymer, has been investigated. Xylan is a polysaccharide present in grains, cereals and angiosperm plants that is specifically degraded on colon region, by the microbiota. An original process consisting in a water-in-water emulsion of xylan in presence of PEG followed by a crosslinking phase using trisodium trimetaphosphate has been developed, making possible the production of xylan-based biocompatible micro- and nanospheres ranging from 380 nm to 4.5 μm, depending on the parameters in the process. This eco-friendly process is free of harmful solvents and has potential application for the delivery of AmB at the colonic level.