Dissertations / Theses on the topic 'Nanodispersion'

L'étude de la dispersion de matériaux inorganiques dans des médias organiques et aqueux est un facteur déterminant pour leur mise en œuvre industrielle. Ces matériaux trouvent leur application dans de nombreux domaines allant de l'industrie pétrolière à celle du polissage. Nous nous sommes intéressés à la stabilité de ces matériaux dans des médias tels que : (i) les solvants aqueux utilisés pour la formulation de suspensions abrasives, (ii) les eaux d'exploitations pétrolières et (iii) le pétrole brut et ses dérivés. Ceci dans le but d'améliorer les propriétés de luminescence de ces matériaux pour l'industrie pétrolière et les propriétés abrasives pour l'industrie du polissageNous avons ainsi développé des formulations stables de complexes de lanthanides (Ln) dérivés du DOTA et de nanoparticules (NPs) de sulfure de zinc dopées au manganèse, dans le but de tracer les eaux d'injections pétrolières. Ces matériaux ont été détecté par fluorescence en temps-résolu (TRF) et ce afin de contourner les problématiques liées à l'auto-fluorescence des produits pétroliers présents dans les eaux d'exploitation. Nous avons également étudié la dispersion de composés spécifiques dans des carburants, dans le but de proposer des nouveaux marqueurs anticontrefaçon pour le pétrole brut et ses dérivés. Enfin, nous avons cherché à améliorer la dispersion de nanoparticules de diamant dans des médias aqueux et ce afin d'optimiser l'efficacité des suspensions abrasives à base diamantées, utilisées pour le polissage du saphir plan a
The dispersion of inorganic materials in organic and aqueous media is a decisive factor for their industrial implementation. These materials are used in many areas from the oil industry to polishing. Herein, we focus on studying the stability of these materials in different media such as: (i) the aqueous solvents used for the formulation of abrasive slurries, (ii) the waters of oil operations and (iii) crude oil and its derivatives. Our objective is to improve the luminescence properties of these materials for the oil industry and the abrasive properties for polishing applications.We have developed stable formulations of lanthanide complexes (Ln) derived from DOTA and nanoparticles (NPs) of zinc sulfide doped with manganese for water-management in oil fields. These materials were characterized by time-resolved fluorescence (TRF) to solve the problems associated with the auto-fluorescence of petroleum products in the operating water.Furthermore, we studied the dispersion of specific compounds in different fuels in order to develop new anti-counterfeiting markers for crude oil and its derivatives. Finally, we tried to improve the dispersion of diamond nanoparticles in aqueous media in order to enhance the efficiency of diamond abrasive suspensions based for polishing sapphire a-plan

A bixina é o principal carotenoide encontrado na superfície externa das sementes de Bixa orellana L., conhecida popularmente como urucum. Os extratos das sementes são largamente utilizados, tradicionalmente como condimento e no preparo de \"remédios\" caseiros para diversos tipos de doenças e sintomas. Industrialmente, é empregado em formulações farmacêuticas, cosméticos e alimentos como corante de origem natural. Contudo, o uso da bixina ainda é limitado pela sua baixa solubilidade em água. Foi observado que a bixina pode ser dispersa em água, por meio de técnica já patenteada pelo nosso grupo, sem uso de suportes e adjuvantes. O presente projeto teve por objetivo obter a bixina purificada de um extrato comercial de semente de urucum, preparar e caracterizar a dispersão de bixina em água e aplicá-la como um carreador de fármacos, utilizando daunorrubicina como modelo e que é utilizada no tratamento de tumores. Bixina foi obtida com 95% de pureza, a partir do extrato de semente de urucum e dispersões a partir deste composto foram preparadas em soluções aquosas e estabilizadas em cerca de duas horas. Foi determinado que a dispersão em água é constituída por partículas esféricas, com diâmetro médio variando de 20 a 150 nm e potencial Zeta de -24,7 mV. A dispersão manteve-se estável quando submetida a concentrações de NaCl de até 50 mmol/L e resistente em pH ácido. Porém, a partir de pH=10, ocorreu hidrólise do éster metílico da bixina, convertendo-a em norbixina. As partículas de bixina foram capazes de incorporar o fármaco daunorrubicina, em proporção molar máxima de bixina/daunorrubicina de 2:1. Nas concentrações testadas, a incorporação da daunorrubicina à dispersão de bixina causou aumento na atividade antiproliferativa, sendo até 60% mais ativa do que a daunorrubicina livre, na concentração de 1 µg/mL. A dispersão de bixina apresentou tolerância a variações em pH e concentração salina e capacidade de incorporar e aumentar a atividade do fármaco daunorrubicina. Sendo assim, constitui-se em um potencial sistema carreador de fármaco derivado de um produto natural.
Bixin is a carotenoid found on surface of Bixa orellana L. seeds, known as annatto. Annatto seeds extracts are popularly used as a condiment for foods and remedy for several diseases and symptoms. It is also applied in pharmaceutical products, cosmetics and food industry as a natural colorant. Nevertheless, the use of bixin is still limited by its poor solubility in water. Our research group has developed and patented a technique to disperse bixin in water, with no use of stabilizers or scaffolds. In this project, we aimed to obtain purified bixin from commercial annatto seeds extracts, prepare and characterize a dispersion of bixin in water and to employ it as a drug delivery system. Daunorubicin was chosen as a drug model to be delivered by bixin dispersion. Bixin was isolated from extracts in 95% purity. Aqueous dispersions of bixin demonstrated to be stable two hours after preparation. Bixin formed spherical particles with mean diameter ranging from 20 to 150 nm and Zeta potential of -24,7 mV. The dispersion was stable in NaCl solution up to 50 mMol/L and resistant to acidic medium. However, above pH=10, hydrolysis of ester termination begins to occur, converting bixin into to norbixin and since norbixin is water-soluble, the particles dissolved in water. Bixin dispersion was able to incorporate daunorubicin, in a bixin:daunorubicin molar proportion of 2:1. In all tested concentrations, daunorubicin delivered by bixin showed higher antiproliferative activity compared to free drug, reaching 60% more acitivity at 1 µg/mL In conclusion, bixin dispersion showed good stability in large range of pH and salt concentrations, ability to incorporate daunorubicin and enhanced the antitumoral activity. Thus, it can be considered a potential drug delivery system derived from a natural product.

Orientador: Marlus Chorilli
Resumo: O câncer de próstata (CP) é a segunda neoplasia mais frequente entre homens no Brasil e é caracterizado por não apresentar sintomas em seus estágios iniciais, sendo diagnosticado em seu estágio avançado, o que muitas vezes dificulta o tratamento. Alguns fatores relacionados podem intensificar sua agressividade como, por exemplo, a superexpressão do receptor do fator de crescimento epidérmico (EGFR) em alguns subtipos de tumores de próstata. Neste contexto, a inibição do EGFR auxilia no combate da neoplasia, função essa que pode ser atribuída ao anticorpo monoclonal quimérico IgG1 (cetuximabe-CTX) que se liga à porção externa do EGFR, inibindo a proliferação celular, angiogênese e metástase, além de promover a apoptose. Dentre as formas de tratamento destacam-se a braquiterapia, a radioterapia e a quimioterapia utilizando o docetaxel (DTX), o qual apresenta vantagem de prolongar a sobrevivência em pacientes com CP metastático resistentes à terapia antiandrogênica. No entanto, a formulação comercial (Taxotere®) causa efeitos colaterais, como febre, anemia, retenção de líquidos, hipersensibilidades, mialgias, mucosite, neuropatias periféricas e toxidade a pele e unhas, tornando necessário o estudo de novas formas de veiculação para este fármaco Deste modo, o objetivo deste trabalho foi desenvolver uma nanodispersão de cristal líquido (NCL) de fase cúbica baseada em álcool cetílico etoxilado 20 e propoxilado 5 como tensoativo (T), ácido oleico, DSPE-PEG-MAL e fosfatidilcolina de ... (Resumo completo, clicar acesso eletrônico abaixo)
Mestre

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Keeping in mind the problematic that photoaging and hyperpigmentation represent, the development of a topical formulation containing vitamin C (ascorbyl palmitate) and açai oil associated to nanoparticules may represent a protection of the cellular membrane against oxidation, due to a better permeation and the synergic acting of both active. This study has as its objective the development and the characterization of cream-gel formulations containing ascorbyl palmitate associated to nanocarriers, in the presence or absence of the açai oil as the oily nucleus. Associated to that, the stability of the freeze-dried nanocoated active was tested after the incorporation in a silicon basis. Suspensions containing the AP associated t o the nanocapsules (NCAP), nanoemulsions (NEAP) and nanodispersions (NDAP) were developed, being the latter used as active protection comparative parameter without the active (NCBC). With the purpose of assessing the influence of the water on the stability of the active, AP was incorporated in a methanolic solution (SMAP), to which was put in an aging study during 30 days in environment temperature (22 ºC + 2 ºC), refrigeration (4 ºC + 2 ºC), greenhouse (40 ºC + 2 ºC) and chamber UVC (254 nm), along with the formulations cited previously. The sample characterizations was carried out soon after its obtaining and on days 7, 15, 21 and 30, as to the AP content quantification, average particle diameter, polydispersity index, zeta potential and pH. The NCAP, NEAP, NCBC suspensions were incorporated in semi-solid basis of Hostacerin SAF® cream-gel and later stored for the accelerated aging study (21 days) and alternated cycles of heating-cooling, which were analysed at the beginning and end of this period through the determination of the particle average diameter, polydispersity index, zeta potential, pH, active content quantification, organoleptic characteristic determination, spreadability determination and rheological behavior, and centrifugation test. NCAP and NCBC suspension samples were freeze-dried and incorporated to the silicon base (CSNCAPLIO and CSNBCLIO, respectively), and stored in heating-freezing cycles. In this same basis, the active in the free form (CSAP) was incorporated. Soon after its obtaining, the suspensions presented active content equal to 97,51%±0,93 to NCAP, 80,68% + 1,25 to NEAP and 83,10% + 3,10 to NDAP, and remaining desirable paramaters to nanometric systems. However, in the 30th day of storing, all the physical-chemical characteristics suffered significant alteration, being evident in the active loss, which was detected only in the NCAP and NEAP samples conditioned under refrigeration (24,42 % + 1,0 e 21,37 % + 1,27,, respectively). Contrasting with these results, the SMAP kept the active stable and in effective concentrations during 90 days of storing, revealing the fragility of the AP structure in the presence of an aqueous environment. As to the cream-gel formulations, all presented physical-chemical parameters appropriate for the nanoparticle formulations and slightly acid pH, satisfactory for the structure of the active in the non-dissociated form (smaller than the AP pKa value), homogeneous aspect of the sensorial optimum. For those, a proper spreadability was obtained, considering the function and action local, intended. The formulation rheogram demonstrated to have a non-Newtonian character and pseudoplastic flux, which is desired in pharmaceutical formulations, for the initial resistance for the semi-solid formulation to flow diminished, reflecting the application easiness. The greatest active concentration for the samples containing the nanocoated active was obtained, suggesting a greater initial protection of the active in the presence of polymer film and the açai oil. However, at the end of the 21-day period, it was verified the destabilization of the formulation containing the active by the total freeze-drying of those, not being possible to carry out the final tests. The cream in silicone basis samples were analyzed only according to the active content soon after the incorporation of the freeze-dried, which demonstrated an average initial loss of 30% under the previous to the freeze-drying quantification, while for the sample containing the free active, the initial content was superior. This result may be justified possibly by the prolonged exposition to humidity and light during the freeze-drying process. After the 7-day storing in ES, the essay was repeated, obtaining for the CSNAPLIO an average of 15 % + 2,03 of AP content, being that for the CSAP the quantification due to the active oxidation total was not possible. These data restate the destabilization of the active faced with the heat, even for the water free formulation. From the obtained results it was possible to verify the AP instability faced with the heat, light and water conditions. However, keeping in mind its therapy potentiality t is considered relevant to the continuity of this research with the purpose of searching for more ascorbyl palmitate stability keeping it viable for topical application. It is suggested the incorporation to the drying of the suspensions for nano-spry-drying and the incorporation of those in different semi-solid basis, with emulsions, silicones and non-ionic bases for the long-term stability study, searching for evidences which take to definite conclusions on the product stability.
Tendo-se em vista a problemática que o fotoenvelhecimento e a hiperpigmentação representam, o desenvolvimento de uma formulação tópica contendo vitamina C (palmitato de ascorbila) e óleo de açaí associada à nanopartículas pode representar a proteção das membranas celulares contra a oxidação, devido à melhor permeação e a atuação sinérgica de ambos os ativos. Este estudo teve por objetivo o desenvolvimento e caracterização de formulações de creme-gel contendo palmitato de ascorbila associado à nanocarreadores, na presença e ausência do óleo de açaí como núcleo oleoso. Associado a isso, foi testada a estabilidade de liofilizados do ativo nanoencapsulado após incorporação em base de silicone. Foram desenvolvidas suspensões contendo o AP associado à nanocapsulas (NCAP), nanoemulsões (NEAP) e nanodispersões (NDAP), tendo sido esta última utilizada como parâmetro comparativo de proteção do ativo pela total ausência de filme polimérico e núcleo oleoso. Como branco utilizou-se uma suspensão de nanocápsulas sem o ativo (NCBC). A fim de avaliar a influência da água sobre a estabilidade do ativo, incorporou-se AP em uma solução metanólica (SMAP), a qual foi colocada em estudo de envelhecimento durante 30 dias em temperatura ambiente (22 ºC + 2 ºC), refrigeração (4 ºC + 2 ºC), estufa (40 ºC + 2 ºC) e câmara climatizada UVC (254 nm), juntamente com as formulações citadas anteriormente. A caracterização das amostras foi realizada logo após sua obtenção e nos dias 7, 15, 21 e 30, quanto à quantificação do teor de AP, diâmetro médio de partícula, índice de polidispersão, potencial zeta e pH. As suspensões de NCAP, NEAP, NCBC foram incorporadas em bases semissólidas de creme-gel Hostacerin SAF® e posteriormente armazenadas para estudo de envelhecimento acelerado (21 dias) em ciclos alternados de aquecimento – resfriamento, as quais foram analisadas ao início e final deste período através das determinações de diâmetro médio de partícula, índice de polidispersão, potencial zeta, pH, quantificação do teor de ativo, determinação das características organolépticas, determinação da espalhabilidade e comportamento reológico, e teste de centrifugação. Amostras de suspensões de NCAP e NCBC foram liofilizadas e incorporadas em base de silicone (CSNCAPLIO e CSNCBCLIO, respectivamente), e armazenadas em ciclos de aquecimento-resfriamento. Nesta mesma base, foi incorporado o ativo na forma livre (CSAP). Logo após sua obtenção, as suspensões apresentaram teor de ativo iguais a 97,51% + 0,93 para NCAP, 80,68% + 1,25 para NEAP e 83,10% + 3,10 para NDAP, e demais parâmetros desejáveis para sistemas nanométricos. Contudo, no 30o dia de armazenamento, todas as características físico-químicas sofreram significativa alteração, sendo evidente a perda de ativo, o qual foi detectado somente apenas nas amostras de NCAP e NEAP acondicionadas sob refrigeração (24,42 % + 1,0 e 21,37 % + 1,27, respectivamente). Contrastando com estes resultados, a SMAP manteve o ativo estável e em concentrações efetivas durante 90 dias de armazenamento, revelando a fragilidade da estrutura do AP em presença de meio aquoso. Quanto às formulações de creme-gel, todas apresentaram parâmetros físico-químicos adequados para formulações de nanopartículas e pH levemente ácido, satisfatório para a estrutura do ativo na forma não-dissociada (menor que o valor de pKa do AP), aspecto homogêneo de ótimo sensorial. Para as mesmas obteve-se espalhabilidade adequada, considerando a função e local de ação, pretendidos. O reograma das formulações demonstrou haver caráter não-newtoniano e fluxo pseudoplástico, o qual é desejado em formulações farmacêuticas, pois a resistência inicial para a formulação semissólida fluir diminui, refletindo a facilidade de aplicação. Obteve-se a maior concentração de ativo para as amostras contendo o ativo nanoencapsulado, sugerindo a maior proteção inicial do ativo em presença do filme polimérico e do óleo de açaí. Contudo, ao final do período de 21 dias, verificou-se desestabilização das formulações contendo o ativo pela total liquefação das mesmas, não sendo possível a realização dos testes finais. As amostras de creme em base de silicone foram analisadas somente segundo o teor de ativo logo após a incorporação do liofilizado, o qual demonstrou uma perda inicial média de 30 % sob a quantificação anterior à liofilização, enquanto que para a amostra contendo o ativo livre, o teor inicial foi superior. Este resultado pode ser justificado possivelmente pela exposição prolongada à umidade e luz durante o processo de liofilização. Após os primeiros 7 dias de armazenamento em ES, o ensaio foi repetido, obtendo-se para o CSNCAPLIO uma média de 15 % + 2,03 de teor de AP, sendo que para o CSAP não foi possível a quantificação devido a total oxidação do ativo. Estes dados reafirmam a desestabilização do ativo frente ao calor, mesmo para formulação isenta de água. A partir dos resultados obtidos foi possível verificar a instabilidade do AP frente a condições de calor, luz e água. Contudo, tendo em vista sua potencialidade terapêutica considera-se relevante a continuidade desta pesquisa de forma a buscar maior estabilidade do palmitato de ascorbila mantendo-o viável para aplicação tópica. Sugere-se a incorporação a secagem das suspensões por nano-spray-drying e incorporação das mesmas em diferentes bases semissólidas, como emulsões, silicones e bases não-iônicas para estudo de estabilidade de longo prazo, buscando indícios que levam a conclusões definitivas sobre a estabilidade do produto.

Les alliages ODS (Oxides Dispersion Strengthened), sont principalement étudiés pour leur capacité à répondre favorablement au cahier des charges de la fonction de gainage combustible pour les réacteurs nucléaire de type RNR-Na (Réacteur à Neutrons Rapides à caloporteur sodium). Elaborés par métallurgie des poudres, mécanosynthèse puis extrusion, ils affichent des propriétés mécaniques, et notamment en fluage, extrêmement intéressantes. Néanmoins, la voie élaboration utilisée induit une forte anisotropie microstructurale. Cette anisotropie se retrouve au niveau de leurs propriétés mécaniques et conduit à une fragilité dans le sens de sollicitation transverse. Le but de cette thèse est d'étudier l'évolution microstructurale de ces matériaux. Les aciers ODS présentent des microstructures ultra fines en termes de grains, de précipités, et de formation d'amas qui conduisent à de grandes difficultés pour en obtenir la recristallisation. De plus, les microstructures obtenues présentent souvent une recristallisation avec croissance anormale. De telles évolutions demandent des investigations à très fine échelle et ont été relativement peu examinées dans le domaine des alliages ODS. Il faut en effet être capable d'une étude structurale la plus quantitative possible de la microstructure des nanograins, ainsi que de la précipitation afin d'étudier les mécanismes d'interaction précipitation / joints de grain. Ceci n'est possible que par un couplage de différentes méthodes : la microscopie électronique en transmission (en particulier avec l'utilisation des outils récemment développés pour l'étude de la nanotexturation, i.e. ACOM-TEM); la diffusion centrale des neutrons ou des rayons X; et enfin la sonde atomique tomographique, à la fois pour apporter les informations sur la morphologie et la chimie des amas et nanoprécipités mais surtout sur la composition chimique aux joints de grains. A partir de l'identification des mécanismes contrôlant la croissance anormale, une modélisation permettant de prédire son apparition dans la microstructure est confrontée à cette caractérisation microstructurale poussée. Cette modélisation prête une attention particulière à la migration des joints de grains couplée à la diffusion et effets d'ancrage préférentiel des joints triples par les précipités ainsi qu'à l'énergie motrice stockée sous forme de densité de dislocation
Oxide Dispersion Steels (ODS) alloys are mainly studied for their ability to fulfil the technical specifications required for Sodium Fast Reactor (SFR) fuel cladding application. Their processing involves powder metallurgy, mechanical alloying and extrusion. Therefore, despite their interesting mechanical creep properties, the extrusion processing involves a high microstructural anisotropy. These particular feature leads to poor mechanical properties in the transverse direction which are worsen by the occurrence of abnormal grain growth. Unfortunately, since internal pressure increases in the tube with the accumulation of gas fission products, the major stress component is precisely applied in the transverse direction. As a result, the material faces a critical risk of failure and control of the microstructure is a key issue. The aim of this thesis is to study the microstructural evolution of ODS ferritic steels. ODS ferritic steels show ultrafine microstructures in terms both grains and precipitates which made the recrystallization very difficult and allow for abnormal grain growth. To observe such evolutions, fine scale microstructure characterization are necessary. This is only possible by coupling different characterization methods: transmission electronic microscopy (in particular with the new developed tools for nanotexturation studies, i.e. ACOM-TEM); neutron and X-ray small angle scattering; and atomic probe tomography. Based on the mechanisms that lead to and control the abnormal grain growth, a model to predict the occurrence of abnormal grain growth is confronted to the experimental results. This model that takes a particular attention to the dislocation stored energy effect to adequately reproduce the observed characterization results

Pullulan is an exopolysaccharide secreted extracellularly by the black yeast-like fungi Aureobasidium pullulans. Due to an alpha-(1-->6) linked maltotriose repeat unit, which interferes with hydrogen bonding and crystallization, pullulan is completely water soluble unlike cellulose. It has also been tested and shown to possess non-toxic, biodegradable, non-mutagenic and non-carcinogenic properties. Chemical modification of polysaccharides to increased hydrophobicity and increase functionality has shown great promise in drug delivery systems. Particularly in amorphous solid dispersion (ASD) formulations, hydrophobicity increases miscibility with hydrophobic, crystalline drugs and carboxy functionality provides stabilization with drug moieties and well as pH specific release. Successful synthesis of cellulose w-carboxyalkanoates have been reported and showed great promise as ASD polymers based on their ability to retard the recrystallization of the HIV drug ritonavir and antibacterial clarithromycin. However, these cellulose derivatives have limitations due to their limited water solubility. Natural pullulan is water-soluble and modification with w-carboxyalkanoate groups would provide a unique set of derivatives with increased solubility therefore stronger polymer-drug interactions in solution. We have successfully prepared novel pullulan w-carboxyalkanoates, which exhibit good solubility in polar aprotic and polar protic solvents. All derivatives exhibit high thermal stability and most recorded high glass transition temperatures. Due to unknown impact of their three dimensional structure on miscibility and stabilization of drug against crystallization, each of these polymers possesses great potential for use in various drug delivery applications.
Master of Science

Exposure to atmospheric conditions results of deterioration in historic monuments. and their stones. Limestone conservation presents many problems that have to be investigated in detail. In this study, limestone deterioration and development of its conservation treatments were investigated through examination of the statues carved from karstic limestones in Nemrut Dag Monument. The decay mechanisms that had major roles in their deterioration during two thousand years of exposure to atmospheric conditions and the development of their conservation treatments involved several types of analyses that were carried out in the field and in the laboratory. Exposed surfaces of limestones having karstic veins, interior crack surfaces were examined and compared with relatively undeteriorated interior parts. Similar limestones from the geological formations nearby were artificially deteriorated by salt crystallization and were also examined for comparison. Standard physical and physicomechanical tests, petrographical analysis, XRD, SEM-EDX and FTIR were used during those examinations. Swelling nature of clays in limestones and their control were quantified by CEC measurements. The micro structure of limestone was observed to be composed of micritic calcite with karstic veins of sparitic calcite crystals. Some karstic zones were found to be preferred sites of dissolution and precipitation of calcium carbonate where swelling action of clays and widening of cracks occurred. Iron oxides that moved through those zones, as well as biological activity were also found to contribute to those phenomena. Preparation of high concentrations of nanodispersive calcium hydroxide solutions was achieved for the conservation treatments of the deteriorated limestone. Success of treatments with nanodispersive Ca(OH)2 solutions targeted to the decay zones were discussed in terms of their ability to control the swelling action of clays, carbonation of nanodispersive solution, and improvement in the physicomechanical properties of treated limestone.

L’utilisation des nanoparticules Janus pour la délivrance de médicaments suscite un grand intérêt depuis quelques années. Cependant, beaucoup d’aspects sont encore à comprendre dans la préparation de tels systèmes, notamment en présence de principes actifs.Le but de cette thèse était d’étudier des nanoparticules compartimentées Janus (JNP) constituées d’excipients lipidiques, et principalement de glycérides polyoxyéthylénés. Les objectifs étaient, d’une part, une meilleure compréhension de la structure et de la physico-chimie de ces objets puis, d’autre part, leur utilisation contre Helicobacter pylori (H. pylori).Ces JNP sont produites facilement par un procédé d’homogénéisation haute pression qui a permis de fabriquer des lots allant de 10 mL à 1 L. Elles ont été caractérisées par cryo-microscopie, calorimétrie, diffraction des rayons X, diffusion quasi-élastique de la lumière notamment et ont montrées une très grande stabilité physique sur plus d’un an. Les résultats les plus intéressants ont été obtenus avec une formulation contenant 20% de phase lipidique (Labrafil® M1944CS ou Labrafil® M2125CS) et 3% d’un mélange de tensioactifs (Gelucire® 50/13 – Phospholipon® 90G 2:1 en masse). Diverses expériences autour de la formulation et du procédé ont permis d’identifier certains paramètres critiques, notamment la nature de la phase lipidique, l’ordre d’incorporation des excipients ou le taux de diesters de PEG 1500.Des essais d’encapsulation dans ces JNP et de protection d’un principe actif instable en milieu acide, l’érythromycine, ont également été menés. Les nanodispersions formées, physiquement stable pendant au moins 6 mois, avaient une taille de l’ordre de 150 nm et une distribution de taille unimodale. Des tests in vitro ont montré que l’érythromycine encapsulé gardait son efficacité anti-microbienne et était plus stable en milieu acide. Par contre, les nanoparticules ont perdu leur compartimentation en présence d’érythromycineEnfin, des études ont été menées pour comprendre l’influence de l’incorporation de principes actifs (API) sur le procédé. Ainsi, il a été montré que l’ajout de caféine, chloroxylénol, quercétine et tricloasan dans la formulation n’altérait pas la morphologie des JNP contrairement à l’érythromycine et à la dioxybenzone dans une moindre mesure.L’ensemble de ces résultats a montré que les nanoparticules compartimentées Janus sont des systèmes prometteurs de transport et de protection de principes actifs
The use of Janus nanoparticles (JNPs) as constituent of drug delivery formulations has been a topic of considerable interest for the past several years. However, the formation of vesicles and nanoparticle-associated drug/active are still unclear.The aim of the present study was a physicochemical characterization of lipid nanodispersions based on polyoxylglycerides, especially focused on lipid-based JNPs. The experiments should lead to a better understanding of structure and behavior of the very interesting carrier system on Helicobacter pylori (the abbreviation is H. pylori).The multicompartment lipid-based JNPs were produced easily at small (10 to 50 ml/batch) and large scale (1 liter/batch) by hot high pressure homogenization method. Samples were very well long-time stable for at least over 12 months with respect to particle size, DSC, XRD, and special particle morphology characteristics. Especially for particles containing 20% of lipid phase (Labrafil® M1944CS or Labrafil® M2125CS) and surfactant mixture of Gelucire® 50/13 – Phospholipon® 90G (2:1), strong adhesion could be observed in the studies by cryo-TEM technique. By changing the formulation components and process parameters, data showed that the formation of the multicompartment lipid-based JNPs depended on the using of suitable oil phase with the surfactant mixture and the method to produce the vesicles under the identified conditions.We also successfully prepared the stable nanodispersions to protect a labile antibiotic, erythromycin. The mean diameter of the ERY-loaded nanodispersions was found approximately 150 nm, and the size distribution was unimodal. The system was physically stable at room temperature for over six months. The test of antimicrobial activity in vitro on H. pylori showed that not only the preparation process did not reduce the antimicrobial activity of erythromycin, but also the stability of erythromycin was also improved in acidic environment.Furthermore, the properties of APIs loaded into the blank vesicles also affected their particle morphologies. We achieved the nanoparticles like JNPs when loading caffeine, chloroxylenol, quercetin, and triclosan into the vesicles. These results demonstrated that the multicompartment lipid-based JNPs are a promising carrier to protect unstable APIs and enhance their stability and solubility, despite the changes in the structure of the JNPs when incorporating with erythromycin or dioxybenzone

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Mikania glomerata Sprengel é uma planta medicinal que possui a cumarina como marcador químico e devido às propriedades broncodilatadoras é extensamente utilizada na medicina tradicional brasileira. O principal objetivo deste trabalho foi avaliar o potencial do extrato etanólico (EE) e nanodispersão (ND) de M. glomerata contra os efeitos citotóxicos e mutagênicos induzidos pelo quimioterápico ciclofosfamida (CPA). Esta avaliação foi realizada por meio do ensaio do micronúcleo em eritrócitos de medula óssea de camundongos albinos Swiss (Mus musculus) in vivo utilizando o protocolo de tratamento simultâneo. Os tratamentos com EE foram realizados nas concentrações 1,0; 0,5; 0,25; 0,008 e 0,004 mg.kg-1 de cumarina e com a ND nas concentrações 0,008 e 0,004 mg.kg-1 de cumarina. Os resultados obtidos demonstraram que a maioria dos tratamentos reduziu os danos citotóxicos promovidos pela CPA. Apenas o tratamento com EE na concentração 0,004mg.kg-1 de cumarina não apresentou este efeito. Na avaliação da antimutagenicidade, os tratamentos com EE (1,0; 0,5 e 0,25 mg.kg-1 de cumarina) e com a ND (0,008 e 0,004 mg.kg-1 de cumarina) demonstraram-se eficazes na redução da frequência de micronúcleos, apresentando-se como agentes antimutagênicos. Esses resultados também sinalizaram que o tratamento com a ND (0,008 mg.kg-1 de cumarina) apresentou maior efetividade, alcançando 60,38% de redução de danos mutagênicos. De modo geral, os componentes ativos de M. glomerata administrados através das nanodispersões foram mais eficientes na ação quimioprotetora, mesmo contendo concentrações muito inferiores do marcador químico, nas condições experimentais. Estes resultados reforçam a relevância da utilização da nanotecnologia para veiculação de compostos naturais e o potencial da tecnologia de extração de compostos através de nanodispersões.

O processo de interferência de RNA refere-se ao silenciamento pós transcricional seqüência-específico de genes em animais e plantas capaz de ser promovido por dsRNA homólogo à seqüência do gene silenciado. Este processo pode ser aplicado como terapia, que apresenta como vantagens a especificidade pelos alvos escolhidos, a possibilidade de tratar uma enorme gama de doenças genéticas, além do fato de ser muito potente e eficaz. Contudo, o principal desafio consiste em manter a estabilidade dos siRNAs nos fluidos biológicos, visto que estes são bastante susceptíveis à excreção renal e a degradação por RNAses. Com isso, reforça-se a necessidade de sistemas de liberação adequados, que sejam capazes de manter a estabilidade dos siRNAs por tempo suficiente para que atinjam os órgãos alvo da terapia e promover sua liberação sustentada. De particular interesse são determinadas proteínas e peptídeos de transdução (PTDs) que podem ser ligados a fármacos hidrofílicos e assim tornam possível com que estes atravessem membranas. Neste sentido, muitos sistemas carreadores não-virais tem sido estudados para a veiculação de siRNA, sendo de cunho inovador o desenvolvimento de sistemas de liberação nanoestruturados baseados em cristais líquidos funcionalizados com peptídeos de transdução de membrana para a veiculação tópica de siRNAs. Desta forma, nanopartículas de cristais líquidos de fase hexagonal contendo ou não os aditivos catiônicos polietileimina (PEI) e oleilamina (OAM) foram funcionalizadas com peptídeos de transdução de membranas TAT (TAT) ou penetratin (PNT). Os sistemas obtidos foram complexados com siRNA por interação eletrostática e caracterizados através de medidas de tamanho de partícula/ polidispersividade, potencial zeta e eficiência de complexação. A citotoxicidade dos sistemas foi avaliada em fibroblastos L929 pelo ensaio do MTT e por citometria de fluxo e a avaliação da transfecção in vitro foi realizada por citometria de fluxo e por microscopia de fluorescência. Os sistemas contendo PEI ou OAM apresentavam potencial zeta positivo e foram capazes de complexar o siRNA adicionado na concentração de 10 ?M. Os estudos em culturas celulares demonstraram que os sistemas contendo ácido oleico (AO) foram mais eficientes quanto à transfecção em células de fibroblastos L929 e esta eficiência de transfecção foi aumentada com a funcionalização com o peptídeo TAT. A partir daí, os sistemas selecionados foram avaliados quanto a penetração cutânea in vivo. Os sistemas nanodispersos formados por MO/AO/PEI proporcionaram uma maior liberação de siRNA na pele e a eficiência de supressão de TNF-alfa em modelo animal de inflamação cutânea foram maiores que formulações controle. Com isso, demonstrou-se que os sistemas desenvolvidos são promissores para a futura aplicação na terapia gênica tópica de doenças cutâneas inflamatórias.
The RNA interference process refers to the sequence-specific posttranscriptional silencing of genes in animals and plants capable of being promoted by dsRNA that are homologous to the sequence of the silenced gene. This process can be applied as therapy, which presents advantages such as the specificity to the chosen targets, the possibility to treat a variety of genetic diseases, besides being very potent and efficacious. However, the major hurdle consists in keeping the siRNAs stability in the biological fluids, because they are susceptible to renal clearance and degradation by RNAses. Thus, there is the need for suitable delivery systems, capable of maintaining the stability of siRNAs for sufficient time so they can reach the target organ in the therapy and promote sustained release. Of particular interest are certain proteins and peptides transduction domains (PTDs) that can be connected to hydrophilic drugs and thus make it possible to cross cell membranes. Within this context, many non-viral vectors have been studied for siRNA vehiculation which makes innovative the present study because it aims at the development of nanostructured delivery systems based on liquid crystals functionalyzed with membrane transduction peptides for the topical vehiculation of siRNAs. Thus, hexagonal phase liquid crystal nanoparticles containing or not the cationinc polyethylenimine (PEI) and oleylamine (OAM) were functionalyzed with membrane transduction peptides TAT (TAT) or penetratin (PNT). The obtained systems were complexed with siRNA by eletrostatic interaction and characterized for particle size, polidispersity, zeta potential and complexation efficiency. The cytotoxicity of the formulations was performed with L929 fibroblasts by MTT assay and flow cytometry and the in vitro transfection was evaluated by flow cytometry and fluorescence microscopy. The systems containing PEI or OAM presented positive zeta potential and could complex siRNA at the concentration of 10 ?M. The cell culture studies demonstrated that the systems containing oleic acid (OA) were the most efficient to transfect L929 cells and the transfection efficiency was enhanced with the functionalization with the TAT peptide. Thereafter, the selected systems were evaluated for the in vivo skin penetration. The nanosdispersed systems composed of MO/OA/PEI functionalyzed with TAT resulted in a higher siRNA penetration and release in the skin, promoting higher TNF alfa supression in animal model of cutaneous inflammation, compared to the control formulations. Hence, we demonstrated that the developed systems are promising for the treatment of inflammatory skin diseases.

O RNA de interferência (RNAi) é um processo envolvido com o silenciamento pós-transcricional de genes, sendo elucidado por moléculas de RNAs de dupla fita de 21-25 nucleotídeos, os small interfering RNA (siRNA), que ocorre naturalmente em uma ampla variedade de animais, plantas e microrganismos. Esse processo têm demonstrado potencial utilização para o tratamento de doenças em que se observa a super-expressão de genes, pois apresentam diversas vantagens como possibilidade de se utilizar esse mecanismo regulatório apenas pelo conhecimento da sequência do gene terapêutico, menores efeitos tóxicos e alta especificidade. Contudo, o principal desafio consiste no desenvolvimento de vetores seguros e eficazes que permitem a utilização do siRNA como terapia, visto que permitem a proteção do siRNA contra degradação enzimática, apresentam meia-vida prolongada na corrente sanguínea e propiciam um efetivo escape endossomal. Nesse sentido, nanopartículas de cristais líquidos associados ao polímero catiônico polietilenoimina (PEI) foram avaliadas como potenciais vetores não-virais para o siRNA específico para a proteína-1 relacionada à tirosinase (TRP-1), como alternativa para o tratamento tópico do vitiligo. Para isso, os cristais líquidos contendo o PEI foram complexados ao siRNA e avaliados quanto à estrutura líquido cristalina por microscopia de luz polarizada e por difração de raios-X a baixo ângulo (SAXS), tamanho de partícula/polidispersividade, potencial zeta e eficiência de complexação. A citotoxicidade dos sistemas foi avaliada em melanócitos melan-A pelo ensaio do MTT e por citometria de fluxo e, a avaliação da transfecção celular foi realizada por microscopia de fluorescência e por citometria de fluxo. Os diferentes sistemas contendo o polímero PEI apresentaram estruturas líquido cristalinas de fase hexagonal e de fase lamelar pela análise por SAXS, no entanto, a análise sob microscopia de luz polarizada, demonstrou estruturas de fase hexagonal, lamelar e isotrópica. A análise do tamanho de partícula mostrou a presença de sistemas nanoestruturados que foram capazes de se complexar ao siRNA na concentração de 10 ?M. Os estudos em cultura de célula demonstraram a maior viabilidade das células melan-A após tratamento com as nanodispersões líquido cristalinas formadas por monoleína (MO), ácido oleico (AO) e PEI, em relação ao polímero catiônico PEI em sua forma livre. Em relação à transfecção celular por microscopia de fluorescência e por citometria de fluxo foi possível observar a elevada eficiência de transfecção das nanodispersões líquido cristalinas formadas pelo sistema MO:AO:PEI. Resultados de inibição da expressão da proteína TRP-1 foram observadas em células melan-A por Western Blotting, após administração das nanodispersões líquido cristalinas associadas ao siRNA-TRP-1 específico. As nanodispersões liquido cristalinas estudas também proporcionaram maior liberação de siRNA na pele em modelo animal. Esses resultados, demonstram a potencial utilização desses sistemas para a terapia anti-sense de doenças cutâneas como o vitiligo, representando assim, uma importante contribuição para a terapia gênica tópica desta doença
The RNA interference (RNAi) is a process involved with the post-transcriptional gene silencing being elucidated by double-stranded RNA molecules of 21-25 nucleotides, the small interfering RNA (siRNA) that occurs naturally in a wide variety of animals, plants and microorganisms. This process has shown potential use for the treatment of diseases in which there is overexpression of genes, as they offer several advantages such as the possibility of using this regulatory mechanism just by knowing the sequence of the therapeutic gene, lower toxicity and high specificity. However, the main challenge is to develop safe and effective vectors that enable the use of siRNA as a therapy, since they allow the protection of siRNA against enzymatic degradation, have prolonged half-life in the bloodstream and provide an effective endosomal escape. Accordingly, liquid crystal nanoparticles associated with the cationic polymer polyethylenimine (PEI) were evaluated as potential non-viral vectors for specific siRNA for the protein related to tyrosinase-1 (TyRP-1) as an alternative for the topical treatment of vitiligo. For this, the liquid crystals containing PEI were complexed to siRNA and evaluated for liquid crystalline structure by polarized light microscopy and X-ray diffraction (SAXS), particle size / polydispersity index, zeta potential and complexation efficiency. The cytotoxicity of the systems was evaluated by MTT assay and flow cytometry in melan-A melanocytes and the evaluation of cellular uptake was performed by fluorescence microscopy and flow cytometry. The different systems containing the polymer PEI exhibited liquid crystalline structures of hexagonal and lamellar phases by SAXS analysis, however, the analysis under polarized light microscopy showed liquid crystalline structures of hexagonal phase, lamellar and isotropic. The analysis of particle size showed the presence of nanostructured systems that were capable of complexing to the siRNA at concentration of 10 ?M. Studies in cell culture demonstrated a higher viability of melan-A cells after treatment with the liquid crystalline nanodispersions formed by monolein (MO), oleic acid (OA) and PEI in relation to the cationic polymer PEI in its free form. Regarding cellular uptake by fluorescence microscopy and flow cytometry was observed the high efficiency in uptake melan-A cells mediated by liquid crystalline nanodispersions formed by system MO:OA:PEI. Results inhibition of the expression of TyRP-1 protein were observed by Western Blotting in melan-A cells, after administration of liquid crystalline nanodispersions associated with specific siRNA-TyRP-1. The liquid crystalline nanodispersions evaluated also provided greater release of siRNA in the skin in an animal model. These results demonstrate the potential use of these systems for antisense therapy of skin diseases such as vitiligo, thus representing, an important contribution to the topical gene therapy for this disease

A terapia gênica por interferência de RNA (RNAi) trata-se de um processo de silenciamento pós-transcricional capaz de suprimir a expressão de um determinado gene. A RNAi é uma proposta terapêutica promissora para o tratamento de muitas doenças severas que ainda não possuem cura ou terapias bem definidas. Porém, é necessário o desenvolvimento de sistemas de liberação clinicamente adequados, seguros e eficazes para se viabilizar essa nova terapêutica, uma vez que obstáculos na administração e distribuição in vivo comprometem o uso clínico dos siRNAs (small interfering RNA). Paralelamente, a liberação tópica de siRNAs surge como uma alternativa promissora para o tratamento de patologias cutâneas. Neste contexto, a presente pesquisa teve como objetivo o desenvolvimento de um sistema de liberação baseado em nanotecnologia para a liberação tópica de siRNAs, visando introduzir a terapia gênica como nova abordagem para o tratamento de patologias cutâneas. Como sistema de liberação, foram desenvolvidas nanodispersões líquido-cristalinas aquosas, compostas por monoleína (MO), um lipídeo polar biocompatível, associadas ou não com ácido oléico (AO). Foram incorporados a esses sistemas os adjuvantes catiônicos polietilenoimina (PEI) e oleilamina (OAM) para obtenção das nanodispersões. Dentre as nanodispersões aquosas desenvolvidas, foram escolhidas as preparações com as menores concentrações de adjuvantes catiônicos, a saber: MO e OAM a 0,4%, MO e PEI a 0,4%, MO, AO e OAM a 2,5% e MO, AO e PEI a 1,0%. Estas formulações apresentaram: reduzido tamanho médio das partículas, baixa polidispersividade, valores de potencial zeta positivos (característica interessante para interação com as moléculas de siRNA que apresentam carga negativa), baixa citotoxicidade in vitro e foram capazes de complexar o siRNA na concentração final de 2,5 ?M. A análise de difração de raios X caracterizou a fase líquido-cristalina desses sistemas como hexagonal, exceto a nanodispersão MO e PEI a 0,4% que foi caracterizada como uma mistura de fase hexagonal e cúbica. As nanodispersões obtidas foram capazes de aumentar a penetração cutânea de siRNA in vitro. Face aos resultados obtidos, podemos concluir que as formulações desenvolvidas são sistemas de liberação de base nanotecnológica promissores para administração tópica de siRNA para o tratamento de patologias cutâneas na terapia gênica.
Gene therapy by RNA interference (RNAi) is a post-transcriptional silencing process that can suppress the expression of a particular gene. The RNAi is a promising therapeutic approach for the treatment of many severe diseases that have no cure or well-defined treatments. However, the development of clinically appropriate, safe and effective delivery systems is necessary to enable this new therapy, since obstacles in the in vivo administration and distribution committed the clinical use of siRNAs (small interfering RNA). In addition, the topical delivery of siRNAs appears as a promising alternative for the treatment of cutaneous pathologies. In this context, this research aimed to develop a delivery system based on Nanotechnology for the topical delivery of siRNAs, aiming to introduce gene therapy as a new approach for the treatment of skin disorders. As a delivery system, liquid-crystalline nanodispersions, composed by monoolein (MO), a polar biocompatible lipid, associated or not with oleic acid (OA) were developed. The cationic adjuvants polyethylenimine (PEI) and oleylamine (OAM) were incorporated into these systems to obtain the nanodispersions. Among the aqueous nanodispersions developed, preparations with lower concentration of cationic adjuvant were chosen, these consisting of: MO and OAM at 0.4%, MO and PEI at 0.4%, MO, OA and OAM at 2.5% and MO, OA and PEI at 1.0%. These formulations presented: reduced average particle size, low polydispersity, positive values of zeta potential (an interesting feature for interacting with the siRNA molecules that have a negative charge), low cytotoxicity in vitro and they were able to complex the siRNA at a final concentration of 2.5 ?M. The X-ray diffraction analysis characterized the liquid crystalline phase of these systems as hexagonal, except the nanodispersion MO and PEI at 0.4% which was characterized as a mixture of cubic and hexagonal phases. The nanodispersions obtained were able to increase the skin penetration of siRNA in vitro. With the results obtained, we can conclude that the formulations developed are delivery systems based on nanotechnology, promising for topical administration of siRNA for the treatment of cutaneous diseases in gene therapy.

Cette thèse porte sur la mise au point de formulations de nanoparticules lipidiques à base de polyoxylglycérides afin d’assurer la protection de principes actifs instables chimiquement et physiquement, la quercétine (un flavonoïde antioxydant fragile) dans le cas présent. Différents systèmes dispersés ont été préparés par homogénéisation haute pression à chaud avec une taille des particules blanches entre 100 - 200 nm. Ces nanodispersions sont très stables sur plusieurs années à température ambiante. L’encapsulation de la quercétine, dans les nanoparticules lipidiques multicompartimentées et la préparation de nanocristaux ont permis d’augmenter fortement sa teneur dans la dispersion et d’améliorer effectivement sa stabilité physico-chimique
This thesis focuses on the development of polyoxylglycérides-based lipid nanoparticles to protect labile APIs, quercetin (a fragile antioxidant flavonoid) in this case. Different nanoparticulate systems were prepared by high pressure homogenization with particle size between 100 to 200 nm. These nanodispersions are very stable over several years at room temperature. Encapsulation of quercetin in compartmented lipid nanoparticles and preparation of nanocrystals have increased significantly its content in the dispersion and effectively improve its physical and chemical stability

In this study, it was aimed to develop conservation methodologies for the historic sandstones using the case of Nemrut Mount Monument to help their survival in open air conditions. The main conservation approach of this study was holistic as well as aiming at minimum intervention targeted to the problem areas. The most important weathering forms of Nemrut Sandstones were material loss due to loss of scales and granular disintegration as well as detachments by scales, back weathering due to loss of scales, cracking, granular disintegration, rounding/notching and discoloration/biological deposition. Deterioration mechanisms of sandstones were studied on deteriorated and relatively sound sandstones by nondestructive methods of UPV and QIRT, and by microstructural analyses using thin section, XRD and SEM-EDX analyses. In addition, the changes in physical and physcomechanical properties such as, color, bulk density, effective porosity, hydric, hygric and thermal dilatation and CEC of clays were determined. Sandstone deterioration was caused by swelling of clay minerals distributed in their matrix and clay accumulations between the detaching scales. Considerable thermal dilatation characteristics was also an important decay factor. Iron oxides caused discoloration at the surfaces, their phase changes was thought to be important in decay. The use of surfactant DAA, to control clay swelling was found to decrease the hydric dilatation by 40%. The consolidation treatments with nanosilica and silicate dispersions namely Funcosil KSE500STE, SytonX3, KSE300 and KSE100 have improved physicomechanical properties as followed by UPV measurements and decreased hydric dilatation. Their long term behaviour needed to be further investigated.

A Terapia Fotodinâmica (TFD) é uma modalidade de tratamento de câncer relativamente nova e promissora. Baseia-se na utilização de uma substância fotossensibilizante e luz para provocar um dano seletivo ao tecido alvo, sendo que a seletividade ao órgão/tecido deve ser reconhecida como uma das vantagens no tratamento do câncer. Atualmente, o desenvolvimento de sistemas de liberação e promotores de absorção visando a otimização da liberação tópica de fotossensibilizadores, apresenta-se como um promissor e inexplorado campo de pesquisa na TFD do câncer de pele. Sistemas de liberação compostos por monoleína, um lipídeo polar biocompatível, e água são capazes de aumentar a permeação de fármacos na pele, além de serem capazes de controlar a liberação dos mesmos. No presente estudo foram desenvolvidas e caracterizadas nanodispersões de cristais líquidos como sistemas de liberação para os fotossensibilizadores Protoporfirina IX (PpIX) e Ftalocianinas de Zinco (ZnPc) e Cloro Alumínio (ClAlPc), objetivando otimizar a penetração cutânea destes na epiderme. As nanodispersões foram avaliadas em relação a sua eficiência de encapsulação, estabilidade física e química dos fotosensibilizadores veiculados, bem como a permeação e retenção cutânea in vitro e in vivo por microscopia de fluorescência. Estudos de pré-tratamento foram realizados, objetivando-se verificar o efeito promotor de absorção cutânea das nanodispersões desenvolvidas. Os experimentos de eficiência de encapsulação e estabilidade mostraram que as nanodispersões são um sistema de liberação adequado a aplicação tópica de fotosensibilizadores. Os experimentos in vitro mostraram que as nanodispersões aumentaram a penetração cutânea da PpIX e ClAlPc no estrato córneo em, respectivamente, 3,4 e 11,7 vezes, quando comparadas a formulação controle. Na epiderme mais derme sem estrato córneo, os aumentos promovidos pelas nanodispersões foram de 6,5 e 9,7 vezes para a PpIX e ClAlPc. No experimento in vivo as nanodispersões aumentaram a penetração cutânea da PpIX e ClAlPc no estrato córneo em, respectivamente, 13,7 e 7,0 vezes, quando comparadas ao controle, sendo que na epiderme mais derme sem estrato córneo somente as nanodispersões foram capazes de promover a penetração cutânea destes fotossensibilizadores nestas camadas da pele. A visualização da penetração cutânea dos fotosensibilizadores por microscopia de florescência confirmou os resultados obtidos nos experimentos in vivo de que as nanodispersões foram superiores ao controle em aumentar a penetração cutânea da PpIX e ClAlPc em camadas mais profundas da pele. Os resultados obtidos mostraram que as nanodispersões desenvolvidas são sistemas de liberação promissores para a PpIX e ClAlPc no tratamento do câncer de pele na TFD uma vez que aumentaram in vitro e in vivo a penetração cutânea destes na epiderme.
Photodynamic therapy (PDT) is a relatively new and promising cancer treatment modality that involves the administration of a photosensitizing drug and its subsequent activation by light to produce activated oxygen species that selectively destroy target cells. Recently, PDT to treat skin cancer is focused on the development of drug delivery systems and penetration enhancers that aim to optimize the topical release of photosensitizers. Drug delivery systems based on monoolein, a biocompatible polar lipid, and water are able to enhance the cutaneous penetration of drugs and control their release. Liquid crystal nanodispersions were developed and characterized in this project as delivery systems for the photosensitizers Protoporphyrin IX (PpIX) and Zinc and Chloroaluminum phthalocyanines (ZnPc and ClAlPc), aiming to increase their topical penetration in the epidermis. The physical stability of the nanodispersions, their encapsulation efficiency and the chemical stability of the photosensitizers incorporated were tested. In vitro and in vivo skin penetration tests were performed to verify the efficacy of the nanodispersions in enhancing the topical delivery of the photosensitizers. In vitro pre-treatment tests were conducted to determine if the nanodispersions are able to increase the skin penetration of the photosensitizers by a penetration enhancing effect. Stability and encapsulation efficiency tests showed that nanodispersions are an adequate topical delivery system for photosensitizers. In vitro experiments showed increased PpIX and ClAlPc penetration in the stratum corneum, respectively, of 6.5- and 9.7-fold for the nanodispersions compared to the control. In the epidermis with dermis, without stratum corneum, the increase promoted by the nanodispersions for PpIX and ClAlPc were, respectively, of 6.5- and 9.7-fold. Experimental retention in vivo confirmed that when the nanodispersions were used as carrier, PpIX and ClAlPc concentrations in the stratum corneum were about 13.7- and 7.0-fold higher, respectively, than control. In the epidermis with dermis, without stratum corneum, only the nanodispersions were able to promote the skin penetration of these photosensitizers. Visualization of PpIX and ClAlPc skin penetration by fluorescence microscopy confirmed that the nanodispersions increased the skin penetration of these photosensitizers in deeper skin layers. The results showed that the nanodispersions are promising topical delivery systems in the PDT of skin cancer once they increased in vitro and in vivo the topical penetration of PpIX and ClAlPc in the epidermis.

O óxido nítrico (NO) é um versátil agente biológico, atuando em diversas partes do organismo, tais como cérebro, artérias, sistema imunológico, fígado e pulmões. Sua natureza radicalar lhe confere grande reatividade e versatilidade, tornando o entendimento de sua bioquímica um desafio. A molécula de NO tende a reagir rapidamente com alguns metais de transição, formando compostos estáveis denominados complexos nitrosilos, os quais podem ser utilizados como fonte geradora de óxido nítrico. A liberação de NO a partir de complexos nitrosilos pode ocorrer por redução química, eletroquímica e fotoquímica. No presente trabalho foi estudada a obtenção, caracterização e permeação cutânea de um complexo nitrosilo de rutênio, o trans-[RuC(cyclam)(NO)]C2 (cyclam-NO), que associado ao fotossensibilizador protoporfirina IX, possui a peculiaridade de absorver na região do visível, podendo então ser aplicados na terapia fotodinâmica (TFD) para tratamento de câncer de pele. A mistura dos compostos foi incorporada em nanodispersões de cristais líquidos, de fase cúbica (DFC) e de fase hexagonal (DFH), e sua penetração/permeação in vitro em pele de modelo animal foi avaliada, assim como o comportamento fotoquímico do sistema, no que se refere à liberação de NO, visando uma futura aplicação em TFD. A atividade citotóxica dos compostos isolados e em mistura foi avaliada frente às linhagens B16F10 e Melan-A, na ausência e presença de luz, mostrando maior atividade da mistura dos compostos quando irradiados em 630 nm. Foram construídos diagramas de fase binário e ternário e, a partir destes, foram escolhidas as formulações a serem estudadas. Estas formulações foram caracterizadas por microscopia de luz polarizada e difração de raios-X e foram avaliados o tamanho médio de partícula e o índice de polidispersividade das nanodispersões obtidas e sua estabilidade por turbidimetria. Também foi analisada a liberação de oxigênio singlete e de NO a partir dos compostos em solução e destes incorporados nas formulações. Foi desenvolvido e validado um método analítico por cromatografia líquida de alta eficiência para a quantificação simultânea dos compostos nos experimentos. A liberação dos compostos a partir das formulações usando membrana de acetato de celulose também foi avaliada, sendo possível detectar apenas o cyclam-NO. O estudo da eficiência de encapsulação mostrou que cerca de 70% da quantidade adicionada dos compostos foi incorporado na DFC e cerca de 80% na DFH. Os experimentos in vitro de permeação e retenção dos compostos em pele de orelha de porco mostraram aumento significativo da concentração dos compostos, comparado a controles contendo os mesmos em PEG. A DFH promoveu um aumento na concentração de PpIX no estrato córneo (EC) de 2,6 vezes e na epiderme+derme se EC ([E+D]) de 3,4 vezes, e para o cyclam-NO de 2,7 vezes no EC e 2,4 vezes na [E+D]. Já a DFC aumentou em 1,6 vezes a quantidade de PpIX no EC e 1,9 vezes na [E+D] e em 4,6 vezes a quantidade de cyclam-NO no EC e em 2,0 vezes na [E+D]. Os resultados obtidos permitem sugerir que estes sistemas são adequados para utilização como potenciais carreadores para a associação cyclam-NO e PpIX na TFD do câncer de pele e que esta associação apresentou efeito sinérgico, sendo mais eficiente que a utilização de apenas um dos compostos.
Nitric oxide (NO) is a versatile biological agent, acting in several parts of the body such as brain, arteries, immune system, liver and lungs. Its radical nature gives great versatility and reactivity, making the understanding of its biochemical a challenge. The NO molecule tends to react quickly with some transition metals, forming stable compounds called nitrosyl complexes, which can be used as a source of nitric oxide. NO release from nitrosyl complexes can occur by chemical, electrochemical or photochemical reduction. In this work, the acquisition, characterization and permeation of a nitrosyl ruthenium complex, trans-[RuC(cyclam)(NO)]C2 (cyclam-NO), which associated with the photosensitizer protoporphyrin IX, has the peculiarity of absorbing in the visible region, and could then be applied in photodynamic therapy (PDT) for treatment of skin cancer, were studied. The mixture of compounds was incorporated into liquid crystal nanodispersions, cubic (DFC) and hexagonal (DFH) phases, and its penetration/permeation in vitro in an animal model skin was evaluated, as well as the photochemical behavior of the system, with regard to NO release, seeking a future application in PDT. The cytotoxic activity of the compounds alone and in combination was evaluated against the B16F10 and Melan-A cell lines, in the absence and in the presence of light. Binary and ternary phase diagrams were constructed, and from these, the formulations to be studied were chosen. These formulations were characterized by polarized light microscopy and X-ray diffraction and were evaluated for particle size and polydispersity index of nanodispersions obtained and their stability by turbidimetry. Also, the release of singlet oxygen and NO from the compounds in solution and incorporated in the formulations was discussed. An analytical method using high efficiency liquid chromatography was developed and validated for simultaneous quantification of compounds in the experiments. The release of compounds from formulations using cellulose acetate membrane was evaluated, and only the cyclam-NO could be detect. The study of the encapsulation efficiency showed that about 70% of the added amount of the compounds was incorporated in the DFC and approximately 80% in DFH. In vitro permeation and retention experiments of the compounds in pig ear skin were performed, showing a significant increase in the concentration of the compounds in the skin layers, compared to controls containing compounds in polyethylene glycol. The DFH promoted an increase in the concentration of PpIX in the stratum corneum (EC) of 2.6 times and in the epidermis + dermis without EC ([E + D]) of 3.4 times, and the cyclam-NO by 2.7 times for EC and 2.4 times in the [E + D]. DFC already increased by 1.6 times the amount of PpIX in EC and 1.9 times at the [E+D] and 4.6 times the amount of cyclam-NO in EC and 2.0 times in the [E + D]. The results may suggest that these systems are suitable for use as potential carriers for the association of cyclam-NO and PpIX for use in skin cancer PDT and that this association showed a synergistic effect, being more efficient than the use of only one of the compounds.

The aim of this thesis was to develop and test novel food-grade nanodispersions, such as nanoemulsions and solid lipid nanoparticles, for the encapsulation, protection and delivery of bioactive lipophilic food components. Initially, the impact of system composition and homogenization conditions on the formation of nanoemulsions using a high pressure homogenizer (microfluidizer) was examined. The mean particle diameter decreased with increasing homogenization pressure and number of passes, with a linear log-log relationship between mean particle diameter and homogenization pressure. Surfactants emulsifiers formed smaller droplets than protein emulsifiers, which was attributed to their ability to rapidly adsorb to the droplet surfaces during homogenization. At low oil phase-to-aqueous phase viscosity ratios, much smaller mean droplet diameters could be achieved for SDS (d ∼ 60 nm) than for β-lactoglobulin ( d ∼ 150 nm). The effectiveness of various biopolymer emulsifiers at forming and stabilizing model beverage emulsions was examined: β-lactoglobulin (BLG); gum arabic (GA); modified starch (MS). Orange oil-in-water nanoemulsions (5% oil) were prepared using high pressure homogenization. Extensive droplet aggregation occurred in BLG-stabilized nanoemulsions around their isoelectric point, at high salt concentrations, and at high temperatures, due to changes in electrostatic and hydrophobic interactions. There was little effect of pH, ionic strength, and temperature on emulsions stabilized by GA or MS, due to strong steric (rather than electrostatic) stabilization. The potential of utilizing oil-in-water (O/W) nanoemulsions stabilized by a globular protein (β-lactoglobulin) for encapsulating and protecting β-carotene was examined. The influence of temperature, pH, ionic strength, and emulsifier type on the physical and chemical stability of β-carotene enriched nanoemulsions was investigated. The rate of color fading due to β-carotene degradation increased with increasing storage temperature (5 to 55 ºC), decreasing pH, and was largely independent of ionic strength (0 to 500 mM of NaCl). β-carotene degradation was considerably slower in β-lactoglobulin-stabilized nanoemulsions than in Tween 20-stabilized ones. The rate of β-carotene degradation decreased upon addition of additional antioxidants. EDTA was found more effective than ascorbic acid, and Coenzyme Q10 was more effective than vitamin E acetate. The utilization of water-soluble and oil-soluble antioxidants in combination (EDTA and vitamin E acetate) was less effective than using them individually. Solid lipid nanoparticles (SLN) were prepared by homogenizing at a temperature (≈ 80°C) exceeding the melting point of the lipid phase (tripalmitin), and then cooling the resulting oil-in-water nanoemulsions to induce lipid droplet crystallization. Blending tripalmitin with low melting point lipids (either medium chain triglycerides or orange oil) prior to homogenization led to a considerable alteration in the phase behavior and stability of SLN. The presence of the carrier oils reduced the crystallization temperature, melting temperature, and melting enthalpy of tripalmitin. The bioaccessibility of β-carotene encapsulated within nanoemulsion-based delivery systems was examined. A non-ionic surfactant (Tween 20) was used as an emulsifier and long chain triglycerides (LCT), medium chain triglycerides (MCT) or orange oil were used as carrier oils. The bioaccessibility of β-carotene was negligible (≈ 0%) in orange oil nanoemulsions because no mixed micelles were formed to solubilize β-carotene, and was relatively low (≈ 2%) in MCT nanoemulsions because the mixed micelles formed were too small to solubilize β-carotene. In contrast, β-carotene bioaccessibility was relatively high (about 66%) in LCT nanoemulsions because the mixed micelles were large enough to solubilize the bioactive molecule. Overall, our results have important implications for the design of effective delivery systems for encapsulation of carotenoids and other lipophilic bioactive components so that they can be incorporated into functional food and beverage products.