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Trabalho apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas As nanopartículas lipídicas como as nanopartículas de lipídeos sólidos (SLN, Solid lipid nanoparticles) baseadas em emulsões múltiplas são novos sistemas terapêuticos versáteis e inovadores para a incorporação de fármacos hidrófilos. O presente estudo reporta um desenho fatorial 33 com o objetivo de desenvolver uma formulação de SLN ótima. As variáveis independentes analisadas foram as concentrações de lipídeo sólido, agente tensioativo hidrófilo e lipófilo. O tamanho hidrodinâmico de partícula, o índice de polidispersão e o potencial zeta foram as variáveis dependentes. O estudo através do desenho fatorial aplicado, permitiu otimizar uma formulação de SLN que é composta por 1.0%(m/m) de lipídeo sólido, 0.25%(m/m) de agente tensioativo lipófilo e 1.5%(m/m) de agente tensioativo hidrófilo. Com o objetivo de aumentar o potencial zeta das partículas, e consequentemente a sua estabilidade, recorreu-se ao uso do alginato de sódio como polímero de revestimento. Para o efeito foram testadas 4 concentrações diferentes (0.25%, 0.5%, 0.75% e 1.0% (m/m), sugerindo que a concentração ideal testada foi 0.75% (m/m). A influência do pH na fase aquosa interna das SLN também foi testada e os resultados sugerem que pH baixos (i.e., 2-3) influenciam fortemente o tamanho hidrodinâmico das partículas produzidas, aumentando da região nano para micro. Contudo, as SLN podem revelar-se úteis na incorporação de fármacos hidrófilos que requerem pH entre 4-10. As SLN baseadas em múltiplas emulsões são uma abordagem promissora para a incorporação de fármacos hidrófilos, e também para proteínas e péptidos. Lipid nanoparticles, namely solid lipid nanoparticles (SLN) based on multiple emulsions are versatile and innovative carriers for hydrophilic biomolecules. This work reports a 33 full factorial design study to optimize SLN formulations enclosing hydrophilic biomolecules in an inner aqueous phase. The concentrations of solid lipid, lipophilic and hydrophilic emulsifiers were set as the 3 independent variables. Mean particle size, polydispersity index and zeta potential were set as the dependent variables. The selected optimized parameters were set as 1.0%wt of solid lipid, 0.25%wt of lipophilic emulsifier and 1.5%wt of hydrophilic emulsifier. The coating of SLN with sodium alginate was found to improve the PZ of the lipid particles and four different concentrations were evaluated (0.25; 0.5; 0.75 and 1.0% wt). The results suggested that the ideal concentration was 0.75%wt. The influence of low pH (i.e. about 2-3) in the inner aqueous phase was stronger than the higher pH values, contributing for the production of larger droplet sizes. Nevertheless, these systems can be useful for the incorporation of biomolecules requiring a pH ranging between 4 and 10. SLN based on multiple emulsions technology were found to be a promising approach for the incorporation of several hydrophilic drugs, such as proteins and peptides.

Background and the purpose of the study Lamotrigine is a broad spectrum anticonvulsant drug widely used as mono- or adjunct- therapy in adults and children. The aim of this study was to develop controlled release liquid formulation of lamotrigine to improve bioavailability and compliance of pediatric and geriatric epileptic patients. Methods Multiple (w/o/w) emulsion was prepared using one step emulsification technique. It was evaluated for entrapment efficiency (EE), morphology, zeta potential (ZP), polydispersity index (PI), rheology, thermal property, in vitro drug release behavior and stability. In vivo studies in albino mice were carried out using maximal electroshock seizure (MES) test and strychnine induced seizure (SIS) pattern test and results were compared with marketed formulation. Results The EE of the formulations varied from 84.37% to 98.11%. The ZP and PI values of the prepared batches were in the range of +23.46 to +28.07 and 0.256 and 0.365, respectively. Microscopic observation clearly indicated the stability of the emulsions during the storage period. All batches exhibited controlled in vitro drug release up to 12 hrs. Batch C11 exhibited significantly longer duration of protection of seizure in mice against MES and exhibited comparable efficacy in SIS as compared to the marketed formulation. Major Conclusion Multiple emulsion of lamotrigine compared to the marketed tablet showed plasma drug concentration within therapeutic range for longer time and comparable efficacy.

The purpose of this work was to investigate the effect of FGF 2 and TGF-β1 on the early steps of dentin regeneration using microencapsulation of theses factors into a microparticles matrix to ensure growth factors protection and to provide bioactive sustained release in contact with dental pulp cells and then the application of the obtained microparticles in direct pulp capping using a culture model of entire tooth. This work involves the optimization of technical means used to achieve encapsulation of TGFβ1, FGF-2 using the poly (lactic-glycolic acid) PLGA. Physicochemical and colloidal characterization of microspheres shows that the microparticles retain their physicochemical characteristics after drying and re-suspended in water. The double emulsion method was used to separately encapsulate (FGF-2) and (TGFβ1). Microparticles morphology, loading, shelf life, potential toxicity and release kinetics were studied. Then the proliferation of dental pulp cells was examined in contact with microparticles. Biological studies show no toxic effect of particles on pulp fibroblasts. Growth factors have kept their specific biological activity. A culture model of human entire tooth was used to achieve the application of microparticles as a dental direct capping material to confirm their biological activities ex vivo. These microparticles can be useful in studies of early steps of dentin regeneration, activation and migration of progenitor cells in dental pulp; Ce travail a été mené afin d’étudier l’effet du FGF 2 et du TGF-β1 sur les étapes précoces de la régénération dentinaire en utilisant la micro-encapsulation de ces facteurs dans une matrice pour les protéger et contrôler leur libération et ensuite l’application des microparticules obtenues en coiffage pulpaire direct dans un modèle de culture de dents entières. Ce travail consiste d’abord à l’optimisation des moyens techniques mis en oeuvre pour réaliser l'encapsulation du TGFβ1, FGF-2 à l'aide de l'acide poly (lactique-glycolique) PLGA. Les études de la caractérisation colloïdal et physico chimique des microparticules montre que les microparticules gardent leurs caractéristiques physicochimiques après séchage et resuspension dans l’eau. La procèdes optimisé a été ensuite utilisé pour encapsuler les facteurs de croissance. L’encapsulation de FGF-2 et TGF-β1 a été obtenue avec une taille, une efficacité d’encapsulation et une profile de libération adaptés au type d’application choisi. Les études biologiques ne montrent aucun effet toxique des particules sur les fibroblastes pulpaires. Les facteurs de croissance ont gardé leur activité biologique spécifique. Un modèle de culture de dent entier humain a été utilisé pour réaliser l’application de nos microparticules comme un matériau de coiffage dentaire pour confirmer leurs activités biologiques ex-vivo. Ces microparticules peuvent être utiles dans les études des étapes précoces de la régénération dentinaire, l'activation et la migration des cellules progénitrices de la pulpe dentaire

WOS: 000254916000010, PubMed ID: 18166519, The purpose of the study was to see the effect of two antiaging agents in one stable multiple emulsion prepared using natural oil. Vitamin C, which is a very unstable ingredient and is decomposed in the presence of oxygen, active as an antioxidant, was entrapped in the inner aqueous phase of w/o/w multiple emulsion. In this way, slow release can be expected and the effect of vitamin C can be increased since it is protected from the external environment by entrapping it in the internal phase. The other ingredient which is a product of wheat proteins was also used as an antiaging agent in the oily phase. Both of the ingredients increase the synthesis of collagen fibers in the dermis. Therefore, a synergystic effect can be produced by using the two ingredients in one formulation. In this study, multiple emulsions were prepared by the two-step method. Basic formulation containing no active material and a stable formulation containing vitamin C in the internal aqueous phase and wheat protein in the oily phase were prepared. The oil used was macadamia nut oil since it contains a high quantity of palmitoleic acid which is the natural ingredient of the young skin. Basic formulation as well as the active formulation after confirming their stabilities, were applied to the cheeks of 11 human volunteers for four weeks. Different parameters of the skin were monitored every week to see any effect produced by these emulsions. The data obtained was evaluated statistically. It was found that the active formulation as well as base increased the moisture of the skin as verified by statistical tests. However, there were no significant variations in other parameters like skin sebum, pH, elasticity, melanin and erythema, concerning the two formulations.

新大院博（工）甲第236号

This paper describes the conditions of preparation of poly(lactide-coglycolide) microspheres with a mean diameter lower than 10 μm obtained by a (w1/o)w2 emulsion solvent evaporation method. Different parameters influencing respectively the size of the inner emulsion and the diameter of the microspheres were determined. V3 BRU, which is a specific immunogenic fraction from GP120 of HIV, was encapsulated in those microspheres. The entrapment efficiency was shown to be superior to that of microspheres prepared according to the single emulsion solvent evaporation method. Electron microscopy observations demonstrated the presence within the microspheres of globules corresponding to the w / o initial inner emulsion in which the peptide was dissolved in the aqueous phase. Analysis of the release kinetics was carried out in phosphate buffer (PBS), in artificial gastric and intestinal medium. V3 BRU release in PBS was slow, reaching a plateau at 24 h corresponding to 25% of drug release. In addition, V3 BRU was not released in gastric medium within 4 h whereas under the same time conditions, 60% of the drug was released in the presence of intestinal medium. These results open up interesting prospects for the use of these microspheres as an oral adjuvant for HIV vaccination.

Se ha demostrado que las microesferas biodegradables de ácido poli L-láctico como portadores de fármacos proporcionan una liberación controlada y mantenida del principio activo, disminuyendo los efectos colaterales que estos pueden provocar. Por este motivo, se propuso evaluar la influencia de parámetros como el rendimiento, la eficiencia de encapsulación y el diámetro medio de las partículas, en el proceso de preparación de microesferas cargadas con doxorubicina, por el método de emulsión múltiple y evaporación del solvente. Se comprobó que la concentración de alcohol polivinílico y cloruro de sodio, así como la baja temperatura en la fase acuosa externa contribuyen a lograr mejores resultados en las características físico-químicas de las microesferas obtenidas.It has been proved that the biodegradable microspheres of poly l-lactic acid as drug carriers provide a controlled and maintained release of the active principle, reducing the side effects they may cause. For this reason, it was proposed to evaluate the influence of parameters such as performance, encapsulation efficiency and mean diameter of the particles, in the process of preparation of microspheres loaded with doxorubicin by the method of multiple emulsion and evaporation of the solvent. It was proved that the concentration of polyvinyl alcohol and sodium chloride, as well as the low temperature in the external aqueous phase, contribute to attain better results in the physical and chemical characteristics of the obtained microspheres