Delivery of a desired ‘active’ compound (for example, starch (as an energy substrate)) to the gastrointestinal (GI) tract is most easily achieved by oral administration. Unfortunately, the efficacy of most actives is greatly reduced due to the aggressive nature of digestive enzymes and processes which occur in this environment. A commonly applied strategy to prevent deactivation of the active prior to absorption at the target site is to encapsulate the active in another ‘sacrificial’ or non-degradable polymer matrix. Traditionally, the active and matrix is processed into a microparticle format for easy oral delivery (dispersed in a liquid or paste). However, established encapsulation methods which rely on bulk-phase processing to produce these microparticles (e.g. emulsification) are far from ideal as they lack control over the final microparticle size, size distribution, composition and shape. The lack of control in the physical properties of the resultant microparticles in turn results in an inherent lack of control over the kinetics of release of the active at the target site. In contrast, recent advances in microfluidic device fabrication and methodology development have firmly proven that these new generation devices can produce monodisperse droplets and microparticles in a continuous, controllable and predictable manner. Their potential as a processing tool for the production of highly tailored microparticles for targeted delivery, however, remains to be fully explored. Both the physical and chemical (physicochemical) properties of microparticles made from a single polymer system may be altered by the deposition of one or more additional polymer layers onto the microparticle surface (for example, alternating layers of oppositely charged polyelectrolytes to produce core-shell like particles), and this method has proven to be favorable with regards to retarding the release of active compounds. However, this addition of alternate layers of oppositely charged polyelectrolytes (so called Layer-by-Layer (LbL) deposition or assembly) does increase the number of processing steps the particles must undergo prior to storage or delivery. Further, the overall effectiveness of this additional processing is still highly dependent on the properties of the original (core) microparticles. In this thesis, a microfluidic technique was developed to encapsulate starch granules in alginate-based microparticles. Using this continuous technique, the size of the microparticles produced were shown to be monodisperse and reproducible. The developed microfluidic device included a drop formation section, followed by a gelation region and a transfer section, where the particles made on-chip are transferred from the carrier oil phase to an aqueous phase prior to collection. The microparticles collected from this microfluidic device were found to be stable for several weeks and in stark contrast to particles produced via a standard bulk emulsification routes, no aggregation was observed over this time frame. The release profile of glucose (as a result of starch hydrolysation) from microparticles produced using both a standard bulk emulsification method and the developed microfluidic-based method were compared. It was found that the monodisperse particles produced using the microfluidic method showed significantly more retardation to release compared to the glucose release profile from bulk-processed particles. This retardation effect was more pronounced when a thin layer of an oppositely charged polyelectrolyte (chitosan) was adsorbed onto the negatively charged surface (alginate is an anionic polyelectrolyte) of the microfluidic-processed microparticle. The microfluidic device developed within this thesis and the resulting tailored microparticles thus show significant potential with regards to offering a new generation of microparticle delivery systems with highly deterministic delivery over extended lifetimes.

Delivery of a desired ‘active’ compound (for example, starch (as an energy substrate)) to the gastrointestinal (GI) tract is most easily achieved by oral administration. Unfortunately, the efficacy of most actives is greatly reduced due to the aggressive nature of digestive enzymes and processes which occur in this environment. A commonly applied strategy to prevent deactivation of the active prior to absorption at the target site is to encapsulate the active in another ‘sacrificial’ or non-degradable polymer matrix. Traditionally, the active and matrix is processed into a microparticle format for easy oral delivery (dispersed in a liquid or paste). However, established encapsulation methods which rely on bulk-phase processing to produce these microparticles (e.g. emulsification) are far from ideal as they lack control over the final microparticle size, size distribution, composition and shape. The lack of control in the physical properties of the resultant microparticles in turn results in an inherent lack of control over the kinetics of release of the active at the target site. In contrast, recent advances in microfluidic device fabrication and methodology development have firmly proven that these new generation devices can produce monodisperse droplets and microparticles in a continuous, controllable and predictable manner. Their potential as a processing tool for the production of highly tailored microparticles for targeted delivery, however, remains to be fully explored. Both the physical and chemical (physicochemical) properties of microparticles made from a single polymer system may be altered by the deposition of one or more additional polymer layers onto the microparticle surface (for example, alternating layers of oppositely charged polyelectrolytes to produce core-shell like particles), and this method has proven to be favorable with regards to retarding the release of active compounds. However, this addition of alternate layers of oppositely charged polyelectrolytes (so called Layer-by-Layer (LbL) deposition or assembly) does increase the number of processing steps the particles must undergo prior to storage or delivery. Further, the overall effectiveness of this additional processing is still highly dependent on the properties of the original (core) microparticles. In this thesis, a microfluidic technique was developed to encapsulate starch granules in alginate-based microparticles. Using this continuous technique, the size of the microparticles produced were shown to be monodisperse and reproducible. The developed microfluidic device included a drop formation section, followed by a gelation region and a transfer section, where the particles made on-chip are transferred from the carrier oil phase to an aqueous phase prior to collection. The microparticles collected from this microfluidic device were found to be stable for several weeks and in stark contrast to particles produced via a standard bulk emulsification routes, no aggregation was observed over this time frame. The release profile of glucose (as a result of starch hydrolysation) from microparticles produced using both a standard bulk emulsification method and the developed microfluidic-based method were compared. It was found that the monodisperse particles produced using the microfluidic method showed significantly more retardation to release compared to the glucose release profile from bulk-processed particles. This retardation effect was more pronounced when a thin layer of an oppositely charged polyelectrolyte (chitosan) was adsorbed onto the negatively charged surface (alginate is an anionic polyelectrolyte) of the microfluidic-processed microparticle. The microfluidic device developed within this thesis and the resulting tailored microparticles thus show significant potential with regards to offering a new generation of microparticle delivery systems with highly deterministic delivery over extended lifetimes.

Delivery of a desired ‘active’ compound (for example, starch (as an energy substrate)) to the gastrointestinal (GI) tract is most easily achieved by oral administration. Unfortunately, the efficacy of most actives is greatly reduced due to the aggressive nature of digestive enzymes and processes which occur in this environment. A commonly applied strategy to prevent deactivation of the active prior to absorption at the target site is to encapsulate the active in another ‘sacrificial’ or non-degradable polymer matrix. Traditionally, the active and matrix is processed into a microparticle format for easy oral delivery (dispersed in a liquid or paste). However, established encapsulation methods which rely on bulk-phase processing to produce these microparticles (e.g. emulsification) are far from ideal as they lack control over the final microparticle size, size distribution, composition and shape. The lack of control in the physical properties of the resultant microparticles in turn results in an inherent lack of control over the kinetics of release of the active at the target site. In contrast, recent advances in microfluidic device fabrication and methodology development have firmly proven that these new generation devices can produce monodisperse droplets and microparticles in a continuous, controllable and predictable manner. Their potential as a processing tool for the production of highly tailored microparticles for targeted delivery, however, remains to be fully explored. Both the physical and chemical (physicochemical) properties of microparticles made from a single polymer system may be altered by the deposition of one or more additional polymer layers onto the microparticle surface (for example, alternating layers of oppositely charged polyelectrolytes to produce core-shell like particles), and this method has proven to be favorable with regards to retarding the release of active compounds. However, this addition of alternate layers of oppositely charged polyelectrolytes (so called Layer-by-Layer (LbL) deposition or assembly) does increase the number of processing steps the particles must undergo prior to storage or delivery. Further, the overall effectiveness of this additional processing is still highly dependent on the properties of the original (core) microparticles. In this thesis, a microfluidic technique was developed to encapsulate starch granules in alginate-based microparticles. Using this continuous technique, the size of the microparticles produced were shown to be monodisperse and reproducible. The developed microfluidic device included a drop formation section, followed by a gelation region and a transfer section, where the particles made on-chip are transferred from the carrier oil phase to an aqueous phase prior to collection. The microparticles collected from this microfluidic device were found to be stable for several weeks and in stark contrast to particles produced via a standard bulk emulsification routes, no aggregation was observed over this time frame. The release profile of glucose (as a result of starch hydrolysation) from microparticles produced using both a standard bulk emulsification method and the developed microfluidic-based method were compared. It was found that the monodisperse particles produced using the microfluidic method showed significantly more retardation to release compared to the glucose release profile from bulk-processed particles. This retardation effect was more pronounced when a thin layer of an oppositely charged polyelectrolyte (chitosan) was adsorbed onto the negatively charged surface (alginate is an anionic polyelectrolyte) of the microfluidic-processed microparticle. The microfluidic device developed within this thesis and the resulting tailored microparticles thus show significant potential with regards to offering a new generation of microparticle delivery systems with highly deterministic delivery over extended lifetimes.

O mirtilo é uma fruta nativa da América do Norte e ainda pouco conhecida no Brasil. O principal fator de interesse desta fruta é seu elevado potencial antioxidante associado à presença de compostos bioativos, em especial as antocianinas, responsáveis por efeitos benéficos para a saúde humana no tratamento de diversas doenças. O processamento do mirtilo para produção de suco gera em torno de 20% de resíduos (bagaço), o qual contém cerca de 60% das antocianinas totais presentes na fruta fresca. Dentro deste contexto, o presente trabalho tem como objetivo principal avaliar a recuperação das antocianinas presentes no bagaço proveniente da produção de suco de mirtilo, bem como o processo para manter sua estabilidade frente a certas condições ambientais para então servirem como aditivo alimentar. Para isso, a partir do bagaço de mirtilo, foram testadas diferentes condições de extração, utilizando água a 1% de ácido cítrico como solvente, nas temperaturas de 60 e 80 °C e tempos de 5, 15 e 45 minutos. Para extração das antocianinas a condição de 80 ºC durante 5 minutos apresentou o melhor resultado, totalizando 1.944 mg de cianidina-3-glicosídeo/100 g de bagaço em base seca, enquanto para extração de compostos fenólicos totais obteve-se o melhor resultado na temperatura de 80 ºC e 45 minutos analogamente à atividade antioxidante. A etapa subsequente à extração foi a microencapsulação das antocianinas presentes no extrato (80 ºC, 5 minutos) pela técnica de secagem por atomização (spray drying), utilizando como material de parede a maltodextrina e a goma arábica na concentração de 15% m/v. Quanto aos parâmetros de secagem, testaram-se as temperaturas do ar de entrada de 140 e 160 °C. Os pós obtidos foram analisados segundo os aspectos físicos de retenção de antocianinas, morfologia, umidade, higroscopicidade, solubilidade em água e atividade de água. Todas as condições testadas apresentaram um microparticulado com características físicas adequadas para sua estabilidade. Para estudo da degradação das antocianinas quando expostas à radiação ultravioleta (UV) foram feitas análises de concentração de antocianinas e variação de cor durante 41 dias, verificando-se duas cinéticas de degradação. Analisando a cinética mais lenta, o tempo de meia vida das micropartículas de antocianinas ficou entre 7,8 e 14,9 meses, sendo o pó obtido com maltodextrina e temperatura de ar de entrada de 140 ºC o qual se mostrou mais estável à radiação UV. / The blueberry is a native fruit from North America and still little known in Brazil. The main interest factor in this fruit is its high antioxidant potential associated with the presence of bioactive compounds, especially anthocyanins, responsible for beneficial effects on human health in the treatment of several diseases. From blueberry juice process, around 20% of waste (pomace) is generated, which contains about 60% of total anthocyanins present in the fresh fruit. This work aims to evaluate the recovery of anthocyanins present in the waste from blueberry juice production and the process to maintain its stability against some environmental conditions to be used as a replacement to synthetic dyes. The blueberry pomace was subjected to different anthocyanins extraction condition, using water with 1% citric acid as solvent, temperatures of 60 to 80 °C and times of 5, 15 and 45 minutes. The best extracted condition was at 80 °C for 5 minutes, yielding a total of 1,944 mg of anthocyanins extracted from pomace cyanidin-3-glucoside/100 g on dry weight basis, while for the phenolic compounds extraction the best result was obtained in the temperature of 80 ºC and 45 minutes analogously to the antioxidant activity. After the extraction, the subsequent step was the microencapsulation of anthocyanins in the extract with the technique of spray drying using as wall material maltodextrin and arabic gum. Regarding the parameters of drying, the inlet air temperature was 140 to 160 °C. The powders were analyzed according to the physical aspects of anthocyanin retention, morphology, moisture content, hygroscopicity, solubility in water and water activity. All conditions tested showed a microparticle with suitable physical characteristics for its stability. To study the anthocyanins degradation when exposed to ultraviolet irradiation, analysis of anthocyanin content and color variation were done during 41 days. Two degradation kinetics where observed. Analyzing the slower kinetics, the half life of anthocyanins micropaticles was between 7.8 and 14.9 months, being the powder obtained with maltodextrin and 140 °C inlet air temperature the most stable to UV radiation.

La drépanocytose est la première maladie génétique en France et notamment en Guadeloupe. Il s’agit d’une maladie du sang qui est due à une mutation ponctuelle au niveau du gène de la β-globine, laquelle entre dans la composition de l’hémoglobine. Ainsi les drépanocytaires ont une hémoglobine anormale appelée « HbS », contrairement à l’hémoglobine normale « HbA ». En condition de faible oxygénation, l’HbS polymérise et forme des fibres à l’intérieur des érythrocytes. Ces fibres rigidifient et fragilisent les globules rouges. Par conséquent ils peuvent bloquer la circulation à cause de leur faible déformabilité, et causer des crises vaso-occlusives douloureuses, complication caractéristique de la drépanocytose. Ce modèle physiopathologique classique a été complété par les résultats plus récents montrant l’importance du rôle des leucocytes dans l’établissement de ces obstructions. Par ailleurs, les globules rouges des drépanocytaires sont plus prompts à l’hémolyse en raison de leur fragilité. En raison de l’hémolyse exacerbée, l’hémoglobine se retrouve dans le plasma et diminue la biodisponibilité du principal vasodilatateur, le monoxyde d’azote. De plus, les globules rouges rigides et déformés entrainent activation de l’endothélium. Il en résulte dans la drépanocytose, un contexte pro-inflammatoire et pro-adhérent mais aussi pro-coagulant.Ce contexte est propice à l’activation des cellules sanguines et notamment à celle des plaquettes et des érythrocytes qui par bourgeonnement de leur membrane, émettent alors en grandes quantités, des vésicules sub-micrométriques appelées microparticules, ou MP. En l’absence de traitement curatif applicable à tous les patients drépanocytaires, nous avons décidé d’étudier le profil mais aussi le rôle des MP de patients drépanocytaires dans le but de mieux comprendre cette maladie et dans l’espoir de peut-être découvrir une nouvelle piste diagnostique ou thérapeutique.Nous avons donc montré que les patients SC ont des concentrations sanguines en MP plus importantes que les sujets AA, mais moindres que les patients SS. Etonnamment les MP SC, qu’elles soient d’origine érythrocytaire ou plaquettaire, ont aussi plus de phosphatidylsérine (PS) à leur surface que les MP SS. Ce phospholipide étant impliqué dans l’activation de la cascade de la coagulation, il serait intéressant d’évaluer l’intensité de cette activation par des MP SS ou SC. On pourrait aussi comparer ces intensités à celle induite par des MP de patients SS sous hydroxyurée. En effet nous avons aussi montré que 2 ans après avoir commencé ce traitement, les MP érythrocytaires des patients ont une taille plus importante et une exposition de la PS diminuée drastiquement.Les MP étant physiologiquement dans le sang, elles peuvent entrer en contact avec les cellules sanguines mais aussi avec l’endothélium vasculaire. Etant donné l’importance des changements que connaît cet endothélium chez les drépanocytaires (pro-adhérent, pro-inflammatoire et pro-coagulant), nous nous sommes ensuite focalisés sur l’impact des MP de drépanocytaires sur les cellules endothéliales. Ces dernières provenaient de la moelle osseuse humaine, territoire fréquemment affecté par les vaso-occlusions. Il ressort de ces travaux que les MP de patients SS et SC induisent, par rapport aux MP de sujets AA, une surexpression dose-dépendante de gènes impliqués dans l’adhérence (ICAM-1, VCAM-1, E-sélectine), dans l’inflammation (IL-6, IL-1β et CD40-I) et dans la coagulation (TF). Au niveau protéique, ICAM-1 est aussi surexprimé. En effet les MP SS induisent dès 4 heures d’incubation, une augmentation de la densité moyenne d’ICAM-1 membranaire, ainsi qu’une augmentation de la proportion de cellules exprimant cette protéine. ICAM-1 étant impliquée dans l’adhérence des leucocytes à l’endothélium (roulement, adhérence ferme et même transmigration). / Sicle cell disease (SCD) is the first genetic disease in France and more specifically in Guadeloupe. It is a blood disorder due to a point mutation in the β-globin gene. The corresponding peptide chain being a part of hemoglobin, SCD patients have an abnormal hemoglobin called “HbS”, contrary to the normal one, so called “HbA”. In hypoxic conditions, HbS forms polymers inside red blood cells (RBCs), thereby making them rigid but also fragile. Consequently, RBCs can stop blood flow due to their low deformability, and so cause a painful vaso-occlusive crisis, which is a complication characterizing SCD. This pathophysiological model has been modified by recent results showing the involvement of leukocytes in the establishing of these occlusions. Besides, sickle RBCs are more prone to hemolysis owing to their being fragile. Due to this exacerbated hemolysis, hemoglobin is released in the plasma and diminishes the bioavailability of the main vasodilator, nitrite monoxide. Moreover, rigid sickled RBCs entail endothelium activation, which results in a pro-inflammatory, a pro-adhesive and a pro-coagulant context. This latter favors blood cells activation, among which are platelets and erythrocytes that bud high quantities of submicrometric membrane vesicles called microparticles, or MPs. In the absence of curative treatment for all patients, we decided to study the profile but also the role of MPs from SCD patients to better understand this disease and hoping to find a new diagnostic or therapeutic pathway. We showed that SC patients have lower MP levels than SS patients, but higher MP levels than AA subjects. Surprisingly, we have observed that SC MPs, whether they derive from RBCs or platelets (PLTs), have higher densities of exposed phosphatidylserine (PS) than SS MPs. Since this phospholipid is involved in the activation of the coagulation cascade, it would be interesting to evaluate the intensity of this activation by SS or SC MPs. One could also compare these intensities to the one induced by MPs from SS patients under hydrocarbamide. Indeed we also showed that 2 years after the beginning of this treatment, erythrocyte-derived MP are larger and expose PS much less.As MPs are physiologically in the blood, they can interact with blood cells but also with the vascular endothelium. Given the known changes of this endothelium in SCD (pro-adhesive, pro-inflammatory and pro-coagluant), we then focused on the impact of SCD MPs on endothelial cells (ECs). These cells came from human bone marrow, a territory frequently affected by vaso-occlusions. These experiments showed that SS and SC MPs, induce, compared to AA MPs, a dose-dependent overexpression of genes involved in adherence (ICAM-1, VCAM-1, E-selectin), in inflammation (IL-6, IL-1β and CD40-I) and in coagulation (TF). At the protein level, ICAM-1 is also overexpressed. Indeed SS MPs induce within 4 hours of incubation, an increase of the mean membrane density of ICAM-1, but also an increased proportion of cells bearing this protein. As ICAM-1 is involved in leukocytes adherence to the endothelium (rolling, firm adhesion and even transmigration), SS MPs may, by triggering ICAM-1 overexpression at the endothelium surface, allow their adherence to the endothelium, thereby promoting RBC adherence and so the occlusion of the vessel and the occurring of a VOC. It would be interesting to determine which type of MP cause the overexpression of ICAM-1 and to evaluate if it is sufficient to increase in vitro adherence of leukocytes to ECs stimulated with MPs. This would allow to evaluate how important MPs are when considering the fight against sickle cell disease.

Orientador: Anselmo Gomes de Oliveira / Banca: Ana Dóris de Castro / Banca: Victor Hugo Vitorino Sarmento / Resumo: As doenças que afetam o globo ocular de maneira geral têm a terapêutica limitada pela grande dificuldade de se atingir e manter níveis efetivos de fármacos nessa região. Isso porque as formas farmacêuticas convencionais para as doenças oculares são destinadas a aplicação tópica, e não proporcionam níveis terapêuticos no corpo vítreo, retina e coróide. Dessa forma, os sistemas de liberação prolongada de fármacos para aplicação intraocular representam um interesse significativo para a terapêutica em oftalmologia. Sendo assim, o presente trabalho teve como objetivo produzir através da técnica de "Spray Drying" micropartículas biodegradáveis a partir do ácido poli-láctico-co-glicólico (PLGA) contendo cetorolaco de trometamina (CETO), um antiinflamatório não-esteróide que tem apresentado resultados relevantes no tratamento pós-operatório de cirurgias oftálmicas, proporcionando maior conforto e diminuindo os efeitos colaterais causados pelos antiinflamatórios esteróides. Utilizando o método proposto foram produzidas micropartículas com diferentes proporções de CETO:PLGA. As micropartículas foram visualizadas através da Microscopia Eletrônica de Varredura (MEV), apresentando formato esférico e uniforme, com superfícies lisas, e o tamanho médio e distribuição de tamanho das partículas foram determinados através do espalhamento de luz. Somente uma das amostras foi descartada, devido a não formação de microesferas. As propriedades físico-químicas de todos os sistemas foram estudadas utilizando espectroscopia de infravermelho (IR), calometria diferencial exploratória (DSC) e termogravimetria/termogravimetria derivada. Os resultados destes estudos mostraram que após a obtenção as estruturas químicas dos componentes foram preservadas... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The diseases that affect the ocular globe generally have limited treatment due their difficulty to achieve and sustain effective levels of drugs in this region. Because conventional pharmaceutical forms for ocular diseases are destined to topical application, and not provide therapeutic levels in vitreous, retina and choroid. In this way, the ocular drug delivery systems represent a strategy for therapy in ophthalmology. Thus, the present work had as aim to produce through Spray drying technique biodegradable microparticles of poly-lactic co-glycolic acid (PLGA) containing ketorolac tromethamine (CETO), a nonsteroidal anti-inflammatory that has presented relevant results in post-operative treating of ophthalmic surgery, providing greater comfort and reducing the adverse effects caused by steroidal anti-inflammatory. Using the considered method were produced microparticles with different ratios of CETO: PLGA. The microparticles were accessed through Scanning Electronic Microscopy (SEM), presenting itself spherical and uniform, smooth surface, and the particle size analysis and mean diameter were determined by Dynamic Light Scattering. Only one non spherical sample was dismissed. The physicochemical properties of all systems were studied using infrared spectroscopy (IR), differential scanning calorimetry (DSC) and thermogrametry/ derivative thermogravimetry (TG/DTG). The results of these studies showed that after produced the chemical structure of components were preserved, only having the necessaries interactions to promote prolonged released. The amounts of encapsulated CETO were determined by UV-Vis spectrophotometry... (Complete abstract click electronic access below) / Mestre

O mirtilo é uma fruta nativa da América do Norte e ainda pouco conhecida no Brasil. O principal fator de interesse desta fruta é seu elevado potencial antioxidante associado à presença de compostos bioativos, em especial as antocianinas, responsáveis por efeitos benéficos para a saúde humana no tratamento de diversas doenças. O processamento do mirtilo para produção de suco gera em torno de 20% de resíduos (bagaço), o qual contém cerca de 60% das antocianinas totais presentes na fruta fresca. Dentro deste contexto, o presente trabalho tem como objetivo principal avaliar a recuperação das antocianinas presentes no bagaço proveniente da produção de suco de mirtilo, bem como o processo para manter sua estabilidade frente a certas condições ambientais para então servirem como aditivo alimentar. Para isso, a partir do bagaço de mirtilo, foram testadas diferentes condições de extração, utilizando água a 1% de ácido cítrico como solvente, nas temperaturas de 60 e 80 °C e tempos de 5, 15 e 45 minutos. Para extração das antocianinas a condição de 80 ºC durante 5 minutos apresentou o melhor resultado, totalizando 1.944 mg de cianidina-3-glicosídeo/100 g de bagaço em base seca, enquanto para extração de compostos fenólicos totais obteve-se o melhor resultado na temperatura de 80 ºC e 45 minutos analogamente à atividade antioxidante. A etapa subsequente à extração foi a microencapsulação das antocianinas presentes no extrato (80 ºC, 5 minutos) pela técnica de secagem por atomização (spray drying), utilizando como material de parede a maltodextrina e a goma arábica na concentração de 15% m/v. Quanto aos parâmetros de secagem, testaram-se as temperaturas do ar de entrada de 140 e 160 °C. Os pós obtidos foram analisados segundo os aspectos físicos de retenção de antocianinas, morfologia, umidade, higroscopicidade, solubilidade em água e atividade de água. Todas as condições testadas apresentaram um microparticulado com características físicas adequadas para sua estabilidade. Para estudo da degradação das antocianinas quando expostas à radiação ultravioleta (UV) foram feitas análises de concentração de antocianinas e variação de cor durante 41 dias, verificando-se duas cinéticas de degradação. Analisando a cinética mais lenta, o tempo de meia vida das micropartículas de antocianinas ficou entre 7,8 e 14,9 meses, sendo o pó obtido com maltodextrina e temperatura de ar de entrada de 140 ºC o qual se mostrou mais estável à radiação UV. / The blueberry is a native fruit from North America and still little known in Brazil. The main interest factor in this fruit is its high antioxidant potential associated with the presence of bioactive compounds, especially anthocyanins, responsible for beneficial effects on human health in the treatment of several diseases. From blueberry juice process, around 20% of waste (pomace) is generated, which contains about 60% of total anthocyanins present in the fresh fruit. This work aims to evaluate the recovery of anthocyanins present in the waste from blueberry juice production and the process to maintain its stability against some environmental conditions to be used as a replacement to synthetic dyes. The blueberry pomace was subjected to different anthocyanins extraction condition, using water with 1% citric acid as solvent, temperatures of 60 to 80 °C and times of 5, 15 and 45 minutes. The best extracted condition was at 80 °C for 5 minutes, yielding a total of 1,944 mg of anthocyanins extracted from pomace cyanidin-3-glucoside/100 g on dry weight basis, while for the phenolic compounds extraction the best result was obtained in the temperature of 80 ºC and 45 minutes analogously to the antioxidant activity. After the extraction, the subsequent step was the microencapsulation of anthocyanins in the extract with the technique of spray drying using as wall material maltodextrin and arabic gum. Regarding the parameters of drying, the inlet air temperature was 140 to 160 °C. The powders were analyzed according to the physical aspects of anthocyanin retention, morphology, moisture content, hygroscopicity, solubility in water and water activity. All conditions tested showed a microparticle with suitable physical characteristics for its stability. To study the anthocyanins degradation when exposed to ultraviolet irradiation, analysis of anthocyanin content and color variation were done during 41 days. Two degradation kinetics where observed. Analyzing the slower kinetics, the half life of anthocyanins micropaticles was between 7.8 and 14.9 months, being the powder obtained with maltodextrin and 140 °C inlet air temperature the most stable to UV radiation.

O mirtilo é uma fruta nativa da América do Norte e ainda pouco conhecida no Brasil. O principal fator de interesse desta fruta é seu elevado potencial antioxidante associado à presença de compostos bioativos, em especial as antocianinas, responsáveis por efeitos benéficos para a saúde humana no tratamento de diversas doenças. O processamento do mirtilo para produção de suco gera em torno de 20% de resíduos (bagaço), o qual contém cerca de 60% das antocianinas totais presentes na fruta fresca. Dentro deste contexto, o presente trabalho tem como objetivo principal avaliar a recuperação das antocianinas presentes no bagaço proveniente da produção de suco de mirtilo, bem como o processo para manter sua estabilidade frente a certas condições ambientais para então servirem como aditivo alimentar. Para isso, a partir do bagaço de mirtilo, foram testadas diferentes condições de extração, utilizando água a 1% de ácido cítrico como solvente, nas temperaturas de 60 e 80 °C e tempos de 5, 15 e 45 minutos. Para extração das antocianinas a condição de 80 ºC durante 5 minutos apresentou o melhor resultado, totalizando 1.944 mg de cianidina-3-glicosídeo/100 g de bagaço em base seca, enquanto para extração de compostos fenólicos totais obteve-se o melhor resultado na temperatura de 80 ºC e 45 minutos analogamente à atividade antioxidante. A etapa subsequente à extração foi a microencapsulação das antocianinas presentes no extrato (80 ºC, 5 minutos) pela técnica de secagem por atomização (spray drying), utilizando como material de parede a maltodextrina e a goma arábica na concentração de 15% m/v. Quanto aos parâmetros de secagem, testaram-se as temperaturas do ar de entrada de 140 e 160 °C. Os pós obtidos foram analisados segundo os aspectos físicos de retenção de antocianinas, morfologia, umidade, higroscopicidade, solubilidade em água e atividade de água. Todas as condições testadas apresentaram um microparticulado com características físicas adequadas para sua estabilidade. Para estudo da degradação das antocianinas quando expostas à radiação ultravioleta (UV) foram feitas análises de concentração de antocianinas e variação de cor durante 41 dias, verificando-se duas cinéticas de degradação. Analisando a cinética mais lenta, o tempo de meia vida das micropartículas de antocianinas ficou entre 7,8 e 14,9 meses, sendo o pó obtido com maltodextrina e temperatura de ar de entrada de 140 ºC o qual se mostrou mais estável à radiação UV. / The blueberry is a native fruit from North America and still little known in Brazil. The main interest factor in this fruit is its high antioxidant potential associated with the presence of bioactive compounds, especially anthocyanins, responsible for beneficial effects on human health in the treatment of several diseases. From blueberry juice process, around 20% of waste (pomace) is generated, which contains about 60% of total anthocyanins present in the fresh fruit. This work aims to evaluate the recovery of anthocyanins present in the waste from blueberry juice production and the process to maintain its stability against some environmental conditions to be used as a replacement to synthetic dyes. The blueberry pomace was subjected to different anthocyanins extraction condition, using water with 1% citric acid as solvent, temperatures of 60 to 80 °C and times of 5, 15 and 45 minutes. The best extracted condition was at 80 °C for 5 minutes, yielding a total of 1,944 mg of anthocyanins extracted from pomace cyanidin-3-glucoside/100 g on dry weight basis, while for the phenolic compounds extraction the best result was obtained in the temperature of 80 ºC and 45 minutes analogously to the antioxidant activity. After the extraction, the subsequent step was the microencapsulation of anthocyanins in the extract with the technique of spray drying using as wall material maltodextrin and arabic gum. Regarding the parameters of drying, the inlet air temperature was 140 to 160 °C. The powders were analyzed according to the physical aspects of anthocyanin retention, morphology, moisture content, hygroscopicity, solubility in water and water activity. All conditions tested showed a microparticle with suitable physical characteristics for its stability. To study the anthocyanins degradation when exposed to ultraviolet irradiation, analysis of anthocyanin content and color variation were done during 41 days. Two degradation kinetics where observed. Analyzing the slower kinetics, the half life of anthocyanins micropaticles was between 7.8 and 14.9 months, being the powder obtained with maltodextrin and 140 °C inlet air temperature the most stable to UV radiation.

Orientador: Joyce Maria Annicchi-Bizzacchi / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-09T02:14:05Z (GMT). No. of bitstreams: 1 Flores-Nascimento_MarianeCristina_M.pdf: 1784393 bytes, checksum: 7247759965458fc5957d9f5617c9fb7e (MD5) Previous issue date: 2007 / Resumo: A Trombose Venosa Profunda (TVP) é uma doença multicausal, mas muitos fatores de risco ainda não estão definidos. Micropartículas (MPs) são pequenas vesículas liberadas da membrana celular durante ativação e apoptose. MPs podem ser um reflexo da dinâmica entre repouso, ativação e morte celular e podem contribuir com a gravidade da doença pois são procoagulantes e pro-inflamatórias. Parece haver associação entre elevado número de MPs e risco de complicações tromboembólicas, que podem ter um papel na patogênese destas doenças. Neste estudo avaliamos e caracterizamos as MPs em pacientes com TPV de membro inferior, [ao diagnóstico (5M/4H, idade média=41,1 anos), após 6 meses de tratamento (7M/3H, idade média=32,9 anos), associada à Síndrome do Anticorpo Antifosfolípide (7M/3H, idade média=33,8 anos)], em portadores assintomáticos do Fator V de Leiden (FVL) (7M, idade média=34 anos), e comparando-as a controles pareados por sexo, idade e etnia. As MPs foram isoladas de sangue periférico citratado, por centrifugação diferencial. A quantificação e caracterização foram feitas por citometria de fluxo usando os anticorpos: CD235, CD61, CD45, CD31, CD14, CD45, anti-TF e Anexina V. A atividade procoagulante plasmática foi investigada pela dosagem do fragmento 1+2 de protrombina (F1+2). A atividade procoagulante das MPs foi analisada pela dosagem de F1+2, de Dímero-D (DD2) e pelo teste de geração de trombina (TGT) em pool de indivíduos saudáveis em presença de MPs, corrigidas ou não por número na amostra (10.000 MPs). A análise estatística empregou os testes Wilcoxon ou U de Mann-Whitney, a=0.05. O número de MPs não estave diminuído em nenhum dos grupos estudados. A porcentagem de MPs estava estatisticamente aumentada nos pacientes com SAF/TVP em relação a seus controles (P=0,007). Observou-se um aumento significativo das MPs plaquetárias nos pacientes com SAF/TVP (P=0,01) e diminuição das MPs endoteliais naqueles com TVP ao diagnóstico (P=0,03), quando comparados aos seus controles. Os pacientes com SAF/TVP apresentaram diminuição significativa do F1+2 plasmático, tanto em relação aos seus controles (P=0,008) como ao CTR total (P=0,002). O F1+2 gerado pelas MPs estava significativamente diminuído em indivíduos com FVL em relação ao CTR total (0,009), e em pacientes com SAF/TVP em relação aos seus controles (P=0,001), e ao CTR total (P=0,008). O DD2 em pool de plasma, independente do número de MPs, estava significativamente aumentado na comparação entre TVP ao diagnóstico e seus controles (P=0,008) e ao CTR total (P=0,0001). O DD2 em pool com número corrigido de MPs apresentava-se aumentado significativamente nos pacientes com TVP ao diagnóstico quando comparados aos seus controles (P=0,008). Os valores do TGT com número corrigido de MPs estavam estatisticamente diminuídos em pacientes com TVP após 6 meses (P=0,01), em comparação ao CTR total. Nossos resultados demonstraram que o número de MPs está alterado em pacientes com TVP, e talvez possam ter um papel, particularmente após o evento trombótico ou em presença de anticorpos antifosfolípides. As MPs demonstraram atividade procoagulante, principalmente ao diagnóstico de TVP, podendo contribuir ou agravar o quadro clínico do paciente / Abstract: Deep Venous Thrombosis (DVT) is a multicausal disease, but many risk factors are not well defined. Microparticles (MPs) are small blebs released from cellular surfaces during activation and apoptosis. MPs may be the consequence of the dynamics between rest, activation and cellular death and can contribute to the seriousness of the illness and are therefore procoagulant and pro-inflammatory. There seems to be an association between high numbers of MPs and risk of thromboembolic complications and these may have a role in pathogenesis of these illnesses. In this study, we evaluated and characterized the MPs in patients with DVT of inferior limbs, [at diagnosis (5M/4H, medium age= 41.1 years), after 6 months of treatment (7M/3H, medium age= 32.9 years), and associated to Antibody Antiphospholipid Syndrome (7M/3H, medium age= 33.8 years)], and asymptomatic carriers of Factor V Leiden (FVL) (7M, medium age 34= years), matched to health controls by sex, age and ethnic origin. The MPs were isolated from citrated peripheral blood, by differential centrifugation. The quantification and characterization were performed by flow cytometry using the antibodies: CD235, CD61, CD45, CD31, CD14, CD45, anti-TF and Annexin V. The plasmatic procoagulantic activity was investigated by prothrombin fragment 1+2 (F1+2) dosage. The MPs procoagulant activities were analyzed by F1+2 dosage, D-dímer (DD2) and Thrombin Generation Test (TGT) in a pool of healthy individuals in the presence of MPs, corrected or not for number in the sample (10.000 MPs). Statistical analysiswas performed by Wilcoxon or the Mann-Whitney tests, a=0.05. The MPs number were not lower in any of the studied groups. The MPs percentage was statistically increased in the SAF/DVT patients compared to their matched controls (P=0.007). A significant increase in the platelet-derived MPs in SAF/DVT patients (P=0.01) and a reduction in the endothelial-derived MPs at diagnosis were observed (P=.03), when compared to their matched controls. The SAF/TVP patients show a significant reduction in plasmatic F1+2, when compared to their matched controls (P=0.008) and to the total CTR (P=0.002). The F1+2 generated by MPs were significantly lower in FVL carriers compared to the total CTR (0.009), and in SAF/DVT patients compared to their controls (P=0.001), and to the total CTR (P=0.008). The DD2 in the pool of plasma, independently of the number MPs, was significantly higher in DVT at diagnosis when compared to their matched controls (P=0.008) and the total CTR (P=0.0001). The DD2 in the pool with corrected MPs number was significantly higher in DVT at diagnosis patients when compared to their matched controls (P=0,008). The values of TGT in corrected MPs number were statistically lower in patients with DVT after 6 months (P=0.01), in comparison to the total CTR. Our results demonstrated that the number of MPs is modified in patients with DVT, and may play a role, particularly after the thrombotic event or in association with antiphospholipid antibodies. The MPs demonstrated procoagulant activity, especially at DVT diagnosis, and were able to contribute or to aggravate the patient¿s clinical situation / Mestrado / Biologia Estrutural, Celular, Molecular e do Desenvolvimento / Mestre em Fisiopatologia Médica

Orientador: Daniel Barrera-Arellano / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-15T08:05:38Z (GMT). No. of bitstreams: 1 Ribeiro_MarileneDeMoriMorselli_M.pdf: 5112345 bytes, checksum: 0515b869d93054892980d08c8bb30fd9 (MD5) Previous issue date: 2010 / Resumo: A facilidade de obtenção de micropartículas lipídicas e a possibilidade de produção em escala industrial aumentam o interesse do mercado alimentício de processar este tipo de material. Contudo, estas micropartículas apresentam desvantagens com relação à baixa encapsulação e à expulsão de material de recheio durante a estocagem. Assim, a finalidade deste trabalho foi estudar o comportamento das microcápsulas lipídicas produzidas pelo processo spray chilling utilizando as seguintes misturas em diferentes proporções: ácidos esteárico (AE) e oléico (AO), óleo de soja totalmente hidrogenado (STH) e ácido oléico (AO), álcool cetoestearílico (ACE) e ácido oléico (AO) como materiais de parede (matriz), lecitina de soja como tensoativo e solução de glicose como recheio. O objetivo foi aumentar a eficiência de encapsulação, verificando o efeito da composição e estrutura da matriz lipídica. Para este propósito, foram caracterizadas as matérias-primas lipídicas em composição de ácidos graxos e triacilgliceróis, bem como, as misturas lipídicas avaliadas por calorimetria diferencial de varredura (DSC), teor de gordura sólida (SFC) e curva de isosólidos. Nas micropartículas, foram avaliadas morfologia de superfície e microestrutura, tamanho e distribuição de partícula, quantidade de glicose superficial (não encapsulada), eficiência de encapsulação e perfil de liberação em solução aquosa. As micropartículas apresentaram formas esféricas e rugosas, com diâmetros médios entre 83 e 115 µm. Os resultados de eficiência de encapsulação nas misturas do AE e STH foram acima de 75% e nas misturas do ACE menor que 9%. A liberação do recheio foi avaliada a cada 30 minutos por 2 horas, obtendo-se valores de 28 a 89% ao término deste período para as misturas do AE e STH. Foi observado, nestas misturas, que a liberação de recheio é inversamente proporcional à quantidade de AO na mistura lipídica. Nas misturas do ACE, a liberação foi de 100% (efeito burst). A adição do lipídio líquido (AO) ao lipídio sólido (AE e STH) foi um fator determinante na modificação da cristalização da mistura lipídica, proporcionando uma alta eficiência de encapsulação / Abstract: The ease to obtain lipid microparticles and the possibility of their production in an industrial scale increase the interest of the food market to process this kind of particles. However, these microparticles present disadvantages with regard to low encapsulation and the expulsion of the core material core during storage. Thus, the purpose of this work was to study the behavior of lipid microcapsules produced by the spray chilling process, using the following mixtures in different proportions: stearic acid (SA) and oleic acid (OA), fully hydrogenated soybean oil (FHSO) and oleic acid (OA), cetostearyl alcohol (CEA) and oleic acid (OA) as the wall material (matrix), soy lecithin as surfactant and glucose solution as core. The objective was to increase the encapsulation efficiency, checking the composition effect and the lipid matrix structure. For this purpose, the lipid materials were characterized as to their fatty acids and triacylglycerol composition and the lipid mixtures were evaluated by differencial scanning calorimetry (DSC), solid fat content (SFC) and iso-solid curve. The surface morphology and microstructure were evaluated in the microparticles, as well as the particle size distribution, amount of core on the surface (not encapsulated), the encapsulation efficiency and controlled-release in an aqueous solution. The microparticles showed spherical and wrinkled shape with average diameters between 83 and 115 µm. The results of encapsulation efficiency of the mixtures with SA and FHSO were over 75% and lower than 9% in the mixtures with CEA. The core release was evaluated every 30 minutes for 2 hours obtaining values from 28 to 89% at the end of this period for the mixtures SA and FHSO. It was observed, in these mixtures, that the core release is inversely proportional to the quantity of AO in the lipid mixture. In the CEA mixtures, the core release was above 100% (burst effect). The addition of the liquid lipid (OA) to the solid lipid (SA and FHSO) was a determining factor in the modification of the crystallization of the lipid mixture providing high encapsulation efficiency / Mestrado / Mestre em Tecnologia de Alimentos