Helicoverpa armigera et Epiphyas postvittana, deux insectes nuisibles pour l'agriculture, ont développé des résistances contre la plupart des insecticides connus. Les lipides étant les constituants majeurs de leur régime alimentaire, les fonctions digestives des lipases deviennent alors une cible privilégiée pour l'élaboration de nouveaux insecticides. L'identification récente de gènes codant pour l'expression de protéines potentiellement actives et inactives apparentées aux lipases pancréatiques (PLRP) dans le tractus digestif de ces deux insectes et dont le niveau de transcription varie en fonction de leur régime alimentaire a ouvert un nouveau champ de recherche. Dans le but de contribuer à cette thématique, nous avons construit cinq lipases recombinantes d'E. postvittana (EpLIPs) et testé leur expression dans trois systèmes différents (E.coli, P.pastoris et bacculovirus). Le tractus digestif de Helicoverpa armigera a été étudié par chromatographie échangeuse d'ions et les différentes protéines séparées ont été testées en spectrométrie de masse et sur pHstat pour l'activité. Les résultats obtenus ont permis de mettre en évidence, pour la première fois, la présence à la fois d'une lipase active et d'une lipase inactive dans le tractus digestif d'un lépidoptère. Par ailleurs, la caractérisation biochimique d'un mutant GPLRP2-Δ β9 a été faite dans le but de comprendre l'effet de cette boucle, partiellement délétée dans les lipases d'insectes, dans la spécificité du substrat. Nous avons pu montrer que cette boucle β9 est essentielle pour la stabilisation de la chaîne acyle durant la réaction de lipolyse. / Helicoverpa armigera and Epiphyas postvittana, two major pest crops, have developed resistances against most of the known insecticides. Lipids being a major component of insect diet, digestive function of lipase are a target of choice for new insecticide design. The recent identification of active and inactive pancreatic lipase related protein (PLRP) genes in those two insects midgut, with a level of transcription depending on the diet, opened the field of insect digestive lipase study. In order to contribute to this thematic, we built five recombinants lipase from E. postvittana (EpLIPs) and tested their expression in three different systems (E.coli, P.pastoris and bacculovirus). Protein structure prediction of EpLIPs allowed us to develop some functional hypothesis enlightening the role of inactive lipase in lipid transport. H. armigera midgut contents were separated through a one step purification chromatography and the different fractions were tested for activity and mass spectrometry. The results obtained gave the first evidence of the presence of both an active and an inactive lipase in lepidopteran midgut. In addition to this work, a biochemical characterisation of a β9 GPLRP2 mutant was carried out to understand the effect of this loop, partially deleted in insect lipase, in substrate specificity. The result shows that β9 loop is essential for stabilizing the leaving acyl chain during the lipolysis reaction.

A gordura abdominal de frango é um subproduto ou resíduo avícola que apresenta grande potencial de utilização na indústria de alimentos, uma vez que possui rica composição lipídica, elevado teor energético, baixo custo e propriedades físico-químicas desejáveis. Nesta pesquisa, a gordura abdominal de frango, obtida no comércio local, foi submetida à fusão e à filtração. A seguir, sofreu fracionamento a seco e mistura com gordura de toucinho. O processo de fracionamento foi realizado a duas temperaturas: 17,5ºC e 20ºC. As análises realizadas na gordura original e nas frações obtidas (oleína e estearina) foram ponto de fusão, consistência, conteúdo de gordura sólida, composição em ácidos graxos e índices de iodo e de saponificação. De acordo com os resultados observou-se que a gordura de frango apresentou 67,2% de ácidos graxos insaturados. A gordura de frango à temperatura de 10ºC apresentou-se plástica e espalhável. Foi obtida relação linear entre a consistência e o conteúdo de gordura sólida. O alto rendimento das oleínas (>84%) sugere a possibilidade de aplicação destas frações como óleo para frituras. As estearinas podem ser utilizadas como componentes na fabricação de gorduras, para aplicação em pastelaria, margarinas para massa folhada e caldos desidratados. Foram realizadas misturas binárias entre as estearinas e a gordura de toucinho e entre a gordura de frango e a gordura de toucinho. As amostras foram analisadas quanto à composição em ácidos graxos, consistência às temperaturas de 10 a 25ºC e quanto ao conteúdo de gordura sólida às temperaturas de 10 a 35ºC. Utilizou-se o analisador de textura TA-XT2 (Stable Micro Systems) e o espectrômetro de ressonância magnética nuclear Maran Ultra Benchtop de 20 MHz para as análises de consistência e de conteúdo de gordura sólida, respectivamente. Foi aplicado um modelo de regressão múltipla do tipo quadrático. Os resultados para a consistência demonstraram que ocorreu o efeito eutético entre as estearinas obtidas no fracionamento da gordura de frango e a gordura de toucinho. Por outro lado, nas misturas entre gordura de frango e gordura de toucinho ocorreu a formação de compostos. Os resultados para o conteúdo de gordura sólida demonstraram que ocorreram os efeitos eutético e de formação de compostos entre as misturas. Nas misturas binárias de gordura abdominal de frango com gordura de toucinho foram determinadas as propriedades de tensão, firmeza e energia de penetração às temperaturas de 10, 15 e 20ºC. Foram verificadas as interações entre estas gorduras. O aumento da temperatura de análise causou diminuição dos valores de firmeza e de energia necessária para a penetração, em função da redução do conteúdo de gordura sólida e alterações na rede cristalina. Por aplicação de um modelo de regressão múltipla do tipo quadrático, verificou-se que, nas misturas entre gordura de frango e gordura de toucinho, ocorreu o efeito sinérgico da formação de compostos. Análises da distribuição estereoespecífica dos ácidos graxos foram realizadas na gordura abdominal de frango e suas frações, e na gordura de toucinho. As amostras foram submetidas à hidrólise com lipase pancreática suína a temperatura de 40°C e posteriormente analisadas quanto à composição em ácidos graxos na posição sn-2 por cromatografia gasosa. A partir dos resultados obtidos foram calculados os grupos de triacilgliceróis nas amostras naturais e em suas frações, utilizando a teoria 1,3-random 2-random. Os resultados demonstraram que houve preferência do ácido oléico pela posição sn-2 para a gordura de frango e suas oleínas. Para a gordura de toucinho, o ácido palmítico mostrou alta preferência pela posição sn-2. Para as estearinas da gordura de frango, também houve preferência do ácido palmítico pela posição central do glicerol. / Abdominal chicken fat from the local commerce was melted, filtered, dry fractionated, and blended with bacon fat. Fractionation process was made at 17,5ºC and 20C. Physical and chemical properties such as melting point, consistency, solid fat content, fatty acid composition, iodine and saponification values of the products from the abdominal chicken fat fractionation were determined. Results showed that chicken fat has 67.2% of unsaturated fatty acids. Chicken fat at 10ºC was plastic and spreadable. A linear relationship between consistency and solid fat content was obtained. High oleins yield (>84%) suggest that these fractions can be used as frying oils. Stearins can be applied as components in the fat manufacturing, in pastry, in puff-pastry margarines and dehidrated soup cubes. Binary mixtures between stearins and bacon fat, and chicken fat with bacon fat were made. The samples were analyzed for the fatty acid composition, consistency at the temperatures of 10 to 25C, and solid fat content at 10 to 35ºC. The texture analyzer TA-XT2 (Stable Micro Systems) and the magnetic nuclear resonance espectrometer Maran Ultra Benchtop with 20 MHz were utilized, respectively. A mathematical model of multiple regression of the quadratic type was applied. The consistency results showed that the eutectic effect was observed between the binary mixtures of stearins and bacon fat. On the other hand, mixtures between abdominal chicken fat and bacon fat showed compound formation. The solid fat content results showed that the eutectic effect and compound formation were observed. The properties of stress, firmness, and penetration energy of binary mixtures of abdominal chicken fat and bacon fat, at 10, 15 and 20ºC. The increase of the analysis temperature leads reduction on the values of firmness and penetration energy, as a function of the reduction of solid fat content and the alteration on the crystal network. The results of mixtures between abdominal chicken fat and bacon fat showed a synergic effect of compounds formation. Stereospecific distribution analysis in chicken fat and its fractions, and in bacon fat were realized. The samples were hydrolyzed with swine pancreatic lipase at 40°C and then analyzed for the fatty acid composition by gas chromatography. From the experimental results, the groups of triacylglycerols in the natural samples and its fractions were calculated. The 1,3-random 2-random theory was utilized. The results showed that oleic acid was mainly esterified to the sn-2 position for abdominal chicken fat and oleins whereas palmitic acid was mainly distributed at this same position for bacon fat. In the case of chicken fat stearins the preference by sn-2 position was also to palmitic acid.

A gordura abdominal de frango é um subproduto ou resíduo avícola que apresenta grande potencial de utilização na indústria de alimentos, uma vez que possui rica composição lipídica, elevado teor energético, baixo custo e propriedades físico-químicas desejáveis. Nesta pesquisa, a gordura abdominal de frango, obtida no comércio local, foi submetida à fusão e à filtração. A seguir, sofreu fracionamento a seco e mistura com gordura de toucinho. O processo de fracionamento foi realizado a duas temperaturas: 17,5ºC e 20ºC. As análises realizadas na gordura original e nas frações obtidas (oleína e estearina) foram ponto de fusão, consistência, conteúdo de gordura sólida, composição em ácidos graxos e índices de iodo e de saponificação. De acordo com os resultados observou-se que a gordura de frango apresentou 67,2% de ácidos graxos insaturados. A gordura de frango à temperatura de 10ºC apresentou-se plástica e espalhável. Foi obtida relação linear entre a consistência e o conteúdo de gordura sólida. O alto rendimento das oleínas (>84%) sugere a possibilidade de aplicação destas frações como óleo para frituras. As estearinas podem ser utilizadas como componentes na fabricação de gorduras, para aplicação em pastelaria, margarinas para massa folhada e caldos desidratados. Foram realizadas misturas binárias entre as estearinas e a gordura de toucinho e entre a gordura de frango e a gordura de toucinho. As amostras foram analisadas quanto à composição em ácidos graxos, consistência às temperaturas de 10 a 25ºC e quanto ao conteúdo de gordura sólida às temperaturas de 10 a 35ºC. Utilizou-se o analisador de textura TA-XT2 (Stable Micro Systems) e o espectrômetro de ressonância magnética nuclear Maran Ultra Benchtop de 20 MHz para as análises de consistência e de conteúdo de gordura sólida, respectivamente. Foi aplicado um modelo de regressão múltipla do tipo quadrático. Os resultados para a consistência demonstraram que ocorreu o efeito eutético entre as estearinas obtidas no fracionamento da gordura de frango e a gordura de toucinho. Por outro lado, nas misturas entre gordura de frango e gordura de toucinho ocorreu a formação de compostos. Os resultados para o conteúdo de gordura sólida demonstraram que ocorreram os efeitos eutético e de formação de compostos entre as misturas. Nas misturas binárias de gordura abdominal de frango com gordura de toucinho foram determinadas as propriedades de tensão, firmeza e energia de penetração às temperaturas de 10, 15 e 20ºC. Foram verificadas as interações entre estas gorduras. O aumento da temperatura de análise causou diminuição dos valores de firmeza e de energia necessária para a penetração, em função da redução do conteúdo de gordura sólida e alterações na rede cristalina. Por aplicação de um modelo de regressão múltipla do tipo quadrático, verificou-se que, nas misturas entre gordura de frango e gordura de toucinho, ocorreu o efeito sinérgico da formação de compostos. Análises da distribuição estereoespecífica dos ácidos graxos foram realizadas na gordura abdominal de frango e suas frações, e na gordura de toucinho. As amostras foram submetidas à hidrólise com lipase pancreática suína a temperatura de 40°C e posteriormente analisadas quanto à composição em ácidos graxos na posição sn-2 por cromatografia gasosa. A partir dos resultados obtidos foram calculados os grupos de triacilgliceróis nas amostras naturais e em suas frações, utilizando a teoria 1,3-random 2-random. Os resultados demonstraram que houve preferência do ácido oléico pela posição sn-2 para a gordura de frango e suas oleínas. Para a gordura de toucinho, o ácido palmítico mostrou alta preferência pela posição sn-2. Para as estearinas da gordura de frango, também houve preferência do ácido palmítico pela posição central do glicerol. / Abdominal chicken fat from the local commerce was melted, filtered, dry fractionated, and blended with bacon fat. Fractionation process was made at 17,5ºC and 20C. Physical and chemical properties such as melting point, consistency, solid fat content, fatty acid composition, iodine and saponification values of the products from the abdominal chicken fat fractionation were determined. Results showed that chicken fat has 67.2% of unsaturated fatty acids. Chicken fat at 10ºC was plastic and spreadable. A linear relationship between consistency and solid fat content was obtained. High oleins yield (>84%) suggest that these fractions can be used as frying oils. Stearins can be applied as components in the fat manufacturing, in pastry, in puff-pastry margarines and dehidrated soup cubes. Binary mixtures between stearins and bacon fat, and chicken fat with bacon fat were made. The samples were analyzed for the fatty acid composition, consistency at the temperatures of 10 to 25C, and solid fat content at 10 to 35ºC. The texture analyzer TA-XT2 (Stable Micro Systems) and the magnetic nuclear resonance espectrometer Maran Ultra Benchtop with 20 MHz were utilized, respectively. A mathematical model of multiple regression of the quadratic type was applied. The consistency results showed that the eutectic effect was observed between the binary mixtures of stearins and bacon fat. On the other hand, mixtures between abdominal chicken fat and bacon fat showed compound formation. The solid fat content results showed that the eutectic effect and compound formation were observed. The properties of stress, firmness, and penetration energy of binary mixtures of abdominal chicken fat and bacon fat, at 10, 15 and 20ºC. The increase of the analysis temperature leads reduction on the values of firmness and penetration energy, as a function of the reduction of solid fat content and the alteration on the crystal network. The results of mixtures between abdominal chicken fat and bacon fat showed a synergic effect of compounds formation. Stereospecific distribution analysis in chicken fat and its fractions, and in bacon fat were realized. The samples were hydrolyzed with swine pancreatic lipase at 40°C and then analyzed for the fatty acid composition by gas chromatography. From the experimental results, the groups of triacylglycerols in the natural samples and its fractions were calculated. The 1,3-random 2-random theory was utilized. The results showed that oleic acid was mainly esterified to the sn-2 position for abdominal chicken fat and oleins whereas palmitic acid was mainly distributed at this same position for bacon fat. In the case of chicken fat stearins the preference by sn-2 position was also to palmitic acid.

La famille des lipases pancréatiques est constituée de trois sous-familles : la lipase pancréatique classique (PL) et les lipases pancréatiques apparentées de type 1 et 2 (PLRP1 et PLRP2). Contrairement à la PL, qui est active seulement sur les triglycérides, la PLRP2 possède trois types d’activités enzymatiques : lipase, phospholipase de type A1 et surtout galactolipase. Cette dernière activité est particulièrement intéressante car les galactolipides, principalement les monogalactosyldiacylglycérols (MGDG) et les digalactosyldiacylglycérols (DGDG) sont les lipides les plus abondants dans la nature. Un test continu, spécifique et fiable de dosage de l’activité galactolipase à été mis au point en utilisant la technique du pH-stat et un galactolipide synthétique à chaîne moyenne comme substrat, le MGDG en C8. Ce nouveau test de dosage nous a permis de mesurer pour la première fois des activités spécifiques importantes avec les PLRP2 humaine (HPLRP2) et du cobaye (GPLRP2) (1786±100 et 5420±85 U/mg, respectivement). Nous avons par la suite utilisé cette même technique pour étudier l’hydrolyse par les PLRP2 des galactolipides naturels à chaînes longues purifiés à partir de feuilles d’épinard. Les activités spécifiques mesurées sont du même ordre de grandeur que celles mesurées avec le MGDG en C8 mais aussi le DGDG en C8 synthétique, montrant ainsi que l’activité galactolipase n’est affectée ni par la longueur de la chaîne acyle ni par la présence d’un ou de deux résidus galactoses. Grâce à ce test nous avons pu réaliser un criblage de lipases microbiennes de différentes origines à la recherche d’activités galactolipase. Dans une deuxième partie, nous avons étudié les propriétés physico-chimiques du bis-monoacylglycérol-phopshate (BMP). Nous avons montré que non seulement ce phospholipide rare est hydrolysé par la PLRP2, mais le BMP et la PLRP2 humaine peuvent être trouvés dans la même cellule, le monocyte, suggérant un nouveau rôle physiologique pour la PLRP2. / The pancreatic lipase family consists of three sub-families : the classical pancreatic lipase (PL) and the pancreatic lipase-related proteins 1 and 2 (PLRP1 and PLRP2). On the contrary to PL, which is active only on triglycerides, the PLRP2 possesses three types of enzymatic activities: lipase activity, phospholipase A1 activity and especially galactolipase activity. This latter activity is particularly interesting because monogalactosyl-diacylglycerol (MGDG) and digalactosyldiacyl-glycerol (DGDG) are the most abundant lipids in nature. A specific, continuous and robust galactolipase assay using the pH-stat technique and a synthetic medium chain MGDG as substrate was developed, and recombinant human (rHPLRP2) and guinea pig (rGPLRP2) pancreatic lipase-related proteins 2 were tested as model enzymes. This new assay allowed us to measure for the first time high specific activities with both PLRP2 (1786±100 and 5420±85 U/mg, respectively). We then used this technique to study the hydrolysis by PLRP2 of natural long chain galactolipids purified from spinach leaves. The length of acyl chains and the nature of the galactosyl polar head of the galactolipid did not have major effects on the specific activities of PLRP2, which were found to be very high on both medium chain and long chain galactolipids. We also used the pH-stat technique and medium chain MGDG and DGDG for screening several microbial lipases for their galactolipase activity.In a separate study, we investigated the physico-chemical properties of bis-monoacylglycorol-phopshate (BMP) and found that this rare phospholipid was hydrolyzed by PLRP2. Moreover, BMP and human PLRP2 could be found in the same cell, the monocyte, suggesting a new physiological role for PLRP2.

<p>  </p> <p>The ever-growing interest in novel food ingredients and their dietary influence on human health and wellbeing has driven the study of bioactive peptides (BAP). BAP are protein-derived fragments composed of (2-20 amino acids) that could positively affect bodily function and chronic diseases. This dissertation explores the health-promoting properties of a novel source of BAPs, namely canary seed, by encompassing three specific aims: 1) evaluate the <em>in vitro </em>potential of hairless canary seed peptides (CSP) as a nutraceutical ingredient, 2) develop an understanding of CSP's bioavailability and molecular interactions with its biological targets, and 3) evaluate CSP's antioxidant and antiobesity activity at the organism level using a nematode (<em>C. elegans</em>) and murine (C57BL/6J mice) model, respectively. First, CSP were generated by implementing mechanical oil extraction followed by commercial enzymatic hydrolysis with Alcalase™. In addition, CSP were also subject to simulated gastrointestinal digestion (SGD) to assess their gastric stability and <em>in vitro</em> bioavailability. The results showed that canary seed proteins were mainly composed of prolamins fractions followed by glutelins, globulins, and albumins. CSP extracts with low molecular weight (< 3 kDa and 3–10 kDa) showed the highest bioactivity. Furthermore, after SGD, CSP inhibitory activity remained stable toward angiotensin-converting enzyme (ACE), dipeptidyl peptidase IV (DPP-IV), and pancreatic lipase but unstable for α-glucosidase. The digested peptides were transported efficiently (>10%) through the Caco-2 monolayer, indicating a potential high absorption capacity through the intestinal epithelium. During kinetic analysis by Lineweaver-Burk plots, it was observed that CSP-SGD interacted by mixed-type inhibition for DPP-IV and α-glucosidase, non-competitive inhibition for ACE, and uncompetitive inhibitor for pancreatic lipase. Furthermore, CSP-SGD were especially potent as antihypertensive (ACE inhibitors) and antiobesity (pancreatic lipase) agents. Consequently, molecular docking and <em>in silico</em> analyses were targeted to understand CSP-SGD interactions with ACE and pancreatic lipase. CSP-SGD with ACE inhibitory activity were found to be rich in proline, glutamine, and cationic residues and could have inhibited ACE by destabilizing the tetrahedral transition state and zinc ions interaction leading to conformational changes in the enzyme structure. For peptides with antiobesity properties, it was also found that arginine, glycine, and hydrophobic amino acids from CSP-SGD hold critical interactions with the lid domain (CYS238-CYS262) of pancreatic lipase, disrupting its proper function and preventing fat hydrolysis.</p> <p>In the second part of this dissertation, the relevant health-promoting properties of CSP were further investigated by testing the effects of peptide supplementation on obesity and oxidative stress animal models. The studies showed that exposure to CSP significantly mitigated the acute and chronic oxidative damage in <em>C. elegans</em>, extending the lifespan of the nematodes by 88 and 61%, respectively. Furthermore, it was established that the CSP prevented oxidative stress by scavenging free radicals and antioxidant gene upregulation. Concerning this, CSP caused a drop in reactive oxygen species (ROS) to safe levels and induce the upregulation of the GST-4 gene encoding antioxidant enzyme Glutathione S-transferase. Concerning antiobesity properties, the daily supplementation with CSP successfully prevented metabolic implications of western diet-induced obesity in C57BL/6J mice, including preventing weight gain by up to 20%, increasing glucose tolerance, and reducing insulin, leptin, and LDL/VLDL levels in plasma. Likewise, CSP promoted a drop in fatty acid uptake gene, LPL, and fatty acid biosynthesis genes FAS and ACC while unaffecting lipid oxidation genes PPAR-α and ACO in the liver. While both moderate and high CSP supplementation levels exhibit hypolipidemic effects, only moderate levels induce satiety and significantly prevent weight gain. Together, these results suggest that CSP's weight gain prevention depended on a dual mechanism involving lipid metabolism retardation to modulate satiety. Overall, the results presented in this dissertation establish the effectiveness of canary seed peptides as nutraceutical ingredients for antioxidant and antiobesity functional food applications.</p>

La prevalencia de los trastornos metabólicos está creciendo a nivel mundial. Botánicos, como el romero (Rosmarinus officinalis L.) y las semillas de fresno (Fraxinus excelsior L.), pueden ser una alternativa para mejorar estos trastornos. El objetivo de esta tesis doctoral fue generar extractos industriales de las partes comestibles de estas plantas, identificar sus compuestos por HPLC, HPLC-MS y RMN, y determinar su eficacia y seguridad. Se desarrollaron tres extractos de romero - estandarizados al 20% de ácido carnósico, 20% de ácido rosmarínico, y 40% de ácido ursólico. El extracto rico en ácido ursólico mostró la menor capacidad antioxidante en modelos ORAC y FRAP in vitro, mientras que los otros dos extractos tuvieron capacidades más elevadas. Sin embargo, el extracto rico en ácido carnósico (RE) fue superior en la inhibición de la oxidación de LDL ex vivo, y fue el único seleccionado para estudios posteriores. Se desarrolló un extracto de semilla de fresno (FE) de acuerdo a su uso tradicional en Marruecos, siendo identificados salidrosido y 9 glucósidos secoiridoides, dos de ellos fueron descubiertos por primera vez: Excelside A y Excelside B. En estudios in vitro, RE y FE activaron receptores nucleares que regulan la homeostasis de la glucosa y la energía, y actuaron sobre mecanismos contra la obesidad y la dislipidemia - RE activa PPAR? e inhibe la lipasa pancreática, y FE estimula PPAR? y evita la diferenciación de preadipocitos. La eficacia de ambos extractos para el control de trastornos metabólicos fue confirmada en ratones C57BL/6J administrados con una dieta baja en grasa, una dieta alta en grasa, o una dieta alta en grasa más 0.5% RE (HFD.RE) o 0.5% FE (FED) durante 16 semanas. HFD.RE limitó el aumento del peso corporal y grasa en el epidídimo en un 69% (P<0.01) y 79% (P<0.001), respectivamente. HFD.RE también redujo la glucemia (72%, P<0.01) y los niveles de colesterol (68%, P<0.001). FED redujo la glucemia (76.52%, P<0.001), insulinemia (53.43%, P< / Ibarra, SA. (2011). OBTENTION AND CHARACTERIZATION OF ROSEMARY AND ASH TREE SEED EXTRACTS AND STUDY OF THEIR PREVENTIVE EFFECTS ON METABOLIC DISORDERS [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/10795

The main objective of this study was the formulation of food-grade water-in-oil (W/O) nano-dispersions based mainly on medium or long-chain triglycerides. Two types of dispersions were formulated and structurally compared, namely emulsions and microemulsions. The systems were used as matrices for encapsulating targeted bioactive molecules with specific characteristics such as antioxidants or peptides. The structural characterization of the formulated systems was investigated using techniques such as Electron Paramagnetic Resonance (EPR) spectroscopy, Dynamic Light Scattering (DLS), Cryogenic Transmission Electron Microscopy (Cryo-TEM) and Small Angle Xray Scattering (SAXS). The existence of swollen inverse micelles was revealed for the case of microemulsions whereas larger droplets still at the nano-scale were observed for the case of emulsions. Structural differences in the presence of the bioactive molecules or induced by the alteration of components were also observed. In order to study the efficacy of the formulations, the proposed loaded systems were assessed either using EPR spectroscopy or Well Diffusion Assay (WDA) depending on the bioactive molecule. It was found that the encapsulated molecules retained their claimed characteristics when encapsulated to the proposed matrices. Finally, some of the formulated dispersions were investigated for their behavior under gastrointestinal (GI) conditions. A two-step digestion model using recombinant Dog Gastric Lipase (rDGL) and Porcine Pancreatic Lipase (PPL) was proposed to simulate lipid hydrolysis in humans. The studies revealed significant decrease of the rDGL specific activity in the presence of the microemulsion while in the presence of lower percent of surfactants (case of emulsion) no alterations were observed.

Submitted by Renata Lopes (renatasil82@gmail.com) on 2016-01-22T10:23:14Z No. of bitstreams: 1 gustavorezendebelleideoliveira.pdf: 2916542 bytes, checksum: 307fa0e3fa1f842c7fbc463dd8a0a712 (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2016-01-25T18:49:31Z (GMT) No. of bitstreams: 1 gustavorezendebelleideoliveira.pdf: 2916542 bytes, checksum: 307fa0e3fa1f842c7fbc463dd8a0a712 (MD5) / Made available in DSpace on 2016-01-25T18:49:32Z (GMT). No. of bitstreams: 1 gustavorezendebelleideoliveira.pdf: 2916542 bytes, checksum: 307fa0e3fa1f842c7fbc463dd8a0a712 (MD5) Previous issue date: 2014-07-25 / FAPEMIG - Fundação de Amparo à Pesquisa do Estado de Minas Gerais / A obesidade é o resultado de diversas características do indivíduo, entre as quais se destacam os aspectos genéticos, ambientais e comportamentais. Sua ocorrência é devido ao acúmulo anormal ou excessivo de gordura, que pode resultar em risco de desenvolvimento de doenças graves e alterações lipídicas como dislipidemias e hipercolesterolemia. Vários tratamentos terapêuticos são utilizados para redução do peso corporal. Medicamentos com ação inibidora da lipase digetiva na região do lumen intestinal são utilizados para controlar o metabolismo lipídico, que propicia a redução da absorção de gordura. As plantas têm sido amplamente estudadas como recurso terapêutico, fundamentado no uso popular. Várias plantas apresentam ação antilipidêmica comprovada, devido à presença dos catecois. A Endopleura uchi, conhecida popularmente como uxi amarelo, apresenta em seus extratos, derivados fenólicos e flavonoídicos (catequinas). Este trabalho teve como objetivo, o estudo de avaliação da eficácia antilipidêmica da casca de Endopleura uchi, empregando método espectrofotométrico. A amostra de casca de E. uchi seca e triturada foi adquirida comercialmente. A metodologia envolveu a obtenção dos extratos; quantificação de derivados flavonoídicos e derivados fenólicos totais; avaliação da atividade antioxidante e determinação do efeito inibidor dos extratos na atividade enzimática da lipase pancreática. Foram preparados três extratos em solventes com polaridades diferentes, a partir de 200g de casca seca triturada de E. uchi. Para a determinação de derivados flavonídicos foi utilizado o método com cloreto de alumínio em meio ácido na presença de piridina e para derivados fenólicos totais o método com reagente de Folin-Ciocalteau. A atividade antioxidante foi realizada pelo ensaio de redução do radical DPPH. O efeito antilipidêmico dos extratos foi avaliado através da inibição enzimática da lipase pancreática utilizando como substrato o p-nitrofenolpalmitato. Os extratos obtidos apresentaram rendimento de 23,4 g (11,7%) para o extrato em água, 34,2g (17,1%) de extrato em etanol 80% e 26,6 g (13,3%) de extrato em acetona 80%. O conteúdo de derivados flavonoídicos equivalente em quercetina por grama de amostra foi de 1,35mg para extrato em água, 1,80mg para extrato em etanol 80% e 2,13 mg para extrato em acetona 80%. O conteúdo de derivados fenólicos expressos em equivalente em ácido tânico por grama de amostra foi de 0,639g para extrato em água, 0,950g para extrato em etanol 80% e 0,930g para extrato em acetona 80%. As concentrações efetivas 50% para o método de DPPH variaram de 12,4 μg/mL, 9,7 μg/mL e 7,9 μg/mL para os extratos em água, etanol 80% e em acetona 80% respectivamente. Foi encontrada forte correlação entre atividade antioxidante e o conteúdo de derivados fenólicos. O coeficiente de Pearson para os três extratos foi r > 0,9. Na avaliação dos extratos frente a inibição da lipase pancreática, o extrato aquoso mostrou atividade inibidora de 47,54% (106,90 UIL/g), em etanol 80% (1:1) 36,88% (96,06 UIL/g) e extrato em acetona 80% (1:1) 49,33% (112,14 UIL/g). Os resultados obtidos mostraram a presença das atividades antilipidêmica e antioxidante dos extratos de casca de E. uchi, importantes para o tratamento de redução de gordura corporal. / Obesity is the result of various individual characteristics, among which stand out the genetic, environmental and behavioral aspects. Its occurrence is due to abnormal or excessive fat accumulation that may result in the risk of developing serious diseases and lipid disorders such as dyslipidemia and hypercholesterolemia. Many therapeutic treatments are used for reducing body weight. Drugs that inhibits the digestive lipase in the intestinal lumen region are used to control lipid metabolism, which promotes the reduction of fat absorption. Plants have been widely studied as a treatment method based on the popular use. Several plants have proven, due to the presence of catechols antilipemic action. The Endopleura uchi, popularly known as yellow uxi presents in their extracts, phenolic derivatives and flavonoids (catechins). This work aimed to study the efficacy assessment antilipemic bark Endopleura uchi, employing spectrophotometric method. A sample of E. uchi bark was dried and crushed commercially acquired. The methodology involved obtaining the extracts; quantification of flavonoid compounds and phenolic derivatives; evaluation of antioxidant activity and determination of the inhibitory effect of the extract on the enzymatic activity of pancreatic lipase. Three extracts were prepared in solvents with different polarities from 200g dry shredded bark of E. uchi. For the determination of the method flavonoid derivative with aluminum chloride in an acid medium in the presence of pyridine and phenolic derivatives with the method of Folin-Ciocalteau reagent was used. The antioxidant activity assay was performed by reduction of DPPH. Antilipemic effect of the extracts was assessed by enzymatic inhibition of pancreatic lipase using a substrate of p-nitrofenylpalmitate. The extracts showed yield 23,4 g (11,7%) for the water extract, 34,2 g (17.1%) of 80% ethanol extract and 26,6 g (13,3%) of 80% acetone extract. The content of flavonoid compounds in quercetin equivalent of gram of sample was 1,35 mg to water extract, 1,80 mg extract in 80% ethanol and 2,13 mg 80% extract acetone. The contents of phenolic derivatives expressed in equivalent of gram of tannic acid was 0,639 g sample to extract in water to 0,950 g in 80% ethanol extract and 0,930 g in 80% acetone extract. The effective concentration for 50% DPPH assay ranged from 12,4 μg/mL, 9,7 μg/mL and 7,9 μg/mL for the extracts with water, 80% ethanol and 80% acetone, respectively. Strong correlation between antioxidant activity and content of phenolic derivatives was found. The Pearson coefficient for the three extracts was r > 0,9. In the evaluation of the extracts against pancreatic lipase inhibition of the aqueous extract showed inhibitory activity of 47.54% (106,90 ALI/g) in 80% ethanol (1:1) 36,88% (96,06 ALI/g) and extract with 80% acetone (1:1) 49,33% (112,14 ALI/g). The results showed the presence of antilipemic and antioxidant activities of the extracts of bark of E. uchi, important for the treatment of body fat reduction.

Le diabète de type 2 et l’obésité affectent la population mondiale et, de façon disproportionnelle, chez la population adulte crie d’Eeyou Istchee. Afin de contribuer à la documentation des activités antidiabétiques de 17 plantes médicinales de la forêt boréale traditionnellement recommandées pour traiter les symptômes du diabète, cette étude procède à l’évaluation de l’activité enzymatique de la lipase pancréatique par différentes espèces de plantes. De plus, une évaluation de l’activité enzymatique de la lipase pancréatique est réalisée avec des espèces antidiabétiques de la pharmacopée urbaine de Niamey (Niger) ainsi qu’avec des baies québécoises. Enfin, des composés phénoliques sont identifiés chez ces deux derniers groupes d’espèces végétales. Les résultats de ces études démontrent que la majorité des espèces évaluées ont un effet inhibiteur sur la lipase pancréatique in vitro. Les espèces ayant démontré les plus fortes activités inhibitrices sur la lipase pancréatique sont Kalmia angustifolia, Gaultheria hispidula et Acacia nilotica. Ainsi, cette étude permet d’appuyer la justesse des connaissances traditionnelles en ajoutant la documentation d’une activité antidiabétique des espèces végétales au répertoire scientifique. / Type 2 diabetes and obesity affect the global population and, disproportionately, the adult population of the Eeyou Istchee Cree. In order to contribute to the documentation of the antidiabetic activities of 17 medicinal plants of the boreal forest traditionally recommended for treating the symptoms of diabetes, this study evaluates the pancreatic lipase enzymatic activity of different plant species. In addition, an evaluation of the enzymatic activity of the pancreatic lipase is carried out with antidiabetic species of the urban pharmacopoeia of Niamey (Niger) as well as with Quebec berries. Finally, phenolic compounds are identified in these last two groups of plant species. Results of these studies demonstrate that the majority of the evaluated species have an inhibitory effect on pancreatic lipase in vitro. Species showing the strongest inhibitory activities towards pancreatic lipase are Kalmia angustifolia, Gaultheria hispidula and Acacia nilotica. Thus, this study supports the accuracy of traditionnal knowledge by adding the documentation of an antidiabetic activity of plant species to the scientific repertoire.