Déterminer la composition en triacylglycérols des huiles végétales est un défi important à relever aussi bien en biologie végétale que dans le domaine médical ou celui de l’industrie agro-alimentaire. La chromatographie en phase liquide à polarité inversée de phases en milieu non aqueux (NARP-LC) est la méthode la plus utilisée, avec cependant une utilisation récurrente de solvants chlorés. Le premier objectif de ce travail a été de proposer des conditions analytiques alternatives n’utilisant pas de solvants chlorés. Pour ce faire, nous avons établi le diagramme de force éluante sur phases stationnaires en C18 des phases mobiles binaires constituées d’acétonitrile comme solvant faible et de divers solvants forts (acétone, iso-propanol, acétate d’éthyle, butanol) à quatre températures différentes (25, 43, 63 et 85°C). La comparaison dans des conditions iso-éluantes de l’analyse de 9 huiles de graines contenant une large gamme de TAG nous a permis de montrer que le mélange MeCN/BuOH 74/26 à 25°C est le meilleur choix, en terme de sélectivité pour l’analyse des TAG. Ce qui répond à notre premier objectif. Dans un second temps, nous avons comparé le potentiel séparatif des phases stationnaires de nouvelle génération à petit diamètre de particules, partiellement ou totalement poreuses. L’optimisation des conditions chromatographiques nous a permis de décrire deux systèmes chromatographiques, très performants, en termes d’efficacité et de rapidité. Le premier permet la séparation de TAG contenant des acides gras polyinsaturés isomères de position en C18 : 3 et C18 : 2. L’identification de ces isomères particuliers a été réalisée grâce à la synthèse d’informations complémentaires, obtenues en GC/MS, LC/MS ainsi que l’utilisation de lois de rétention chromatographiques. En outre, ce travail nous a permis de proposer un tableau récapitulatif regroupant un très grand nombre de TAG, qui n’ont jamais été décrits à notre connaissance. Le second permet une analyse rapide de l’huile d’olive, en moins de 5 min, tout en respectant les consignes qualitatives imposées par l’organisme «Conseil Oléique International (COI)». Comparée aux méthodes officielles couramment utilisées, elle mérite d’être proposée comme méthode de référence pour le contrôle de qualité de l’appellation de ces huiles. / The determination of triacylglycerols in vegetable oils is an important challenge in plant biology, in the medical field, and in food industry. Nowadays, non-Aqueous Reversed Phase Liquid Chromatography (NARP-LC) using chlorinated solvents is commonly used for this purpose. The first objective of this work was to develop alternative analytical conditions that can avoid using chlorinated solvents. In a first step, by using C18 stationary phases, we have established the eluotropic solvent strength scale as a function of temperature of several binary mobile phases consisting of acetonitrile as weak solvent and various strong solvents including acetone, isopropanol, ethyl acetate, and butanol. The comparison of the results obtained under iso-eluotropic conditions for nine seed oils containing a wide range of TAG enabled us to show that the MeCN/BuOH (74/26, v/V) mixture operating at 25 °C are the best mobile phase conditions for TAG analysis, in terms of selectivity, thus avoiding the use of chlorinated solvents. In a second step, we compared the separation of TAGs on new generation of fully or partially porous stationary phase particles of small diameter. After optimizing the separating conditions, the obtained data allowed us to propose two highly efficient chromatographic systems. The first system enables the efficient separation of C18:3 and C18:2 positional isomers of C18 polyunsaturated fatty acids containing TAG. For the identification of these TAG isomers, it was necessary to combine the data obtained by GC-MS and LC -MS as well as the data obtained by the application of some chromatographic retention laws. Taken together, these results allowed us to provide a list containing a large number of TAG unknown to date. The second system allows rapid analysis of olive oil in less than 5 min. This system obeying the guidelines of the International Olive Council can be proposed as a candidate reference method for rapid quality control of olive oils.

La capacité des microalgues à accumuler des quantités importantes de lipides de réserve font de ces organismes de bons candidats pour envisager une production durable de biocarburants (biodiesel). Cependant, des verrous d’ordre technologique et biologique persistent avant d’atteindre une production économiquement viable. Dans le but de mieux comprendre les mécanismes et biosynthèse et d’accumulation des lipides chez les microalgues et de proposer des voies d’amélioration biotechnologiques, nous avons développé deux approches expérimentales complémentaires en utilisant la microalgue Chlamydomonas reinhardtii comme modèle. La première a été de caractériser par des techniques de protéomique et de lipidomique la composition des gouttelettes lipidiques s’accumulant en réponse à une carence en azote. Les données de protéomique nous ont permis de montrer que les gouttelettes lipidiques étaient des structures cellulaires dynamiques impliquées non seulement dans le stockage, mais aussi dans la biosynthèse, la remobilisation et le « trafficking » des lipides. Les protéines identifiées au cours de cette étude nous fournissent des gènes cibles d’intérêt pour mieux comprendre les voies de biosynthèse des triacylglycérols et accroître l’accumulation d’huile. La seconde approche, de génétique formelle, a consisté à rechercher puis à caractériser des mutants isolés à partir d’une banque de mutants d’insertion de C. reinhardtii. Deux mutants d’intérêt, l’un affecté dans la composition en acides gras (crfad7) et l’autre capable d’accumuler des lipides en l’absence de stress (coa1, pour constitutive oil accumulator 1), ont été isolés. / The ability of microalgae to accumulate high amounts of reserve lipids makes these organisms good candidates for the production of sustainable biofuel (biodiesel). However, both technological and biological bottlenecks remain to be overcome before profitable production is reached. With the aim to better understand lipid metabolic pathways in microalgae and further propose new strategies for biotechnological improvement, we have developed two complementary experimental approaches in the model microalga Chlamydomonas reinhardtii. As a first approach, we performed a proteomic and lipidomic characterization of oil bodies isolated from nitrogen-deprived cells. Based on proteomic data, we have concluded that oil bodies are dynamic structures involved not only in the storage, but also in oil biosynthesis, degradation and lipid homeostasis. The proteins identified in this study should provide useful targets for genetic studies aiming at increasing our understanding of triacylglycerol synthesis and further improve intracellular oil accumulation. The second approach, based on the development of a forward genetic screen, aimed at searching and further characterizing mutants isolated from a C. reinhardtii insertion library. Two mutants of interest, one affected in the fatty acid composition (crfad7), the other (coa1, for constitutive oil accumulator 1) able to accumulate reserve lipids in the absence of stress, have been isolated. The crfad7 mutant, affected in the expression of the unique ω3 fatty acid desaturase present in the C. reinhardtii genome, has been complemented and subjected to extensive phenotypical characterization.

The role of dietary fatty acids and their subsequent effects on metabolism has received considerable attention in mammalian species. It is becoming increasingly clear that fatty acids have metabolic roles over and above their influence on energy density of the diet. Recent studies have linked changes in the fatty acyl composition of the plasma membrane, induced by the dietary fat profile, to alterations in both lipid and glucose metabolism. These dietary induced changes have profound effects on insulin action, glucose transport and enzyme activity that regulate triglyceride and fatty acid synthesis, factors that ultimately influence protein and lipid deposition of animals. Because of their high growth rate, broiler chickens have a high requirement for energy and the use of triglycerides as a major energy source has resulted in a fat carcass. A change in the glucose-insulin balance has been suggested as being the main reason for differences in adiposity between broilers selected for fatness or leanness. The hypotheses of this thesis is based on the finding that dietary polyunsaturated fatty acids (PUFA�s) increase the sensitivity of muscle tissue to insulin and this would presumably augment insulin-stimulated glucose uptake into muscle cells. Therefore, increasing the capacity of broiler muscle tissue to utilise glucose as its principal energy substrate would reduce the bird�s reliance on triglycerides and this inturn would result in a leaner carcass. The aims of this study are firstly to explore the role that dietary PUFA�s from the n-3 and n-6 series have on the growth and body composition of broiler chickens and secondly to determine the relationships between dietary fatty acid profile, tissue insulin sensitivity and lipid deposition. Because dietary fatty acids have been implicated in the modulation of hormones important for the growth and development of animals, a third aim of this thesis is to determine the effects of dietary n-3 and n-6 polyunsaturated fatty acids on pituitary and adrenal sensitivity. Since the modulation of metabolism by dietary fatty acids has been attributed to changes in the fatty acyl composition of the plasma membrane, the final aim of this study will be to investigate and characterise the molecular species of the breast muscle choline and ethanolamine phospholipids. Feeding either fish oil a source of n-3 PUFA�s or sunflower oil a source of n-6 PUFA�s fatty acids to broiler chickens resulted in a significant (P<0.01) reduction in the abdominal fat pad mass and a modest increase in breast muscle mass when compared to broilers fed edible tallow. Associated with the changes in carcass composition was an alteration in energy substrate utilisation. This was reflected by lower respiratory quotients and reduced triglyceride and insulin concentrations for the chickens fed the two PUFA diets. Coupled to the shift in energy metabolism was a significant (P<0.05) increase in the proportion of PUFA�s incorporated into the abdominal fat pad and breast muscle. The dietary fat supplements resulted in the incorporation of specific fatty acid subtypes. Feeding fish oil significantly increased the proportion of long-chain n-3 PUFA�s whereas feeding sunflower oil significantly increased the proportion of long-chain n-6 PUFA�s compared to tallow feeding whose tissues were dominated by a higher proportion of saturated fatty acids. It was further shown that dietary n-3 and n-6 PUFA�s enhanced glucose /insulin action. Feeding either fish oil or sunflower oil to broiler chickens increased insulin action when examined by an intravenous glucose tolerance test. The maximal insulin release in response to the glucose infusion was higher in the tallow fed group compared to either the sunflower oil or fish oil groups. To estimate the disappearance rate of glucose from the plasma and its incorporation into tissues, 2-deoxy-D-3H glucose was infused into each chicken. There were no significant differences in the clearance rate of 2-deoxy-D-3H glucose from the plasma. However, when measured under steady state conditions, the labelled glucose incorporation into the breast muscle was greater in birds fed fish oil compared to either tallow or sunflower oil feeding. The dietary fatty acid induced increase in insulin action suggests that the sensitivity of muscle cells to insulin was enhanced. This modulation of tissue sensitivity by dietary fatty acids was also shown to occur at the level of the pituitary. To provide an estimate of pituitary sensitivity, bolus GnRH and GHRH infusions were given on different days to chickens fed the three dietary treatments. Feeding sunflower oil (n-6 PUFA�s) increased the level of GH that was released in response to the GHRH infusion when compared to birds fed either tallow or fish oil (n-3 PUFA�s). This dietary fatty acid modulation appears to be specific to certain pituitary cell types as there was no effect on LH secretion following the GnRH infusion. Dietary fatty acid modulation of endocrine gland sensitivity is particular to the gland type. Although the dietary treatments mediated a distinct pattern in pituitary sensitivity to GHRH infusion, these same three diets did not influence adrenal sensitivity, as there was no difference in the corticosterone profile following either ACTH or CRF infusion. The previously observed physiological changes for the three dietary groups was expected to be positively correlated to an alteration of the plasma membrane phospholipids induced by the dietary fatty acids. Supplementation with fish oil (n-3 PUFA�s) significantly increased levels of both eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) into the choline (PC) and ethanolamine (PE) breast muscle phospholipids compared to either sunflower oil (n-6 PUFA�s) or tallow supplementation. The increase in n-3 PUFA incorporation was associated with a corresponding decrease in the proportion of arachidonic acid (AA; 20:4n-6) an event that would presumably alter substrate availability for the 1- and 2-series eicosanoids. However, feeding sunflower oil or tallow gave a molecular species profile that was remarkably similar in both fatty acid subtype and proportion. This suggests that the plasma membrane dynamics would be similar for these two dietary groups. Therefore, it is appears that factors other than a change in the fatty acyl- composition of the plasma membrane may be responsible for modifying the physiology of the broiler.

The role of dietary fatty acids and their subsequent effects on metabolism has received considerable attention in mammalian species. It is becoming increasingly clear that fatty acids have metabolic roles over and above their influence on energy density of the diet. Recent studies have linked changes in the fatty acyl composition of the plasma membrane, induced by the dietary fat profile, to alterations in both lipid and glucose metabolism. These dietary induced changes have profound effects on insulin action, glucose transport and enzyme activity that regulate triglyceride and fatty acid synthesis, factors that ultimately influence protein and lipid deposition of animals. Because of their high growth rate, broiler chickens have a high requirement for energy and the use of triglycerides as a major energy source has resulted in a fat carcass. A change in the glucose-insulin balance has been suggested as being the main reason for differences in adiposity between broilers selected for fatness or leanness. The hypotheses of this thesis is based on the finding that dietary polyunsaturated fatty acids (PUFA�s) increase the sensitivity of muscle tissue to insulin and this would presumably augment insulin-stimulated glucose uptake into muscle cells. Therefore, increasing the capacity of broiler muscle tissue to utilise glucose as its principal energy substrate would reduce the bird�s reliance on triglycerides and this inturn would result in a leaner carcass. The aims of this study are firstly to explore the role that dietary PUFA�s from the n-3 and n-6 series have on the growth and body composition of broiler chickens and secondly to determine the relationships between dietary fatty acid profile, tissue insulin sensitivity and lipid deposition. Because dietary fatty acids have been implicated in the modulation of hormones important for the growth and development of animals, a third aim of this thesis is to determine the effects of dietary n-3 and n-6 polyunsaturated fatty acids on pituitary and adrenal sensitivity. Since the modulation of metabolism by dietary fatty acids has been attributed to changes in the fatty acyl composition of the plasma membrane, the final aim of this study will be to investigate and characterise the molecular species of the breast muscle choline and ethanolamine phospholipids. Feeding either fish oil a source of n-3 PUFA�s or sunflower oil a source of n-6 PUFA�s fatty acids to broiler chickens resulted in a significant (P<0.01) reduction in the abdominal fat pad mass and a modest increase in breast muscle mass when compared to broilers fed edible tallow. Associated with the changes in carcass composition was an alteration in energy substrate utilisation. This was reflected by lower respiratory quotients and reduced triglyceride and insulin concentrations for the chickens fed the two PUFA diets. Coupled to the shift in energy metabolism was a significant (P<0.05) increase in the proportion of PUFA�s incorporated into the abdominal fat pad and breast muscle. The dietary fat supplements resulted in the incorporation of specific fatty acid subtypes. Feeding fish oil significantly increased the proportion of long-chain n-3 PUFA�s whereas feeding sunflower oil significantly increased the proportion of long-chain n-6 PUFA�s compared to tallow feeding whose tissues were dominated by a higher proportion of saturated fatty acids. It was further shown that dietary n-3 and n-6 PUFA�s enhanced glucose /insulin action. Feeding either fish oil or sunflower oil to broiler chickens increased insulin action when examined by an intravenous glucose tolerance test. The maximal insulin release in response to the glucose infusion was higher in the tallow fed group compared to either the sunflower oil or fish oil groups. To estimate the disappearance rate of glucose from the plasma and its incorporation into tissues, 2-deoxy-D-3H glucose was infused into each chicken. There were no significant differences in the clearance rate of 2-deoxy-D-3H glucose from the plasma. However, when measured under steady state conditions, the labelled glucose incorporation into the breast muscle was greater in birds fed fish oil compared to either tallow or sunflower oil feeding. The dietary fatty acid induced increase in insulin action suggests that the sensitivity of muscle cells to insulin was enhanced. This modulation of tissue sensitivity by dietary fatty acids was also shown to occur at the level of the pituitary. To provide an estimate of pituitary sensitivity, bolus GnRH and GHRH infusions were given on different days to chickens fed the three dietary treatments. Feeding sunflower oil (n-6 PUFA�s) increased the level of GH that was released in response to the GHRH infusion when compared to birds fed either tallow or fish oil (n-3 PUFA�s). This dietary fatty acid modulation appears to be specific to certain pituitary cell types as there was no effect on LH secretion following the GnRH infusion. Dietary fatty acid modulation of endocrine gland sensitivity is particular to the gland type. Although the dietary treatments mediated a distinct pattern in pituitary sensitivity to GHRH infusion, these same three diets did not influence adrenal sensitivity, as there was no difference in the corticosterone profile following either ACTH or CRF infusion. The previously observed physiological changes for the three dietary groups was expected to be positively correlated to an alteration of the plasma membrane phospholipids induced by the dietary fatty acids. Supplementation with fish oil (n-3 PUFA�s) significantly increased levels of both eicosapentaenoic acid (EPA; 20:5n-3) and docosahexaenoic acid (DHA; 22:6n-3) into the choline (PC) and ethanolamine (PE) breast muscle phospholipids compared to either sunflower oil (n-6 PUFA�s) or tallow supplementation. The increase in n-3 PUFA incorporation was associated with a corresponding decrease in the proportion of arachidonic acid (AA; 20:4n-6) an event that would presumably alter substrate availability for the 1- and 2-series eicosanoids. However, feeding sunflower oil or tallow gave a molecular species profile that was remarkably similar in both fatty acid subtype and proportion. This suggests that the plasma membrane dynamics would be similar for these two dietary groups. Therefore, it is appears that factors other than a change in the fatty acyl- composition of the plasma membrane may be responsible for modifying the physiology of the broiler.

Dietary lipids determine the incorporation and fatty acid composition of the organism, especially in organs most susceptible to its modulating action on membrane phospholipids esterified fatty acids, such as the liver. Ketogenic diets, widely used to treat various disorders, often use long-chain triacylglycerols, however, medium chain triglycerides are considered an alternative source, since they are digested, absorbed, oxidized and removed from circulation more rapidly than long chain triacylglycerols. In addition, it lead to the formation of ketone bodies quickly, without the influence of hormonal or nutritional factors. To contribute to the discussion, this paper presents a review chapter, entitled Mitochondrial dysfunction in nonalcoholic fatty liver disease, which gathers information on oxidative stress and consequent mitochondrial dysfunction in the context of liver disease. It also presents an original article, entitled Fatty acid profile in liver tissue of rats subjected to ketogenic diet rich in trienantin, which discusses the effects of a ketogenic diet based on trienantin, a homogeneous triacylglycerol composed of enantic (heptanoic) acid, or based on soybean oil on this profile. The livers of animals fed the ketogenic diet showed a significant reduction in the concentration of monounsaturated fatty acids, 9-hexadecenoic and 9-octadecenoic compared to control group, suggesting a potential reduction in the activity of the enzyme Stearoil-CoA Desaturase 1. Additionally, the group fed trienantin showed increased levels of octadecanoic acid compared to control group, which corroborated the hypothesis of decreased enzyme activity in this group. Thus trienantin in ketogenic proportions exerted modulatory effects similar to soybean oil, widely used conventional oil, despite the differences in fatty acid composition between these two fat sources. The study showed that rats fed a ketogenic diet experienced modulation of fatty acid profile of hepatic lipids and that in the conditions set forth herein, this modulation was due especially to the nature of the ketogenic diet itself and not to the type of oil used in their preparation. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Os lipídios dietéticos determinam a incorporação e a composição em ácidos graxos do organismo, especialmente em órgãos mais susceptíveis a sua ação moduladora sobre os ácidos graxos esterificados a fosfolipídios de membrana, como, por exemplo, o fígado. Dietas cetogênicas, amplamente utilizadas para o tratamento de diversas desordens, normalmente utilizam triacilgliceróis de cadeia longa, porém, triacilgliceróis de cadeia média são considerados uma fonte alternativa, visto que são digeridos, absorvidos, oxidados e removidos da circulação mais rapidamente que os de cadeia longa. Além disso, levam à formação de corpos cetônicos de forma acelerada, não sofrendo a influência de fatores hormonais ou nutricionais. Visando contribuir com a discussão, a presente dissertação apresenta um capítulo de revisão, intitulado Disfunção mitocondrial na doença do fígado gorduroso não alcoólica, que reúne informações sobre o estresse oxidativo e a conseqüente disfunção mitocondrial, no contexto desta doença hepática. E, um artigo original, intitulado Perfil em ácidos graxos do tecido hepático de ratos submetidos à dieta cetogênica rica em trienantina, que discute os efeitos de dieta cetogênica à base de trienantina, um triacilglicerol homogêneo do ácido enântico (heptanóico), ou de óleo de soja, sobre tal perfil. Os fígados dos animais alimentados com as dietas cetogênicas apresentaram redução significativa na concentração de ácidos graxos monoinsaturados, 9-hexadecenóico e 9-octadecenóico, quando comparados ao grupo Controle, sugerindo uma possível redução na atividade da enzima Estearoil-CoA Dessaturase 1. Adicionalmente, o grupo alimentado com trienantina apresentou aumento dos níveis de ácido octadecanóico, quando comparado ao grupo Controle. Assim, a trienantina, em proporções cetogênicas, exerceu efeito modulador similar ao do óleo de soja, um óleo convencional largamente utilizado, apesar das diferenças na composição em ácidos graxos entre essas duas fontes lipídicas. Neste estudo constatou-se que os ratos alimentados com dieta cetogênica sofreram modulação do perfil em ácidos graxos dos lipídios hepáticos e que, nas condições aqui estabelecidas, tal modulação se deveu especialmente à natureza cetogênica da dieta per si e não ao tipo de óleo empregado em sua elaboração.

Les microalgues accumulent de grandes quantités d’huile, et sont de bons candidats pour la production de biocarburants. Mais des verrous techniques et biologiques doivent être levés pour une production rentable. Augmenter la teneur en huile par cellule et découvrir des protéine clés du métabolisme des triglycérides sont des objectifs importants. Nous avons mis en place une approche de génétique classique ciblée sur l’isolement de mutants d’insertion affectés dans la remobilisation des lipides de réserves suite à la re-supplémentation en azote après carence. Nous avons mis au point un protocole de criblage à haut débit basé sur la semi-quantification de ces lipides qui nous a permis d’isoler plus de 30 mutants. Nous avons identifié les loci d’insertion pour certains en utilisant la méthode Genome Walker. Le marqueur de résistance à l’antibiotique a été trouvéé dans des gènes codant pour des kinases, une protéine de type polycystine avec répétitions d’un domaine homologue de type lipoxygénase, des protéines du métabolisme de l’amidon, ou encore une méthyltransférase. Ces mutants forment un set de candidats devant être validés par complémentation pour une meilleure compréhension du métabolisme des lipides. Nous avons vu que la plupart des mutants défectueux dans la remobilisation des lipides de réserve sont aussi affectés dans celle de l’amidon. Ce lien potentiel entre les 2 processus est renforcé par le fait que dans 2 mutants connus de synthèse de l’amidon nous avons pu mettre en évidence un défaut de remobilisation des triglycérides. Ainsi, nous avons montré un lien d’interdépendance entre les dégradations des 2 formes majoritaires de réserves carbonées des microalgues. / Microalgae are able to accumulate high amounts of oil reserves, which make them promising candidates for biofuel production. Nevertheless, some technical and biological bottlenecks have to be overcome before a profitable industrial production. Increasing oil content per cell and discovering key proteins of oil metabolism is a major goal. We took a forward genetic approach and focused on isolating insertional mutants affected in oil remobilization following nitrogen resupply after a starvation phase. We setup and developed a medium- to highthroughput semi-quantitative oil content screening protocol, which has enabled isolation of >30 mutants. We identified the insertion loci in some of these mutants through the “genome walker” PCR-based method. The antibiotic marker was found to be inserted in genes encoding various proteins including serine-threonine kinases, a polycystin-related protein containing repetitions of a lipoxygenase homology domain, an E3 ubiquitin ligase, a starch metabolism protein and a methyltransferase. Mutants isolated provide a first set of candidate genes that remain to be validated by complementation and should contribute to a better understanding of lipid homeostasis in green microalgae. During the course of this work, we observed that most mutants defected in oil remobilization were also impaired in starch degradation. The occurrence of a link between the degradation of starch and oil was further strengthened by the fact that in two known starch-less mutants the oil remobilization process was found to be defected. This is the first evidence of an interdependency between the degradation processes of the major types of carbon reserves in microalgae.

Submitted by Maike Costa (maiksebas@gmail.com) on 2017-06-21T12:55:32Z No. of bitstreams: 1 arquivototal.pdf: 4313605 bytes, checksum: edb0f819c041917885291efd9ed6923a (MD5) / Made available in DSpace on 2017-06-21T12:55:32Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 4313605 bytes, checksum: edb0f819c041917885291efd9ed6923a (MD5) Previous issue date: 2016-02-02 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Terminalia catappa linn also known as castanets is belonging to the family of combretaceae and is found throughout the Brazilian coast. Although it is not a native species, it has ease germination and high resistance to salinity that lead it to become invasive. The fruits of castanets collected around UFPB campus showed almonds rich in oil with yield around 26% obtained by mechanical extraction. The triacylglycerides in the oil have showed a direct influence in the physical and chemical properties as acid value, iodine, peroxide, specific gravity and viscosity rheological factor. The results of these analyzes proved that the samples were in good condition of conservation. Furthermore, the results obtained from analytical thermogravimetry technics showed good thermooxidative stability to decomposition of the samples. The gas chromatography analysis coupled with mass spectrometer suggested that the palmitate, elaidate, linoleate and stearate as major methyl esters constituents of the triacylglycerides responsible for the evaluated properties.These esters were confirmed by the most important spectroscopic techniques to discuss structural elucidation of organic molecules such as FTIR, 1H and 13C NMR, COSY and HSQC. / A Terminália catappa linn também conhecida como castanhola, pertencente à família das combretaceae, é encontrada em todo litoral brasileiro. Apesar de não ser uma espécie nativa, sua facilidade em germinar e alta resistência à salinidade, tornou-a invasiva. Os frutos coletados desta espécie nas imediações da UFPB, apresentam amêndoas ricas em óleo e o rendimento obtido por extração mecânica foi em torno de 26 %. Os triacilglicerídeos que o compõe influenciam diretamente nos parâmetros físico e químicos avaliados como índice de acidez, iodo, peróxido, massa especifica e o fator reológico viscosidade. Estas análises comprovam que a amostra estudada apresentou bom estado de conservação. Este resultado foi corroborado com as técnicas analíticas de termogravimetria e através da oxidação acelerada por fluxo de calor, evidenciando que a amostra em estudo apresenta boa estabilidade em relação à decomposição termo-oxidativas. A análise em cromatografia gasosa acoplada ao espectrômetro de massas sugere os ésteres metílicos majoritários palmitato, elaidato, linoleato e estearato como constituintes dos triacilglicerídeos responsáveis pelas propriedades avaliadas. Estes foram confirmados por técnicas espectroscópicas como infravermelho, RMN 1H, RMN 13C, COSY e HSQC que são as mais importantes quando se discute sobre elucidação estruturais de moléculas orgânicas.

Orientador: Marcos Nogueira Eberlin / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Química / Made available in DSpace on 2018-08-17T04:42:07Z (GMT). No. of bitstreams: 1 Simas_RosineideCosta_D.pdf: 5859215 bytes, checksum: bfba079c3f7e2a4ec510da69e7858d40 (MD5) Previous issue date: 2010 / Resumo: A caracterização química de óleos de forma rápida, eficiente e com mínimo preparo de amostras é um desafio para a química analítica moderna. Este estudo abrangeu a caracterização química de 15 amostras de óleos vegetais e 6 amostras de petróleo, e demonstrou a versatilidade da técnica de espectrometria de massas (MS) nesta área. A caracterização de óleos vegetais foi feita por uma técnica de ionização ambiente desenvolvida no Brasil, denominada Easy Ambient Sonic-Spray Ionization (EASI) e utilizando como analisador de massas um equipamento monoquadrupolar, que possui resolução unitária. Foi obtido, para os óleos vegetais, a sua composição de triacilglicerídeos (TAG), ácidos graxos livres e hidroperóxidos utilizando apenas uma gota de óleo e nenhum preparo de amostra. A análise direta de óleos vegetais possibilitou ainda propor um método quantitativo por EASI-MS para determinação do teor de triacilglicerídeos, que demonstrou concordância com os métodos de referência. Já a caracterização do petróleo empregou a altíssima exatidão e resolução da espectrometria de massas por ressonância ciclotrônica de íons com transformada de Fourier, Fourier Transform Ion Cyclotron Resonance Mass Spectrometry (FT-ICR MS) com as técnicas de ionização EASI e Electrospray (ESI). O presente trabalho representou ainda a primeira tese de doutorado no Brasil em petroleômica, descrevendo um método de análise que dispensa uma série de procedimentos analíticos dispendiosos e demorados, de maneira a conferir um alto grau de refinamento e desempenho à análise de petróleo. Foram estudados os compostos polares do petróleo que contêm os heteroátomos nitrogênio (N), enxofre (S) e oxigênio (O) na sua composição, sendo possível caracterizá-los por distribuição em porcentagem de classes de heteroátomos, número de carbonos e insaturações por double bond equivalents (DBE). Foi apresentada uma proposta de estudo de precisão para a metodologia de petroleômica-MS por meio da estimativa de repetitividade e reprodutibilidade, além do estudo comparativo de eficiência das técnicas de ionização ESI e EASI / Abstract: The chemical characterization of oils in a fast way and with minimal sample preparation is a challenge for modern analytical chemistry. The present study comprised the chemistry characterization of 15 vegetal oil samples and 6 samples of petroleum, demonstrating the versatility of the mass spectrometry (MS) technique in this area. Vegetal oils characterization was performed by an ambient ionization technique developed in Brazil, named Easy Ambient Sonic-Spray Ionization (EASI), and using a monoquadrupole mass analyzer with unitary resolution. EASI-MS detected for vegetable oils the composition of triacylglycerols (TAG), free fat acids and hydroperoxides using only one drop of oil and without sample preparation. The direct analysis provided by EASI-MS allowed us to propose also a quantitative method based on EASI MS data to determine the level of triacylglycerols of the vegetal oils. This method was in agreement with reference methods. For petroleum characterization, we used ultra-high accuracy and resolution of the Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) with EASI and electrospray (ESI) ionization. The present work describes results from the first doctorate thesis in Brazil dealing with petroleomic MS, a method of analysis that precludes a series of expensive and time-consuming analytical procedures and that can confer a high degree of refinement and performance to petroleum analysis. We have studied polar components of the petroleum containing nitrogen (N), sulphur (S) and oxygen (O) as heteroatoms in their composition. These components were classified according to the percentage distribution of heteroatoms, carbon number and unsaturations, as well as double bond equivalent (DBE). The study also evaluated precision for petroleomic MS via estimations of repeatability and reproducibility. The efficiencies of ESI and EASI for the petroleum analysis were also compared / Doutorado / Quimica Analitica / Doutor em Ciências

Orientador: Antonio Jose de Almeida Meirelles / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-05T02:09:23Z (GMT). No. of bitstreams: 1 Rabelo_Juliana_M.pdf: 2293566 bytes, checksum: 7592ac89859a7882bd6ce62626eb622a (MD5) Previous issue date: 2005 / Resumo: Com o objetivo de predizer e descrever a viscosidade de diferentes compostos graxos e suas misturas, este trabalho desenvolveu um procedimento de cálculo baseado em métodos de contribuição de grupos. Um novo modelo para predição da viscosidade de substâncias graxas puras foi desenvolvido, inspirado no modelo proposto por CERIANI e MEIRELLES (2004) para predição da pressão de vapor. O presente trabalho ajustou parâmetros de contribuição de grupos para o modelo proposto, tomando como base dados experimentais de compostos graxos (ácidos graxos, álcoois graxos, ésteres graxos e triacilgliceróis) presentes na literatura. Utilizando o método MARQUARD, o ajuste dos parâmetros envolveu a minimização de uma função objetivo que representava o somatório dos desvios relativos entre os valores experimentais e calculados de viscosidade dinâmica de compostos graxos puros. Tal processo foi realizado através de programas desenvolvidos no SAS ® e de planilhas eletrônicas em EXCEL, nas quais podiam ser feitas análises ponto a ponto dos ajustes. O trabalho envolveu ainda a determinação de um conjunto extenso de dados experimentais de viscosidade de misturas graxas, a caracterização química de óleos vegetais utilizados no teste da capacidade preditiva do modelo e teste dos modelos de estimativa de densidade e de viscosidade de misturas com parâmetros disponíveis na literatura. Confirmada a sua eficiência, este modelo será de interesse para o desenvolvimento de processos e projetos de equipamentos industriais, uma vez que dados de viscosidade de compostos graxos são escassos na literatura / Abstract: This work aims to describe and predict the viscosity of different fatty compounds and their mixtures by developing a predictive model based on the concept of group contribution. A new model developed in this work was based in the method suggested by CERIANI e MEIRELLES (2004) for vapor pressure prediction. All the group parameters have been adjusted based on experimental data for fatty acids, fatty alcohols, fatty esters and triacylglycerols. Using the method of MARQUARD, the adjustment of parameters have minimized an objective function which is based on the sum of the relative deviation between the experimental and calculated values of the dynamic viscosity of fatty compounds. The softwares used for this work was SAS ® and EXCEL. This work has also determined an extensive experimental data bank of viscosities of fatty compounds mixtures. Some models of densities and viscosities found in the literature have been tested. The method suggested in this work can be a valuable tool for the design of industrial equipments of the oil industry, since such experimental data are relatively scarce in the literature / Mestrado / Mestre em Engenharia de Alimentos

Orientador: Daniel Barrera Arellano / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-19T21:15:47Z (GMT). No. of bitstreams: 1 Fernandes_GabrielDeschamps_M.pdf: 1854677 bytes, checksum: e57833484e8f9ad4a136187bed59d8d2 (MD5) Previous issue date: 2012 / Resumo: Os fosfolipídios são definidos como o grupo de moléculas que contém um grupamento fosfato. Por apresentarem características anfipáticas, este grupo de moléculas se organiza naturalmente em bicamadas, originando as membranas dos seres vivos. Industrialmente são capazes de estabelecer interfaces óleo/água, possibilitando a formação e estabilização de emulsões. Este grupo de moléculas é bastante diverso quimicamente, sendo os principais componentes a fosfatidilcolina, fosfatidiletanolamina, fosfatidilserina, fosfatidilinositol, ácido fosfatídico e esfingomielina. A determinação e quantificação desses compostos é bastante laboriosa tanto nos meios industriais como acadêmicos, envolvendo, entre outras, etapas de digestão ácida e incineração. A espectrometria de massas desponta como uma técnica bastante favorável à análise de lipídios, englobando desde estudos clínicos até de biocombustíveis. Mais recentemente, as técnicas de espectrometria de massas com ionização ambiente facilitaram o acesso a este tipo de tecnologia, diminuindo os custos de implantação e principalmente de operação. A ionização ambiente por sonic-spray (EASI, easy ambient sonic-spray ionization) denota-se como uma técnica adequada à análise de lipídios, uma vez que não aplica alta voltagem e alta temperatura, prevenindo, portanto possíveis degradações destas moléculas. Este trabalho teve como objetivo, estudar a ionização de fosfolipídios (PL) e triacilgliceróis (TAG) frente à técnica EASI-MS, bem como, estudar a viabilidade técnica da caracterização de lecitinas comerciais por meio da técnica EASI-MS. Quanto à ionização dos lipídios, foi possível observar, nas condições de estudo, que dentro de uma mesma classe (PL ou TAG) a intensidade de ionização diminui com o aumento da cadeia dos ácidos graxos e aumenta com o aumento das insaturações. Para o estudo de caracterização foram utilizadas seis amostras de lecitina de soja comercial, obtidas por diferentes processos. As amostras foram diluídas em clorofórmio e submetidas à análise de EASI-MS, nos modos positivo e negativo. Nos espectros de EASI(+)-MS, os íons mais representativos foram os íons correspondentes à fosfatidilcolina e aos triacilgliceróis, enquanto que, nos espectros de EASI(-)-MS os íons mais representativos corresponderam à fosfatidiletanolamina, aos ácidos graxos livres e aos glicofosfolipídios. A técnica EASI-MS mostrou-se eficiente na caracterização das lecitinas comerciais. Sendo uma técnica rápida e que não exige preparo de amostra / Abstract: Phospholipids are defined as the group of molecules containing a phosphate grouping. As they have amphipathic characteristics, this group of molecules naturally organizes bilayer, origin the membranes of living organism and are able to establish an industrial oil / water interface, allowing the formation and stabilization of emulsions. This group of molecules is very chemically different; the main components are phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, phosphatidic acid and sphingomyelin. The determination and quantification of these compounds is very laborious for the academic and industrial circles, involving, among others, several steps, like acid digestion and incineration. Mass spectrometry is emerging as a very favorable tool of lipids analysis, since clinical and biofuel studies. Recently, the techniques of ambient mass spectrometry have facilitated the access to this type of technology, reducing deployment costs and especially the operation. Easy ambient sonic-spray ionization (EASI) denotes as a suitable technique to analyze the lipids, since it does not apply high voltage and high temperature, and thereby prevent possible degradation of these molecules. This work aimed to study the ionization of phospholipids (PL) and triacylglycerols (TAG) in EASIMS technique, as well as studying the technical feasibility of the characterization of commercial lecithins by EASI-MS. On the lipid ionization, it was observed, under the conditions of the study, that within the same class (TAG or PL) the ionization intensity decreases with increasing of fatty acids chains and increases with increasing of unsaturation. For characterization studies were used six samples of commercial soy lecithin, obtained by different processes. Samples were diluted in chloroform and analyzed for EASI-MS in positive and negative ion modes. In the spectra of EASI (+)- MS, the most representing ions are corresponding to triglycerides and phosphatidylcholine, whereas in the spectra of EASI (-)-MS the most representative ions correspond to the phosphatidylethanolamine, the free fatty acids and glicophospholipidios. The EASI-MS technique was efficient in the characterization of commercial lecithins. As a fast technique and does not require sample preparation / Mestrado / Tecnologia de Alimentos / Mestre em Tecnologia de Alimentos