This diploma thesis deals with the determination of selected antioxidants, namely rosmarinic acid, caffeic acid, cinnamic acid and t-4-hydroxycinnamic acid in aqueous and ethanolic extracts from Melissa officinalis. The aim of the work was to find out whether the above- mentioned antioxidants were present in the extracts prepared at home. A new UHPLC method with UV and MS detection was developed to monitor the content of rosmarinic acid, caffeic acid, cinnamic acid and t-4-hydroxycinnamic acid in the extracts. A BEH C18 column (2,1 mm × 100 mm, particle size 1,7 µm) was selected for separation, the mobile phase consisted of methanol (component A) and 0.1% aqueous formic acid solution at pH 2.6 (component B), the ratio of components being changed according to the gradient program. The method was validated and repeatability, limits of detection and limits of quantification, linearity, yield and robustness were determined after optimization. Analysis of 20 aqueous and 20 ethanolic extracts was performer differing in extraction time, type of solvent and lighting conditions. Based on the comparison of retention time and MS detection, the presence of 3 of 4 selected phenolic acids was confirmed, namely rosmarinic acid, caffeic acid and cinnamic acid in ethanolic extracts. Unfortunately, the presence of...

La cosmétique valorise des extraits de plantes en les intégrant dans des produits de soin. Ces extraits constituent les ingrédients actifs de la formule cosmétique. Les plantes sont diverses, riches et contiennent de nombreux composés d’intérêt biologique. La phytochimie s’intéresse à décrire le contenu métabolique d’extraits de plantes. Mais la caractérisation moléculaire de ces matrices complexes reste un défi majeur aujourd’hui. En effet, les étapes de traitements de données sont longues et laborieuses. Un outil de profilage métabolique, GAINS, a ainsi été développé afin de traiter de façon automatisée les données issues d’analyses en chromatographie liquide couplée à la spectrométrie de masse haute résolution. Il constitue un support au phytochimiste car le traitement automatisé des données permet de gagner du temps par rapport à un traitement manuel. Cet outil associé à une vaste base de données de produits naturels permet d’attribuer des candidats potentiels aux pics détectés. GAINS fait aussi appel à un module de fragmentation in silico afin d’étayer les propositions de candidats. Cela permet de comparer le spectre de fragmentation modélisé des candidats avec le spectre de fragmentation expérimental.L’ensemble des études phytochimiques réalisées pour identifier ou isoler des composés va de pair avec l’étude du potentiel d’effet biologique des extraits sur la peau, organe ciblé par les cosmétiques. Cela sert à découvrir les actions bénéfiques que peut avoir l’extrait. En connaissant le contenu phytochimique, on peut expliquer et rationaliser les tests d’activités biologiques. Le développement d’un actif anti-âge à partir de la salicaire, une plante de la région Centre-Val de Loire, en est l’exemple. / The cosmetic field valorizes plant extracts by integrating them in care products. These extracts constitute the active ingredients of the cosmetic formulation. Plants are diverse, rich and contain numerous compounds of biological interest. Phytochemistry is a way to describe the metabolic content of plant extracts. But molecular characterization of these complex matrices remains a major challenge nowadays. Indeed,steps of data treatment are time-consuming and laborious. In this way, a tool of metabolic profiling, GAINS, has been developed in order to treat in an automatized way data from analyses performed in liquid chromatography coupled with high-resolution mass spectrometry. It constitutes a real support for phytochemists because automatized data treatment allows gaining time compared to manual treatment. This tool, associated with a large database of natural compounds make possible to assign potential candidates to detected peaks. GAINS appeals a module of in silico fragmentation for holding candidates assignments up.This permits to compare modeled spectrum of fragmentation of candidates with experimental spectrum off ragmentation.The whole set of phytochemical studies realized to identify or isolate compounds goes hand in hand with the study of potential biological effects of extracts to the skin, targeted organ by skin-care products. This allows the discovery of beneficial actions that the extract could have. By knowing the phytochemical content, it is possible to explain and rationalize assays about biological activities. The development of an anti-aging ingredient from purple loosestrife, a plant occurring in the region Centre-Val de Loire, is an example of it.

A família Orchidaceae é a maior família entre as monocotiledôneas e representa uma das maiores famílias de plantas floríferas com um número estimado de cerca de 30 mil espécies. A complexidade taxonômica de um dos gêneros dentro dessa família, o Catasetum, tem despertado muito interesse por parte dos cultivadores e pesquisadores em Orchidaceae, especialmente devido aos novos táxons que estão sendo descobertos no gênero. Visando aprimorar a classificação de espécies do gênero, uma ferramenta a ser utilizada é a análise dos constituintes químicos do metabolismo dessas plantas, onde fica evidenciada a necessidade de estudos utilizando métodos analíticos mais sensíveis, rápidas e seletivas, como a análise metabolômica. A obtenção da impressão digital metabólica é a obtenção detalhada dos metabólitos do organismo sob determinada condição, produzindo um cromatograma que é característico daquela planta, como uma impressão digital. Através disso, é possível determinar diferenças metabólicas em amostras, por meio de ferramentas estatísticas. Para o refinamento dos dados, é necessário o uso de softwares específicos que extraem as informações úteis para as análises. Foram analisadas 42 espécies distintas do gênero Catasetum e algumas duplicatas, totalizando 51 amostras por GC-MS e UHPLC-MS(ESI-Orbitrap). As \"impressões digitais\" metabólicas foram processadas nos softwares MetAlignTM e MSClust, e as matrizes geradas foram analisadas por análise de componentes principais (PCA), análise de cluster hierárquico (HCA) e análise discriminante ortogonal por mínimos quadrados parciais (OPLS-DA). Os métodos utilizados para as análises tanto por GC-MS quanto por UHPLC-MS resultaram em dados satisfatórios para análise metabolômica, gerando 57 substâncias voláteis identificadas a partir da fragrância das flores, sendo acetato de nerila, sabineno, ?-mirceno, ?-pineno e ?-pineno as mais frequentes. Os resultados das análises metabolômicas apresentaram proximidade com a filogenia, principalmente em níveis mais específicos. Os grupos gerados pelas análises, de modo geral, foram coerentes quanto à semelhança metabólica entre as espécies que os compõem, sugerindo a possibilidade do uso de impressões digitais metabólicas como uma boa ferramenta para análise quimiotaxonômica / The Orchidaceae family is the largest family among monocotyledons and represents one of the largest families of flowering plants with an estimated 30,000 species. The taxonomic complexity of one of the genera within this family, the Catasetum, has aroused much interest from growers and researchers in Orchidaceae, especially due to the new taxa being discovered in the genus. Aiming to improve the classification of species of the genus, a tool to be used is the analysis of the chemical constituents of the metabolism of these plants, where it is evidenced the need for studies using analytical methodologies more sensitive, fast and effective, such as the metabolomics analysis. Obtaining the metabolic fingerprint is the detailed obtaining of the metabolites of the organism under certain condition, producing a chromatogram that is characteristic of that plant, like a fingerprint. Through this, it is possible to determine metabolic differences in samples, through statistical tools. For the refinement of the data, it is necessary to use specific softwares that extract the useful information from analyzes data. The information generated can be used with different applications. A total of 51 samples were analyzed by GC-MS and UHPLC-MS (ESI-Orbitrap), 42 different species of the genus Catasetum and some duplicates. Metabolic \"fingerprints\" were processed in the MetAlignTM and MSClust softwares, and the generated matrices were analyzed by Principal Component Analysis (PCA), hierarchical cluster analysis (HCA) and partial least squares orthogonal discriminant analysis (OPLS-DA). The methods used for analysis by both GC-MS and UHPLC-MS resulted in satisfactory data for metabolomics analysis, generating 57 volatile substances identified from the flower fragrance, being neryl acetate, sabinene, ?-myrcene, ?-pinene and ?-pinene the most frequent. The results of the metabolic analyzes showed proximity to the phylogeny, mainly in more specific levels. The groups generated by the analyzes, in general, were coherent regarding the metabolic similarity between the species that compose them, suggesting the possibility of the use of metabolic fingerprints as a good tool for chemotaxonomic analysis

O Brasil é um dos principais produtores de frutas, porém esta produção está pautada em frutas na maioria das vezes exóticas. Existe uma diversidade de frutas nativas brasileiras ainda desconhecidas e pouco exploradas. Tendo em vista a grande importância da Myrtaceae no que diz respeito à conservação da Mata Atlântica; o potencial econômico e nutricional da produção de polpa de frutas nativas; e considerando que o processamento de alimentos pode exercer efeitos sobre o nível de nutrientes, particularmente no que tange ao teor de compostos bioativos e sua atividade antioxidante, este trabalho teve como objetivo gerar conhecimento sobre a composição e capacidade antioxidante das polpas de frutas, com o intuito de ampliar seu consumo, potencial de comercialização e agregar valor as mesmas, bem como, avaliar o efeito da radiação gama nas propriedades bioativas, durante o período de armazenamento refrigerado das polpas e, por fim, a identificação e quantificação dos compostos fenólicos. As polpas de uvaia (Eugenia pyriformis Camb.), cambuci (Campomanesia phaea), feijoa (Acca sellowiana), grumixama (Eugenia brasiliensis) e cereja do rio grande (Eugenia involucrata), foram divididas em 5 lotes, 4 deles foram submetidos ao tratamento de irradiação (0, 2, 4, e 6 kGy) e 1 lote a pasteurização (85°C/5min.). Os resultados demonstraram que as polpas possuem quantidades expressivas de compostos com atividade antioxidante, sendo que, a polpa de uvaia destacou-se pelos teores de ácido ascórbico, que variaram de 89,99 mg.100 g-1 na pasteurizada a 64,74 mg.100 g-1 a 6 kGy, e pelos carotenoides, que variaram de 24,7 ?g.g-1 no controle a 15,94 ?g.g-1 a 4 kGy. Na feijoa destacam-se os flavonoides no controle, 10,21 mg de ác. gálico.mL-1, nas irradiadas com 2; 4 e 6 kGy, 9,75; 11,58 e 12,79 mg de ác. gálico.mL-1, respectivamente, e na pasteurizada 12,64 mg de ác. gálico.mL-1. As antocianinas foram encontradas em maiores quantidades nas polpas de grumixama (pasteurizadas, 106,01 mg eq. cianidina-3-glicosideo.L-1, irradiadas a 2, 4 e 6 kGy, 52,93; 32,36 e 48,64, respectivamente e no controle 59,75) e cereja do rio grande (pasteurizadas 136,33 mg eq. cianidina-3-glicosideo.L-1, controle 77,11 mg eq. cianidina-3-glicosideo.L-1 e nas irradiadas 74,91; 77,7 e 69,47 mg eq. cianidina-3-glicosideo.L-1, respectivamente a 2, 4 e 6 kGy). Na polpa de cambuci não foram encontrados teores de flavonoides e antocianinas e apresentaram valores relativamente baixos dos outros compostos estudados. O composto fenólico majoritariamente encontrado na polpa de uvaia foi o ácido gálico, na feijoa os ácidos elágico e quínico, no cambuci, o ácido quínico, na grumixama o ácido elágico e a quercetina e na cereja do rio grande o ácido quínico e a rutina. Apesar da radiação gama ser um eficiente método de conservação e poder ser utilizada sem prejuízo dos compostos bioativos, nas condições estudadas, as doses entre 2 e 6 kGy, não foram efetivas para evitar as perdas dos compostos bioativos de interesse, no período de armazenamento, apresentando resultados inferiores em relação à pasteurização quanto às antocianinas, compostos fenólicos totais e a atividade antioxidante, medida por ABTS e DPPH / Brazil is a major producer of exotic native fruits. There is a diversity of native fruits still unknown and slightly researched. Taking into consideration the great importance of Myrtaceae regarding the conservation of the Atlantic Forest; the economic and nutritive potential of the production of native fruit pulp; and considering that the food processing may affect the level of nutrients, particularly with regard to the bioactive compound content and its antioxidant activity, the purpose of this study is to generate knowledge on the composition and the antioxidant capability of Myrtaceae pulps. The study aims at enlarging the consumption of Myrtaceae pulps, their marketing potential and their value-added, as well as at evaluating the effect of gamma radiation on bioactive properties during the cold storage period of pulps, and finally, the identification and quantification of the phenolic compounds. The pulps of uvaia (Eugenia pyriformis Camb.), cambuci (Campomanesia phaea), feijoa (Acca sellowiana), grumixama (Eugenia brasiliensis) and cereja do rio grande (Eugenia involucrata) were divided into 5 batches, 4 of them were exposed to irradiation (0, 2, 4, e 6 kGy) and one batch pasteurization (85°C/5min.) treatments. The results showed that the pulps have an expressive quantity of compounds with antioxidant activity. Uvaia pulp stood out by the ascorbic acid contents, which decreased from 89,99 mg.100 g-1 in pasteurization to 64,74 mg.100 g-1 by 6 kGy, and by the carotenoids, which varied from 24,7 ?g.g-1 in the control sample to 15,94 ?g.g-1 by 4 kGy. Feijoa presented high levels of flavonoids in the control sample, 10,21 mg of gallic acid.mL-1, in irradiation 2; 4 and 6 kGy, 9,75; 11,58 e 12,79 mg of gallic acid.mL-1, respectively, and in pasteurization 12,64 mg of gallic acid.mL-1. The anthocyanins were found in major quantities in the grumixama pulps (in pasteurization, 106,01 mg eq. cianidina-3-glicosideo.L-1, irradiated by 2, 4 e 6 kGy, 52,93; 32,36 e 48,64, respectively and in the control sample 59,75) and cereja do rio grande (in pasteurization 136,33 mg equ. cianidina-3-glicosideo.L-1, in the control sample 77,11 mg equ. cianidina-3-glicosideo.L-1 and irradiated 74,91; 77,7 and 69,47 mg equ. cianidina-3-glicosideo.L-1, respectively by 2, 4 e 6 kGy). Flavoniods and anthocyanins were not found in cambuci pulps. Also, there were low values of the others researched compounds in cambuci pulps. The phenolic compound majorly found in the uvaia pulp was the gallic acid, in the feijoa was the ellagic and quinic acids, in cambuci was the quinic acid, in the grumixama was the ellagic acid and quercetin, and in the cereja do rio grande was the quinic acid and the rutin. Although the gamma radiation revealed to be an efficient conservation method and may be used without prejudice of bioactive compounds, based on the conditions researched, the doses between 2 and 6 kGy were not effective to prevent losses of bioactive compounds of interest in the cold storage period, presenting lower results in relation to the pasteurization treatment as anthocyanins, total of phenolic compounds and the antioxidant activity measured by ABTS and DPPH

Le diabète de type 2 est souvent associé à un stress oxydant pouvant entrainer plusieurs complications métaboliques. Au Sénégal, en plus du traitement médicamenteux, les plantes médicinales restent encore très utilisées dans la prise en charge de cette pathologie. Le but de ce travail a été de contribuer à la valorisation de la biodiversité sénégalaise et plus particulièrement d’approfondir les connaissances phytochimiques de Parinari macrophylla Sabine, utilisée traditionnellement pour le traitement du diabète. Pour cela, l’activité anti-radicalaire totale et au niveau moléculaire des feuilles et écorces a été déterminée par les méthodes TEAC, ORAC et HPLC - ABTS•+ online. De plus un screening phytochimique (réactions de précipitation, CCM) a été réalisé et a permis de mettre en évidence plusieurs groupes chimiques de composés tels que des flavonoïdes, des tanins, des terpènes, des anthracènes, des saponosides, des hétérosides cardiotoniques et des alcaloïdes. Pour l’identification des composés actifs, la RMN et l’approche par réseau moléculaire (UHPLC-MS), ont permis d’identifier l’acide chlorogénique, l’hyperoside, la procyanidine B2 et d’autres procyanidines. L’étude in vitro réalisée sur un modèle cellulaire RINm5F a montré une faible activité anti-radicalaire des extraits de plante vis-à-vis d’un stress induit à l’HX-XO. Toutefois les composés identifiés dans cette étude sont connus pour leurs effets antioxydants et inhibiteurs sur les enzymes digestives responsables du métabolisme du glucose. Cela pourrait justifier l’utilisation de cette plante dans le traitement du diabète de type 2 par les tradithérapeutes Sénégalais. / Type 2 diabetes is often associated with oxidative stress that can lead to several metabolic complications. In Senegal, in addition to drug treatment, medicinal plants are still widely used in the treatment of this pathology. The aim of this work was to contribute to the enhancement of Senegalese biodiversity and more particularly to improve the phytochemical properties of Parinari macrophylla Sabine, traditionally used for the treatment of diabetes. For this, the total antiradical activity and at the molecular level of leaves and bark has been determined by the TEAC, ORAC and HPLC - ABTS-+ online methods. In addition, a phytochemical screening (precipitation reactions, CCM) was carried out and revealed several chemical groups of compounds such as flavonoids, tannins, terpenes, anthracenes, saponosides, cardiotonic heterosides and alkaloids. For the identification of active compounds, NMR and the molecular network approach (UHPLC-MS), identified chlorogenic acid, hyperoside, procyanidin B2 and other procyanidins. The in vitro study carried out on a RINm5F cellular model showed a low anti-radical activity of plant extracts against HX-XO-induced stress. However, the compounds identified in this study are known for their antioxidant and inhibitory effects on digestive enzymes responsible for glucose metabolism. This could justify the use of this plant in the treatment of type 2 diabetes by Senegalese traditional therapists.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / In recent years, the indiscriminate use of pesticides and non-adoption of good agricultural practices have been evidenced by the results of analysis of pesticide residues in food, such as sweet pepper, one of Brazilian record holders with active ingredients that are banned or above the recommended limits. Thus, the present study aimed to develop and validate a rapid method for the determination of 83 pesticides in sweet pepper using modified QuEChERS method and Ultra-high Performance Liquid Chromatography coupled to tandem Mass Spectrometry (UHPLC-MS/MS). In the sample preparation extraction/partition procedures, and different sorbents for clean-up were tested, such as diatomaceous earth and polymeric sorbent. It was used the 24 factorial design for optimization of the method of extraction, which consisted of the extraction of 10.0 g sweet pepper with 10.0 mL of acetonitrile. In the partition step 1.5 g of NaCl, 4.0 g MgSO4 were added, carrying out shaking and centrifugation. It was carried out the clean-up step of 1.0 ml of extract with 40.0 mg of PSA and 150.0 mg of MgSO4, followed by shaking and centrifugation. The extract was filtered, diluted and analyzed by UHPLC-MS/MS. Most compounds showed linear range (r2 ≥ 0.99).The matrix effect was compensed by using analytical curves prepared in blank matrix-matched. The accuracy of the method was verified by testing recovery at four concentration levels, yielding values between 70.0 and 120.0% for most compounds evaluated. Values of LODm and LOQm were 3 or 7.5 μg kg-1 and 10 or 25 μg kg-1, respectively. RSD values lower than 20% ensured good precision of the method. The method was performed on 20 samples of sweet pepper, in which residues of acetamiprid, azoxystrobin, boscalid, carbendazim, chlorpyrifos ethyl, clothianidin, difenoconazole, fenpropathrin, pyraclostrobin and pyrimethanil were quantified in concentrations ranging from 10 to 294 μg kg-1. The proposed modified QuEChERS method for the determination of 81 from 83 pesticide residues in sweet pepper using UHPLC-MS/MS shown to be effective and can be applied in routine analysis. / Nos últimos anos, o uso indiscriminado de agrotóxicos e a não adoção das boas práticas agrícolas têm sido evidenciados pelos resultados de análises de resíduos de agrotóxicos em alimentos, como o pimentão, um dos recordistas brasileiros com princípios ativos proibidos ou acima dos limites recomendados. Assim, o presente trabalho teve como objetivo desenvolver e validar um método rápido para a determinação de 83 agrotóxicos em pimentão, utilizando método QuEChERS modificado e Cromatografia Líquida de Ultra Eficiência acoplada à Espectrometria de Massas em série (UHPLC-MS/MS). Na etapa de preparo de amostra testou-se procedimentos de extração/partição, além de diferentes sorventes para a etapa de limpeza, como terra diatomácea e sorvente polimérico. Utilizou-se o planejamento fatorial 24 para otimização do método de extração, o qual foi constituído da extração de 10,0 g de pimentão com 10,0 mL de acetonitrila. Na etapa de partição adicionou-se 1,5 g de NaCl e 4,0 g de MgSO4, seguindo-se agitação e centrifugação. Realizou-se a etapa de limpeza de 1,0 mL do extrato com 40,0 mg de PSA e 150,0 mg de MgSO4, seguido de agitação e centrifugação. O extrato foi filtrado, diluído e analisado por UHPLC-MS/MS. A maioria dos compostos apresentou faixa linear (r2≥ 0,99). O efeito matriz foi compensado utilizando-se curvas analíticas preparadas no extrato branco da matriz. A exatidão do método foi verificada com ensaios de recuperação em quatro níveis de concentração, obtendose valores entre 70,0 e 120,0% para a maioria dos compostos avaliados. Valores de LODm e LOQm foram de 3 ou 7,5 μg kg-1 e 10 ou 25 μg kg-1, respetivamente.Valores de RSD inferiores a 20% garantiram a boa precisão do método. A aplicação do método foi realizada em 20 amostras de pimentão, nas quais resíduos de acetamiprido, azoxistrobina, boscalida, carbendazim, clorpirifós etílico, clotianidina, difenoconazol, fempropatrina, piraclostrobina e pirimetanil foram quantificados, em concentrações que variaram de 10 a 294 μg kg-1. O método QuEChERS modificado proposto para a determinação de resíduos de 81 dos 83 agrotóxicos em pimentão utilizando UHPLC-MS/MS mostrou ser eficaz, podendo ser aplicado em análises de rotina.

The use of pesticides in agriculture and urban areas has always been associated with advantages and disadvantages. As benefits, the increase of the productivity of crops and the control of vectors that cause diseases can be highlighted. However, excessive use of these substances has been causing human health damage as well as environmental degradation, especially regarding to water resources. In the present work, a method for the determination of 62 pesticides residues in surface water using rotating disk sorptive extraction (RDSE) as sample preparation and determination by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) was optimized and validated. Several parameters were evaluated during the development of the extraction method: time and rotational disk velocity in the extraction step, types and amounts of sorbents, sample pH, ionic strength, time and velocity of the rotating disk in the desorption step of the analytes and different desorption solvents. The best results were obtained using 50 mL of acidified sample at pH 2.0 and 2.5 g of NaCl. The velocity of rotation disk in the extraction stage was 1600 rpm in a time of 80 min. Inside the disk cavity the polymeric sorbent Oasis® HLB (20 mg) used. The desorption step of the analytes was performed with 3 mL of methanol for 60 min using a rotation velocity of 1600 rpm. The analytical curves showed linearity between 0.05 or 0.1 to 2 μg L-1, with r2 values greater than 0.99 for all compounds. The method LOQ values ranged from 0.05 to 0.1 μg L-1. The method showed suitable accuracy and precision, with recoveries between 70.1 and 119.9% and RSD ≤20% for the spiked levels at 0.05; 0.1; 0.5 and 2 μg L-1. The validated method was applied to 11 surface water samples from different river basins of the Rio Grande do Sul state. Only one sample presented no pesticide residues. The other samples presented residues of at least one of the following pesticides: atrazine, azoxystrobin, clomazone, difenoconazole, epoxiconazole, propoxur, simazine and tebuconazole. From the six hydrographic basins, all presented pesticides residues and the concentrations ranged between 0.06 and 0.35 μg L-1. Therefore, the results indicate that the proposed method is suitable for the analysis of pesticide residues in surface waters, allowing the preparation of several samples simultaneously with low material consumption. / O uso de agrotóxicos tanto na agricultura quanto na área urbana, sempre esteve associado a vantagens e desvantagens. Pode-se destacar como benefícios o aumento da produtividade das lavouras e o controle de vetores causadores de doenças. No entanto, a utilização excessiva destas substâncias vem causando malefícios à saúde humana e ao meio ambiente, principalmente no que diz respeito aos recursos hídricos. No presente trabalho, um método para a determinação de resíduos de 62 agrotóxicos em águas superficiais, utilizando extração sortiva em disco rotatório (RDSE) para o preparo de amostra e determinação por cromatografia líquida de ultra-alta eficiência acoplada à espectrometria de massas em série (UHPLC-MS/MS) foi otimizado e validado. Na otimização do método de extração foram avaliados diversos parâmetros como: diferentes tempos e velocidades de rotação do disco na etapa de extração, tipos e quantidades de sorventes, pH da amostra, força iônica, tempo e velocidade de rotação do disco na etapa de dessorção dos analitos e diferentes solventes de dessorção. Os melhores resultados foram obtidos utilizando 50 mL de amostra acidificada em pH 2 e adicionando 2,5 g de NaCl. A velocidade de rotação do disco na etapa de extração foi de 1600 rpm por um tempo de 80 min. Na cavidade do disco foi utilizado o sorvente polimérico Oasis® HLB (20 mg). A dessorção dos analitos foi realizada com 3 mL de metanol por 60 min usando uma velocidade de rotação de 1600 rpm. As curvas analíticas apresentaram linearidade entre 0,05 ou 0,1 a 2 μg L-1, com valores de r2 maiores que 0,99 para todos os compostos. O limite de quantificação do método variou entre 0,05 e 0,1 μg L-1. O método apresentou exatidão e precisão adequadas, com recuperações entre 70,1 e 119,9% e RSD ≤20% para níveis de fortificação 0,05; 0,1; 0,5 e 2,0 μg L-1. O método validado foi aplicado em 11 amostras de água superficiais de diferentes bacias hidrográficas do estado do Rio Grande do Sul. Apenas uma amostra não apresentou resíduos de agrotóxicos. As demais apresentaram resíduos de pelo menos um dos seguintes agrotóxicos: atrazina, azoxistrobina, clomazona, difenoconazol, epoxziconazol, propoxur, simazina e tebuconazol. Das seis bacias hidrográficas amostradas todas apresentaram resíduos de agrotóxicos e as concentrações variaram entre 0,06 e 0,35 μg L-1. Os resultados indicam que o método proposto é adequado para análise de resíduos de agrotóxicos em águas superficiais, permitindo o preparo de várias amostras simultaneamente com baixo consumo de material.

O Brasil é um dos principais produtores de frutas, porém esta produção está pautada em frutas na maioria das vezes exóticas. Existe uma diversidade de frutas nativas brasileiras ainda desconhecidas e pouco exploradas. Tendo em vista a grande importância da Myrtaceae no que diz respeito à conservação da Mata Atlântica; o potencial econômico e nutricional da produção de polpa de frutas nativas; e considerando que o processamento de alimentos pode exercer efeitos sobre o nível de nutrientes, particularmente no que tange ao teor de compostos bioativos e sua atividade antioxidante, este trabalho teve como objetivo gerar conhecimento sobre a composição e capacidade antioxidante das polpas de frutas, com o intuito de ampliar seu consumo, potencial de comercialização e agregar valor as mesmas, bem como, avaliar o efeito da radiação gama nas propriedades bioativas, durante o período de armazenamento refrigerado das polpas e, por fim, a identificação e quantificação dos compostos fenólicos. As polpas de uvaia (Eugenia pyriformis Camb.), cambuci (Campomanesia phaea), feijoa (Acca sellowiana), grumixama (Eugenia brasiliensis) e cereja do rio grande (Eugenia involucrata), foram divididas em 5 lotes, 4 deles foram submetidos ao tratamento de irradiação (0, 2, 4, e 6 kGy) e 1 lote a pasteurização (85°C/5min.). Os resultados demonstraram que as polpas possuem quantidades expressivas de compostos com atividade antioxidante, sendo que, a polpa de uvaia destacou-se pelos teores de ácido ascórbico, que variaram de 89,99 mg.100 g-1 na pasteurizada a 64,74 mg.100 g-1 a 6 kGy, e pelos carotenoides, que variaram de 24,7 ?g.g-1 no controle a 15,94 ?g.g-1 a 4 kGy. Na feijoa destacam-se os flavonoides no controle, 10,21 mg de ác. gálico.mL-1, nas irradiadas com 2; 4 e 6 kGy, 9,75; 11,58 e 12,79 mg de ác. gálico.mL-1, respectivamente, e na pasteurizada 12,64 mg de ác. gálico.mL-1. As antocianinas foram encontradas em maiores quantidades nas polpas de grumixama (pasteurizadas, 106,01 mg eq. cianidina-3-glicosideo.L-1, irradiadas a 2, 4 e 6 kGy, 52,93; 32,36 e 48,64, respectivamente e no controle 59,75) e cereja do rio grande (pasteurizadas 136,33 mg eq. cianidina-3-glicosideo.L-1, controle 77,11 mg eq. cianidina-3-glicosideo.L-1 e nas irradiadas 74,91; 77,7 e 69,47 mg eq. cianidina-3-glicosideo.L-1, respectivamente a 2, 4 e 6 kGy). Na polpa de cambuci não foram encontrados teores de flavonoides e antocianinas e apresentaram valores relativamente baixos dos outros compostos estudados. O composto fenólico majoritariamente encontrado na polpa de uvaia foi o ácido gálico, na feijoa os ácidos elágico e quínico, no cambuci, o ácido quínico, na grumixama o ácido elágico e a quercetina e na cereja do rio grande o ácido quínico e a rutina. Apesar da radiação gama ser um eficiente método de conservação e poder ser utilizada sem prejuízo dos compostos bioativos, nas condições estudadas, as doses entre 2 e 6 kGy, não foram efetivas para evitar as perdas dos compostos bioativos de interesse, no período de armazenamento, apresentando resultados inferiores em relação à pasteurização quanto às antocianinas, compostos fenólicos totais e a atividade antioxidante, medida por ABTS e DPPH / Brazil is a major producer of exotic native fruits. There is a diversity of native fruits still unknown and slightly researched. Taking into consideration the great importance of Myrtaceae regarding the conservation of the Atlantic Forest; the economic and nutritive potential of the production of native fruit pulp; and considering that the food processing may affect the level of nutrients, particularly with regard to the bioactive compound content and its antioxidant activity, the purpose of this study is to generate knowledge on the composition and the antioxidant capability of Myrtaceae pulps. The study aims at enlarging the consumption of Myrtaceae pulps, their marketing potential and their value-added, as well as at evaluating the effect of gamma radiation on bioactive properties during the cold storage period of pulps, and finally, the identification and quantification of the phenolic compounds. The pulps of uvaia (Eugenia pyriformis Camb.), cambuci (Campomanesia phaea), feijoa (Acca sellowiana), grumixama (Eugenia brasiliensis) and cereja do rio grande (Eugenia involucrata) were divided into 5 batches, 4 of them were exposed to irradiation (0, 2, 4, e 6 kGy) and one batch pasteurization (85°C/5min.) treatments. The results showed that the pulps have an expressive quantity of compounds with antioxidant activity. Uvaia pulp stood out by the ascorbic acid contents, which decreased from 89,99 mg.100 g-1 in pasteurization to 64,74 mg.100 g-1 by 6 kGy, and by the carotenoids, which varied from 24,7 ?g.g-1 in the control sample to 15,94 ?g.g-1 by 4 kGy. Feijoa presented high levels of flavonoids in the control sample, 10,21 mg of gallic acid.mL-1, in irradiation 2; 4 and 6 kGy, 9,75; 11,58 e 12,79 mg of gallic acid.mL-1, respectively, and in pasteurization 12,64 mg of gallic acid.mL-1. The anthocyanins were found in major quantities in the grumixama pulps (in pasteurization, 106,01 mg eq. cianidina-3-glicosideo.L-1, irradiated by 2, 4 e 6 kGy, 52,93; 32,36 e 48,64, respectively and in the control sample 59,75) and cereja do rio grande (in pasteurization 136,33 mg equ. cianidina-3-glicosideo.L-1, in the control sample 77,11 mg equ. cianidina-3-glicosideo.L-1 and irradiated 74,91; 77,7 and 69,47 mg equ. cianidina-3-glicosideo.L-1, respectively by 2, 4 e 6 kGy). Flavoniods and anthocyanins were not found in cambuci pulps. Also, there were low values of the others researched compounds in cambuci pulps. The phenolic compound majorly found in the uvaia pulp was the gallic acid, in the feijoa was the ellagic and quinic acids, in cambuci was the quinic acid, in the grumixama was the ellagic acid and quercetin, and in the cereja do rio grande was the quinic acid and the rutin. Although the gamma radiation revealed to be an efficient conservation method and may be used without prejudice of bioactive compounds, based on the conditions researched, the doses between 2 and 6 kGy were not effective to prevent losses of bioactive compounds of interest in the cold storage period, presenting lower results in relation to the pasteurization treatment as anthocyanins, total of phenolic compounds and the antioxidant activity measured by ABTS and DPPH

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.

I den här studien har metoder för att separera immunoglobulin (IgG) utvecklats med hjälp av kapillärelektrofores (CE) och Ultra högupplösande vätskekromatografi-Mass spektrometri (UHPLC-MS). Vid analysen av peptider från IgG spjälkat av trypsin (IgG TD) detekterades 26 glykopepeptider med UHPLC-MS genom att använda olika gradienter och mobilfaser. Dessutom analyserades fyra olika koncentrations av IgG TD för att testa detektionsgränsen. Resultatet visade att glykopeptider kunde detekteras i alla fyra koncentrationer men antalet detekterade glykopeptider minskade med minskande koncentration. Den sista delen av UHPLC-MS analysen var en selektivitetsanalys där en blandning av bovint serumalbumin spjälkat av trypsin (BSA TD) och IgG TD analyserades i olika koncentrationer. Selektivitetsanalysen visade att ungefär samma antal IgG glykopeptider detekterades som vid analys av IgG TD. Vid CE-analysen användes tre olika kapillärer, varav två av dem var polyvinylalkohol-coated och olika buffertlösningar (BGE) användes. De slutgiltiga resultaten visade 5 toppar som skulle kunna representera IgG gykopeptider men eftersom ingen MS utfördes kunde inga slutsatser dras. Slutligen, användes Matrix-Assisted Laser desorption/ionisation-time-of-flight (MALDI-TOF) för att analysera proverna innan analys och för att jämföra resultaten med UHPLC-MS. / In this study, methods to separate immunoglobulin G (IgG) glycopeptides were developed using capillary electrophoresis (CE) and ultra-high-performance liquid chromatography-Mass spectrometry (UHPLC-MS). In the UHPLC-MS- analysis, 26 glycopeptides were detected during the analysis of trypsin digested IgG (IgG TD) using different gradients and mobile phases. Moreover, a limit of detection study with four different concentrations (0.01, 0.1, 0.5 and 2mg/mL) was performed. Although the number of detected glycopeptides decreased with decreasing concentration, glycopeptides were detected in the 0.01 mg/mL IgG TD sample. Lastly, a selectivity study was performed, in which two trypsin digested Bovine serum albumin (BSA TD) and IgG TD mixtures with different concentrations were analysed. The results showed that approximately the same number glycopeptides were detected as in the IgG sample suggesting that the BSA sample did not interfere with the detection of glycopeptides. Moreover, the CE-analysis included three different capillaries and several background electrolytes (BGEs). Two of the capillaries were polyvinyl alcohol (PVA) coated. The final electropherogram showed five peaks possibly representing IgG glycopeptides but no MS were performed on the CE part leading to no conclusions for the CE-part. Matrix-Assisted Laser Desorption/Ionisation- time-of flight (MALDI-TOF) was performed throughout the project to control the samples and for comparison to the UHPLC-MS analysis.