Yes / We investigated the protective role of myricetin bulk and nanoforms, against reactive oxygen species (ROS)-induced oxidative stress caused by hydrogen peroxide and tertiary-butyl hydro peroxide in lymphocytes in vitro from healthy individuals and those from pre-cancerous patients suffering with monoclonal gammopathy of undetermined significance (MGUS). The change in intracellular reactive oxygen species was measured once cells were treated with myricetin bulk forms and nanoforms with and without either hydrogen peroxide or tertiary-butyl hydro peroxide co-supplementation. The direct and indirect antioxidant activity of myricetin was spectrofluometrically measured using the fluorescent dye 2',7'-dichlorofluorescin diacetate and using the Comet assay, respectively. Hydrogen peroxide (50 µM) and tertiary-butyl hydro peroxide (300 µM) induced a higher level of reactive oxygen species-related DNA damage and strand breaks. Addition of myricetin nanoform (20 µM) and bulk (10 µM) form could, however, significantly prevent hydrogen peroxide- and tertiary-butyl hydro peroxide-induced oxidative imbalances and the nanoform was more effective. Glutathione levels were also quantified using a non-fluorescent dye. Results suggest that myricetin treatment had no significant effect on the cellular antioxidant enzyme, glutathione. The current study also investigates the effect of myricetin on the induction of double-strand breaks by staining the gamma-H2AX foci immunocytochemically. It was observed that myricetin does not induce double-strand breaks at basal levels rather demonstrated a protective effect.

Yes / 2-Amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP) is a central dietary mutagen, produced when proteinaceous food is heated at very high temperatures potentially causing DNA strand breaks. This study investigates the protective potential of a well-researched flavonoid, myricetin in its bulk and nano-forms against oxidative stress induced ex vivo/in vitro by PhIP in lymphocytes from pre-cancerous monoclonal gammopathy of undetermined significance (MGUS) patients and those from healthy individuals. The results from the Comet assay revealed that in the presence of myricetin bulk (10 µM) and myricetin nano (20 µM), the DNA damage caused by a high dose of PhIP (100 µM) was significantly (P < 0.001) reduced in both groups. However, nano has shown better protection in lymphocytes from pre-cancerous patients. Consistent results were obtained from the micronucleus assay where micronuclei frequency in binucleated cells significantly decreased upon supplementing PhIP with myricetin bulk (P < 0.01) and myricetin nano (P < 0.001), compared to the PhIP treatment alone. To briefly determine the cellular pathways involved in the protective role of myricetin against PhIP, we studied gene expression of P53 and ATR kinase (ATM- and Rad3-related), using the real-time PCR technique.

Yes / The present study investigated the genoprotective and genotoxic effects of myricetin bulk (10 μM) and nano forms (20 μM) in the lymphocytes from pre-cancerous, monoclonal gammopathy of unknown significance (MGUS) patients and healthy individuals using the Comet and micronucleus assays. The study also evaluated the effect of myricetin on P53 expression levels, using the Western blot technique. Results showed that throughout the in-vitro treatment, lymphocytes from the patients group had higher levels of baseline DNA damage compared to the healthy group. Myricetin in both forms induced significant DNA damage, only at higher concentrations (>40 μM). The micronucleus assay showed a significant reduction (P < 0.01) in the frequency of micronuclei in mono-nucleated cells in the patient group treated with the nano form of myricetin at the non-toxic dose of 20 μM. There was a significant increase in both gene and protein P53 levels in lymphocytes isolated from healthy individuals and pre-cancerous patients. These results suggested a protective effect of myricetin and indicated its nutritional supplement potential for protection against cancer development among patients suffering from MGUS. / This study was kindly funded by Mr Nasir Qayyum, Bradford, UK.

No description available.

Submitted by Rosivalda Pereira (mrs.pereira@ufma.br) on 2017-05-22T16:51:30Z No. of bitstreams: 1 SamiraAbdala.pdf: 3051742 bytes, checksum: 816c5c802400bcf5a75deb6a99de0617 (MD5) / Made available in DSpace on 2017-05-22T16:51:30Z (GMT). No. of bitstreams: 1 SamiraAbdala.pdf: 3051742 bytes, checksum: 816c5c802400bcf5a75deb6a99de0617 (MD5) Previous issue date: 2017-04-07 / Fundação de Amparo à Pesquisa e ao Desenvolvimento Científico e Tecnológico do Maranhão (FAPEMA) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPQ) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Platelets, the blood cells involved in maintaining hemostasis, play a key role in the development of acute ischemic coronary, cerebrovascular events and are critically involved in the thrombosis process. In response to vascular injury, changes in blood flow or chemical stimuli, platelets trigger three functional mechanisms: adhesion, activation and aggregation. After platelet capture, a rapid stabilization of adhesion is required for thrombus formation to occur. Platelet activation results from conformational changes dependent of the protein disulfide isomerase (PDI), so that it has recently been proposed as a molecular target in platelet antiaggregant activity. The use of plant species rich in phenolic compounds as a source of bioactive substances is a promising strategy for the development of new therapeutic alternatives for thromboembolic diseases. Previously, we have shown that Syzygium cumini (L.) Skeels leaf contains multiple polyphenols, which support its use for antiplatelet purposes. Therefore, this study sought to evaluate the effects of polyphenol-rich extract (PESc) from S. cumini leaf on platelet activation and aggregation, as well as on PDI reductase activity. Platelet-rich plasma from healthy volunteers (n=5) was incubated with PESc (10-1000 μg/mL), for 25 min, before activation with ADP, thrombin or PMA. To analyze PESc effect on integrin αIIbβ3 activation, flow citometry protocols were conducted in washed platelets pre-treated with PESc (10-1000μg/mL) and activated with thrombin before tagging with PAC–1 antibody. Finally, PESc (0.1-100 μg/mL) effects on PDI reductase activity were assessed in absence or presence of polyphenolic standards gallic acid, myricetin and quercetin. PESc dose-dependently inhibited platelet aggregation despite the agonist used, even though lower agonist concentration potentiated PESc inhibitory effects to a maximal 77% inhibition at 2.5 μM ADP. Similarly, PESc dose-dependently reduced the proportion of activated αIIbβ3 molecules per platelet up to one third of control at 1000 μg/mL. These effects correlated with the strong inhibitory action of PESc on PDI activity, an effect synergically augmented in presence of standards. Therefore, our data show that PESc reduces platelet aggregation and activation, probably through PDI inhibition, strengthening its prominent antiplatelet activity. / As plaquetas, células sanguíneas envolvidas na manutenção da hemostase, exercem uma função essencial no desenvolvimento de eventos isquêmicos agudos coronarianos, cerebrovasculares e estão criticamente envolvidas no processo de formação da trombose. Em resposta à lesão vascular, às alterações no fluxo sanguíneo ou a estímulos químicos, as plaquetas desencadeiam três mecanismos funcionais: adesão, ativação e agregação. Após a captura da plaqueta, uma rápida estabilização da adesão é necessária para que ocorra a formação do trombo. A ativação plaquetária resulta de alterações conformacionais dependentes da proteína dissulfeto isomerase (PDI), de tal modo, que recentemente foi proposto o seu uso como alvo molecular na atividade antiagregante plaquetária. O uso de espécies vegetais ricas em compostos fenólicos como fonte de substâncias bioativas apresenta-se como uma estratégia promissora para o desenvolvimento de novas alternativas terapêuticas das doenças tromboembólicas. Anteriormente, temos mostrado que as folhas de Syzygium cumini (L.) Skeels contém múltiplos polifenóis, que apoiam a sua utilização para fins antiplaquetários. Portanto, este estudo procurou avaliar os efeitos do extrato rico em polifenóis (ERP) da folha de S. cumini sobre a ativação e agregação plaquetária, bem como, sobre a atividade redutase da PDI. Para tanto, plasma rico em plaquetas de voluntários saudáveis foi incubado com ERP (10 - 1000μg/mL), durante 25 min, antes da ativação com ADP, trombina ou PMA. Para analisar o efeito de ERP sobre a ativação da integrina αIIbβ3, os protocolos de citometria de fluxo foram conduzidos em plaquetas lavadas pré-tratadas com ERP (10 -1000 μg/mL) e ativadas com trombina antes da marcação com anticorpo PAC-1. Finalmente, os efeitos de ERP (0,1-100 μg/mL) na atividade redutase da PDI foram avaliados na ausência ou presença de padrões polifenólicos de ácido gálico, miricetina e quercetina. ERP inibiu a agregação plaquetária dependente da dose apesar do agonista utilizado, embora uma menor concentração de agonistas potencializasse os efeitos inibitórios de ERP até uma inibição máxima de 77% a 2,5 μM de ADP. De modo semelhante, o ERP reduziu a dose proporcionalmente a proporção de moléculas de αIIbβ3 ativadas por plaquetas até um terço do controle a 1000μg/mL. Estes efeitos correlacionaram-se com a forte ação inibitória de ERP na atividade da PDI, um efeito sinergicamente aumentado na presença de padrões. Portanto, nossos dados mostram que o ERP reduz a agregação e ativação plaquetária, provavelmente através da inibição da PDI, fortalecendo sua proeminente atividade antiplaquetária.

Cancer is one of the leading causes of death across the globe which needs appropriate and cost-effective treatment. Several recent studies have suggested that dietary intake of various flavonoids such as myricetin have a protective effect against different types of cancers and cardiovascular diseases. The present study was conducted to investigate the genoprotective and genotoxic effects of myricetin nano and bulk forms on the lymphocytes from pre-cancerous and multiple myeloma cancer patients compared to those from healthy individuals. Also, to investigate the protective potential of myricetin bulk and nano against the oxidative stress produced in vitro by 2- amino-1-methyl-6 phenylimidazo [4, 5-b] pyridine and reactive oxygen species- induced DNA damage using the Comet assay, micronucleus assay, cellular reactive oxygen species and glutathione detection assay, Western blotting, real-time polymerase chain reaction and immunofluorescence. Lymphocytes from the patient groups showed significantly higher levels of basal DNA damage compared to the lymphocytes from healthy individuals which was observed throughout the in vitro treatment. Myricetin in both forms has not induced any significant DNA damage in all of the investigative groups at selective lower concentrations; in fact, the results demonstrate a reduction in DNA damage upon treating with myricetin nano in lymphocytes from pre-cancerous patients demonstrated by significant reduction in micronuclei formation in mononucleated cells. DNA repair capacity of myricetin bulk and nano was determined by co-treating the drugs with hydrogen peroxide. Myricetin significantly reduced the oxidative stress related damage caused by hydrogen peroxide, where myricetin nano seemed to be more effective employing the Comet assay. In the presence of myricetin bulk and nano, the damaging effects of 2- amino-1-methyl-6 phenylimidazo [4,5-b] pyridine were considerably decreased, where myricetin nano was more effective. This could be because nanoparticles have a larger surface area which could improve their reactivity and also the reduction in size of the particles could improve the anti-cancer properties of this compound. Myricetin has shown genoprotective and anti-oxidant effects by demonstrating the potential to reduce DNA damage caused by over-production of reactive oxygen species and oxidative stress. It has also shown anti-cancer potential in the lymphocytes from multiple myeloma patients by regulating the apoptosis related proteins, dependent on oxidative stress. Therefore, this study suggests that myricetin supplementation in our regular diet with enhanced bioavailability could have potential health beneficial effects and possibly protect against various diseases including cancer.

This work concerns the amount of certain phenols in some of the representatives of families Alliaceae, Lamiaceae and Apiaceae. Phenols in plants are widely represented and highly concentrated. Flavonoids are one of the smallest, yet quite significant classes of phenols. Ingestion of food containing flavonoids can prevent certain diseases such as Arteriosclerosis. cardiovascular and tumor diseases. This work focuses, for their special biological effects, on five flavonoids: Kaempferol, Quercetin, Myricetin, Apigenin and Luteolin. For determining the content of phenols a method of High-performance liquid chromatography (HPLC) was used. Three representative of the family Alliaceae, two representatives of the family Lamiaceae and three representatives of the family Apiaceae were analysed. Only edible parts of the plants were used for the analysis and the amount of phenols was compared in certain plants planted in beddings to the plants planted in greenhouses. At first, the qualitative representantion of phenols was ascertained by the HPLC method. The result is chromatographic profiles, which were then used in calculating the amount of particular phenols. Then the total amounts of kaempferol, quercetin, myricetin, apigenin and luteolin were found out thanks to HPLC method. The biggest amount of total kaempferol was determined in petroselium hortense planted in beddings (588 mg/kg of fresh sample) and in a greenhouse (340 mg/kg of fresh sample). The biggest amount of quercetin was determined in red onion planted in a greenhouse (773 mg/kg of fresh sample) and in ocimum basilicum planted in beddings (535 mg/kg of fresh sample). The biggest amount of apigenin was determined in petroselium hortense planted in a greenhouse (1790 mg/kg of fresh sample) and in petroselium chrispum planted in beddings (3690 mg/kg of fresh sample).

[ES] La actual problemática medioambiental está demandando sistemas de producción más sostenibles, como los de la agricultura ecológica. Una buena alternativa para estos sistemas son las variedades tradicionales de pimiento, con rasgos de calidad característicos y de amplia variabilidad genética, fundamental en los programas de mejora, concretamente para la adaptación a la producción ecológica. En la presente tesis se ha estudiado la calidad nutricional y organoléptica del cultivo del pimiento (Capsicum sp.) en los dos estados de madurez comercial y bajo diferentes prácticas culturales (sistemas ecológico y convencional). Se han analizado los principales compuestos nutricionales de los frutos de pimiento: el ácido ascórbico, los compuestos fenólicos y los carotenoides; así como la fracción volátil, y se ha estudiado su variación en una amplia colección varietal. Adicionalmente, se ha estudiado la fertilidad biológica del suelo a través de las actividades enzimáticas de la fosfatasa alcalina y la catalasa en interacción con el cultivo del pimiento bajo los sistemas de cultivo convencional y ecológico. El estudio de la calidad nutricional mostró amplios rangos de variación según el factor genotipo, aunque el estado de maduración y el sistema de cultivo también contribuyeron a las diferencias observadas. En general, los frutos maduros presentaron niveles superiores de compuestos bioactivos, especialmente para el ácido ascórbico. Asimismo, el cultivo ecológico proporcionó, en general, niveles más altos de ácido ascórbico y compuestos fenólicos en estado maduro. La notable interacción genotipo × sistema de cultivo para el contenido en ácido ascórbico y fenoles totales, y en menor intensidad para el contenido en flavonoides individuales, permitió la selección de varias accesiones con alto rendimiento en estos compuestos, bajo condiciones de cultivo ecológico en ambos estados de maduración. Por el contrario, los efectos del sistema de cultivo y la interacción genotipo × sistema de cultivo no resultaron significativos en los niveles de carotenoides, cuya variación se debió principalmente al factor genotipo. El estudio de la composición volátil permitió encontrar amplias diferencias cuantitativas y cualitativas para este factor de calidad organoléptica en los frutos de pimiento, dependiendo principalmente del factor genético y del estado de maduración, mientras que el sistema de cultivo afectó en menor medida y significativamente a algunos compuestos volátiles individuales. Se encontraron un total de 124 y 122 volátiles en estado inmaduro y maduro, respectivamente. Los cambios en el perfil volátil con la maduración fueron evidentes y muy diversos según el genotipo. En general, el sistema de cultivo ecológico favoreció la acumulación de compuestos volátiles en ambos estados de maduración y durante el proceso de maduración. Las actividades enzimáticas fosfatasa alcalina y catalasa aumentaron durante la fase de cultivo, con actividades máximas generales en la fase intermedia, especialmente para la fosfatasa alcalina. El sistema de cultivo tuvo una mayor contribución a la variación, resultado significativo y superior para el sistema convencional en fase intermedia para ambas actividades y estando generalmente seguido del factor genotipo, significativo para la fosfatasa alcalina en fase intermedia. La interacción genotipo × sistema de cultivo permitiría seleccionar variedades, con capacidad para que los suelos acumularan mayores valores de las actividades enzimáticas fosfatasa alcalina y catalasa, en el sistema de cultivo ecológico. En definitiva, los resultados de este trabajo proporcionarán información de especial utilidad para programas de investigación en hortalizas de alto valor añadido y programas de mejora de la calidad en agricultura ecológica, así como para el control de autenticidad de variedades tradicionales. / [CA] L'actual problemàtica mediambiental està demandant sistemes de producció més sostenibles, com els de l'agricultura ecològica. Una bona alternativa per a aquests sistemes són les varietats tradicionals de pebrot, amb trets de qualitat característics i d'àmplia variabilitat genètica, fonamental en els programes de millora, concretament per a l'adaptació a la producció ecològica. En la present tesi s'ha estudiat la qualitat nutricional i organolèptica del cultiu del pebrot (Capsicum sp.) en els dos estats de maduresa comercial i sota diferents pràctiques culturals (sistemes ecològic i convencional). S'han analitzat els principals compostos nutricionals dels fruits de pebrot: l'àcid ascòrbic, els compostos fenòlics i els carotenoides; així com la fracció volàtil, i s'ha estudiat la seva variació en una àmplia col·lecció varietal. Addicionalment, s'ha estudiat la fertilitat biològica de terra a través de les activitats enzimàtiques de la fosfatasa alcalina i la catalasa en interacció amb el cultiu del pebrot sota els sistemes de cultiu convencional i ecològic. L'estudi de la qualitat nutricional mostrar amplis rangs de variació segons el factor genotip, encara que l'estat de maduració i el sistema de cultiu també van contribuir a les diferències observades. En general, els fruits madurs van presentar nivells superiors de compostos bioactius, especialment per a l'àcid ascòrbic. Així mateix, el cultiu ecològic va proporcionar, en general, nivells més alts d'àcid ascòrbic i compostos fenòlics en estat madur. La notable interacció genotip × sistema de cultiu per al contingut en àcid ascòrbic i fenols totals, i en menor intensitat per al contingut en flavonoides individuals, va permetre la selecció de diverses accessions amb alt rendiment en aquests compostos, sota condicions de cultiu ecològic en els dos estats de maduració. Per contra, els efectes de el sistema de cultiu i la interacció genotip × sistema de cultiu no van resultar significatius en els nivells de carotenoides, la variació es va deure principalment al factor genotip. L'estudi de la composició volàtil va permetre trobar àmplies diferències quantitatives i qualitatives per aquest factor de qualitat organolèptica en els fruits de pebrot, depenent principalment del factor genètic i de l'estat de maduració, mentre que el sistema de cultiu va afectar en menor mesura i significativament a alguns compostos volàtils individuals. Es van trobar un total de 124 i 122 volàtils en estat immadur i madur, respectivament. Els canvis en el perfil volàtil amb la maduració van ser evidents i molt diversos segons el genotip. En general, el sistema de cultiu ecològic va afavorir l'acumulació de compostos volàtils en els dos estats de maduració i durant el procés de maduració. Les activitats enzimàtiques fosfatasa alcalina i catalasa van augmentar durant la fase de cultiu, amb activitats màximes generals en la fase intermèdia, especialment per a la fosfatasa alcalina. El sistema de cultiu va tenir una major contribució a la variació, resultat significatiu i superior per al sistema convencional en fase intermèdia per a ambdues activitats i estant generalment seguit del factor genotip, significatiu per a la fosfatasa alcalina en fase intermèdia. La interacció genotip × sistema de cultiu permetria seleccionar varietats, amb capacitat perquè els sòls acumulessin grans valors de les activitats enzimàtiques fosfatasa alcalina i catalasa, en el sistema de cultiu ecològic. En definitiva, els resultats d'aquest treball proporcionaran informació d'especial utilitat per a programes d'investigació en hortalisses d'alt valor afegit i programes de millora de la qualitat en agricultura ecològica, així com per al control d'autenticitat de varietats tradicionals. / [EN] The current environmental problem is demanding more sustainable production systems, such as organic farming. A good alternative for these systems are the traditional varieties of pepper, with characteristic quality traits and wide genetic variability, essential in breeding programs, specifically for adaptation to organic production. In this thesis, the nutritional and organoleptic quality of the pepper crop (Capsicum sp.) at the two ripening stages and under different cultural practices (ecological and conventional systems) has been studied. The main nutritional compounds of pepper fruits have been analysed: ascorbic acid, phenolic compounds and carotenoids; as well as the volatile fraction, and its variation has been studied in a wide varietal collection. Additionally, the biological fertility of the soil has been studied through the enzymatic activities of alkaline phosphatase and catalase in interaction with pepper cultivation under conventional and ecological growing systems. The study of nutritional quality showed wide ranges of variation according to the genotype factor, although the ripening stage and the growing system also contributed to the differences observed. In general, fully ripe fruits presented higher levels of bioactive compounds, especially for ascorbic acid. Also, organic farming provided, in general, higher levels of ascorbic acid and phenolic compounds at the fully ripe stage. The remarkable genotype × growing system interaction for the content of ascorbic acid and total phenolics, and to a lesser extent for the content of individual flavonoids, allowed the selection of several accessions with high yield in these compounds, under organic farming conditions at both ripening stages. On the contrary, the effects of the growing system and the genotype × growing system interaction were not significant in carotenoid levels, whose variation was mainly due to the genotype factor. The study of the volatile composition allowed to find wide quantitative and qualitative differences for this organoleptic quality factor in pepper fruits, depending mainly on the genetic factor and the ripening stage, while the growing system affected to a lesser extent and significantly some individual volatile compounds. A total of 124 and 122 volatiles were found at unripe and fully ripe stages, respectively. The changes in the volatile profile with ripening were evident and very diverse depending on the genotype. In general, the organic growing system favoured the accumulation of volatile compounds at both ripening stages and during the ripening process. Alkaline phosphatase and catalase enzymatic activities increased during the growing phase, with overall peak activities in the intermediate phase, especially for alkaline phosphatase. The growing system had a greater contribution to variation, a significant and superior result for the conventional system in the intermediate phase for both activities and being generally followed by the genotype factor, significant for alkaline phosphatase in the intermediate phase. The genotype × growing system interaction would allow the selection of varieties, with the capacity for soils to accumulate higher values of the enzymatic activities alkaline phosphatase and catalase, in the organic growing system. In conclusion, the results of this work will provide information of special use for research programs in vegetables with high added value and quality breeding programs in organic farming, as well as for the authenticity control of traditional varieties. / Authors thank seeds providers included in Table 1, like P.W. Bosland, S. Lanteri, François Jourdan, I. Ruiz de Galarreta and the different Regulatory Boards of the PDOs and GPIs included in this study. Authors also thank LA UNIÓ for the availability of experimental fields and the technical support from Manuel Figueroa, Rafael Hurtado, Antonio Muñoz and Ricard Ballester in the trials. Finally, the advice of Prof. Jaime Prohens in statistical procedures is also thanked. / Ribes Moya, AM. (2020). Selección y mejora de ecotipos de pimiento (Capsicum sp.) para agricultura sostenible y calidad nutricional y organoléptica [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/159886 / TESIS