Dissertations / Theses on the topic 'Hibiscus sabdariffa'

A systematic study on extraction of Hibiscus sabdariffa was carried out for the first time using different solvents (water, methanol, ethyl acetate and hexane) in the presence and absence of formic acid, using different extraction times and temperatures. The extracts were analysed for total phenol content, antioxidant capacity using DPPH, FRAP and TEAC assays, and total monomeric anthocyanin content. In addition, specific anthocyanins were determined using HPLC and LC-MS. The results showed the highest antioxidant capacities were obtained by extracting using water, with or without formic acid, for 10 min at 100 °C. These extracts provided the highest concentrations of cyanidin 3-sambubioside and delphinidin 3-sambubioside. Commercially available herbal teas containing H. sabdariffa were analysed. The study found that contents of total phenols, anthocyanins and antioxidant capacity were higher when using the optimal extraction procedure, suggesting that putative health benefits could be increased by altering processing methods. The partition coefficients (log p) of anthocyanins found in H. sabdariffa, were measured showed that aglycone and glucoside forms of hibiscus anthocyanins behave differently when in the presence of cell wall material. Such behaviour could, in vivo, affect the absorption and bioactivity of these anthocyanins, and therefore, their efficacy. A human crossover study investigated the effect of daily consumption for 8 weeks of a H. sabdariffa juice for 8 weeks on the blood pressure of healthy subjects (n= 29). Cranberry juice was used as the control. A significant reduction was found in systolic blood pressure (but not diastolic) compared to the baseline. No significant effect on blood pressure was seen with cranberry juice. The study suggests that regular consumption of extracts of H. sabdariffa may reduce the risk of cardiovascular disease, on the other hand people with low blood pressure should consumed it very carefully due to hypotensive effect of the extract.

L’oseille de Guinée (Hibiscus sabdariffa L. ) est une plante importante en Afrique de l’Ouest pour son utilisation au niveau nutritionnel, industriel et médical. Cependant, hormis quelques études de phytotechnie, peu de travaux concernent la sélection de cette plante. En outre, aucune étude de régénération par embryogenèse somatique n’est disponible. L’objectif principal de ce travail est de caractériser la collection d’oseille de Guinée en vue à long terme, de mettre en oeuvre un programme de sélection de génotypes vigoureux, haut producteurs et résistants à certaines maladies. Par ailleurs, des perspectives de conservation et de sortie variétale de génotypes élites, nécessitent le développement de méthodes de régénération par culture in vitro. La première partie de l’étude portant sur la caractérisation morphologique et agronomique de la collection d’oseille de Guinée, a confirmé l’organisation de cette espèce en deux variétés sabdariffa et altissima dont les phénotypes sont complémentaires. H. Sabdariffa var. Sabdariffa est caractérisée par une forte vigueur végétative juvénile, une faible productivité de calices et fruits, une sensibilité faible à la photopériode et une couleur rouge des organes. H. Sabdariffa var. Altissima a une forte vigueur végétative adulte, une productivité plus élevée, une grande sensibilité à la photopériode et une couleur verte des organes. Dans une deuxième partie, les variétés sont évaluées pour leurs réactions de défense vis-à-vis des champignons pathogènes (Fusarium oxysporum et Phoma sabdariffae). Des études sur l’impact des attaques sur la croissance des plantes et certains paramètres physiologiques montrent que les individus de la variété sabdariffa sont plus résistants aux deux champignons que ceux de la variété altissima. Aucune incidence n’est observée sur la photosynthèse chez les deux variétés. Suite aux inoculations, la variété sabdariffa accumule une quantité croissante de polyphénols mais ce taux diminue chez la variété altissima. La dernière partie aborde les méthodes utilisées pour induire la formation de cals embryogènes, précédées de la mise au point d’un protocole efficace de décontamination des explants. La technique des couches cellulaires minces transversales (CCMTs) mise en oeuvre sur des plantules, permet d’obtenir des cals embryogènes après l’étude de nombreux facteurs déterminants comme le génotype (sabdariffa altissima), le milieu de culture (MS, DKW), la nature de l’expIant (racine, hypocotyle, feuille cotylédonaïre), le type (auxines, cytokinines) et la teneur des régulateurs de croissance. Les CCMTs d’hypocotyle et de feuilles cotylédonaires mis en culture dans le milieu DKW contenant les combinaisons 1/0,5 (mg/l) et 4/1 (mg/l) de 2,4-D/TDZ constituent les meilleures conditions d’induction de cals embryogènes
Roselle (Hibiscus sabdariffa L. ) is an important crop in West Africa for its nutritional, industrial and medicinal uses. However, except some studies on phytotechny, there are a few report on the selection of this plant. Moreover, no study on thie in vitro regeneration by somatic embryogenesis is available. The main aim of this work is to characterize the collection of roselle for in long-term sight, to implement a programme of selection for vigorous, high yielding and resistant genotypes te certain diseases. Moreover, die conservation ami seeds production ofgood genotypes, require the development of methods of regeneration by in vitro culture. The first part of the study relating to the morphological and agronomical characterization of the collection of roselle, confirmed the organization of this species into two varieties such as sabdariffa and altissima whose phenotypes were complementary. H. Sabdariffa var. Sabdariffa was characterized by a strong early vegetative vigour, a low yielding of calyx and fruits, a low photoperiod susceptibility and a red color of plants. H. Sabdariffa var. Altissima had a strong adult vegetative vigour, a higher yielding, a high photoperiod susceptibility and a green color of the plant. In a second part, the varieties were evaluated for their defense reactions against to pathogenic fungi (Fusarium oxysporum and Phoma sabdariffae). Studies ou the impact of the attacks on the growth of the plants and certain physiological parameters, showed that the individuals of the sabdariffa variety were more resistaut to both fungi than those of the altissima variety. No incidence was observed on both varieties photosynthesis. Following die inoculations, the sabdariffa variety accumulated an increasing quantity of polyphenols but this rate decreases in the altissima variety. The last part describes the methods used to induce the formation of embryogenic calli, preceded by the development of an efficient protocol of decontamination of explants. The technique of the thin Transversal Cell Layers (tTCLs) implemented on seedlings, enabled to obtain some embryogenic calli after the study of many determining factors such as the genotype (sabdariffa, altissima), the culture medium (MS, DKW), the nature of the explant (root, hypocotyle, cotyledon), the type (auxines, cytokinines) and the content of the growth regulaters. The tTCLs from hypocotyl and cotyledon in culture medium DKW containing combinations 1/0,5 (mg/l) and 4/1 (mg/l) of 2,4-D/TDZ constituted the best conditions of induction of the embryogenic calli

Les calices séchés d’Hibiscus sabdariffa L. sont traditionnellement utilisés en Afrique pour la production de boissons par macération dans l’eau. Cette matière première, riche en anthocyanes, est aussi connue dans l’industrie agro-alimentaire comme source de colorants naturels. Toutefois, la couleur des extraits obtenus évoluent rapidement au cours du temps ce qui nuit au développement de ces produits. Afin de mieux caractériser et de mieux comprendre l’instabilité de la couleur, ce travail propose une étude cinétique et mécanistique de la dégradation des anthocyanes durant la conservation d’un extrait d’H. sabdariffa. La caractérisation biochimique de l’extrait a notamment permis d’identifier les polyphénols majoritaires (delphinidine 3-O-sambubioside Del-3Sb, cyanidine 3-O-sambubioside Cya-3Sb, acides caféoylquiniques) et à mis en évidence la présence de métaux connus comme potentiels catalyseurs d’oxydation (Fe, Cu, Mn). Les cinétiques de dégradation des anthocyanes dans l’extrait ont été suivies par CLHP-DAD entre 4 et 37°C pendant 60 j. Elles peuvent être représentées par un modèle d’ordre 1. La Del-3Sb (Ea= 90 kJ.mol-1) est plus sensible à l’augmentation de la température que la Cya-3Sb (Ea= 80 kJ.mol-1). L’influence de différents facteurs sur la dégradation des anthocyanes a été étudiée dans des solutions modèles simplifiées. La constante de vitesse de dégradation de la Del-3Sb en solution modèle contenant du Fe (III), Cu (II) et Mn (II) à des concentrations similaires à celles de l’extrait, est proche de celle mesurée dans l’extrait. Celle de la Cya-3Sb est 40 % plus faible que dans l’extrait. L’augmentation de la concentration en Fe de 1 à 13 mg.L-1 multiplie environ 3 fois les vitesses de dégradation des anthocyanes. L’ajout d’acide chlorogénique à une solution modèle contenant du Fe augmente la vitesse de dégradation de la Cya-3Sb de 42 %. Un effet inverse est observé dans le cas de Del-3Sb (diminution de 47%). La teneur en oxygène dissous n’a pas d’influence sur les vitesses de dégradation. Deux mécanismes de dégradation des anthocyanes ont été mis en évidence : le mécanisme de scission, représentant une voie minoritaire (10% des anthocyanes dégradés) avec production de phloroglucinaldéhyde, d’acide gallique (issu de Del-3Sb) et protocatéchique (issu de Cya-3Sb) et un mécanisme de condensation avec formation de polymères bruns qui est favorisé par l’augmentation de la concentration en Fe et le mélange Fe, Cu et Mn. Les conclusions de l’ensemble du travail aboutissent à des perspectives sur l’étude de l’impact d’autres composés présents dans l’extrait (sucres, acides aminés) ainsi qui l’identification et la quantification des polymères bruns
Dried calyx of Hibiscus sabdariffa L. are traditionally used in Africa for beverage production by maceration in water. This raw material, rich in anthocyanins, is also known in the food industry as a source of natural dyes. However, the color of extracts obtained quickly changes over time which affects the development of these products. To better characterize and understand the instability of color, this work provides a mechanistic and kinetic study of the degradation of anthocyanins during the conservation of an extract from H. sabdariffa. Biochemical characterization of the extract allowed to identify predominant polyphenols (delphinidin 3-O-sambubioside Del-3Sb, cyanidin 3-O-sambubioside Cya-3Sb and caffeoylquinic acids) and highlighted the presence of metals known as potential oxidation catalysts (Fe, Cu, Mn). Kinetics of degradation of anthocyanins in the extract were followed by HPLC-DAD between 4 and 37 ° C for 60 days. It can be represented by a first order model. Del-3Sb (Ea = 90 kJ mol-1) is more sensitive to temperature increase than Cya-3Sb (Ea = 80 kJ.mol-1). Influence of different factors on the degradation of anthocyanins was studied in simplified model media. Rate constant of degradation (k) of Del-3Sb on model media containing Fe (III), Cu (II) and Mn (II) at similar concentrations to those of the extract was similar to that measured in the extract. For Cya-3Sb it was 40% lower than in the extract. Increasing Fe concentration of 1 to 13 mg.L 1 increased by about 3-fold the k of anthocyanins. Adding chlorogenic acid in a model media containing Fe increased k of Cya-3Sb 42%. Opposite effect was observed for Del-3Sb (47% decrease). Dissolved oxygen content has no influence on k of anthocyanins. Two mechanisms of degradation of anthocyanins have been identified: the scission mechanism, representing a minority pathway (10% of degraded anthocyanins) with production of phloroglucinaldehyde, gallic acid (from Del-3Sb) and protocatechuic (from Cya -3Sb). The other mechanism is condensation with formation of brown polymers, it is promoted by increasing of concentration of Fe was and mixture of Fe, Cu and Mn. Conclusions led to perspectives on study of impact of other compound in the extract (sugar, amino-acids) as well as identification and quantification of brown polymers

Hibiscus Sabdariffa (HS) is a plant of the Malvacee family commonly cultured in tropical and subtropical countries. It is mainly known as the main ingredient for the preparation of cold drink called Karkadè. Calices and leaves of HS plant are also used in folk medicine thanks to their antioxidant and anti-inflammatory properties. In recent years, HS has also gained great interest as a possible antitumoral agent. In the present PhD project, we evaluated the antitumoral effects of HS against multiple mye-loma cells in vitro. Multiple myeloma is the most frequent hematological malignancy world-wide. In recent years, new drugs have increased the survival expectancy of patients. Despite this, new therapeutic approaches are necessary, especially for high multiple myeloma hetero-geneity and for relapsed or refractory multiple myeloma. The project was organized in three distinct phases: 1- Evaluation of antitumoral effects of HS against RPMI 8226 human multiple myeloma cells. We demonstrated by MTT and Trypan blue assays that a total HS extract (HSE) and one of its fraction obtained by liquid-liquid extraction (HSEC) were able to impair cell viability of human multiple myeloma RPMI 8226 in a dose and time dependent manner. HSE cell viability reduction was due to a cytostatic action, while HSEC was more cytotoxic and induced a caspase dependent apoptosis. Moreover, both HSE and HSEC impaired cell migration and invasion of RPMI 8226 cells in a Boyden chamber as-say. We also demonstrated in in vitro model of neurotoxicity (dorsal root ganglia model) that HSE and HSEC concentrations used in our experiments were not neurotoxic. In RPMI 8226 cells autophagy and proteasome activity were impaired by both HSE and HSEC. MAPK p38 activation was observed in the first 6h of treatment, while ERK 1 and ERK 2 activation occurred between 16 and 48h. 2- Evaluation of combinations between Bortezomib (BTZ) and HSE or HSEC against RPMI 8226 multiple myeloma cells. We evaluated several combinations between BTZ and HSE or HSEC (simultaneous, not-simultaneous treatment and pretreatment) using MTT assay to assess their effectiveness. Among all evaluated combinations, only the 24h BTZ 1nM pretreatment followed by HSE or HSEC treatment resulted more effective than BTZ or HSE or HSEC single treatment in reducing cell viability. This combination was not neurotoxic in the dorsal root ganglia model. 3- Isolation and characterization of HSEC molecules responsible of the antitumoral ef-fect. Using a bioguided method, we isolated several fractions from HSEC. Fractions were obtained by flash column chromatography or by resin use. Molecular characterization was performed by HPLC, NMR or mass spectrometry, while biological activity was screened in human multiple myeloma RPMI 8226 cells by MTT and proteasome activity assay. We isolated three fractions with a first fractionating process that keep their activ-ity on RPMI 8226 cells, but that are not completely pure. In a second fractionating, we isolated a highly pure fraction with a significant activity on RPMI 8226 cell viability. We also characterized this fraction by both a molecular and biological point of view. In conclusion, we demonstrated the antitumoral effect of HS in vitro against RPMI 8226 multiple myeloma cells. We also found a new therapeutic combination between BTZ and HS that enhanced their antitumoral effect when compared to single treatments. Moreover, we also isolated one of the molecules that are involved in the antitumoral effect of HS. The results of my PhD project showed that HS could be a promising chemotherapeutic agent, but further studies are still needed.
Hibiscus Sabdariffa (HS) is a plant of the Malvacee family commonly cultured in tropical and subtropical countries. It is mainly known as the main ingredient for the preparation of cold drink called Karkadè. Calices and leaves of HS plant are also used in folk medicine thanks to their antioxidant and anti-inflammatory properties. In recent years, HS has also gained great interest as a possible antitumoral agent. In the present PhD project, we evaluated the antitumoral effects of HS against multiple mye-loma cells in vitro. Multiple myeloma is the most frequent hematological malignancy world-wide. In recent years, new drugs have increased the survival expectancy of patients. Despite this, new therapeutic approaches are necessary, especially for high multiple myeloma hetero-geneity and for relapsed or refractory multiple myeloma. The project was organized in three distinct phases: 1- Evaluation of antitumoral effects of HS against RPMI 8226 human multiple myeloma cells. We demonstrated by MTT and Trypan blue assays that a total HS extract (HSE) and one of its fraction obtained by liquid-liquid extraction (HSEC) were able to impair cell viability of human multiple myeloma RPMI 8226 in a dose and time dependent manner. HSE cell viability reduction was due to a cytostatic action, while HSEC was more cytotoxic and induced a caspase dependent apoptosis. Moreover, both HSE and HSEC impaired cell migration and invasion of RPMI 8226 cells in a Boyden chamber as-say. We also demonstrated in in vitro model of neurotoxicity (dorsal root ganglia model) that HSE and HSEC concentrations used in our experiments were not neurotoxic. In RPMI 8226 cells autophagy and proteasome activity were impaired by both HSE and HSEC. MAPK p38 activation was observed in the first 6h of treatment, while ERK 1 and ERK 2 activation occurred between 16 and 48h. 2- Evaluation of combinations between Bortezomib (BTZ) and HSE or HSEC against RPMI 8226 multiple myeloma cells. We evaluated several combinations between BTZ and HSE or HSEC (simultaneous, not-simultaneous treatment and pretreatment) using MTT assay to assess their effectiveness. Among all evaluated combinations, only the 24h BTZ 1nM pretreatment followed by HSE or HSEC treatment resulted more effective than BTZ or HSE or HSEC single treatment in reducing cell viability. This combination was not neurotoxic in the dorsal root ganglia model. 3- Isolation and characterization of HSEC molecules responsible of the antitumoral ef-fect. Using a bioguided method, we isolated several fractions from HSEC. Fractions were obtained by flash column chromatography or by resin use. Molecular characterization was performed by HPLC, NMR or mass spectrometry, while biological activity was screened in human multiple myeloma RPMI 8226 cells by MTT and proteasome activity assay. We isolated three fractions with a first fractionating process that keep their activ-ity on RPMI 8226 cells, but that are not completely pure. In a second fractionating, we isolated a highly pure fraction with a significant activity on RPMI 8226 cell viability. We also characterized this fraction by both a molecular and biological point of view. In conclusion, we demonstrated the antitumoral effect of HS in vitro against RPMI 8226 multiple myeloma cells. We also found a new therapeutic combination between BTZ and HS that enhanced their antitumoral effect when compared to single treatments. Moreover, we also isolated one of the molecules that are involved in the antitumoral effect of HS. The results of my PhD project showed that HS could be a promising chemotherapeutic agent, but further studies are still needed.

The beverage industry is tending towards the production of healthier, low calorie beverages containing only naturally derived ingredients. The use of these ingredients presents an opportunity for a rise in functional beverages but could also present beverage quality challenges. Using Hibiscus sabdariffa (Roselle) extracts, known for its functional properties, this research investigates some quality considerations that may result due to relevant changes to the production, storage and consumption of the beverage to suit clean label considerations. These issues relate to processing, calories reduction and stability. Several research questions have been generated under these categories and addressed in subsequent chapters of the thesis. From a Roselle beverage perspective, pasteurisation (in-bottle, 85oC for 20 minutes) and cold storage (below 4oC) were satisfactory for processing of the beverage. Stevia Rebaudioside A (SRA) was explored as a sugar replacer and displayed promising results such as improving the sensory attributes of the unsweetened Roselle extract. In physical and chemical tests, it matched the attributes of unsweetened beverages. However, it did not perform as well as sugar in either consumer studies and when combined with spice flavours. The spices (cinnamon, ginger and cloves) used for their flavouring and preserving properties did not improve consumer liking of the beverages in general but cinnamon demonstrated stabilising effects on the anthocyanins in a sugar sweetened Roselle beverage through a suspected co-pigmentation mechanism. Furthermore, cinnamon and ginger were deemed congruent with the Roselle flavour profile based on their performance in the consumer study. Further investigative work on spice and sweetener synergies and the understanding of their mechanisms were recommended.

O hibisco (Hibiscus sabdariffa L.) além de possuir propriedades antioxidantes e antimicrobianas, é utilizado como planta medicinal e alimento funcional nos países tropicais. Através de Testes de Diluição em Sistema de Tubos Múltiplos determinou-se a Intensidade de Atividade de Inibição Bacteriana (IINIB/Bacteriostasia) e a Intensidade de Atividade de Inativação Bacteriana (IINAB/Bactericidia) de extrato alcoólico de dois acessos de hibisco, a saber: Palmares do Sul/RS e Porto Alegre/RS sobre inóculos padronizados de Enterococcus faecalis (ATCC 19433), Escherichia coli (ATCC 11229), Salmonella Enteritidis (ATCC 11076) e Staphylococcus aureus (ATCC 25923). Paralelamente, o teor de polifenóis totais e de antocianinas nos cálices e nos frutos com sementes do hibisco foi determinado. A atividade antimicrobiana do extrato alcoólico de cálices, em ambos os acessos, apresentou diferença positiva significativa quando relacionada ao extrato alcoólico dos frutos com sementes. Salmonella Enteritidis foi a bactéria mais sensível ao extrato alcoólico de cálices do hibisco enquanto a mais resistente foi Staphylococcus aureus, independente da variável acesso, considerando somente a estrutura vegetal. Em relação ao extrato alcoólico dos frutos com sementes, Escherichia coli demonstrou a maior sensibilidade e Staphylococcus aureus a maior resistência. Os valores de polifenóis totais e de antocianinas do extrato alcoólico de cálices apresentaram diferença significativa entre si e foram superiores ao extrato alcoólico dos frutos com sementes. Possivelmente existe uma relação direta entre a concentração de antocianina e a atividade antibacteriana em diferentes estruturas vegetais do hibisco.
The hibiscus (Hibiscus sabdariffa L.) has antioxidant and antimicrobial properties and it is utilized as functional food and medicinal plant in tropical countries. Through of Dilution Testing in Multiple Tubes System, it were determined the intensity of bacterial inhibition activity (IINIB/ Bacteriostasy) and the intensity of bacterial inactivation activity (IINAB/ Bactericidie) of alcoholic extracts of two accesses of hibiscus, known as: Palmares do Sul/RS and Porto Alegre/RS on standardized inoculum of Enterococcus faecalis (ATCC 19433), Escherichia coli (ATCC 11229), Salmonella Enteritidis (ATCC 11076) and Staphylococcus aureus (ATCC 25923). At the same time, the total content of polyphenols and anthocyanins in the calyxes and fruits with seeds hibiscus was determined. The antimicrobial activity of alcoholic extract of the calyxes in both accesses showed a significant positive difference when related to the alcoholic extract of the fruits with seeds. Salmonella enteritidis was the most sensitive bacteria to the alcoholic extract of calyxes of the hibiscus while the most resistant was Staphylococcus aureus, independent of the variable access, considering only the plant structure. In relation to the alcoholic extract of the fruits with seeds, Escherichia coli showed the highest sensitivity and Staphylococcus aureus the highest resistence. Total polyphenols and anthocyanins of alcoholic extract of calyxes‟s values showed a significant difference and they were superior to alcohol extract of fruits with seeds. Possibly there is a direct relationship between the concentration of anthocyanin and antibacterial activity in different structures of the hibiscus plant.

O interesse pela extração dos compostos bioativos, a partir de fontes naturais, para o emprego na produção de alimentos funcionais tem aumentado, devido, principalmente, à crescente demanda por parte dos consumidores, por produtos mais saudáveis e que possam trazer benefícios à saúde. Dentre as fontes naturais de compostos bioativos, destaca-se o hibisco (Hibiscus sabdariffa L.), que é rico em antocianinas, flavonoides, ácidos fenólicos, carotenoides, dentre outros. Entretanto, quando os compostos bioativos são separados de suas matrizes, estes se tornam altamente instáveis frente a diversos fatores ambientais, necessitando serem protegidos. O recobrimento por microencapsulamento é uma alternativa para retardar a degradação desses compostos. Este estudo teve como objetivo a extração, identificação, quantificação e microencapsulamento por atomização e liofilização dos compostos bioativos dos cálices do hibisco. Primeiramente, foi realizada a extração exaustiva total dos carotenoides e compostos fenólicos por meio de solventes orgânicos, para a identificação e quantificação desses compostos. Também foi estudada a extração de antocianinas e demais compostos fenólicos por meio de solvente aquoso acidificado (ácido cítrico 2 %, p/v). A partir do melhor tratamento de extração, o extrato obtido foi microencapsulado mediante atomização e liofilização, empregando goma arábica (GA), goma guar parcialmente hidrolisada (GGPH) e polidextrose (PD) como agentes encapsulantes, na concentração de 10%. Os carotenoides e compostos fenólicos foram identificados e quantificados por HPLC-DAD-MS/MS (cromatografia líquida de alta eficiência com detecção por arranjo de diodos e espectrometria de massa). Vinte e um carotenoides foram encontrados, dos quais, quinze foram identificados. O total de carotenoides nos cálices de hibisco foi de 641,38 ± 23,61 μg/100 g massa fresca, sendo a all-trans-luteína e o all-trans--caroteno os compostos majoritários, representando 49 e 23%, respectivamente. Para os compostos fenólicos, foram encontrados vinte compostos, dos quais, catorze foram identificados. As antocianinas foram os compostos majoritários nos cálices de hibisco, sendo que a delfinidina 3-sambubiosídeo e cianidina 3-sambubiosídeo representaram 41 e 13% do total de compostos fenólicos, respectivamente. Dentre os ácidos fenólicos, os componentes majoritários foram o ácido 3-cafeoilquínico e ácido 5-cafeoilquínico, representando 15 e 13% do total de compostos fenólicos, respectivamente. Para a extração aquosa acidificada, foi utilizado um planejamento experimental fatorial fracionado (24-1), com quatro fatores: concentração de enzima, temperatura, velocidade de agitação e tempo de extração. A partir da ANOVA, os efeitos principais e de interação foram avaliados, tendo como respostas Chroma, antocianinas monoméricas totais (TMA), capacidade redutora, ABTS e compostos fenólicos. A partir dos resultados, o melhor tratamento foi: 55 °C, 50 μL de enzima/1000 g extrato, 400 rpm e 4 horas de extração, obtendo-se nessa condição de extração 3,82 mg/g extrato em base seca para TMA e 17,59 mg/g de extrato em base seca para compostos fenólicos totais, que resultou em capacidade antioxidante de 7,72 μmol Eq. Trolox/g de extrato em base seca, avaliado por ABTS e de 3,96 mg GAE/g de de extrato em base seca, avaliado pela capacidade redutora. Este extrato foi empregado no estudo de encapsulamento, por atomização (140 ºC) e liofilização (-68 ºC por 24 horas), utilizando GA, GGPH e PD como encapsulantes. Observou-se que o melhor tratamento foi por liofilização empregando GA como encapsulante, resultando em 2,83 mg/g amostra em base seca para TMA, capacidade antioxidante de 2,98 mg GAE/g amostra em base seca e 5.67 μmol Eq. Trolox/g amostra em base seca, avaliados por capacidade redutora e ABTS, respectivamente. Entretanto, quando foram avaliadas as propriedades físicas e morfológicas dos pós, as amostras elaboradas por atomização e usando GA e GGPH apresentaram os melhores desempenhos, onde os valores de solubilidade, higroscopicidade e umidade foram de 95,8 e 95,2%, 31,3 e 28,9%, 1,9 e 2,4%, respectivamente. Para a temperatura de transição vítrea (Tg), os tratamentos que utilizaram GA e GGPH nos dois métodos de encapsulamento, tiveram os maiores valores de Tg, variando de 10,9 a 17,4 ºC. Já para os tratamentos que utilizaram a PD como material de parede, os valores foram de (0,7 °C), tanto na atomização como na liofilização. Na microscopia também foi observado um melhor desempenho nas micropartículas atomizadas usando GA e GGPH, as quais mostraram partículas mais esféricas e sem tendência de atração e aderência entre si. Em relação ao diâmetro médio de partícula (D[4, 3]), os tratamentos liofilizados tiveram partículas maiores que os atomizados, variando de 101,7 a 143,1 μm para os liofilizados, e de 5,4 a 7,3 μm para os atomizados. Quanto ao span, o qual avalia distribuição de tamanho de partícula, variou de 1,90 a 2,00 para as amostras atomizadas e de 3,06 a 3,19 para as amostras liofilizadas, indicando que houve uma boa uniformidade na distribuição de tamanho de partícula. Conclui-se que o hibisco é uma matriz com ampla composição de compostos bioativos e tem potencial para aplicação em alimentos.
The interest in the extraction of bioactive compounds from natural sources, for use in the production of functional foods has increased, mainly due to the growing demand by consumers for healthier products and can bring health benefits. Among the natural sources of bioactive compounds, stands out the hibiscus (Hibiscus sabdariffa L.), which is rich in anthocyanins, flavonoids, phenolic acids, carotenoids, among others. However, when the bioactive compounds are separated from their matrix, they become highly unstable against various environmental factors and need to be protected. The coating by microencapsulation is an alternative to slow the degradation of these compounds. This study aimed at the extraction, identification, quantification and microencapsulation by spray drying and freeze drying of bioactive compounds of hibiscus calyces. Firstly, a thorough exhaustive extraction of carotenoids and phenolic compounds by organic solvents was performed for identification and quantification of these compounds. The extraction of anthocyanins was also studied along with other phenolic compounds by an aqueous solvent acidified (2% citric acid, w/v). From the best treatment for extraction, the extract obtained was microencapsulated by spray drying and freeze drying using Arabic gum (GA), partially hydrolyzed guar gum (PHGG) and polydextrose (PD) as encapsulating agents in a concentration of 10%. Carotenoids and phenolic compounds were identified and quantified by HPLC-DAD-MS/MS (high-performance liquid chromatography with diode array detection and mass spectrometry). Twenty-one carotenoids were found, of which fifteen were identified. The total carotenoids in hibiscus calyces was 641.38 ± 23.61 mg/100 g fresh weight, with the all-trans-lutein and all-trans-β-carotene the major compounds, representing 49 and 23%, respectively. Regarding the phenolic compounds it was found twenty of those, of which fourteen have been identified. Anthocyanins were the main components in the hibiscus calyces, and delphinidin and cyanidin 3-sambubioside 3-sambubioside represented 41 and 13% of total phenolic compounds, respectively. Among the phenolic acids, the major components were the 3-caffeoylquinic acid and 5-caffeoylquinic acid, representing 15 and 13% of total phenolic compounds, respectively. For acidified aqueous extraction, we used a fractional factorial design (24-1) with four factors: enzyme concentration, temperature, stirring speed and extraction time. From the ANOVA, the main and interaction effects were assessed as answers: Chroma, total anthocyanins monomeric (TMA), reducing capacity, ABTS and phenolic compounds. From the results, the best treatment was with 55 °C, 50 μL of enzyme/1000 g extract, 400 rpm and 4 hours of extraction, it was obtained in this extraction condition 3.82 mg/g extract on a dry basis for TMA and 17.59 mg/g extract on a dry basis for phenolic compounds, which resulted in antioxidant capacity of 7.72 μmol Eq. Trolox/g extract on a dry basis, evaluated by ABTS and 3.96 mg GAE/g extract on a dry basis, assessed by reducing capacity. This extract was used for the encapsulation study, by spray drying (140 °C) and freeze drying (-68 ° C for 24 hours) using GA, PHGG, and PD as encapsulants. It was observed that the best treatment is by freeze drying using GA as encapsulant, resulting in 2.83 mg/g sample on dry basis for TMA, antioxidant capacity of 2.98 mg GAE/g sample on dry basis and 5.67 μmol Eq. Trolox/g sample on dry basis, evaluated by reducing capacity and ABTS, respectively. However, when we evaluated the physical and morphological properties of powders, samples prepared by spray drying and using GA and PHGG showed the best performance, and the values for solubility, hygroscopicity and moisture were 95.8 and 95.2%, 31.3 and 28.9%, 1.9 and 2.4%, respectively. For the glass transition temperature (Tg), treatments with GA and PHGG on both encapsulation methods had high Tg values ranging from 10.9 to 17.4 °C. As for treatments of PD as wall material, the values were (0.7 °C), both the spray drying as in freeze drying. In microscopy was also observed improved performance in spray-dried microparticles using GA and PHGG, which showed more spherical particles and with no tendency to attract and adhere to each other. Regarding the average particle diameter (D [4, 3]), the freeze-dried treatments had higher spray-dried particles ranging from 101.7 to 143.1 μm for freeze-dried, and 5.4 to 7.3 μm for spray-dried. As the span, which assesses particle size distribution ranged from 1.90 to 2.00 for spray-dried samples and 3.06 to 3.19 for the freeze-dried samples, indicating that there was a good uniformity in the size in the distribution of the size of the particle. It follows that hibiscus is a matrix with broad composition and bioactive compounds have potential for application in foods.

Os cálices do hibisco possuem uma grande quantidade de compostos bioativos responsáveis pela sua atividade antioxidante. O presente trabalho teve como objetivos a obtenção de extratos contendo esses compostos bioativos em solução aquosa com 2 % de ácido cítrico por extração assistida por micro-ondas (EAM) e o encapsulamento desses extratos por atomização e liofilização utilizando polidextrose (PD), proteína isolada do soro de leite (WPI) e a mistura destes na concentração de 10 %. Previamente foram estudados três métodos de extração, o primeiro usando somente EAM a 200, 300 e 700 W de potência, e tempos de 2, 5 e 8 minutos; o segundo consistiu de dois períodos, a extração aquosa ácida com tempos de 1, 2, 4, 6, 18 e 24 horas seguida de EAM nas potências de 200, 300 e 700 W; o terceiro consistiu de EAM seguida de extração aquosa ácida, nos mesmos tempos e potências citados para o segundo método. Os resultados indicaram que a melhor condição do primeiro método foi a 700 W e 8 min, do segundo método realizada a 6 horas de extração aquosa ácida, seguida de EAM a 700 W por 8 min e do terceiro método, EAM a 700 W por 8 min seguida de 6 horas de extração aquosa ácida. Quando os três métodos foram comparados, a melhor condição de extração foi aquela obtida no terceiro método: 1,63 mg delfinidina-3-sambubiosídeo · g-1; 29,62 mg EAG · g-1; 133,25 μmol ET · g-1 para antocianinas, fenólicos totais e atividade antioxidante por ABTS, respectivamente. Para avaliar o efeito da extração obtida somente por EAM, os extratos obtidos por extração exaustiva com metanol por 25 min e EAM a 700 W e 8 min foram quantificados por HPLC-DAD-ESI-MS/MS, sendo encontrados 13 compostos (6 ácidos fenólicos, 2 antocianinas e 5 flavonóides derivados da quercetina, kaempferol e miricetina). Os compostos fenólicos majoritários foram o ácido 3-cafeoilquínico (2,58 e 1,32 mg · g-1) e ácido 5-cafeoilquínico (1,71 e 0,90 mg · g-1) para extração exaustiva e EAM, respectivamente. Esse mesmo extrato (700 W e 8 min) foi encapsulado por atomização (160 °C) e liofilização (- 68 °C por 54 horas). Os pós obtidos foram avaliados quanto aos teores de compostos fenólicos totais, antocianinas monoméricas totais, atividade antioxidante (ABTS, DPPH e HRSA), medidas por análises espectrofotométricas, atividade de água, umidade, higroscopicidade, solubilidade, eficiência de encapsulação, cor, análise termogravimétrica, temperatura de transição vítrea, espectroscopia de infravermelho com transformada de Fourier (FTIR) e microestrutura (MEV). Os pós atomizados tiveram menor atividade de água (0,14 a 0,17), umidade (3,4 a 4,5 %), higroscopicidade (23,9 a 34,1 %), solubilidade (86 a 98,2 %) e eficiência de encapsulação (51,62 a 84,52 %) do que os pós liofilizados. Os resultados do FTIR mostraram que os encapsulantes não interagiram quimicamente, visto que não foram observados mudanças na frequência dos picos; as provas termogravimétricas indicaram que os pós apresentaram a mesma tendência nas perdas de massa. Na análise de microestrutura foi observado um melhor desempenho nas micropartículas atomizadas com PD, as quais mostraram partículas mais esféricas e sem tendência de atração e aderência entre si. Foram obtidas retenções de 38 a 77 % para antocianinas monoméricas totais, 42 a 89 % para compostos fenólicos totais, e entre 33 e 90 % para atividade antioxidante nos pós obtidos. O pó encapsulado liofilizado com 10 % de polidextrose mostrou uma maior retenção de antocianinas (77 %), atividade antioxidante por DDPH (90 %) e HRSA (74 %), entretanto com maior higroscopicidade (39,4 %). As provas aceleradas de estocagem (umidades relativas de 75 e 90 % em temperaturas de 40 e 60 °C) realizadas em todos os pós encapsulados, após 30 dias, indicaram que o tratamento liofilizado com 10 % de PD foi o que apresentou melhores resultados a essas condições, retendo 75 % dos compostos fenólicos, com atividades antioxidantes medidas por ABTS, DPPH e HRSA de 75, 90 e 74 %, respectivamente, existentes no extrato original. O pó obtido tem potencial para aplicação em alimentos, portanto, devido ao hibisco ser uma matriz com ampla composição de compostos bioativos.
The hibiscus calyces contend a high quantity of bioactive compounds responsible for their antioxidant activity. The present paper was aimed the production of extracts containing those bioactive compounds in acidified aqueous solution 2 % of citric acid by microwave assisted extraction (MAE) and the encapsulation of those extracts by spray drying and freeze-drying using polydextrose (PD), whey protein isolate (WPI) and their mixture in the concentration of 10 %. Previously three methods of extraction were studied, the first using only MAE at 200, 300 and 700 W of power, and times of 2, 5 and 8 minutes; the second consisted of two steps, the acid aqueous extraction with times of 1, 2, 4, 6, 18 and 24 hours followed by MAE at the powers of 200, 300 and 700 Watts; the third consisted of MAE followed by acid aqueous extraction, in the same times and powers mentioned for the second method. The results indicated that the best condition of the first method was 700 W and 8 minutes, the second method performed at 6 hours of acid aqueous extraction, followed by MAE at 700 W for 8 minutes and the third method, MAE at 700 W for 8 minutes followed by 6 hours of acid aqueous extraction. When the three methods are compared, the best condition of extraction was obtained in the third method: 1.63 mg delphinidin-3-sambubioside · g-1; 29.62 mg GAE · g-1; 133.25 μmol TE · g-1 for total monomeric anthocyanins, total phenolic compounds and antioxidant activity by ABTS, respectively. To evaluate the effect of the extraction obtained only by MAE, the extracts obtained by exhaustive extraction with methanol for 25 minutes and MAE at 700 W and 8 minutes were quantified by HPLC-DAD-ESI-MS/MS, was found 13 compounds (6 phenolic acids, 2 anthocyanins and 5 flavonoids derived from quercetin, kaempferol and myricetin). The phenolic compounds majorities were acid 3-caffeoylquinic (2.58 e 1.32 mg · g-1) and acid 5-caffeoylquinic (1.71 e 0.90 mg · g-1) for exhaustive extraction and MAE, respectively. That same extract (700 W and 8 minutes) was encapsulated by spray drying (160 ºC) and freeze-drying (- 68 °C for 54 hours). The obtained powders were evaluated about the levels of total phenolic compounds, total monomeric anthocyanins, antioxidant activity (ABTS, DPPH e HRSA), measured by spectrophotometric analysis, water activity, moisture, hygroscopicity, solubility, encapsulation efficiency, color, thermogravimetric analysis, glass transition temperature, Fourier transform infrared spectroscopy (FTIR) and microstructure (MEV). The spray dried powders had lower water activity (0.14 to 0.17), moisture (3.4 to 4.5 %), hygroscopicity (23.9 to 34.1 %), solubility (86 to 98.2 %) and encapsulation efficiency (51.62 to 84.52 %) than the freeze-dried powders. The results of FTIR showed that the encapsulants did not interact chemically, since changes were not observed on the frequency of the peaks; the thermogravimetric tests indicated that the powders presented the same tendency on the mass loss. On the microstructure analysis a better perform was observed on the spray dried microparticles with PD, which showed more spherical particles and with no tendency of attraction and adherence between them. Were obtained retentions of 38 to 77 % for total monomeric anthocyanins, 42 to 89 % for total phenolic compounds and between 33 and 90 % for antioxidant activity in the obtained powders. The encapsulated power by freeze-drying, with 10 % of polydextrose, was showed higher retention of anthocyanins (77 %), antioxidant activity by DDPH (90 %) and HRSA (74 %), however with higher hygroscopicity (39.4 %). The accelerated tests of storage (relative humidity of 75 and 90 % in temperatures of 40 and 60 ºC) performed in all the encapsulated powders, after 30 days, indicated that the freeze-drying treatment with 10 % of PD has the best behavior in those conditions, retaining 75 % of the phenolic compounds, with antioxidant activities measured by ABTS, DPPH and HRSA of 75, 90 and 74 %, respectively, present in the original extract. The obtained powder has potential for application in foods, therefore, due to the hibiscus being a matrix with ample composition of bioactive compounds.

Hibiscus sabdariffa L. est une plante vivace herbacée localisée dans les zones tropicales et subtropicales des deux hémisphères. Au Sénégal, où elle a été introduite au 19ème siècle, H. sabdariffa, encore appelé bissap ou roselle, est cultivé sur l'ensemble du territoire et exploité principalement pour ses calices. Une boisson désaltérante est traditionnellement élaborée à partir des calices séchés par extraction à l'eau. La caractérisation physico-chimique et biochimique des calices nous a tout d'abord permis de mieux évaluer le potentiel de ce produit. Ainsi, les calices contiennent entre 767 et 1480 mg.100g -1 d'anthocyanes et entre 1347 et 3736 mg.100g-1 de polyphénols totaux. Ils présentent un pouvoir antioxydant voisin de 260 µmolTrolox.g-1. L'acide malique y est l'acide organique majoritaire avec une teneur pouvant atteindre 2,3 g.100g-1. De part ces caractéristiques, l'extrait aqueux de calices d'H. sabdariffa constitue une excellente source d'anthocyanes. Le travail s'est ensuite focalisé sur le procédé d'obtention de cet extrait. L'étude de l'extraction solide-liquide a été réalisée à partir de calices séchés et d'eau à des températures comprises entre 25 et 90 °C. Un rendement d'extraction en anthocyanes maximum de 88 % a été obtenu à 25 °C avec un ratio calice/eau de 1/25. La réduction de la taille des particules et l'augmentation de la température diminuent le temps d'extraction mais n'affectent pas le rendement d'extraction. Un modèle d'extraction solide-liquide incluant les cinétiques de dégradation thermique des anthocyanes a été proposé puis validé. La microfiltration tangentielle sur membrane céramique de diamètre moyen de pores 0,2 µm a permis de produire un extrait stérilisé à froid, stable pendant 90 j à 20 °C. Nous avons montré à ce niveau que la pression transmembranaire optimisée à un facteur de réduct ion volumique (FRV) égal à 1 n'est plus optimale lorsque le FRV augmente. Pour concentrer les anthocyanes présents dans l'extrait, l'ultrafiltration sur une membrane de seuil de coupure de 5 kDa peut être utilisée malgré une rétention incomplète (95-97 %). Par contre, la nanofiltration offre une rétention totale des anthocyanes. Le procédé, mis en uvre dans des conditions semi-industrielles, a permis d'augmenter la teneur en extrait sec soluble (ESS) de 40 à 250 g.kg-1 à 35 °C, à une pression transmembranaire comprise entre 20 et 30 bar avec un flux moyen de 15 kg.h-1.m-2. L'évaporation osmotique, testée sur un pilote équipé d'une membrane en polypropylène de 10 m2, a permis de concentrer l'extrait anthocyanique jusqu'à 620 g d'ESS.kg-1. Cet extrait concentré conserve toutes les qualités physico-chimiques, biochimiques, nutritionnelles et aromatiques de l'extrait initial. Le flux évaporatoire est directement corrélé à la teneur en extrait sec solub le du produit à concentrer. Enfin, divers couplages d'opérations unitaires ont été comparés pour obtenir un extrait concentré à partir de calices d'H. sabdariffa. Le procédé qui associe une extraction à froid, une pré-concentration par nanofiltration et une concentration par évaporation osmotique est la meilleure voie tant en terme de coûts d'investissement et de fonctionnement que de qualité de l'extrait
Hibiscus sabdariffa L. is a herbaceous plant, cultivated largely in tropical and subtropical areas of both hemispheres. In Senegal, where it was introduced in the 19th century, H. sabdariffa L. also called bissap or roselle is grown throughout the territory and operated primarily for its calyx. A refreshing drink is traditionally made from dried calyx by water extraction. The physical-chemical and biochemical characterization of the calyx allowed us to assess better the potential of this product. Thus, the calyx contained between 767 and 1480 mg.100g-1 of anthocyanins and between 1347 and 3736 mg.100g-1 of total polyphenols. They have an antioxidant capacity closed to 260 μmolTrolox.g-1. Malic acid is the major organic acid with content up to 2.3 g.100g-1. Because of these characteristics, the aqueous extract of H. sabdariffa calyx is an excellent source of anthocyanins. The work was then focused on the process for obtaining this extract. The study of solid-liquid extraction was performed from dried calyx with water at temperature range from 25 and 90 ° C. A maximum extraction yield of anthocyanin of 88 % was obtained at 25 ° C with a solid-to-solvent ratio of 1/25. Reducing the particle size and increasing the temperature decreased the extraction time but do not affect the extraction efficiency. A model of solid-liquid extraction which included the thermal anthocyanins degradation kinetics has been proposed and validated. Crossflow microfiltration with tubular ceramic 0.2 pore size membrane has produced a cold sterilized extract, stable during 90 days at 20 °C. We have shown here that the transmembrane pressure optimized at a volume reduction ratio (VRR) equal to 1 is no longer optimum when the VRR is increasing. To concentrate the anthocyanins present in the extract, the ultrafiltration membrane with a cutoff of 5 kDa can be used despite incomplete retention (95-97%). By cons, nanofiltration offers a tot al retention of anthocyanins. The method, implemented in semi-industrial conditions, has increased the content of total soluble solids (TSS) from 40 to 250 g.kg-1 at 35 °C at a transmembrane pressure between 20 and 30 bar with an average flow of 15 kg.h-1.m-2. The osmotic evaporation, tested on a pilot equipped with a polypropylene membrane of 10 m2, has concentrated anthocyanin extract up to 620 g TSS. kg-1. This concentrated extract retains all the physical-chemical, biochemical, nutritional and aromatic quality of the original extract. The evaporation flux is directly correlated with the total soluble solids content of the product to concentrate. Finally, various couplings of unit operations were compared to obtain a concentrated extract from H. sabdariffa calyx. The process which combines a cold extraction, a pre-concentration by nanofiltration and concentration by osmotic evaporation is the best way both in terms of investment costs and operating and quality of the ext ract

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Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES
Vegetable of African origin, the hibiscus (Hibiscus sabdariffa. L) is fully adapted to the climatic conditions of the state of Maranh?o its leaves are the basis for many dishes Maranh?o. The vinegar has high nutritional value, low cost, easy to grow especially for lowincome families who produce and consume a product containing functional nutrients such as minerals and vitamins, like iron and vitamin C and thus acquiring great socioeconomic importance, however in its marketing there is a significant loss, which is linked primarily to biological factors that indicate numbers to producers marketing a fresh hibiscus anthocyanins, total phenolics, sugars, non-reducing sugars, fiber, minerals and calories , microbiological analyzes evaluated the products and they were hygienic and sanitary conditions. Sensory analysis showed an average of more than seven in a nine-point hedonic scale, which represents the impression "liked moderately", indicating that the flavor of the vegetable vinegar was well received by consumers. The percentage of panelists who demonstrated purchase intent for jelly and sweet vinegar cut was fairly representative, showing the ability to market products based on vinegar, increasing the shelf life and value of vegetables
Hortali?a de origem africana, a vinagreira (Hibiscus sabdariffa. L) est? plenamente adaptada as condi??es edafoclim?ticas do estado do Maranh?o suas folhas servem de base para v?rios pratos da culin?ria maranhense. A vinagreira possui alto valor nutritivo, baixo custo, f?cil cultivo principalmente para as fam?lias de baixa renda que produzem e consomem um produto que contem nutrientes funcionais como minerais e vitaminas, a exemplo do ferro e vitamina C e assim, adquirindo grande import?ncia socioecon?mica, entretanto, na sua comercializa??o existe uma perda significativa, a qual esta ligada principalmente a fator biol?gico que indicam n?meros preocupantes para os produtores que comercializam a vinagreira in natura antocianinas, compostos fen?licos totais,a??cares redutores,a??cares n?o redutores,fibras,minerais e valor cal?rico,as an?lises microbiol?gicas avaliaram os produtos e estes apresentaram condi??es higi?nicos sanit?rias satisfat?rias.A analise sensorial apresentou m?dias superiores a sete em uma escala hed?nica de nove pontos,que representa a impress?o ?gostei moderadamente? indicando que o sabor caracter?stico da hortali?a vinagreira foi bem recebido pelos consumidores. O percentual de provadores que demonstrou inten??o de compra para geleia e doce de corte de vinagreira foi bastante representativo, evidenciando a possibilidade de introduzir no mercado produtos a base de vinagreira, aumentando a vida de prateleira e o valor agregado da hortali?a

Les composés phénoliques sont des métabolites secondaires chez les végétaux, qui possèdent un large éventail de propriétés biologiques et fonctionnelles, par exemple antioxydantes, antimicrobiennes. Comme ils sont très répandus dans le règne végétal, les sous-produits issus de la transformation végétale représentent une source intéressante et peu coûteuse des polyphénols qui pourraient être utilisés comme additifs naturels, inoffensifs et multifonctionnels, avec une valeur de marché ajoutée. Toutefois, leur polarité limite leur utilisation dans des systèmes émulsionnés complexes, tels que ceux rencontrés en formulations cosmétiques, alimentaires, ou pharmaceutiques. Sur cette base, la lipophilisation s’est avérée être une stratégie efficace pour améliorer le caractère hydrophobe de ces composés bioactifs et, par conséquent, leurs propriétés fonctionnelles.Le principal objectif de ce travail a été d'ajouter de la valeur à des sources végétales riches en polyphénols par l'extraction, caractérisation et enfin, par lipophilisation des principaux polyphénols issus de ces sources. Dans ce contexte, Hibiscus sabdariffa L. ou roselle, cultivée dans le monde entier et considérée comme une importante source d’anthocyanes, avec des propriétés colorantes uniques, ainsi que des propriétés antioxydantes et antimicrobiennes, a été utilisée pour l'extraction assistée par ultrasons, suivi d’une seule étape de fractionnement par chromatographie en phase liquide à basse pression, pour isoler ces principaux anthocyanes. Les composés ainsi obtenus ont été utilisés pour évaluer leur possibilité à être lipophilisés en utilisant différentes longueurs de chaînes alkyles. Egalement, les propriétés colorantes des anthocyanes issues d’hibiscus ont été calculées par la méthode de pH jumps. Ainsi, les constantes de pseudo équilibre (pK'a 2,9), d’acidité (pKa 3,7) et d’hydratation (pK'h 3,0) de la delphinidine-3-O-sambubioside, ont été estimées.La delphinidine-3-O-sambubioside et la cyanidine-3-O-sambubioside ont été les principaux anthocyanes obtenus à partir de l’extrait d'hibiscus. Ces composés ont été lipophilisés avec succès pour la première fois, par voie chimique. Les produits ont été caractérisés par des techniques de chromatographie et spectrométrie. L'acide protocatéchique (PA), qui peut être extrait de la fleur d'hibiscus, a été utilisé comme molécule modèle, et il a été lipophilisé par voie chimique avec une série homologue d'alcools. Afin d'évaluer l'impact de la lipophilisation sur leurs propriétés fonctionnelles, leur activités antioxydantes (DPPH, ORAC, CAT), et antifongique (méthode de microdilution) ont été évaluées avant et après lipophilisation. De manière générale, les molécules estérifiées avec des alcools de chaîne courte et moyenne (de C1 à C6) ont montré des activités améliorées par rapport au composé non estérifié.En conclusion, cette étude a montré que la lipophilisation, en utilisant une longueur de chaîne optimale, pourrait avoir un impact positif sur les propriétés antioxydantes et antifongiques des composés phénoliques. Ces travaux de thèse ont également démontré que cette stratégie peut être appliquée à différentes familles de composés phénoliques, tels que les anthocyanes et les acides phénoliques, pour l’obtention des molécules pouvant être potentiellement utilisées comme additifs dans des produits de formulation
Phenolic compounds are a group of secondary plant metabolites with a broad range of inherent biological and functional properties e.g. antioxidants and antimicrobial agents, all bundled within the phenol functional group. Since they are one of the most widely distributed molecules in the vegetal kingdom, by-products issued from the vegetal processing represent an interesting and inexpensive source of polyphenols that could be used as natural, innocuous and multifunctional ingredients, with an added market value. However, the polarity of these phenolic compounds limits their use in complex emulsified systems, such as food, pharmaceutical or cosmetic formulations. Based on this, lipophilization has showed to be an efficient strategy to enhance the hydrophobicity of these bioactive compounds and, consequently, their functional properties. The aim of this work was to add value to polyphenol-rich plant sources by the extraction, characterization and finally, lipophilization of their predominant polyphenols. In this context, Hibiscus sabdariffa L. or roselle flower, an inexpensive, worldwide produced source of anthocyanins, with unique color, and, antioxidant and antimicrobial properties, was used for ultrasound-assisted extraction and single-step fractionation by low-pressure liquid chromatography, to isolate its main anthocyanins. The obtained compounds were used to evaluate their suitability for lipophilization with different chain length alkyl donors. Moreover, the color properties of hibiscus anthocyanins were studied by the pH jumps method, which allowed the calculation of the pseudo equilibrium (pK'a 2.9), acidity (pKa 3.7) and hydration constants (pK'h 3.0).The obtained hibiscus anthocyanins (delphinidin-sambubioside and cyanidin-sambubioside) were successfully chemically lipophilized. The products were characterized by means of LC/ESI-MS. Moreover, protocatechuic acid (PA), which can be extracted from hibiscus flowers, was used as model molecule, and it was chemically lipophilized with a homologous series of alcohols. In order to evaluate the impact of the lipophilization on their functional properties, protocatechuates were assessed for their antioxidant (DPPH, ORAC, CAT), and antifungal (microdilution method) activity, before and after chemical lipophilization. In general, protocatechuates with short to medium alkyl chain lengths (from C1 to C6) showed improved activities compared to the parent compound. In conclusion, this study showed that, lipophilization using the optimal chain length could have a positive impact in the functional properties, e.g. antioxidant and antifungal activities, of phenolic compounds. Here was also demonstrated that this strategy can be applied to different families of phenolic compounds, such as anthocyanins and phenolic acids, representing an alternative to synthesize molecules that might have potential use as additives in formulated products

Thesis (MTech (Biomedical Technology))--Cape Peninsula University of Technology, 2015.
Cancer is the leading cause of death in both developed and developing countries. In particular, breast cancer is regarded as the most common neoplastic disease in females and accounts for the high mortality rates in women. Increased mortality rates could be attributed to ineffective current cancer treatment modalities that have been implicated to cause multidrug resistance, high toxicity and induction of several side effects. In addition, oxidative stress appears to play a role in the development of breast cancer. Therefore, current cancer research aims to search for plant based anticancer compounds with less side effects and toxicity towards the human body. An example of such a plant is Hibiscus sabdariffa also known as roselle and is reported to have bioactive compounds that exhibit anticancer and antioxidant effects. However, the effects of Hibiscus sabdariffa on breast cancer in relation to oxidative stress and apoptosis have not been investigated. In this research study, the aim was to evaluate the cytotoxic and antioxidant effects of water and methanolic extracts of Hibiscus sabdariffa (HS) on cancerous MCF7 and non-cancerous MCF12A breast cell lines with special reference to oxidative stress and apoptosis. This was done based on the fact that HS has been documented for its traditional use against cancer and other ailments.

Dissertação (Mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Agrárias.
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Este trabalho foi realizado com o objetivo de pesquisar a estabilidade das antocianinas de Hibiscus sabdariffa L. frente a determinadas condições e avaliação de sua composição físico-química para um possível aproveitamento desta malvácea como fonte de nutrientes e corante para a indústria de alimentos. Os resultados da composição físico-química demonstraram que vinagreira (Hibiscus sabdariffa L.), pode ser utilizada como fonte de alimento, constituindo-se numa boa fonte de nutrientes, destacando-se a quantidade de cálcio determinada (659 mg/100g), situando-se acima dos valores encontrados para hortaliças em geral. A avaliação aminoacídica demonstrou que o cálice desta malvácea possui a maioria dos aminoácidos essenciais, com exceção de cistina e tirosina. A estabilidade de antocianinas foi estudada através do efeito da luz, oxigênio, pH, adição de ácido tânico e solução de b-ciclodextrina. A solução extratora mais eficiente foi metanol - HCl 0,01%. Os fatores que mais contribuíram para a degradação da cor das antocianinas de Hibiscus sabdariffa L. foram a incidência da luz e variações de pH. A adição de b-ciclodextrina não resultou num efeito protetor, enquanto que o ácido tânico resultou numa considerável estabilidade.

Magister Scientiae Dentium - MSc(Dent)
The antimicrobial and antifungal efficacy of indigenous plant extracts against Streptococcus mutans , Escherichia coli and Candida albicans .Aim: To determine the antimicrobial and antifungal efficacy of indigenous plant extracts, Tamarindus Indica (T. ndica), Hibiscus sabdariffa (H. sabdaraffi), Adansonia digitata (A.digitata) and Moringa oleifera (M. oleifera) against Streptococcus mutans ( S. mutans ), Escherichia coli ( E. coli ) and Candida albicans (C. albicans) objectives:The objectives of this study were to: 1.Measure the zones of growth inhibition by T. indicia , A. digitata , M.oleifera and H. sabdariffi extracts against, S.mutans , E.coli and C.albicans . 2.Compare the size of inhibition zones of different bacteria or fungus, S. mutans ,E.coli and C. albicans, around the same plant extract.3.Compare the size of inhibition zones for the same bacteria in different plant extracts T.indicia ,A.digitata H.sabdariffi and M.oleifera Methodology .The antimicrobial and antifungal effect of the ethanolic extracts of T. indica , H.sabdariffa, A.digitata and M.oleifera was performed using the disc diffusion method against S. mutans ,E.coli and C.albicans. The antibacterial and antifungal activity of the plants was determined by measuring the diameter of the inhibition zones. esults and conclusion: The results showed that.T.indica and H.sabdariffa ethanolic extracts have an antibacterial effect against S.mutans and E.coli.However, H.sabdariffi showed a significantly higher antibacterial effect against E.coli and S.mutans, with a range of 14.50mm to 12.01mm and 16.41 mm to 14.39 mm compared to T.indica ,with a range of 11.41 mm to 7.04mm and 6.88mm to 10.40mm, respectively.Furthermore, the statistical multiple pairwise test (Conover Iman procedure/Two -tailed test) omputed that the effect of H.sabdariffi is significantly (critical value >7.229) greater for the G ram positive S.mutans than the G ram negative E.coli.On the other hand,T.indica showed a similar antibacterial effect against S.mutans and E.coli,respectively.In contrast, M. oleifera and A.digitata ethanolic plant extracts showed no antibacterial effect against E.coli and S.mutans. All the indigenous plants tested, T.indica ,H.sabdariffa, M. oleifera and A.digitata had no antifungal activity on C.albicans

Hibiscus sabdariffa (H. sabdariffa) is a rich source of bioactive phytochemicals with potential health benefits. Based on its reported use in folk medicine for the treatment of hypertension and recently type 2 diabetes, the plant has gained considerable commercial importance as a functional food. Three varieties of H. sabdariffa were analyzed for their phytochemical content and inhibitory potential on carbohydrate-digesting enzymes as a basis for selecting a variety for wine production. The dark red variety was chosen as it was highest in phenolic content and partially inhibited α–glucosidase (maltase), with delphinidin 3-O-sambubioside, cyanidin 3-O-sambubioside and 3-O-caffeoylquinic acid accounting for 65 % of this activity. None of the varieties significantly inhibited α–amylase. Regarding H. sabdarifa wine, the effect of fermentation (20 and 30 ˚C) and ageing (6, 15 and 30 °C) temperatures on the phytochemical composition and bioactivity of Roselle wine was investigated for a period of 40 days and 12 months respectively. The main change in phytochemical composition observed during fermentation was the hydrolysis of 3-O-caffeolquinic acid and the concomitant increase of caffeic acid irrespective of fermentation temperature. Wine fermented at 20 ˚C was slightly more active for α–glucosidase inhibition with more fruity aromas, but there were more flowery notes at 30 ˚C. Although ageing temperature did not produce an obvious trend in concentrations of most of the volatiles analysed, its impact was clearly demonstrated in the final concentrations of diethyl succinate which increased with higher ageing temperature. The data on the overall quality rating of roselle wine by a sensory panel showed that neither fermentation nor ageing temperatures produced any significant difference (p ≥ 0.05) in the wine samples. This study demonstrates that processing of H. sabdariffa into wine represents a promising alternative to expanding the functional properties of this crop.

Con el fin de generar una nueva fibra dietética antioxidante y prebiótica, el ácido gálico (AG) se injertó en inulina nativa por el método de radicales libres generados mediante el par Redox peróxido de hidrógeno/ácido ascórbico (H2O2/AA). A través de Resonancia Paramagnética Electrónica (EPR), se evaluaron diversas relaciones molares H2O2/AA para determinar el efecto de la oxidación de la inulina y medir la eficiencia en el injerto Inulina- ácido gálico (IGA). El injerto se confirmó mediante UV, espectroscopia de infrarrojo (FT-IR) y difracción de rayos X en polvos. Además, la actividad antioxidante fue evaluada por métodos espectroscópicos y la actividad prebiótica del IGA se determinó por el crecimiento In vitro de Lactobacillus acidophilus.
Con el fin de generar una nueva fibra dietética antioxidante y prebiótica, el ácido gálico (AG) se injertó en inulina nativa por el método de radicales libres generados mediante el par Redox peróxido de hidrógeno/ácido ascórbico (H2O2/AA). A través de Resonancia Paramagnética Electrónica (EPR), se evaluaron relaciones molares (RM) a 9, 20, 39 y 49 moles de H2O2/AA para determinar el efecto de la oxidación de la inulina y medir la eficiencia en el injerto Inulina- ácido gálico (IGA). El injerto se confirmó mediante UV, espectroscopia de infrarrojo (FT-IR) y difracción de rayos X en polvos. Además, la actividad antioxidante fue evaluada por métodos espectroscópicos y la actividad prebiótica del IGA se determinó por el crecimiento In vitro de Lactobacillus acidophilus. La concentración más alta de macro-radical en la inulina se obtuvo con relaciones molares de H2O2/AA de 20 y 49. El macro-radical de la inulina se forma por la eliminación de un átomo de hidrógeno de los grupos metilo en los monómeros de fructosa. Los espectros UV muestran picos de absorbancia a 214 y 266-268 nm que evidencian la presencia de anillos aromáticos en el injerto IGA y los espectros FT-IR muestran una banda a 1743 cm-1, que confirma el enlace covalente entre la inulina y el AG. Las concentraciones equivalentes de AG en IGA fueron; 30.4 y 16.3 mg AG/ g IGA, con RM = 9 M de H2O2/AA. Los nuevos polímeros exhiben capacidad de inhibir radicales libres tales como DPPH, 1O2, y para reducir la peroxidación de lípidos. La inulina muestra una significativa capacidad de estimular el crecimiento de L. acidophilus que no se ve afectada por la presencia de moléculas de AG injertado. Este trabajo demuestra que es posible proporcionar capacidad de inhibición de radicales libres a los fructo-oligosacáridos, evitando que sus propiedades prebióticas disminuyan, lo que podría extender su potencial como alimentos funcionales.
Consejo Nacional de Ciencia y Tecnología México

L'objectif de l'étude était d'évaluer les potentialités aromatiques et médicinales de deux plantes (Hibiscus sabdariffa et Artemisia annua). Les performances et rendement de plusieurs méthodes d'extraction et de solvants sur des calices et des graines d'hibiscus, issus de trois pays (Vietnam, Sénégal et Mexique) ont été comparés. Les profils en composés volatifs diffèrent selon les extraits. La teneur en acide gras, stérols et tocophérols des huiles des graines d'hibiscus est comparable à celle de l'huile de coton et de soja. Une seconde partie du projet portant sur l'Artemisia du Cambodge et du Sénégal, consiste à établir les conditions de production et la teneur en artémisinine, en terme de stade de développement, date de récolte, type de site. Le bourgeon accumule le maximum d'artémisinine. La teneur en principe actif chute rapidement avec le développement des fleurs. L'Artémisia cultivée au Sénégal présente des taux d'artémisinine moins élevés que l'Artemisia du Cambodge.

Present days lifestyles increase the number of free radicals in our body and predispose people to oxidative stress, responsible for the development of several diseases as well as aging. The need for inhibitors of oxidative processes has become essential. Plant-derived antioxidant compounds are preferred. Hibiscus sabdariffa L. belongs to the Malvaceae family. Its calyces contain anthocyanins, responsible for the antioxidant activity reported for the extracts. Moringa oleifera Lam. belongs to the Moringaceae family. Its antioxidant properties are mainly attributed to its phenolic compounds and, recently, to its isothiocyanates, deriving from the enzymatic hydrolysis of glucosinolates. Both the plants are source of oils with an interesting composition in antioxidant compounds. This research work was divided into three lines: The first line focused on H. sabdariffa calyces. It was performed the selection of a variety of calyces rich in anthocyanins. It was optimized an extraction method suitable for the recovery of anthocyanins in high yield but transferable on large scale, too. It were tried different procedures to obtain an extract enriched in anthocyanins using polyvinylpolypyrrolidone. The second line was devoted to M. oleifera tissues. This comparative study concerns plant material grown in Paraguay and therefore extends the information available on the chemical composition of M. oleifera plants cultivated in different areas of the world. As far as we know, this is the first study about the characterization of both direct and indirect antioxidant compounds in different tissues of M. oleifera. Branches and the patented matrix “EDYNEA” are investigated besides seeds and leaves. In the first part of the study, it was compared the total phenolic content (TPC), the direct total antioxidant capacity (TAC) and the glucomoringin content (GMG) of aqueous extracts of different tissues of M. oleifera. Two different drying treatments (freeze-drying and oven drying, 40°C) were performed on the samples before extraction, to verify their influence on the chemical composition of the final extracts. The study was continued with the selection of an extraction condition able to maintain a high extraction yield of TPC and suitable for the conversion of GMG into its bioactive isothiocyanate (GMG-ITC) inside the extracts that were obtained from the tissues selected in the first part of the study because of their richness in GMG or TPC. A further objective was the determination of GMG, GMG-ITC, TPC and TAC in the extracts. In the applied extraction conditions, the branches (that have a lower TPC and TAC than leaves) could bring to an extract with an equal TPC but a higher TAC than the extract of leaves, becoming a valid alternative to leaves in the preparation of extracts rich in antioxidant compounds. The matrix “EDYNEA” showed to be an interesting alternative to leaves for the preparation of extracts richer in GMG-ITC. The third project was dedicated to oilseeds and oily extracts. The perspective was the production of an oily extract enriched in antioxidant compounds, with a higher TAC than the pure oil. It was decided to use M. oleifera oil and H. sabdariffa oils for the preparation of the extracts together with two commonly used oils in the preparation of cosmetics and for human consumption (refined olive oil and sunflower oil). The oil of H. sabdariffa was selected between different varieties of H. sabdariffa oils. The oleolites were prepared by ultrasonic extraction. The main classes of antioxidant compounds were determined as well as the TAC. The production of an oily extract higher in antioxidant components has not lead to a better TAC compared to the pure oil, at least in the extraction conditions used. Among the oils, the more active was the one extracted from the selected variety of H. sabdariffa seeds.
Gli stili di vita attuali causano un aumento del numero di radicali liberi nel nostro corpo e predispongono le persone allo stress ossidativo, responsabile dello sviluppo di diverse patologie e dell'invecchiamento. La necessità di inibitori dei processi ossidativi è diventata essenziale. Si preferiscono i composti antiossidanti derivati da piante. L’Hibiscus sabdariffa L. appartiene alla famiglia delle Malvaceae. I suoi calici contengono antocianine, responsabili dell'attività antiossidante riportata per gli estratti. La Moringa oleifera Lam. appartiene alla famiglia delle Moringaceae. Le sue proprietà antiossidanti sono principalmente attribuite ai suoi composti fenolici e, recentemente, ai suoi isotiocianati, derivanti dall'idrolisi enzimatica dei glucosinolati. Entrambe le piante sono fonte di oli con una composizione interessante in composti antiossidanti. Questo lavoro di ricerca è diviso in tre linee: la prima linea è incentrata sui calici di H. sabdariffa. È stata eseguita la selezione di una varietà di calici ricchi di antocianine. È stato ottimizzato un metodo di estrazione adatto al recupero di antocianine ad alto rendimento ma trasferibile anche su larga scala. Sono state provate diverse procedure per ottenere un estratto arricchito in antocianine usando il polivinilpolipirrolidone. La seconda linea è dedicata ai tessuti di M. oleifera. Questo studio comparativo riguarda il materiale vegetale coltivato in Paraguay e quindi amplia le informazioni disponibili sulla composizione chimica delle piante di M. oleifera coltivate in diverse aree del mondo. Per quanto ne sappiamo, questo è il primo studio sulla caratterizzazione di composti antiossidanti diretti e indiretti in diversi tessuti di M. oleifera. I rami e la matrice brevettata "EDYNEA" sono stati studiati oltre a semi e foglie. Nella prima parte dello studio, è stato confrontato il contenuto fenolico totale (TPC), la capacità antiossidante totale diretta (TAC) e il contenuto di glucomoringina (GMG) di estratti acquosi di diversi tessuti di M. oleifera. Due diversi trattamenti di essiccazione (liofilizzazione e asciugatura in forno, 40 ° C) sono stati eseguiti sui campioni prima dell'estrazione, per verificarne l'influenza sulla composizione chimica degli estratti finali. Lo studio è proseguito con la selezione di una condizione di estrazione in grado di mantenere un alto rendimento di estrazione di TPC e adatta per la conversione di GMG nel suo isotiocianato bioattivo (GMG-ITC) all'interno degli estratti che sono stati ottenuti dai tessuti selezionati nella prima parte dello studio per la loro ricchezza in GMG o TPC. Un ulteriore obiettivo è la determinazione di GMG, GMG-ITC, TPC e TAC negli estratti. Nelle condizioni di estrazione applicate, i rami (che hanno un TPC e TAC inferiori rispetto alle foglie) portano a un estratto con un TPC uguale ma un TAC superiore rispetto all'estratto di foglie, diventando una valida alternativa alle foglie nella preparazione di estratti ricchi in composti antiossidanti. La matrice "EDYNEA" si è rivelata un'alternativa interessante alle foglie per la preparazione di estratti più ricchi in GMG-ITC. Il terzo progetto è dedicato ai semi oleosi e agli estratti oleosi. L’obiettivo è la produzione di un estratto oleoso arricchito in composti antiossidanti, con un TAC superiore rispetto all'olio puro. Si è deciso di utilizzare l'olio di M. oleifera e gli oli di H. sabdariffa per la preparazione degli estratti insieme a due oli comunemente usati nella preparazione di cosmetici e per il consumo umano (olio di oliva raffinato e olio di girasole). L'olio di H. sabdariffa è stato selezionato tra diverse varietà di oli di H. sabdariffa. Gli oleoliti sono stati preparati mediante estrazione a ultrasuoni. Le principali classi di composti antiossidanti sono state caratterizzate così come il TAC. La produzione di un estratto oleoso più ricco in componenti antiossidanti non ha portato a un TAC migliore rispetto all'olio puro, almeno nelle condizioni di estrazione utilizzate. Tra gli olii, il più attivo è quello estratto dalla varietà selezionata di semi di H. sabdariffa.

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CAPES
Vegetables are a source of nutrients essential to human health and have low caloric value. Among these, non-conventional vegetables also have other advantages such as hardiness and the lowest price, but lacks information on these species which prevents an increase in its production and consumption. Thus, the objective of this study was to cultivate the species Hibiscus sabdariffa, better known as vinagreira, characterize some of its constituents and estimate its shelf life potential when stored in polypropylene packaging with different perforations and stored at 5 ? 1 ?C. The vinagreira remained for approximately 120 days in field conditions in the Horticultural Sector UFRRJ. The branches were collected and transported to the Embrapa Food, which were washed, selected, arranged in bunches of about 150g and subjected to treatment consisting of the conical packaging polypropylene, perforated (P), not perforated (NP), micro perforated (MP) and control (C) in which the packs were stored without use of packaging. They were carried physical, physicochemical and chemical every two days storage. The design was completely randomized in split plots, and treatments in the plots and the days of storage in the subplot. The results were analyzed using SPSS. It was the homogeneity and normality of the data, followed by analysis of variance and significance according to Tukey test and/or regression analysis. For control, the analyzes were performed only until the fourth day of storage due to reduction of apparent quality such as wilting and color changes of the leaves, while those packaged in decreased the apparent quality, with symptoms of senescence the eighth day of storage. The leaves have a high levels of titratable acidity and carotenoids, being the predominant malic acid and the major carotenoid, the ?-carotene. It was concluded that vinagreira is well adapted Climate conditions Serop?dica-RJ, showing the source of compounds with antioxidant activity. With regard to cold storage, the use of plastic polypropylene packaging provided greater potential life for vegetables, because in these conditions the bunches could be sold for up to seven days after harvest without loss of quality, while the control bunches potential shelf life was less four days.
As hortali?as s?o fonte de nutrientes essenciais ? sa?de humana e possuem baixo valor cal?rico. Dentre estas, as hortali?as n?o-convencionais apresentam ainda outras vantagens como a rusticidade e o menor pre?o, por?m faltam informa??es sobre estas esp?cies o que impossibilita um aumento da sua produ??o e consumo. Desse modo, o objetivo do presente trabalho foi cultivar a esp?cie Hibiscus sabdariffa, mais conhecida como vinagreira, caracterizar alguns dos seus constituintes e estimar a sua vida ?til potencial, quando acondicionada em embalagens de polipropileno com diferentes perfura??es e armazenadas a 5 ? 1 ?C. A vinagreira permaneceu por aproximadamente 120 dias em condi??es de campo no Setor de Horticultura da UFRRJ. Os ramos foram colhidos e transportados para a Embrapa Agroind?stria de Alimentos, onde foram lavados, selecionados, arranjados em ma?os de aproximadamente 150 g e submetidos aos tratamentos, constitu?dos pelas embalagens c?nicas de polipropileno, perfurada (P), n?o perfurada (NP), micro perfurada (MP) e o controle (C) em que foram armazenados sem o uso de embalagens. Foram realizadas an?lises f?sicas, f?sico-qu?micas e qu?micas a cada dois dias do armazenamento. O delineamento utilizado foi o inteiramente casualizado, em esquema de parcelas subdivididas, tendo os tratamentos nas parcelas e os dias de armazenamento na subparcela. Os resultados foram submetidos a an?lises estat?sticas pelo programa SPSS. Verificou-se a homogeneidade e normalidade dos dados, com posterior an?lise de vari?ncia e de acordo com a signific?ncia procedeu-se o teste Tukey e/ou an?lise de regress?o. Para o controle, as an?lises foram realizadas somente at? o quarto dia do armazenamento, devido ? redu??o da qualidade aparente, como murchamento e altera??es de cor das folhas, enquanto para os ma?os acondicionados nas embalagens houve redu??o da qualidade aparente e sintomas de senesc?ncia ao oitavo dia do armazenamento. As folhas apresentaram elevado teor de acidez total titul?vel e de carotenoides, sendo o ?cido predominante o m?lico e o carotenoide majorit?rio, o ?-caroteno. Concluiu-se que a vinagreira se adapta bem as condi??es clim?ticas de Serop?dica-RJ, mostrando-se fonte de compostos com atividade antioxidante. Em rela??o ao armazenamento refrigerado, a utiliza??o de embalagens pl?sticas de polipropileno proporcionou maior vida ?til potencial para a hortali?a, pois nestas condi??es os ma?os poderiam ser comercializados por at? sete dias ap?s a colheita, sem preju?zo da qualidade, enquanto os ma?os do controle a vida ?til potencial foi inferior a quatro dias.

L’extraction des polyphénols de feuilles de Carapa procera, de fleurs de Delonix regia et de calices d Hibiscus sabdariffa est optimisée à l'échelle du laboratoire pour obtenir des extraits rouge riches en antioxydants. Les anthocyanes, les flavonols et les acides phénols de ces 3 plantes endémiques de Côte d'Ivoire ont été caractérisés par CL-SM et RMN pour la première fois dans les 2 premières plantes et confirmés dans la troisième. La macération 12-16h dans l' eau d échantillons séchés localement, la macération 1h avec assistance par ultrasons, la microfiltration et la concentration des extraits à l'échelle pilote a permis la production de 250L de microfiltrat/plante avec des débits moyens de 100 à 200 L. H-1. M-2. B-1 à 0,6 bar, de concentrer de 100 à 120 fois le volume de ces microflltrats à 16 et 20 L. H-1. M-2 à 40 bar et a 20° C et de produire par atomisation, avec des rendements de 5 à 10% des poudres d' extraits polyphénoliques (30 à 160 [micro]mol. G-1 AG eq), dont on a déterminé la capacité antioxydante par DPPH et ABTS (de 8 à 30 mmol. L-1 AG eq pour les concentrés d'OI et 220 à 500 mmol. G-1 TEAC pour les poudres)
The extraction of the polyphenols of Carapa procera leaves, of Delonix regia flowers and of Hibiscus sabdariffa calyx was optimized at laboratory scale to obtain a red extracts rich in antioxidants. Anthocyanins, flavonols and phenolic acids of these 3 endemic plants from Ivory Coast were characterized by LC-MS and NMR for the first time in the 2 first plants and confirmed in the third. Water maceration, 12-16h time, of locally dried samples, 1hour time ultrasound-assisted maceration, microfiltration and concentration of the extracts at pilot plant scale allowed the production of 250L of microfiltrate/plant processed with average MFT flows of 100 to 200 L. H-1. M-2. B-1 at 0,6 bar, the volume concentration of the microfiltrates of 100 to 120 times with 16 and 20 L. H -1. M -2 fluxes at 40 bar and at 20°C and to produce powders by atomization, with a 5 to 10 % yield, of polyphenolic extracts (30 in 160 [micro]mol. L-1 AG eq), of which the antioxidant capacity was determined by DPPH and ABTS (from 8 to 30 mmol. L-1 AG eq for RO concentrates and from 220 to 500 mmol. G-1 TEAC for powders)

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La dalia (Dahlia coccinea Cav.) es una planta que ha sido cultivada y mejorada genéticamente con fines ornamentales; no obstante, su sistema radicular almacena carbohidratos de reserva bajo la forma de inulina y otros fructanos. La inulina forma parte de la fibra dietética de diversos vegetales y es considerada como un compuesto prebiótico. Este polisacárido se extrae principalmente de la achicoria y la alcachofa mediante métodos de separación muy variados, siendo la raíz de la dalia una opción interesante de explotación industrial. El propósito de este estudio fue proponer un método simple de extracción de inulina de la raíz de dalia y evaluar el efecto del cultivo de dalias silvestres sobre su contenido de inulina, fructanos, fructosa y glucosa. Se realizaron colectas de dalia silvestre y se propuso un método de extracción de inulina. Las semillas de los ejemplares recolectados fueron cultivadas y a las raíces obtenidas se les determinó la concentración de fructanos, inulina, fructosa y glucosa y se estimó el grado de polimerización. La relación tiempo-temperatura afecta significativamente la concentración de inulina y fructanos extraídos, siendo la combinación óptima 80° C durante 60 minutos. La influencia del pH resultó no significativa para este mismo propósito. Por otro lado, el cultivo de las dalias silvestres aumenta significativamente el contenido de inulina y fructanos de sus raíces, aunque el grado de polimerización, estimado a partir de la relación fructosa/glucosa, disminuye. La liberación de polifenoles encapsulados en inulina en una solución isotónica sigue una cinética de primer orden con una constante de velocidad (basada en la absorbancia del medio isotónico) de 0.0148 unidades por minuto y una liberación máxima de 0.4166 unidades de absorbancia que se observaron a las 8 horas de permanencia de las partículas en la solución isotónica.
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L'Hibiscus sabdariffa (hibiscus) est une plante dont les calices comestibles, contiennent des polyphénols et des molécules d'anthocyane qui sont à la fois des composés antioxydants et responsables de leur couleur rouge attrayante pour le consommateur. Ces particularités constituent un fort potentiel santé qui répond aux demandes actuelles du marché et un atout économique pour les industries. Cependant, cette plante tropicale et saisonnière dont les calices riches en eau, sont très périssables, sensibles à l'humidité et à la chaleur, et les molécules d'intérêt (anthocyanes) sont thermosensibles et instables en milieu aqueux. Dans ce contexte, la transformation des calices en poudre permet, en plus d'optimiser le coût du transport et l'espace de stockage, de stabiliser les calices en assurant une longue durée de conservation du produit, de faciliter l'accessibilité aux biomolécules et donc à ses bienfaits pour la santé, ceci, à condition que le procédé soit maîtrisé. L'un des meilleurs moyens de préserver les propriétés nutritionnelles d'origine (teneur en anthocyanes, activité antioxydante) des calices est d'obtenir des produits peu transformés ou « minimally processed products » en combinant le séchage à l'étuve (contrôlé), le broyage et le fractionnement par tamisage. Ce procédé a été étudié dans ce travail et les propriétés physicochimiques (granulométrie, forme, structure, porosité, proportion de fibre, teneur en anthocyanes, activité antioxydante) et fonctionnelles des poudres (écoulement, reconstitution, extractibilité des biomolécules) ont été systématiquement évaluées dans un but de formulation de boisson. L'intérêt d'un tel procédé est la facilité de sa mise en œuvre, applicable à tous végétaux, la facilité d'usage de la poudre, l'amélioration de la disponibilité et l'accessibilité des biomolécules. Ce travail a permis d'identifier l'impact du séchage solaire et du broyage sur les propriétés physicochimiques des poudres, ces dernières impactant directement les propriétés fonctionnelles. La substitution du séchage solaire par le séchage contrôlé à l'étuve et un fractionnement par tamisage supplémentaire après le broyage ont permis d'améliorer les propriétés fonctionnelles des poudres. Ces propriétés singulières des poudres ainsi obtenues, ont permis de distinguer des applications qui leur sont propres
Hibiscus sabdariffa (hibiscus) is a plant with edible calyxes containing polyphenol and anthocyanin molecules that are both antioxidant compounds and responsible for their attractive red color for consumers. These particularities constitute a good health potential that meets the current market demands, and an economic potential for industries. However, this tropical and seasonal plant whose water-rich calyxes are highly perishable, sensitive to humidity and heat, and whose the interesting molecules (anthocyanins) are heat-sensitive and unstable in aqueous medium. In this context, the calyx transformation into powder allows, besides optimizing the transport cost and storage space, to stabilize the calyxes by ensuring a long product shelf-life, and to improve the biomolecule accessibility. This allows to benefit to its health assets provided that the process is controlled. One of the best ways to preserve the calyx original nutritional properties (anthocyanin content, antioxidant activity) is to obtain minimally processed products by combining controlled oven-drying, grinding and fractionation by sieving. This process was studied in this work and the powder physicochemical properties (particle size and shape, structure, porosity, fiber proportion, anthocyanin content, antioxidant activity) and functional properties (flowability, reconstitution, biomolecule extractability) were systematically evaluated for drink formulation. The interest of such a process is the ease of its implementation applicable to all plants, the powder ease of use, the improvement of the biomolecule availability and accessibility. This work allows identifying the impact of sun-drying and grinding on the powder physicochemical properties, the latter impacting the functional properties. The substitution of sun-drying by controlled oven-drying and an additional fractionation by sieving preceded by grinding allowed improving the powder functional properties. These powder functional properties were singular and allowed to distinguish specific applications for each powder type (fine or coarse powders)

RIASSUNTO E SCOPO DEL LAVORO Esiste una stretta correlazione tra obesità e osteoporosi: nell'obesità si riscontra una maggiore fragilità ossea ed un ridotto assorbimento di calcio a livello intestinale, mentre l'osteoporosi è spesso accompagnata da un aumento dell'adipogenesi midollare. In questi ultimi anni un'area della ricerca scientifica si è concentrata sullo studio di nuovi farmaci/sostanze in grado di agire sull'adipogenesi o sull'osteoblastogenesi in quanto è fondamentale trovare nuove terapie che prevengano tali patologie. Conoscendo la stretta relazione tra il differenziamento adipogenico e osteogenico, un farmaco contro l'obesità non dovrebbe avere effetti sul metabolismo osseo, mentre contro l'osteoporosi non dovrebbe avere effetti sull'adipogenesi. A tale proposito gli studi in vitro rappresentano un punto importante sia per valutare l’effetto di tali sostanze che per comprendere i meccanismi alla base delle loro azioni. La maggior parte degli studi condotti sul differenziamento adipocitico, e sulla sua inibizione, sono stati condotti su linee di preadipociti murini (3T3-L1) o umani, mentre sono state utilizzate linee murine MC3T3-E1 e C3H10T1/2 per il differenziamento osteogenico. Nonostante la rilevanza dei dati ottenuti grazie a questi modelli cellulari, i risultati sono spesso controversi, anche in considerazione del fatto che i processi differenziativi nell’uomo e nei roditori possono essere regolati a livello molecolare in modo significativamente diverso. Inoltre questi modelli sono basati su cellule già indirizzate verso il differenziamento adipocitico o osteogenico, e quindi non prendono in considerazione il processo di determinazione che porta la cellula progenitrice ed indifferenziata verso un lineage mesengenico. Le cellule staminali mesenchimali (MSC) in questo contesto rappresentano un modello cellulare particolarmente utile in quanto possono essere efficacemente indotte da uno stato indifferenziato allo stato di adipociti o osteoblasti, di cui peraltro rappresentano fisiologicamente i precursori biologici. Lo scopo di questa tesi è stato quello di valutare l'effetto di Acido Valproico (AVP), Valpromide (VPM), Berberina (BRB), Hibiscus Sabdariffa (HIB) e Resveratrolo (RESV) sul differenziamento adipogenico e osteogenico di cellule staminali mesenchimali umane (hMSC). Le hMSC utilizzate in questo studio sono state caratterizzate e corrispondono ai criteri minimi stabiliti dalla letteratura (Dominici et al., 2006) per poterle definire MSC. Tre le condizioni necessarie: capacità di aderire alla plastica assumendo una morfologia simil-fibroblastica; negatività per i marker ematopoietici CD34 e CD45 e positività per i marker CD29, CD90, CD105 e CD73; capacità, sotto opportune condizioni di coltura, di differenziare nei lineage mesengenici ovvero in osteoblasti, condroblasti ed adipociti. Questo lavoro si è articolato in 5 fasi: 1) Valutazione della tossicità delle diverse sostanze. Per valutare la tossicità delle sostanze precedentemente elencate, le hMSC indifferenziate e le MSC differenziate in senso adipogenico e osteogenico sono state trattate con ciascuna delle diverse sostanze a differenti concentrazioni e sono stati effettuati saggi di vitalità. Sono stati condotti due tipi di test: MTT assay, che valuta l'attività mitocondriale come indicatore della vitalità cellulare e SRB assay, che valuta il contenuto proteico per stimare il numero delle cellule vitali. Sulla base dei risultati ottenuti sono state individuate le concentrazioni non tossiche dei diversi composti da utilizzare negli esperimenti successivi. 2) Effetto delle diverse sostanze sul differenziamento adipogenico e osteogenico. A partire da hMSC indifferenziate è possibile studiare l’intero processo adipogenico trattando le cellule con un terreno (DMEM high glucose) di induzione contenente fattori adipogenici (desametasone 1μM, indometacina/isobutilmetilxantina 100μM/500μM, insulina 10μg/ml) per i primi 10 giorni e successivamente, fino al 28° giorno, con un terreno di mantenimento (contenente insulina 10μg/ml) per la maturazione in senso adipogenico. Le diverse sostanze sono state somministrate alle hMSC sin dal momento dell'induzione e portate fino al 28° giorno di differenziamento adipogenico. L'effetto anti-adipogenico è stato valutato mediante colorazioni istologiche specifiche per i trigliceridi presenti nei depositi lipidici degli adipociti: la colorazione con Oil Red O per osservare la morfologia cellulare mentre la colorazione Nile Red, esaminata mediante citofluorimetro a flusso, per determinare il numero di cellule differenziate. I risultati dimostrano che AVP, BRB, HIB e RESV presentano un effetto anti-adipogenico, mentre VPM non ha dimostrato avere effetto sul differenziamento adipogenico. Successivamente è stato valutato l'effetto di AVP, VPM e HIB aggiunte dopo 3, 7, 10, 14 e 21 giorni dall'induzione adipogenica e analizzate tutte al 28°giorno. Con questi dati abbiamo dimostrato che solo AVP ha un effetto anti-adipogenico, anche quando viene aggiunto dopo 3 o 7 giorni dall'induzione. L'aggiunta di VPM e HIB a tempi diversi dall'induzione non ha portato a delle variazioni significative rispetto alle hMSC indotte con solo il terreno adipogenico. Per il differenziamento osteogenico le hMSC sono state trattate con uno specifico terreno (DMEM low glucose) contenente fattori osteogenici (desametasone 100nM, acido ascorbico 2-fosfato 50μM e βglicerolo 2-fosfato10mM) fino al 28° giorno. La valutazione del differenziamento osteogenico è stata effettuata mediante la colorazione istologica Alizarin red S, che evidenzia la deposizione di calcio mineralizzato. Le diverse sostanze, alle dosi precedentemente scelte, sono state aggiunte inizialmente al terreno osteogenico delle hMSC e portate fino al 28°giorno di trattamento. Da questi esperimenti abbiamo ottenuto risultati differenti per le diverse sostanze valutate. AVP e VPM determinano un precoce differenziamento osteogenico delle hMSC rispetto alle cellule trattate con solo il terreno di differenziamento. Infatti in presenza di AVP e VPM il differenziamento si osserva già dopo 10gg rispetto alle cellule trattate con solo terreno osteogenico. BRB e HIB presentano invece un effetto anti-osteogenico, poichè con l’aggiunta di queste sostanze nel terreno non si osserva deposito di matrice mineralizzata e la colorazione istologica risulta negativa. Le cellule trattate con RESV non mostrano differenze significative rispetto alle cellule trattate con solo terreno osteogenico. Successivamente per AVP, VPM e HIB è stato valutato l'effetto della sostanza aggiunta dopo 3, 7, 10, 14 e 21 giorni dall'induzione osteogenica e analizzate tutte al 28°giorno. Da questi dati abbiamo dimostrato che HIB ha un effetto anti-osteogenico anche quando aggiunta dopo 3, 7, 10 e 14 giorni dall'induzione. L'aggiunta di AVP e VPM a tempi diversi dall'induzione non ha portato variazioni significative rispetto alle hMSC indotte con solo il terreno osteogenico. 3) Valutazione dell'effetto delle sostanze sulla fosforilazione di ERK1/2 durante il differenziamento adipogenico e osteogenico. Mediante esperimenti di immunoblotting abbiamo studiato la fosforilazione (forma attiva) di ERK1/2 per poter capire i meccanismi molecolari alla base delle azioni di tali sostanze nei due diversi differenziamenti. Nelle hMSC, trattate per il differenziamento adipogenico con l'aggiunta di AVP, VPM, BRB, HIB e RESV non si sono osservate variazioni significative nella fosforilazione di ERK1/2 rispetto alle hMSC indotte con solo il terreno adipogenico. Nel differenziamento osteogenico abbiamo osservato a 10giorni una riduzione significativa della forma fosforilata di ERK1 nelle cellule trattate con VPM o BRB rispetto alle cellule trattate con solo terreno osteogenico; la forma fosforilata di ERK2 non varia. A 14giorni solo le cellule trattate con terreno osteogenico e BRB presentano una riduzione significativa di entrambe le forme pERK1/2 rispetto alle cellule trattate con solo terreno osteogenico. 4) Valutazione dell'effetto di Acido Valproico, Valpromide e Hibiscus Sabdariffa sui livelli di fosforilazione di GSK3β e di espressione di βcatenina. Per comprendere meglio il meccanismo d'azione di AVP, VPM e HIB abbiamo studiato, mediante esperimenti di immunoblotting, i livelli di fosforilazione (forma inattiva) di GSK3β e i livelli di espressione di βcatenina nelle hMSC trattate per il differenziamento adipogenico e osteogenico con l'aggiunta delle tre sostanze. Per quanto riguarda il differenziamento adipogenico solo a 3giorni di induzione abbiamo osservato nelle hMSC indotte al differenziamento con l'aggiunta di AVP un aumento significativo di pGSK3β, rispetto alle hMSC trattate con solo il terreno adipogenico. Analizzando i livelli di βcatenina a 12ore, 1, 3giorni non abbiamo rilevato variazioni significative tra le hMSC indotte al differenziamento adipogenico sia in presenza che in assenza delle sostanze. Analizzando i dati relativi al confronto tra le hMSC indotte al differenziamento osteogenico, in presenza o assenza delle sostanze, non abbiamo riscontrato differenze significative nei livelli di pGSK3β per tutti i tempi analizzati. Abbiamo osservato una riduzione significativa nei livelli di βcatenina a 3, 10 e 14giorni in hMSC trattate per il differenziamento osteogenico con l'aggiunta di AVP, mentre solo a 10giorni si osserva una riduzione significativa di βcatenina anche per le hMSC trattate con VPM rispetto alle hMSC indotte con solo terreno osteogenico. 5) Valutazione dell'effetto di Acido Valproico, Valpromide e Hibiscus Sabdariffa sull'espressione genica di C/EBPβ, PPARγ, RUNX2 e OSTERIX. Infine è stato valutato, mediante qPCR, l’effetto di AVP, VPM e HIB sull'espressione genica di C/EBPβ, PPARγ, RUNX2 e OSTERIX (importanti fattori trascrizionali adipogenici e osteogenici) durante il differenziamento adipogenico e osteogenico di hMSC. Dai risultati ottenuti si osserva che le hMSC indotte al differenziamento adipogenico con l'aggiunta di AVP hanno dei livelli di espressione significativamente maggiori per C/EBPβ e significativamente minori per PPARγ rispetto alle hMSC indotte con solo terreno adipogenico. Durante il differenziamento osteogenico si osserva un aumento significativo di Runx2 nelle hMSC trattate con AVP rispetto alle hMSC indotte con solo terreno osteogenico. L'analisi dei nostri risultati ha evidenziato che la sostanza più promettente per un eventuale uso terapeutico sembra essere l’Acido Valproico perché, oltre ad inibire il differenziamento adipogenico, sembra avere un effetto pro-osteogenico sulle hMSC indotte al differenziamento osteogenico anche se saranno necessari ulteriori studi per comprendere a pieno il meccanismo d'azione di tale sostanza.

As bebidas à base de extratos de hibisco são ricas em ácidos orgânicos, compostos fenólicos e polissacáridos, substâncias com potenciais benefícios para a saúde, sendo tradicionalmente consumidas na Africa Ocidental, na Ásia e na parte sul da América do Norte. A sua divulgação na Europa e dos Estados Unidos tem sido lenta e dirigida sobretudo a nichos de mercado, no entanto, o crescente interesse dos consumidores europeus e norte-americanos em bebidas com ingredientes únicos ou exóticos, com potenciais benefícios para a saúde, criaram recentemente novas oportunidades de mercado para bebidas de hibisco de alta qualidade. Entre 2010 e 2015, o projeto Europeu African Food Tradition rEvisited by Research (AFTER) investigou a produção, a comercialização e o consumo de bebidas de hibisco (Hibiscus sabdariffa var. sabdariffa ruber) no Senegal e na Europa, com o objetivo de otimizar a sua produção industrial, minimizando a degradação dos compostos fitoquímicos. Pouco se sabia sobre o perfil sensorial destes produtos, na sua forma tradicional ou melhorada e sobre como as mesmas eram avaliadas pelos consumidores, fatores essenciais para o sucesso destes produtos em particular fora dos seus mercados tradicionais. Esta dissertação teve como objetivo avaliar o impacto da otimização dos processos de produção industrial das bebidas de hibisco levadas a cabo pelo projeto AFTER (1) nas propriedades físico-químicas dos produtos resultantes com impacto na qualidade sensorial; (2) no seu perfil sensorial; (3) na perceção e aceitação por parte dos consumidores em mercados tradicionais (Senegal) e potenciais (França, Portugal e Reino Unido). O uso de cálices de hibisco secos à sombra e triturados das cultivares Vimto e Koor (50:50) e de condições moderadas de tempo e temperatura de extração e pasteurização, deu origem a novas bebidas com uma cor mais intensa e mais vermelha, maior concentração em antocianas monoméricas e polifenois e consequentemente maior capacidade antioxidante do que as bebidas convencionais, maior equilíbrio entre a concentração de açúcares e de ácidos e maior intensidade aromática. O estudo efetuado com consumidores no Senegal (N=150), mostrou que as novas bebidas foram positivamente avaliadas e melhor apreciadas que bebidas convencionais, sem que a perceção do caracter tradicional tenha sido alterada. Três segmentos de consumidores com distintos padrões de aceitação das bebidas foram identificados. A avaliação hedónica foi influenciada por preferências pessoais relativamente à cor, aroma e equilíbrio doçura-acidez, por características sociodemográficas e hábitos de consumo. Vinte e duas bebidas de hibisco foram avaliadas por painéis de provadores no Senegal e em Portugal. Léxicos em Português, Inglês e Francês contendo 21 descritores definidos, e referenciados foram desenvolvidos. As características distintivas das bebidas de hibisco eram a cor vermelha, o aroma floral (hibisco) e frutado (aronia/arando, ginja e framboesa) e o sabor predominantemente ácido (infusões de hibisco e bebidas preparadas a partir de concentrados) ou doce (bebidas preparadas a partir de xaropes). Eram ainda percetíveis aromas de chá preto, passa de uva, mel e feno, assim como sabor amargo e adstringência. O vocabulário usado por profissionais não-treinados (N=30) e por consumidores foi estudado usando entrevistas semiestruturadas (N=75) e questões Check-all-that-apply (N=490) no Senegal, em França, Portugal e Reino Unido. Léxicos gráficos contendo 27 atributos sensoriais e 15 termos gerais foram desenvolvidos. Diferenças com origem na cultura e familiaridade com o produto foram observadas. A comparação dos perfis sensoriais obtidos com provadores treinados e consumidores permitiu uma melhor compreensão da linguagem destes últimos. Estudos com consumidores em França, Portugal e Reino Unido (N=403) mostraram padrões similares de aceitação e perceção pelos consumidores. Diferenças importantes foram observadas relativamente ao Senegal. Na generalidade, os consumidores na Europa, gostaram menos das bebidas do que no Senegal, embora avaliações hedónicas médias positivas tenham sido observadas para todas as bebidas. No Senegal os consumidores avaliaram mais favoravelmente uma maior intensidade de cor e maior doçura e de forma mais depreciativa menor acidez. Na Europa e caracter floral e frutado das bebidas constituíram importantes fatores de aceitação das mesmas. Segmentos de consumidores foram identificados. A aceitação e perceção foram influenciadas pela familiaridade com o produto e com produtos com características de algum modo similares na Europa (caso dos sumos de frutos vermelhos); por características sociodemográficas; e por preferências individuais inatas ou culturalmente adquiridas. Foram observadas diferenças entre segmentos de consumidores na Europa relativamente ao perfil sensorial de uma bebida ideal, mas não no Senegal.
Hibiscus beverages are rich in organic acids, phenolic compounds and polysaccharides, which have been demonstrated to be linked to important health benefits. They are traditionally consumed in West African countries, in Asia and in the Southern part of the North American continent. In Europe and United States of America, hibiscus beverages have slowly penetrated niche markets. Due to the rising number of health conscious consumers, along with the growing interest of North Americans and Europeans in beverages made from unique or exotic ingredients, new mainstream market opportunities for high quality hibiscus beverages have been recently created. Between 2010 and 2015, the African Food Tradition rEvisited by Research (AFTER) project, investigated the production, trading and consumption of hibiscus beverages made from dried hibiscus (Hibiscus sabdariffa var. sabdariffa ruber) calyx extracts in Senegal and Europe. Key improvements were proposed to increase production process efficiency and minimize degradation of phytochemical compounds. The aims of this thesis encompassed the evaluation of the impact of the reengineering processes held under AFTER project scope (1) in the main chemical-sensory properties of hibiscus beverages; (2) on their sensory profile; (3) on consumer acceptance in traditional markets (Senegal) and potential markets (France, Portugal and United Kingdom). The selection of a blend of shade dried ground hibiscus calyces (Vimto:Koor cultivars 50:50) and the use of mild water extraction and pasteurization conditions, led to the production of beverages with a more intense redder colour, higher monomeric anthocyanin, total phenolic content and higher antioxidant capacity than their conventional counterparts. Colour evaluation by trained panelists was mainly linked to colour density and to anthocyanin/polyphenol content, while flavour assessments were associated to titratable acidity and sugar-to-acid ratio. New beverages presented also a more intense aroma than conventional ones. Results of hedonic assessments held in Senegal (N=150), indicated that all improved beverages were liked by roughly three-quarters of consumers and were better accepted than a traditional one (baseline comparison). Three clusters of consumers with distinct overall liking patterns were identified. Acceptance was driven personal preferences regarding colour intensity, aromatic character, balance between sweetness and acidity but also by gender, age frequency of consumption and education level. Importantly, new beverages were still perceived by consumers as traditional products. Given the scarcity of information about hibiscus beverages sensory profile, sensory lexicons were developed in French, Portuguese and English. Twenty-two samples, including freshly prepared and ready-to-drink infusions, syrups, concentrates and an instant tea, were evaluated by trained panelists, resulting in 21 defined and referenced descriptors, subsequently assembled in a sensory wheel. A distinctive red colour, a floral (hibiscus) and fruity aroma (aronia/cranberry, sour cherry and raspberry) and a predominant acid (in case of infusions and beverages prepared from concentrates) or sweet taste (for beverages prepared from syrups) were the main sensory attributes. Also moderate or low intensities of black tea, honey, raisin and hay odours and of bitter taste and astringency were perceived in all products. The vocabulary used by untrained panelists was also investigated in Europe and Senegal through professional meetings (N=30), consumer focus groups (N=75) and check-all-that-apply questions (N=490), resulting in graphical lexicons with 27 sensory and 15 overall impression terms. Training, food culture and product familiarity influenced results. Trained panelists used precise terms to describe the aromatic profile of hibiscus beverages (e.g., reminiscent of dried hibiscus flowers, with berry, raisin, hay and sweet notes), whereas untrained ones were more prolific in general expressions (e.g., strong in hibiscus in Senegal; floral and fruity in Europe). Consumer studies in France, Portugal, United Kingdom (N=403), showed similar patterns of acceptance and perception for hibiscus beverages in European countries. Positive mean overall liking ratings were observed for all beverages. Major differences were observed relatively to Senegal where consumers favored higher intensities of red colour and sweetness. In Europe fruity and floral character constituted important drivers of liking. Consumer segments were identified. Perception and acceptance was influenced by consumers’ sociodemographic characteristics and by innate and culturally acquired preferences. Besides exposure to product category, also exposure and liking of familiar products, with somewhat similar characteristics to hibiscus beverages (like red fruit beverages), was shown to influence liking in Europe. Differences across consumer segments in the profile of an ideal beverage were observed in Europe but not in Senegal.

碩士
靜宜大學
化粧品科學系
104
Hibiscus sabdariffa also known as Roselle, it is often used as a plant health. Roselle at different pH environment due to the contents of the different enzymes and showing a different color. Therefore, this study will be of chitosan as the shell material, and Hibiscus sabdariffa extract as a core material, made into chitosan microcapsules. The Hibiscus sabdariffa extract was covered, and may be used in the skin care and cosmetics products. It has a natural pigment and sustained effect. In this study, morphological appearance Roselle microcapsules, particle size, entrapment efficiency, pH value changes, antioxidant capacity, and percutaneous infiltration to be explored. The microscopic morphology and entrapment efficiency calculated that the experimental data results chitosan has been successfully prepared Roselle microcapsules. When the Roselle extract was added in an amount of 35ml, the entrapment efficiency can up to 70 percent. In the microscopic observation that microcapsules showing spherical and having a dark periphery. The dark periphery was the Roselle natural pigment. After chitosan coated Roselle extract, the color can be changed with different pH value. In this study, the pH was adjusted from pH 4 to pH 11, in color will from red turn green and then yellow. Therefore it can be used to adjust pH to change color. And when tripolyphosphate added as a curing agent, that significantly reduced the average particle diameter of the microcapsules. No curing agent microcapsule particle size of about 2.0μm, but after adding the curing agent is less than 1.0μm. It confirmed that chitosan does with sodium tripolyphosphate as cross-linked to form colloidal particles. It becomes smaller particles will more readily penetrate into the skin and be absorbed. Determination by the percutaneous absorption confirmed this result that polysaccharide chitosan microcapsules after curing to increase the efficiency of the percutaneous penetration. The test can enhance the absorption of almost 5 times after 120 minutes. In terms of antioxidant will have a small amount to enhance the effect, there is no phenomenon of recession. Whereby it can be utilized on cosmetic products to enhance the polyphenols contained within Roselle into the skin to reach the antioxidant capacity.

碩士
中山醫學大學
生化暨生物科技研究所
95
Acetaminophen (AAP) possessed analgesic and antipyretic properties; but it could induce hepatic necrosis in centrilobular cell. The major cause of hepatic damage was contributed by the reactive intermediate N-acetyl-p-benzoquinone imine (NAPQI). NAPQI accumulation was due to protein arylation and severe oxidative stress to cause liver injury. It was demonstrated that Hibisus Sabdariffa (HS) potentiated antioxidative effect in previous studies. In this study, we investigated HS extracts against AAP induced liver injury in BABL/c mice. In vivo, BABL/c mice were fed orally with HS polyphenols extract (HPE) (100, 200, 300 mg/kg) or HS aqueous extract (HSE) (200, 400, 600 mg/kg) for two weeks; and then 1000mg/kg of AAP was injected i.p. At 6h after APAP injection, mice were decapitated. In vitro, BABL/c normal liver (BNL) cell was pretreated with AAP 5mM or HPE/HSE (0.05, 0.1, 0.5, 1 mg/kg) for 48 hrs. Base on that, we investigated the possible mechanism. The results showed that after AAP administration, pretreatment of HPE (100, 200, 300 mg/kg) or HSE (200, 400, 600 mg/kg) decreased the levels of lipid peroxidation to 63.2%, 70.1%, 83.1% (HPE) or 70.6%, 65.1%, 54.1%(HSE) ；catalase increased to 22%, 24.4%, 16.2% (HPE) or 45.7%, 44.3%, 55.8%(HSE), glutathione increased to 43.4%, 25.2%, 27.9%(HPE) or 41.5%, 45.7%, 54.2%(HSE). In histopathological evaluation of liver, HSE or HPE can decrease AAP induced liver injury. Both in vivo and in vitro, HSE or HPE can decreased the expressions of JNK, iNOS, Bax, Bid. Futhermore in vitro assay, we also found that AAP induced cell injury could be decreased by JNK inhibitor as well as HSE/HPE. In conclusion, we have demonstrated that HPE or HSE can protect liver from AAP induced injury. The protecting mechanism could involve inhibition of (1) JNK1/2 activation coupled with inhibition of Bax/Bid activation; (2) induction of oxidative stress.

碩士
中山醫學大學
生化暨生物科技研究所
96
Diabetic ophthalmopathy is the leading cause of acquired blindness in the world. Diabetes sustained hyperglycemia circumstances, produced abnormal metabolism products to influence eye normal function, besides decreased antioxidant system, and furthermore produced free radical to damage eye. It was demonstrated that Hibisus Sabdariffa potentiated antioxidative effect in previous studies. In this study, we further examined the effect of HSE or HPE on anti-ophthalmopathy. In Sprague-Dawley (SD) Rat experiment, after diabetes induction completed, HSE (100,200 and 300mg/kg), HPE (100,200 and 300 mg/kg) were fed by oral tube for nine weeks to observe the effects on diabetic rats. The results showed that treatment of HSE&HPE (100, 200 and 300 mg/kg) did not restrain the blood glucose. However, 100, 200 and 300mg/kg of HSE increased catalase activity to 13.2%, 10.1%, 13.8% ; and glutathione was increased to 9.2%, 8.2%, 8.4%. In the groups treated with 200 and 300mg/kg of HPE, catalase activity was increased to 4.4%, 3.3%; and glutathione was increased to 28%, 78%. In lens, HSE could decelerate the development of diabetic cataract by empiricism. In histopathological evalution of retina, HSE and HPE decreased diabetes induced retinal injury. We also found that in eye tissues, HSE or HPE cannot affect phosphorylation of Akt, but downregulate phosphorylation of extracellular signal-regulated kinase (ERK), p38 and JNK to reduce apoptosis. In retinal pigment epithelial (RPE) cell experiment, cell was pretreated with 200 μM of H2O2 accompanied with HSE or HPE (0.5, 1, 2, 3 mg/mL) for 6 hrs. The results showed that HPE could protect RPE cell from H2O2 induced damage . We also found HPE cannot affect phosphorylation of Akt, but downregulate phosphorylation of extracellular signal-regulated kinase (ERK) to reduce apoptosis. In conclusion, we have demonstrated that HPE or HSE could have potential benefits in inhibiting diabetes induced ophthalmopathy.

Submitted in complete fulfilment for the Degree of Master in Food Science and Technology, Durban University of Technology, 2017.
Hibiscus sabdariffa is an under-utilised plant that has been reported to have great potential in the pharmaceutical and nutraceutical industries. The vibrant red pigment indicates a source of anthocyanins that could be produced into a food colourant with additional nutritional benefits however stability is a hindering factor. The crude anthocyanins were extracted from dried calyces by means of four different acidified ethanol and methanol solvent systems to determine the maximum crude anthocyanin yield. The crude extracts were analysed under the following parameters; heat, light, pH stability and degradation kinetics, which included thermal degradation and DPPH radical scavenging ability. Two synthetic colourants were analysed based on the stability parameters; heat, light and pH and compared with the natural H. sabdariffa crude extracts. Each of the four crude extracts were analysed for the total phenolic content using Folin Ciocalteu’s method. The DPPH and FRAP assays were used to determine the radical scavenging activity of the extract with the highest yield. The identification and quantification of the crude anthocyanins were carried out using HPLC-DAD. The highest crude anthocyanin yield of 19.92% was observed by HCl acidified ethanol extract Acetic acid/water/methanol extract produced the lowest yield of 8.72%. The stability results showed that pigment retention of samples heated at 80˚C had a greater decrease over time than those heated at 50˚C. The pH stability of samples incubated for 7 days indicated that crude anthocyanins degraded slower at acidic pH, which is in keeping with reported literature therefore this extract, should be added to foods with lower pH. Light stability showed slower degradation in dark incubated samples resulting in 84% pigment retention after a 10 day period. Synthetic colourants proved to be superior, as they had showed better stability than the natural colourant under the same conditions. Half-life of thermally treated samples showed a decrease upon heating, colour was also affected as samples became dull and murky. DPPH of thermal treated samples showed a decline in radical scavenging activity from 70 to 85˚C and thereafter an increase was observed between 85 and 90˚C, this could be due to the release of degradation products that have antioxidant capability. Solvent systems did not have an effect on the total phenolic content of crude extracts as no significant difference was observed by each of the H. sabdariffa crude extracts contained an average of 54.67 mg/ml GAE. The radical scavenging ability assessed by the DPPH and FRAP assays showed 53.75% and 57.51% radical scavenging ability respectively. Although the synthetic colourants showed better stability, a natural food colourant from H. sabdariffa can still be beneficial as it has potential to be applied into foods that contain low pH such as jelly and yoghurt. The additional benefits that natural food colourants possess aid in the marketability of the product.
M

碩士
中山醫學大學
營養學研究所
101
Melanogenesis has many important physiological functions, including photo-protection of human skin from UV irradiation. Melanin synthesis is stimulated by α-melanocyte stimulating hormone (α-MSH) and cyclic AMP (cAMP) elevating agents. However, abnormal melanogenesis causes hyperpigmentation in the skin, which results in serious aesthetic issues and increases the risk of skin cancer. Previous studies have indicated Hibiscus sabdariffa leaf extract (HLE) has antioxidant and anticancer activities. Therefore, in this study we examined the effect of HLE on melanogenesis and signaling pathways in vitro and in vivo. Results showed that non-cytotoxic doses of HLE and its main composition ellagic acid (EA) reduced α-MSH-induced both tyrosinase activity and melanin production in mouse melanoma B16F0 cells. Western blotting data showed HLE and EA inhibited the expression of tyrosinase and TRP-1 (tyrosinase-related protein 1), cooperative with intracellular cAMP content. Further, HLE also suppressed the nuclear levels of CREB causing the consequent disturbed activation of MITF in α-MSH-stimulated B16F0 cells. The HLE-inhibited α-MSH-induced tyrosinase expression appeared be a consequence of MITF inactivation, because its DNA binding activity was suppressed by HLE. Besides, HLE inhibited melanin production in the dorsal skin of hairless mice exposed to UVB and the molecules involved in the melanogenesis. These findings suggested that HLE might be used as a potential natural whitening agent.

碩士
中山醫學大學
生化暨生物科技研究所
94
In Taiwan, Hibiscus Sabdariffa Linnaeus (HS), a well-known beverage, was demonstrated to represent antioxidant effect in previous studies. To further exploit the extracts, polyphenols and anthrocynins from HS (HPE and HA), we investigated the anti-inflammatory of effect of target sample in RAW264.7 cell on inflammatory mediators secretions (Nitrite and PGE2). Sequentially, animal model study was also preformed to confirm the anti-inflammatory effect of HPE and HA on lipopolysaccharide (LPS)-induced acute hepatic inflammation. The results showed that HPE and HA (0.01-0.5mg/ml) reduced the xanthine oxidase activity to 94.6% and 10.5% in the highest concentration in vitro, and also decreased the nitrite and PGE2 secretion in LPS-activated cells. The in vivo assay showed that pretreatment of HPE (10-40 mg/kg) or HA (20-60 mg/kg) for 5 days significantly decreased the levels of AST and ALT induced by LPS. Therefore also evaluate the antioxidant enzyme or antioxidant molecule, such as catalase and glutathione (GSH) and decrease the level of lipid peroxidation in liver. In histopathological evaluation of rat livers, it revealed that HPE and HA reduced the incidence of liver lesion such as inflammatory cell and infiltration induced by LPS. Furthermore, we also found that pretreated with HPE and HA decreased the hepatic protein expressions of COX-2, p-JNK and p-ERK induced by LPS in rats. These results showed that HPE and HA represented anti-inflammatory potential in vitro and in vivo; and they could be developed to nature, non-toxic anti-inflammatory materials.

碩士
國立屏東科技大學
生物科技系所
105
In this study, we evaluated the short-term cytotoxicity, intracellular reactive oxygen scavenging capacity, wound repairing, tyrosinase-induced melanogenesis and scavenging free radical scavenging ability of the human skin fibroblasts CCD-966SK which treated with Antrodia cinnamomea sample A (ACA), Antrodia cinnamomea sample B (ACB) and extraction of Hibiscus sabdariffa (EHS). The results of DPPH free radical scavenging assay, The clearance rate of EHS (1000 g/ml), ACB (5000 g/ml) and ACA (10 g/ml) were 80%, 75% and 23% respectively. For MTT assay, the survival rate of CCD-966SK cell were 90% when treated with ACA (100 g/ml), ACB (100 g/ml) and EHS (1000 g/ml) for 72 hours. For wound healing assay of each treatments, treated with ACA (100 g/ml) haven’t shown cell growth and migration at 12-48 hours, but treated with ACB (100 g/ml) and EHS (100 g/ml and 500 g/ml) shown wound healing. Moreover, the wound healing rate were reached 100% at 36 hours which negitive control only reached 40%. For tyrosinase inhibition assay, the melanin inhibition rate of ACA (50 g/ml), ACB (500 g/ml) and EHS (2000 g/ml) were 56%, 50% and 64%, respectively. The EHS shown most effect melanin inhibition rate compare to other groups. For Cell oxidative stress assay, the ROS inhibition rate of ACA (1 g/ml), ACB (500 g/ml) and EHS (2000 g/ml) were 36%, 49% and 62%, respectively.

碩士
中華醫事科技大學
醫學檢驗生物技術系碩士班
100
Phenolic compounds are the popular studies in the present day , cause of antioxidants. In this day, many people are using phenolic compounds to prevent disease from Plant extract. More and more people eat plant extract products , because it is not only can antiseptic or anti-virus but also inhibit LDL oxidation and prevention cardiovascular disease. High performance liquid chromatography with UV separation of these phenolic compounds are possible on C-18 column, using 0.1% formic acid and acetonitrile with 0.1% formic acid as mobile phase. Flow-rate, detection wavelength and temperature were adjusted to 1.0 mL/min, 285 & 345nm and RT, respectively. In this study, Roselle’s leaf, extract by water, detection Catechin 6.41 ppm and EA 3.37 ppm , extract by methanol detection Protocatechuic acid 0.67 ppm、Catechin 53.4 ppm、Caffeic acid 14.6 ppm、Epigallocatechin 11.1 ppm、Ferulic acid 15.4 ppm、Quercetin 1.94 ppm with 285 nm and EA 113.8 ppm with 345 nm. Nelumbinis Receptaculum,are extracted by water, detection Catechin 5.93 ppm、Procyanidin B2 1.92 ppm、ρ- Coumaric acid 0.45 ppm、Epigallocatechin 74.1 ppm and extract by methanol detection Catechin 329 ppm、Procyanidin B2 7.16 ppm、Caffeic acid 3.6 ppm、ρ- Coumaric acid 7.63 ppm、Epigallocatechin 14.8 ppm with 285 nm. Graptopelaum Paraguayense, are extracted by water, detection ρ- Coumaric acid 1.34ppm with 285 nm, extract by methanol detection Gallic acid 5.92 ppm、Caffeic acid 0.51 ppm、ρ- Coumaric acid 1.17 ppm、Epigallocatechin 25.1 ppm、Ferulic acid 0.55 ppm、Gossypetin 1.93 ppm with 285 nm. In the future, the unknown peaks would be detected by gas chromatography – mass spectrometry(GC-MS).

碩士
國立臺灣大學
化學工程學研究所
93
This work focuses on the separation, purification and identification of anthocyanins, mulberry (Morus spp.) and roselle (Hibiscus Sabdariffa L.) grown in Taiwan. Their antioxidative activities were also determined and compared. Extracts from the fruit samples were fractionated by flash chromatography using 25 and 50% methanol (with 1% acetic acid each). Qualitatative and quantitative analysis of fractionated products were carried out by using HPLC and identified the structures by employing HPLC-PDA, LC-Mass as well as NMR. The results showed that the mulberry contained cyanidin 3-glucoside (237.0±14.26mg/100g FW), cyanidin 3-rutinoside (158.8±8.80mg/100g FW) and a trace amount of plargonidin 3-rutinoside. 8.7±0.05mg/g DW of anthocyanins were isolated form the dried roselle. The method of Ferric Reducing Ability of Plasma (FRAP) and the inhibition of H2O2-induced lipid peroxidation in the rat brain homogenates were employed to assess their antioxidant activities. The ID 50 values of F1-25 and F1-50 were 2.0 and 1.7 μg, respectively. Comparing to the Trolox (ID 50 4.3μg), the inhibition capability of lipid peroxidation of mulberry was remarkable

博士
中山醫學大學
生化暨生物科技研究所
94
Hibiscus sabdariffa L., a local soft drink material and medicinal herb, was usually used effectively in native medicines against hypertension, pyrexia and liver disorders. Here in our research found that an extract of Hibiscus sabdariffa contains lot of polyphenolic acids, flavonoids and anthocyanins. In our report, HSE was able to prevent LDL oxidation as evidenced by its inhibition on electrophoresis, ApoB fragmentation, cholesterol degradation, and TBARS of LDL. Taken together, HSE showed a strong potency to inhibit the production of oxidized LDL induced by copper. Furthermore, we examined the effect of HSE on the serum lipids in animal. Firest, we used high fructose (HFD)- or high cholesterol (HCD)- fed rats. HSE specifically reduce the serum triglyceride in the HFD-fed rats and the serum cholesterol in the HCD-fed animals. The level of LDL and the ratio of LDL-cholesterol (LDL-C) to HDL-cholesterol (HDL-C) were declined by HSE in both models. Then we also designed another animal model to test whether HSE exhibit hypolipidemia and antiatherosclerotic effect in rabbits with experimental atherosclerosis. New ZeaLane White (NZW) rabbits were fed with a normal diet, high cholesterol (1.3%), lard oil (3%) diet (HCD) with or without 0.5 or 1 % HSE for 10 weeks. The level of triglyceride, cholesterol and low-density lipoprotein - cholesterol (LDL-C) were lower in the serum of rabbits fed HCD plus HSE than in the serum of rabbits fed HCD. Feeding HSE (0.5 and 1% in the diet) to rabbits significantly reduced severe atherosclerosis in the aorta. Histopathological examination showed that HSE reduced foam cell formation, inhibited smooth muscle cell migration and calcification in the blood vessel of rabbits. In addition, we used hamster hyperlipidemia model found that, HSE not only reduced serum cholesterol, LDL-C and triglyceride. It can reduced hepatic cholesterol and triglyceride concentration and increase lipase activity in hamster serum and liver tissure. In Hep G2 cell, we found that, HSE can increase antioxidative enzyme expression such as catalase and SOD, suppress FAS espression via PI-3 kinase signaling pathwy, and activated Ras/ERK kinase increase PPAR expression. In mRNA expression, we found that HSE can increase LDL receptor expression and suppress HMG-Co A reductase and SREBP-1c expression. These results suggest that HSE inhibits serum lipids increase via inhibited FAS expression and increase PPAR avtivation and shows an antiatherosclerotic activity.

Submitted in fulfillment of the requirements for the award of the degree of Master of Health Sciences: Dental Technology, Durban University of Technology, Durban, South Africa, 2017.
The poor mechanical properties of denture base acrylic resins can cause dentures to fracture either through impact or stress fatigue. Various metal reinforcing materials such as powders, nets, plates and wires together with synthetic fibres are therefore used to improve the strength properties of denture base acrylic resins. Despite their benefits, they are difficult to apply during fabrication, are costly, non-recyclable, and not environmentally friendly. The suitability of Hibiscus sabdariffa (H. sabdariffa) as a fibre reinforcement material for denture base acrylic resins was therefore investigated. A quantitative research paradigm and a two-phased experimental research design was used. The preliminary study used Fourier transform infrared (FT-IR) spectroscopy, thermogravimetric analysis (TGA) and field emission scanning electron microscope (FE-SEM) to assess the characteristics of, and the effect of, mercerisation on H. sabdariffa fibres. In determining the suitability of H. sabdariffa fibres, fibre diameter, fibre density, moisture content and absorption were examined. The main study investigated the mechanical and physical properties of two sample groups namely: the unreinforced (control) group and the H. sabdariffa fibre-reinforced (test) group denture base acrylic resins. The flexural strength, impact strength, specific wear rate, hardness and water absorption behaviour were determined in line with ASTM D 790 and D 256, G 99, D 2583 and D 570 methods of testing, respectively. Parametric and non-parametric tests were used to analyse the mean differences (p = 0.05) between the unreinforced and the H. sabdariffa fibre-reinforced denture base acrylic resins. The preliminary study revealed that H. sabdariffa fibres favourably compared to other lignocellulosic fibres in terms of fibre diameter, fibre density, water content and absorption behaviour. In the main study, a t-test revealed that there were significant differences in flexural strength at 0oC (p < 0.0001) and at 65oC (p < 0.0025) between the two sample groups. Similarly, there were significant differences in the impact strength between the two sample groups at 0oC (p < 0.031) and at 65oC (p < 0.0001). In contrast to the 200g load, there were significant differences (p < 0.044) in the specific wear rate of the unreinforced and the H. sabdariffa fibre-reinforced denture base acrylic resins at the 500g load. The hardness values revealed no significant differences (p > 0.707) between the two sample groups. There were however, significant differences (p < 0.041) in the water absorption rate between the two sample groups. Overall, the prominent features of this study showed that H. sabdariffa fibre is a suitable reinforcement fibre material for denture base acrylic resins and is in compliance with ISO 1567: 1999. Although this study found that H. sabdariffa fibres can be used as reinforcement material for denture base acrylic resins, the results cannot be generalised to clinical conditions, therefore in vitro and in vivo investigations are recommended.
M

Hibiscus sabdariffa (HS) is a plant of the Malvaceae family that has anti-obesity, antihypertensive, hypocholesterolaemic, anti-oxidant and anti-cancer properties. Dietary manipulations during the suckling period cause precocious maturation of the gastrointestinal tract (GIT). Events in early life affect the individual‘s metabolic health in later life. This study investigated the effects of early administration of the aqueous calyx extracts of HS on the growth performance, general health and the GIT of neonatal rats and whether it conferred protection or predisposition to the development of metabolic dysfunction in adolescence. The study was carried out in two phases. The first phase was to determine the effects of HS aqueous calyx extracts on the growth performance, metabolic substrates and the development of the neonatal rat GIT. In the second phase, the effects of the early administration of the HSE on the response of the pups to diet induced metabolic dysfunction were investigated. In phase one, forty two 4-day old Sprague Dawley pups of both sexes were randomly assigned to three treatment groups. Each group consisted of 7 males and 7 females. The control group received distilled water at 10ml.kg-1 b.w while the other two groups received either a low (50mg.kg-1) or high dose (500mg.kg-1) of the HS aqueous calyx extracts via oral gavage daily for 9 consecutive days. The rats were euthanased and their tissues harvested and analyzed. Pups that were administered with the high dose HSE had significantly heavier small intestines relative to the body mass when compared to those on the low dose HSE (P<0.01) and the control group (p<0.001). Pups in the high dose HSE group had significantly heavier caeca (p<0.05) than those in the low dose HSE group. In the second phase, eighty five 4-day old Sprague Dawley rat pups were used. They were initially divided randomly into three groups and received similar treatments as in phase one up to postnatal day (PND) 14. There was no intervention from PND14 to PND 21 when the pups were weaned. The rats in each of the treatment groups were further divided into a control group that continued on their normal rat chow diet and a test group that received high fructose (20% w/v) in their drinking water for 30 days in order to induce metabolic dysfunction. Each of the six study groups had at least 5 male and 5 female rats. The male rats in each of the treatment groups gained more body mass than their corresponding female counterparts in the control and treatment groups (p<0.001). Female rats that received high dose HSE in the neonatal period had significantly greater visceral fat pad (p≤0.05) than the males in the groups. There were no negative effects on the rats‘ general health. At the end of the study, features of metabolic syndrome did not manifest in the control or any of the treatment groups. Hibiscus sabdariffa aqueous calyx extracts did not exhibit any long term effects and therefore may be considered safe for consumption in the neonatal age group.

碩士
南華大學
自然醫學研究所
93
Due to the aging of population more and more serious in recent years, people pay attention to their own health earlier and earlier. The issues such as how to avoid growing old and how to strengthen the anti-oxidant are therefore in prevailing. Malignant tumour, cerebrovascular disease, cardiovascular disease, and diabetes are among ten major death causes in Taiwan. Current researches verified that these diseases not only related to aging structure, but also showed that they may all had something to do with the injury of oxidizing the human body caused by the free radical or the active oxygen. 　 　　The purpose of this research is therefore to probe into anti-oxidant composition and activation included in the following four kinds of folk prescription, namely: Chrysanthemum morifolium, Hibiscus sabdariffa, Lycium chinese, and Ziziphus jujube. In the meantime, they were also reconfigured as the drink of formula (compound prescription) according to the statistical methodology in order to investigate their optimal result of anti-oxidizing effects. 　 　　In this regard, this research examined separately four kinds of folk prescriptions and formula mentioned above; and analyzed their scavenging capacity of DPPH, total antioxidant activity, reduction ability, flavonoid contents, and phenol contents. In addition, the relation among flavonoids, phenol, and anti-oxidant ability with SPSS were also analyzed. Analysis results showed that: 1. In the order of flavonoid contents, they were Chrysanthemum morifolium (12.38), Hibiscus sabdariffa (11.60), formula (5.41), Ziziphus jujube (1.59), and Lycium chinese (0.77 mg quechitin/g dw); 2. In the order of phenol contents, they were Chrysanthemum morifolium (25.83), Hibiscus sabdariffa (23.05), formula (11.54), Lycium chinese (5.22), and Ziziphus jujube (2.83 mg gallic acid /g dw). 　 　　Their flavonoid contents and phenol contents had linear relations with the anti-oxidant activation in separate. Moreover, the scavenging capacity of DPPH, total antioxidant activity and reduction ability of all samples were raised with the increase of their concentrations. 　 　　Chrysanthemum morifolium, hibisus, and formula had better anti-oxidant efficiency: 1. In the order of their effective concentrations (EC50) in scavenging capacity of DPPH, they were Chrysanthemum morifolium (0.09), Hibiscus sabdariffa (0.90), and formula (0.94 g/L). 2. In the order of the effective concentrations in reduction ability (OD1.0), they were Chrysanthemum morifolium (0.81), formula (1.18), Hibiscus sabdariffa (1.21), and Lycium chinese (1.25g/L). 3. In the order of effective concentrations (EC50) in total anti-oxidant ability, they were Chrysanthemum morifolium (0.75), Hibiscus sabdariffa (0.93), and formula (1.37 g/L). Based on the above research results, the Chrysanthemum morifolium, hibiscus, and formula had better anti-oxidant efficiency because they had more flavonoids and total phenol compositions. Although Ziziphus jujube didn’t have too much anti-oxidant efficiency, it could increase the flavor in the drink of formula; nevertheless, it has the functions of mixing with hundreds of other Chinese herbal medicines. The Hibiscus sabdariffa had very good color and luster, which not only can beautify the appearance of the formula drink, but also sometimes produce amazing effects and make it additional curative. If the Hibiscus sabdariffa could be further processed, it would be helpful for developing the mechanism of natural food. Furthermore, it would elevate the economic benefits and application value of the Chinese herbal medicine. The Hibiscus sabdariffa will notably play a major role in developing its anti-oxidant efficiency to prevent people from growing old and to promote human health.

碩士
靜宜大學
食品營養研究所
98
Although the antioxidant and antihypertensive effects of Roselle (Hibiscus Sabdariffa L.) was confirmed, its anti-aging mechanism is still indeterminate. The purpose of this study was to investigate the effects of roselle extracts on learning and memory ability, and antioxidant status in senescence-accelerated mice (SAMP8). Three and six-month-old SAMP8 male and female mice were used in this study. SAMP8 mice were randomly divided into five groups, where each group contained 10 mice, including control group, 3.30 g/kg BW, 6.59 g/kg BW, 13.19 g/kg BW and 32.97 g/kg BW freeze-dried roselle extract powder group. This study period lasted for 12 weeks. Before the experiment , open field activity test was recorded in evaluation for behavior. Besides food intake, the body weight was also measured. One week before scarification, the aging scores were measured, and single-trail passive avoidance test and active shuttle avoidance test were performed independently. The mice were sacrificed to analyze the biochemical parameters of sera. The antioxidant ability and the lipid peroxidation of liver. The results showed that there was no significant difference in the body weight, food intake, and locomotion among each group. The aging score of control groups was significantly higher than experimental groups in 3 and 6-month old mice (p&lt;0.05). Results from ingle-trail passive avoidance test and active shuttle avoidance test showed that experimental groups had significantly better learning memory ability (p&lt;0.05). Activities of antioxidant ability, including SOD, GPx, catalase, and total thiol , in the experimental groups were significantly higher than that of control groups (p&lt;0.05). The MDA and protein carbonyl of liver in the experimental groups were significantly lower than the control groups (p&lt;0.05). Besides, the β-amyloid protein deposition of brain in male and female mice were significantly lower in the experimental groups than control groups (p&lt;0.05) In summary, the supplement of roselle extract may promote the antioxidant defense system, reduce oxidative stress-induced damage,reduce the accumulation of β-amyloid deposition, and improve learning and memory ability. The underlying anti-aging mechanism needs further investigation.

碩士
中州科技大學
保健食品系
103
The roselle (Hibiscus sabdariffa L.) is a annual shrub, related report showed that it has antioxidant, anti-glycation, anti-hyper- tension and prevent cardiovascular diseases. This study of raw material is Taiwan’s roselle, which is extracted useing different solvent as (30℃, 80℃, 90℃) water and (50%, 70%) ethanol, and is to evaluate and analyze its functional components content (polyphenols, total flavonoids, total anthocyanine, polysaccharides, organic acid) and (antioxidant, anti-glycation and inhibit bacterial growth capacity). Finally, we select the best of 50% ethanol of crude extracts and is separated and purified by the XAD-7 column chromatography. Respectively, obtained the (0.2%, 20%) Delphini- din-3-O-sambubioside of fractionates, and to assess the differences of antioxidant capacity, anti-glycation, inhibit bacterial growth capacity of unhydrolyzed and hydrolyzed fractionates. The study showed (A) the antioxidant capacity : (1) Trolox equivalent antioxidant capacity : the hydrolyzed fractionates (0.2%, 20%-Delphinidin) ( 359.7, 400.7uM) are the best than unhydrolyzed fractionates (313.3, 400.7 uM)(2)DPPH free radical scavenging capacity: unhydrolyzed fractionate (0.2%-Delphinidin-3-o-sambubioside) (EC50：64.7 ug/g) is the best than hydrolyzed fractionate (EC50：94.1g/g), but the hydrolyzed fractionate (20%-Delphinidin) (EC50：138.4ug/g) is the best than the unhydrolyzed fractionate (EC50：242.1 ug/g). In addition, (B). anti-glycation capacity: unhydrolyzed fractionates (0.2%, 20%-Delphinidin-3-O-sambubioside) are the best (9.1-25.3%, 10.2-53.0%) than the hydrolyzed fractionates (0.0%, 0.0%). (C). Inhibition of bacterial growth capacity (Escherichia coli, Staphylococcus aureus) : In 0.5mg/ml of sample, the hydrolyzed fractionate (20%-Delphinidin) (100%、100%) is the best than the unhydrolyzed fractionate 20%-Delphinidin-3-O-sambubioside (87.7、 50.1%). the hydrolyzed fractionate (0.2%-Delphinidin) (98.4%、42.7%) is the best than the unhydrolyzed fractionate (16.0%、39.4%).The results studied that the hydrolyzed fractionate (20%- Delphinidin) is the best in antioxidant and inhibit bacterial growth capacity.But unhydrolyzed fractionate (20%-Delphinidin-3-O-sam- bubioside) is the best in anti-glycation capacity, it was showed the unhydrolyzed (20%-Delphinidin-3-O-sambubioside) and hydrolyzed (20%-Delphinidin) fractionates are the better in antioxidant, anti-glycation and inhibit bacterial growth capacity. Finally, it is proven to the higher content of anthocyanin of ethanol extract that can be increased the Roselle’s functional, and have a potential as the development of health food products.

碩士
中山醫學大學
生化暨生物科技研究所
93
In developed countries, atherosclerosis and its complications are becoming a major cause of death. Several studies showed that free radical-mediated oxidation of low density lipoprotein leading to the production of oxidized LDL (OxLDL) is a key event in initiating endothelial cells proliferation and atherosclerosis plaque. Some natural antioxidants such as vitamin C, vitamin E, β-carotene have been revealed clearly to prevent the atherosclersis. As shown in previous studies, we found that the aqueous extract of Hibiscus Sabdariffa ( HSE ) and protocatechuic acid ( PCA ) could be able to inhibit the apoptosis of macrophage induced by OxLDL. In this study, we used the human monocyte, THP-1, to further detect the effects induced by OxLDL, and what kind of role could HSE and PCA play. Using TBAR, migration assay, flow cytometric DNA fluorescence and detecting apoptosis-related proteins, our results showed that increasing the dosage of HSE or PCA could inhibit monocyte apoptosis caused by OxLDL. At the high concentration of HSE and PCA, the concentrations of MDA produced from lipid peroxidation were inhibited up to 91% and 85%; the inhibitory percentage on migration were 70% and 76%; and the level of apoptosis was reduced to 81% and 93%. Taken the above, we concluded that the inhibitory mechanisms of HSE or PCA on atherosclerosis could also be contributed to the other pathway by inhibiting monocyte apoptosis and differentiation.

碩士
中山醫學大學
醫學檢驗暨生物技術學系碩士班
101
Many studies have shown that polyphenols prevent and inhibit the growth and metastasis of cancer cells. In previous study, Hibiscus sabdariffa leaf was demonstrated be rich in flavonoid, and possess anti-cancer potential. Recently, the data of our laboratory presented the evidence of H. sabdariffa leaf extract (HLE) as an apoptosis inducer in human prostate cancer LNCaP cells. Therefore, the object of the study was to examine the anticancer properties and molecular mechanisma of H. sabdariffa leaf polyphenolic extract (HLP) on LNCaP cells. Trypan blue assay and BrdU assay showed HLP, its main composition epicatechin gallate (ECG) and β-sitosterol, a plant sterol, inhibit the LNCaP cell growth. Our results revealed that the cells presented DAPI-positive morphology, and had an increase in the distribution of hypodiploid phase after a 48-h treatment with HLP. This effect of HLP in LNCaP cells might be mediated via the death receptor (FasL-mediated caspase-8) and/or partially mitochondria (Bax-mediated caspase-9) pathways. In addition, HLP and β-sitosterol could induce mainly cellular autophagy via PI3K class III/Beclin-1/LC3 signaling. Molecular data showed these influences of HLP might be mediated via apoptosis and autophagy pathways, as demonstrated by the treatment of FasL antagonist (NOK-1) or autophagic inhibitor (3-MA), respectively. Based on the above, HLP induced LNCaP cell apoptosis and autophagy. These results suggested that HLP potentially could be developed as an anti-cancer agent, and may open interesting perspectives to the strategy in human prostate cancer treatment.

El cáliz de la jamaica contiene cantidades significativas de compuestos antioxidantes cuya ingesta habitual, a través de una infusión, previene y controla enfermedades crónico-degenerativas. Sin embargo, se ha documentado su baja biodisponibilidad en el cuerpo humano, debido a la inestabilidad de sus compuestos fenólicos durante su trayecto a través del tracto digestivo. En el presente trabajo se propuso preservar la actividad antioxidante de los compuestos fenólicos de este producto vegetal para lograr una mayor bioaccesibilidad empleando polímeros comestibles con dos tecnologías de encapsulación, a) secado por aspersión y b) coacervación. En el primer caso se mezcló el extracto de la jamaica con los polímeros: maltodextrina, pectina, gelatina, carboximetilcelulosa, proteína de suero, carragenina y goma arábiga obteniendo polvos para su análisis. Los coacervados se elaboraron con grenetina y se encapsularon con una y dos coberturas de alginato de sodio gelificado por inmersión en una solución de CaCl2 durante 10, 20, 30 y 40 minutos. Se evaluó la cinética de liberación de los productos obtenidos y se ajustó, en todos los casos, a una ecuación de Weibull-Prmer Orden. La constante de velocidad de liberación (ks) de los coacervados disminuyó linealmente con el número de coberturas de alginato y con el aumento en el tiempo de inmersión en la solución de CaCl2. Los resultados obtenidos muestran que la liberación de los compuestos fenólicos de la jamaica puede controlarse en función del número de coberturas de alginato y del incremento en el tiempo de inmersión en la solución de CaCl2. Los polímeros utilizados en el proceso de secado por aspersión mostraron diferentes parámetros cinéticos. Derivado de la cinética de liberación, se propuso un método para estimar el tiempo de disolución de las partículas obtenidas.

碩士
建國科技大學
機械工程系暨製造科技研究所
99
Supercritical carbon dioxide (SC-CO2) can provide a benefit of lower temperature and high diffusion ability in nature product extraction process, not only preserved the thermo unstable but also keep the bioactivity of the extracts. Owing to this, we use supercritical carbon dioxide extract the caffeic acid (CA), a kind of polyphenol, from Roselle. Caffeic acid has abilities with antioxidant, radical catching and tyrosinase inhibition. We also use high performance liquid chromatography analysis the components differences between water extraction, alcohol extraction and supercritical carbon dioxide extraction. The result show that caffeic acid was complete extracted by water and alcohol extraction, but lower in supercritical carbon dioxide extraction. The components of supercritical carbon dioxide extraction are much simple. Following, we add water and alcohol as the co-solvent to modify the polarity of the supercritical carbon dioxide, and use this modified supercritical carbon dioxide in the caffeic acid extraction with differ critical temperatures, critical pressures and flow rates. In the results of the further analysis by high performance liquid chromatography, the best optimization in modified supercritical carbon dioxide extraction of caffeic acid needs critical pressure at 250 bar, critical temperature at 40℃, Co-solvent 85% ethanol as co-solvent at the flow rate 1.2ml/min. In conclusion, add water and alcohol as co-solvent into supercritical carbon dioxide can optimize the caffeic acid extraction efficiency from Roselle. Actually, add only water into supercritical carbon dioxide will make supercritical unstable and become sub-supercritical. Adding alcohol can provide a better stability when water is a necessary co-solvent to the supercritical carbon dioxide extraction. Both adding water and alcohol as co-solvents are the key point of increasing the efficiency of caffeic acid supercritical carbon dioxide extraction from Roselle.

碩士
國立屏東科技大學
農園生產系所
105
The Roselle(Hibiscus sabdariffa L.) calyx is one of the most important medicinal plants in many parts of the world for its edible, medicinal uses as health foods. Roselle calyx can produce considerable amounts of antioxidant such as, polyphenols, flavonoids and anthocyanins. Arbuscular mycorrhizal fungi (AMF) are a beneficial microorganism in the terrestrial ecosystem , It can enhance the absorption of nutrients and improve the secondary metabolite accumulation in crops. In this study, these are six species of AMF, such as Glomus clarum, G. etunicatum, G. fasciculatum, Gigaspora sp., G. mosseae, and Acaulospora sp. are used to screened out which one is the optimal mycorrhizal fungi and also to evaluate the effects of secondary metabolites and antioxidant capacity of Roselle calyx. The results showed that all six AMF species are able to form mycorrhizal structure with the host Roselle, the mycorrhizal formation rates from 71.67% to 78.89%, indicating that these AMF present high compatibility with the host roselle. After cultivation 210 days, the plant height inoculated with AMF was from 154.98 cm to169.98 cm, which was enhanced by inoculating with Gm, Asp, Gg, Ge, Gc, and the stem diameter of plant inoculated with AMF from 12.23 mm to 14.74 mm, which was enhanced through inoculating with Gg, Asp, Gm, Gf, Ge. In pot trials for screening AMF species, Asp, Gg species were screened out and mixed with 1: 1 for next experiments. In following study, Roselle was inoculated with mix AMF(Gigasporasp.,Acaulosporasp.), after cultivation 210 days, the mycorrhizal formation rates were AMF 93.33%, control 0.00%; the plant heights were AMF 153.88 cm, control 134.57 cm and the stem diameters were AMF 22.13 mm, control 15.17 mm, they indicated that the treatment of mycorrhizal fungi increased significantly. And the total chlorophyll contents were AMF 2.26 mg/g, control 1.71 mg/g; the soluble carbohydrate contents were AMF 5.06 mg/g, control 3.04 mg/g; soluble protein contents were AMF 17.59 mg/g, control 10.46 mg/g. It illustrated that the Roselle leaves with AMF inoculation had substantially higher chlorophyll content as well as soluble carbohydrate and soluble protein compared with those in control group. The Roselle calyxes of polyphenols contents were AMF 27.37 mg/g, control 20.53 mg/g; the total flavonoids contents were AMF 8.03 mg/g, control 7.40 mg/g; total anthocyanin contents were AMF 11.58 μmole/g, control 9.77 μmole/g, all significantly enhanced by inoculating with AMF. The phenylalanine ammonia lyase (PAL) activities of inoculated and control groups were 0.96 OD290˙30min-1g-1 FW and 0.68 OD290˙30min-1g-1 FW. In the antioxidant capacity of the methanol extraction, the DPPH radical scavenging ability of Roselle residue obtained the highest ability 90.95% at a concentration of 1 mg/ml of inoculation AMF treatment, and control were 89.90%, enhanced significantly by inoculating with AMF. In the antioxidant capacity of the methanol extract, the DPPH radical scavenging ability of Roselle residue obtained the highest ability 0.418 at a concentration of 1 mg/ml of inoculation AMF treatment, and control were 0.316, enhanced significantly by inoculating with AMF. The Roselle calyxes with AMF inoculation also had substantially higher secondary metabolite contents such as polyphenols, lavonoids, and anthocyanin, and substantially higher PAL activities and antioxidant ability. Therefore, AMF can be used to develop for Roselle cultivation and production. Keyword : antioxidant capacity, arbuscular mycorrhizal fungi (AMF), phenylalanine ammonia lyase(PAL), Roselle(Hibiscus sabdariffa L.)

博士
中山醫學大學
生化暨生物科技研究所
103
Hibiscus sabdariffa leaf has been previously shown to possess hypoglycemic, hypolipidemic, and antioxidant effects, and induce tumor cell apoptosis. However, the molecular mechanisms involved in the anticancer activity of Hibiscus sabdariffa leaf polyphenolic extract (HLP) and H. sabdariffa leaf extract (HLE) are poorly understood. The object of the study was to examine the anti-cancer potential of HLP and HLE. First, HLP was exhibited to be rich in epicatechin gallate (ECG) and other polyphenols. Apoptotic and autophagic activities of HLP and ECG were further evaluated by DAPI stain, cell cycle analysis, and acidic vascular organelle (AVO) stain. Our results revealed that both of HLP and ECG induced the caspases cleavages, Bcl-2 family proteins regulation, and Fas/FasL activation in human melanoma A375 cells. In addition, HLP could increase the expressions of autophagy-related proteins autophagy-related gene 5 (ATG5), Beclin1 and light chain 3-II (LC3-II), and induce autophagic cell death in A375 cells. Second, we want to examine the anti-invasive potential of HLE. The results of wound-healing assay and in vitro transwell assay revealed that HLE dose-dependently inhibited the migration and invasion of human prostate cancer LNCaP cells under non-cytotoxic concentrations. Molecular data showed all these influences of HLE might be mediated via inhibition of protein kinase B (PKB, also known as Akt)/NF-kB/MMP-9 cascade pathway. Finally, the inhibitory effect of HLE was proven by its inhibition on the growth of LNCaP cells and the expressions of metastasis-related molecular proteins in vivo. These data indicated that the anti-cancer effect of HLP and HLE. HLP and HLE potentially could be developed as an anti-cancer agent.

博士
中山醫學大學
生物化學研究所
91
The natural antioxidants, such as Vit E, Vit C and beta-carotene, has been found to possess the ability to avoid LDL oxidation or reduce oxidized LDL (oxLDL) uptaken by macrophage; and furthermore, they were able to prevent the formation of atherosclerosis. In our previous studies, we explored that both protocatechuic acid (PCA) and the aqueous extract of its original plant, Hibiscus Sabdariffa (HSE), showed strong inhibitory effects on the negative charge increasing of protein moiety in oxLDL (lipogel electrophoresis) and on the lipid peroxidation of lipid moiety in oxLDL (TBARS). At present, we further investigate the effects of PCA and HSE on LDL oxidation。We incubated 10 mM CuSO4 and 100 mg protein/ml of LDL at 37℃ for 12 hrs to induce the degradation of cholesterol. When cotreating with various concentrations of PCA or HSE, the results showed that 0.1 mg/ml of HSE recover the degradation of cholesterol by 18.15% and 0.5 mg/ml of HSE by 67.25% compared to native LDL. On the other side, the more concentrated PCA showed more significant recovery of cholesterol degradation. At the concentration of 0.05 mg/ml, the degradation of cholesterol was recovered by 74.96 %. On the investigation of ApoB fragmentation, HSE recovered the fragmentation gradually depending on the increasing concentrations. Moreover than, the ApoB fragmentation recovered almost by 100% at the concentration of 0.5 mg/ml. When cotreating with PCA, the similar situation also occurred. At the concentration of 0.03 and 0.05 mg/ml, ApoB fragmentation was recovered by 100 %. Taken above together, we suggested that PCA or HSE should play an important role on preventing atherosclerosis via inhibiting LDL oxidation. In part I, we found that protocatechuic acid (PCA) and the aqueous extract of Hibiscus Sabdariffa (HSE) possessed the abilities to inhibit LDL oxidation. Because of that, in this study, OxLDL was added to RAW264.7 macrophages to further investigate the effects of PCA or HSE. When treating with 100 mg/ml OxLDL, the cell was induced severe death. Under the demonstrations of Leukostat stain and DNA fragmentation (TUNEL assay), we proved that the cell death was apoptosis. When co-treating with PCA or HSE, the apoptosis was able to reduce in a dose-dependent. At the concentration of 1.0 mM PCA or 0.5 mg/ml, the cell morphology showed almost same as the control. We further detect PARP protein cleavage and Caspase-3 activity to confirm the inhibitory effects of PCA or HSE on macrophage apoptosis. The levels of PARP protein cleavage was able to decrease to 100% by 1.0 mM PCA and 93% by 0.5 mg/ml HSE; and Caspase-3 activity was also inhibited by 74.2% or 72.3% when co-treating with 1.0 mM PCA or 0.5 mg/ml HSE. In migration assay, we found that the inhibitory effect of PCA or HSE attributed to reduce the uptake of OxLDL by macrophage, and some interactions between PCA/HSE and macrophage. Some researchers pointed that the OxLDL-laden macrophages possessed the ability to digest OxLDL. However, when macrophage uptaking too much OxLDL, they would die, and they would not digest OxLDL. Furthermore, the abundant death macrophages could accelerate the formation of fatty streak to process to atheroma. According to these results, we suggested that PCA or HSE was able to protect macrophage from dead when uptaking OxLDL; and that could make macrophage to digest OxLDL and further decelerate atherosclerosis. Under the circumstance, PCA and HSE were able to prevent atherosclerosis. According to the above, we observed the inhibitory effects of HSE and PCA on OxLDL-induced macrophage apoptosis. Based on that, we further investigated the mechanisms here. Firstly, we surveyed the changes of apoptotic proteins, such as mitochondrial proteins, MAPK proteins and P53. When treating with OxLDL in macrophages, the mitochondrial potent lowered to 27.4% compared to control (the group of native LDL). Interestingly, the group co-treating with HSE and PCA in highest concentrations recovered the mitochondrial potent to 100% and 104%. At this moment, the mitochondrial proteins, MAPK proteins and P53 were changed because of the extreme oxidative stress, and more over than, this stress would make cell to trigger to apoptosis. However, co-treated with HSE or PCA (especially the groups in high concentrations) would change the expressions of these proteins and make cell survive. In addition to, we also detected the contents of vitamin E and free radicals existing in cell inside and in media. By the way, either in cell inside or in media, we examined the inhibitory level of OxLDL by co-treating HSE or PCA. We found that high concentrations of HSE and PCA were able to decelerate the level of LDL oxidation, and the inhibitory effects could be contributed by increasing vitamin E and by decreasing the content of free radicals. Taken the above together, these two antioxidants possessed the abilities to reduce oxidative stress in either cell inside or environment (media). These inhibitory effects would slow down the macrophage apoptosis induced by OxLDL. Under the circumstance, HSE and PCA were able to prevent the blood vessel from forming atherosclerotic lesions, and further prevent vascular diseases.

碩士
中山醫學大學
生化暨生物科技研究所
101
In recent years, a natural plant extract an effective ingredient, thereby preventing or inhibiting the occurrence of cancer research has been the current trend. Roselle polyphenols Hibiscus sabdariffa Polyphenol-rich extract (HPE), one of the component in Roselle’s function, our previous studies indicate that HPE can effectively promote apoptosis on gastric cancer and colon cancer cell, the impact on tumor metastasis has not been clarified, so the next one probe into whether HPE inhibit cancer cell metastasis, and to explore the mechanism. We conducted MTT assay, using the DLD-1 colon cancer cells treated with different doses of HPE, observations DLD-1 cell viability, followed by DLD-1 cell as the experimental model, a fine cells induced lethal dose (IC50) of the HPE (1,2,3 mg / ml) for the following experiments. First observation of DLD-1 cells in wound healing experiments, we found that the higher the dose with the addition, DLD-1 cells crawling ability weaker; followed by cell invasion and metastasis experiments by boyden chamber and transwell assay results observed with the addition of the higher dose, DLD-1 cells, invasion and metastasis ability are increasingly inhibited. Zymography we used the experimental observations and related MMP and uPA expression that MMP-2 and uPA activity of HPE in a significant inhibitory effect, and a concentration-dependent manner. In order to clarify the molecular mechanism, we analysis cytoskeletal proteins, adhesion molecules, and the specificity of inhibition of Akt signal pathway to confirmed, HPE via regulating CD44 signaling pathway effectively inhibited DLD-1 colon cancer cell metastasis and invasion effect.