Made available in DSpace on 2014-12-17T14:03:32Z (GMT). No. of bitstreams: 1 CinthiaBST.pdf: 1239764 bytes, checksum: b0f986e10d6574ec55e837255b634b41 (MD5) Previous issue date: 2010-02-11 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / The exopolysaccharides are extracellular compounds produced by some species of fungi and bacteria. It is suggested that these molecules, even when in the form of complex polysaccharide-peptide, are the main bioactive molecules of many fungus. Some of the biological activities displayed by these compounds can be accentuated and others may arise when you add chemically polar or nonpolar groups to polysaccharides. The fruiting body of Pleurotus sajor-caju produces a heteropolysaccharide with antineoplastic and antimicrobial activity, but other biological activities of this polymer have not been evaluated. In this work the exopolysaccharide of Pleurotus sajor-caju was sulfated chemically and structurally characterized. We also evaluated the antiproliferative, antioxidant and anticoagulant activities from native exopolysaccharide (PN) and its sulfated derivated (PS). Polyacrylamide gel electrophoresis, infrared spectroscopy and nuclear magnetic resonance (??C) proved successful in sulfation of PN to obtain PS. Analysis by gas chromatography-mass spectroscopy showed that PN and PS are composed of mannose, galactose, 3-O-methyl-galactose and glucose in proportion percentage of 44,9:16,3:19,8:19 and 49, 7:14,4:17,7:18,2, respectively. The percentage of sulfate found in PS was 22.5%. Antioxidants assays revealed that the sulfation procedure affects differently the activities of exopolysaccharides, while the total antioxidant capacity, the scavenging activity of superoxide radical and ferric chelating were not affected by sulfation, on the other hand the chemical modification of PN enhanced the scavenging activity of hydroxyl radical and reducing power. PS also showed anticoagulant activity in a dose-dependent manner and clotting time was 3.0 times higher than the baseline value in APTT at 2 mg/mL. The exopolysaccharide not presented antiproliferative activity against HeLa tumor cells, but PS affects the cellular proliferation in a time-dependent manner. After 72 h, the inhibition rate of PS (2.0 mg/mL) on HeLa cells was about 60%. The results showed that PN sulfation increase some of their activities. / Os exopolissacar?deos s?o compostos extracelulares produzidos por algumas esp?cies de fungos e bact?rias. ? sugerido que estas mol?culas, inclusive quando na forma de complexo polissacar?deo-pept?deo, s?o as principais mol?culas bioativas de v?rios fungos. Muitas das atividades biol?gicas apresentadas por esses compostos podem ser acentuadas e outras podem surgir quando se adiciona quimicamente aos polissacar?deos grupamentos polares ou apolares. O corpo de frutifica??o de Pleurotus sajor-caju produz um heteropolissacar?deo com atividade antioneopl?sica e antimicrobiana, contudo outras atividades biol?gicas desse pol?mero ainda n?o foram avaliadas. Neste trabalho o exopolissacar?deo de Pleurotus sajor-caju foi sulfatado quimicamente e caracterizado estruturalmente. Tamb?m foram avaliadas as atividades antiproliferativa, antioxidante e anticoagulante do exopolissacar?deo nativo (PN) e de seu derivado sulfatado (PS). Eletroforese em gel de agarose, espectroscopia de infravermelho e resson?ncia magn?tica nuclear (??C) comprovaram o sucesso na sulfata??o de PN para a obten??o de PS. An?lise por cromatografia gasosa acoplada a espectroscopia de massa mostrou que PN e PS s?o constitu?dos de manose, galactose, 3-O-metil-galactose e glicose na propor??o percentual de 44,9:16,3:19,8:19 e 49,7:14,4:17,7:18,2, respectivamente. O percentual de sulfato encontrado em PS foi de 22,5%. Testes antioxidantes revelaram que o processo de sulfata??o influencia de forma diferente nas atividades do exopolissacar?deo. Enquanto a capacidade antioxidante total, a capacidade de seq?estro de radical super?xido e a quela??o f?rrica n?o foram influenciadas pela sulfata??o, essa potencializou a atividade seq?estradora de radicais hidroxila e o poder redutor do exopolissacar?deo. Ap?s o processo de sulfata??o o exopolissacar?deo passou a apresentar atividade anticoagulante de forma dose-dependente, triplicando o tempo de coagula??o em rela??o ao controle numa concentra??o de aproximadamente 2 mg/mL. O exopolissacar?deo n?o apresentou atividade antiproliferativa frente ?s c?lulas tumorais HeLa, por?m ap?s sulfata??o ele passou a apresentar essa atividade de forma tempo- ependente, chegando a inibir em 60% a taxa de prolifera??o das c?lulas com 2 mg/mL, ap?s 72 h de exposi??o. Os resultados aqui obtidos mostraram que a sulfata??o do exopolissacar?deo potencializou algumas de suas atividades.

Made available in DSpace on 2014-12-17T14:03:32Z (GMT). No. of bitstreams: 1 RaniereFMS.pdf: 3115798 bytes, checksum: 664aaedacd292e8b8c2d08a15e193378 (MD5) Previous issue date: 2010-02-25 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / The corn cob is an agricultural by-product still little used, this in part due to the low knowledge of the biotechnological potential of their molecules. Xylan from corn cobs (XSM) is a polysaccharide present in greater quantity in the structure of plant and its biotechnology potential is little known. This study aimed to the extraction, chemical characterization and evaluation of biological activities of xylan from corn cobs. To this end, corncobs were cleaned, cut, dried and crushed, resulting in flour. This was subjected to a methodology that combines the use of alkaline conditions with waves of ultrasound. After methanol precipitation, centrifugation and drying was obtained a yield of 40% (g/g flour). Chemical analysis indicated a high percentage of polysaccharides in the sample (60%) and low contamination by protein (0.4%) and phenolic compounds (> 0.01%). Analysis of monosaccharide composition indicated the presence of xylose:glucose:arabinose:galactose:mannose:glucuronic acid in a molar ratio 50:20:15:10:2.5:2.5. The presence of xylan in the sample was confirmed by nuclear magnetic resonance (?H and ??C) and infrared spectroscopy (IR). Tests were conducted to evaluate the antioxidant potential of XSM. This showed a total antioxidant capacity of 48.45 EAA/g sample. However, did not show scavenging activity of superoxide and hydroxyl radical and also reducing power. But, showing a high capacity chelating iron ions with 70% with about 2 mg/mL. The ability to XSM to influence cell proliferation in culture was also evaluated. This polymer did not influence the proliferation of normal fibroblast cells (3T3), however, decreased the rate of proliferation of tumor cells (HeLa) in a dose-dependent, reaching an inhibition of about 50% with a concentration around 2 mg/mL. Analyzing proteins related to cell death, by immunoblotting, XSM increases the amount of Bax, Bcl-2 decrease, increase cytochrome c and AIF, and reduce pro-caspase-3, indicating the induction of cell death induced apoptosis dependent and independent of caspase. XSM did not show anticoagulant activity in the PT test. However, the test of activated partial thromboplastin time (aPTT), XSM increased clotting time at about 5 times with 600 ?g of sample compared with the negative control. The presence of sulfate on the XSM was discarded by agarose gel electrophoresis and IR. After carboxyl-reduction of XSM the anticoagulant activity decreased dramatically. The data of this study demonstrate that XSM has potential as antioxidant, antiproliferative and anticoagulant compound. Future studies to characterize these activities of XSM will help to increase knowledge about this molecule extracted from corn and allow their use in functional foods, pharmaceuticals and chemical industries. / O sabugo de milho ? um subproduto agr?cola ainda pouco utilizado, isto se deve em parte ao baixo conhecimento do potencial biotecnol?gico de suas biomol?culas. Xilana de sabugo de milho (XSM) ? um polissacar?deo presente em maior quantidade na estrutura do vegetal e seu potencial biotecnol?gico ? pouco conhecido. Este trabalho teve como objetivo a extra??o, caracteriza??o qu?mica e avalia??o de atividades biol?gicas de XSM. Sabugos de milho foram limpos, cortados, desidratados e triturados, dando origem a uma farinha. Esta foi submetida a uma metodologia que combina o uso de meio alcalino com ondas de ultra-som. Ap?s precipita??o metan?lica, centrifuga??o e secagem obteve-se um rendimento de 40% (g/g de farinha). An?lises qu?micas indicaram um alto percentual de polissacar?deos na amostra (60%) e baixa contamina??o por prote?nas (0.4%) e compostos fen?licos (>0.01%). An?lises da composi??o monossacar?dica por cromatografia em papel e por cromatografia l?quida de alta performance (CLAE) indicaram a presen?a de xilose:glicose:arabinose:galactose:manose:?cido glucur?nico em uma propor??o molar de 50:20:15:10:2,5:2,5. A presen?a de xilana na amostra foi confirmada por resson?ncia magn?tica nuclear (13C e 1H) e por espectroscopia de infravermelho (IR). Testes foram realizados para avalia??o do potencial antioxidante de XSM. Esta mostrou uma capacidade antioxidante total de 48.45 EAA/g de amostra. Contudo, a mesma n?o mostrou atividade sequestradora de super?xido, radical hidroxila bem como poder redutor. Em contra partida, apresentou 70% de atividade quelante de ?ons de ferro na concentra??o de 2 mg/mL. A capacidade de XSM em influenciar a prolifera??o celular em cultura tamb?m foi avaliada. Este polissacar?deo n?o influenciou a prolifera??o de c?lulas fibrobl?sticas normais (3T3), entretanto, diminuiu a taxa de prolifera??o de c?lulas tumorais (HeLa) de maneira dose-dependente, chegando a uma inibi??o de aproximadamente 50% com concentra??o em torno de 2 mg/mL. Analisando prote?nas relacionadas ? morte celular, atrav?s de immunoblotting, XSM aumenta a quantidade de Bax, citocromo c e AIF e diminui Bcl-2 e procaspase- 3, indicando a indu??o de morte celular por apoptose dependente e independente de caspase. XSM n?o apresentou atividade anticoagulante pelo teste de PT. Todavia, no teste de tempo de tromboplastina parcial ativada (aPTT), XSM aumentou o tempo de coagula??o em cerca de 5 vezes utilizando 600 ?g de amostra, quando comparadas com o controle negativo. A presen?a de sulfato ligado a XSM foi descartada por eletroforese em gel de agarose e por IR. Ap?s carboxirredu??o de XSM a atividade anticoagulante diminuiu drasticamente. Os dados deste trabalho demonstram que XSM apresenta potencial como composto antioxidante, antiproliferativo e anticoagulante. Estudos futuros de caracteriza??o dessas atividades do XSM contribuir?o para aumentar o conhecimento sobre esta mol?cula extra?da de milho e permitir?o a sua utiliza??o em alimentos funcionais, produtos farmac?uticos e ind?strias qu?micas.