Made available in DSpace on 2015-03-03T13:58:17Z (GMT). No. of bitstreams: 1 CybelleTM.pdf: 889951 bytes, checksum: bcf6eb5adb7011fa19c2ab4cd178326c (MD5) Previous issue date: 2007-09-05 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / ?Fucans, sulfated polysaccharides extracted from brown algae and some echinoderms, have been extensively studied for its diverse biological activities and because of its interference with molecular mechanisms of cell to cell recognition, including leukocyte trafficking from blood vessels into sites of inflammation mediated by selectin, a family of adhesion molecules. In the present study, we examined structural features of a heterofucan extracted from brown algae Padina gymnospora and its effect on the leukocyte migration to the peritoneum. The sulfated polysaccharides were extracted from the brown seaweed by proteolysis with the proteolytic enzyme maxatase. The presence of protein and uronic acid contamination was detected in the crude polysaccharide extract. Fractionation of the crude extract with growing concentrations of acetone produced five fractions with different concentrations of fucose, xylose, uronic acid, galactose, glucose and sulfate. The fraction precipitated with 1.5 volumes of acetone was characterized by infrared and nuclear magnetic resonance, through which can be observed the presence of sulfate groups in the C4 of -L-fucose. The anti-inflammatory action of this composite was assessed by a sodium thioglycollate-induced peritonitis assay and through nitric oxide production by the peritoneal macrophages using Griess reagent. Fraction F1.5 was efficient in reducing leukocyte influx into the peritoneal cavity when 10 mg/kg and 25mg/kg were used, resulting in a decrease of 56 and 39%, respectively. A decrease of nitric oxide production occurred when high concentrations of fucana were used. The cytotoxicity of the composite was also assessed using the reduction of 3-(4,5 dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide (MTT). Fraction F1.5 had no cytotoxicity when 500 ?g/mL of the fraction was used. This study suggests the use of fraction F1.5 (heterofucan) as an anti-inflammatory / Fucanas, polissacar?deos sulfatados extra?dos de alga marrom e alguns equinodermos, t?m sido extensivamente estudadas devido as suas diversas atividades biol?gicas e pela sua interfer?ncia com mecanismos moleculares de reconhecimento celular, incluindo o tr?fego de leuc?citos dos vasos sangu?neos para o interior de s?tios inflamat?rios mediado pela fam?lia das mol?culas de ades?o denominada selectinas. Neste presente estudo, n?s examinamos a estrutura de uma heterofucana extra?da da alga marrom Padina gymnospora e seu efeito sobre a migra??o de leuc?citos para o perit?nio. Os polissacar?deos sulfatados foram extra?dos da alga marrom por prote?lise com a enzima proteol?tica maxatase. A presen?a de contamina??o por prote?nas e ?cido ur?nico foi detectada no extrato bruto dos polissacar?deos. Fracionamento do extrato bruto com concentra??es crescentes de acetona produziu cinco fra??es com diferentes concentra??es de fucose, xilose, ?cido ur?nico, galactose, glicose e sulfato. A fra??o precipitada com 1,5 volume de acetona foi caracterizada por infravermelho e resson?ncia magn?tica nuclear, atrav?s das quais se p?de constatar a presen?a de grupos sulfato no C4 da -L-fucose. A a??o antiinflamat?ria deste composto foi avaliada atrav?s do ensaio de peritonite induzida por tioglicolato de s?dio e atrav?s da produ??o de ?xido n?trico por macr?fagos peritoneais utilizando-se o reagente de Griess. A fra??o F1,5 mostrou-se eficiente na redu??o do influxo leucocit?rio para a cavidade peritoneal quando utilizados 10 mg/Kg e 25 mg/Kg resultando em uma diminui??o na ordem de 56 e 39 %, respectivamente. Uma diminui??o na produ??o de ?xido n?trico ocorreu quando altas concentra??es de fucana foram usadas. A citotoxicidade do composto tamb?m foi avaliada utilizando-se como par?metro a redu??o do 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). A fra??o F1,5 n?o apresentou citotoxicidade quando utilizados 500 ?g/ml da fra??o. Este estudo sugere o uso da fra??o F1,5 (heterofucana) como um antiinflamat?rio

Made available in DSpace on 2014-12-17T14:03:41Z (GMT). No. of bitstreams: 1 TarcianaCGA.pdf: 634423 bytes, checksum: 54bd90fdc2abea42aaa8c6e698915963 (MD5) Previous issue date: 2006-07-18 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Fucans are a family of sulfated homo and teropolysaccharides respectively, composed mainly of a- (1?2) and a- (1?3) linked by L-fucose residues. Properties such as the ability to act as an anti-contraceptive, to reduce cholesterol levels, and to act as an anti-tumor agent are much related. We have focused our attention on the anticoagulant properties, platelet aggregation, hemorrhagic activity and complement system in vitro of commercial fucoidan (F) and their purified fractions (F1, F2 and F3) from Fucus vesiculosus obtained from fractionation of the fucoidan with different concentrations of acetone 1, 2 and 3v. These compounds were chemically characterized and the fucoidan (F) was modified by desulfation. The anticoagulant activity of the compounds was assessment by activated partial thromboplastin time (APTT) and prothrombine time assay (PT) using citrated normal human plasma. The results of APPT test showed that F, F1 and F2 have high anticoagulants activities 240.0 s (5 ?g). The F3 showed 73.7 s in the same concentrations. The results obtained with PT test to F, F1, F2 and F3 were 81.5 s, 120.0 s, 57.1 and 32.5 s respectively with 50 ?g. The dessulfated polymer showed a decrease in the anticoagulant activity in these two tests. Platelet aggregation assay was measured turbidimetrically with platelet aggregometer by method of Born. The aggregation platelet with F and fractions F1, F2 and F3 exhibited a two-phase answer in the concentration of 5 mg/mL with maximum aggregation of 76.36 ? 10.3% ; 69.54 ? 9.40%; 75.94 ? 9.01%; 51.13 ? 9.59% respectively. However, was observed a hipoaggregate profile F (15.17 ? 5.2%), F1 (7.40 ? 3.04 %), F2 (19.1 ? 5.41%) and F3 (5.09 ? 3.02%) at 0.1 mg/mL. The hemorrhagic activity assay was carried in Wistar rats and showed that these compounds have low hemorrhagic effect when compared to heparin. The complement system ( alternative pathway was made using non-sensibilized rabbit red blood cells The results of complement system essay showed that F , F2 and F3 have action inhibitory in relation to the group control 0.544, 0.697, 0.622 and 0.958 respectively The results showed that these compounds have action on this system. Interaction of the polisaccharides with proteins C3 and C4 showed that the fraction F1 stimulated the activity assay hemolytic using red blood cells / Fucanas s?o uma fam?lia de homo e hetero polissacar?deos sulfatados, formadas por uma cadeia central com liga??es α-(1→2) ou α-(1→3) unidas atrav?s de res?duos de L-fucose. Propriedades como a habilidade para agir como um anticoncepcional, reduzir n?veis de colesterol, e agir como um agente anti-tumoral foram relatadas. N?s focalizamos nossa aten??o nas propriedades anticoagulantes, agrega??o plaquet?ria, atividade anti-hemost?tica e sistema de complemento in vitro do fucoidan comercial e das fra??es F1, F2 e F3 de Fucus vesiculosus obtidas atrav?s do fracionamento do fucoidan por precipta??o com acetona (1, 2 e 3v). A atividade anticoagulante dos compostos foi avaliada pelo tempo de tromboplastina parcial ativado (APTT) e o tempo de protrombina (PT) usando plasma humano citratado. Os resultados do teste de APPT mostraram que o Fucoidan (F) e as fra??es F1 e F2 t?m altas atividades anticoagulantes 240,0 s (5 ?g), enquanto que a F3 mostrou 73,7s nas mesma concentra??o. Os resultados obtidos com PT para o F, F1, F2 e F3 foram 81,5 s, 120,0 s, 57,1 e 32,5 s respectivamente usando a massa de 50 ?g. A dessulfata??o do Fucoidan demonstrou uma diminui??o da atividade anticoagulante nos dois testes. O ensaio de agrega??o plaquet?ria foi realizado no agregometro de acordo com o m?todo de Born. A agrega??o plaquet?ria induzida pelo fucoidam e pelas fra??es de F1, F2 e F3 exibiram uma resposta bif?sica na concentra??o de 5 mg/mL com amplitude m?xima de agrega??o de 76,36% ? 10,3%; 69,54% ? 9,40%; 75,94% ? 9,01%; 51,13% ? 9,59% respectivamente. Por?m, na concentra??o 0,1 mg/mL foi observada um perfil hipoagregante para o Fucoidan (15,17% ? 5,2) e para as fra??es F1 (7,4% ? 3,04%), F2 (19,1% ? 5,41) e F3 (5,09% ? 3,02%). A atividade anti-hemost?tica foi realizada com ratos machos da linhagem Wistar e demonstrou que estes compostos t?m efeito hemorr?gico residual menor do que o da heparina. A a??o dos polissacar?deos fucosilados sulfatados na via alternativa do sistema complemento foi realizada atrav?s da utiliza??o de eritr?citos de coelhos n?o sensibilizados. Os resultados para o Sistema Complemento mostraram que o F, F2 e F3 t?m a??o neste sistema, apresentando efeito inibit?rio em rela??o ao grupo controle 0,544, 0,697, 0,622 e 0,958 respectivamente. Intera??es dos polissacar?deos com as prote?nas C3 e C4 demonstraram que a fra??o F1 estimula a atividade hemol?tica do complemento usando eritr?citos de coelho. Conclu?mos que estes a??cares t?m atividade anticoagulante e atuam como inibidores do sistema complemento sendo uma valiosa droga que pode ser empregada em doen?as relacionadas com inflama??o e coagula??o sang??nea

Made available in DSpace on 2014-12-17T14:03:44Z (GMT). No. of bitstreams: 1 ThiagoGC_DISSERT.pdf: 2736665 bytes, checksum: 120a3ba44fefe1ccd9373aeb0ff1629f (MD5) Previous issue date: 2014-02-06 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Seaweeds are a major source of biologically active compounds . In the extracellular matrix of these organisms are sulfated polysaccharides that functions as structural components preventing it against dehydration. The fraction 0.9 (FucB) rich in sulfated fucans obtained from brown seaweed Dictyota menstrualis was chemical characterized and evaluated for pharmacological activity by testing anticoagulant activity, stimulatory action on the synthesis of an antithrombotic heparan sulfate, antioxidant activity and its effects in cell proliferation. The main components were FucB carbohydrates (49.80 ? 0.10 %) and sulfate (42.30 ? 0.015 %), with phenolic compounds ( 3.86 ? 0.016 %) and low protein contamination ( 0.58 ? 0.001 % ) . FucB showed polydisperse profile and analysis of signals in the infrared at 1262, 1074 and 930 cm -1 and 840 assigned to S = O bonds sulfate esters , CO bond presence of 3,6- anhydrogalactose , β -D- galactose non- sulfated sulfate and the axial position of fucose C4 , respectively. FucB exhibited moderate anticoagulant activity , the polysaccharides prolonged time (aPTT ) 200 ug ( > 90s ) partial thromboplastin FucB no effect on prothrombin time (PT), which corresponds to the extrinsic pathway of coagulation was observed. This stimulation promoted fraction of about 3.6 times the synthesis of heparan sulfate (HS) by endothelial cells of the rabbit aorta ( RAEC ) in culture compared with cells not treated with FucB . This has also been shown to compete for the binding site with heparin. The rich fraction sulfated fucans exhibited strong antioxidant activity assays on total antioxidant (109.7 and 89.5 % compared with BHT and ascorbic acid standards ) , reducing power ( 71 % compared to ascorbic acid ) and ferric chelation ( 71 , comparing with 5 % ascorbic acid). The fraction of algae showed cytostatic activity on the RAEC cells revealed that the increase of the synthesis of heparan sulfate is not related to proliferation. FucB showed antiproliferative action on cell lines modified as Hela and Hep G2 by MTT assay . These results suggest that FucB Dictyota menstrualis have anticoagulant , antithrombotic , antioxidant potential as well as a possible antitumor action, promoting the stimulation of the synthesis of antithrombotic HS by endothelial cells and is useful in the prevention of thrombosis, also due to its inhibitory action on species reactive oxygen ( ROS ) in some in vitro systems , being involved in promoting a hypercoagulable state / Algas marinhas s?o uma das principais fontes de compostos biologicamente ativos. Na matriz extracelular desses organismos existem os polissacar?deos sulfatados que funcionam como componente estrutural prevenindo-a contra desidrata??o. A fra??o 0,9 (FucB) rica em fucanas sulfatadas obtida da alga marrom Dictyota menstrualis foi caracterizada quimicamente e avaliada quanto a atividade farmacol?gica por meio de ensaios de atividade anticoagulante, a??o estimulat?ria sobre a s?ntese de heparam sulfato antitromb?tico, atividade antioxidante e seus efeitos na prolifera??o celular. Os principais componentes da FucB foram carboidratos (49,80 ? 0,10%) e sulfato (42,30 ? 0,015%), apresentando compostos fen?licos (3,86 ? 0,016%) e baixa contamina??o prot?ica (0,58 ? 0,001%). FucB mostrou perfil polidisperso e sinais na an?lise de infravermelho em 1262, 1074 e 930 e 840 cm-1 atribu?dos a liga??es S=O de ?steres de sulfato, presen?a de liga??o C-O de 3,6-anidrogalactose, β-D-galactose n?o sulfatada e sulfato na posi??o axial do C4 da fucose, respectivamente. FucB exibiu moderada atividade anticoagulante, este polissacar?deo prolongou o tempo de tromboplastina parcial activada (aPTT) a 200 ug (>90s) n?o foi observado qualquer efeito de FucB sobre o tempo de protrombina (PT), que corresponde a via extr?nseca da coagula??o. Esta fra??o promoveu estimula??o cerca de 3,6 vezes na s?ntese de heparam sulfato (HS) pelas c?lulas endoteliais da aorta de coelho (RAEC), em cultura, quando comparadas com as c?lulas n?o tratadas com FucB. Esta tamb?m demonstrou competir pelo s?tio de liga??o com a heparina. A fra??o rica em fucanas sulfatadas exibiu forte a??o antioxidante sobre os ensaios de antioxidante total (109,7 e 89,5% comparados com padr?es BHT e ?cido asc?rbico), poder redutor (71% comparado ao ?cido asc?rbico) e quela??o f?rrica (71,5% comparando com ?cido asc?rbico). A fra??o dessa alga mostrou atividade citost?tica sobre as c?lulas RAEC revelando que o aumento da s?ntese de heparan sulfato n?o est? relacionado ? prolifera??o. FucB apresentou a??o antiproliferativa sobre linhagens celulares modificadas como Hela e Hep G2 pelo ensaio de MTT. Esses resultados sugerem que FucB de Dictyota menstrualis tem potencial anticoagulante, antitromb?tico, antioxidante bem como uma poss?vel a??o antitumoral, promovendo a estimula??o da s?ntese de HS antitromb?tico pelas c?lulas endoteliais, sendo ?til na preven??o da trombose, devido tamb?m a sua a??o inibit?ria sobre as esp?cies reativas do oxig?nio (ROS) em alguns sistemas in vitro, estando envolvidos na promo??o de estado de hipercoagulabilidade