Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-02-05T23:00:04Z No. of bitstreams: 1 LiviaDeLourdesDeSousaPinto_DISSERT.pdf: 1563914 bytes, checksum: afff2cde0e6703ffa7b4fae03c2036ac (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-02-16T22:03:58Z (GMT) No. of bitstreams: 1 LiviaDeLourdesDeSousaPinto_DISSERT.pdf: 1563914 bytes, checksum: afff2cde0e6703ffa7b4fae03c2036ac (MD5) / Made available in DSpace on 2016-02-16T22:03:58Z (GMT). No. of bitstreams: 1 LiviaDeLourdesDeSousaPinto_DISSERT.pdf: 1563914 bytes, checksum: afff2cde0e6703ffa7b4fae03c2036ac (MD5) Previous issue date: 2015-03-13 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / Condroitim sulfato (CS) ? um glicosaminoglicano natural presente na matriz extracelular de tecidos conectivos, podendo ser extra?do e purificado desses tecidos. O CS est? envolvido em diversas fun??es biol?gicas, o que pode estar relacionado ? variabilidade estrutural que possui, apesar da simplicidade da cadeia linear dessa mol?cula. Pesquisas na ?rea biotecnol?gica e farmacol?gica com rejeitos provenientes da aquicultura vem sendo desenvolvidas no Brasil. Nas ?ltimas d?cadas, a til?pia (Oreochromis niloticus), peixe nativo da ?frica, tem sido uma das esp?cies mais cultivadas em v?rias regi?es do mundo, inclusive no Brasil. A tilapicultura ? uma atividade economica cujo principal inconveniente ? a grande quantidade de res?duos que s?o descartados pelos produtores. Entende-se que o material descartado pode ser aproveitado em pesquisas como fonte de mol?culas com importante aplica??o biotecnol?gica, o que tamb?m contribui na redu??o de impactos ambientais e favorece o desenvolvimento dessa atividade de maneira ecologicamente correta. Dessa forma, v?sceras de til?pias nil?tica foram submetidas ? prote?lise, em seguida os glicosaminoglicanos foram complexados com resina de troca i?nica (Lewatit), fracionados com volumes crescentes de acetona e purificados atrav?s da cromatografia de troca i?nica DEAE-Sephacel. Na sequ?ncia, a fra??o foi analisada atrav?s de eletroforese em gel de agarose e resson?ncia magn?tica nuclear (RMN). O perfil eletrofor?tico do composto, a an?lise dos espectros 1H de RMN e a correla??o do HSQC permitem afirmar que o composto corresponde a uma mol?cula de condroitim sulfato. O ensaio de MTT foi utilizado para avalia??o da viabilidade celular na presen?a do CS isolado de til?pia e mostrou que o composto n?o ? citot?xico a c?lulas normais provenientes do fibroblasto de embri?o de camundongos (3T3). Ent?o, o composto foi testado quanto a capacidade de reduzir o influxo de leuc?citos em modelo de peritonite aguda (in vivo) induzida por tioglicolato de s?dio. Nesse contexto, foi realizada a contagem total e diferencial de leuc?citos do sangue e l?quido peritoneal coletadas, respectivamente, da veia cava e do lavado peritoneal de cada animal submetido ao experimento. O condroitim sulfato, pela primeira vez isolado de til?pia (CST ), foi capaz de reduzir a migra??o de leuc?citos ? cavidade peritoneal de camundongos inflamados em at? 80,4% na dose de 10?g/kg. Os resultados mostram ainda que houve redu??o significativa (p<0,001) da popula??o de polimorfonucleares do lavado peritoneal nas tr?s doses testadas (0,1?g/kg; 1?g/kg e 10?g/kg) quando comparado ao controle positivo (apenas tioglicolato). Portanto, uma vez que a estrutura e o mecanismo de a??o do CST tenham sido totalmente elucidados, esse composto pode apresentar potencial para uso terap?utico em doen?as inflamat?rias. / Chondroitin sulfate (CS) is a naturally glycosaminoglycan found in the extracellular matrix of connective tissues and it may be extracted and purified those tissues. CS is involved in various biological functions, which may be related to the having structural variability, despite the simplicity of the linear chain structure from this molecule. Researches in biotechnology and pharmaceutical field with wastes from aquaculture has been developed in Brazil. In recent decades, tilapia (Oreochromis niloticus), native fish from Africa, has been one of the most cultivated species in various regions of the world, including Brazil. The tilapia farming is a cost-effective activity, however, it generates large amount of wastes that are discarded by producers. It is understood that waste from tilapia can be used in research as a source of molecules with important biotechnological applications, which also helps in reducing environmental impacts and promote the development of an ecofriendly activity. Thus, nile tilapia viscera were subjected to proteolysis, then the glycosaminoglycans were complexed with ion exchange resin (Lewatit), it was fractionated with increasing volumes of acetone and purified by ion exchange chromatography DEAE-Sephacel. Further, the fraction was analyzed by agarose gel electrophoresis and nuclear magnetic resonance (NMR). The electrophoretic profile of the compound together the analysis of 1H NMR spectra and the HSQC correlation allow to affirm that the compound corresponds to a molecule like chondroitin sulfate. MTT assay was used to assess cell viability in the presence of CS tilapia isolated and showed that the compound is not cytotoxic to normal cells such as cells from the mouse embryo fibroblast (3T3). Then, this compound was tested for the ability to reduce the influx of leukocytes in model of acute peritonitis (in vivo) induced by sodium thioglycolate. In this context, it was done total and differential leukocytes counting in the blood and peritoneal fluid collected respectively from vena cava and the peritoneal cavity of the animals subjected to the experiment. The chondroitin sulfate for the first time isolated from tilapia (CST ) was able to reduce the migration of leukocytes to the peritoneal cavity of inflamed mice until 80.4 per cent at a dose 10?g/kg. The results also show that there was a significant reduction (p<0.001) of the population of polymorphonuclear leukocytes from peritoneal cavity in the three tested doses (0.1?g/kg; 1?g/kg and 10?g/kg) when it was compared to the positive control (just thioglycolate). Therefore, since the CST structure and mechanism of action has been completely elucidated, this compound may have potential for therapeutic use in inflammatory diseases