Mild-acid hydrolysis of a native polysulfated fraction from Acanthophora muscoides generates sulfated oligosaccharides displaying in vitro thrombin generation inhibition / Hidrólise ácida branda de uma fração polissulfatada de Acanthophora muscoides gera oligossacarídeos sulfatados mostrando inibição de geração de trombina in vitro

Acanthophora muscoides (Rhodophyta) has three polysulfated fractions    (Am-1, Am-2 and Am-3). Am-2 displayed anti-inflammation and serpin-independent anticoagulation; however, no effect of oligomers on thrombin-generation (TG) assay has been demonstrated. This study employed mild-acid hydrolysis to obtain low-molecular-size derivatives from Am-2 and compared in vitro inhibitory effects between intact Am-2 and its hydrolysates on a TG assay. The enzymatic extract was fractionated by DEAE-cellulose that revealed Am-2 eluted with 0.75-M NaCl containing sulfate (23%), hexoses (51%) and devoid of proteins, and indicating, by one-dimension nuclear magnetic resonance, structure of galactan similar to that of extract. The depolymerization with HCl (0.02 or 0.04-M, 60C) for different times progressively reduced the charge density and the molecular-size of Am-2 based on electrophoreses in agarose and polyacrylamide gels, respectively, where at higher acid concentration and critical time up to 5 h yielded fragment of   10-kDa similar to that of unfractionated heparin (UHEP). Regarding the TG assay, intact Am-2 inhibited concentration-dependent the intrinsic pathway, whereas its hydrolysates abolished it like UHEP, when in 60-fold diluted human plasma using chromogenic method by a continuous system. The results reveal an alternative approach for producing oligosaccharides(AU)

A rodofícea A. muscoides possui três frações polissulfatadas (-1, -2 e Am-3). Am-2 mostrou efeito anti-inflamação e anticoagulação independente de serpina. Entretanto, não se demonstrou efeito de oligômeros sobre ensaio de geração de trombina (GT). Este estudo empregou hidrólise ácida branda para obter derivados de tamanho molecular baixo de Am-2 e os efeitos inibitórios in vitro entre Am-2 intacta e hidrolisados comparados sobre um ensaio de GT. O extrato polissacarídico, fracionado por DEAE-celulose, revelou Am-2 eluída com NaCl-0,75M contendo sulfato (23%), hexoses (51%) e destituída de proteínas. E, ainda, por ressonância magnética nuclear-unidimensional, indicando galactana semelhante a do extrato. A depolimerização com HCl (0,02 ou 0,04-M; 60C) reduziu, progressivamente durante tempos diferentes, a densidade de carga e o tamanho molecular de Am-2 baseada nas eletroforeses em géis de agarose e de poliacrilamida, respectivamente, em que, concentração ácida elevada e tempo crítico de até 5h renderam fragmento de 14-kDa semelhante ao da heparina não fracionada (HEPNF). Já no ensaio de GT, Am-2 intacta, quando em plasma humano diluído 60 vezes, usando método cromogênico por meio de sistema contínuo, quem inibiu a via intrínseca dependente de concentração, ao passo que seus hidrolisados aboliram como HEPNF, exceto fragmento análogo (inibição 92,87%). Os resultados revelam(AU)

Análise de duas metodologias de precipitação sobre o rendimento, características estruturais e atividade de polissacarídeos sulfatados de Gracilaria cornea (Rhodophyta) / Analysis of two precipitation methods on the yield, structural features and activity of sulfated polysaccharides from Gracilaria cornea (Rhodophyta)

A demanda global de produtos naturais de algas marinhas tem aumentado mundialmente. Entretanto, a obtenção de polissacarídeos sulfatados (PSs) com álcool isoamílico (AIA) não é relatada. Investigou-se a eficiência de dois métodos (M) de precipitação de PSs da alga marinha vermelha Gracilaria cornea. Os PSs isolados enzimaticamente foram concentrados com cloreto cetilpiridimínio (M I) ou AIA (M II). Os extratos foram examinados, segundo seu rendimento, características estruturais e efeitos in vitro sobre o tempo de tromboplastina parcial ativada (TTPA) usando plasma humano normal e heparina padrão (193 UI mg-1). A diferença nos rendimentos foi 12,99% e semelhante determinação quantitativa de sulfato foi obtida entre os métodos ( 26%). A eletroforese em gel de agarose revelou diferenças em termos de densidade de cargas entre os extratos. Enquanto ambos os extratos revelaram agarocoloides, o método M II também se mostrou alternativo para lipídios, proteínas e ácidos nucleicos de acordo com a análise de infravermelho. Os extratos praticamente não modificaram o TTPA (1,95 e 2 UI mg-1 para M I e M II, respectivamente). Os resultados revelaram AIA como um solvente alternativo para obtenção de PSs da alga marinha vermelha G. cornea, dependendo do critério de utilização na indústria.(AU)

The global demand for natural products from seaweeds has increased worldwide; however, no description of the use of isoamly alcohol (IAA) for obtaining of sulfated polysaccharides (SPs) has been reported. We investigated the efficiency of two precipitation methods (M) in obtaining SPs from the red seaweed Gracilaria cornea. SPs enzymatically isolated were concentrated with cetylpyridinium chloride (M I) or IAA (M II) and extracts were examined with regard to their yield, structural features and in vitro effects on the activated partial thromboplastin time (APTT) using normal human plasma and standard heparin (193 IU mg-1). Yield difference reached 12.99%. Quantitative determination of sulfate was similar between the two methods ( 26%), but extracts revealed different pattern on charge density by agarose gel electrophoresis. Whereas both extracts revealed as agarocolloids, alternative M II was also efficient for lipids, proteins and nucleic acids according to the infrared analysis. Extracts had virtually no effect on APPT (1.95 and 2 IU mg-1 for M I and M II, respectively). The results revealed IAA as an alternative solvent for obtaining SPs from the red seaweed G. cornea, depending on the industry usage criterion.(AU)

Analysis of two drying methods on the yield and activity of sulfated polysaccharides extracted from Halymenia sp. (Rhodophyceae) - doi: 10.4025/actascibiolsci.v34i1.6961

Sulfated polysaccharides (SPs) have attracted growing interest for various biotechnological applications. We evaluated the efficiency of two methods of drying SPs (M I and II) extracted from Halymenia sp Rhodophyceae in order to compare the yield, purification and anticoagulant activity. The total SPs (TSPs) were first extracted with papain in 100 mM sodium acetate (pH 5.0) containing cysteine and EDTA (5 mM). The TSPs obtained were dried in an oven (M I) or lyophilized (M II) and then examined by ion exchange chromatography (DEAE-cellulose) using the NaCl gradient technique. The fractions were analyzed by 0.5% agarose gel electrophoresis and the in vitro anticoagulant activity was evaluated by the activated partial thromboplastin time test using normal human plasma and compared to heparin (HEP) (193.00 IU mg-1). There was a difference in TSP yield of 19.05% and similar chromatographic SP profiles. Electrophoresis revealed fractions with distinct resolutions. The fractions eluted with 0.75 M of salt (M I and II) were the most active, measuring 27.40 and 72.66 IU mg-1, respectively, when compared to HEP. Therefore, obtaining SP with anticoagulant activity from Halymenia sp. is more efficient by freeze-drying.

Analysis of two drying methods on the yield and activity of sulfated polysaccharides extracted from Halymenia sp (Rhodophyceae) / Análise de dois métodos de secagem sobre o rendimento e atividade de polissacarídeos sulfatados extraídos da rodofícea Halymenia sp

Sulfated polysaccharides (SPs) have attracted growing interest for various biotechnological applications. We evaluated the efficiency of two methods of drying SPs (M I and II) extracted from Halymenia sp Rhodophyceae in order to compare the yield, purification and anticoagulant activity. The total SPs (TSPs) were first extracted with papain in 100 mM sodium acetate (pH 5.0) containing cysteine and EDTA (5 mM). The TSPs obtained were dried in an oven (M I) or lyophilized (M II) and then examined by ion exchange chromatography (DEAE-cellulose) using the NaCl gradient technique. The fractions were analyzed by 0.5% agarose gel electrophoresis and the in vitro anticoagulant activity was evaluated by the activated partial thromboplastin time test using normal human plasma and compared to heparin (HEP) (193.00 IU mg-1). There was a difference in TSP yield of 19.05% and similar chromatographic SP profiles. Electrophoresis revealed fractions with distinct resolutions. The fractions eluted with 0.75 M of salt (M I and II) were the most active, measuring 27.40 and 72.66 IU mg-1, respectively, when compared to HEP. Therefore, obtaining SP with anticoagulant activity from Halymenia sp. is more efficient by freeze-drying.

Os polissacarídeos sulfatados (PS) têm despertado interesse crescente para diversas aplicações em biotecnologia. Avaliou-se a eficiência de duas metodologias de secagem de PSs (M I e II) extraídos da rodofícea Halymenia sp. a fim de se comparar o rendimento, a purificação e a atividade anticoagulante. Inicialmente, os PS totais (PSTs) foram extraídos com papaína em tampão acetato de sódio 100 mM (pH 5,0) contendo cisteína e EDTA (5 mM). Os PSTs foram secados em estufa (M I) ou liofilizados (M II), seguido por cromatografia de troca iônica (DEAE-celulose) utilizando um gradiente de NaCl. As frações foram analisadas por eletroforese em gel de agarose a 0,5% e a atividade anticoagulante "in vitro" avaliada por meio do teste do tempo de tromboplastina parcial ativada usando plasma humano normal e comparadas à heparina (HEP) (193.00 UI mg-1). Verificaram-se diferença no rendimento de PSTs (19,05%) e semelhantes perfis cromatográficos de PS, revelando, por eletroforese, frações de PS distintas em grau de resolução entre os métodos. As frações eluídas com 0,75 M de sal (M I e II) foram as mais ativas, com valores de atividade da ordem de 27,40 e 72,66 UI mg-1, respectivamente, em relação à HEP. Portanto, a secagem por liofilização seria uma forma mais eficaz na obtenção de PSs com atividade anticoagulante de Halymenia sp.

Isolamento, fracionamento e atividade anticoagulante de iota-carragenanas da Solieria filiformis

This study aimed to isolate, fractionate and evaluate the anticoagulant potential of iota-carrageenans (i-CARs) from Solieria filiformis when two extraction methods (M I and M II) were used. i-CARs were isolated with papain in 0.1M sodium acetate (pH 5.0) containing 5mM cystein and 5mM EDTA (M I) or water (80°C) (M II), and then their chemical composition of total carbohydrates, free sulfate (FS) and contaminant proteins were determined. i-CARs were submitted to anion-exchange chromatography (DEAE-cellulose) using a sodium chloride gradient,being evaluated the activated partial thromboplastin time (APTT) and prothrombin time of obtained fractions and compared to heparin (193IU mg-1). A rich fraction of anticoagulant was also submitted to 0.5% agarose gel electrophoresis procedure. The difference of yield between methods was 10.14%. The chemical composition of FS (29.40%) and the fractionation by DEAE-cellulose showed M I more effectiveness in the obtaining of i-CARs compared to M II. The APTT was also modified for i-CARs from M I. However, the in vitro anticoagulant activity of a rich fraction (8.52IU mg-1) was inferior to heparin.

Este estudo teve como objetivo isolar, fracionar e avaliar o potencial anticoagulante de iota-carragenanas (i-CARs) da rodofícea Solieria filiformis, quando obtidas por dois métodos de extração (M I e M II). As i-CARs foram isoladas com papaína bruta em tampão acetato de sódio 0,1M (pH 5,0), contendo cisteína 5mM e EDTA 5mM (M I) ou água (80°C) (M II) e, em seguida, determinada sua composição química de carboidratos totais, sulfato livre (SL) e proteínas contaminantes. As i-CARs foram submetidas à cromatografia de troca iônica (DEAE-celulose) usando um gradiente de cloreto de sódio, sendo avaliado o tempo de tromboplastina parcial ativada (TTPA) e tempo de protrombina das frações obtidas e comparadas à heparina (193UI mg-1). Uma fração anticoagulante também foi submetida ao procedimento de eletroforese em gel de agarose a 0,5%. A diferença no rendimento de i-CARs entre os métodos foi 10,14%. A composição química de SL (29,40%) e o fracionamento, por DEAE-celulose, indicaram o M I mais eficiente na obtenção de i-CARs, comparado ao M II. O TTPA também foi somente alterado para as i-CARs do M I. Contudo, a atividade anticoagulante in vitro de uma fração rica (8,52UI mg-1) foi inferior à da heparina.

Isolamento, fracionamento e atividade anticoagulante de iota-carragenanas da Solieria filiformis

This study aimed to isolate, fractionate and evaluate the anticoagulant potential of iota-carrageenans (i-CARs) from Solieria filiformis when two extraction methods (M I and M II) were used. i-CARs were isolated with papain in 0.1M sodium acetate (pH 5.0) containing 5mM cystein and 5mM EDTA (M I) or water (80°C) (M II), and then their chemical composition of total carbohydrates, free sulfate (FS) and contaminant proteins were determined. i-CARs were submitted to anion-exchange chromatography (DEAE-cellulose) using a sodium chloride gradient,being evaluated the activated partial thromboplastin time (APTT) and prothrombin time of obtained fractions and compared to heparin (193IU mg-1). A rich fraction of anticoagulant was also submitted to 0.5% agarose gel electrophoresis procedure. The difference of yield between methods was 10.14%. The chemical composition of FS (29.40%) and the fractionation by DEAE-cellulose showed M I more effectiveness in the obtaining of i-CARs compared to M II. The APTT was also modified for i-CARs from M I. However, the in vitro anticoagulant activity of a rich fraction (8.52IU mg-1) was inferior to heparin.

Este estudo teve como objetivo isolar, fracionar e avaliar o potencial anticoagulante de iota-carragenanas (i-CARs) da rodofícea Solieria filiformis, quando obtidas por dois métodos de extração (M I e M II). As i-CARs foram isoladas com papaína bruta em tampão acetato de sódio 0,1M (pH 5,0), contendo cisteína 5mM e EDTA 5mM (M I) ou água (80°C) (M II) e, em seguida, determinada sua composição química de carboidratos totais, sulfato livre (SL) e proteínas contaminantes. As i-CARs foram submetidas à cromatografia de troca iônica (DEAE-celulose) usando um gradiente de cloreto de sódio, sendo avaliado o tempo de tromboplastina parcial ativada (TTPA) e tempo de protrombina das frações obtidas e comparadas à heparina (193UI mg-1). Uma fração anticoagulante também foi submetida ao procedimento de eletroforese em gel de agarose a 0,5%. A diferença no rendimento de i-CARs entre os métodos foi 10,14%. A composição química de SL (29,40%) e o fracionamento, por DEAE-celulose, indicaram o M I mais eficiente na obtenção de i-CARs, comparado ao M II. O TTPA também foi somente alterado para as i-CARs do M I. Contudo, a atividade anticoagulante in vitro de uma fração rica (8,52UI mg-1) foi inferior à da heparina.

Natural heparinoids isolated from Halymenia sp. (Rhodophyceaes) delivery on the Ceará coast - doi: 10.4025/actascibiolsci.v32i3.6282 / Heparinoides naturais isolados de rodofíceas (Halymenia sp.) arribadas na costa cearense - doi: 10.4025/actascibiolsci.v32i3.6282

The increasing demand for heparin (HEP) has led to a search for alternative sources of natural anticoagulants. This study aimed to evaluate the anticoagulant activity of sulfated polysaccharides (SP) isolated from a Halymenia rhodophyceae genus native to the coast of Ceará, Brazil. Total SP were obtained by consecutive digestions with papain in 0.1 M sodium acetate buffer (pH 5.0) containing 5 mM cysteine and 5 mM EDTA, followed by ion-exchange chromatography on DEAE-cellulose column. The obtained fractions were concentrated by lyophilization and submitted to 0.5% agarose gel electrophoresis. Anticoagulant activity was evaluated by the activated partial thromboplastin time (APTT) using plasma from rabbits and a standard HEP (100 IU mg-1) curve. The extractions (53.96%) showed marked differences during the fractionation and in the degree of purification of SP. The species SP showed higher activity anticoagulant than that of HEP. However, the APTT of the fractions changed sharply among the extractions, expressing itself in a dose-dependent manner and increasing from 110.40 (1st extraction) to 143.10 IU mg-1 (3rd extraction). The results suggest that the anticoagulant activity of SP isolated from Halymenia sp. was promoted by inhibition of the intrinsic and/or common pathway of the coagulation cascade. The changes on APTT possibly will be elucidated through the mechanisms of actio

A crescente carência de heparina (HEP) motiva a busca por fontes alternativas de novos anticoagulantes naturais. Objetivou-se avaliar a atividade anticoagulante dos polissacarídeos sulfatados (PS) isolados de uma rodofícea do gênero Halymenia, nativa do litoral cearense, Brasil. Os PS totais foram obtidos por digestões consecutivas com papaína em tampão acetato de sódio 0,1 M (pH 5,0), contendo cisteína 5 mM e EDTA 5 mM, seguidas por cromatografia de troca iônica em coluna de DEAE-celulose. As frações obtidas foram concentradas por liofilização e submetidas à eletroforese em gel de agarose a 0,5%. Os ensaios anticoagulantes foram realizados pelo tempo de tromboplastina parcial ativada (TTPA), usando-se plasma de coelho e uma curva padrão de HEP (100 UI MG-1). As extrações (53,96%) mostraram diferenças marcantes durante o fracionamento e no grau de resolução dos PS. A espécie apresentou PS com atividade anticoagulante superior a HEP. O TTPA das frações modificou-se acentuadamente entre as extrações, expressando-se de maneira dose-dependente e sofrendo um acréscimo de 110,40 (1ª extração) para 143,10 UI MG-1 (3ª extração). Os resultados sugerem que a atividade anticoagulante dos PS isolados de Halymenia sp. foi promovida pela inibição da via intrínseca e/ou comum da cascata de coagulação. As modificações no TTPA possivelmente serão elucidadas pelos mecanismos de

Heparinoides naturais isolados de rodofíceas (Halymenia sp.)arribadas na costa cearense / Natural heparinoids isolated from Halymenia sp. (Rhodophyceaes) delivery on the Ceará coast.

A crescente carência de heparina (HEP) motiva a busca por fontes alternativas de novos anticoagulantes naturais. Objetivou-se avaliar a atividade anticoagulante dos polissacarídeos sulfatados (PS) isolados de uma rodofícea do gênero Halymenia, nativa do litoral cearense, Brasil. Os PS totais foram obtidos por digestões consecutivas com papaína em tampão acetato de sódio 0,1 M (pH 5,0), contendo cisteína 5 mM e EDTA 5 mM, seguidas por cromatografia de troca iônica em coluna de DEAE-celulose. As frações obtidas foram concentradas por liofilização e submetidas à eletroforese em gel de agarose a 0,5%. Os ensaios anticoagulantes foram realizados pelo tempo de tromboplastina parcial ativada (TTPA), usando-se plasma de coelho e uma curva padrão de HEP (100 UI mg-1). As extrações (53,96%) mostraram diferenças marcantes durante o fracionamento e no grau de resolução dos PS. A espécie apresentou PS com atividade anticoagulante superior a HEP. O TTPA das frações modificou-se acentuadamente entre as extrações, expressando-se de maneira dose-dependente e sofrendo um acréscimo de 110,40 (1a extração) para 143,10 UI mg-1 (3a extração). Os resultados sugerem que a atividade anticoagulante dos PS isolados de Halymenia sp. foi promovida pela inibição da via intrínseca e/ou comum da cascata de coagulação. As modificações no TTPA possivelmente serão elucidadas pelos mecanismos de ação envolvidos na coagulação e caracterização estrutural desses compostos. Portanto, a rodofícea Halymenia sp. é uma boa fonte de heparinoides e sugerem-se estudos relacionados ao cultivo da espécie, em proteção aos bancos de algas.(AU)

The increasing demand for heparin (HEP) has led to a search for alternative sources of natural anticoagulants. This study aimed to evaluate the anticoagulant activity of sulfated polysaccharides (SP) isolated from a Halymenia rhodophyceae genus native to the coast of Ceará, Brazil. Total SP were obtained by consecutive digestions with papain in 0.1 M sodium acetate buffer (pH 5.0) containing 5 mM cysteine and 5 mM EDTA, followed by ion-exchange chromatography on DEAE-cellulose column. The obtained fractions were concentrated by lyophilization and submitted to 0.5% agarose gel electrophoresis. Anticoagulant activity was evaluated by the activated partial thromboplastin time (APTT) using plasma from rabbits and a standard HEP (100 IU mg-1) curve. The extractions (53.96%) showed marked differences during the fractionation and in the degree of purification of SP. The species SP showed higher activity anticoagulant than that of HEP. However, the APTT of the fractions changed sharply among the extractions, expressing itself in a dose-dependent manner and increasing from 110.40 (1st extraction) to 143.10 IU mg-1 (3rd extraction). The results suggest that the anticoagulant activity of SP isolated from Halymenia sp. was promoted by inhibition of the intrinsic and/or common pathway of the coagulation cascade. The changes on APTT possibly will be elucidated through the mechanisms of action involved in coagulation and structural characterization of these compounds. Therefore, the red alga Halymenia sp. is a good source of heparinoids, and studies are suggested on the cultivation of this species and on the protection of natural algae banks.(AU)

Analysis of two drying methods on the yield and activity of sulfated polysaccharides extracted from Halymenia sp. (Rhodophyceae) - doi: 10.4025/actascibiolsci.v34i1.6961

Sulfated polysaccharides (SPs) have attracted growing interest for various biotechnological applications. We evaluated the efficiency of two methods of drying SPs (M I and II) extracted from Halymenia sp Rhodophyceae in order to compare the yield, purification and anticoagulant activity. The total SPs (TSPs) were first extracted with papain in 100 mM sodium acetate (pH 5.0) containing cysteine and EDTA (5 mM). The TSPs obtained were dried in an oven (M I) or lyophilized (M II) and then examined by ion exchange chromatography (DEAE-cellulose) using the NaCl gradient technique. The fractions were analyzed by 0.5% agarose gel electrophoresis and the in vitro anticoagulant activity was evaluated by the activated partial thromboplastin time test using normal human plasma and compared to heparin (HEP) (193.00 IU mg-1). There was a difference in TSP yield of 19.05% and similar chromatographic SP profiles. Electrophoresis revealed fractions with distinct resolutions. The fractions eluted with 0.75 M of salt (M I and II) were the most active, measuring 27.40 and 72.66 IU mg-1, respectively, when compared to HEP. Therefore, obtaining SP with anticoagulant activity from Halymenia sp. is more efficient by freeze-drying. 

Analysis of two drying methods on the yield and activity of sulfated polysaccharides extracted from Halymenia sp. (Rhodophyceae) - doi: 10.4025/actascibiolsci.v34i1.6961

Sulfated polysaccharides (SPs) have attracted growing interest for various biotechnological applications. We evaluated the efficiency of two methods of drying SPs (M I and II) extracted from Halymenia sp Rhodophyceae in order to compare the yield, purification and anticoagulant activity. The total SPs (TSPs) were first extracted with papain in 100 mM sodium acetate (pH 5.0) containing cysteine and EDTA (5 mM). The TSPs obtained were dried in an oven (M I) or lyophilized (M II) and then examined by ion exchange chromatography (DEAE-cellulose) using the NaCl gradient technique. The fractions were analyzed by 0.5% agarose gel electrophoresis and the in vitro anticoagulant activity was evaluated by the activated partial thromboplastin time test using normal human plasma and compared to heparin (HEP) (193.00 IU mg-1). There was a difference in TSP yield of 19.05% and similar chromatographic SP profiles. Electrophoresis revealed fractions with distinct resolutions. The fractions eluted with 0.75 M of salt (M I and II) were the most active, measuring 27.40 and 72.66 IU mg-1, respectively, when compared to HEP. Therefore, obtaining SP with anticoagulant activity from Halymenia sp. is more efficient by freeze-drying. 

Natural heparinoids isolated from Halymenia sp. (Rhodophyceaes) delivery on the Ceará coast - doi: 10.4025/actascibiolsci.v32i3.6282 / Heparinoides naturais isolados de rodofíceas (Halymenia sp.) arribadas na costa cearense - doi: 10.4025/actascibiolsci.v32i3.6282

The increasing demand for heparin (HEP) has led to a search for alternative sources of natural anticoagulants. This study aimed to evaluate the anticoagulant activity of sulfated polysaccharides (SP) isolated from a Halymenia rhodophyceae genus native to the coast of Ceará, Brazil. Total SP were obtained by consecutive digestions with papain in 0.1 M sodium acetate buffer (pH 5.0) containing 5 mM cysteine and 5 mM EDTA, followed by ion-exchange chromatography on DEAE-cellulose column. The obtained fractions were concentrated by lyophilization and submitted to 0.5% agarose gel electrophoresis. Anticoagulant activity was evaluated by the activated partial thromboplastin time (APTT) using plasma from rabbits and a standard HEP (100 IU mg-1) curve. The extractions (53.96%) showed marked differences during the fractionation and in the degree of purification of SP. The species SP showed higher activity anticoagulant than that of HEP. However, the APTT of the fractions changed sharply among the extractions, expressing itself in a dose-dependent manner and increasing from 110.40 (1st extraction) to 143.10 IU mg-1 (3rd extraction). The results suggest that the anticoagulant activity of SP isolated from Halymenia sp. was promoted by inhibition of the intrinsic and/or common pathway of the coagulation cascade. The changes on APTT possibly will be elucidated through the mechanisms of actio

A crescente carência de heparina (HEP) motiva a busca por fontes alternativas de novos anticoagulantes naturais. Objetivou-se avaliar a atividade anticoagulante dos polissacarídeos sulfatados (PS) isolados de uma rodofícea do gênero Halymenia, nativa do litoral cearense, Brasil. Os PS totais foram obtidos por digestões consecutivas com papaína em tampão acetato de sódio 0,1 M (pH 5,0), contendo cisteína 5 mM e EDTA 5 mM, seguidas por cromatografia de troca iônica em coluna de DEAE-celulose. As frações obtidas foram concentradas por liofilização e submetidas à eletroforese em gel de agarose a 0,5%. Os ensaios anticoagulantes foram realizados pelo tempo de tromboplastina parcial ativada (TTPA), usando-se plasma de coelho e uma curva padrão de HEP (100 UI MG-1). As extrações (53,96%) mostraram diferenças marcantes durante o fracionamento e no grau de resolução dos PS. A espécie apresentou PS com atividade anticoagulante superior a HEP. O TTPA das frações modificou-se acentuadamente entre as extrações, expressando-se de maneira dose-dependente e sofrendo um acréscimo de 110,40 (1ª extração) para 143,10 UI MG-1 (3ª extração). Os resultados sugerem que a atividade anticoagulante dos PS isolados de Halymenia sp. foi promovida pela inibição da via intrínseca e/ou comum da cascata de coagulação. As modificações no TTPA possivelmente serão elucidadas pelos mecanismos de