Comparative study of sulfated polysaccharides from Caulerpa spp. (Chlorophyceae). Biotechnological tool for species identification? - doi: 10.4025/actascibiolsci.v34i4.8976

Studies on macromolecules isolated from marine algae suggested sulfated polysaccharides (SPs) as possible molecular markers for species. We evaluated isolated and fractionated SPs from the green marine algae Caulerpa cupressoides, C. prolifera and C. racemosa collected at Pacheco Beach, as possible taxonomic molecular indicators. Total SPs were extracted with papain in 100 mM sodium acetate buffer (pH 5.0) containing cysteine and EDTA (both 5 mM), followed by ion-exchange chromatography on DEAE-cellulose using a NaCl gradient. The obtained fractions were analyzed by 0.5% agarose gel electrophoresis. Anticoagulant assays employing normal human plasma and standard heparin (193 IU mg-1) by the activated partial thromboplastin time (APTT) test were also performed as comparison parameters. Low yields, and similar chromatographic profiles were found among species SPs, but electrophoresis revealed distinct SPs resolution patterns. The changes in APTT of SP fractions were dependent on charge density as showed by electrophoresis profiles. Activities were 17.37 (C. cupressoides), 22.17 (C. racemosa) and 25.64 (C. prolifera) IU mg-1, respectively, similar to a previous study using the first and second species. The results suggest that comparative studies of SPs isolated from seaweeds may be an important tool for the identification of Caulerpaceae.

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.

Glycosaminoglycans isolated from the skin of palombeta (Chloroscombrus chrysurus) and guaiúba (Ocyurus chrysurus): features and biological implications - doi: 10.4025/actascibiolsci.v34i2.6056 / Glicosaminoglicanos isolados da pele de palombeta (Chloroscombrus chrysurus) e guaiúba (Ocyurus chrysurus): características e implicações biológicas - doi: 10.4025/actascibiolsci.v34i2.6056

A great number of pharmacological compounds is found in the Brazilian marine diversity. This study evaluated the anticoagulant potential of glycosaminoglycans (GAGs) isolated from the skin of palombeta Chloroscombrus chrysurus and guaiúba Ocyurus chrysurus. GAGs were extracted with crude 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 chemical composition (contaminant proteins and total carbohydrates) and the analysis by 0.5% agarose gel electrophoresis of fractions were also determined. Anticoagulant assays were performed by activated partial thromboplastin time (APTT) using normal human plasma and standard heparin (193.00 IU mg-1). The obtaining and fractionation procedures of GAGs were effective and similar chromatographic profiles were verified between the species. A similar mobility to dermatan sulfate was revealed for C. chrysurus. This GAG also showed a low activity of 3.30 IU mg-1.

O Brasil abriga uma das maiores biodiversidades marinhas do mundo, favorecendo a descoberta de fontes alternativas de compostos farmacológicos. Desta forma, objetivou-se avaliar o potencial anticoagulante de glicosaminoglicanos (GAGs) isolados das peles da palombeta (Chloroscombrus chrysurus) e guaiúba (Ocyurus chrysurus). Os GAGs foram extraídos com papaína bruta em tampão acetato de sódio 0,1 M (pH 5,0) contendo cisteína 5 mM e EDTA 5 mM, seguido por cromatografia de troca iônica do extrato total em coluna de DEAE-celulose. As frações obtidas foram analisadas quanto à composição química (proteínas contaminantes e carboidratos totais) e os GAGs identificados por eletroforese em gel de agarose a 0,5%. Os ensaios de atividade anticoagulante foram realizados por meio do tempo de tromboplastina parcial ativada (TTPA) usando plasma humano normal e heparina-padrão (193,00 UI mg-1). O procedimento de obtenção e fracionamento dos GAGs mostrou-se eficiente, indicando semelhantes perfis cromatográficos entre as espécies avaliadas e, revelando para C. chrysurus, bandas com mobilidades semelhantes ao dermatam sulfato e com atividade de apenas 3,30 UI mg-1.

An antithrombin-dependent sulfated polysaccharide isolated from the green alga Caulerpa cupressoides has in vivo anti- and prothrombotic effects

Red algae sulfated polysaccharides (SPs) have been widely described as anticoagulant and antithrombotic agents; however no description of antithrombotic activity regarding green algae SPs has been reported. Caulerpa cupressoides (Chlorophyta) has three different SPs fractions (SP1, SP2 and SP3). We investigated the effects of SP2 on thrombin activity by antithrombin and in an experimental model of venous thrombosis in rats. The inhibition of thrombin assay was evaluated using antithrombin (AT) in the presence of SP2 and the antithrombotic activity was investigated in rats with thromboplastin as the thrombogenic stimulus. The anticoagulant effects of SP2 are suggested be due to the potentiation of thrombin inhibition by antithrombin (IC50 ~ 10.0µg mL-1) and this mechanism of interaction is different when compared to other studied Caulerpa polysaccharides. SP2 exhibited antithrombotic effects at doses of 1.0 and 2.0mg kg-1 body weight, but at higher doses (>2.0mg kg-1 body weight) this polysaccharide revert the antithrombotic property. No hemorrhagic effect (2.0mg kg-1) was observed. As occurs with red algae SPs, these results indicate that green algae SPs are also capable of exhibiting different in vivo properties.

Os polissacarídeos sulfatados (PSs) de algas vermelhas têm sido relatados mundialmente como agentes anticoagulantes e antitrombóticos. Entretanto, nenhuma descrição de atividade antitrombótica tem sido relacionada com os PSs de algas verdes. A clorofícea Caulerpa cupressoides possui três frações de PSs (PS1; PS2 e PS3). Dessa forma, objetivou-se investigar os efeitos da fração PS2 sobre a atividade da trombina por antitrombina e usando um modelo experimental de trombose venosa em ratos. O ensaio de inibição da trombina foi avaliado usando a antitrombina (AT) na presença de PS2 e a atividade antitrombótica foi investigada em ratos, usando a tromboplastina como o estímulo trombogênico. Os efeitos anticoagulantes de PS2 devem-se provavelmente à sua potência de inibir a trombina mediada pela AT (IC50 ~ 10,0µg mL-1) e esse mecanismo de interação é diferente, comparado ao de outros polissacarídeos de Caulerpa estudados. PS2 exibiu efeitos antitrombóticos nas doses de 1,0 e 2,0mg kg-1 peso corpóreo, mas em doses mais elevadas (>2,0mg kg-1 peso corpóreo) esse polissacarídeo exibe efeitos pró-trombóticos. Também não foi observado nenhum efeito hemorrágico (2,0mg kg-1). Assim como ocorre com os PSs de algas vermelhas, os resultados indicam que os PSs de algas verdes também possuem atividades biológicas distintas in vivo.

An antithrombin-dependent sulfated polysaccharide isolated from the green alga Caulerpa cupressoides has in vivo anti- and prothrombotic effects

Red algae sulfated polysaccharides (SPs) have been widely described as anticoagulant and antithrombotic agents; however no description of antithrombotic activity regarding green algae SPs has been reported. Caulerpa cupressoides (Chlorophyta) has three different SPs fractions (SP1, SP2 and SP3). We investigated the effects of SP2 on thrombin activity by antithrombin and in an experimental model of venous thrombosis in rats. The inhibition of thrombin assay was evaluated using antithrombin (AT) in the presence of SP2 and the antithrombotic activity was investigated in rats with thromboplastin as the thrombogenic stimulus. The anticoagulant effects of SP2 are suggested be due to the potentiation of thrombin inhibition by antithrombin (IC50 ~ 10.0µg mL-1) and this mechanism of interaction is different when compared to other studied Caulerpa polysaccharides. SP2 exhibited antithrombotic effects at doses of 1.0 and 2.0mg kg-1 body weight, but at higher doses (>2.0mg kg-1 body weight) this polysaccharide revert the antithrombotic property. No hemorrhagic effect (2.0mg kg-1) was observed. As occurs with red algae SPs, these results indicate that green algae SPs are also capable of exhibiting different in vivo properties.

Os polissacarídeos sulfatados (PSs) de algas vermelhas têm sido relatados mundialmente como agentes anticoagulantes e antitrombóticos. Entretanto, nenhuma descrição de atividade antitrombótica tem sido relacionada com os PSs de algas verdes. A clorofícea Caulerpa cupressoides possui três frações de PSs (PS1; PS2 e PS3). Dessa forma, objetivou-se investigar os efeitos da fração PS2 sobre a atividade da trombina por antitrombina e usando um modelo experimental de trombose venosa em ratos. O ensaio de inibição da trombina foi avaliado usando a antitrombina (AT) na presença de PS2 e a atividade antitrombótica foi investigada em ratos, usando a tromboplastina como o estímulo trombogênico. Os efeitos anticoagulantes de PS2 devem-se provavelmente à sua potência de inibir a trombina mediada pela AT (IC50 ~ 10,0µg mL-1) e esse mecanismo de interação é diferente, comparado ao de outros polissacarídeos de Caulerpa estudados. PS2 exibiu efeitos antitrombóticos nas doses de 1,0 e 2,0mg kg-1 peso corpóreo, mas em doses mais elevadas (>2,0mg kg-1 peso corpóreo) esse polissacarídeo exibe efeitos pró-trombóticos. Também não foi observado nenhum efeito hemorrágico (2,0mg kg-1). Assim como ocorre com os PSs de algas vermelhas, os resultados indicam que os PSs de algas verdes também possuem atividades biológicas distintas in vivo.

Isolamento, fracionamento e avaliação toxicológica in vivo de polissacarídeos sulfatados de Hypnea musciformis

This study aimed to isolate, fractionate and evaluate the in vivo toxicity of sulfated polysaccharides (SPs) from Hypnea musciformis (Rhodophyta), when obtained by three extraction methods (M I, M II and M III). SPs were extracted with papain in 100mM sodium acetate (pH 5.0) containing cysteine and EDTA (5mM) (M I) or water (25-80°C (M II), 80°C (M III)), and then their chemical composition of total carbohydrates, free sulfate (FS) and contaminant proteins (CPs) was determined. SPs were submitted to ion-exchange chromatography (DEAE-celulose) using a sodium chloride gradient, being the degree of homogeneity and charge density evaluated by agarose gel electrophoresis of the fractions obtained and compared to heparin. The in vivo assay was performed using groups (n=6) of male and female Swiss mice (24-33g), which received: SPs (9mg kg-1, i.p.) absence of CPs (M I) and 0.9% saline (0.1mL 10g-1, i.p.), for 14 consecutive days. On the 15th day, collect blood and organs for biochemical dosages and corporal mass correlation, respectively, from the animals anesthetized and sacrificed were performed. The sulfate content of FS (31.05±0.53%) (P 0.05) and the fractionation by DEAE-cellulose showed M I more effectiveness in obtaining SPs compared to M II and M III. The animals were tolerable to SPs from M I, and it wasn't observed hepatic or renal alteration (P>0.05)

Objetivou-se isolar, fracionar e avaliar a toxicidade in vivo dos polissacarídeos sulfatados (PSs) da rodofícea Hypnea musciformis, quando obtidos por três métodos de extração (M I; M II e M III). Os PSs foram extraídos com papaína em tampão acetato de sódio 100mM (pH 5,0), contendo cisteína e EDTA (5mM) (M I) ou água (25-80°C (M II); 80°C (M III)) e, em seguida, determinados sua composição química de carboidratos totais, sulfato livre (SL) e proteínas contaminantes (PCs). Os PSs foram submetidos à cromatografia de troca iônica (DEAE-celulose) usando um gradiente de cloreto de sódio, sendo avaliado o grau de homogeneidade e densidade de carga por eletroforese em gel de agarose das frações obtidas e comparadas à heparina. O ensaio in vivo foi realizado em grupos (n=6) de camundongos Swiss machos e fêmeas (24-33g), os quais receberam: PSs (9mg kg-1; i.p.) isentos do PCs (M I) e salina 0,9% (0,1mL 10g-1; i.p.), durante 14 dias consecutivos. No 15o dia, os animais foram anestesiados e sacrificados para coletas de sangue e órgãos, os quais foram utilizados para dosagens bioquímicas e correlações com suas massas corpóreas, respectivamente. O teor de SL (31,05±0,53%) (P 0,05) e o fracionamento, em DEAE-celulose, indicaram o M I mais eficiente na obtenção de PSs, comparado ao M II e M III. Os animais mostraram-se tolerantes aos PSs do M I e não se observou alteração de ordem hepática ou renal (P>0,05).

Isolamento, fracionamento e avaliação toxicológica in vivo de polissacarídeos sulfatados de Hypnea musciformis

This study aimed to isolate, fractionate and evaluate the in vivo toxicity of sulfated polysaccharides (SPs) from Hypnea musciformis (Rhodophyta), when obtained by three extraction methods (M I, M II and M III). SPs were extracted with papain in 100mM sodium acetate (pH 5.0) containing cysteine and EDTA (5mM) (M I) or water (25-80°C (M II), 80°C (M III)), and then their chemical composition of total carbohydrates, free sulfate (FS) and contaminant proteins (CPs) was determined. SPs were submitted to ion-exchange chromatography (DEAE-celulose) using a sodium chloride gradient, being the degree of homogeneity and charge density evaluated by agarose gel electrophoresis of the fractions obtained and compared to heparin. The in vivo assay was performed using groups (n=6) of male and female Swiss mice (24-33g), which received: SPs (9mg kg-1, i.p.) absence of CPs (M I) and 0.9% saline (0.1mL 10g-1, i.p.), for 14 consecutive days. On the 15th day, collect blood and organs for biochemical dosages and corporal mass correlation, respectively, from the animals anesthetized and sacrificed were performed. The sulfate content of FS (31.05±0.53%) (P 0.05) and the fractionation by DEAE-cellulose showed M I more effectiveness in obtaining SPs compared to M II and M III. The animals were tolerable to SPs from M I, and it wasn't observed hepatic or renal alteration (P>0.05)

Objetivou-se isolar, fracionar e avaliar a toxicidade in vivo dos polissacarídeos sulfatados (PSs) da rodofícea Hypnea musciformis, quando obtidos por três métodos de extração (M I; M II e M III). Os PSs foram extraídos com papaína em tampão acetato de sódio 100mM (pH 5,0), contendo cisteína e EDTA (5mM) (M I) ou água (25-80°C (M II); 80°C (M III)) e, em seguida, determinados sua composição química de carboidratos totais, sulfato livre (SL) e proteínas contaminantes (PCs). Os PSs foram submetidos à cromatografia de troca iônica (DEAE-celulose) usando um gradiente de cloreto de sódio, sendo avaliado o grau de homogeneidade e densidade de carga por eletroforese em gel de agarose das frações obtidas e comparadas à heparina. O ensaio in vivo foi realizado em grupos (n=6) de camundongos Swiss machos e fêmeas (24-33g), os quais receberam: PSs (9mg kg-1; i.p.) isentos do PCs (M I) e salina 0,9% (0,1mL 10g-1; i.p.), durante 14 dias consecutivos. No 15o dia, os animais foram anestesiados e sacrificados para coletas de sangue e órgãos, os quais foram utilizados para dosagens bioquímicas e correlações com suas massas corpóreas, respectivamente. O teor de SL (31,05±0,53%) (P 0,05) e o fracionamento, em DEAE-celulose, indicaram o M I mais eficiente na obtenção de PSs, comparado ao M II e M III. Os animais mostraram-se tolerantes aos PSs do M I e não se observou alteração de ordem hepática ou renal (P>0,05).

Isolamento, fracionamento e avaliação toxicológica in vivo de polissacarídeos sulfatados de Hypnea musciformis

This study aimed to isolate, fractionate and evaluate the in vivo toxicity of sulfated polysaccharides (SPs) from Hypnea musciformis (Rhodophyta), when obtained by three extraction methods (M I, M II and M III). SPs were extracted with papain in 100mM sodium acetate (pH 5.0) containing cysteine and EDTA (5mM) (M I) or water (25-80°C (M II), 80°C (M III)), and then their chemical composition of total carbohydrates, free sulfate (FS) and contaminant proteins (CPs) was determined. SPs were submitted to ion-exchange chromatography (DEAE-celulose) using a sodium chloride gradient, being the degree of homogeneity and charge density evaluated by agarose gel electrophoresis of the fractions obtained and compared to heparin. The in vivo assay was performed using groups (n=6) of male and female Swiss mice (24-33g), which received: SPs (9mg kg-1, i.p.) absence of CPs (M I) and 0.9% saline (0.1mL 10g-1, i.p.), for 14 consecutive days. On the 15th day, collect blood and organs for biochemical dosages and corporal mass correlation, respectively, from the animals anesthetized and sacrificed were performed. The sulfate content of FS (31.05±0.53%) (P 0.05) and the fractionation by DEAE-cellulose showed M I more effectiveness in obtaining SPs compared to M II and M III. The animals were tolerable to SPs from M I, and it wasn't observed hepatic or renal alteration (P>0.05)

Objetivou-se isolar, fracionar e avaliar a toxicidade in vivo dos polissacarídeos sulfatados (PSs) da rodofícea Hypnea musciformis, quando obtidos por três métodos de extração (M I; M II e M III). Os PSs foram extraídos com papaína em tampão acetato de sódio 100mM (pH 5,0), contendo cisteína e EDTA (5mM) (M I) ou água (25-80°C (M II); 80°C (M III)) e, em seguida, determinados sua composição química de carboidratos totais, sulfato livre (SL) e proteínas contaminantes (PCs). Os PSs foram submetidos à cromatografia de troca iônica (DEAE-celulose) usando um gradiente de cloreto de sódio, sendo avaliado o grau de homogeneidade e densidade de carga por eletroforese em gel de agarose das frações obtidas e comparadas à heparina. O ensaio in vivo foi realizado em grupos (n=6) de camundongos Swiss machos e fêmeas (24-33g), os quais receberam: PSs (9mg kg-1; i.p.) isentos do PCs (M I) e salina 0,9% (0,1mL 10g-1; i.p.), durante 14 dias consecutivos. No 15o dia, os animais foram anestesiados e sacrificados para coletas de sangue e órgãos, os quais foram utilizados para dosagens bioquímicas e correlações com suas massas corpóreas, respectivamente. O teor de SL (31,05±0,53%) (P 0,05) e o fracionamento, em DEAE-celulose, indicaram o M I mais eficiente na obtenção de PSs, comparado ao M II e M III. Os animais mostraram-se tolerantes aos PSs do M I e não se observou alteração de ordem hepática ou renal (P>0,05).

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.

Comparative study of sulfated polysaccharides from Caulerpa spp. (Chlorophyceae). Biotechnological tool for species identification? - doi: 10.4025/actascibiolsci.v34i4.8976

Studies on macromolecules isolated from marine algae suggested sulfated polysaccharides (SPs) as possible molecular markers for species. We evaluated isolated and fractionated SPs from the green marine algae Caulerpa cupressoides, C. prolifera and C. racemosa collected at Pacheco Beach, as possible taxonomic molecular indicators. Total SPs were extracted with papain in 100 mM sodium acetate buffer (pH 5.0) containing cysteine and EDTA (both 5 mM), followed by ion-exchange chromatography on DEAE-cellulose using a NaCl gradient. The obtained fractions were analyzed by 0.5% agarose gel electrophoresis. Anticoagulant assays employing normal human plasma and standard heparin (193 IU mg-1) by the activated partial thromboplastin time (APTT) test were also performed as comparison parameters. Low yields, and similar chromatographic profiles were found among species SPs, but electrophoresis revealed distinct SPs resolution patterns. The changes in APTT of SP fractions were dependent on charge density as showed by electrophoresis profiles. Activities were 17.37 (C. cupressoides), 22.17 (C. racemosa) and 25.64 (C. prolifera) IU mg-1, respectively, similar to a previous study using the first and second species. The results suggest that comparative studies of SPs isolated from seaweeds may be an important tool for the identification of Caulerpaceae.  

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. 

Glycosaminoglycans isolated from the skin of palombeta (Chloroscombrus chrysurus) and guaiúba (Ocyurus chrysurus): features and biological implications - doi: 10.4025/actascibiolsci.v34i2.6056 / Glicosaminoglicanos isolados da pele de palombeta (Chloroscombrus chrysurus) e guaiúba (Ocyurus chrysurus): características e implicações biológicas - doi: 10.4025/actascibiolsci.v34i2.6056

A great number of pharmacological compounds is found in the Brazilian marine diversity. This study evaluated the anticoagulant potential of glycosaminoglycans (GAGs) isolated from the skin of palombeta Chloroscombrus chrysurus and guaiúba Ocyurus chrysurus. GAGs were extracted with crude 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 chemical composition (contaminant proteins and total carbohydrates) and the analysis by 0.5% agarose gel electrophoresis of fractions were also determined. Anticoagulant assays were performed by activated partial thromboplastin time (APTT) using normal human plasma and standard heparin (193.00 IU mg-1). The obtaining and fractionation procedures of GAGs were effective and similar chromatographic profiles were verified between the species. A similar mobility to dermatan sulfate was revealed for C. chrysurus. This GAG also showed a low activity of 3.30 IU mg-1.

O Brasil abriga uma das maiores biodiversidades marinhas do mundo, favorecendo a descoberta de fontes alternativas de compostos farmacológicos. Desta forma, objetivou-se avaliar o potencial anticoagulante de glicosaminoglicanos (GAGs) isolados das peles da palombeta (Chloroscombrus chrysurus) e guaiúba (Ocyurus chrysurus). Os GAGs foram extraídos com papaína bruta em tampão acetato de sódio 0,1 M (pH 5,0) contendo cisteína 5 mM e EDTA 5 mM, seguido por cromatografia de troca iônica do extrato total em coluna de DEAE-celulose. As frações obtidas foram analisadas quanto à composição química (proteínas contaminantes e carboidratos totais) e os GAGs identificados por eletroforese em gel de agarose a 0,5%. Os ensaios de atividade anticoagulante foram realizados por meio do tempo de tromboplastina parcial ativada (TTPA) usando plasma humano normal e heparina-padrão (193,00 UI mg-1). O procedimento de obtenção e fracionamento dos GAGs mostrou-se eficiente, indicando semelhantes perfis cromatográficos entre as espécies avaliadas e, revelando para C. chrysurus, bandas com mobilidades semelhantes ao dermatam sulfato e com atividade de apenas 3,30 UI mg-1.