A fucan from the brown seaweed Spatoglossum schröederi inhibits Chinese hamster ovary cell adhesion to several extracellular matrix proteins

Fucans, a family of sulfated polysaccharides present in brown seaweed, have several biological activities. Their use as drugs would offer the advantage of no potential risk of contamination with viruses or particles such as prions. A fucan prepared from Spatoglossum schröederi was tested as a possible inhibitor of cell-matrix interactions using wild-type Chinese hamster ovary cells (CHO-K1) and the mutant type deficient in xylosyltransferase (CHO-745). The effect of this polymer on adhesion properties with specific extracellular matrix components was studied using several matrix proteins as substrates for cell attachment. Treatment with the polymer inhibited the adhesion of fibronectin to both CHO-K1 (2 x 10(5))()and CHO-745 (2 x 10(5) and 5 x 10(5)) cells. No effect was detected with laminin, using the two cell types. On the other hand, adhesion to vitronectin was inhibited in CHO-K1 cells and adhesion to type I collagen was inhibited in CHO-745 cells. In spite of this inhibition, the fucan did not affect either cell proliferation or cell cycle. These results demonstrate that this polymer is a new anti-adhesive compound with potential pharmacological applications.

Efeito antitrombótico de uma fucana não anticoagulante extraída da alga Spatoglossum Schröederi

Fucan is a term used to denominate a family of sulfated L-fucose-rich polysaccharides. The brown alga Spatoglossum schröederi (Dictyotaceae) has three heterofucans namely fucan A, B and C. The 21 kDa fucan A is composed of a core of β (1-3) glucuronic acid-containing oligosaccharide of 4.5 kDa with branches at C4 of fucose chains α (1-3) linked. The fucose is mostly substituted at C4 with a sulfate group and at C2 with chains of β (1-4) xylose. This fucan has neither anticoagulant (from from 0.1 to 100µg) nor hemorrhagic activities (from 50 to 800 µg/mL). The antithrombotic test in vivo showed the fucan A has no activity in any of the concentrations (from 0.2 to 20µg/g/day) tested 1h after polysaccharide administration. However, when fucan A was injected endovenously 24h before the ligature of the venae cavae, we observed a dose-dependent effect, reaching saturation at around 20g/g of rat weight. In addition, this effect is also time-dependent, reaching saturation around 16h after fucan administration. In addition, regardless of administration pathway, fucan A displayed antithrombotic action. The exception was the oral pathway. Of particular importance was the finding that fucan A stimulates the synthesis of an antithrombotic heparan sulfate from endothelial cells like heparin. The hypothesis has been raised that in vivo antithrombotic activity of fucan A is related to the increased production this heparan. Taken together with the fact that the compound is practically devoid of anticoagulant and hemorrhagic activity suggests that it may be an ideal antithrombotic agent in vivo

Análise toxicológica in vitro e in vivo de uma fucana antitrombótica da alga marrom Spatoglossum schdöederi

Fucan is a term used to denominate a family of sulfated polysaccharides rich in L-fucose. They are extracted mainly from the extracellular matrix of brown algae and echinoderms. The brown alga Spatoglossum schröederi (Dictyotaceae) has three heterofucans named A, B and C. Our research group have been extracted non anticoagulant heterofucan from S. schröederi which possess antithrombotic activity in vivo. However, their toxicity in vitro and in vivo has not yet been determined. For the results in toxicity in vitro, we observed that the fucan A at 20, 500 and 1000 μg/plate showed no mutagenic activity in Kado test (Microsuspension), when the bacterial strains TA97a, TA98, TA100 and TA102, with and without S9 were used. The comet assay showed that fucan A (from 20 to 1000 μg/mL) did not cause any genotoxic effect on CHO cells. There was no damage to the DNA of these cells, as evidenced by the tail length and tail moment, which were similar to that found for the negative control. The fucan A from S. schröederi was administered at 20 μg/g of rat (dose which it showed high antithrombotic activity) during two months. After that, the animals were killed and examined. The data showed that fucan A did not cause any change in biochemistry and hematological parameters, as well as, in the morphology and size of the rat s organs analyzed. In conclusion, this study indicates that fucan is a compound with potential pharmacological that has no toxicity

Síntese e avaliação da atividade antitumoral de nanogéis de fucana A da alga marrom Spatoglossum schöederi (C. Agardh) Kützing

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Análise toxicológica in vitro e in vivo de uma fucana antitrombótica da alga marrom Spatoglossum schdöederi

Fucan is a term used to denominate a family of sulfated polysaccharides rich in L-fucose. They are extracted mainly from the extracellular matrix of brown algae and echinoderms. The brown alga Spatoglossum schröederi (Dictyotaceae) has three heterofucans named A, B and C. Our research group have been extracted non anticoagulant heterofucan from S. schröederi which possess antithrombotic activity in vivo. However, their toxicity in vitro and in vivo has not yet been determined. For the results in toxicity in vitro, we observed that the fucan A at 20, 500 and 1000 μg/plate showed no mutagenic activity in Kado test (Microsuspension), when the bacterial strains TA97a, TA98, TA100 and TA102, with and without S9 were used. The comet assay showed that fucan A (from 20 to 1000 μg/mL) did not cause any genotoxic effect on CHO cells. There was no damage to the DNA of these cells, as evidenced by the tail length and tail moment, which were similar to that found for the negative control. The fucan A from S. schröederi was administered at 20 μg/g of rat (dose which it showed high antithrombotic activity) during two months. After that, the animals were killed and examined. The data showed that fucan A did not cause any change in biochemistry and hematological parameters, as well as, in the morphology and size of the rat s organs analyzed. In conclusion, this study indicates that fucan is a compound with potential pharmacological that has no toxicity

Sulfated fucan as support for antibiotic immobilization

Xylofucoglucuronan from Spatoglossum schröederi algae was tested as a support for antibiotic immobilization. The polysaccharide (20 mg in 6 ml) was first activated using carbodiimide, 1-ethyl-3-(3-dimethylamino-propyl)carbodiimide methiodide (20 mg in 2 ml), under stirring for 1 h at 25ºC and pH from 4.5 to 5.0. After adjusting the pH to 8.0, either gentamicin or amikacin (62.5 mg in 1.25 ml) was then immobilized on this chemically modified polysaccharide with shaking for 24 h in a cold room. Infrared spectra of the activated carbodiimide xylofucoglucuronan showed two bands to carbonyl (C = O at 1647.9 and 1700.7 cm-1) and to amide (CÝ-NH2) groups (1662.8 and 1714.0 cm-1). Microbial characterization of the derivatives was carried out by the disk diffusion method using Staphylococcus aureus or Klebsiella pneumoniae incorporated in Müller Hinton medium. Inhibition halos of bacterial growth were observed for the antibiotics immobilized on this sulfated heteropolysaccharide before and after dialysis. However, the halos resulting from the samples after dialysis were much smaller, suggesting that dialysis removed either non-covalently bound antibiotic or other small molecules. In contrast, bacterial growth was not inhibited by either xylofucoglucuronan or its activated form or by gentamicin or amikacin after dialysis. An additional experiment was carried out which demonstrated that the sulfated heteropolysaccharide was hydrolyzed by the microorganism. Therefore, the antibiotic immobilized on xylofucoglucuronan can be proposed as a controlled drug delivery system. Furthermore, this sulfated heteropolysaccharide can be extracted easily from sea algae Spatoglossum schröederi.

Bioprospecção de polissacarídeos sulfatados de macroalgas marinhas do litoral do Rio Grande do Norte: caracterização de uma heterofucana extraída da alga marron sargassum filipendula que induz apoptose em células hela

The coast of Rio Grande do Norte has more than 100 species of seaweed, mostly unexplored regarding their pharmacological potential. The sulfated polysaccharides (PS) are by far the more seaweed compounds studied, these present a range of biological properties, such as anticoagulant activity, anti-inflammatory, antitumor and antioxidant properties. In this study, we extract sulfated polysaccharide rich-extracts of eleven algae from the coast of Rio Grande do Norte (Dictyota cervicornis; Dictiopterys delicatula; Dictyota menstruallis; Dictyota mertensis; Sargassum filipendula; Spatoglossum schröederi; Gracilaria caudata; Caulerpa cupresoides; Caulerpa prolifera; Caulerpa sertularioides e Codim isthmocladum), and these were evaluated for the potential anticoagulant, antioxidant and antiproliferative. All polysaccharide extracts showed activity for anticoagulant, antioxidant and/or antiproliferative activity, especially D. delicatula and S. filipendula, which showed the most prominent pharmacological potential, thereby being chosen to have their sulfated polysaccharides extracted. By fractionating method were obtained six fractions rich in sulfated polysaccharides to the algae D. delicatula (DD-0,5V, DD-0, 7V, DD-1,0v, DD-1,3v, DD-1,5v and DD-2,0) and five fractions to the alga S. filipendula (SF-0,5V, SF-0,7V, SF-1,0v, SF-1,5v and SF-2,0v). For the anticoagulant assay only the fractions of D. delicatula showed activity, with emphasis on DD-1, 5v that presented the most prominent activity, with APTT ratio similar to clexane® at 0.1 mg/mL. When evaluated the antioxidant potential, all fractions showed potential in all tests (total antioxidant capacity, hydroxyl and superoxide radicals scavenging, ferrous chelation and reducing power), however, the ability to chelate iron ions appears as the main mechanism antioxidant of sulfated polysaccharides from seaweed. In antiproliferative assay, all heterofucanas showed dose-dependent activity for the inhibition of cell proliferation of HeLa, however, with the exception of SF-0,7V, SF- 1,0v and SF-1,5v, all fractions showed antiproliferative activity against MC3T3, a normal cell line. The heterofucana SF-1,5V had its antiproliferative mechanism of action evaluated. This heterofucan induces apoptosis in HeLa cells by a pathway caspase independent, promoting the release of apoptosis Inducing Factor (AIF) in the cytosol, which in turn induces chromatin condensation and DNA fragmentation into 50Kb fragments. These results are significant in that they provide a mechanistic framework for further exploring the use of SF-1.5v as a novel chemotherapeutics against human cervical cancer.

Fucana ativa via das Map quinases e inibe a proliferação de células de ovário de hamster chinês (CHO)

Fucan is a term used to denominate L-fucose rich sulfated polysaccharides. The fucans have been studied due their pharmacological activities like antithrombotic, antiproliferative and antioxidant. We have extracted three fucan fractions from the brown seaweed Spatoglossum schröederi. These fucans were denominated Fuc B 1, Fuc B 1.5 and Fuc B 2. The chemical analyzes show that the fucans have very similar composition as demonstrated by agarose electrophoresis gel, sugar and sulfate content. The antiproliferative effect was determined by MTT and BrdU methodologies in CHO cells. The inhibition of proliferation effect of the three fractions was about 40%. Therefore this we proceed just with the Fuc B 2 due the higher yield. There is no apoptosis indication using the anexin V/propidium iodide test. We found a cell cycle phase G1 arrest. The western blotting show that the PKC; pFAK; pERK 1/2 are activated when the cells were treated with fucans. The treatement with inhibitor of MAPK PD98059 extinguished the fucan effect. These results indicates that fucan act by the ERK pathway inducing the cell death.

Bioprospecção de extratos metanólicos de macroalgas marinhas do litoral do Rio Grande do Norte: avaliação das atividades anticoagulante, antioxidante e antiproliferativa

Seaweeds are organisms known to exhibit a variety of biomolecules with pharmacological properties. The coast of Rio Grande do Norte has over 100 species of seaweeds, most of them not yet explored for their pharmacological potential. Sugars and phenolic compounds are the most studied of these being assigned a range of biological properties, such as anticoagulant , antiinflammatory, antitumor and antioxidant activities. In this work, we obtained methanolic extracts from thirteen seaweeds of the coast of Rio Grande do Norte (Dictyota cervicornis; Dictiopterys delicatula; Dictyota menstruallis; D. mertensis; Sargassum filipendula; Spatoglossum schröederi; Acanthophora specifera; Botryocladia occidentalis; Caulerpa cupresoides; C. racemosa; C. prolifera; C. sertularioides e Codium isthmocladum). They were evaluated as anticoagulant and antioxidant drugs, as well as antiproliferative drugs against the tumor cell line HeLa. None of the methanolic extracts showed anticoagulant activity, but when they were evaluated as antioxidant drugs all of extracts showed antioxidant activity in all tests performed (total antioxidant capacity, sequestration of superoxide and hydroxyl radicals, ferric chelation and reductase activity), especially the algae D. mentrualis, D. cilliolata and C. prolifera, who had the greatest potential to donate electrons.In addition, the ability of iron ions chelation appears as the main antioxidant mechanism of the methanolic extracts of these seaweeds mainly for the extract of the C. racemosa seaweed, which reached almost 100% activity. In the MTT assay, all extracts showed inhibitory activity at different levels againts HeLa cells. Moreover, D. cilliolata (MEDC) and D. menstrualis (MEDM) extracts showed specific activity to this cell line, not inhibiting the viability of 3T3 normal cell line, so they were chosen for detailing the antiproliferative mechanism of action. Using flow cytometry, fluorescence microscopy and in vitro assays we demonstrated that MEDC and MEDM induced apoptosis in HeLa cells by activation of caspases 3 and 9 and yet, MEDC induces cell cycle arrest in S phase. Together, these results showed that the methanolic extracts of brown seaweed D. menstrualis and D. cilliolata may contain agents with potential use in combatting cells from human uterine adenocarcinoma. This study also points to the need for more in-depth research on phytochemical and biological context to enable the purification of biologically active products of these extracts

Bioprospecção de polissacarídeos sulfatados de macroalgas marinhas do litoral do Rio Grande do Norte: caracterização de uma heterofucana extraída da alga marron Sargassum filipendula que induz apoptose em células HeLa

The coast of Rio Grande do Norte has more than 100 species of seaweed, mostly unexplored regarding their pharmacological potential. The sulfated polysaccharides (PS) are by far the more seaweed compounds studied, these present a range of biological properties, such as anticoagulant activity, anti-inflammatory, antitumor and antioxidant properties. In this study, we extract sulfated polysaccharide rich-extracts of eleven algae from the coast of Rio Grande do Norte (Dictyota cervicornis; Dictiopterys delicatula; Dictyota menstruallis; Dictyota mertensis; Sargassum filipendula; Spatoglossum schröederi; Gracilaria caudata; Caulerpa cupresoides; Caulerpa prolifera; Caulerpa sertularioides e Codim isthmocladum), and these were evaluated for the potential anticoagulant, antioxidant and antiproliferative. All polysaccharide extracts showed activity for anticoagulant, antioxidant and/or antiproliferative activity, especially D. delicatula and S. filipendula, which showed the most prominent pharmacological potential, thereby being chosen to have their sulfated polysaccharides extracted. By fractionating method were obtained six fractions rich in sulfated polysaccharides to the algae D. delicatula (DD-0,5V, DD-0, 7V, DD-1,0v, DD-1,3v, DD-1,5v and DD-2,0) and five fractions to the alga S. filipendula (SF-0,5V, SF-0,7V, SF-1,0v, SF-1,5v and SF-2,0v). For the anticoagulant assay only the fractions of D. delicatula showed activity, with emphasis on DD-1, 5v that presented the most prominent activity, with APTT ratio similar to clexane® at 0.1 mg/mL. When evaluated the antioxidant potential, all fractions showed potential in all tests (total antioxidant capacity, hydroxyl and superoxide radicals scavenging, ferrous chelation and reducing power), however, the ability to chelate iron ions appears as the main mechanism antioxidant of sulfated polysaccharides from seaweed. In antiproliferative assay, all heterofucanas showed dose-dependent activity for the inhibition of cell proliferation of HeLa, however, with the exception of SF-0,7V, SF- 1,0v and SF-1,5v, all fractions showed antiproliferative activity against MC3T3, a normal cell line. The heterofucana SF-1,5V had its antiproliferative mechanism of action evaluated. This heterofucan induces apoptosis in HeLa cells by a pathway caspase independent, promoting the release of apoptosis Inducing Factor (AIF) in the cytosol, which in turn induces chromatin condensation and DNA fragmentation into 50Kb fragments. These results are significant in that they provide a mechanistic framework for further exploring the use of SF-1.5v as a novel chemotherapeutics against human cervical cancer.

Fucana ativa via das Map quinases e inibe a proliferação de células de ovário de hamster chinês (CHO)

Fucan is a term used to denominate L-fucose rich sulfated polysaccharides. The fucans have been studied due their pharmacological activities like antithrombotic, antiproliferative and antioxidant. We have extracted three fucan fractions from the brown seaweed Spatoglossum schröederi. These fucans were denominated Fuc B 1, Fuc B 1.5 and Fuc B 2. The chemical analyzes show that the fucans have very similar composition as demonstrated by agarose electrophoresis gel, sugar and sulfate content. The antiproliferative effect was determined by MTT and BrdU methodologies in CHO cells. The inhibition of proliferation effect of the three fractions was about 40%. Therefore this we proceed just with the Fuc B 2 due the higher yield. There is no apoptosis indication using the anexin V/propidium iodide test. We found a cell cycle phase G1 arrest. The western blotting show that the PKC; pFAK; pERK 1/2 are activated when the cells were treated with fucans. The treatement with inhibitor of MAPK PD98059 extinguished the fucan effect. These results indicates that fucan act by the ERK pathway inducing the cell death.

Efeito antiinflamatório de fucana extraída da alga Parda spatoglossum schroederii em modelos experimentais de Peritonite, choque não séptico e colite

Fucans is a name used for sulfated polysaccharides, which is most characteristic structure of the presence of sulfated L-fucose, are found in brown seaweed (Phaeophyceae) and echinoderms (sea urchins and sea cucumbers). These polysaccharides have been reported to possess anticoagulant, antitumor, anti-viral, anti-proliferative and anti-inflammatory activities. Therefore, in the present study was evaluate the effect of the fucan from the brown seaweed Spatoglossum schroederii in models of peritonitis and non-septic shock induced by zymosan, as well as in a murine model of colitis induces by DSS. So, the mice treatment by intravenous route with the fucan was able to reduce the exudate formation and the cell migration in the model of acute peritonitis induced by zymosan during the kinetic of 6, 24 and 48 hours. Similarly, in the model of non-septic shock induced by zymosan the fucan demonstrated a protector effect to inhibited the cellular migration to the peritoneo, to decrease the levels of IL-6 in the serum and in the peritoneal exudate, to attenuate the lose of weight in the mice; beside to reduce the serum levels of hepatic transaminases and as well as the liver injury. In the model of murine colitis, the treatment with the fucan reduced the lose of weight of the animals, decreased the levels of IL-17 and IFN- produced in the gut and decrease the intestinal lesion induced by DSS. In conclusion, the fucan used in this study presented a significant protector effect in the murine models of inflammation

Contribuição para o conhecimento da flora algológica marinha do estado do Paraná

Esta nota é a primeira contribuição para a distribuição especial das algas marinhas do litoral paranaense (Caiobá). São considerados três tipos de litoral: a) Litoral rochoso sujeito à ação de arrebentação. b) Litoral rochoso mais ou menos abrigado. c) Manguesais. Dentro do esquema proposto, o primeiro tipo é subdividido em 2 zonas: 1.º) Zona dos borrifos, situada acima do limite médio da maré alta (fig. 2 n.º 1), correspondente à "Splash zone" ou à "Sprit zone" dos autores estrangeiros. 2.º) Zona de arrebentação (Ressaca), situada entre os níveis médios das marés baixa e alta, correspondendo à "Interdital zone" ou à "Gezeitenzone" dos autores estrangeiros, (fig. 2 n.º 2). A primeira zona só conta com uma alga, Lyngbya confervoides e pelo menos mais duas espécies de moluscos do gênero Littorina. A segunda zona, a mais rica tanto em algas como em animais, é essencialmente caracterizada em Caiobá pelas algas seguintes: Levringia sp. Chaetomorpha media e Centroceras clavulatum na parte mais alta, associadas a balanoides (craca) e a Mytilus perna (marisco). Mais abaixo domina Pterosiphonia pennata, Hypnea musciformis e Bryocladia thyrsigera. São apresentados também mais três tipos secundários dependendo das condições locais dos rochedos. Esta sucessão de Littorina, Balanus e Mytilus é a mesma existente no sul da África, segundo se depreende dos trabalhos de Stephenson (11,12) e de vários de seus colaboradores. O segundo tipo apresenta também duas zonas, sendo a segunda a mais rica e variada, aparecendo aqui como dominante, uma associação na qual, Callithamnion, Laurencia papulosa, Gigartina Teedii e Sargassum cymosum são as mais abundantes. É sugerida a existência de uma zona abaixo do limite inferior das marés baixas, zona essa representada por Rhodymenia Palmetta, Plocamium brasiliensis e Spatoglossum Schroederi pelo menos. É feita uma rápida enumeração das algas dos manguesais.

Avaliação da atividade antimalárica de extratos obtidos de algas marinhas no litoral do Rio Grande do Norte

Malaria is a major parasitic disease worldwide, accounting for about 500 million cases and causing 2 million to 3 million deaths annually. Four species are responsible for transmitting this disease to humans: Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae and Plasmodium ovale. The parasite resistance to antimalarial drugs and the usual limitations of the vector control implications are contributing to the spread of the disease. The most of significant advances in the search for new antimalarial drugs is based on natural components, the main ones being currently used antimalarial drugs derived from plants. Research on natural products of marine origin (particularly algae) show that some species possess antiplasmodial activity. Knowing that the coast of Rio Grande do Norte is home to several species of algae, the present study was to evaluate, for the first time, the antimalarial activity of ethanolic extracts of seaweed Spatoglossum schroederi, Gracilaria birdiae and Udotea flabellum against Plasmodium falciparum 3D7 strain tests and in vitro using the murine model (Plasmodium berghei) for evaluation in vivo. These species were ground, macerated with ethanol for 24 hours and the extracts concentrated in rotaevaporador (45 ° C ± 5 ° C). For in vitro tests, the extracts were diluted and tested at concentrations between 100 and 1.56 μg/ml (seven concentrations in triplicate), in order to obtain IC50 of each extract. The cytotoxicity tests with macrophages and BGM were performed using the MTT colorimetric assay. BGM macrophages and cells were distributed in 96 wells per plate (1x 105 to macrophages and 1x104 cells per well for BGM) and incubated for 24h at 37 ° C. The ethanol extracts were diluted and tested at concentrations of 100 to 1,56 μg/ml (seven concentrations in triplicate). After periods of 24 hours of incubation with the extracts, 100 μg of MTT was added to each well, and 3 hours elapsed, the supernatant was removed and added 200 μl of DMSO in each well. The absorbance of each well was obtained by reading on a spectrophotometer at 570 nm filter. To evaluate the acute toxicity in vivo, Swiss mice received a single dose (oral) 2000 mg/kg/animal of each extract tested. The parameters of acute toxicity were observed for 8 days. For in vivo tests, Swiss mice were inoculated with 1x105 erythrocytes infected with P. berghei. The treatment was given first to fourth day after infection with 0.2 ml of the extracts in doses of 1000 and 500 mg//g animal. The negative control group received 0.2 ml of 2% Tween-20, whereas the positive control group received sub-dose of chloroquine (5 mg/kg/animal). The assessment of antimalarial activity was done by suppressing suppressing the parasitemia at 5 and 7 days after infection. The growth inhibition of parasites was determined relative to negative control (% inhibition = parasitaemia in control - parasitemia in sample / parasitemia control x 100), the mortality of animals was monitored daily for 30 days The results showed that algae Spatoglossum schroederi and Udotea flabellum showed antimalarial activity in vitro, with reduced parasitemia of 70.54% and 54, respectively. The extracts of the three algae tested showed moderate to high cytotoxicity. Algae S. schroederi and U. flabellum were active against P. berghei only at doses of 500 mg / kg with reduction ranging from 54.58 to 52.65% for the fifth day and from 32.24 to 47.34% for the seventh day, respectively. No toxicity was observed in vivo at the dose tested, over the 8 days of observation. Although preliminary data, the bioactive components in those possible seaweed may be promising for the development of new anti-malarial drugs