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

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

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Avaliação das atividades antinociceptiva e anti-inflamatória de uma heterofucana da alga Dictyota menstrualis

Fucana é um termo que define uma família de hetero e homopolissacarídeos que contem L-fucose em sua estrutura. Neste trabalho uma hererofucan F 2,0v da alga Dictyota menstrualis foi avaliada como agente antinociceptivo e antiinflamatório. A fucana F 2,0v inibiu a migração de leucócitos em até 100% (20.0 mg/kg) para a cavidade peritoneal após estimulo químico, porém, não alterou a expressão das interleucinas IL-1β, IL-6 e de TNF-α. Com relação a sensação dolorosa a F 2,0v (20.0 mg/kg) possui atividade antinociceptiva periférica com potência semelhante à dipirona. Por outro lado não apresentou efeito no teste da placa quente. Análises de microscopia confocal e citometria de fluxo mostram que a F 2,0v se liga a superfície dos leucócitos. O conjunto de resultados apresentados pela fucana F 2,0v sugerem que o mecanismo de ação está relacionado com sua capacidade de inibir a migração de leucócitos para o local da injúria tecidual. Em resumo os dados mostram que F 2,0v apresenta grande potencial como composto antinociceptivo e antiinflamatório. Estudos futuros serão realizados para caracterizar melhor o mecanismo de ação da F 2,0v

Atividade anticoagulante, antioxidante e antitumoral de heterofucanas da alga Dictyopteris delicatula (Lamouroux, 1809)

In the present study, six families of sulfated polysaccharides were obtained from seaweed Dictyopteris delicatula (Lamouroux, 1809) and their anticoagulant, antioxidant and antitumor activities were evaluated. All fractions showed anticoagulant activity on aPTT assay, but not on PT assay. Fractions also exhibited total antioxidant activity, superoxide radical scavenging capacity and ferric chelating property. Thus, six fractions (F0.5v, F0.7v, F1.0v, F1.3v, F1.5v e F2.0v) we obtained by proteolytic digestion, followed by acetone fractionation and molecular sieving on Sephadex G-100. Chemical analyses demonstrated that all polysaccharides contain heterofucans composed mainly of fucose, xylose, glucose, galactose, uronic acid, and sulfate. Any fractions changed the PT. However, all fractions were able on double the aPPT on a dose-dependent manner. The heterofucans F0.7v and F1.0v showed low anticoagulant activity while F1.5v presented the most prominent anticoagulant activity .When compared to Clexane®, a low molecular weight heparin, at same concentration F1.5v presented similar anticoagulant activity. The fucans F0.5v and F0.7v at 1.0 mg/mL showed high ferric chelating activity (~45%), whereas fucans F1.3v (0.5 mg/mL) showed considerable reducing power, about 53.2% of the activity of vitamin C. The fucan F1.5v presented the most prominent anticoagulant activity. The best antiproliferative activity was found with fucans F1.3v and F0.7v. However, F1.3v activity was much higher than F0.7v inhibiting almost 100% of HeLa cell proliferation. These fucans have been selected for further studies on structural characterization as well as in vivo experiments, which are already in progress

Efeito de extratos da alga marinha caulerpa mexicana em modelos de inflamação em murinos

The regulation of the inflammatory response is essential to maintain homeostasis. Several studies have been performed to search new drugs that can contribute to avoiding or minimizing an excessive inflammatory process. The aim of this study was to evaluate the effect of extracts of green algae Caulerpa mexican in models of inflammation. In mice, the model of peritonitis induced inflammatory zymosan pretreatment of mice with aqueous and methanol extracts of C. mexican was able to reduce cell migration to the peritoneal cavity. Treatment of mice with extracts of C. mexican also reduced the ear edema induced by xylene and exerted inhibitory action on the migration of leukocytes in inflammation-induced zymosan the air pouch, and timedependent for the extracts tested in the model of ulcerative colitis induced by DSS 3%, the extract methanol, but not the aqueous C. mexican, significantly reduced the clinical symptoms of colitis, as well as the production of proinflammatory cytokines in the culture of mouse colon, in the histological analysis there was a slight reduction of inflammation in the intestinal mucosa. We concluded that the administration of the extracts resulted in the reduction of cell migration to different sites as well as reducing the edema formation induced by chemical irritant. This study demonstrates for the first time the antiinflammatory effect of aqueous and methanolic extracts from green marine algae Caulerpa mexican

Processo de desidratação da alga Gracilaria birdiae beneficiada em laboratório e artesanalmente: análise da atividade antioxidante e composição centesimal

The main aim of this study was to compare the procedure for dehydration of Gracilaria birdiae prepared handmade and laboratory, collected in the northern coast of Rio Grande do Norte. The sample was collected in the Rio do Fogo beach in march 2009. The sample collected followed by two processing, the first the material prepared in laboratory was air-dried at 50°C for 24 hours in air-flow oven. The second the handmade sample was air-dried on the sun during three days. The extract was prepared in three different solvents: ethanol, hydroethanol and water, resulting in ethanol, hidroethanol and aqueous extracts from handmade and laboratory sample. In according with results only the ethanol extract was fractionated yielding the fractions hexane, dichloromethane and ethyl acetate fractions. The different process to obtain Gracilaria birdiae resulted in the samples with different shades. The soluble solids content was higher in the laboratory sample. The chemical composition the both samples were characterized by presenting a considerable amounts of carbohydrates, with amior percentage protein and ash, respectively, in the handmade and laboratory sample. In two samples showed a low content of lipids and the lipid profile showed a higher proportion of monounsaturated fatty acids, with the absence polyunsaturated handmade sample. The phytochemical screening by chemical reactions showed the presence of flavonoids, tannins, alkaloids and saponins the laboratory sample, presenting a greater diversity of bioactive compounds. Through of the analysis by thin layer chromatography was possible to identify the phytosterols β-sitosterol and stigmasterol the both samples, also suggest the presence of β-carotene and chlorophyll α the laboratory sample. The levels of total phenolics and flavonoids were more significant in the ethanol extract of the laboratory sample. The in vitro lethality showed that extracts of the laboratory sample and handmade from 125 to 500 μg/ mL, respectively, were highly lethal. In the evaluation of antioxidant capacity by the system β-carotene/ácido linoleic method and by DPPH radical scavernging assay, the ethanol extract from the laboratory process showed significantly greater activity than the other extracts, being and the first and second methods, respectively, lower and equivalent to the synthetic antioxidant BHT. The handmade ethanol extract has not demonstrated skill in deactivating free radicals, but showed activity in inhibiting lipid peroxidation, although the values were significantly lower than the laboratory sample. We conclude that the dehydration process in the laboratory is the most efficient technique to maintenance of the chemical composition present in the seaweed, providing beneficial properties such as antioxidant capacity. We emphasize that this property can be explored with the objective of adding commercial value to the final product, which will promote the expansion of production of this seaweed in the community of Rio do Fogo

Biossorção utilizando alga marinha (sargassum sp.) aplicada em meio orgânico

In this work, biosorption process was used to remove heavy metals from used automotive lubricating oils by a bus fleet from Natal-RN-Brazil. This oil was characterized to determine the physical-chemistry properties. It was also characterized the used oil with the aim of determining and quantifying the heavy metal concentration. Fe and Cu were the metals existent in large concentration and these metals were choused to be studied in solubilization process. For the biosorption process was used the seaweed Sargassum sp for the study of influencing of the metals presents separately and with other metals. It was also studied the effect of the protonation treatment of alga with the objective to know the best efficiency of heavy metals removal. The study of the solubilization showed that the presence of more than a metal favors the solubilization of the metals presents in the oil and consequently, it favors the biosorption process, what becomes interesting the perspective application in the heavy metals removal in lubricating oils used, because the presence of more than a heavy metal favors the solubility of all metals present. It was observed that the iron and copper metals, which are present in large concentration, the protonated biosorbtent was more effective. In this study we used as biomass the marine alga Sargassum sp to study the influence of agitation velocity, temperature and initial biomass concentration on the removal of iron and copper from used lubricant oils. We performed an experimental design and a kinetic study. The experiments were carried out with samples of used lubricant oil and predetermined amounts of algae, allowing sufficient time for the mixture to obtain equilibrium under controlled conditions. The results showed that, under the conditions studied, the larger the amount of biomass present, the lower the adsorption capacity of the iron and of the copper, likely due to a decrease in interface contact area. The experimental design led us to conclude that a function can be obtained that shows the degree of influence of each one of the system variables

Atividade antiinflamatória de uma heterofucana da alga marrom Padina gymnospora

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 μg/mL of the fraction was used. This study suggests the use of fraction F1.5 (heterofucan) as an anti-inflammatory

Avaliação de uma fração polissacaridica da alga Lobophora variegata (Lamouroux) em modelo de artrite induzida em ratos por Zymosan

Fucans seaweed Lobophora variegata estructures are known for their chemical and biological properties. In this study, we analyzed, the action of fucans L. variegata and the fractions purified with acetone in Zymosan-induced arthritis. After differential fractionation with acetone, six fractions were obtained and named F0.3, F0.5, F0.8, F1, F1.5 and F2. The results showed that the fraction F1 showed high yield (51.9%) and was chosen for studies of antioxidant activity and induced arthritis. Nuclear magnetic resonance (NMR) of 13C showed signals at 103.3 and 15.78 ppm that are assigned to links β13 galactose and of the C6 methyl fucose, respectively. The infrared (IR) showed absorbance at 1238 and 850 cm-1 which are attributed to sulfate. The fraction F1 showed antioxidant activities in vitro. For analysis of inflammatory parameters chosen the polysaccharide was administered in different doses (25, 50 and 75 mg / kg ip, per body weight) and diclofenac sodium (5 mg / kg ip) and L-NAME (25 mg / kg ip) in groups of animals (n = 6). After 6 h, were analyzed for cellular influx and levels of nitrite. In experiment five days, were made analysis of swelling and serum TNF-α. Histopathological analysis were performed for confirmation of results. The fraction F1 (25, 50 and 75 mg / kg ip) reduced the cellular influx (52.1 to 96.7%) and nitric oxide levels (27.2 - 39%) compared to control group. The reduction of edema (63.4 - 100%) and serum TNF-α (p <0.001) were observed when the polysaccharide F1 administered at a dose (50 mg / kg) These results suggest that these heterofucanas of Lobophora variegata have besides the activity antioxidant and potential anti-inflammatory activity in arthritis induced by zymosan

Purificação e caracterização estrutural de homogalactanas sulfatadas anticoagulantes da alga verde Codium isthmocladum

Conselho Nacional de Desenvolvimento Científico e Tecnológico

Propriedades bioativas dos polissacarídeos sulfatados da alga comestível Gracilaria birdiae

Red marine algae of the genus Gracilaria synthesize sulfated polysaccharides (PS) bioactive. But many of these PS were not properly assessed, as is the case of PS synthesized by edible seaweed Gracilaria birdiae. Previous studies showed that sulfated galactans this alga has anti-inflammatory effect. In this work, a galactan (GB) of G. birdiae was obtained and evaluated by different tests. GB showed anticoagulant activity in APTT assay. GB showed no toxicity to normal cells (3T3), but inhibited the survival of cells of adenocarcinoma of the cervix (HeLa) and human pancreatic cancer (Panc-1) 80% (1.5 mg / ml). GB was not able to hijack the OH radical or the superoxide radical. However, showed activity electron donor in two different tests and presented iron chelator activity (70% and 1.0 mg / ml) and Copper (70% at 0.5 mg / ml). The presence of a higher GB promotes formation of crystals of calcium oxalate dihydrate small size, which is less aggressive, because GB is able to interact with and stabilize the crystal that form. Furthermore, GB (2.0 mg / mL) was not cytotoxic to human renal cells (HEK-293). The data lead us to propose that GB has a great potential for the treatment of urolithiasis

Otimização da extração de polissacarídeos sulfatados da alga vermelha Gracilaria birdiae e análise da atividade anticoagulante e antioxidante

The sulfated polysaccharides (SP) from the edible red seaweed Gracilaria birdiae were obtained using five different condition extraction (GB1: Water; GB1p: Water/proteolysis; GB1s: Water/sonication; GB1sp: Water/sonication/proteolysis; GB2s: NaOH/sonication; GB2sp: NaOH/sonication/proteolysis. The yield (g) increased in the following order GB2sp>GB1sp>GB1p>GB2s>GB1s>GB1. However, the amount of SP extracted increased in different way GB2sp>GB1p>GB1>GB1sp>GB1s>GB2s. Infrared and electrophoresis analysis showed that all conditions extracted the same SP. In addition, monosaccharide composition showed that ultrasound promotes the extraction of other polysaccharides than SP. In the prothrombin time (PT) test, which evaluates the extrinsic coagulation pathway, none of the samples showed anticoagulant activity. While in the activated partial thromboplastin time (aPTT) test, which evaluates the intrinsic coagulation pathway, all samples showed anticoagulant activity, except GB2s. The aPTT activity decreased in the order of GB1sp>GB2sp>GB1p>GB1>GB1s>GB2s. Total capacity antioxidant (TCA) of the SP was also affected by condition extraction, since GB2s and GB1 showed lower activity in comparison to the other conditions. In conclusion, the conditions of SP extraction influence their biological activities and chemical composition. The data showed NaOH/sonication/proteolysis was the best condition to extract anticoagulant and antioxidant SPs from Gracilaria birdiae.

Caracterização estrutural e atividades farmacológicas do alginato obtido da alga Dictyopteris delicatula (J.V. Lamouroux., 1809) e seu derivado sulfatado

Marine algae are rich sources of various structural compounds which recently has been increasingly studied as a new source of bioactive substances. The alginate, as come as fucans, are considered the main acidic polysaccharides found in brown seaweed. This molecule consists a linear natural polysaccharide, non-sulfated, and presents monosaccharides: acid β-D-mannuronic (M) and α-L-guluronic acid (G); in a vast amount compositions and threads. Alginate has been widely applied in food and pharmaceutical industries because of its ability to retain water, forming films and gels as well as thickening, stabilizing and form emulsions. In this work we aimed to extract, structurally characterize, compare and analyze the possible pharmacological activities of native alginate molecule obtained from brown seaweed Dyctiopteris delicatula (DYN), and its chemically sulfated derivative (DYS). The alginate structure and composition molecule can be proven through chemical dosing, that showed low protein contamination and high sugar level, existence and separation of M and G blocks in the descending paper chromatography, infrared spectroscopy and nuclear magnetic resonance. Molecule sulfation was proven with sulphate dosage, resulting in 28.56% sulphate in molecule; electrophoresis, verify metachromasia with toluidine blue; and infrared spectroscopy, that showed a characteristic band at 1221cm-1 corresponding a sulfate group vibration. For the pharmacological activities the tests was: antioxidant activity, changes in cell function (MTT test) and anticoagulant test. In the antioxidant activity we observed that DYN showed better results in the kidnapping of hydroxyl radicals and ferric chelation compared to DYS, this had the best result in the total antioxidant capacity. Both showed similar activity in reducing power and the kidnapping radicals DPPH. In MTT test DYN and DYS had not proliferative and cytotoxic activity in fibroblast cells (3T3) and showed antiproliferative and cytotoxic activity in cancer cell lines HeLa and B16 melanoma. In anticoagulant assay DYN showed good activity in the intrinsic pathway of blood coagulation, and a small activity in the extrinsic pathway, in the other hand DYS showed only a very small activity in the extrinsic pathway, but cannot come to be regarded as an anticoagulant agent. From these results it can be concluded that the alginate was extracted and sulfated, revealing a potential compound to be used in the pharmaceutical industry as an anticoagulant agent, antioxidant and antitumor and the sulfation has not been conclusively important to performance in the tested pharmacological activities

Homogalactanas sulfatadas da alga Codium isthmocladum com atividade anticoagulante

Since the first description of sulfated polysaccharides from seaweeds, the biological activities of these compounds have been evaluated under different aspects and experimental procedures. Among the broad biological activities presented by seaweed polysaccharides, anticoagulant action appears as a promising function. In this present study we have obtained sulfated polysaccharides from the green seaweed Codium isthmocladium by proteolytic digestion, followed by separation into five fractions (0.3, 0.5, 0.7, 0.9 and 1.2) by sequential acetone precipitation. The chemical analyses have demonstrated that all fractions are composed mainly by sulfated polysaccharides. The anticoagulant activity of these fractions was determined by activated partial thromboplastin time (aPTT) and prothrombin time test (PT) using citrate normal human plasma. None fraction has shown anticoagulant activity by PT test. Furthermore, all of them have shown anticoagulant activity by aPTT test. These results indicated that the molecular targets of these sulfated polysaccharides are mainly in the intrinsic via of the coagulation cascade. Agarose gel electrophoresis in 1,3-diaminopropane acetate buffer, pH 9.0, stained with 0.1% toluidine blue showed the presence of two or three bands in several fractions while the fraction 0.9 showed a single spot. By anion exchange chromatography, the acid polysaccharides from the 0.9 acetone fraction were separated into two new fractions eluted respectively with 2.0 and 3.0 M NaCl. These compounds showed a molecular weight of 6.4 and 7.4 kDa respectively. Chemical analyses and infrared spectroscopy showed that Gal 1 and Gal 2 are sulfated homogalactans and differ one from the other in degree and localization of sulfate groups. aPPT test demonstrated that fractions 2,0 and 3,0M (Gal1 and Gal 2, respectively) have anticoagulant activity. This is the first time that anticoagulant sulfated homogalatans have been isolated from green algae. To prolong the coagulation time to double the baseline value in the aPTT, the required amount of sulfated galactan 1 (6,3mg) was similar to low molecular heparin Clexane®, whereas only 0,7mg of sulfated galactan 2 was needed to obtain the same effect. Sulfated galactan 2 in high doses (250mg) induces platelet aggregation. These results suggest that these galactans from C. isthmocladum have a potential application as an anticoagulant drug

Atividades biológicas de polissacarídeos sulfatados extraídos da alga vermelha Gracilaria caudata

In recent years, sulfated polysaccharides (SP) from marine algae have emerged as an important class of natural biopolymers with potential pharmacology applications. Among these, SP isolated from the cell walls of red algae have been study due to their anticoagulant,antithrombotic and anti-inflammatory activities. In the present study, three sulfated polysaccharides fractions denominated F1.5v, F2.0v and F3.0v were obtained from seaweed G. caudate by proteolysis followed to acetone fractionation. Gel electrophoresis using 0.05 M 1,3-diaminopropane-acetate buffer, pH 9,0, stained with 0.1% toluidine blue, showed the presence of SP in all fractions. The chemical analysis demonstrated that all the fractions are composed mainly of galactose. These compounds were evaluated in anticoagulant, antioxidant and antiproliferative activities. In anticoagulant activity evaluated through aPTT and PT tests, no one fractions presented anticoagulant activity at tested concentrations (0.1 mg/mL; 1.0 mg/mL; 2.0 mg/mL).The antioxidant activities of the three fractions were evaluated by the following in vitro systems: Total antioxidant capacity, superoxide and hydroxyl radical scavenging, ferrous chelating activity and reducing power. The fractions were found to have different levels of antioxidant activity in the systems tested. F1.5v shows the highest activity, especially in the ferrous chelating system, with 70% of ferrous inhibiting at 1.0 mg.mL-1. Finally, all the fractions showed dose-dependent antiproliferative activity against HeLa cells. The fractions F1.5v and F2.0v presented the highest antiproliferative activity at 2.0 mg/mL with 42.7% and 37.0% of inhibition, respectively. Ours results suggests that the sulfated polysaccharides from seaweed G. caudata are promising compounds in antioxidant and/or antitumor therapy