Anticoagulant activity of sulfated polysaccharide isolated from fermented brown seaweed Sargassum fulvellum

A sulfated polysaccharide with anticoagulant properties was isolated from the fermented brown seaweed Sargassum fulvellum. Freeze-dried S. fulvellum was fermented in an incubator for 10(th) week at 25 degrees C to convert seaweed macromolecules into anticoagulant sulfated polysaccharides (ASP). Anticoagulant activity was determined by an activated partial thromboplastin time (APTT) test using citrated human blood plasma. The 8(th) week S. fulvellum crude seaweed extract (SWE) exhibited the highest blood anticoagulant activity. Therefore, 8(th) week crude SWE was used for purification of ASP by two steps; DEAE cellulose anion-exchange followed by Sepharose 4B chromatography. The isolated ASP showed a single spot on agarose gel electrophoresis, which confirmed the purification status of our ASP. Polyacrylamide gel electrophoresis (PAGE) analysis showed that the molecular mass of the purified ASP was between 8 and 20 kDa. Polysaccharide and sulfate concentrations of the purified ASP were 180 and 29.70 mu g mL(-1) respectively. ASP recovery was 1.32% (w/w) from the crude polysaccharide applied to the DEAE column. Purified ASP had a pH of 3.86 and was considered an acidic polysaccharide. Moreover, both ASP and heparin showed a relative clotting factor of 27.47 at the concentrations of 180 and 60 mu g mL(-1) respectively. Therefore, S. fulvellum ASP can be considered a weaker anticoagulant than heparin. Results of the APTT, PT, and TT clotting assays showed that ASP was able to inhibit both intrinsic and extrinsic blood coagulation pathways. Finally, this study established a feasible and simple experimental protocol to isolate anticoagulant from fermented seaweeds leading to potential further development of anticoagulant agent for the pharmaceutical industry.