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A dermatan sulfate isolated from the shark Scyliorhinus canicula skin by enzymatic digestion followed by purification with anion exchange chromatography was identified by chondroitinase and nitrous acid treatment and partially characterized by Fourier-transform infrared spectroscopy. Dermatan sulfate was the major glycosaminoglycan and represented 75% of the polysaccharide fraction in the sharkskin. This dermatan sulfate had a 38.6 kDa average molecular weight and 23% sulfate content. The anticoagulant action of this dermatan sulfate was checked by several coagulometric and colorimetric assays such as the activated partial thromboplastin time, thrombin time, thrombin generation and heparin cofactor II and antithrombin-mediated inhibition of thrombin and compared with that of porcine intestinal mucosa dermatan sulfate. The effects on platelet activation and aggregation were investigated using flow cytometry and aggregometry, respectively. The dermatan sulfate prolonged activated partial thromboplastin time and thrombin time, delayed and inhibited thrombin generation in a concentration-dependent manner. The specific anticoagulant activity of the sharkskin dermatan sulfate was 43 UI/mg. The anticoagulant effect of sharkskin dermatan sulfate was higher than that of the porcine dermatan sulfate and was due to the potentiation of thrombin inhibition by heparin cofactor II. Moreover, it had no effect on platelet aggregation and activation induced by various agonists and thereby constitutes a potentially useful drug of interest in anticoagulant therapy.