Antithrombotic and antiplatelet activities of new isohopane triterpene from the roots of Rubia akane

Abstract

A new isohopane triterpenoid (1) and two known triterpenoids (2-3) were isolated from the roots of Rubia akane (Rubiaceae). The molecular formula C30H46O4 of 1 was determined by HRESIMS. Detailed NMR spectroscopic data analysis suggested that compound 1 is a new isohopane triterpenoid with a ketone moiety at C-16. Based on the key NOE correlations of H-3/H-5 and H-21/H3-28, compound 1 was determined as 3β-hydroxy-16-oxo-21β-isohop-22(29)-en-24-oic acid. The anticoagulant activities of new isohopane 1 were evaluated by monitoring activated partial thromboplastin time (aPTT), prothrombin time (PT), and the activities of thrombin (Factor IIa, FIIa) and activated factor X (FXa). The effects of 1 on expression of plasminogen activator inhibitor type 1 (PAI-1) and tissue-type plasminogen activator (t-PA) were evaluated in tumor necrosis factor-α activated human umbilical vein endothelial cells. Treatment with 1 (200 μM) resulted in the prolongation of aPTT and PT and the inhibition of relative thrombin (28%) and FXa (29%) activities. In addition, 1 inhibited thrombin-catalyzed fibrin polymerization (18% inhibition at 100 μM) and platelet aggregation (21.8% inhibition at 100 μM). Compound 1 also elicited anticoagulant effects in mice in a dose-dependent manner ranging from 18.8 to 94.0 μg/mouse. In addition, treatment with 1 resulted in significant reduction of the PAI-1 to t-PA ratio (25% decreased). Collectively, the new isohopane triterpenoid 1 possesses antithrombotic activities and offers a basis for the development of a new anticoagulant agent.