Oleanolic acid, a triterpenoid known for its anti-inflammatory properties, is commonly present in several medicinal plants. The present study evaluated the effect of oleanolic acid on sPLA (2), a key enzyme in inflammatory reactions. Oleanolic acid inhibited sPLA (2) activities of human synovial fluid (HSF), human pleural fluid (HPF) and VIPERA RUSSELLI (VRV-PL-V) and NAJA NAJA (NN-PL-I) snake venoms in a concentration-dependent manner. The IC (50) values of sPLA (2) from these sources ranged from 3.08 to 7.78 muM. Increasing calcium (Ca (2+)) concentrations from 2.5 to 15 mM and substrate concentration up to 180 nM did not affect the level of inhibition. Oleanolic acid enhanced the relative intrinsic fluorescence intensity of sPLA (2) (VRV-PL-V). In the presence of oleanolic acid, an apparent shift in the far UV-CD spectrum of sPLA (2) was observed. These studies indicate direct interaction with the enzyme and formation of an sPLA (2)-oleanolic acid complex. The complex formed resulted in irreversible inhibition of sPLA (2). Oleanolic acid inhibited indirect hemolytic activity and mouse paw edema induced by sPLA (2). Inhibition of IN VITRO and IN VIVO sPLA (2) activity by oleanolic acid explains the observed anti-inflammatory properties of several oleanolic acid-containing medicinal plants.

In the present study we compared the clot inducing and dissolving properties of Calotropis gigantea R. Br. (Asclepiadaceae), Synadenium grantii Hook. f. (Euphorbiaceae) and Wrightia tinctoria R. Br. (Apocynaceae) latex extracts. All the three latex extracts hydrolyzed casein, fibrinogen and crude fibrin dose-dependently. The proteolytic action on fibrinogen subunity was in the order of Aalpha > Bbeta > gamma. All extracts exhibited procoagulant activity as assayed by re-calcification time. However, thrombin like activity is restricted to C. gigantea. In addition, the extracts dose-dependently hydrolyzed blood and plasma clots. Furthermore, the hydrolyzing pattern of fibrin in the plasma clot was substantiated by SDS-PAGE. The extracts hydrolyzed all the subunits (alpha polymer, alpha-chains, gamma-gamma dimer and beta-chain) of fibrin efficiently. Both fibrinogenolytic and fibrinolytic activity potency of the extracts were in the order of C. gigantea > S. grantii > W. tinctoria. Among the three latices, C. gigantea is toxic with a minimum hemorrhagic dose (MHD) of > 75 microg, whereas S. grantii and W. tinctoria latex extracts were non-toxic and did not induce any hemorrhagic effect at the tested dose (> 200 microg). The proteolytic activity of C. gigantea latex extract on different substrates was inhibited by IAA. On the other hand, the proteolytic activities of S. grantii and W. tinctoria were inhibited by PMSF. Thus, this study provides the basis for the probable action of plant latex proteases to stop bleeding and effect wound healing as exploited in folk medicine.

In the present study we evaluated the presence of cysteine protease from the latex of four plants Asclepias curassavica L., Calotropis gigantea R.Br., Pergularia extensa R.Br. and Cynanchum puciflorum R.Br. belongs to the family Asclepiadaceae. Cysteine proteases from these plants latex exhibited both thrombin and plasmin like activities. Latex enzyme fraction in a concentration dependent manner induced the formation of clot in citrated blood plasma. Direct incubation of fibrinogen with latex enzyme fraction resulted in the formation of fibrin clot similar to thrombin enzyme. However prolonged incubation resulted in degradation of the formed fibrin clot suggesting plasmin like activity. Latex enzyme fraction preferentially hydrolyzed Aalpha and Bbeta chains of fibrinogen to form fibrin clot. Latex enzyme fraction also hydrolyzed the subunits of fully cross linked fibrin efficiently, the order of hydrolysis was alpha-polymer > alpha-chains > beta-chain and gamma-gamma dimer. Cysteine proteases from all the four Asclepiadaceae plants latex exhibited similar action on fibrinogen and fibrin. This study scientifically validate the use of plant latex in stop bleeding and wound healing by traditional healers all over the world.

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