Neutralization Of Fibrinogenolytic And Coagulant Activities From Bothrops moojeni Snake Venom By Schizolobium parahyba (Caesalpinoideae) Crude Aqueous Extract And Fractions. 

¹Vale, L. H. F.; ²Nascimento, E. A.; ¹Hamaguchi, A.; ¹Homsi-Brandeburgo, M. I.

1 Instituto de Genética e Bioquímica, 2 Instituto de Química, Universidade Federal de Uberlândia, Minas Gerais, Brasil.

Plant extracts constitute an extremely rich source of biologically active compounds, and a number of extracts have been shown to possess antivenom activity. This study reports the ability of the crude aqueus extract (AE) and fractions of Schizolobiumparahyba to inhibit two proteolytic activities from Bothropsmoojeni snake venom. The AE was prepared from the leaves by mixing in destiled water, centrifuged and the supernatant was lyophilized and stored at -20°C. AE (1mg) was submitted to fractionation on Sephadex LH-20 and eluted with solvents of growing polarity. A methanolic fraction (F1), one fraction 50% methanol and water (F2) and a aqueous fraction (F3) were collected. Proteolytic activity upon fibrinogen was determinate by using 75µL of bovine fibrinogen (1mg/mL PBS) incubated at 37ºC for 2h, pH 8.0. 1) positive control: crude venom (CV -5 mg);2) negative control AE, F1, F2 and F3 up to 1,25mg; 3) tests tubes: CV + AE or CV + Fractions in different proportions up to 1:250. The reactions were interrupted by add of the buffer containing denaturant, redutor agents, and analyzed by 14% (w/v) SDS-PAGE. Crude Venom was able to degrade totally the A and B chains of fibrinogen, while CV+AE resulted in the total protection of the Bb chain and partial of the Aa chain of the fibrinogen. The F1 fraction protected totally the fibrinogen chains while F2 and F3 didn’t protect the fibrinogen. The coagulant activity on the bovine plasma was tested at 37°C at the coagulometter apparatus by using 10mg of  CV (positive control); 0,5 mg of AE (negative control); 45 mL CaCl2 0,5 M (blanc); 45 mL CaCl2 0,5 M + 0,5 mg de EA (blanc + EA); CV + EA or CV + Fractions in different proportions up to 1:50. The statistical test of KrusKal-Wallis and Tukey-Type was aplicated to the results of the coagulant activity and revealed significantly increase in the clotting time for the test with AE and  fraction F3. F1 and F2 didn’t exhibit effect of the inhibition. We concluded that S. parahyba aqueous extract is able to neutralize fibrinogenolytic and coagulant activities from B. moojeni venom, which actives principles were separated by fractionation on LH-20 in two groups of compounds of different polarities.

Support: CAPES, FAPEMIG, CNPq and UFU.

Correspondence to: homsi@ufu.br