In vitro antileishmanial activity of hydroalcoholic extract, fractions, and compounds isolated from leaves of Piper ovatum Vahl against Leishmania amazonensis

The medicinal plant P. ovatum is used popularly as an anesthetic [1] and anti-inflammatory [2]. We assessed the biological activity of a crude extract, a mixture of several fractions, and a pure compound obtained from Piper ovatum Vahl against promastigote and amastigote forms of Leishmania amazonensis. This study included the extraction process and bioassay-guided fractionation by the adsorption chromatography and Sephadex LH-20 method. A progressive increase in the antileishmanial effect was observed in the course of fractionation. The 50% inhibitory concentration (IC₅₀) for dichloromethane-ethyl acetate (1:1 v/v) fraction was 2.1µg/ml and 24µg/ml; mixture of piperovatine: piperlongumune (2:3) 0.9µg/ml and 24µg/ml; piperovatine (1) 9.5µg/ml and 10µg/ml; and piperlonguminine (2) 2.5µg/ml and 9.0µg/ml, for promastigote and amastigote forms, respectively. Cytotoxicity analysis indicated that these toxic concentrations were much higher for J774G8 macrophages and Vero cells than for the protozoans. The mixture of piperovatine:piperlongumune (2:3) showed important antiprotozoal activity against the amastigote and promastigote forms of L. amazonensis, and it produced morphological changes in promastigotes and amastigotes at 0.9µg/ml and 24µg/ml (50% growth inhibition concentration), respectively, including intense cytoplasmic vacuolization, mitochondrial swelling, and mitochondrial damage, as revealed by transmission electron microscopy.

Acknowledgements The authors are grateful to CNPq for providing a research grant and fellowships.

References: [1] Correa, M.P. (1984) Dicionario das Plantas Uteis do Brasil e das Exoticas Cultivadas, vol. 1. Instituto Brasileiro de

Desenvolvimento Florestal, Rio de Janeiro.

[2] Rodrigues-Silva, D. et al. (2008)J. Ethnopharmacol. 116:569–573.