- Several anti-leishmanial drugs of choice are of plant origin. Many of the available drugs against the disease are toxic and in certain cases parasite drug resistance is developed. The development of new compounds is urgently required. Aims of the study To determine the leishmanicidal activity of the Nuphar lutea plant extract against Leishmania major in vitro . Materials and methods The leishmanicidal activity of methanolic plant extract against L. major free living promastigotes and intracellular amastigotes was evaluated, using microscopic examinations and the enzymatic XTT assay. Results Methanolic extract of N. lutea was highly effective against both Leishmania promastigotes and L . amastigotes (IC 50 =2±0.12 μg/ml; ID 50 =0.65±0.023 μg/ml; LD 50 =2.1±0.096 μg/ml, STI=3.23). The extract at 1.25 μg/ml totally eliminated the intracellular parasites within 3 days of treatment. Also, a synergistic anti-leishmanial activity was demonstrated with N. lutea extract combined with the anti-leishmanial drug – paromomycin. The partially purified N. lutea active component was found to be a thermo-stable alkaloid(s) with no electrical charge and is resistant to boiling and to methanol, dichloromethane and xylene treatment. Conclusions The present study suggests that N. lutea might be a potential source of anti-leishmanial compounds.