Abstract
Leishmaniasis is a parasitic disease affecting millions of people in Africa, Asia, and South America. It is considered as a major public health problem causing morbidity and mortality worldwide. Actually, the luck of vaccines and effective and less expensive antileishmanial therapy, have oriented pharmacological researches on natural antileishmanial drugs from medicinal plants. Hence, the purpose of this work is to optimize extraction parameters such as the percentage of aqueous ethanol mixture, the temperature and the time through the ultrasound bath equipment using a Box–Behnken Design (BBD) for maximal antileishmanial activity extraction from Moringa oleifera leaves. The cytotoxicity of the optimum extract was evaluated on RAW 264.7 macrophage cell line. The synergism between M. oleifera optimal extract and amphotericin B was also investigated using checkboard method. The optimum extract exhibiting the highest percent inhibition of Leishmania major promastigotes (99.15 ± 0.34%) was obtained using 16%v/v ethanol, at 60 °C for 20 min. Interestingly, this extract showed the best IC50 value of 6.87 ± 0.32 and 9.31 ± 0.72 μg/mL against promastigote and amastigote forms, respectively with no cytotoxicity on macrophage cells Raw 264.7 (SI = 17.53). Moreover, this extract showed a synergistic effect with amphotericin B (FIC = 0.375). HPLC analysis showed that at least six phenolic compounds were identified. Resorcinol, luteolin7-O-glucoside and syringic acid were the most active ones against L. major with IC50 values 3.788 ± 0.48, 10.70 ± 0.59 and 13.41 ± 0.59 μg/mL, respectively. Rutin and ferulic acid were further identified and showed moderate antileishmanial activity with IC50 values 78.51 ± 1.09 and 89.34 ± 1.22 μg/mL, respectively. Kaempferol 3-O-rutinoside was less active with IC50 value 204.4 ± 1.63 μg/mL.
•This study aims to optimize the antileishmanial activity of M. oleifera using BBD.•Optimal extraction conditions are 16% hydroalcoholic percentage at 60 °C for 20 min.•Optimized extract showed an effective antiprotozoal activity with no cytotoxicity.•High synergistic combination of optimized extract with amphotericin B was revealed.•Identified phenolic compounds by HPLC showed an effective antileishmanial property.