The in vitro effect of alcohlic extract of Laurus nobilis leaves on Leishmania major promastigote stage by colorimetric assay

Leishmaniasis is one of the most prevalent parasitic unicellular diseases worldwide, which is common between human and animal. At present, antiviral compounds such as glucantime and amphotericin B are used to treat cutaneous leishmaniasis. In many cases, herbal remedies do not have any side effects and are both affordable and inexpensive. Therefore, finding a herbal medicine to treat leishmaniasis is one of the global goals. The aim of this study was to evaluate the anti-leishmanial activity of alcoholic extract of leaves of bay laurel (Laurus nobilis) on promastigote stage of Leishmania major by in vitro colorimetric method.

Different concentrations of leaf extract of bay laurel were studied against L. major promastigotes compared to amphotericin B. L. major promastigotes were cultured in RPMI-1640 medium supplemented with 10 Percent FBS. Optical absorption (OD) was measured by ELISA method to determine 50 Percent inhibitory concentration (IC50) of drugs. Promastigotes were added to the 96-well plate and incubated for 72 h with different concentrations of bay laurel leaf extract and amphotericin B (800, 600, 400, 200, 50, 25 and 10 μg ml-1). Flow cytometry was used to evaluate apoptosis. All concentrations were incubated in MTT assay for 3 min and flow cytometric for 2 h. Statistical data were analyzed by Duncan, ANOVA and SAS software at P≤0.05.

Maximum amount of apoptesis (97.35 Percent) in amphotericin B treatments as positive control was related to treatment 800 μg ml-1 concentration.  In leaf extract treatments of bay laurel, greatest amount of apoptosis (63.04 Percent) was related to treatment of 800 μg ml-1 concentration. CC50 of amphotericin B and leaf extract of bay laurel treatments was obtained in 129.6 and 200.5 μg ml-1 concentrations, respectively. Inhibitory concentration (IC50) of amphotericin B and leaf extract of bay laurel treatments was 18.5 and 589.5 μg ml-1 on L. major promastigotes.

The results were shown increased dose-dependent apoptosis based on flow cytometric.  The results of this study showed that proximity of L. major promastigotes and amphotericin B concentration treatments leads to apoptosis-like cell death, which increases with increasing drug concentration. According to the IC50 results of leaf extract of bay laurel and high its apoptetic efficiency, further studies are needed to evaluate its effective combinants of leaf extract on Leishmania parasite and also, to design clinical setting for this herbal drug.