Development and evaluation of miltefosine-loaded nanoemulsions on in vitro culture of Toxoplasma gondii

Toxoplasma gondii is a common parasite in the world. Pharmaceutical options for toxoplasmosis treatment are limited. Several studies have been conducted on the anti-infectious properties of miltefosine (MLF). We investigated the effectiveness of nanoemulsion miltefosine (NEM) in tachyzoites of T. gondii, RH strain. Various NEM formulations were designed considering pseudo-ternary phase diagrams. Physicochemical properties of the developed nanoemulsions (NEs), including pH, polydispersity index (PDI), droplet size, and refractive index (RI) were evaluated. The considered formulation was analyzed for dilution and stability tests. MTT assay was performed on Vero cells for calculation CC50 and on Vero cells with RH strain tachyzoite for calculation IC50. Sulfadiazine (SDZ) and pyrimethamine (PYR) were positive controls. The trypan blue method was used to investigate the effect of drugs (NEM, MLF, SDZ, PYR) in reducing the number of infected Vero cells and reducing the intracellular proliferation of tachyzoites. Next, the viability of tachyzoites was measured in the tube in the direct vicinity of different drug concentrations. The final particle size of NEM was calculated to be 17.463 nm by DLS and TEM. The CC50 of NEM (75.7 µg/mL) indicated lower toxicity than the other drugs. IC50 obtained by trypan blue, MTT, and test tube methods showed that NEM (28.43 µg/mL) has a suitable IC50 against Toxoplasma tachyzoites. The calculated selectivity index (SI) demonstrated that NEM (SI=2.66) is a more suitable drug candidate than the MLF and positive controls. The trypan blue assay indicated the excellent reduction effect of NEM on T. gondii intracellular proliferation rate and the number of infected cells.