Leishmaniasis, one of the main concerns of the World Health Organization, is a parasitic disease caused by Leishmania species. The main objective of this study was to prepare a topical drug delivery system that can deliver glucantime to the site of cutaneous Leishmania wounds. Using the electrospinning method, a core-shell nanofibrous mat composed of macromolecules including polyethylene oxide, gelatin, poly (vinyl alcohol) and chitosan was prepared. The prepared nanofibers were characterized by scanning electron microscopy (SEM), transmission electron microscopy, Fourier transform infrared spectroscopy (FT-IR), tensile test and in vitro drug release test. The anti-Leishmania activities of drug-loaded nanofibers against Leishmania promastigotes and its cytotoxicity on fibroblasts were determined respectively by flow-cytometry and indirect MTT methods. Results of morphological studies showed that uniform nanofibers were prepared without any bead with average diameter of 404 nm. The TEM investigation confirmed the core-shell structure of the fibers. The in-vitro drug release assay was executed using Franz diffusion cell, which indicted 84% of glucantime was released during the first 9 h. The results indicated that 4 and 6 cm(2) of nanofibers mat were significantly killed promatigotes up to 78%. Moreover, the MTT assay also showed that the fabricated nanofibers do not possess any cytotoxicity towards fibroblast cells. (C) 2020 Elsevier B.V. All rights reserved.