Leishmaniasis is a neglected disease that is increasing globally at an alarming rate. Glucantime has been the therapy of choice for more than 50 years. A recent study reported the antileishmanial activity of copaiba oil against Leishmania amazonensis. These results led us to investigate morphological and ultrastructural changes in L. amazonensis treated with copaiba oil, using electron microscopy and flow cytometry to assess specific organelles as targets for copaiba oil. In the promastigote and axenic amastigote forms, this copaiba oil caused notable morphological and ultrastructural changes, including extensive mitochondrial damage and denaturation of the plasma membrane. Copaiba oil treatment also induced a decrease in Rh123 fluorescence, suggesting interference with the mitochondrial membrane potential and loss of cell viability with an increase in plasma membrane permeability, as observed by flow cytometry after staining with propidium iodide. In conclusion, copaiba oil could be exploited for the development of new antileishmanial drugs.