Leishmaniasis is characterized as a diverse set of infectious diseases caused by protozoan parasites of the genus Leishmania. In this eukaryotic cell, lipophosphoglycan (LPG) is the main glycoconjugate present on the outer surface of the plasma membrane. This molecule has important intra- and interspecific polymorphism, being crucial for the development and survival of the parasite in its hosts. Leishmaniasis treatments are based on drugs that have considerable side effects and are high cost. In this scenario, natural products emerge as relevant alternative sources of new compounds that have biotechnological applications. Within this context, lectins stand out, proteins of non-immune origin capable of binding in a specific and reversible way to carbohydrate residues or glycoconjugates free or attached to cell surfaces. The present study aimed to evaluate the anti-promastigote activity of the Canavalia ensiformis lectin (ConA) against Leishmania amazonensis strains. Promastigote forms of L. amazonensis strains PH8 and Josefa were incubated with different concentrations of ConA (298 - 4.65 μg/mL) and cell viability was determined by counting in a Neubauer chamber, as well as the participation of the carbohydrate recognition domain (DRC) on lectin activity. Fluorescence assays with 2',7'-dichlorodihydrofluorescein diacetate (DCFH2-DA), Propidium Iodide (IP) and Rhodamine 123 (Rho123) were carried out in order to verify the production of reactive oxygen species (ROS), damage cell membrane integrity and mitochondrial membrane potential (ΔΨm), respectively. Catalase assay was applied to evaluate the effect of extracellular H2O2 on lectin action. ConA inhibited the growth of promastigotes in a concentration- and time-dependent manner, with IC50/24h = 56.48 ± 0.02 μg/mL; IC50/48h = 54.34 ± 0.03 μg/mL; IC50/72h = 39.61 ± 0.02 μg/mL, for Josefa strain, and IC50/24h = 21.05 ± 0.02 μg/mL; IC50/48h = 14.08 ± 0.01 μg/mL; IC50/72h = 13.90 ± 0.01 μg/mL, for strain PH8. The anti-promastigote activity of ConA is due to its ability to recognize and bind to glycoconjugates present on the surface of L. amazonensis strains, leading to the production of ROS and damage to the integrity of the parasite membrane, however, without the participation of extracellular H2O2 and without causing damage to the mitochondrial membrane potential. The results suggest that the ConA lectin may be a promising alternative for the treatment of leishmaniasis, as well as for in vivo tests related to L. amazonensis strains.