Abstract A new sesterterpene lactone, Hedyosulide, together with five previously described sesquiterpene lactones were isolated from the leaves of Hedyosmum brasiliense Mart. ex Miq. (Chloranthaceae). Their structures were established by NMR analysis and HRESIMS data. The isolated compounds were evaluated for antiprotozoal activity against trypomastigote and amastigote forms of Trypanosoma cruzi. Hedyosulide displayed the highest activity against the amastigote forms, in which the parasite takes in the vertebrate host, with IC50 of 21.6 μmol/L with a selectivity index (SI) higher than 9. Oxyonoseriolide showed moderate activity against the intracellular form with IC50 of 60.2 μmol/L and a weak SI (2.7), while the four other sesquiterpene lactones were inactive. Therefore, sesterterpene lactones, a relatively rare group of terpenoids, might be considered as a scaffold for developing new trypanosomicidal agents that are effective against T. cruzi. Also, this work brings an important contribution on the chemical/biological aspects of H. brasiliense due to the isolation of a sesterterpene lactone selectively active against both clinical forms of T. cruzi.