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This study assessed the in vitro anthelmintic activity of eugenol, thymol and tannic acid against the acanthocephalan endoparasite Neoechinorhynchus buttnerae. Tambaqui fish Colossoma macropomum intestines were collected and dissected for parasite removal. The anthelmintic activity of the compounds studied was tested in vitro using minimum essential medium (MEM) to maintain parasite viability. One control group and from 8 to 10 treatments were carried out in triplicate. Ten N. buttnerae specimens were introduced to each MEM plate, and after exposure to the compounds, parasite motility was observed every 15 min. The LC50 value of eugenol after 2, 6 and 24 h exposure was 1.01, 0.79 and 0.41 mg.mL-1, respectively; to thymol the values were 1.97, 0.96 and 0.92 mg.mL-1, and to tannic acid were 4.68, 1.79 and 1.30 mg.mL-1. Among the compounds tested, eugenol was the most efficient against N. buttnerae, eliminating 100% of the parasites after 15 min exposure to the three highest concentrations tested (1.56, 3.125 and 6.25 mg.mL-1). Thymol eliminated over 90% of the parasites after 2h exposure to the three highest levels tested (0.78, 1.56 and 3.125 mg.mL-1), and tannic acid showed the lowest activity, eliminating 100% of parasites after 6h exposure to doses between 6.25 and 50 mg.mL-1. [ABSTRACT FROM AUTHOR]