Abstract In a continuation of our search for potential antiprotozoal agents from plants, we found that a crude extract from the roots of Lepidium virginicum exhibited antiprotozoal activity against Entamoeba histolytica trophozoites (IC 50 of 100.1 µg/mL). Bioassay‐guided fractionation resulted in the isolation of one known glucosinolate responsible for such activity. This compound was identified as benzyl glucosinolate. It showed in vitro activity against Entamoeba histolytica strain HM1‐IMSS (IC 50 of 20.4 µg/mL). The results support the anecdotal reports for the traditional use of L. virginicum roots in the control of diarrhoea and dysentery in the highlands of Chiapas, Mexico. Copyright © 2003 John Wiley & Sons, Ltd.