Objective To determine the structure of triterpenoid isolated from avocado seeds and the cytotoxic effect on MCF-7 and HepG2 cells. Methods The powder sample was macerated with ethanol, followed with separation of the extract by column chromatography. The target compound was monitored on thin layer chromatography plate and reagent Lieberman–Buchard. The isolated compound was characterized by spectral analysis, mainly ultraviolet, infrared, and liquid chromatography-mass spectroscopy and their spectroscopic data with those reported in literature were compared. In vitro cytotoxic activity was investigated against Vero, MCF-7, and HepG2 cell lines using MTT assay. Results A triterpenoid compound was isolated from ethanol extract. The extracts, fraction (F