A phytochemical investigation of the stem bark of Diospyros cuneata (Ebenaceae) together with in vitro cytotoxic evaluation of the pure compounds in four human cancer cell lines KB, Hep-2, HeLa and SiHa led to the isolation of plumbagin, elliptinone, lupeol, atraric acid methyl ester, maritinone, betulin and betulinaldehyde. The structural determination of the compounds was established by nuclear magnetic resonance, electron ionisation mass spectrometric analysis as well as comparison with data from the literature. Plumbagin exhibited a potent cytotoxic activity (CC50 = 3.56 µg mL(-1)) against KB cell lines whereas maritinone displayed cytotoxic activity against Hep-2 (CC50 = 17.30 µg mL(-1)), cervical cancer (CC50 = 21.10 µg mL(-1)) and the KB cell lines (CC50 = 20.30 µg mL(-1)). This is the first report on a phytochemical and biological evaluation of D. cuneata.