Objective:To isolate and characterize the cytotoxic compounds from Diospyros quercina(Baill.)G.E.Schatz&Lowry(Ebenaceae).Methods:An ethno-botanical survey was conducted in the south of Madagascar from July to August 2010.Bio-guided fractionation assay was carried out on the root bark of Diospyros quercina,using cytotoxicity bioassay on murine P388 leukemia cell lines as model.The structures of the cytotoxic compounds were elucidated by 1D and 2D NMR spectroscopy and mass spectrometry.Results:Biological experiments resulted in the isolation of three bioactive pure compounds(named TR-21,TR-22,and TR-23)which exhibited very good in vitro cytotoxic activities with the IC_(50)values of(0.017 5±0.0060)μg/mL,(0.089±0.005)μg/mL and(1.027±0.070)μg/mL respectively.Thus,they support the claims of traditional healers and suggest the possible correlation between the chemical composition of this plant and its wide use in Malagasy folk medicine to treat cancer.Conclusions:The ability of isolated compounds in this study to inhibit cell growth may represent a rational explanation for the use of Diospyros quercina root bark in treating cancer by Malagasy traditional healers.Further studies are,therefore,necessary to evaluate the in vivo antineoplastic activity of these cytotoxic compounds as effective anticancer drugs. Objective: To isolate and characterize the cytotoxic compounds from Diospyros quercina (Baill.) GESchatz & Lowry (Ebenaceae). Methods: An ethno-botanical survey was conducted in the south of Madagascar from July to August 2010. The Bio-guided fractionation assay was carried out on the root bark of Diospyros quercina, using cytotoxicity bioassay on murine P388 leukemia cell lines as model. The structures of the cytotoxic compounds were elucidated by 1D and 2D NMR spectroscopy and mass spectrometry. Results: Biological SCIENCES results in the isolation of three bioactive pure compounds (named TR-21, TR-22, and TR-23) which exhibited very good in vitro cytotoxic activities with the IC 50 values ​​of (0.017 5 ± 0.0060) μg / mL, (0.089 ± 0.005) μg / mL and (1.027 ± 0.070) μg / mL respectively.Thus, they support the claims of traditional healers and suggest the possible correlation between the chemical composition of this plant and its wide use in Malagasy folk medicine to treat cancer. Conclusions: The ability of isolate d compounds in this study to inhibit cell growth may represent a rational explanation for the use of Diospyros quercina root bark in treating cancer by Malagasy traditional healers. Further studies are, therefore, necessary to evaluate the in vivo antineoplastic activity of these cytotoxic compounds as effective anticancer drugs.