Rheum turkestanicum Janischew. (Polygonaceae) is a plant that grows in central Asia and in north-east of Iran. Traditionally, people use roots of R. turkestanicum as an anti-diabetic and anti-hypertensive as well as anticancer agent. In this study the cytotoxicity and apoptogenic properties of ethyl acetate (EtOAc), n-hexane and H2O extracts from Rheum turkestanicum Janischew. (Polygonaceae) root were determined against HeLa and MCF-7 cell lines and human blood lymphocytes. Malignant and non-malignant cells were cultured in RPMI 1640 medium and incubated with different concentrations of plant extracts. Cell viability was measured by MTS assay. Apoptotic cells were evaluated using PI staining of DNA fragmentation by flow cytometry (sub-G1 peak). The degree of DNA fragmentation was analyzed using agarose gel electrophoresis based on the formation of inter-nucleosomal units. The expression of apoptosis-related protein Bax and PARP cleavage were detected by Western blotting. EtOAc and n-hexane extracts decreased cell viability in malignant but not in non-malignant cells, as a concentration and time dependent manner. EtOAc extract induced a sub-G1 peak in flow cytometry histogram of treated cells compared to the control. DNA fragmentation indicating apoptotic cell death was involved in R. turkestanicum induced toxicity and cleaved PARP fragment was also detected. In conclusion, this is the first report on the cytotoxic effects of R. turkestanicum in which apoptosis played an important role. However, further evaluations are needed to fully understand the possible anti-tumor properties.