The antitumour antibiotic pingyangmycin (PYM; bleomycin A5) was isolated from many components of bleomycin (BLM) produced by Streptomyces pingyangensisn. PYM has a similar chemical structure to that of BLM but the terminal amine moiety is different. Therefore, it would be of significance to demonstrate the antitumour effect and action mechanism of PYM on cultivated tumour cells. In this study, we used the cell growth curve, plating efficiency, and DNA synthesis inhibition assay to demonstrate the cytotoxicity of PYM on cultured KB cells. In the meantime, the morphological variations of drug-treated cells were also observed. In addition, we used the DNA precipitation assay, a simple and rapid assay, for detecting DNA damage caused by PYM on cultured KB cells for potential genotoxicity. Our results indicate that the effect of PYM significantly inhibits the cell growth, colonyforming ability, and DNA synthesis of KB cells in a dose-dependent manner. Furthermore, when treated with 5 micrograms/ml of PYM for 24 h on cultured KB cells, DNA strand breaks can be induced (P < 0.05). Therefore, it is considered that the action mechanism of PYM is due to its ability to inhibit the synthesis of DNA and split the DNA chains.