Berberine iodide (IK-1) and acetoneberberine (IK-2) showed higher cytotoxicity against five human oral squamous cell carcinoma (HSC-2, HSC-3, HSC-4, NA, CA9-22) and one human promyelocytic leukemia (HL-60) cell lines, than against normal human oral tissue-derived cells (gingival fibroblast HGF, pulp cell HPC, periodontal ligament fibroblast HPLF), producing a tumor specificity index of 4.0 and 3.6, respectively. IK-1 was more potent than IK-2 in inducing the production of apoptotic cells, internucleosomal DNA fragmentation, the activation of caspases-3, -8 and -9, and the increased expression of proapoptotic BAD protein, with a corresponding decrease in the expression of anti-apoptotic Bcl-2 protein in HL-60 cells. These compounds did not induce internucleosomal DNA fragmentation (only producing larger DNA fragment), nor increased the Bad protein expression in HSC-2 cells. The present study demonstrated the tumor-specific cytotoxicity and apoptosis-inducing activity of berberines, suggesting their possible antitumor potentiaL