We further used the propidium iodide (PI) cell cycle assay based on flow cytometry to examine how coumarin-30 affects the proliferation of cultured human cancer cells (Fig. ). Treatment of cells with nocodazole was used as a positive control. Coumarin-30 inhibited mitosis in submicromolar concentrations (Fig. , IC50 = 0.15 ± 0.01 μM). Using time-lapse phase contrast microscopy, we confirmed that coumarin-30, like nocodazole, induced a prolonged mitotic arrest in A549, MCF-7 and HeLa cells after 12 hours of treatment (Fig. , Fig. ).