In the present study, extracts of CHCl(3), n-BuOH and water of Lomatogonium carinthiacum were tested for their possible anticancer effects on human lung adenocarcinoma A549 cell line, human erythroleukaemia K562 cell line and human cervical carcinoma HeLa cell line. The inhibitory effect of the extracts on cell proliferation was assessed by MTT colourimetric assay in vitro. A high-performance liquid chromatography-electrospray-mass spectrometric (HPLC-EIS-MS/MS) method was developed for the determination of the constituents of the extracts. According to HPLC-EIS-MS/MS data, the chemical structures of 21 constituents of L. carinthiacum were identified on-line without time-consuming isolation. The L. carinthiacum extracts showed inhibitory effects on the abovementioned cell lines. Extracts of CHCl(3) were found to be the most inhibitory, with IC(50) values of 0.13, 0.75 and 0.60 µg mL(-1) on A549, K562 and HeLa, respectively. According to the IC(50) values, the order of sensitivity of the cell lines was A549 > HeLa > K562 and the inhibitory effects to the cell lines of these extracts were in the order CHCl(3) extract > water extract > n-BuOH, as xanthones > iridoids and secoiridoids > flavonols. The present study showed inhibitory activity of L. carinthiacum extracts on tumour cells.