Gas chromatography coupled with mass spectrometry (GC/MS), using both electron impact (EI) and chemical ionization (CI) detection modes on apolar and polar stationary phases, led to the determination of the volatile composition of the essential oil obtained from tubers of Cyperus rotundus (Cyperaceae). In this study, more than 33 compounds were identified and then compared with the results obtained in our previous work. Cyperene, alpha-cyperone, isolongifolen-5-one, rotundene, and cyperorotundene were the principal compounds comprising 62% of the oil. An in vitro cytotoxicity assay with MTT indicated that this oil was very effective against L1210 leukaemia cells line. This result correlates with significantly increased apoptotic DNA fragmentation. The oxidative effects of the essential oil were evaluated using the 1,1-diphenyl-2-picrylhydrazyl (DPPH), xanthine/xanthine oxidase assays, and the scavenging of superoxide radical assay generated by photo-reduction of riboflavin. The antimutagenic activity of essential oil has been examined by following the inhibition of H(2)O(2) UV photolysis which induced strand-break formation in pBS plasmid DNA scission assay. Based on all these results, it is concluded that C. rotundus essential-oil composition established by GC/MS analysis, in EI- and CI-MS modes, presents a variety of a chemical composition we were not able to detect with only GC/MS analysis in our previous work. This essential oil exhibited antioxidant, cytotoxic, and apoptotic properties.