Cytotoxic Effects of Bifora testiculata (L.) Spreng. Essential Oil and Its Main Component on Cancer Cell Lines

Bifora testiculata (L.) Spreng. (Apiaceae), an understudied species endemic to the Mediterranean and the only representative species of the genus Bifora in Sicily, was investigated for the first time for its essential oil (EO) chemical composition and cytotoxic properties. The EO was obtained via hydrodistillation and analyzed using GC-MS, revealing an aldehyde-rich profile (86.10%), dominated by trans-2-dodecenal (67.49%). Comparative analysis with previous studies on B. testiculata from Greece confirmed a similar aldehyde-rich profile, although minor compositional differences suggest potential chemotype variation. Given the biological relevance of trans-2-dodecenal and related aldehydes, further investigations into the cytotoxic properties of the EO of B. testiculata (Bt) and its main constituent against cancer cell lines were undertaken. Three human tumor cell lines (MDA-MB 231, A375, and CaCo2) and a human non-tumor cell line (HEK293) were subjected to viability tests using the MTT assay. The EO and trans-2-dodecenal exhibited remarkable cytotoxic activity against all cell lines, with IC₅₀ values ranging between 7.93 and 14.41 µg/mL for Bt and between 1.88 and 5.29 µg/mL for trans-2-dodecenal. AO/BE fluorescent staining and Hoechst nuclear staining showed the presence of apoptotic bodies in the treated cells. N-acetyl-L-cysteine was able to invert the effects of Bt and trans-2-dodecenal on cell lines, suggesting ROS involvement in cytotoxic activity. The results demonstrated that the Bt cytotoxic activity was mainly due to the presence of trans-2-dodecenal.