Thirty-six essential oils from Verbenaceae and Asteraceae family plants along with their major constituents were evaluated to determine cytotoxicity on Jurkat, HeLa, HepG2 and Vero cell lines. The plants were collected in different regions of Colombia, and their essential oils were extracted through microwave-assisted hydrodistillation and then characterized through GC/MS and GC/FID analyses. Cytotoxic activity testing was carried out using a tetrazolium-dye (MTT) assay. The essential oils from Lippia alba Citral and Carvone chemotypes showed the lowest IC50 and selectivity index (IS) > 10. The essential oils from Asteraceae family were not selective against the tumor cell lines tested. Citral chemotype L. alba essential oils showed values up to 27.2, and Carvone chemotype L. alba essential oils showed values up to 30.7. Citral, showed the highest cytotoxic activity, with an SI value of 1241, and Limonene and Linalool showed SI values of 6.97 and 10.1, respectively, on HeLa cells.