Abstract: Cytotoxicity-guided fractionation of a methanol extract of the leaves and twigs of Rolandra fruticosa using the HT-29 human colon cancer cell line led to the isolation of seven sesquiterpene lactones, including the hitherto unknown isorolandrolide, 13-methoxyisorolandrolide (1), and bourbonenolide, 2α,13-diacetoxy-4α-hydroxy-8α-isobutyroyloxybourbonen-12,6α-olide (2), as well as five known compounds, 13-acetoxyrolandrolide (3), 8-desacyl-13-acetoxyrolandrolide-8-O-tiglate (4), 2-epi-glaucolide E (5), 2α,13-diacetoxy-4α-hydroxy-8α-methacryloyloxybourbonen-12,6α-olide (6), and 2α,13-diacetoxy-4α-hydroxy-8α-tigloyloxybourbonen-12,6α-olide (7). The structures of the two sesquiterpenes were elucidated on the basis of spectroscopic methods. All isolates were evaluated for their cytotoxicity using the HT-29 cell line, and only 13-acetoxyrolandrolide (3) was found to possess a potent inhibitory effect against this cell line. Compounds 3, 5 and 6 were also tested in a NF-κB (p65) inhibition assay, and 3 was assessed in an in vivo hollow fiber assay. [Copyright &y& Elsevier]