Stability study on a cytotoxic dienynone isolated from Echinacea pallida

Recent studies have demonstrated that some of the secondary metabolites (polyacetylenes and polyenes) isolated from the raw lipophilic extract of Echinacea pallida roots have good cytotoxic activity on cancer cell lines3. In particular, pentadeca-(8Z,13Z)-dien-11-yn-2-one was found to be the most active constituent, with an IC50 value of 32.17 ± 3.98 uM on pancreatic MIA PaCa-2 cancer cells and 2.34 ± 0.33 uM on colonic COLO320 cancer cells, after 72 h of exposure. This compound is present in the roots in a not-oxidized form but, during the drying process and the subsequent storage, a change in its concentration was observed, which has been attributed to a possible oxidation process. As part of our ongoing research on bioactive metabolites from Echinacea, the aim of this research was the evaluation of the stability of this secondary metabolite by RP-HPLC-DAD2 under the experimental conditions applied during the biological assays. The study was initially carried out on the lipophilic crude extract obtained from E. pallida roots and then it was focused on the individual component isolated and purified from the plant material by preparative chromatographic techniques. The results showed a moderate change in the HPLC profiles of E. pallida crude extract in 72 h, with an increase of hydroxylated derivatives. The oxidation kinetic of the most active compound, pentadeca-(8Z,13Z)-dien-11-yn-2-one, was found to be quite slow, since the % area of the intermediate hydroperoxide was about 25% after 72 h of exposure. The final product of the oxidation process, i.e. the hydroxilated derivative (i.e. 8-hydroxy-pentadeca-(9E,13Z)-dien-11-yn-2-one, available thanks to a synthetic approach), was not observed in the HPLC chromatogram after 72 h. Considering that during the first 24 h of exposure, the % area of the hydroperoxide intermediate is below 15%, the observed cytotoxic activity can be mainly attributed to the genuine not oxidized compound.