Cytotoxic activity of ethanol extracts of in vitro grown Cistus creticus subsp. creticus L. on human cancer cell lines

Cistus creticus subsp. creticus is a native plant of the Mediterranean region and it has been used since ancient times for its medicinal properties. The leaves secrete labdane type diterpenes that exhibit considerable antibacterial and cytotoxic activities. However there are no data about implementation of in vitro culture of C. creticus in order to obtain applicable production of labdane diterpenes. We successfully established in vitro culture of shoots and roots on solid or liquid MS (Murashige and Skoog) medium without addition of plant growth regulators. GC/FID and GC/MS detections were used to carry out characterization of ethanol extracts of shoots and roots from in vitro culture. Labdane diterpenes were the most abundant compounds in shoot extract, but absent from the root extract. Total phenolic and flavonoid contents of C. creticus shoot and root ethanol extracts were also determined. In order to evaluate possible cytotoxic activity of extracts, sulforhodamine B (SRB) assay was performed on five human cancer cell lines. Only shoot extract had cytotoxic activity on HeLa (cervix), MDA-MB-453 (breast) and FemX (melanoma) cancer cells with IC50 reaching 80.83 μg/ml (HeLa), 76.18 μg/ml (MDA-MB-453), and 87.52 μg/ml (FemX) respectively. This is the first report of cytotoxic effect of labdane diterpenes from Cistus on HeLa cells.

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► Successfully established in vitro culture of Cistus creticus on MS medium without plant growth regulators.
► GC/MS analysis revealed the presence of labdane diterpenes in shoot ethanol extract and free-fatty acids in root ethanol extract of C. creticus from in vitro culture.
► Shoot extract had cytotoxic activity on HeLa (cervix), MDA-MB-453 (breast) and FemX (melanoma) cell lines.