Thujone-Rich Fraction of Thuja occidentalis Demonstrates Major Anti-Cancer Potentials: Evidences from In Vitro Studies on A375 Cells

Crude ethanolic extract of Thuja occidentalis (Fam: Cupressaceae) is used as homeopathic mother tincture (TO Phi) to treat various ailments, particularly moles and tumors, and also used in various other systems of traditional medicine. Anti-proliferative and apoptosis-inducing properties of TO Phi and the thujone-rich fraction (TRF) separated from it have been evaluated for their possible anti-cancer potentials in the malignant melanoma cell line A375. On initial trial by S-diphenyltetrazolium bromide assay, both TO Phi and TRF showed maximum cytotoxic effect on A375 cell line while the other three principal fractions separated by chromatography had negligible or no such effect, because of which only TRF was further characterized and subjected to certain other assays for determining its precise anti-proliferative and apoptotic potentials. TRF was reported to have a molecular formula of C10H16O with a molecular weight of 152. Exposure of TRF of Thuja occidentalis to A375 cells in vitro showed more cytotoxic, anti-proliferative and apoptotic effects as compared with TO Phi, but had minimal growth inhibitory responses when exposed to normal cells (peripheral blood mononuclear cell). Furthermore, both TO Phi and TRF also caused a significant decrease in cell viability, induced inter-nucleosomal DNA fragmentation, mitochondrial transmembrane potential collapse, increase in ROS generation, and release of cytochrome c and caspase-3 activation, all of which are closely related to the induction of apoptosis in A375 cells. Thus, TRF showed and matched all the anti-cancer responses of TO Phi and could be the main bio-active fraction. The use of TO Phi in traditional medicines against tumors has, therefore, a scientific basis.