Phytochemical and bioactivity studies on two endemic sideritis species

In this thesis study, phytochemical and bioactivity studies were carried out on Sideritis congesta P.H. Davis et Hub-Mor and Sideritis stricta Boiss. & Held. apud Bentham, two species belonging to Sideritis genus, which has a high endemism ratio for our country. The aerial parts of the two species were first macerated with methanol (MeOH) separately to get the main extracts. Then the main extracts were subjected to liquid - liquid re-extraction to get the sub-extracts of petroleum ether (PE), dichloromethane (DCM), ethyl acetate (EtOAc) and butanol (BuOH). Cytotoxic activity in selected extracts (MeOH, PE, DCM and BuOH) was tested using WST-1 cell proliferation assay on MCF-7 and MDA-MB-231 breast cancer cell lines and MCF-10A fibrocystic breast epithelial cell line. Comparison of IC50 values showed that DCM extract for S. congesta, DCM and PE extracts for S. stricta had more antiproliferative activity than the rest extracts. Since S. stricta BuOH extract had the highest extraction yield among all extracts and S. congesta DCM extract was found to be more active than other extracts, they were subjected to isolation by column and thin layer chromatography techniques. Linearol, 7-epicandicandiol, and foliol were isolated from S. congesta dichloromethane extract while four flavonoid glycosides 4′-O-methylhypolaetin 7-O-[6′′′-O-acetyl-β-D-allopyranosyl-(1 → 2)- 6″-O-acetyl-β-D-glucopyranoside] (F1), isoscutellarein 7-O-[6′′′-O-acetyl-β-D-allopyranosyl-(1 → 2)- 6″-O-acetyl-β-D-glucopyranoside] (F2), 4′-O-methylisoscutellarein 7-O-[6′′′-O-acetyl-β-D-allopyranosyl-(1 → 2)- 6″-O-acetyl-β-D-glucopyranoside] (F3), 3′-O-methylhypolaetin 7-O-[6′′′-O-acetyl-β-D-allopyranosyl-(1 → 2)- 6″-O-acetyl-β-D-glucopyranoside] (F4) were isolated from S. stricta butanol extract. Structure elucidation of the isolated compounds was made with 1D- and 2D NMR and MS techniques. By using LC-HRMS, identification and quantification of phenolic compounds was performed in the PE, DCM, EtOAc and BuOH extracts obtained by liquid-liquid extraction. EtOAc extracts followed by BuOH extracts were found to have high concentrations of phenolic content in both species. While p-coumaric acid and verbacoside were found to be in large quantities in S. congesta extracts, S. stricta extracts were rich in verbascoside and chlorogenic acid, in general. WST-1 test is also performed for linearol and 7-epicandicandiol isolated from S. congesta DCM extract. Although linearol was present to be high quantities in S. congesta DCM extract, it was not found to be active on the tested cell lines, showing that the activity of DCM extract can not be attributed to linearol. In contrast 7-epicandicandiol showed moderate activity on MCF-7, MDA-MB-231 and MCF-10A cell lines after 72 hours of incubation (IC50 values were found to be 12.39, 31.96 and 11.90 µM, respectively). Cytotoxic activity assay in purified diterpenes (linearol, 7-epicandicandiol and foliol) and F2, F3 and F4 flavonoid glycosides was performed by means of resazurin assay on CCRF-CEM and multi-drug resistant CEM/ADR5000 leukemia cell lines. The flavonoid glycosides were not found to be active on the selected cell lines, however 7-epicandicandiol among the three kaurene diterpenes was found to be the most active diterpene (IC50 16.83 ± 1.00 µM and 31.52 ± 3.00 µM for two cell lines, respectively). Within the scope of the thesis by using the extracts of Sideritis congesta, Zingiber officinale (ginger), Tilia sp. (linden) and Cinnamomum sp. (cinnamon) a syrup formulation was prepared. In accordance with the traditional usage of the plants in the formulation the syrup can be used to reduce the symptoms of upper respiratory tract infections.