Methods to isolate the chemical constituents responsible for the cytotoxicity of Ligusticum grayi towards breast cancer cells

Thesis (M.S., Chemistry (Biochemistry)) -- California State University, Sacramento, 2009.The taproot of the perennial herb Ligusticum grayi has been used by Native Americans for many generations to treat common colds, respiratory infections, menstrual pain, and many other ailments. More recently, it was also reported to exhibit cytotoxic properties against breast cancer. An in vitro cell assay of hexane, ethanol, and aqueous root extracts showed that both the hexane and ethanol extracts are cytotoxic towards both breast cancer cells overexpressing estrogen and progesterone receptors (BT-474, LC50 = 14 + 4 ???g/mL) and those not overexpressing estrogen and progesterone receptors (MDA-MB-231, LC50 = 15 + 5 ???g/mL). A hexane-soluble fraction was made by extracting the roots with ethanol, drying the extract, then re-dissolving in hexane. This hexane-soluble fraction was stored at -20 ??C and used for all further studies. In an attempt to isolate the constituent(s) responsible for the cytotoxic properties, various separation techniques were employed: Separation via column chromatography, liquid-liquid partitioning, preparative thin layer chromatography, and liquid-liquid extraction followed by column chromatography did not successfully isolate the cytotoxic compound(s). Although the compounds were not successfully isolated, differences in peak abundance were observed as a result of these separations, and elimination of non-cytotoxic constituents were accomplished. The cytotoxicity assay had shown that osh?? roots do not exhibit any estrogenic activity, and chromatograms of GC and HPLC suggest one or a group of compounds are responsible for the cytotoxicity. Of the methods attempted, liquid-liquid extraction followed by column chromatography is the best scheme for the isolation of cytotoxic constituents from L. grayi roots.\ud \ud However, preparative HPLC with fraction collector is believed to be a better isolation method as shown by an efficient separation of the chemical constituents and the ability to collect fractions with relative close retention times.Chemistry (Biochemistry