Separation identification and quantification of selected withanolides in plant extracts of Withania somnifera by HPLC-UV(DAD) -: Positive ion electrospray ionisation-mass spectrometry

This paper describes a method for separation, identification, and quantification of selected withanolides in Withania somnifera plant extracts by HPLC-UV(DAD)-Mass Spectrometry (HPLC-MS). Withaferin-A (WS-3), 12-deoxywithastramonolide (WS-12DS), Withanolide A (WS-1), and Withanone (WS-2) were used as external standards. The compounds were isolated from Withania somnifera by repeated column chromatography of the root extract and their identity was established by H-1- and C-13-NMR and mass spectral data. The compounds were chromatographed on a Merck (250 x 4.6 mm ID, 5 mum) column and analyzed by Electrospray Ionization on a mass spectrometer in Selected Ion Mode (SIM). For quantification, [M + Na](+) ions were monitored. Linear calibration curves were obtained in the concentration range of 1.50 mug/mL to 6.5 mug/mL. The method was applied successfully to the detection and quantification of the said withanolides in a number of samples.