Identification and Quantification of three Tubeimosides in Rhizoma Bolbostematis by High Performance Liquid Chromatography with Evaporative Light Scattering Detection and Electrospray Mass Spectrometric Detection

Abstract

Tubeimoside I, tubeimoside II and tubeimoside III were simultaneously determined and identified by high performance liquid chromatography coupled with electrospray tandem mass spectrometry (HPLC-ESI/MS), and a novel and sensitive high performance liquid chromatography - evaporative light scattering detection (HPLC-ELSD) in a 70% methanol extract of Rhizoma Bolbostematis. The chromatographic separation was performed on a Zorbax Extend C₁₈ analytical column using gradient elution with a solution of acetonitrile and 0.5% acetic acid. The method was validated with acceptable linearities (r > 0.9992) and recoveries (98.6 to 102.4 %). The limits of detection of these three tubeimosides were as low as 0.05 μg. The intra- and inter-day precisions of the method were evaluated and were less than 3.6%. The method was successfully used to analyze 15 batches of Rhizoma Bolbostematis. The content of tubeimosides in the plant material varied from habitat to habitat confirming the necessity to control the quality of Rhizoma Bolbostematis during its preparation and application in the clinic.