Qualitative and Quantitative Analysis of Steroidal Alkaloids in Fritillaria Species by Solid-Phase Extraction followed by Rapid Resolution LC/TOF-MS

Steroidal alkaloids are naturally occurring nitrogen-containing compounds in many edible or medicinal plants, such as potato, tomato, Fritillaria and American hellebore, which possess a variety of toxicological and pharmacological effects on humans. Such biological effects of these compounds create a critical demand for developing a sensitive and selective analytical method to accurately evaluate the presence and content of the major and minor steroidal alkaloids in these plants. In this report, we present a high-selective and sensitive method for rapid analysis of steroidal alkaloids in Fritillaria species, utilizing selective solid-phase extraction and rapid resolution liquid chromatography/time-of-flight mass spectrometry (SPE-RRLC/TOF-MS). The selective solid-phase extraction step was developed using a mixed-mode cation-exchange/reversed-phase cartridge (Oasis MCX). The strong cation exchange capacity of MCX can selectively capture basic analytes and remove acidic and neutral compounds in the plant extract, thereby reducing the matrix effect and improving the MS detection sensitivity. The sample recoveries on Oasis MCX cartridges were found to be > 80%. The analysis of steroidal alkaloids was carried out by RRLC/TOF-MS. The use of RRLC can shorten analytical time and improve chromatographic resolution, and TOF-MS provides abundant structure information by accurate mass measurements for each molecular ion and fragment ions at different fragmentor voltage. As a result, the SPE-RRLC/TOF-MS was successfully used for simultaneous determination of 26 steroidal alkaloids in different Fritillaria species in a single run within 18 min (Fig. 1), which is 5-times faster than conventional HPLC/TOF-MS method [1]. Acknowledgements: Financial support for this research from the cultivation fund of the key scientific and technical innovation project, Ministry of Education of China (No. 2004-295). References: [1] Zhou JL, Li P, (2008) J Chromatogr A, 1177: 126–137.

Fig. 1 Representative total ion chromatograms (TIC) of 26 steroidal alkaloids and internal standard. For those with poor separation, extraction ion mode was used to achieve reliable quantification, because they had different molecular weight.