High-speed separation and characterization of major constituents in Radix Paeoniae Rubra by fast high-performance liquid chromatography coupled with diode-array detection and time-of-flight mass spectrometry.

A fast high-performance liquid chromatography (HPLC) method coupled with diode-array detection (DAD) and electrospray ionization time-of-flight mass spectrometry (ESI-TOFMS) has been developed for rapid separation and sensitive identification of major constituents in Radix Paeoniae Rubra (RPR). The total analysis time on a short column packed with 1.8-microm porous particles was about 20 min without a loss in resolution, six times faster than the performance of a conventional column analysis (115 min). The MS fragmentation behavior and structural characterization of major compounds in RPR were investigated here for the first time. The targets were rapidly screened from RPR matrix using a narrow mass window of 0.01 Da to restructure extracted ion chromatograms. Accurate mass measurements (less than 5 ppm error) for both the deprotonated molecule and characteristic fragment ions represent reliable identification criteria for these compounds in complex matrices with similar if not even better performance compared with tandem mass spectrometry. A total of 26 components were screened and identified in RPR including 11 monoterpene glycosides, 11 galloyl glucoses and 4 other phenolic compounds. From the point of time savings, resolving power, accurate mass measurement capability and full spectral sensitivity, the established fast HPLC/DAD/TOFMS method turns out to be a highly useful technique to identify constituents in complex herbal medicines.