Simultaneous Determination of 10 Components in Bu-Zhong-Yi-Qi Wan by Solid Phase Extraction-High Performance Liquid Chromatography with Diode Array Detection and Evaporative Light Scattering Detection

An effective, accurate and reliable method for the simultaneous separation and determination of 10 major components in Chinese medicine Bu-Zhong-Yi-Qi Wan (BZYQW) was developed and validated using solid phase extraction column-high performance liquid chromatography-diode array detection-evaporative light scattering detection (SPE-HPLC-DAD-ELSD). The chromatographic separation was performed on a Spursil™ C18 column (250 mm × 4.6 mm, 5 µm) at 30°C with an acetonitrile-water gradient as mobile phase. The DAD detection wavelength 254 nm was utilized for the quantitative analysis. The drift tube temperature and the carrier gas flow rate of the ELSD detection was set at 110.5°C and 3.1 mL/min. The total run time is 103 min, these determined components peak out within 81 min. Excellent linear behaviors over the investigated concentration ranges were observed with the values of r(2) higher than 0.9990 for all the analytes. The Linear range over hesperidin, senkyunolide I, senkyunolide H, ononin, calycosin, formononetin, ligustilide, butylene phthalide, astragaloside IV, astragaloside I is 4.50-94.50 µg/mL, 22.75-364.00 µg/mL, 2.30-45.00 µg/mL, 11.76-125.14 µg/mL, 4.62-50.35 µg/mL, 1.90-28.93 µg/mL, 1.29-159.00 µg/mL, 2.90-36.00 µg/mL, 35.40-192.40 µg/mL, 41.40-96.60 µg/mL, respectively. The method was validated by its repeatability [relative standard deviation (RSD) < 3.54%] and intra-day (RSD < 2.11%) and inter-day precision (RSD < 3.45%). The limits of detection and quantification of each component were in the ranges of 0.04-10.24 and 0.12-39.22 µg/mL, respectively. The average recovery yields of the 10 compounds ranged from 95.79 to 101.25%. The validated method was successfully applied to the simultaneous determination of these principal components in 10 commercial samples of BZYQW from different manufacturers.