Determination of nine active components in Radix Hedysari and Radix Astragali using capillary HPLC with diode array detection and MS detection.

A capillary HPLC (cHPLC) coupled with diode array detection (DAD) and MS method was developed for the simultaneously qualitative and quantitative determination of nine components, namely vanillic acid, calycosin-7-O-beta-D-glucoside, (6alphaR,11alphaR)-9,10-dimethoxypterocarpan-3-O-beta-D-glucoside, ononin, calycosin, (3R)-2'-hydroxy-3',4'-dimethoxyisoflavan-7-O-beta-D-glucoside, isoliquiritigenin, formononetin, (3R)-8,2'-dihydroxy-7,4'-dimethoxyisoflavan, in Radix Hedysari (Hongqi) and Radix Astragali (Huangqi). Simultaneous separation of these nine compounds was achieved on a Zorbax C18 microcolumn (5 microm, 150 x 0.3 mm). The mobile phase consisted of (A) 0.3% aqueous formic acid and (B) ACN with a gradient elution. The identification of nine compounds in both Hongqi and Huangqi was confirmed by TOF-MS. All calibration curves showed good linearity (R(2) >0.998) within test ranges. This method showed good repeatability for the quantification of these nine components in Hongqi and Huangqi with intra- and inter-day variations of less than 1.89 and 3.13%, respectively. The validated method was successfully applied to quantify nine investigated components in eighteen samples of Hongqi and Huangqi. Hierarchical cluster analysis of 18 samples was performed using the peak area of nine analytes on cHPLC chromatograms. The result showed that Hongqi and Huangqi are significantly different, though the two species of Astragalus are very similar.