Simultaneous Separation and Determination of Five Bioactive Components in Traditional Chinese Medicinal Formula, Guanxin II, by HPLC

Abstract

A reversed phase high performance liquid chromatographic method was established for the simultaneous determination of the bioactive constituents in traditional Chinese medicinal (TCM) formula Guanxin II. Danshensu, protocatechuic acid, protocatechualdehyde, paeoniflorin, and ferulic acid were successfully separated on an Agilent Zorbax extend‐C₁₈narrow‐bore column (150 × 2.1 mm i.d., 5 µm) with a guard column (12.5 × 2.1 mm i.d., 5 µm) packed with the same material at 25°C. The mobile phase was a mixture of methanol and 0.5% acetic acid employing gradient elution at a flow rate of 0.15 mL/min. Detection was accomplished with a diode‐array detector and chromatograms were recorded at 230 nm, 262 nm, 280 nm, and 322 nm. The compounds were identified by comparing their retention times and UV spectra in the 200–400 nm range with authentic standards. Regression equations revealed good linear relationship (correlation coefficients: 0.99938–0.99996) between the peak areas of the constituents and their concentrations. The relative standard deviations (n = 5) of retention time and peak area were less than 0.73% and 1.13%, respectively. The average recoveries (n = 3) were between 97.57% and 100.68%. The proposed method has been successfully applied to the simultaneous determination of the above bioactive constituents in Guanxin II decoction.

Acknowledgment

The authors thank the Modern Research Center for Traditional Chinese Medicine of Peking University for financial support.