Development of a Validated HPLC Method for Analysis of Astragaloside II, Paeonol, and Osthole in Snake Wine

A gradient-elution RP-HPLC method has been established for simultaneous determination of astragaloside II, paeonol, and osthole in the snake wine. The compounds were separated on a Polaris C₁₈ column (150 mm × 4.6 mm i.d., 5-µm particles); compounds were eluted with a gradient prepared from acetonitrile and 0.01 M phosphate buffer, pH 6.8. The detection wavelengths were 205, 274, and 323 nm from 0–10 min, 10–13 min, and 13–20 min, respectively. Linear quantitative calibration plots were obtained over the concentration ranges 2–200 µg mL^?1 for astragaloside II and 0.1–50 µg mL^?1 for paeonol and osthole. Average recoveries for astragaloside II, paeonol, and osthole were 96.32%, 96.89%, and 97.12%, respectively, and the respective RSD were 1.45%, 1.13%, and 1.17%. The method was found to be precise with RSD values in the ranges 0.87–1.91% for intra-day assays and 1.03–2.43% for inter-day assays. This method was efficient with high reproducibility and accuracy. Different batches of snake wine were analyzed by use of the method.