Simultaneous determination of principal components and related substances of raw material drug of ammonium glycyrrhizinate by reversed-phase high performance liquid chromatography

An analytical method for the simultaneous determination of 18 alpha-glycyrrhizic acid, 18 beta-glycyrrhizinic acid, related substances A and B and drug quality standard by reversed-phase high performance liquid chromatography (RP-HPLC) was established. The assay was carried out on a Durashell-C-18 column (250 mm x 4.6 mm, 5 mu m) with 10 mmol/L ammonium perchlorate (the pH value was adjusted to 8.20 with ammonia)-methanol (48 : 52, v/v) as mobile phase at a flow rate of 0.80 mL/min, and the detection wavelength was set at 254 nm. The column temperature was 50 degrees C and the injection volume was 10 mu L. Under the separation conditions, the calibration curves of the analytes showed good linearities within the mass concentrations of 0.50 - 100 mg/L (r > 0.999 9). The detection limits for 18 alpha-glycyrrhizic acid, 18 beta-glycyrrhizinic acid, related substances A and B were 0.15, 0.10, 0.10, 0.15 mg/L, respectively. The average recoveries were between 97.32% and 99.33% (n = 3) with the relative standard deviations (RSDs) between 0.05% and 1.06%. The method is sensitive, reproducible, and the results are accurate and reliable. The method can be used for the determination of principal components and related substances of ammonium glycyrrhizinate for the quality control of raw material drug of ammonium glycyrrhizinate.