Development of an HPLC method for asperulosidic acid quantification in the powder prepared from the spinal degeneration remedy of physician Nguyen Thien Chung

Abstract

Nguyen Thien Chung’s remedy which consists of 13 medicinal herbs had been demonstrated its analgesic and anti-inflammatory effects on animal experiments. This remedy has further been developed into powder form. This study was conducted to develop a method to quantify asperulosidic acid, the important marker in BT powder, by high-performance liquid chromatography (HPLC). Sample processing conditions and quantitative HPLC conditions were screened. The HPLC method was validated according to the ICH guidelines for system suitability, specificity, linearity, precision and accuracy. Methanol was chosen as the solvent for sample preparation and the ratio between BT powder and methanol was 300:25 (mg/ml). The selected HPLC conditions included a C18 Shimpack GIST column (250×4.6 mm, 5 μm), a detection wavelength of 236 nm, a column temperature of 30^oC, a flow rate of 1 ml/min, and an injection volume of 10 μl. The mobile phase was a mixture of acetonitrile and 0.1% H₃PO₄ with a ratio of 9.5:90.5 (v/v). The quantitative method showed a good linear correlation between the peak area and the concentration of asperulosidic acid (r²=0.9987). The RSD value of the intermediate precision was 1.24%. The recovery was found in a range of 91.22-99.76%. The HPLC method for asperulosidic acid quantification in BT powder met the validation requirements and could be applied to standardise and evaluate the quality of BT powder.