RP-HPLC Determination of Pseudohypericin in Hypericum Attenuatum Choisy

The sample (10.000 0 g) was extracted ultrasonically for 35 min with 300 mL of 80% (φ) ethanol solution at 70℃ (ultrasonic power 225 W). An aliquot of 10% of total mass of the concentrated extract was dissolved and made its volume up to 100 mL with 80% ethanol, and filtered through 0.45 μm PTEF microporous filter membrane. The diluted solution was separated by RP-HPLC using VP-ODS C₁₈ chromatographic column as the stationary phase and a mixture of 0.006 mol·L^-1 disodium hydrogen phosphate-phosphoric acid solution (pH 6.5) and methanol in various ratios as mobile phase in gradient elution. Linear relationship between values of peak area and mass concentration of pseudohypericin was found in the range of 0.21-200.0 mg·L^-1, with detection limit (3S/N) of 0.040 mg·L^-1. Test for recovery was made by standard addition method, giving results in the range of 95.3%-101%, with RSDs (n=6) all less than 3.5%.