Simultaneous Determination of the Five Marker Compounds in Melandrium firmum using High-Performance Liquid Chromatography with Photodiode-Array Detection

In Korean folk medicine, Melandrium firmum Rohrbach (Caryophyllaceae) has been used to treat various diseases, including anuria, breast cancer, and gonorrhea. In this study, a high-performance liquid chromatography method with photodiode-array UV detection (HPLC-PDA) was established and then validated for the simultaneous determination of five flavonoids in M. firmum: schaftoside, homoorientin, cytisoside, vitexin, and isovitexin. Separation of the five compounds was performed on a Gemini C18 column at 40°C by gradient elution of two mobile phases. The flow rate and injection volume were 1.0 mL/min and 10 μL, respectively. The HPLC-PDA method showed excellent linear regression with r2 values of 0.9999 within the test ranges. The limits of detection and quantification of all components were 0.01–0.11 and 0.04–0.34 μg/mL, respectively. The extraction recovery of the five analytes was 97.6—105.6% and the RSD values were < 2.0% The relative standard deviation values of intra- and interday precision for all analytes were < 1.71 and < 1.46%, respectively. As determined using the established and validated HPLC-PDA method described here, the amounts of the five marker compounds in M firmum were 0.02–2.54 mg/g.