Development and validation of a HPLC-UV-ESI-MS method for the simultaneous quantitation of ten bioactive compounds in Dahuang Fuzi Tang

AIM:To develop a high performance liquid chromatography(HPLC) coupled with electrospray ionization mass spectrometry(ESI-MS) and ultraviolet(UV) detector method for the acid-alkaline simultaneous determination of ten bioactive compounds,and analyze the effect of compatible medicinal plants on the concentration of components in Dahuang Fuzi Tang(DFT).METHOD:The chromatographic separation was performed on a Hypersil BDS C18 analytical column by gradient elution with acetonitrile and formate buffer(containing 0.15% formic acid,V/V) at 25℃ with a flow rate of 1.0 mLmin-1 and UV detection at 280 nm.Four of the ten compounds in DFT were identified and their MS fragments were elucidated by HPLC-ESI-MS,and the contents of the six compounds were determined by HPLC-UV.RESULTS:All calibration curves showed good linear regression(r20.9990).The limits of detection and limits of quantification were 0.021-0.155 gmL-1 and 0.076-0.520 gmL-1,respectively.Overall precision RSD(intra-day and inter-day) were less than 2.96%,and the average recoveries were 98.35%-101.45%,with RSD ranging from 1.54% to 3.01% for the analytes.CONCLUSION:The developed method can be applied for the quality control and provide analytical evidence on the chemical basis and combinational principles of DFT.