Simultaneous determination of ten Aconitum alkaloids in rat tissues by UHPLCMS/MS and its application to a tissue distribution study on the compatibility of Heishunpian and Fritillariae thunbergii Bulbus

Abstract A rapid, sensitive and selective ultra-high performance liquid chromatography with tandem mass spectrometry (UHPLC⿿MS/MS) method was developed and validated for simultaneous determination of ten Aconitum alkaloids in rat tissues. The tissue samples were prepared by a simple procedure protein precipitation with acetonitrile containing 0.1% acetic acid and separated on an Agilent XDB C18 column (4.6 mm y 50 mm, 1.8 μm) using gradient elution with a mobile phase consisting of water and acetonitrile (both containing 0.1% formic acid) at a flow rate of 0.3 mL/min. The quantitive determination was performed on an electrospray ionization (ESI) triple quadrupole tandem mass spectrometer using selective reaction monitoring (SRM) under positive ionization mode. The established method was fully validated according to the USA Food and Drug Administration (FDA) bioanalytical method validation guidance and the results demonstrated that the method was sensitive and selective with the lowest limits of quantification (LLOQ) at 0.025 ng/mL in rat tissue homogenates. Meanwhile, the linearity, precision, accuracy, extraction recovery, matrix effect and stability were all within the required limits of biological sample analysis. After method validation, the validated method was successfully applied to the tissue distribution study on the compatibility of Heishunpian (HSP, the processed product of Aconitum carmichaelii Debx) and Fritillariae thunbergii Bulbus (Zhebeimu, ZBM). The results indicated that the distribution feature of monoester diterpenoid aconitines (MDAs), diester diterpenoid aconitines (DDAs) and non-ester alkaloids (NEAs) were inconsistency, and the compatibility of HSP and ZBM resulted in the distribution amount of DDAs increased in tissues. What⿿s more, the results could provide the reliable basis for systematic research on the substance foundation of the compatibility of the herbal pair.