QUANTITATIVE ANALYSIS OF TEN DITERPENOIDS IN RAT BILE AFTER ORAL ADMINISTRATION OF Isodon rubescens EXTRACT BY HIGH PERFORMANCE LIQUID CHROMATOGRAPHY-ELECTROSPRAY IONIZATION TANDEM MASS SPECTROMETRY

Abstract

Diterpenoids are the primary bioactive ingredients in Isodon rubescens named Dong Ling Cao in China which possess many pharmacological activities, including anti-tumor, anti-microbial, anti-inflammatory, anti-catastrophe, anti-oxidation, and anti-inflammatory effects. We employed a sensitive and selective high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry (HPLC–ESI–MS/MS) method for the quantification of ten diterpenoids in rat bile. The ten diterpenoids are effusanin A (1), lasiodonin (2), oridonin (3), epinodosin (4), nodosin (5), ponicidin (6), rabdoternin A (7), enmenol (8), lasiokaurin (9), and lasiokaurinol (10). The sulfamethoxazole was used as internal standard (IS). The bile samples were pretreated by liquid-liquid extraction with ethyl acetate (EtOAc). Chromatographic separation was performed on a C₁₈ column with linear gradient elution using water and methanol at a flow rate of 0.8 mL/min. The precursor and product ions of analytes were monitored on a hybrid quadrupole linear ion trap mass spectrometer equipped with a turbo ion spray interface in positive and negative mode, respectively, using multiple–reaction monitoring (MRM). The specificity, linearity, accuracy, precision, recovery, matrix effect, and several of stabilities have been validated for diterpenoids in rat bile samples. The results showed that this method is robust, specific, and sensitive and it can successfully fulfill the requirement of excretion study of the ten diterpenoids in Isodon rubescens.

Keywords:

• bile
• diterpenoids
• HPLC-ESI-MS
• Isodon rubescens
• MRM

ACKNOWLEDGMENT

This work received financial support from the Natural Science Foundation of Hebei Province- CSPC Pharmaceutical Joint Research Fund, China (C2011206174).