Quantitative Assessment of the Influence of Rhizoma Zingiberis on the Level of Aconitine in Rat Gut Sacs and Qualitative Analysis of the Major Influencing Components of Rhizoma Zingiberis on Aconitine Using UPLC/MS.

Yang Xin,Shuying Liu,Andrew C Gill

doi:10.1371/journal.pone.0124110

Abstract

This study attempted to clarify the material basis for the detoxification of Rhizoma Zingiberis (RZ) on aconitine, an analgesic drug, by quantitatively assessing the influence of RZ on the in vitro intestinal concentration of aconitine using an everted gut sac model and by qualitatively identifying the components in the RZ extract. To quantify aconitine in rat everted gut sacs, both an accurate processing method and a sensitive detection method were required. We developed a three-step sample processing method to protect the components from decomposition and applied ultra-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UPLC/TQMS) to quantify aconitine, glucose and digoxin. In addition, ultra-performance liquid chromatography coupled with linear ion trap mass spectrometry (UPLC/ITMS) was applied to detect the potential antidotal components in the RZ extract. Finally, the RZ extract reduced the level of aconitine in everted gut sacs, and eleven gingerols were successfully identified, which could be considered potential antidotal components for aconitine. This study demonstrated the application of two UPLC/MS methods for analyzing the material basis for the reciprocity between Chinese medicine components in everted gut sacs.

Highlights

The P1 value of aconitine for each gut sac decreased after the Rhizoma Zingiberis extract was added, and the decrease in the duodenum was most significant, which can be observed from its lowest ER value. These results indicate that the Rhizoma Zingiberis extract reduced the level of aconitine in the rat gut sacs, and this reduction was greatest in the duodenum; the mechanism could not be elucidated from the above results
The findings demonstrate that aconitine is a potential substrate of intestinal P-gp and that the Rhizoma Zingiberis extract is a potential revulsant of P-glycoprotin; in addition, the Rhizoma Zingiberis extract reduced the permeation of aconitine by inducing P-glycoprotein, and gingerols were the main Rhizoma Zingiberis components responsible for the effect of Rhizoma Zingiberis on aconitine transport in the intestine
Large numbers of phenols that contain hydroxy groups are present in the molecular structures of Rhizoma Zingiberis extract components; these hydroxy groups competed with aconitine to form hydrogen bonds with P-gp

Summary

Methods

The reference standards for aconitine and 6-gingerol were purchased from the Institute for the Control of Pharmaceutical and Biological Products of China (Beijing, China). Rhizoma Zingiberis was purchased from the Jilin Pharmacy (Changchun, China). Verapamil and digoxin were purchased from the Sigma Chemical Co. Methanol and acetonitrile were of HPLC grade. Distilled water was prepared using a Millipore water purification system. Aconitine solutions were prepared with a high concentration, an intermediate concentration and a low concentration of 0.3, 0.03 and 0.003 mg/mL, respectively. A digoxin solution was prepared with a concentration of 0.8 mg/mL. A verapamil solution was prepared with a concentration of 1.0 mg/mL. A 10 mL aqueous extract of 10 g Rhizoma Zingiberis was prepared using the decocting method

Results

Discussion

Conclusion