Simultaneous multi-component quantitation of Chinese herbal injection Yin-zhi-huang in rat plasma by using a single-tube extraction procedure for mass spectrometry-based pharmacokinetic measurement

• A new “SMIPE” procedure developed for straightforward bio-sample preparation.
• One-pot multifunction impurity removal for matrix-matched external standard calibration.
• Mass spectrometric multiple reaction monitoring of Yin-zhi-huang herbal prescription in vivo.
• Simultaneous multi-component pharmacokinetic study of Yin-zhi-huang injection for the first time.

Ying-zhi-huang injection (YZH-I) is an injectable multi-herbal prescription derived from the ancient Chinese remedy “Yin-chen-hao-tang”, which is widely used in the clinic for the treatment of jaundice and chronic liver diseases. To date, little information is available on the pharmacokinetic properties of this poly-herbal formulation. Herein, we reported a simple, rapid and sensitive liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for quantitative multiple reaction monitoring (MRM) of eight major ingredients of YZH-I (including baicalin, baicalein, wogonoside, geniposide, geniposidic acid, chlorogenic acid, neochlorogenic acid, and caffeic acid) in rat plasma. A fast single-tube multi-impurity precipitation extraction (“SMIPE”) procedure was introduced for straightforward plasma preparation, based on one-pot deproteinization precipitation with acidified methanol extraction and in-situ multifunction impurity removal by a solid sorbent mixture (anh. magnesium sulfate plus octadecylsilane). Particularly, the addition of ascorbic acid in methanol (10 mg/mL) was found to exhibit a pronounced protective effect and significantly increase extraction effectiveness of the herbal phenolic components. Some pretreatment variables (protein precipitating solvent, acidifying agent and sorbent) were optimized with acceptable matrix effect (−18 to 7.7%), extraction recovery (65–88%) and process efficiency (62–91%) for the SMIPE-based LC-MRM multi-analyte quantitation using matrix-matched calibration (5–1000 ng/mL) without using internal standard. Mean accuracies were obtained in the range of 83–114% at three different fortification levels, with intra- and inter-day variations within 13%. This validated method was successfully applied to the simultaneous measurement and pharmacokinetic investigation of the chemical constituents in rats following an intravenous administration of YZH-I.