Pharmacokinetic comparison of 18 constituents from Qingxin Zishen Prescription in normal and ovariectomized rats via a validated HPLC-MS/MS method.

Qingxin Zishen Prescription (QZP) is an empirical formula for treating menopausal syndrome, yet the pharmacokinetic behavior of its constituents under pathological conditions remains poorly understood. In the present work, we developed and validated a sensitive high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method to simultaneously quantify 18 representative bioactive constituents of QZP in rat plasma, and applied it to a comparative pharmacokinetic study in normal and ovariectomized (OVX) rats. Sample preparation involved protein precipitation with acetonitrile followed by concentration. Separation was achieved on an Agilent ZORBAX SB C18 column (4.6 ×100 mm, 1.8 μm), and detection was performed on a triple quadrupole mass spectrometer in both positive and negative modes. The method showed excellent linearity (r > 0.9965) with lower limits of quantification ranging from 0.015 to 25 ng/mL. The within-run and between-run precision (relative standard deviation, RSD) were within 8.58% and 10.37%, respectively, and the accuracy ranged from 85% to 115% at all quality control levels. Matrix effects ranged from 1.98% to 14.57% (RSD), and extraction recoveries ranged from 85.54% to 113.29%, both within acceptable limits. Stability under different storage conditions was also verified. The comparative pharmacokinetic study revealed that the t1/2 of rhynchophylline, corynoxeine, hirsuteine, magnoflorine, and catalpol were significantly prolonged in OVX rats, whereas that of demethyleneberberine was significantly shortened. In addition, hirsuteine showed significantly increased Cmax and AUC0-t, suggesting enhanced absorption in the pathological state. The results clarify the systemic exposure behavior of the main bioactive components of QZP and highlight marked pharmacokinetic differences between healthy and OVX rats. These findings indicate that estrogen deficiency significantly modulates the elimination and systemic exposure of multiple QZP constituents, suggesting that dose adjustment may be necessary when administering herbal medicines to menopausal patients.