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A HPLC-DAD-MS/MS Method for Simultaneous Determination of Six Active Ingredients of Salviae Miltiorrhizae and Ligustrazine Hydrochloride Injection in Rat Plasma and its Application to Pharmacokinetic Studies

Author(s):

Jianyang Pan, Luquan Zhang, Difeifei Xiong, Bailing Li and Haibin Qu*   Pages 1 - 10 ( 10 )

Abstract:


Aims: Pharmacokinetic Study of Salviae Miltiorrhizaeand Ligustrazine Hydrochloride injection. For the evaluation of mechanism of action, safety and clinical rational use of Salviae Miltiorrhizae and Ligustrazine Hydrochloride injection.

Background: Salviae Miltiorrhizae and Ligustrazine Hydrochloride injection is a compound preparation consisted of Salvia Miltiorrhiza extract and ligustrazine hydrochloride for the treatment of cardiovascular and cerebrovascular diseases in China.

Objective: To develop a rapid and sensitive high-performance liquid chromatography-diode array detector-tandem mass spectrometry (HPLC-DAD-MS/MS) method for simultaneous determination of six major active ingredients of Salviae Miltiorrhizae and Ligustrazine Hydrochloride injection, namely danshensu, protocatechuic aldehyde, rosmarinic acid, lithospermic acid, salvianolic acid A, and ligustrazine hydrochloride, in rat plasma.

Methods: Plasma samples were precipitated with methanol, which was spiked with ascorbic acid and the supernatant was separated on a Waters Cortecs C18 column, by using a gradient mobile phase system of acetonitrile-water containing 0.05% formic acid (v/v). For internal standards, puerarin was selected for the five salvianolic acids, while isofraxidin was used for ligustrazine hydrochloride. Besides, electrospray ionization in negative mode and multiple-reaction monitoring were used to identify and quantify the five salvianolic acids, whereas ligustrazine hydrochloride was quantified at 310 nm using the diode array detector.

Results: Noticeably, all calibration curves showed good linearity (R2>0.99) over the concentration range, with a lower limit of quantification between 0.00411 and 0.0369μg/mL for salvianolic acids, and 1.74 μg/mL for ligustrazine hydrochloride. Next, the precision of the developed method was evaluated by intra-and inter-day assays, and the percentage of relative standard deviation was within 10%. Although the extraction efficiency of some salvianolic acids were not very satisfactory, the sensitivity of the analytical method met the analysis requirements of rat plasma samples. Moreover, the validated method was successfully applied to a pharmacokinetic study of Salviae Miltiorrhizae and Ligustrazine Hydrochloride injection in the rat model.

Conclusion: Linear pharmacokinetic characteristics were observed for the six active ingredients after intravenous infusion administration in rats, within the dose range examined here. In summary, our study proposed a HPLC-DAD-MS/MS method in the simultaneous determination of multiple ingredients, and demonstrated its applicability in pharmacokinetic studies.

Keywords:

HPLC-DAD-MS/MS, Pharmaco kinetic, Salviae Miltiorrhizae and Ligustrazine Hydrochloride Injection, Danshensu, Ligustrazine hydrochloride

Affiliation:

Pharmaceutical Informatics Institute, College of Pharmaceutical Sciences, Zhejiang University, 866#, Yuhangtang Road, Hangzhou,310058, Guizhou Baite Pharmaceutical Co., Ltd., Guiyang, 550014, Guizhou Baite Pharmaceutical Co., Ltd., Guiyang, 550014, Guizhou Baite Pharmaceutical Co., Ltd., Guiyang, 550014, Pharmaceutical Informatics Institute, College of Pharmaceutical Sciences, Zhejiang University, 866#, Yuhangtang Road, Hangzhou,310058



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