Pharmaceutical Informatics Institute, College of Pharmaceutical Sciences, Zhejiang University, 866, Yuhangtang Road, Hangzhou, 310058, China.
Guizhou Baite Pharmaceutical Co., Ltd., Guiyang, 550014, China.
Curr Drug Metab. 2021;22(1):60-69. doi: 10.2174/1389200221999200819143230.
This is a pharmacokinetic study of Salviae miltiorrhizae and ligustrazine hydrochloride injection. The study aimed to evaluate the mechanism of action, safety and rational clinical use of Salviae miltiorrhizae and ligustrazine hydrochloride injection.
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.
The study aimed 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.
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 multiplereaction monitoring were used to identify and quantify the five salvianolic acids, whereas ligustrazine hydrochloride was quantified at 310 nm using the diode array detector.
Noticeably, all calibration curves showed good linearity (R2>0.99) over the concentration range, with a lower limit of quantification between 0.00411 and 0.0369 μg/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 was 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.
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-DADMS/ MS method with the simultaneous determination of multiple ingredients, and demonstrated its applicability in pharmacokinetic studies.
本研究旨在对丹参川芎嗪注射液进行药代动力学研究，旨在评估丹参川芎嗪注射液的作用机制、安全性和临床合理应用。
丹参川芎嗪注射液是一种复方制剂，由丹参提取物和盐酸川芎嗪组成，用于治疗中国的心血管和脑血管疾病。
本研究旨在建立一种快速灵敏的高效液相色谱-二极管阵列检测器-串联质谱法（HPLC-DAD-MS/MS），用于同时测定丹参川芎嗪注射液中 6 种主要活性成分的含量，即丹参素、原儿茶醛、迷迭香酸、紫草酸、丹酚酸 A 和盐酸川芎嗪。
采用甲醇沉淀法处理血浆样品，以抗坏血酸为内标，上清液在 Waters Cortecs C18 柱上进行梯度洗脱，流动相为乙腈-水（含 0.05%甲酸，v/v）。对于 5 种丹酚酸，选择葛根素作为内标，而对于盐酸川芎嗪，则选择异嗪皮啶作为内标。此外，采用电喷雾离子化负离子模式和多反应监测模式进行鉴定和定量分析，二极管阵列检测器用于定量检测盐酸川芎嗪。
所有校准曲线在浓度范围内均显示出良好的线性关系（R2>0.99），丹酚酸的定量下限在 0.00411 至 0.0369 μg/mL 之间，盐酸川芎嗪的定量下限为 1.74 μg/mL。接下来，通过日内和日间试验评估了所建立方法的精密度，相对标准偏差的百分比在 10%以内。虽然某些丹酚酸的提取效率不是很理想，但分析方法的灵敏度满足了大鼠血浆样品分析的要求。此外，该验证方法成功应用于丹参川芎嗪注射液在大鼠体内的药代动力学研究。
在本研究考察的剂量范围内，静脉注射后 6 种活性成分呈现线性药代动力学特征。综上所述，本研究建立了一种同时测定多种成分的 HPLC-DAD-MS/MS 方法，并证明了其在药代动力学研究中的适用性。
