基于超高效液相色谱-四级杆/静电场轨道阱高分辨质谱的益心舒片中多种成分定量测定研究

Quantitative research of multiple active components in Yixinshu Tablet based on ultra performance liquid chromatography-quadrupole/orbitrap high resolution mass spectrometry

目的建立基于超高效液相色谱-四级杆/静电场轨道阱高分辨质谱的益心舒片中多成分(绿原酸、芦丁、咖啡酸、阿魏酸、迷迭香酸、丹酚酸B、丹酚酸A、丹参酮Ⅰ、隐丹参酮、欧当归内酯A、丹参酮Ⅱ_A)的定量分析方法。方法采用BEH C_(18)色谱柱(50 mm×2.1 mm,1.7μm),以乙腈-0.1%甲酸水为流动相,梯度洗脱,体积流量为0.2 m L/min;质谱采用ESI离子源,平行反应监测(PRM)的扫描方式进行检测。结果在优化的色谱质谱条件下,绿原酸、芦丁、咖啡酸、阿魏酸、迷迭香酸、丹酚酸B、丹酚酸A、丹参酮Ⅰ、隐丹参酮、欧当归内酯A、丹参酮Ⅱ_A分别在0.3～3.0、0.000 5～0.005 0、0.01～0.10、0.04～0.40、0.1～1.0、5.0～50.0、0.2～2.0、0.1～1.0、0.1～1.0、0.004～0.040、5.0～50.0μg/m L线性关系良好(r≥0.999 5);精密度、重复性及稳定性良好;加样回收率在99%～102%,RSD均小于3%;本研究所测成分在各批次样品中的质量分数依次为绿原酸217.80～502.25μg/g、芦丁0.50～0.59μg/g、咖啡酸8.45～13.54μg/g、阿魏酸2.61～3.56μg/g、迷迭香酸66.70～188.56μg/g、丹酚酸B 4 002.53～8 793.80μg/g、丹酚酸A 158.04～321.65μg/g、丹参酮Ⅰ 127.72～612.48μg/g、隐丹参酮75.56～285.70μg/g、欧当归内酯A 2.68～3.98μg/g、丹参酮Ⅱ_A 2 921.86～6 085.88μg/g。结论本实验建立的方法灵敏度高且准确性好,方法学考察结果符合测定要求,可用于益心舒片中多种活性成分的快速测定;并为其质量评价提供新的科学依据和参考。

Objective To establish a quantitative method of 11 active components （chlorogenic acid, rutin, caffeic acid, ferulic acid, rosmarinic acid, salvianolic acid B, salvianolic acid A, tanshinone I, cryptotanshinone, levistolide A, and tanshinone IIA） in Yixinshu Tablet based on ultra performance liquid chromatography-quadrupole/orbitrap high resolution mass spectrometry. Methods The column was BEH C^s （50 mmx 2.1 mm, 1.7 gm） and the mobile phase was consisted of acetonitrile-water （containing 0.1% formic acid） at a flow rate of 0.2 mL/min with gradient elution; Mass spectrometer conditions： electrospray ionization source （ESI） and parallel reaction monitoring （PRM） were used to perform the determination. Results Under the optimized conditions, chlorogenic acid, rutin, caffeic acid, femlic acid, rosmarinic acid, salvianolic acid B, salvianolic acid A, tanshinone I, cryptotanshinone, levistolide A, and tanshinone IIg all showed good liners （r 〉 0.999 5） in the ranges of 0.3--3.0, 0.000 5--0.005 0, 0.01--0.10, 0.04--0.40, 0.1--1.0,5.0--50.0, 0.2--2.0, 0.1--1.0, 0.1--1.0, 0.004--0.040, and 5.0--50.0μg/mL, respectively; The results of the accuracy, the repeatability and stability all reached the standards; The recoveries were ranged from 99%--102% and RSDs were below 3%; The results of the content in different batches were 217.80--502.25 μg/g （chlorogenic acid）, 0.50--0.59μg/g （rutin）, 8.45--13.54μg/g （caffeic acid）, 2.61--3.56 μg/g （ferulic acid）, 66.70--188.56 μg/g （rosmarinic acid）, 4 002.53--8 793.80 μg/g （salvianolic acid B）, 158.04--321.65 μg/g （salvianolic acid） A, 127.72--612.48 μg/g （tanshinone I）, 75.56--285.70 μg/g （cryptotanshinone）, 2.68--3.98 μg/g （levistolide A）, and 2 921.86--6 085.88 μg/g （tanshinone IIA, orderly. Conclusion The methods established are proved to have a high sensitivity and accuracy; The results of methodology can conform to the relevant requirements and the methods can have a rapid determination for the multiple active components in Yixinshu Tablet; The research also provides a new scientific basis and reference for the quality assessment at the same time.