UHPLC-MS/MS同时测定活血化瘀类中成药中非法添加的30个化学药物

Simultaneous Determination of 30 Compounds Illegally Added in Chinese Patent Medicine for Promoting Blood Circulation and Removing Blood Stasis Medicine by UHPLC-MS/MS

关键词:  活血化瘀  舒筋活血片  非法添加  抗心绞痛药  抗凝药  抗血小板药  抗炎镇痛药  超高效液相色谱-串联质谱法

Key words:  promoting blood circulation and removing blood stasis  Shujin Huoxue tablets  illegally added  antianginal drugs  anticoagulant  anti-platelet aggregators  anti-inflammatory analgesics  UHPLC-MS/MS

摘要:

目的 建立超高效液相色谱-串联质谱法同时测定活血化瘀类中成药中非法添加抗心绞痛、抗凝、抗血小板聚集以及抗炎镇痛等30个化学药物的方法。方法 样品采用60%甲醇超声提取，以0.1%甲酸溶液-乙腈为流动相进行梯度洗脱，经色谱柱Phenomenex Kinetex C₁₈(2.1 mm×100 mm，2.6 μm)分离，流速为0.3 mL·min^–1，柱温为40 ℃。质谱采用电喷雾电离，正负离子同时扫描，多反应监测的模式采集数据。结果 30个化合物在考察的浓度范围内线性关系良好，相关系数均>0.999，各化合物的检出限为1.3~238.4 ng·g^–1，平均回收率为63.7%~108.2%，RSD为1.0%~6.7%。采用该方法对90批舒筋活血片进行检测，共有3批检出对乙酰氨基酚，其余化合物均未检出。对乙酰氨基酚检出量为每片0.16~0.93 μg，远低于对应药品日服用的最小剂量，排除人为故意添加。可能的原因是企业共线生产清洁验证不彻底引起的残留。结论 该方法快速准确，灵敏度高，高选择性，可用于活血化瘀类中成药非法添加30个化合物的定性筛查和定量检测。

Abstract:

OBJECTIVE To establish an ultra-performance liquid chromatography-tandem mass spectrometry method for the simultaneous determination of 30 compounds(including antianginal drugs, anticoagulant, anti-platelet aggregators and anti-inflammatory analgesics) illegally added in Chinese patent medicine for promoting blood circulation and removing blood stasis. METHODS The sample was extracted by ultrasonication with 60% methanol. The analysis was performed on Phenomenex Kinetex C₁₈ column (2.1 mm×100 mm, 2.6 μm) with gradient elution using 0.1% formic acid solution and acetonitrile as the mobile phase at the flow rate of 0.3 mL·min^–1, and the column temperature was 40 ℃. Mass spectrometry was collected using electrospray ionization (ESI), simultaneous scanning of positive and negative ions, and multiple reaction monitoring mode. RESULTS The linearity of the 30 compounds was good in the mass range examined, with the correlation coefficients all >0.999. The limits of detection of each compound ranged from 1.3 to 238.4 ng·g^–1 and the average recoveries ranged from 63.7% to 108.2% with the RSDs of 1.0% to 6.7%. The method was used to test 90 batches of Shujin Huoxue tablets. Paracetamol was detected in 3 batches, while the rest of the compounds were not detected. The amount of paracetamol detected ranged from 0.16 to 0.93 μg per tablet, which was much lower than the minimum daily dose of the corresponding drug, excluding intentional addition. The probable explanation was the residues caused by incomplete cleaning validation of the enterprise’s collinear production. CONCLUSION This method is rapid, accurate, highly sensitive, and highly selective, and can be used for qualitative screening and quantitative determination of 30 chemical compounds illegally added in Chinese patent medicine for promoting blood circulation and removing blood stasis.