目的 建立含阿胶中成药阿胶补血口服液、阿归养血颗粒中阿胶类皮源检测方法,包括阿胶的专属性检测方法,以及猪皮源、马皮源、牛皮源掺伪检查研究。方法 采用胰蛋白酶对用0.01 g·mL-1NH4HCO3溶解的阿归养血颗粒及阿胶补血口服液中阿胶进行酶解,利用超高效液相色谱-三重四极杆质谱(UPLC-QqQ-MS)对样品进行检测。色谱柱为 Waters ACQUITY UPLC HSS T3-C18(2.1 mm×100 mm,1.8 μm),流动相为体积分数0.1%甲酸溶液(A)-乙腈(B),梯度洗脱(0~25 min,95%A→80%A;25~40 min,80%A→50%A),流速0.3 mL·min-1,柱温30 ℃,进样量5 μL;离子化模式为ESI+,进行多反应监测,选择阿胶特征分子离子峰 m/z 539.8(双电荷)→612.4和m/z 539.8(双电荷)→923.8;马源寡肽A特征分子离子峰m/z 386.4(双电荷)→322.3和m/z 386.4(双电荷)→377.3;新阿胶特征分子离子峰m/z 774.5(双电荷)→799.8和m/z 774.5(双电荷)→1 034.6;黄明胶特征分子离子峰m/z 641.3(双电荷)→726.2和m/z 641.3(双电荷)→783.3作为检测离子对。结果 10批市售阿归养血颗粒和10批阿胶补血口服液样品中均可检出阿胶的特征肽段,其中的3批阿归养血颗粒样品和2批阿胶补血口服液样品中同时检出了马皮特征肽。结论 所建立的方法经方法学验证,专属性强,可用于中成药中阿胶的检测及掺伪检查。
Abstract
OBJECTIVE To establish an analysis method for detection of animal-based gelatin in Chinese patent medicines like Ejiaobuxue oral liquid and Eguiyangxue granules, including the specific identification of dongkey-hide gelatin contained in Chinese patent medicines and the examination of pig-hide gelatin,horse-hide gelatin and bovine-hide gelatin aduteration. METHODS Samples were dissolved in water containing 0.01 g·mL-1NH4HCO3 and digested by trypsin. Ultra-high performance liquid chromatography(UPLC)coupled with triple quadruple mass spectrometry(QqQ-MS)was used to detect the samples. Determination was carried out with the application of Waters ACQUITY UPLC HSS T3-C18(2.1 mm×100 mm,1.8 μm) column at 30 ℃. The mobile phase was composed of water containing 0.1% formic acid(A)and acetonitrile(B). Gradient elution(0-25 min,95%A→80%A;25-40 min,80%A→50%A) was conducted at a flow rate of 0.3 mL·min-1. Electrospray ionization(ESI) was performed in multiple reaction monitoring(MRM) mode with the transitions of m/z 539.8(double charge)→612.4,m/z 539.8(double charge)→923.8,m/z 386.4(double charge)→322.3 and m/z 386.4(double charge)→377.3, m/z 774.5(double charge)→799.8 and m/z 774.5(double charge)→1 034.6, m/z 641.3(double charge)→726.2 and m/z 641.3(double charge)→783.3. RESULTS The characteristic peptides of donkey-hide gelatin could be detected in 20 batches of samples collected from market. Horse skin marker peptide was detected in 3 batches of Eguiyangxue granules and 2 batches of Ejiaobuxue oral liquid. CONCLUSION The established method is specific, which is suitable for identification of donkey-hide gelatin in Chinese patent medicine.
关键词
阿归养血颗粒 /
阿胶补血口服液 /
特征肽段 /
专属性鉴别 /
超高效液相色谱-三重四极杆质谱
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Key words
Eguiyangxue Granula /
Ejiaobuxue oral liquid /
marker peptide /
specificity identification /
UPLC-QqQ-MS
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中图分类号：
R284
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脚注
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基金
国家药品监督管理局中药材及饮片质量控制重点实验室项目资助(2020GSMPA-KL15)
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