毛细管电泳-无鞘流电喷雾质谱用于药物分析

doi:10.3724/SP.J.1123.2022.07015

摘要/Abstract

摘要：

毛细管电泳-质谱联用技术具有分离效率高、检测灵敏度高、样品消耗量少,可同时提供样品的结构信息等优点,成为复杂样品分离分析的强有力工具。但是,毛细管电泳与质谱联用的接口技术依然未能很好的解决。为了拓展我们发展的金箔包裹的毛细管电泳分离柱尖端直接作为喷雾电极和无鞘流质谱接口的应用,本文报道了用无鞘流接口毛细管电泳-电喷雾质谱联用(CE-ESI-MS)分析5种酪氨酸激酶抑制剂(舒尼替尼、甲磺酸伊马替尼、吉非替尼、达沙替尼、埃罗替尼)的研究结果。这种接口集分离与电喷雾离子化于一根毛细管中,制作简单,成本低廉,且可批量制作。实验发现采用非水毛细管电泳分离模式不仅可以对5种酪氨酸激酶抑制剂实现基线分离,而且可以获得稳定的质谱信号。考察了电解质溶液组成对分离效果的影响,得到优化的背景电解质组成,即含2%(v/v)乙酸及5 mmol/L乙酸铵的乙腈-甲醇(80:20, v/v)混合溶剂。在优化的条件下,5种激酶抑制剂可以得到基线分离,无鞘接口也可以长时间保持稳定的电喷雾,分析物的保留时间日内、日间重复性(RSD值)分别小于0.5%和0.8%,接口批次间的RSD值小于2.6%。与水相分离条件下的CE-MS对比,非水相条件下的5种酪氨酸激酶抑制剂的分离柱效更高,检测灵敏度更高,绝对检出限达到amol级。此外,采用无鞘流CE-MS分析了各类有机酸(千层纸素A、丹酚酸C和迷迭香酸)和脂溶性的大环内酯类抗生素(阿奇霉素、红霉素和环孢素A),均可以获得良好的分离效果和质谱检测结果。

关键词: 无鞘流接口, 非水毛细管电泳-质谱, 金箔包裹的电喷雾电极, 酪氨酸激酶抑制剂

Abstract:

Capillary electrophoresis-mass spectrometry (CE-MS) combines the advantages of capillary electrophoresis, such as the high separation efficiency and low sample consumption, and the high detection sensitivity of mass spectrometry and the ability for providing the structural information for structure elucidation of unknown components. However, the interface technology for coupling capillary electrophoresis and mass spectrometry is still not well resolved. In the present work, we explored the application of the sheathless CE-MS interface which was prepared by gold foil-wrapped CE separation column tip directly as a spray electrode for the analysis of five tyrosine kinase inhibitors, namely sunitinib, imatinib mesylate, gefitinib, dasatinib and erlotinib. This interface integrates separation and electrospray ionization in one capillary, which is easy to manufacture, low in cost, and can be produced in batches. We found that using the nonaqueous CE separation mode can not only achieve baseline separation of five tyrosine kinase inhibitors, but also obtain stable mass spectrometry signals. First, we investigated the effect of the electrolyte solution composition on the separation. The optimized background electrolyte composition was obtained: 2% (v/v) acetic acid and 5 mmol/L ammonium acetate in acetonitrile-methanol (80:20, v/v). Under optimized conditions, the five kinase inhibitors could be baseline separated, meantime, the sheathless interface could also maintain stable electrospray for a long time. The relative standard deviation (RSD) values of the intraday and interday repeatability of the analyte retention times were less than 0.5% and 0.8%, respectively, and the RSD value between interface batches is less than 2.6%. Compared with CE-MS with aqueous phase, the separation column efficiency of the five tyrosine kinase inhibitors under nonaqueous phase conditions is higher, the detection sensitivity is higher, and the absolute detection limit reaches amol level. In addition, we evaluated the sheathless interface with various organic acids, such as palaflin A, salvianolic acid C, and rosmarinic acid, as well as hydrophobic macrolide antibiotics, azithromycin, erythromycin, and sporin A, good separation effect and mass spectrometric detection results can be obtained.

Key words: sheathless interface, nonaqueous capillary electrophoresis-mass spectrometry (NACE-MS), gold foil-wrapped electrospray ionization emitter, tyrosine kinase inhibitors

中图分类号:

O658

张含智, 李凤, 康经武. 毛细管电泳-无鞘流电喷雾质谱用于药物分析[J]. 色谱, 2023, 41(2): 160-167.

ZHANG Hanzhi, LI Feng, KANG Jingwu. Coupling capillary electrophoresis-electrospray ionization mass spectrometry with sheathless interface for drug analysis[J]. Chinese Journal of Chromatography, 2023, 41(2): 160-167.

图/表 7

图1 5种酪氨酸激酶抑制剂的化学结构式

Fig. 1 Chemical structures of the five tyrosine kinase inhibitors

图2 (a)无鞘流NACE-ESI-MS系统示意图及(b)氢氟酸刻蚀方法得到的对称型石英毛细管喷针的SEM图

Fig. 2 (a) Schematic diagrams of sheathless interface and NACE-ESI-MS system and (b) SEM image of the symmetrical fused silica capillary emitter observed by HF acid etching method NACE-ESI-MS: nonaqueous capillary electrophoresis-electrospray ionization mass spectrometry.

图3 (a)甲醇、(b)乙酸及(c)乙酸铵含量对NACE分离5种酪氨酸激酶抑制剂的影响

Fig. 3 Effect of (a) methanol,(b) acetic acid and (c) ammonium acetate contents on the NACE separation for the five tyrosine kinase inhibitors Background electrolyte (BGE): a. acetonitrile-methanol containing 5 mmol/L ammonium acetate and 2%(v/v) acetic acid; b. acetonitrile-methanol (80:20, v/v) containing 5 mmol/L ammonium acetate and acetic acid; c. acetonitrile-methanol (80:20, v/v) containing 2%(v/v) acetic acid and ammonium acetate. Peak identifications: 1. sunitinib; 2. imatinib; 3. gefitinib; 4. dasatinib; 5. erlotinib.

图4 (a)NACE-ESI-MS与(b)水相CE-ESI-MS分离5种激酶抑制剂的电泳图

Fig. 4 Electropherograms of the five kinase inhibitors by (a) NACE-ESI-MS and (b) aqueous CE-ESI-MS Conditions: capillary, 70 cm×50 μm I. D., 360 μm O. D.; applied separation voltage, 12 kV; ESI voltage, 2.2 kV; hydrodynamic injection, 1 kPa×2.0 s. BGE: a. 2%(v/v) acetic acid and 5 mmol/L ammonium acetate in acetonitrile-methanol (80:20, v/v); b. 20 mmol/L ammonium formate in 50% acetonitrile aqueous solution (pH 3.0). The mass concentrations of analytes are all 10 μg/mL. Peak identifications are the same as those in Fig. 3.

表1 5种酪氨酸激酶抑制剂的检出限及迁移时间的重复性

Table 1 LODs of the five tyrosine kinase inhibitors and repeatabilities(RSDs)of migration time

Compound	LOD/ amol	RSDs of migration time
Compound	LOD/ amol	Intraday (n=3)	Interday (n=3)	Batch-to-batch (n=3)
Sunitinib	370	0.48	0.75	2.5
Imatinib	298	0.42	0.27	1.5
Gefitinib	330	0.21	0.32	1.3
Dasatinib	302	0.34	0.51	1.3
Erlotinib	374	0.50	0.74	1.7

图5 NACE-ESI-MS分离酸类化合物的电泳图及质谱图

Fig. 5 Electropherogram and mass spectra of acid compounds separation by NACE-ESI-MS

图6 NACE-ESI-MS分离抗生素类化合物的电泳图和质谱图

Fig. 6 Electropherogram and mass spectra of antibiotic compounds separation by NACE-ESI-MS

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