Automated coupling of capillary-HPLC to matrix-assisted laser desorption ionization mass spectrometry for the analysis of small molecules utilizing a reactive matrix
Automated coupling of capillary-HPLC to matrix-assisted laser desorption ionization mass spectrometry for the analysis of small molecules utilizing a reactive matrix
复制标题
DOI：
10.1007/s00216-003-2024-9
复制
发表时间：
2003-07-01
影响因子：
4.3
通讯作者：
Volmer, DA
中科院分区：
文献类型：
--
作者：
Brombacher, S;Owen, SJ;Volmer, DA
本研究描述了一种新型反应基质在毛细管-高效液相色谱-基质辅助激光解吸电离(MALDI)联用分析类固醇化合物中的应用。类固醇的质谱分析是在色谱峰全自动沉积到MALDI目标上后完成的。作为受试化合物的七种皮质类固醇是：曲安奈德、泼尼松、可的松、氟可的松、地塞米松、脱氧皮质酮和布地奈德。采用PepMap C-18色谱柱(粒径3微米，孔径100埃)，分离浓度分别为25、15、7.5、2.5和1 ng/MU，检测波长237 nm。柱流出物与2，4-二硝基苯肼(DNPH)直接混合，在UV检测器下进行检测。然后，利用由紫外光探测器信号触发的机器人微量收集器，将色谱峰沉积到MALDI目标上。一种特殊的疏水表面涂层允许在一个样点上沉积多达4微升(高达色谱峰体积的90%)。用MALDI质谱仪对化合物进行了鉴定。根据分析物的性质，自由基阳离子([M](+.))在正离子模式下检测类固醇的钠加成离子([M+Na](+))以及质子化的类固醇-二硝基苯肼衍生物([M-D+H](+))。毛细管进样时，MALDI-TOF-MS的检出限为0.5~15 ng，手动沉淀时，MALDI-TOF的检出限为0.3~3 ng。
This study describes the application of a novel, reactive matrix for the mass spectral analysis of steroids by capillary-high performance liquid chromatography (capillary-HPLC) coupled to matrix-assisted laser desorption/ionization (MALDI). The mass spectral analysis of steroids was accomplished after fully automated peak deposition of chromatographic peaks onto MALDI targets. The seven corticosteroids used as test compounds were: triamcinolone, prednisone, cortisone, fludrocortisone, dexamethasone, deoxycorticosterone, and budesonide. They were separated using a PepMap C-18 (3 mum particle size, 100 Angstrom pore width) column at five different concentration levels of 25, 15, 7.5, 2.5 and 1 ng/muL, and the peaks were detected at a wavelength of 237 nm. The column effluent was mixed with 2,4-dinitrophenylhydrazine (DNPH) directly following the UV detector. The chromatographic peaks were then deposited onto the MALDI target with a robotic micro-fraction collector triggered by the UV detector signals. A special hydrophobic surface coating allowed the deposition of up to 4 muL (up to 90 % of the chromatographic peak volume) onto one sample spot. The compounds were then identified by MALDI mass spectrometry. Depending on the nature of the analyte, radical cations ([M](+.)) and sodium adduct ions ([M+Na](+)) of the steroids as well as protonated steroid-dinitrophenylhydrazone derivatives ([M-D+H](+)) were detected in positive ion mode. The detection limits were between 0.5 and 15 ng injected with capillary-HPLC-MALDI-TOF-MS and between 0.3 and 3 ng on target with MALDI-TOF when deposited manually.