微波辅助衍生-UPLC-MS/MS联用技术及在手性药物分析中的应用

Sample pretreatment, which is time-consuming and laborious, has become the bottleneck of the separation and analysis procedure. The development of fast, effective and sensitive coupling technique of sample pretreatment and chromatography has become the frontier in the current analysis science. In this project, an on-line coupling technique based upon microwave-assisted derivatization and ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) is proposed to be established, and will be applied to the fast, highly sensitive and selective online analysis of chiral drugs. The realization of this online technique would not only solve the current chiral analysis problems such as long analysis time, low sensitivity, but also improve the selectivity of the achiral columns and solve the problem of the common achiral UPLC columns in separating the enantiomers of chiral drugs. This online system is realized by the online mixing, six-port valve switching and the column-head stacking, and microwave is used to accelerate the chiral derivatization. The features of the online system and the online analytical method would be systematically investigated and compared with the offline derivatization method and the method of separation with common chiral columns. This online system would be applied to the analysis of chiral drugs in biological samples. It is strongly believed that this research is of great importance for the rapid, high sensitive analysis of chiral drugs and the automation of the chiral analysis.

耗时、费力的样品前处理已经成为分离分析过程的瓶颈，发展快速、高效、灵敏的样品前处理与色谱分析联用技术成为当前分析科学领域的前沿课题。本项目拟发展一种微波辅助衍生-超高效液相色谱-二级质谱（UPLC-MS/MS）在线联用技术，并应用于手性药物的快速、高灵敏、高选择性在线分析。这一联用技术的实现不仅可以解决手性分析速度慢、灵敏度低的问题，提高手性分析的自动化程度，还可以提高非手性柱对手性对映体的选择性，解决普通非手性UPLC柱难以分离手性药物对映体的问题。该技术采用微波加速手性衍生化速率，通过在线混合、六通阀切换和柱头聚焦实现在线联用，并对联用系统和联用方法进行系统考察，同时与离线衍生化和手性柱分离的方法进行对比，最后应用于生物样品中手性药物的分离分析。本研究对于推动手性药物的高灵敏度快速检测、实现手性分析的自动化具有十分重要的意义。

生物样品中被测物质浓度低、成分复杂、基质干扰严重，其前处理往往耗时、费力，发展生物样品前处理与色谱质谱分析联用技术已经成为近年来分析科学领域关注的重点。本项目针对生物样品中的手性药物、药物残留以及抗精神病药物的分析测定，开展了基于样品前处理-UPLC-MS/MS联用技术的研发及其应用研究，主要内容如下： 1、研发了微波辅助衍生-UPLC-MS/MS联用装置，建立了高效、高灵敏度的β2-激动剂类手性药物班布特罗的手性分析方法，系统考察了微波辅助衍生-UPLC-MS/MS联用技术的特性。在微波辅助作用下，手性衍生化时间和色谱分离时间都仅需3 min，最低定量下限达2.5 pg/mL，回收率为90.1-93.0%，与普通加热衍生化方法和直接手性柱分离方法相比，本联用方法分析时间短、灵敏度高。该方法成功应用于β2-激动剂班布特罗的人体药代动力学研究和手性转化研究。2、研发了柱前稀释大体积进样UPLC-MS/MS联用系统，该在线系统可实现200 μL强溶剂直接进样，成功应用于24种药物残留的分析，定量限在0.00074-0.8 ng/kg之间，回收率范围为63.3-109%之间，RSD小于8.1%。与普通UPLC-MS/MS大体积进样进行了对比，本在线系统强溶剂大体积进样不影响色谱峰形，灵敏度和精密度显著提高。3、研发了SPE-UPLC-MS/MS联用系统，系统考察了该联用系统特性，成功应用于人血浆中10种抗精神病药物的分析，血浆样品离心后即可直接上样，分析时间仅为10 min，定量限为0.00321-2.75 μg/L，回收率在75.9-122%之间，与离线固相萃取结合UPLC-MS/MS的方法相比，本联用方法分析时间短、灵敏度高。以上研究工作分别在Journal of Chromatography A、Analytical and Bioanalytical Chemistry等刊物上发表，标注基金资助的SCI论文7篇，并申请了1项发明专利。