Institut für Analytische Chemie, Universität Leipzig, Linnéstraße 3, 04103 Leipzig, Germany.
Anal Chem. 2013 Sep 3;85(17):8150-7. doi: 10.1021/ac4010937. Epub 2013 Aug 22.
In this study, we introduce time-resolved fluorescence detection with two-photon excitation at 532 nm for label-free analyte determination in microchip electrophoresis. In the developed method, information about analyte fluorescence lifetimes is collected by time-correlated single-photon counting, improving reliable peak assignment in electrophoretic separations. The determined limits of detection for serotonin, propranolol, and tryptophan were 51, 37, and 280 nM, respectively, using microfluidic chips made of fused silica. Applying two-photon excitation microchip separations and label-free detection could also be performed in borosilicate glass chips demonstrating the potential for label-free fluorescence detection in non-UV-transparent devices. Microchip electrophoresis with two-photon excited fluorescence detection was then applied for analyses of active compounds in plant extracts. Harmala alkaloids present in methanolic plant extracts from Peganum harmala could be separated within seconds and detected with on-the-fly determination of fluorescence lifetimes.
在这项研究中，我们引入了使用 532nm 双光子激发的时间分辨荧光检测，用于微流控芯片电泳中的无标记物分析物测定。在开发的方法中，通过时间相关的单光子计数收集关于分析物荧光寿命的信息，从而改善电泳分离中可靠的峰分配。使用熔融石英制成的微流控芯片，分别确定了血清素、普萘洛尔和色氨酸的检测限为 51、37 和 280nM。应用双光子激发微芯片分离和无标记物检测还可以在硼硅酸盐玻璃芯片中进行，这证明了在非紫外透明器件中进行无标记荧光检测的潜力。然后，将双光子激发荧光检测的微芯片电泳用于植物提取物中活性化合物的分析。从 Peganum harmala 的甲醇植物提取物中存在的哈尔马生物碱可以在几秒钟内分离出来，并通过荧光寿命的实时测定进行检测。
