Ultrafast determination of drugs in biological samples by column-switching LC/MS

茶碱和咖啡因是广泛摄取的天然生物碱，在人体中表现出多种药理作用。咖啡因是咖啡和其他饮料的成分，也包含在许多药物中。茶碱是一种支气管扩张剂，也是人体咖啡因的代谢物；由于茶碱的安全治疗剂量范围相对较窄，因此在法医毒理学领域偶尔会遇到茶碱中毒。采用气相色谱（GC）、气相色谱-质谱（MS）、高效液相色谱(LC)-质谱（MS）等方法对其进行了分析。在现有的药物分析方法中，通常需要对样品进行液-液萃取、固相萃取、过滤或离心等预处理，以去除生物基质中含有的蛋白质等杂质。耗时的多步骤样品预处理等分析方法不适用于人体标本中有毒物质的快速检测。为了避免这种样品前处理，开发了能够直接进样粗生物样品的专用高效液相色谱柱；我们的小组将它们应用于法医毒理学兴趣的化合物。此外，使用一种特殊的高效液相色谱柱的反冲洗切换系统已成为同时测定生物样品中各种药物的有力分析工具。在本研究中，我们建立了反冲洗柱切换和整体分离柱的高效液相色谱-质谱在线超快速测定人血清中茶碱和咖啡因的方法。据我们所知，本研究提出的方法是使用常规HPLC仪器测定人血清中茶碱和咖啡因的最快MS测定技术。

Theophylline and caffeine are natural alkaloids widely being ingested, and show various pharmacological effects in humans. Caffeine is a constituent of coffee and other beverages and included in many medicines. Theophylline is used as a bronchodilator and also formed as a metabolite of caffeine in humans ; theophylline poisoning is occasionally encountered in the field of forensic toxicology, because of its relatively narrow safe range of therapeutic doses. They have been analyzed by various methods, such as gas chromatography (GC), GC-mass spectrometry (MS), high-performance (HP) liquid chromatography (LC)-MS. In the existing analytical methods for the drugs, the sample pretreatments such as liquid-liquid extraction, solid-phase extraction, filtration or centrifugation were usually required to remove proteins and other impurities contained in biological matrices. The analytical methods including time-consuming multi-step sample pretreatments are unsuitable for rapid detection of poisonous materials in human specimens. To avoid such sample pretreatments, special HPLC columns, which enabled direct injection of crude biological samples, were developed ; our group applied them to compounds of forensic toxicological interest. Furthermore, backflush column-switching system using one of special HPLC columns have become a powerful analytical tool for simultaneous determinations of various drugs in biological samples. In this study, we have established an automated on-line method for ultra-fast determination of theophylline and caffeine in human serum by HPLC-MS with backflush column-switching and a monolithic separation column. To our knowledge, our method presented in this study is the fastest MS determination technique for theophylline and caffeine in human serum using the conventional HPLC instrument.