Single-drop microextraction (SDME) was in-line coupled with capillary electrophoresis-mass spectrometry to provide sample cleanup and enrichment simultaneously. Since there is no outlet vial in a conventional capillary electrophoresis-electrospray ionization-mass spectrometry (CE-ESI-MS) configuration, it is not easy to hang a single drop in the capillary inlet for extraction. We overcame the difficulty of coupling SDME and CE-MS by using a temporary outlet reservoir. Basic drugs such as methamphetamine, amphetamine, phenethylamine, methoxyphenamine, and mephentermine were extracted from a basic sample solution to an acidic acceptor drop covered with a thin octanol layer formed at the capillary inlet tip. Compared to the CE-MS method in the multiple reaction monitoring (MRM) mode, the in-line SDME-CE-MS/MS technique showed 130 similar to 150-fold enrichment in 10 min. The relative standard deviations (RSDs) of peak height ranged from 9 to 13 %. RSDs can be reduced from 4 to 6 % using mephentermine as an internal standard. We examined the pretreatment of sample with and without SDME from human urine under the full-scan mode, which confirmed that many metabolites were cleaned up by the selective extraction method of SDME. Even if the analytes from human urine were analyzed under the MRM mode used as a mass filter, there was an isobaric compound causing a disturbance to the analysis. However, in-line SDME-CE-MS/MS made it possible to perform a sample cleanup as well as sample enrichment. The research is extremely advantageous in that it is rapid, convenient, and highly sensitive for the analysis of biological samples using a commercially available instrument.
