Highly Sensitive Analysis in Capillary Electrophoresis Using Large-volume Sample Stacking with an Electroosmotic Flow Pump Combined with Field-amplified Sample Injection.

Abstract

To achieve highly sensitive analysis without labor-intensive experimental procedures in capillary electrophoresis (CE), large-volume sample stacking with an electroosmotic flow pump (LVSEP)-field-amplified sample injection (FASI) was combined with a dynamic coating technique. In this study, poly(vinyl pyrrolidone) (PVP) was employed for the dynamic coating additive. Since a standard fluorescent dye, fluorescein, was well concentrated in a conventional LVSEP, the PVP dynamically-coated capillaries can be also applied to the LVSEP-FASI analysis. In our home-made CE apparatus, however, current breakdown was often caused, especially at a longer electrokinetic injection time due to bubble formation. To avoid the interference of bubble formation, the distance between the tips of the electrode and the capillary in the vertical direction was changed from 0 to 2.5 cm under the magnetic stirring condition. This allowed for a long electrokinetic injection time of up to 20 min, resulting in a sensitive enhancement factor (SEF) of 34900 for fluorescein. The developed method was applied to the chiral analysis of amino acids in CE. As a result, leucine (Leu) was successfully separated in LVSEP-FASI with SEFs of 6420 and 4500 for the D- and L-Leu peaks, respectively.