Measurement of adenosine by capillary zone electrophoresis with on-column isotachophoretic preconcentration

Abstract

An on-column isotachophoretic (ITP)-capillary electrophoresis (CE) system capable of preconcentrating polyhydroxyl species is reported. The ITP-CE system utilizes borate complexation of the neutral diol species to form anionic compounds that can be directly separated by CE. Borate buffer fuctions as both the terminating electrolyte for the ITP preconcentration and the operating buffer for the subsequent CE separation. Isotachophoretic preconcentration allows injection volumes as large as 50% of the column volume, without compromising separation integrity, to yield detection limits about 70-fold lower than direct CE separation (with borate operating buffer). In this paper we also present an application of the ITP-CE system, with laser-induced fluorescence (LIF) detection, to the quantitative analysis of adenosine from urine. Nanomolar concentration levels of adenosine are successfully derivatized with chloroacetaldehyde (CAA) to form a fluorescent derivative whose spectral characteristics match the HeCd laser. The technique is shown to be capable of quantitative measurement of adenosine as low as 10 −9 M, the levels expected in plasma and urine.