Two-dimensional fluorescence correlation in capillary electrophoresis for peak resolution and species identification

A new spectroscopic dimension-fluorescence intensity correlation--is introduced to enhance peak resolution and species identification in capillary electrophoresis. In two-dimensional correlation CE, a conventional electropherogram is spread into two dimensions through cross-correlation analysis of fluorescence time response. A laser that is sinusoidally modulated in intensity is used as the excitation source. Three channels of information are collected during a CE run: the steady-state intensity, the ac amplitude, and the phase-resolved fluorescence intensity. The correlation between two chosen channels is then evaluated. A two-dimensional correlation electropherogram consists of a plot of the correlation intensity versus two axes of migration time. Through correlation analysis, species discrimination and peak resolution are significantly enhanced without having to physically separate the solutes. Two-dimensional correlation CE showed complete resolution between two overlapping sample peaks with a resolution of 0.28 in the conventional one-dimensional electropherogram. In separations of polycyclic aromatic hydrocarbons by micellar electrokinetic chromatography (MEKC), two-dimensional correlation analysis resolved all overlapping elution peaks unseparable by one-dimensional MEKC, demonstrating the utility of 2D correlation in separation method development. The capability of 2D correlation CE in species identification is demonstrated with a sequence of 39 consecutively injected peaks containing four fluorescent dyes. Species identification in sequencing is achieved without complex data treatment in two-dimensional correlation CE.