Comprehensive Two-Dimensional Separations with an Electrodriven Component

6 COMPREHENSIVE TWO-DIMENSIONAL SEPARATIONS WITH AN ELECTRODRIVEN COMPONENT 

The most successful multidimensional electrodriven separations to date have been performed by James Jorgenson and his group at the University of North Carolina at Chapel Hill. This group has accomplished several successful comprehensive 

7 MICROCOLUMN REVERSE PHASE HIGH PEREORMANCE LIQUID CHROMATOGRAPHY-CAPILLARY ZONE ELECTROPHORESIS 

In 1990, Bushey and Jorgenson developed the first automated system that eoupled HPLC with CZE (19). This orthogonal separation teehnique used differenees in hydrophobieity in the first dimension and moleeular eharge in the seeond dimension for the analysis of peptide mixtures. The LC separation employed a gradient at 20 p.L/min volumetrie flow rate, with a eolumn of 1.0 mm ID. The effluent from the ehromatographie eolumn filled a 10 p.L loop on a eomputer-eontrolled, six-port miero valve. At fixed intervals, the loop material was flushed over the anode end of the CZE eapillary, allowing eleetrokinetie injeetions to be made into the seeond dimension from the first. 

The HPLC elution time was typieally under 260 min, and the CZE analysis took plaee in 60 s, whieh led to an overall run time of about 4 h. The 1 min CZE sampling interval was problematie, as the LC eolumn was probably slightly undersampled. A shorter CZE analysis time, whieh would provide a more frequent sampling rate, would improve this system a great deal. The seeond-dimension analysis time must be short relative to the first dimension, lest resolution in the first dimension be saeri-fieed. 

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