Fast analytical-scale separations by capillary electrophoresis and liquid chromatography.

Separations are generally considered a slow step in any analytical methodology. HPLC and electrophoresis, the mainstay of separations of nonvolatile compounds in the modern laboratory, take anywhere from tens of minutes to hours to complete depending upon the complexity of the sample, the desired information, and selectivity available. This time requirement has limited the types of problems that separations can be asked to solve. For instance, separation methods are not typically used in highthroughput analyses or for monitoring applications. The time-consuming nature of separations has driven many researchers to investigate methods of improving the speed of separations techniques. In this review, we examine recent trends in improving the speed of liquid-phase analytical-scale separations based on chromatography and electrophoresis. While the present review is limited to liquid-phase separations, we note that recent advances have been made in the speed of gas chromatography separations as well, and this work has recently been reviewed elsewhere. Because many of the technical advances that have improved the speed of separations have occurred since 1990, our review emphasizes the past 9 years. In developing a review on fast separations, we were faced with the dilemma of how to define a fast separation. Our approach was to consider articles in which the emphasis was on use of theory and technology which were developed specifically to improve the speed of separation over conventional practice. Conventional practice, while difficult to define, in HPLC includes separations performed in 1-4.6 mm bore by 10-25 cm long columns packed with 3-5 μm porous particles. Thus, innovations such as nonporous particles, particle diameters <2 μm, perfused particles, elevated temperature, or open tubular columns and their application are discussed. For electrophoresis, we considered only capillary electrophoresis (CE). CE, by its very nature, is “faster” than gel electrophoresis; however, the rapid development of CE has meant that it would be impractical to simply review CE relative to gel electrophoresis. (Furthermore, exhaustive reviews of CE are available.) Rather, we consider innovations in CE that led to speed over conventional practice. For CE, conventional practice was defined as separations in columns with 50-100 μm bore and 50-100 cm length and electric field (E) < 300 V/cm. Use of miniaturized systems, rapid and automated injection schemes, high E, and certain gels and additives are innovations which can dramatically improve the speed of the separation and are included here. Microfabricated systems have been extensively utilized in achieving fast separations, especially for CE. A separate review on microfabricated systems for analysis and separation has been commissioned for this special issue of Chemical Reviews; therefore, while we acknowledge such advances, we limit our discussion of these important techniques relative to other innovations. Further information on microfabricated systems can be found in this other review for a more complete picture of fast separation systems. In addition to descriptions of the technical innovations, examples of the applications are included. We have emphasized applications which were made qualitatively possible by the improvement in speed. Such examples include on-line in vivo monitoring, using a fast separation as a second-dimension in twodimensional separations, detection of short-lived spe* To whom correspondence should be addressed. E-mail: rtkenn@ chem.ufl.edu. 3081 Chem. Rev. 1999, 99, 3081−3131