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When a truck becomes a motorcycle: The impact of sample load on a chiral capillary electrophoresis separation using mixtures of neutral and sulfated cyclodextrins

Chiral capillary electrophoresis (CE) separations are useful for monitoring the presence of a minor isomer at low levels (e.g., <0.5%) in the presence of the major form. In order to quantitate these low levels, it is necessary to inject large amounts of sample. Separations which appear to have more than enough resolution (“big enough to drive a truck through”) for dilute, equal‐concentration mixtures of isomers can become inadequately resolved when the necessary amount of sample is injected. This paper addresses some important considerations in maintaining adequate resolution at high sample loads for chiral separations involving a dual‐cyclodextrin (CD) system. For hydrophobic compounds, the use of both a neutral and a sulfated CD can be helpful in achieving a chiral separation. In such a system, the migration time and resolution can be controlled by varying the ratio of neutral to charged CD concentrations. It is demonstrated here that not only the ratio, but also the total CD concentration can significantly affect the separation. In this paper, the impact of the total CD concentration in a dual‐CD system (with the concentration ratio constant) is examined with respect to peak shape and resolution. The influences of temperature, capillary diameter, and current are also considered. The corresponding impact on the amount of sample which can be loaded and successfully separated determines the limit of quantitation of the minor isomer. Thus, this information is important in making such chiral separations applicable to determinations of low levels of minor isomer in the presence of large amounts of the major form.