Some chiral compounds show over- loading at lower concentrations than achiral compounds. One reason is the different amounts of cyclodextrin (5- 50%) dissolved in the stationary phase. Unlike the classical fronting peaks of normal stationary phases, the characteristic of an overloaded peak on cyclodextrin stationary phases is indicated by a tailing peak. Overloading chiral compounds results in loss of resolution, even when column capacity has not been exceeded. The first example is linalool separation on the Rt-βDEXsp (Figure 1). The amount is about 20 ng on-column (Figure 1A). Figure 1B shows the same components at a higher concentration of 200 ng on-column. Note that the linalool enantiomers are beginning to tail, and there is a small loss in chiral resolution. Even though the maximum sample capacity for 0.32mm ID capillary columns is normally 400- 500 ng per component, the peak shapes of chiral compounds indicate overload at one-third of the sample amount. Again, there is much less cyclodextrin for which a chiral compound can interact. Figure 1C shows pronounced overloading at 2 µg on-column. Extreme tailing and complete loss in resolution are the result.
Figure 1: Overloading of linalool on Rt-βDEXsp
The second example is 2,3-butanediol on Rt-βDEXsm (Figure 2). We see here a similar trend of overloading and tailing between 500 ng on-column (Fig 2A) and 25 ng on-column (Fig 2B). Due to a very good separation between the isomers, the loss of resolution isn’t as obvious as it was in the previous example, even though we can see more overloading.
Figure 2: Overloading of butanediol on Rt-βDEXsm