Factors Influencing Cyclodextrin-Mediated Chiral Separations - Chiral Separations by Capillary Electrophoresis - page 94

derivatives of amino acids using S- -CD in the negative polarity mode [8]. The most

critical factor affecting the enantioseparation of the studied analytes was found to
be the BGE pH. As can be seen in Figure 4.3, a pH increase led to a complete loss
of CBI-ser enantiomers resolution, the migration times were not strongly modii ed.
A similar trend was observed for two other analytes (CBI-ala and CBI-glu) while
for CBI-arg, the opposite effect was observed. The ionization of CBI-amino acids
carboxylic groups depends on pH. CBI-ser having a p K a value of 3, at pH 2, is
mainly present in uncharged form and the enantiomers were completely resolved.
On the other hand, at pH 3, the unionized and the anionic forms equally coexist
and there was no enantioresolution. It is worth noting that, despite the fact that both
the analyte and the chiral selector exhibit (at least partially) a negative charge at
pH 3, CBI-ser strongly interacts with S-

β

β -CD. Indeed, at pH 3, CBI-ser migration

time in the absence of S- β -CD is much longer than that observed in the presence

of 2% S- -CD is strongly affected by the

analyte ionization state. The opposite trend observed for CBI-arg can be related to
the solute charge. In contrast to the other studied analytes, CBI-arg carries a posi-
tive charge over the investigated pH range (due to the protonation of its guanidine
group). Therefore, at pH 2, this analyte is positively charged and will be in zwit-
terionic form at higher pH.
The pH also determines the magnitude and the direction of the EOF which sig-
nii cantly inl uence the apparent selectivity and the enantioresolution. As illustrated
in the review of Rizzi, the optimal selector concentration is shifted to slightly lower
values with a comigrating EOF and to slightly higher values in the case of a weak
countermigrating EOF [3]. A reversal of migration order and a shift of the maximum
concentration to lower values are observed in the presence of a strong countermi-
grating EOF. If the value of the countermigrating EOF is comprised between the

β -CD. Therefore, the complex analyte/S- β

R s = 0

pH 3.00

R s = 0.78

pH 2.50

R s = 1.28

pH 2.25

R s = 1.72

pH 2.00

550 575 600

Migration time, s

625 650 675

FIGURE 4.3 Inl uence of BGE pH on chiral resolution of cyanobenz[ f ]isoindole-ser. BGE:
2% S-β-CD, 25 mM phosphate. (Reproduced from Kirschner, D.L. et al., Anal. Chem ., 79,
736, 2007. With permission.)