Phosphorescence for sensitive enantioselective detection in chiral capillary electrophoresis.

Enantioselective phosphorescence lifetime detection was combined with chiral cyclodextrin-based electrokinetic chromatography for the analysis of camphorquinone (CQ). A time-gated detection system based on a pulsed light-emitting diode for excitation at 465 nm was developed for the online lifetime determination. The background electrolyte for the chiral separation consisted of 20 mM alpha-cyclodextrin (alpha-CD), 10 mM carboxymethyl-beta-CD, and 25 mM borate buffer at pH 9.0. The separation of (+)-CQ and (-)-CQ is caused by a difference in association constants of these enantiomers with alpha-CD. Under the separation conditions, different phosphorescence lifetimes were obtained for (+)-CQ and (-)-CQ (tau = 384 +/- 8 and 143 +/- 5 micros, respectively), which could be used to distinguish the enantiomers. This selectivity in detection is based on a difference in protection of the enantiomers against phosphorescence quenching after their complexation with alpha-CD. Concentration detection limits were 2 x 10(-7) and 1 x 10(-6) M for (+)-CQ and (-)-CQ, respectively. After correction for the lifetime shortening by triplet-triplet annihilation at higher CQ concentrations, a linear dynamic range was obtained from the detection limit up to 2 mM. The system was used to determine the enantiomeric impurity levels of commercial samples of (+)-CQ and (-)-CQ; 0.2% and 0.1%, respectively.