Cyclodextrin-Mediated Enantiomeric Separation of Idelalisib: A Validated Capillary Electrophoresis and NMR Study

Idelalisib (IDE) is a marketed chiral anticancer drug administered as the S-enantiomer, requiring sensitive monitoring of the R-enantiomer to ensure enantiomeric purity. However, no dedicated capillary electrophoresis (CE) method has been reported for trace-level quantification of R-IDE. In this study, a cyclodextrin-mediated CE method was developed for reliable detection of the R-enantiomer at the 0.1% level (LOD 2 µg/mL; LOQ 5 µg/mL). Systematic screening identified hydroxypropyl-β-cyclodextrin (HP-β-CD) with an intermediate degree of substitution (DS~6.8) as the optimal chiral selector, providing efficient enantioseparation (Rs up to 4.3). The method was validated according to ICH Q2(R2) guidelines, demonstrating suitable precision, accuracy, and robustness. Complementary NMR studies revealed hindered rotation of the 3-phenyl moiety and elucidated the molecular basis of enantioselectivity. Complexation with β-CD and HP-β-CD produced clear diastereomeric differentiation in both 1H and 19F NMR spectra, while the simplified 19F NMR profiles enabled direct enantiomer discrimination. NOESY and ROESY experiments demonstrated distinct inclusion modes, with HP-β-CD accommodating both the fluorinated aromatic ring and the 3-phenyl moiety. These interactions may account for the superior enantioseparation observed with HP-β-CD of intermediate DS. Our validated CE method addresses the distomer determination while NMR insights provide mechanistic understanding of the chiral recognition.