We present high resolution fast, cost-effective and sensitive Capillary zone electrophoresis (CZE) methods for determination of enantiomeric compounds of Kynurenine pathway, i.e. D, L-Kynurenine (KYN), in human serum and urine samples by cationic-β-CD and its synergistic dual chiral selector system (SD-CSs) with α-CD in 50 mM borax borate buffer (pH 9.0) as BGE. Acid-mediated stacking enrichment by HCl delivered 15 nM limit of detection (LOD) and 50 nM limit of quantification (LOQ). The methods gave advantages of linearity in the concentration range of 50 nM-1000 nM, reproducibility (RSD ≤ 3.35), selectivity against interfering amino acids, and remarkable recoveries. SD-CSs delivered resolution of D, L-KYN twice that of individual chiral selectors (CSs) under similar conditions. The binding constants (Kb) and electrophoretic mobilities (µeff) of D, L-KYN with different concentrations of CSs were calculated to find the migration order of enantiomers. The chiral recognition mechanism was investigated by molecular docking and molecular mechanics, which revealed strong hydrogen bonding between Kynurenine enantiomers and the SD-CSs as compared to individual CS as the key player in binding, formation of stable complexes which led to the ultimate separation.
Keywords: Capillary electrophoresis; D, L-Kynurenine; Kynurenine enantiomers; Stacking.
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