Simultaneous determination of the enantiomers of leucine and [H7]leucine in plasma by capillary gas chromatography–mass spectrometry

A method for the stereoselective assay of - and -enantiomers of both leucine and ²H₇]leucine in rat plasma was developed using gas chromatography–mass spectrometry–selected-ion monitoring. -²H₃]leucine was used as an internal standard. The method involved purification by cation-exchange chromatography using BondElut SCX cartridge and derivatization with hydrochloric acid in methanol to form methyl ester followed by subsequent chiral derivatization with (+)-α-methoxy-α-trifluoromethylphenylacetyl chloride to form diastereomeric amide. The derivatization made the separation of the leucine enantiomers possible with good gas chromatographic behavior. Quantitation was performed by selected-ion monitoring of the quasi-molecular ions of the diastereomers on the chemical ionization method. The sensitivity, specificity, accuracy and reproducibility of the method were demonstrated to be satisfactory for application to pharmacokinetic studies of leucine enantiomers.