Quantification of 1‐13C‐l‐methionine in rat serum with hydrophilic interaction liquid chromatography coupled with tandem mass spectrometry and its application in pharmacokinetic study

A rapid, selective and sensitive hydrophilic interaction liquid chromatography (HILIC) coupled with tandem mass spectrometry (MS/MS) method was developed to determine 1-13C-l-methionine in rat serum. Proteins in serum were precipitated using acetonitrile and the supernatant was separated after centrifugation. 1-13C-l-phenylalnine was used as the internal standard. HILIC–tandem mass spectrometry analysis was performed on a hydrophilic interaction silica column (TSK-GEL AMIDE-80) using a linear gradient elution system, acetonitrile−5 mm ammonium acetate containing 0.1% formic acid and multiple reaction monitoring mode for 1-13C-l-methionine and 1-13C-l-phenylalnine. The assay was validated with a linear range between 10 and 150 ng mL−1 (r ≥ 0.99) and a lower limit of quantification of 10 ng mL−1, calculated with weighted (1/x2) least squares linear regression. The RSD of intra-day precision was smaller than 3.6% and the inter-day RSD less than 6.5%, while the average recovery was 100.48% with an RSD of accuracy within 2.9%, determined from quality control samples. The HILIC-MS/MS method was fully validated and successfully applied to the in vivo pharmacokinetic study of stable-isotope 1-13C-l-methionine in rats. Copyright © 2011 John Wiley & Sons, Ltd.