ABSTRACT
BACKGROUND: We optimized a quantitative amino acid method with pre-column derivatization, norvaline (nva) internal standard and reverse phase ultra-performance liquid chromatography by replacing the ultraviolet detector with a single quadrupole mass spectrometer (MSnva). METHOD: We used 13C15N isotopically labeled amino acid internal standards and a C18 column with 1.6µm particles to optimize the chromatography and to confirm separation of isobaric compounds (MSlis). We compared the analytical performance of MSnva with MSlis and the original method (UVnva) with clinical samples. RESULTS: The chromatography time per sample of MSnva was 8min, detection capabilities were <1µmol/L for most components, intermediate imprecisions at low concentrations were <10% and there was negligible carryover. MSnva was linear up to a total amino acid concentration in a sample of approximately 9500µmol/L. The agreements between most individual amino acids were satisfactory compared to UVnva with the latter prone to outliers and suboptimal quantitation of urinary arginine, aspartate, glutamate and methionine. MSnva reliably detected argnininosuccinate, ß-alanine, citrulline and cysteine-s-sulfate. CONCLUSION: MSnva resulted in a more than fivefold increase in operational efficiency with accurate detection of amino acids and metabolic intermediates in clinical samples.
