Automated Assay for the Determination of Methylmalonic Acid, Total Homocysteine, and Related Amino Acids in Human Serum or Plasma (original) (raw)

2014

Abstract

lonic acid (MMA) and total homocysteine (tHcy) in serum or plasma is useful in diagnosing and distin-guishing between cobalamin and folate deficiencies. We developed and validated an isotope-dilution gas chro-matography–mass spectrometry (GC-MS) method with automated sample workup for the determination of MMA, tHcy, and the related amino acids Met, total cysteine (tCys), Ser, and Gly in serum or plasma. Methods: Serum or plasma samples (100 L) were treated with a reductant (dithioerythritol), deprotein-ized with ethanol, and derivatized and extracted in a single step by the addition of methylchloroformate and toluene. All liquid handling was performed in 96-well (1 mL) microtiter plates by a robotic workstation. The N(S)-methoxycarbonyl ethyl ester derivatives were ana-

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