title: Semi-automated quantification of methylmalonic acid in human serum by LC-MS/MS.
abstract: BACKGROUND
  Methylmalonic acid (MMA), a sensitive biomarker of functional vitamin B12 deficiency, is commonly determined by gas chromatography-mass spectrometry (GC-MS) or liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods using manual extraction and derivatization of MMA to reduce polarity prior to separation.
  
  
  METHODS
  In the present study we introduce a semi-automated extraction on a strong anion exchanger, HPLC separation on a BEH-amide column to separate serum MMA from its abundant isoform, succinic acid, followed by MS/MS detection and quantification.
  
  
  RESULTS
  The extraction of MMA plus internal standard provides full recovery and the method is linear between 0.03 μmol/L and 20.0 μmol/L (r(2) = 1.0) with intra-and inter-assay imprecision of 2.2%. Agreement with other laboratories has been demonstrated in external proficiency testing. Compared to both conventional GC-MS and LC-MS/MS methods, the correlation is r(2) > 0.99.
  
  
  CONCLUSIONS
  The use of robotic pipetting, elimination of derivatization and improved separation by the BEH-amide column combined with HILIC chromatographic conditions significantly improve sample throughput compared to conventional methods. Using a single pipetting robot and LC-MS/MS instrument, this method is currently performing 180 analyses per day from 10 regional hospitals and several additional distant sites.
authors.affiliation: Clinical Chemistry, Laboratory Medicine of Skåne, Sweden.
authors.forename: Dick
authors.initials: D
authors.lastname: Nelson
authors.forename: Ning
authors.initials: N
authors.lastname: Xu
authors.forename: Joyce
authors.initials: J
authors.lastname: Carlson
mesh_terms: D001331:Automation; D002853:Chromatography, Liquid / Q000379:methods*; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008764:Methylmalonic Acid / Q000097:blood*; D015203:Reproducibility of Results
chemical_list: D008764:Methylmalonic Acid