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Semi-automated quantification of methylmalonic acid in human serum by LC-MS/MS

Nelson, Dick LU ; Xu, Ning LU and Carlson, Joyce LU (2012) In Scandinavian Journal of Clinical & Laboratory Investigation 72(6). p.441-446
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 mu mol/L and 20.0 mu mol/L (r(2) = 1.0) with intra-and... (More)
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 mu mol/L and 20.0 mu mol/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. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Chromatography, high performance liquid, tandem mass spectrometry, gas, chromatography-mass spectrometry, methylmalonic acid, vitamin B12
in
Scandinavian Journal of Clinical & Laboratory Investigation
volume
72
issue
6
pages
441 - 446
publisher
Informa Healthcare
external identifiers
  • wos:000308941400001
  • scopus:84866413491
ISSN
1502-7686
DOI
10.3109/00365513.2012.679963
language
English
LU publication?
yes
id
c3c52c62-c48e-4615-abd2-bf6850337e81 (old id 3190060)
date added to LUP
2012-12-03 07:10:14
date last changed
2017-09-10 03:54:19
@article{c3c52c62-c48e-4615-abd2-bf6850337e81,
  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 mu mol/L and 20.0 mu mol/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.},
  author       = {Nelson, Dick and Xu, Ning and Carlson, Joyce},
  issn         = {1502-7686},
  keyword      = {Chromatography,high performance liquid,tandem mass spectrometry,gas,chromatography-mass spectrometry,methylmalonic acid,vitamin B12},
  language     = {eng},
  number       = {6},
  pages        = {441--446},
  publisher    = {Informa Healthcare},
  series       = {Scandinavian Journal of Clinical & Laboratory Investigation},
  title        = {Semi-automated quantification of methylmalonic acid in human serum by LC-MS/MS},
  url          = {http://dx.doi.org/10.3109/00365513.2012.679963},
  volume       = {72},
  year         = {2012},
}