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Clinical Chemistry and Laboratory Medicine (CCLM)

Published in Association with the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM)

Editor-in-Chief: Plebani, Mario

Ed. by Gillery, Philippe / Greaves, Ronda / Lackner, Karl J. / Lippi, Giuseppe / Melichar, Bohuslav / Payne, Deborah A. / Schlattmann, Peter


IMPACT FACTOR 2018: 3.638

CiteScore 2018: 2.44

SCImago Journal Rank (SJR) 2018: 1.191
Source Normalized Impact per Paper (SNIP) 2018: 1.205

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1437-4331
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Volume 44, Issue 4

Issues

Quantitative determination of erythrocyte folate vitamer distribution by liquid chromatography-tandem mass spectrometry

Desirée E.C. Smith
  • Department of Clinical Chemistry, ICaR-VU, VU University Medical Center, Amsterdam, The Netherlands
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Robert M. Kok
  • Department of Clinical Chemistry, ICaR-VU, VU University Medical Center, Amsterdam, The Netherlands
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Tom Teerlink
  • Department of Clinical Chemistry, ICaR-VU, VU University Medical Center, Amsterdam, The Netherlands
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Cornelis Jakobs
  • Department of Clinical Chemistry, ICaR-VU, VU University Medical Center, Amsterdam, The Netherlands
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Yvo M. Smulders
  • Department of Internal Medicine, ICaR-VU, VU University Medical Center, Amsterdam, The Netherlands and Institute for Cardiovascular Research, ICaR-VU, VU University Medical Center, Amsterdam, The Netherlands
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
Published Online: 2011-09-21 | DOI: https://doi.org/10.1515/CCLM.2006.085

Abstract

Background: Given the role of folate in many disorders, intracellular distribution of folate vitamers is of potential clinical importance. In particular, accumulation of non-methyltetrahydrofolates due to altered partitioning of folate metabolism at the level of methylenetetrahydrofolate is of interest.

Methods: We describe a positive-electrospray liquid chromatography tandem mass spectrometry (LC-MS/MS) method that allows determination of erythrocyte folate vitamer distribution by accurately measuring both 5-methyltetrahydrofolate (5-methylTHF) and non-methyl folate vitamers. Whole blood lysates are deconjugated in ascorbic acid solutions, deproteinized, purified using folate-binding protein affinity columns, concentrated by solid-phase extraction (SPE) and evaporation, and separated on a C18 column within 6min.

Results: The limit of quantification for both 5-methylTHF and non-methylTHF was 0.4nmol/L (signal-to-noise >10). Intra- and inter-assay CVs for 5-methylTHF were 1.2% and 2.8%, respectively. Intra- and inter-assay CVs for non-methylTHF as a group were 1.6% and 1.5%, respectively. Recovery results were 97–107%. We measured 8–72% non-methyl folate vitamers in volunteers (n=5) with the methylenetetrahydrofolate reductase (MTHFR) 677 TT genotype. Concentrations ranged from 117 to 327nmol/L and 23 to 363nmol/L for 5-methylTHF and non-methylTHF vitamers, respectively. We measured 0–2% non-methylTHF vitamers in MTHFR 677 CC genotype volunteers. In addition, we found that storage of whole-blood samples in ascorbic acid at low pH resulted in 53–90% loss of the non-methylTHF fraction.

Conclusion: This LC-MS/MS method accurately determines erythrocyte 5-methylTHF and non-methyl folate vitamers.

Keywords: erythrocyte; folate; folate vitamer; liquid chromatography; mass spectrometry

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About the article

Corresponding author: Dr. Yvo M. Smulders, Department of Internal Medicine, VU University Medical Center, P.O. Box 7057, 1007 MB Amsterdam, The Netherlands Phone: +31-20-4444307, Fax: +31-20-4444313


Received: 2005-11-08

Accepted: 2006-01-16

Published Online: 2011-09-21

Published in Print: 2006-04-01


Citation Information: Clinical Chemistry and Laboratory Medicine (CCLM), Volume 44, Issue 4, Pages 450–459, ISSN (Online) 1437-4331, ISSN (Print) 1434-6621, DOI: https://doi.org/10.1515/CCLM.2006.085.

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