Assay of whole blood (6S)-5-CH3-H4folate using ultra performance liquid chromatography tandem mass spectrometry

被引:0
作者
Susanne H. Kirsch
Wolfgang Herrmann
Jürgen Geisel
Rima Obeid
机构
[1] Saarland University Hospital,Department of Clinical Chemistry and Laboratory Medicine
来源
Analytical and Bioanalytical Chemistry | 2012年 / 404卷
关键词
Folate; Red blood cells; 5-Methyltetrahydrofolate; Folic acid; Homocysteine; Tandem mass spectrometry;
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摘要
Folates act as essential coenzymes in many biological pathways, including the synthesis and methylation of DNA. Low folate concentration in serum and whole blood (WB) is associated with several disease conditions. We describe a stable-isotope-dilution ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the quantification of (6S)-5-CH3-H4folate (where H4folate is tetrahydrofolate) and non-CH3-H4folate [sum of HCO-H4folate, (6R)-5,10-CH+-H4folate, (6R)-5,10-CH2-H4folate, (6S)-H4folate, dihydrofolate, and folic acid] in WB. The assay includes a solid-phase extraction procedure after the hemolysis and deconjugation. The method was linear over the concentration range from 0.2 to 200 nmol/L. The limits of detection were 0.40 nmol/L or lower for the folate forms. The interassay coefficients of variation were 7.4 % for (6S)-5-CH3-H4folate and 15.4 % for non-CH3-H4folate. For the folate forms, the recoveries were between 97.1 % and 102.7 %. Sample preparation caused the generation of artificial folic acid in WB and serum in a dose-dependent manner, which can lead to misinterpretation of the results. The use of antioxidants could not prevent the formation of folic acid. The median fasting WB folate concentrations from 42 nonsupplemented and nonfortified adults were 576 nmol/L (6S)-5-CH3-H4folate and 73.6 nmol/L non-CH3-H4folate, and 1,206 nmol/L (6S)-5-CH3-H4folate and 155 nmol/L non-CH3-H4folate for 35 adults who had taken 500 μg of folic acid, 50 mg of vitamin B6, and 500 μg of vitamin B12 per day orally for 6 months. In conclusion, the UPLC-MS/MS method is fast and has a good sensitivity and selectivity for WB folates. We observed a dose-dependent oxidation of (6S)-H4folate, which resulted in the formation of artificial folic acid in serum and WB. To minimize this effect, we recommend a fast sample preparation.
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页码:895 / 902
页数:7
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  • [1] Fazili Z(2005)undefined Clin Chem 51 2318-2325
  • [2] Pfeiffer CM(2006)undefined Clin Chem Lab Med 44 450-459
  • [3] Zhang M(2008)undefined Rapid Commun Mass Spectrom 22 2403-2412
  • [4] Jain R(2010)undefined J Chromatogr B Anal Technol Biomed Life Sci 878 68-75
  • [5] Smith DE(2005)undefined Anal Chim Acta 553 36-42
  • [6] Kok RM(2006)undefined J Chromatogr B Anal Technol Biomed Life Sci 830 293-300
  • [7] Teerlink T(2004)undefined Clin Chem 50 2378-2381
  • [8] Jakobs C(2008)undefined Clin Chem 54 197-201
  • [9] Smulders YM(2010)undefined Anal Biochem 398 150-160
  • [10] Huang Y(1979)undefined J Food Sci 44 717-722