Determination of human reference values for serum total 1, 25-dihydroxyvitamin D using an extensively validated 2D ID-UPLC–MS/MS method. Issue 164 (November 2016)

• Record Type: Journal Article
• Authors: Dirks, Niek F.; Martens, Frans; Vanderschueren, Dirk; Billen, Jaak; Pauwels, Steven; Ackermans, Mariëtte T.; Endert, Erik; Heijer, Martin den; Blankenstein, Marinus A.; Heijboer, Annemieke C.
• Is Part Of: Journal of steroid biochemistry and molecular biology. Issue 164(2016)
• Page Start: 127
• Page End: 133
• Publication Date: 2016-11
• Subjects: DEQAS vitamin D External Quality Assessment Scheme -- 25OHD 25-hydroxyvitamin D -- 24, 25diOHD 24, 25-dihydroxyvitamin D -- 1, 25diOHD 1, 25-dihydroxyvitamin D -- VDR vitamin D receptor -- RIA radioimmunoassay -- 2D ID-UPLC–MS/MS two-dimensional isotope dilution ultra pressure liquid chromatography–tandem mass spectrometry -- FA formic acid -- BSA bovine serum albumin -- PBS phosphate-buffered saline -- PTAD 4-phenyl-1, 2, 4-triazole-3, 5-dione -- CV coefficient of variation -- LOQ limit of quantitation -- SNR signal-to-noise ratio -- OCP oral contraceptive; LC–MS/MS -- 1, 25-dihydroxyvitamin D -- Reference values -- Comparison; Steroid hormones -- Periodicals; Biochemistry -- Periodicals; Hormones -- Periodicals; Molecular Biology -- Periodicals; Hormones stéroïdes -- Périodiques; Steroid hormones; Periodicals; 572.579
• DOI: 10.1016/j.jsbmb.2015.12.003
• Languages: English
• ISSNs: 0960-0760

Abstract

Highlights: We have developed a LC–MS/MS method able to measure both 1, 25dihydroxyvitamin D3 and D2 . The method was extensively validated with regard to sensitivity, specificity and robustness and correlated well with another published LC–MS/MS method. Human reference values for 1, 25-dihydroxyvitamin D3 and D2 were 59–159 pmol/L and <17 pmol/L, respectively. Premenopausal women taking oral contraceptive pills show significantly higher 1, 25-dihydroxyvitamin D3 values compared to postmenopausal women. Abstract: Background: To assess a patient's vitamin D status the precursor metabolite 25-hydroxyvitamin D can be determined. However, measurement of 1, 25-dihydroxyvitamin D is required when disorders of 1a-hydroxylation, extrarenal 1a-hydroxylation, or vitamin D receptor defects are suspected. Methods: The aim of this study was to determine reference values for 1, 25-dihydroxyvitamin D3 and D2 using a 2D ID-UPLC–MS/MS method. Results: The LC–MS/MS method, able to measure picomolar concentrations of both 1, 25-dihydroxyvitamin D3 and D2 in human serum, was extensively validated. Intra-assay variations were <5% and 8.5% and <7.5% and 11%, for 1, 25-dihydroxyvitamin D3 and D2, respectively, over the whole dynamic range (3.1–376 and 3.1–652 pmol/L). Limit of quantitation was 3.4 pmol/L for both compounds. Our method correlated well with a published LC–MS/MS method ( r = 0.87) and with the average 1, 25-dihydroxyvitamin D3 results of the vitamin D External Quality Assessment Scheme (DEQAS) determined with LC–MS/MS ( r = 0.93). Reference ranges, determined in 96 plasma samples of healthy volunteers were 59–159 pmol/L and <17 pmol/L for respectively 1, 25-dihydroxyvitamin D3 and D2 . The female part of the reference group showed a statistically significant decrease of 1, 25-dihydroxyvitamin D3 concentrations with age. The presence of significantly higher average 1, 25-dihydroxyvitamin D3 levels in premenopausal women taking oral contraceptive pills compared to postmenopausal women suggests that this effect is estrogen-related, as estrogens lead to a higher vitamin D binding protein. Conclusions: The major finding of the present study is a reference interval of 59–159 pmol/L for 1, 25-dihydroxyvitamin D3 determined with a highly sensitive and precise LC–MS/MS method.