Quantitative determination of vitamin D metabolites in plasma using UHPLC-MS/MS |
| |
Authors: | Shujing Ding Inez Schoenmakers Kerry Jones Albert Koulman Ann Prentice Dietrich A Volmer |
| |
Institution: | 1. Medical Research Council–Human Nutrition Research, Elsie Widdowson Laboratory, Fulbourn Road, Cambridge, CB1 9NL, UK 2. Medical Research Council–Keneba Field Site, P. O. Box 273, Banjul, Keneba, The Gambia 3. Institute of Bioanalytical Chemistry, Saarland University, Campus B2 2, Raum 2.27, 66123, Saarbrücken, Germany
|
| |
Abstract: | Vitamin D is an important determinant of bone health at all ages. The plasma concentrations of 25-hydroxy vitamin D (25-OH
D) and other metabolites are used as biomarkers for vitamin sufficiency and function. To allow for the simultaneous determination
of five vitamin D metabolites, 25-OH D3, 25-OH D2, 24,25-(OH)2 D3, 1,25-(OH)2 D3, and 1,25-(OH)2 D2, in low volumes of human plasma, an assay using ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS)
was established. Plasma samples were spiked with isotope-labeled internal standards and pretreated using protein precipitation,
solid-phase extraction (SPE) and a Diels–Alder derivatization step with 4-phenyl-1,2,4-triazoline-3,5-dione. The SPE recovery
rates ranged from 55% to 85%, depending on the vitamin D metabolite; the total sample run time was <5 min. Mass spectrometry
was conducted using positive ion electrospray ionization in the multiple reaction monitoring mode on a quadrupole–quadrupole-linear
ion trap instrument after pre-column addition of methylamine to increase the ionization efficiency. The intra- and inter-day
relative standard deviations were 1.6–4.1% and 3.7–6.8%, respectively. The limit of quantitation for these compounds was determined
to be between 10 and 20 pg/mL. The 25-OH D results were compared with values obtained for reference materials (DEQAS). In
addition, plasma samples were analyzed with two additional Diasorin antibody assays. All comparisons with conventional methods
showed excellent correlations (r
2 = 0.9738) for DEQAS samples, demonstrating the high degree of comparability of the new UHPLC-MS/MS technique to existing
methods. |
| |
Keywords: | |
本文献已被 SpringerLink 等数据库收录! |
|