Flynn, N;
Lam, F;
Dawnay, A;
(2014)
Enhanced 3-epi-25-hydroxyvitamin D3 signal leads to overestimation of its concentration and amplifies interference in 25-hydroxyvitamin D LC-MS/MS assays.
Annals of Clinical Biochemistry: An international journal of biochemistry and laboratory medicine
, 51
(3)
352 - 359.
10.1177/0004563213497691.
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Abstract
Background 3-epi-25-hydroxyvitamin D3 (3-epi-25OHD3) interferes in most liquid chromatography-tandem mass spectrometry (LC-MS/MS) assays for 25-hydroxyvitamin D (25OHD). The clinical significance of this is unclear, with concentrations from undetectable to 230 nmol/L reported. Many studies have quantified 3-epi-25OHD3 based on 25OHD3 calibrators or other indirect methods, and we speculated that this contributes to the observed variability in reported 3-epi-25OHD3 concentrations. Methods We compared continuous MS/MS infusions of 3-epi-25OHD3 and 25OHD3 solutions, spiked both analytes into the same serum matrix and analysed patient samples to assess the effect of three different quantitation methods on 3-epi-25OHD3 concentration. Experiments were performed on an LC-MS/MS system using a phenyl column which does not resolve 3-epi-25OHD3, and a modified method utilizing a Zorbax SB-CN column that chromatographically resolves 3-epi-25OHD3 from 25OHD3. Results A greater 3-epi-25OHD3 signal, compared with 25OHD3, was observed during equimolar post-column continuous infusion of analyte solutions, and following analysis of a serum pool spiked with both analytes. 3-epi-25OHD3 signal enhancement was dependent on mobile phase composition. Compared with 3-epi-25OHD3 calibrators, indirect quantitation methods resulted in up to 10 times as many samples having 3-epi-25OHD3 concentrations ≥ 10 nmol/L, and an approximately fourfold increase in the maximum observed 3-epi-25OHD3 concentration to 95 nmol/L. Conclusions Enhanced 3-epi-25OHD3 signal leads to overestimation of its concentrations in the indirect quantitation methods used in many previous studies. The enhanced signal may contribute to greater interference in some 25OHD LC-MS/MS assays than others. We highlight that equimolar responses cannot be assumed in LC-MS/MS systems, even if two molecules are structurally similar.
| Type: | Article |
|---|---|
| Title: | Enhanced 3-epi-25-hydroxyvitamin D3 signal leads to overestimation of its concentration and amplifies interference in 25-hydroxyvitamin D LC-MS/MS assays |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1177/0004563213497691 |
| Publisher version: | http://dx.doi.org/10.1177/0004563213497691 |
| Language: | English |
| Additional information: | This article is distributed under the terms of the Creative Commons Attribution-NonCommercial 3.0 License (http://www.creativecommons.org/licenses/by-nc/3.0/) which permits non-commercial use, reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and Open Access page(http://www.uk.sagepub.com/aboutus/openaccess.htm). |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences |
| URI: | https://discovery.ucl.ac.uk/id/eprint/1469489 |
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