Mass Spectrometry
Online ISSN : 2186-5116
Print ISSN : 2187-137X
ISSN-L : 2186-5116
Original Article
A Method for Simultaneous Determination of 25-Hydroxyvitamin D3 and Its 3-Sulfate in Newborn Plasma by LC/ESI-MS/MS after Derivatization with a Proton-Affinitive Cookson-Type Reagent
Tatsuya HigashiMai YokotaAyaka GotoKenji KomatsuTakahiro SugiuraShoujiro OgawaMamoru SatohFumio Nomura
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2016 Volume 5 Issue 2 Pages S0051


A method for the simultaneous determination of 25-hydroxyvitamin D3 [25(OH)D3] and its 3-sulfate [25(OH)D3S] in newborn plasma, which is expected to be helpful in the assessment of the vitamin D status, using stable isotope-dilution liquid chromatography/electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) has been developed and validated. The plasma was pretreated based on the deproteinization and solid-phase extraction, then subjected to derivatization with 4-(4-dimethylaminophenyl)-1,2,4-triazoline-3,5-dione (DAPTAD). The derivatization enabled the accurate quantification of 25(OH)D3 without interference from 3-epi-25(OH)D3 and also facilitated the simultaneous determination of the two metabolites by LC/positive ESI-MS/MS. Quantification was based on the selected reaction monitoring with the characteristic fragmentation of the DAPTAD-derivatives during MS/MS. This method was reproducible (intra- and inter-assay relative standard deviations of 7.8% or lower for both metabolites) and accurate (analytical recovery, 95.4–105.6%). The limits of quantification were 1.0 ng/mL and 2.5 ng/mL for 25(OH)D3 and 25(OH)D3S, respectively, when using a 20-μL sample. The developed method was applied to the simultaneous determination of plasma 25(OH)D3 and 25(OH)D3S in newborns; it was recognized that the plasma concentration of 25(OH)D3S is significantly higher than that of 25(OH)D3, and preterm newborns have lower plasma 25(OH)D3S concentrations than full-term newborns.

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© 2016 Tatsuya Higashi, Mai Yokota, Ayaka Goto, Kenji Komatsu, Takahiro Sugiura, Shoujiro Ogawa, Mamoru Satoh, and Fumio Nomura. This is an open access article distributed under the terms of Creative Commons Attribution License, which permits use, distribution, and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.
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