Measurement of serum 3-epi-25-hydroxyvitamin D_3, 25-hydroxyvitamin D₃ and 25-hydroxyvitamin D₂ in infant, paediatric and adolescent populations of Korea using ultra-performance liquid chromatography-tandem mass spectrometry

Abstract

Background We evaluated the performance of ultra-performance liquid chromatography-tandem mass spectrometry to measure serum 3-epi-25-hydroxyvitamin D, 25-hydroxyvitamin D and 25-hydroxyvitamin D concentrations in 519 infant, paediatric and adolescent serum samples in Korea. Methods We used a Kinetex XB-C18 column and isocratic methanol/water (77.5/22.5, v/v) with 0.025% (v/v) high-performance liquid chromatography solvent additive flowing at 0.25 mL/min, yielding an 11 min/sample run time. A TQD triple quadrupole mass spectrometer in electrospray ionization positive ion mode with multiple reaction monitoring transition via an MSMS vitamin D kit was used to evaluate precision, carryover, ion suppression and linearity. Samples were prepared using the 4-phenyl-1,2,4-triazoline-3,5-dione derivatization method. Results Intra- and inter-run precisions were 1.23-13.28% and 1.02-10.08%, respectively. Group carryovers were -0.27% and 0.10%, respectively. There was no ion suppression. The calibration curve showed good linearity from calibrator Level 1 (11.75 nmol/L) to 6 (375 nmol/L) with R> 0.9999. The 3-epi-25-hydroxyvitamin D and 25-hydroxyvitamin D peaks were clearly separated in the extracted ion chromatogram. Infant serum samples 3-epi-25-hydroxyvitamin D concentrations were significantly higher than paediatric and adolescent concentrations. Conclusions The ultra-performance liquid chromatography-tandem mass spectrometry assay performed acceptably, clearly separating 3-epi-25-hydroxyvitamin D from 25-hydroxyvitamin D. High 3-epi-25-hydroxyvitamin D concentrations were observed in infant but not in paediatric and adolescent serum samples.

Simultaneous Determination of Vitamin D and Its Hydroxylated and Esterified Metabolites by Ultrahigh-Performance Supercritical Fluid Chromatography–Tandem Mass Spectrometry

“…Here, analytes were directly infused in the system individually at a concentration of 2 mg/L. Scan spectra were acquired in both positive and negative modes using MeOH, 0.1% formic acid in MeOH, and 0.2% ammonium formate in MeOH as infusion solvents based on the conditions previously reported in the literature. , Results indicated that positive mode provided better ionization for both ion sources since no signal was found for most of the analytes under negative mode (data not shown). Based on that, subsequent single ion monitoring (SIM), product ion (PI), and MRM studies were carried out with the aim to obtain MS/MS data of all target analytes in positive mode using both ESI and APCI.…”

“…In the past years, targeted analysis of these compounds have been done usually by liquid chromatography (LC) combined with conventional detectors and/or mass spectrometry (MS) . All these methods have been focused on the determination of D 3 or hydroxylated metabolites. − However, the analysis of ester forms has not been carried out so far in any type of sample, as described in a recent review article . The advantages of supercritical fluid chromatography (SFC) in terms of selectivity, comprehensiveness, and analysis speed together with its compatibility with a large range of different detectors have identified this technique as a promising alternative to LC .…”