Sung Eun Cho (Presenter)
LabGenomics Clinical Laboratories
Bio: Director of LabGenomics Clinical Laboratories M.D., Ph.D.
Authorship: Sung Eun Cho (1), Hyojung Lee (1), Sollip Kim (2), Young Dae Kim (3), Dong Hee Seo (1), Junghan Song (4)
(1) LabGenomics Clinical Laboratories (2) Laboratory Medicine, Ilsan Paik Hospital, (3) Pediatrics, Ilsan Paik Hospital, (4) Laboratory Medicine, Seoul National University Bundang Hospital, Korea
| Short Abstract We evaluated the performance of Ultra-Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC-MS/MS) method to measure serum 3-epi-25-hydroxyvitamin D3 (epi-25(OH)D3), 25-hydroxyvitamin D3 (25(OH)D3) and 25-hydroxyvitamin D2 (25(OH)D2). Using Kinetex XB-C18 column (Phenomenex, USA) and isocratic methanol/water (77.5/22.5, v/v) flowing at 0.25 mL/min, run time was 13 minutes/sample. TQD triple quadrupole mass spectrometer (Waters, USA) with MRM transitions, MSMS Vitamin D kit (PerkinElmer, Finland) and PTAD derivatization method were used. Intra- and inter-run precisions were less than 15 %. The carryover was 0.10%, -0.27% and 0.10%, respectively. There was no ion suppression. The linearity was good with R2>0.9999. The peak of epi-25(OH)D3 was clearly separated from that of 25(OH)D3 in the XIC. The performance of UPLC-MS/MS for epi-25(OH)D3, 25(OH)D3, and 25(OH)D2 was acceptable. |
Long Abstract
Introduction: We evaluated the performance of Ultra-Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC-MS/MS) method to measure serum 3-epi-25-hydroxyvitamin D3 (epi-25(OH)D3), 25-hydroxyvitamin D3 (25(OH)D3) and 25-hydroxyvitamin D2 (25(OH)D2).
Methods: Using Kinetex XB-C18 column (Phenomenex, USA) and isocratic methanol/water (77.5/22.5, v/v) with 0.025 % (v/v) HPLC solvent additive flowing at 0.25 mL/min, run time was 13 minutes per sample. TQD triple quadrupole mass spectrometer (Waters, USA) in the ESI positive ion mode with multiple reaction monitoring (MRM) transitions using MSMS Vitamin D kit (PerkinElmer, Finland) was used to evaluate the precision, carryover, ion suppression and linearity. We performed 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) derivatization method for sample preparation.
Results: Intra- and inter-run precisions were between 1.23 and 13.28 % and, between 1.02 and 10.08 %, respectively. The carryover was 0.10%, -0.27% and 0.10%, respectively. There was no ion suppression. The calibration curve showed good linearity from calibrator level 1 (4.7 ng/mL) to calibrator level 6 (150 ng/mL) with R2>0.9999. The peak of epi-25(OH)D3 was clearly separated from that of 25(OH)D3 in the extracted ion chromatogram.
Conclusions: The epi-25(OH)D3 was clearly separated from 25(OH)D3 and the performance of UPLC-MS/MS assay for epi-25(OH)D3, 25(OH)D3, and 25(OH)D2 was acceptable.
References & Acknowledgements:
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