Analytical and Bioanalytical Chemistry

, 391:1917

Metabolic profiling of major vitamin D metabolites using Diels–Alder derivatization and ultra-performance liquid chromatography–tandem mass spectrometry

  • Pavel A. Aronov
  • Laura M. Hall
  • Katja Dettmer
  • Charles B. Stephensen
  • Bruce D. Hammock
Original Paper

DOI: 10.1007/s00216-008-2095-8

Cite this article as:
Aronov, P.A., Hall, L.M., Dettmer, K. et al. Anal Bioanal Chem (2008) 391: 1917. doi:10.1007/s00216-008-2095-8

Abstract

Biologically active forms of vitamin D are important analytical targets in both research and clinical practice. The current technology is such that each of the vitamin D metabolites is usually analyzed by individual assay. However, current LC-MS technologies allow the simultaneous metabolic profiling of entire biochemical pathways. The impediment to the metabolic profiling of vitamin D metabolites is the low level of 1α,25-dihydroxyvitamin D3 in human serum (15–60 pg/mL). Here, we demonstrate that liquid–liquid or solid-phase extraction of vitamin D metabolites in combination with Diels–Alder derivatization with the commercially available reagent 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) followed by ultra-performance liquid chromatography (UPLC)–electrospray/tandem mass spectrometry analysis provides rapid and simultaneous quantification of 1α,25-dihydroxyvitamin D3, 1α,25-dihydroxyvitamin D2, 24R,25-dihydroxyvitamin D3, 25-hydroxyvitamin D3 and 25-hydroxyvitamin D2 in 0.5 mL human serum at a lower limit of quantification of 25 pg/mL. Precision ranged from 1.6–4.8 % and 5–16 % for 25-hydroxyvitamin D3 and 1α,25-dihydroxyvitamin D3, respectively, using solid-phase extraction.

Keywords

1α,25-Dihydroxyvitamin D325-Hydroxyvitamin D324R,25-Dihydroxyvitamin D3UPLCLC-MSMetabolic profilingDerivatization

Supplementary material

216_2008_2095_MOESM1_ESM.pdf (186 kb)
ESM 1(PDF 185 kb)

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • Pavel A. Aronov
    • 1
  • Laura M. Hall
    • 2
  • Katja Dettmer
    • 3
  • Charles B. Stephensen
    • 2
  • Bruce D. Hammock
    • 4
  1. 1.Department of EntomologyUniversity of CaliforniaDavisUSA
  2. 2.Department of Nutrition and USDA Western Human Nutrition Research CenterUniversity of CaliforniaDavisUSA
  3. 3.Institute of Functional GenomicsUniversity of RegensburgRegensburgGermany
  4. 4.Department of Entomology and U.C. Davis Cancer Research CenterUniversity of CaliforniaDavisUSA