The determination of 2beta-(3-hydroxypropoxy)-1alpha,25-dihydroxy vitamin D3 (ED-71) in human serum by high-performance liquid chromatography-electrospray tandem mass spectrometry

J Chromatogr B Analyt Technol Biomed Life Sci. 2005 Sep 5;823(2):61-8. doi: 10.1016/j.jchromb.2004.10.066. Epub 2004 Nov 19.

Abstract

A liquid chromatography-electrospray ionization tandem mass spectrometric (LC/ESI-MS/MS) method for the determination of 2beta-(3-hydroxypropoxy)-1alpha,25-dihydroxy vitamin D3 (ED-71) in human serum has been developed. ED-71 in human serum was extracted using two solid-phase extraction steps on Bond Elut C18 and NH2 cartridge. The separation of ED-71 and preED-71 isomer was attained by LC using 2 mmol/L ammonium acetate-methanol (15:85, v/v) as a mobile phase on a Symmetry C18 column (5 microm, 150 mm x 2.1mm i.d.). ESI-MS/MS analysis was operated using selected reaction monitoring (SRM) in positive ion mode. The method achieved a lower limit of quantitation of 25 pg/mL. The calibration curve (25-3200 pg/mL) gave acceptable linearity (r>0.9964). Intra-assay precision ranged from 2.3 to 9.7%. Inter-assay precision ranged from 1.0 to 3.4%. The accuracy was within 90.8-107.0%. This highly sensitive and reproducible method is able to determine only biologically active ED-71 by separating it from preED-71, which is considered to be applicable for the determination of serum samples from pharmacokinetic studies in human.

Publication types

  • Comparative Study

MeSH terms

  • Calcitriol / analogs & derivatives*
  • Calcitriol / blood
  • Calibration
  • Chromatography, High Pressure Liquid / methods*
  • Humans
  • Reproducibility of Results
  • Spectrometry, Mass, Electrospray Ionization / methods*
  • Temperature
  • Time Factors
  • Vitamin D / analogs & derivatives

Substances

  • Vitamin D
  • Calcitriol
  • eldecalcitol