title: Automated method for the determination of vitamin D3 hydroxymetabolites in serum.
abstract: An almost automated method for the determination of hydroxymetabolites of vitamin D(3) (cholecalciferol) in human serum is reported. The method consists of three steps: 1) a batch liquid-liquid extraction step with 2-propanol and hexane, and drying of the extract and reconstitution with phosphate buffer. 2) A cleanup and preconcentration step based on solid-phase extraction using Prospekt equipment, with CN group cartridges and elution with the chromatographic mobile phase. 3) A chromatographic step for individual separation of the target analytes starting with a 90:10 methanol-water mixture, then a linear gradient to obtain 100% methanol; followed by photometric detection. The method provides a linear range between 1.0 and 100 ng mL(-1) for 24,25-(OH)(2) vitamin D(3) and for 25-(OH)(2) vitamin D(3), and between 1.5 and 100 ng mL(-1) for 1,25-(OH) vitamin D(3), with correlation coefficients ranging between 0.993 and 0.987, repeatability between 1.9% and 4.8% and within-laboratory reproducibility between 2.8% and 8.8%.
authors.affiliation: Department of Analytical Chemistry, Annex C-3, Campus of Rabanales, University of Córdoba, 14071 Córdoba, Spain.
authors.forename: J M
authors.initials: JM
authors.lastname: Mata-Granados
authors.forename: A
authors.initials: A
authors.lastname: Caballo-López
authors.forename: M D
authors.initials: MD
authors.lastname: Luque de Castro
authors.forename: J M
authors.initials: JM
authors.lastname: Quesada
mesh_terms: D001331:Automation; D002762:Cholecalciferol / Q000097:blood*; D006863:Hydrogen-Ion Concentration; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D002762:Cholecalciferol