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Development and validation of a new LC‐MS/MS method for simultaneous detection and quantification of Vitamin D related metabolites
Author(s) -
Huang Jianjie,
Peacock Munro,
Adamec Jiri,
Fleet James,
Burgess John,
Teegarden Dorothy,
Ferruzzi Mario,
Weaver Connie M.
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.731.1
Subject(s) - atmospheric pressure chemical ionization , chromatography , mass spectrometry , chemistry , analyte , triple quadrupole mass spectrometer , extraction (chemistry) , analytical chemistry (journal) , selected reaction monitoring , chemical ionization , tandem mass spectrometry , ionization , organic chemistry , ion
Vitamin D (VD) and its metabolites appear in tissues and biological fluids at low concentrations. When combined with complexity of tissue matrices, quantification of VD metabolites in biological samples is challenging. An LC‐MS/MS method for the comparative high throughput quantification of four biologically important VD metabolites was developed using isocratic elution condition coupled with an Atmospheric Pressure Chemical Ionization (APCI) and Multiple Reaction Monitoring (MRM) on triple quadrupole mass spectrometer. To optimize LC ‐ MS/MS conditions, 25‐Hydroxyvitamin D3 (25OH‐D3) was spiked into serum and recovered by liquid‐liquid extraction. Standard curves for all tested analytes displayed liner relationships in the range of 10 ‐ 2500 ng/mL with correlation coefficients of 0.9912 (25OH‐D2), 0.9901 (25OH‐D3), 0.9982 (VD2) and 0.9992 (VD3). Detection limits of quantification (LOQ) for all molecules were determined at 10 ng/mL with signal to noise (s/n) ratios: 10 (25OH‐D2), 14 (25OH‐D3), 8 (VD2), and 14 (VD3) from serum. Relative standard errors of pooled spiked serum samples were less than 3.5%. Future work will focus on extraction from various tissue types.