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 D 3, 25-hydroxyvitamin D3 and 25-hydroxyvitamin D 2 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 D 3 and 1α,25-dihydroxyvitamin D3, respectively, using solid-phase extraction.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1917-1930 |
| Number of pages | 14 |
| Journal | Analytical and Bioanalytical Chemistry |
| Volume | 391 |
| Issue number | 5 |
| DOIs | |
| State | Published - Jul 2008 |
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Keywords
- 1α,25-Dihydroxyvitamin D
- 24R,25-Dihydroxyvitamin D
- 25-Hydroxyvitamin D
- Derivatization
- LC-MS
- Metabolic profiling
- UPLC
ASJC Scopus subject areas
- Analytical Chemistry
- Clinical Biochemistry
Cite this
Metabolic profiling of major vitamin D metabolites using Diels-Alder derivatization and ultra-performance liquid chromatography-tandem mass spectrometry. / Aronov, Pavel A.; Hall, Laura M.; Dettmer, Katja; Stephensen, Charles B.; Hammock, Bruce D.
In: Analytical and Bioanalytical Chemistry, Vol. 391, No. 5, 07.2008, p. 1917-1930.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Metabolic profiling of major vitamin D metabolites using Diels-Alder derivatization and ultra-performance liquid chromatography-tandem mass spectrometry
AU - Aronov, Pavel A.
AU - Hall, Laura M.
AU - Dettmer, Katja
AU - Stephensen, Charles B.
AU - Hammock, Bruce D.
PY - 2008/7
Y1 - 2008/7
N2 - 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 D 3, 25-hydroxyvitamin D3 and 25-hydroxyvitamin D 2 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 D 3 and 1α,25-dihydroxyvitamin D3, respectively, using solid-phase extraction.
AB - 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 D 3, 25-hydroxyvitamin D3 and 25-hydroxyvitamin D 2 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 D 3 and 1α,25-dihydroxyvitamin D3, respectively, using solid-phase extraction.
KW - 1α,25-Dihydroxyvitamin D
KW - 24R,25-Dihydroxyvitamin D
KW - 25-Hydroxyvitamin D
KW - Derivatization
KW - LC-MS
KW - Metabolic profiling
KW - UPLC
UR - http://www.scopus.com/inward/record.url?scp=45849133058&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=45849133058&partnerID=8YFLogxK
U2 - 10.1007/s00216-008-2095-8
DO - 10.1007/s00216-008-2095-8
M3 - Article
C2 - 18437365
AN - SCOPUS:45849133058
VL - 391
SP - 1917
EP - 1930
JO - Fresenius Zeitschrift fur Analytische Chemie
JF - Fresenius Zeitschrift fur Analytische Chemie
SN - 0016-1152
IS - 5
ER -