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A rapid method for the separation of vitamin D and its metabolites by ultra-high performance supercritical fluid chromatography-mass spectrometry.

Jumaah, Firas LU ; Larsson, Sara LU ; Essén, Sofia LU ; Cunico, Larissa LU ; Holm, Cecilia LU ; Turner, Charlotta LU and Sandahl, Margareta LU (2016) In Journal of chromatography. A 1440. p.191-200
Abstract
In this study, a new supercritical fluid chromatography-mass spectrometry (SFC-MS) method has been developed for the separation of nine vitamin D metabolites within less than eight minutes. This is the first study of analysis of vitamin D and its metabolites carried out by SFC-MS. Six columns of orthogonal selectivity were examined, and the best separation was obtained by using a 1-aminoanthracene (1-AA) column. The number and the position of hydroxyl groups in the structure of the studied compounds as well as the number of unsaturated bonds determine the physiochemical properties and, thus the separation of vitamin D metabolites that is achieved on this column. All D2 and the D3 forms were baseline separated with resolution values>1.5.... (More)
In this study, a new supercritical fluid chromatography-mass spectrometry (SFC-MS) method has been developed for the separation of nine vitamin D metabolites within less than eight minutes. This is the first study of analysis of vitamin D and its metabolites carried out by SFC-MS. Six columns of orthogonal selectivity were examined, and the best separation was obtained by using a 1-aminoanthracene (1-AA) column. The number and the position of hydroxyl groups in the structure of the studied compounds as well as the number of unsaturated bonds determine the physiochemical properties and, thus the separation of vitamin D metabolites that is achieved on this column. All D2 and the D3 forms were baseline separated with resolution values>1.5. The effects of pressure, temperature, flow rate and different gradient modes were studied. Electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) were compared in positive mode, both by direct infusion and after SFC separation. The results showed that the sensitivity in APCI(+) was higher than in ESI(+) using direct infusion. In contrast, the sensitivity in APCI(+) was 6-fold lower than in ESI(+) after SFC separation. The SFC-MS method was validated between 10 and 500ng/mL for all analytes with coefficient of determination (R(2))≥0.999 for all calibration curves. The limits of detection (LOD) were found to range between 0.39 and 5.98ng/mL for 24,25-dihydroxyvitamin D3 (24,25(OH)2D3) and 1-hydroxyvitamin D2 (1OHD2), respectively. To show its potential, the method was applied to human plasma samples from healthy individuals. Vitamin D3 (D3), 25-hydroxyvitamin D3 (25OHD3) and 24,25(OH)2D3 were determined in plasma samples and the concentrations were 6.6±3.0ng/mL, 23.8±9.2ng/mL and 5.4±2.7ng/mL, respectively. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of chromatography. A
volume
1440
pages
10 pages
publisher
Elsevier
external identifiers
  • pmid:26931428
  • wos:000372943400021
  • scopus:84975678710
ISSN
1873-3778
DOI
10.1016/j.chroma.2016.02.043
language
English
LU publication?
yes
id
c9c23efc-4bbd-47e1-8bda-0fb4c866c044 (old id 8856606)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/26931428?dopt=Abstract
date added to LUP
2016-03-15 10:47:28
date last changed
2017-11-12 03:40:57
@article{c9c23efc-4bbd-47e1-8bda-0fb4c866c044,
  abstract     = {In this study, a new supercritical fluid chromatography-mass spectrometry (SFC-MS) method has been developed for the separation of nine vitamin D metabolites within less than eight minutes. This is the first study of analysis of vitamin D and its metabolites carried out by SFC-MS. Six columns of orthogonal selectivity were examined, and the best separation was obtained by using a 1-aminoanthracene (1-AA) column. The number and the position of hydroxyl groups in the structure of the studied compounds as well as the number of unsaturated bonds determine the physiochemical properties and, thus the separation of vitamin D metabolites that is achieved on this column. All D2 and the D3 forms were baseline separated with resolution values>1.5. The effects of pressure, temperature, flow rate and different gradient modes were studied. Electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) were compared in positive mode, both by direct infusion and after SFC separation. The results showed that the sensitivity in APCI(+) was higher than in ESI(+) using direct infusion. In contrast, the sensitivity in APCI(+) was 6-fold lower than in ESI(+) after SFC separation. The SFC-MS method was validated between 10 and 500ng/mL for all analytes with coefficient of determination (R(2))≥0.999 for all calibration curves. The limits of detection (LOD) were found to range between 0.39 and 5.98ng/mL for 24,25-dihydroxyvitamin D3 (24,25(OH)2D3) and 1-hydroxyvitamin D2 (1OHD2), respectively. To show its potential, the method was applied to human plasma samples from healthy individuals. Vitamin D3 (D3), 25-hydroxyvitamin D3 (25OHD3) and 24,25(OH)2D3 were determined in plasma samples and the concentrations were 6.6±3.0ng/mL, 23.8±9.2ng/mL and 5.4±2.7ng/mL, respectively.},
  author       = {Jumaah, Firas and Larsson, Sara and Essén, Sofia and Cunico, Larissa and Holm, Cecilia and Turner, Charlotta and Sandahl, Margareta},
  issn         = {1873-3778},
  language     = {eng},
  month        = {02},
  pages        = {191--200},
  publisher    = {Elsevier},
  series       = {Journal of chromatography. A},
  title        = {A rapid method for the separation of vitamin D and its metabolites by ultra-high performance supercritical fluid chromatography-mass spectrometry.},
  url          = {http://dx.doi.org/10.1016/j.chroma.2016.02.043},
  volume       = {1440},
  year         = {2016},
}