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Development of a Candidate Reference Measurement Procedure for the Determination of 25-Hydroxyvitamin $D_{3}$ and 25-Hydroxyvitamin $D_{2}$ in Human Serum Using Isotope-Dilution Liquid Chromatography−Tandem Mass Spectrometry
| Content Provider | Scilit |
|---|---|
| Author | Tai, Susan S.-C. Bedner, Mary Phinney, Karen W. |
| Copyright Year | 2010 |
| Abstract | Vitamin D exists in two major forms, vitamin $D_{3}$ and vitamin $D_{2}$. Vitamin D helps the body absorb calcium and promote optimal bone health. Both forms of vitamin D are metabolized to 25-hydroxyvitamin D in the body, and the levels of 25-hydroxyvitamin $D_{3}$ $[25(OH)D_{3}$] and 25-hydroxyvitamin $D_{2}$ $[25(OH)D_{2}$] in serum are considered the best indicators of vitamin D status. A candidate reference measurement procedure for serum $25(OH)D_{3}$ and $25(OH)D_{2}$ has been developed and critically evaluated. The deuterated compounds $25(OH)D_{3}-d_{3}$ and $25(OH)D_{2}-d_{3}$ are used as internal standards for $25(OH)D_{3}$ and $25(OH)D_{2}$, respectively. The $25(OH)D_{3}$ and $25(OH)D_{2}$ and their respective labeled internal standards are simultaneously extracted from serum using liquid−liquid extraction prior to reversed-phase liquid chromatography−tandem mass spectrometry (LC−MS/MS). Chromatographic separation was performed using a cyano (CN) column for both $25(OH)D_{3}$ and $25(OH)D_{2}$. Atmospheric pressure chemical ionization (APCI) in the positive ion mode and multiple reaction monitoring (MRM) were used for LC−MS/MS. The accuracy of the method was evaluated by recovery studies of measuring 25-hydroxyvitamin D [25(OH)D] in spiked samples with known 25(OH)D levels. The recoveries of the added $25(OH)D_{3}$ and $25(OH)D_{2}$ ranged from 99.0% to 101.0%. The absolute recoveries with this method were 97% and 92% for $25(OH)D_{3}$ and $25(OH)D_{2}$, respectively. Excellent precision was obtained with between-set coefficients of variation (CVs) of 0.2−0.6% for 25(OH)D levels >1 ng/g and within 2% for the level of <1 ng/g. Chromatographic separation of $25(OH)D_{3}$ and $25(OH)D_{2}$ from their respective isomers $3-epi-25(OH)D_{3}$ and $3-epi-25(OH)D_{2}$ was achieved. The limit of detection at a signal-to-noise ratio of ∼3 was 40 pg of 25(OH)D on column (or ∼0.15 ng/g as expressed as a concentration). This candidate reference measurement procedure for serum $25(OH)D_{3}$ and $25(OH)D_{2}$ demonstrates good accuracy and precision and low susceptibility to interferences. It can be used to provide an accuracy base to which clinical methods for $25(OH)D_{3}$ and $25(OH)D_{2}$ can be compared and that will serve as a standard of higher order for measurement traceability. |
| Related Links | http://europepmc.org/articles/pmc2838390?pdf=render https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2838390/pdf |
| Ending Page | 1948 |
| Page Count | 7 |
| Starting Page | 1942 |
| ISSN | 00032700 |
| e-ISSN | 15206882 |
| DOI | 10.1021/ac9026862 |
| Journal | Analytical Chemistry |
| Issue Number | 5 |
| Volume Number | 82 |
| Language | English |
| Publisher | American Chemical Society (ACS) |
| Publisher Date | 2010-02-05 |
| Access Restriction | Open |
| Subject Keyword | Journal: Analytical Chemistry Biochemical Research Analytical Chemistry 25-hydroxyvitamin D3 25-hydroxyvitamin D2 Reference Measurement Procedure Atmospheric Pressure Chemical Ionization Liquid Chromatography/tandem Mass Spectrometry Liquid-liquid Extraction |
| Content Type | Text |
| Resource Type | Article |
| Subject | Analytical Chemistry |
