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  • Title: Determination of vitamins A, D and E in a small volume of human plasma by a high-throughput method based on liquid chromatography/tandem mass spectrometry.
    Author: Midttun Ø, Ueland PM.
    Journal: Rapid Commun Mass Spectrom; 2011 Jul 30; 25(14):1942-8. PubMed ID: 21698677.
    Abstract:
    We have developed an automated high-throughput assay for the determination of vitamin A (retinol), ergocalciferol (25-OH D2), cholecalciferol (25-OH D3) and vitamin E (α-tocopherol) in a small volume of human plasma. Sample preparation involved mixing 50 μL of plasma with 100 μL of ethanol containing isotope-labelled internal standards, followed by mixing with isooctane/chloroform (3:1, 300 μL). The organic phase was evaporated, and the sample reconstituted in 50 μL methanol. The analysis was performed using reversed-phase liquid chromatography with a gradient mobile phase containing water, methanol and ammonium formate. Chromatographic run-time was 5 min, and positive mode electrospray tandem mass spectrometry (MS/MS) was used for detection. The limits of detection were 0.10 μM for all-trans retinol and 3.3 nM for 25-OH D2 and 25-OH D3. Recoveries were 91.9-105.0%, and within- and between-day coefficients of variance (CVs) 2.4-5.3 and 3.1-8.2, respectively. The assay is presently being used in large-scale studies.
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