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An LC/MS/MS method for stable isotope dilution studies of β-carotene bioavailability, bioconversion and vitamin A status in humans

Lookup NU author(s): Dr Anthony Oxley, Philip Berry, Gordon Taylor, Dr Joe Cowell, Dr Michael Hall, Professor John Edward Hesketh, Professor Georg Lietz, Professor Alan Boddy


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Isotope dilution is currently the most accurate technique in humans to determine vitamin A status and bioavailability/bioconversion of provitamin A carotenoids such as β-carotene. However, limits of MS detection, coupled with extensive isolation procedures, have hindered investigations of physiologically-relevant doses of stable isotopes in large intervention trials. Here, a sensitive liquid chromatography-tandem mass spectrometry (LC/MS/MS) analytical method was developed to study the plasma response from coadministered oral doses of 2 mg [13C10]β-carotene and 1 mg [13C10]retinyl acetate in human subjects over a 2 week period. A reverse phase C18 column and binary mobile phase solvent system separated β-carotene, retinol, retinyl acetate, retinyl linoleate, retinyl palmitate/retinyl oleate, and retinyl stearate within a 7 min run time. Selected reaction monitoring of analytes was performed under atmospheric pressure chemical ionization in positive mode at m/z 537→321 and m/z 269→93 for respective [12C]β-carotene and [12C] retinoids; m/z 547→330 and m/z 274→98 for [13C10]β-carotene and [13C5] cleavage products; and m/z 279→100 for metabolites of [13C10]retinyl acetate. A single one-phase solvent extraction, with no saponification or purification steps, left retinyl esters intact for determination of intestinally-derived retinol in chylomicrons versus retinol from the liver bound to retinol binding protein. Coadministration of [13C10]retinyl acetate with [13C10]β-carotene not only acts as a reference dose for inter-individual variations in absorption and chylomicron clearance rates, but also allows for simultaneous determination of an individual's vitamin A status.

Publication metadata

Author(s): Oxley A, Berry P, Taylor GA, Cowell J, Hall MJ, Hesketh J, Lietz G, Boddy AV

Publication type: Article

Publication status: Published

Journal: Journal of Lipid Research

Year: 2014

Volume: 55

Issue: 2

Pages: 319-328

Print publication date: 24/10/2013

ISSN (print): 0022-2275

ISSN (electronic): 1539-7262

Publisher: American Society for Biochemistry and Molecular Biology


DOI: 10.1194/jlr.D040204


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