title: A liquid chromatography-mass spectrometry method for the quantification of bioavailability and bioconversion of beta-carotene to retinol in humans.
abstract: A method based on high-performance liquid chromatography-atmospheric pressure chemical ionization mass spectrometry (APCI LC-MS) was developed for the quantification of the bioavailability of retinyl palmitate and beta-carotene and the bioconversion of beta-carotene to retinol in humans. Following oral administration of [8,9,10,11,12,13,14,15,19,20-13C10]-retinyl palmitate and [12,13,14,15,20,12',13',14',15',20'-13C10]-beta-carotene at physiological doses to children between 8 and 11 years of age, blood samples were drawn and serum was prepared. Retinol and beta-carotene were extracted from 0.2- and 1.0-mL serum samples, respectively, and analyzed using reversed-phase HPLC with a C30 column interfaced to an APCI mass spectrometer. Unlike other LC-MS assays for carotenoids, no additional purification steps were necessary, nor was any derivatization of retinol or beta-carotene required. APCI LC-MS showed a linear detector response for beta-carotene over 4 orders of magnitude. Using selected ion monitoring to record the elution profile of protonated circulating beta-carotene at m/z 537 and [13C10]-beta-carotene at m/z 547, the limit of detection was determined to be 0.5 pmol injected on-column. To assess the ratio of labeled to unlabeled retinol, selected ion monitoring was carried out at m/z 269, 274, and 279. These abundant fragment ions corresponded to the loss of water from the protonated molecule of circulating retinol, [13C5]-retinol (metabolically formed from orally administered [13C10]-beta-carotene), and [13C10]-retinol (formed by hydrolysis of [13C10]-retinyl palmitate). The ratios of labeled to unlabeled retinol and the ratio of labeled to unlabeled beta-carotene were calculated. Combined with standard HPLC measurement of beta-carotene and retinol concentration and a mathematical model, these results showed that this simple LC-MS method can be used to quantify beta-carotene bioavailability and its bioconversion to retinol at physiologically relevant doses.
authors.affiliation: Department of Medicinal Chemistry and Pharmacognosy, University of Illinois at Chicago, 60612, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.forename: X
authors.initials: X
authors.lastname: Xu
authors.forename: M
authors.initials: M
authors.lastname: van Lieshout
authors.forename: C E
authors.initials: CE
authors.lastname: West
authors.forename: J
authors.initials: J
authors.lastname: Lugtenburg
authors.forename: M A
authors.initials: MA
authors.lastname: Verhoeven
authors.forename: A F
authors.initials: AF
authors.lastname: Creemers
authors.lastname: Muhilal
authors.forename: R B
authors.initials: RB
authors.lastname: van Breemen
mesh_terms: D001682:Biological Availability; D002648:Child; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D014801:Vitamin A / Q000378:metabolism*; D019207:beta Carotene / Q000493:pharmacokinetics*
chemical_list: D019207:beta Carotene; D014801:Vitamin A