A sensitive and specific method for measurement of multiple retinoids in human serum with UHPLC-MS/MS

Retinol (vitamin A) circulates at 1-4 mu M concentration and is easily measured in serum. However, retinol is biologically inactive. Its metabolite, retinoic acid (RA), is believed to be responsible for biological effects of vitamin A, and hence the measurement of retinol concentrations is of limited value. A UHPLC-MS/MS method using isotope-labeled internal standards was developed and validated for quantitative analysis of endogenous RA isomers and metabolites. The method was used to measure retinoids in serum samples from 20 healthy men. In the fed state, the measured concentrations were 3.1 +/- 0.2 nM for at RA, 0.1 +/- 0.02 nM for 9-cis RA, 5.3 +/- 1.3 nM for 13-cis RA, 0.4 +/- 0.4 nM for 9,13-dicisRA, and 17.2 +/- 6.8 nM for 4oxo-13-cis RA. The concentrations of the retinoids were not significantly different when measured after an overnight fast (3.0 +/- 0.1 nM for at RA, 0.09 +/- 0.01nM for 9-cis RA, 3.9 +/- 0.2 nM for 13-cis RA, 0.3 +/- 0.1 nM for 9,13-dicis RA, and 11.9 +/- 1.6 nM for 4oxo-13-cis RA). 11-cisRA and 4OH-RA were not detected in human serum.jlr The high sensitivity of the MS/MS method combined with the UHPLC separation power allowed detection of endogenous 9-cis RA and 4oxo-atRA for the first time in human serum.-Arnold, S. L. M., J. K. Amory, T. J. Walsh, and N. Isoherranen. A sensitive and specific method for measurement of multiple retinoids in human serum with UHPLC-MS/MS. J. Lipid Res. 2012. 53: 587-598.