Determination of Lutein and β-Carotene in Infant Formula and Adult Nutritionals by Ultra-High-Performance Liquid Chromatography: Single-Laboratory Validation, First Action 2016.13.

An ultra-HPLC method for the determination of lutein and β-carotene in infant formula and adult nutritionals was validated using both unfortified and fortified samples provided by the AOAC Stakeholder Panel on Infant Formula and Adult Nutritionals (SPIFAN). All experiments showed separation of all-trans-lutein and β-carotene from their major cis isomers, apocarotenal, α-carotene, lycopene, and zeaxanthin. Samples spiked with all-trans-lutein and β-carotene showed no isomerization during sample preparation. Linearity of the calibration solutions correlated to approximately 0.8-45 μg/100 g (reconstituted basis) for samples prepared for the lowest sample concentrations. With dilutions specified in the method, the range can be extended to approximately 2250 μg/100 g. The LOD for both lutein and β-carotene was 0.08 μg/100 g, and the LOQ for both was 0.27 μg/100 g. For all measurements in the range of 1-100 μg/100 g, repeatability RSD was ≤5.8% for lutein and ≤5.1% for β-carotene. For measurements >100 μg/100 g, repeatability RSD was ≤1.1% for lutein and ≤1.7% for β-carotene. Accuracy was determined by recovery from spiked samples and ranged from 92.3 to 105.5% for lutein and from 100.1 to 107.5% for β-carotene. The data provided show that the method meets the criteria specified in the Standard Method Performance Requirements for carotenoids (SMPR 2014.014).