An interlaboratory-verified method for the determination of vitamins A and E in milk- and soy-based infant formula by liquid chromatography with matrix solid-phase dispersion extraction

Abstract

An interlaboratory-verified, liquid Chromatographic (LC) method is presented for determination of all-racemic α-tocopheryl acetate and retinyl palmitate in infant formula. The extraction procedure uses matrix solid-phase dispersion. A sample is mixed with C18, and the mixture is packed into a reservoir and eluted with selective solvents to extract the analytes. After evaporation and filtration, the sample extract is injected directly into a normal-phase LC system with fluorescence detection. All-racemic α-tocopheryl acetate and retinyl palmitate are quantitated isocratically with a mobile phase of hexane containing isopropanol at 0.2% (v/v) and 0.125% (v/v), respectively. A nonfortified zero control reference material (ZRM) was spiked at 5 levels, with 5 replicate analyses of 1/2x, x, 2x, 4x, and 16x where "x" represents the minimum levels of 250 IU/100 kcal (vitamin A) and 0.7 IU/100 kcal (vitamin E) as specified in Title 21 of the Code of Federal Regulations, part 107.100. Recoveries of retinyl palmitate ranged from 83.8 to 107%, and those of all-racemic α-tocopheryl acetate ranged from 87.7 to 108%. Two additional laboratories analyzed the ZRM samples at 4 spiking levels with 6 replicates. Recoveries of retinyl palmitate and all-racemic α-tocopheryl acetate ranged from 92.2 to 104% and from 91.7 to 101%, respectively, in the second laboratory. Recoveries of retinyl palmitate and all-racemic α-tocopheryl acetate ranged from 85.3 to 97.0% and from 86.6 to 110%, respectively, in the third laboratory. Relative standard deviations for all 3 laboratories ranged from 0.2 to 7.5% with an average of 2.9%. In addition, each laboratory analyzed a commercial milk- and commercial soy-based infant formula. Excellent agreement in results was obtained between the 3 laboratories for vitamins A and E in all matrixes.