Development of Vitamin D Determination in Infant Formula by Column-Switching HPLC with UV Detector

This study was carried out to develop an analytical method for the determination of vitamin D in infant formula. Vitamin D was determined by column-switching high-performance liquid chromatography (HPLC) equipped with a reversed phase column and UV detector after saponification and extraction of the formula with an organic solvent. A preseparation column ($C_8$), focusing column ($C_{18}$), analytical column ($C_{18}$) and UV-Vis detector (254 nm) were used. The limits of detection (LOD) and the limits of quantification (LOQ) for vitamin D were estimated to be $1.51{\mu}g/kg$ and $4.95{\mu}g/kg$, respectively. The linearity, recovery, precision and accuracy of the analytical method for vitamin D were evaluated through the application of a SRM (Standard Reference Material) 1846 (National Institute of Standard & Technology, USA). The linearity of this method was calculated with a value of the coefficient of determination ($r^2$) ${\geq}0.9999$. The recovery of vitamin D was $85.20{\pm}3.00%$. The intra-assay precision for vitamin D was between $1.68{\pm}0.03%$ and $5.75{\pm}0.33%$, and the inter-assay precision for vitamin D ranged from $1.73{\pm}0.03%$ to $2.96{\pm}0.09%$. The intra-assay accuracy for vitamin D was between $100.03{\pm}2.77%$ and $102.01{\pm}0.59%$, and the inter-assay accuracy for vitamin D ranged from $99.00{\pm}1.53%$ to $102.01{\pm}3.04%$. The proposed method is optimal for the separation and quantification of vitamin D from infant formula.