HPLC-MS/MS Determination of Vitamin D₂ and Vitamin D₃ in Food

HPLC-MS/MS was applied to the determination of vitamin D2 (VD2) and vitamin D3 (VD3) in food. The sample was saponifed with KOH and extracted with ethyl ether, and the organic phase obtained was dehydrated with anhydrous sodium sulfate and evaporated to dryness. The residue was dissolved with 1 mL of ethanol and used for HPLC-MS/MS determination, using Agilent Zorbax XDB C18 column (2.1 mm×50 mm, 3.5 μm) as chromatographic column, and a mixture of methanol and 10 mmol·L-1 ammonium acetate solution (mixed in the ratio of 90 to 10 by vol.) as mobile phase. Positive electrospray ionization as well as multiple reaction monitoring mode was used in the detection. Linearity ranges of VD2 and VD3 were found same between 50-500 μg·L-1 with their detection limits (3S/N) of 10 μg·L-1. The proposed method was used for the determination of VD2 and VD3 in milk powder and sour milk, giving values of recovery in the range of 76.8%-83.9%.