This paper proposes a simple HPLC method for the identification of vitamin D3 in fortified milk powder. The method was applied to the determination of the vitamin D3 (as cholecalciferol) concentration in commercial products containing known amounts of vitamin D, under the following chromatographic conditions: C 18, 5 µrn, 120 A, 4.6 x 150 mm column, a mobile phase consisted of methanol and UV (DAD) set at 265 nm. The linearity range was established between 0.01-0.2 μg/ml of cholecalciferol prior to the analysis. The extraction was performed with dichloromethane: methane 1:1 (v/v) prior to the analysis. The limit of quantification was 50 ng/ml. Rapid determination of vitamin D through the chromatographic method represents a good solution for its quantification in the fortified milk powder.