Abstract: A study on the simultaneous determination of yellow 2G, butter yellow and Citrus Red 2 by HPLC with diode array detector after purification with gel permeation chromatography was reported in this paper. Samples with high fat were extracted directly with a mixture of cyclohexane-athylacetate (1+1) and purified by passing through a column packed with Bio-beads SX3 gel, to remove interfering high molecular fatty substances and natural colorants in food samples. HPLC was carried out on the C18 column (4.6mm×250mm, 5μm) with MeOH+H2O as mobile phase in the gradient elution. The diode array detection was applied in the determination of the 3 compounds mentioned above. Linear relationships in the range from 0.5mg/L to 10mg/L of any of the 3 compounds were obtained with correlation coefficients (r) larger than 0.999. RSD’s obtained were in the range of 1.23% to 4.20%, and average recoveries were in the range of 78.6% to 97.6%. Detection limits were found to be 2.0, 1.0 and 2.0mg/kg for Yellow 2G, butter yellow and Citrus Red 2 respectively.