Sudan IV (Red) is a fat-soluble dye that stains lipids red. Two parameters are expected in positive results of the Sudan IV test:

  • Layers: You should see two layers. Two layer indicates the presences of water-insoluble substances.

  • Colors: You should see red color on the top layer. The Sudan IV will migrate to the top layer and color it red. This indicates that the top layer is a fat.

A negative result will only exhibit one layer instead of two.

There are two test tubes. The first test tube is the negative result and contains an orange solution that is a darker orange near the solution's surface.The second test tube is the positive result and contains a solution made of two layers. The thin top layer near the solution’s surface is a dark orange color, and the thicker bottom layer is white.

Figure 1: (-) is negative result; (+) is positive result.

If the sample doesn't contains lipid, the Sudan IV will form small micelles and disperse throughout the solution.

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