A new and simple method for Cd, Fe, Mn, Ni, Pb, and Zn determination in fish oil by inductively coupled plasma optical emission spectrometry (ICP-OES) was developed using reversed-phase dispersive liquid-liquid microextraction (RP-DLLME) for sample preparation. The RP-DLLME method was based on the simultaneous extraction and pre-concentration of analytes in fish oil, using 10.0 g of sample preheated (75 °C), with the addition of a dispersant/extractant mixture (n-propanol/diluted HNO3 solution). After stirring and centrifugation (5600g, 10 min), the aqueous phase was used for analyte determination. The RP-DLLME method was performed using only 700 μL of n-propanol and 300 μL of 2.0 mol L−1 HNO3 solution. The recoveries for Cd, Mn, Ni, and Zn using the spikes of 100 and 500 μg kg−1 ranged from 86.4 to 93.4% and 90.4 to 97.3%, respectively. For Fe and Pb, the recoveries ranged from 85.7 and 90.0% and 85.3 and 94.4% for the same spikes. The limits of quantification (LOQs) were 0.41, 1.98, 0.33, 0.35, 1.52, and 1.39 μg kg−1 for Cd, Fe, Mn, Ni, Pb, and Zn, respectively. The proposed RP-DLLME sample preparation method allowed analyte extraction for further determination by ICP-OES using a small amount of diluted HNO3 solution avoiding applying a sample decomposition step. The extract was suitable for direct introduction in the ICP-OES equipment and reached LOQs values lower than similar methods from literature. Finally, the proposed analytical method was viable and this is the first application of RP-DLLME for animal source fat combined with ICP-OES determination.