An extraction method employing dispersive liquid–liquid microextraction (DLLME) and ICP-OES has been developed for rapid separation, pre-concentration and determination of ultra-trace amount of Au(III). The extraction of the analyte was performed in the presence of a quaternary ammonium cation, N-Methyl-N, N, N-trioctylammonium chloride, (Aliquat 336) as an extractant based on ion-association extraction system. 1-octanol and acetonitrile were used as extraction and disperser solvents, respectively. The variables affecting the extraction conditions were optimized. Calibration curve in the range of 0.3–100 ng mL−1, the detection limit of 0.09 ng mL−1, enrichment factor of 150 and extraction recovery of 74% were obtained. The precision (R.S.D. %) of the method was 6% for 5 replicates and recoveries of 10 ng mL−1 Au(III). The combined DLLME method with ICP-OES can readily determine Au(III) at trace (μg L−1) level using only 10 mL of sample solution (tap, lake, and mining water) and ore sample without interference by the matrices. This methodology is simple, fast, and low cost which can be used in routine analytical laboratories.