A cloud point extraction procedure was presented for the preconcentration and determination of copper(II) ion in various real samples. After complexation by amino-4-(3-nitrophenylazo)pyridine-3-ol, (ANPAP), copper(II) ions was quantitatively recovered in Triton X-114 after centrifugation. 0.4 mL of ethanol acidified with 1.0 M HNO3 was added to the surfactant-rich phase prior to its analysis by specrophotometry at λmax 618. The influence of analytical parameters including complexing agent, Triton X-114, buffer solutions, bath temperature, heating time, centrifuge rate and time were optimized. Under the optimum conditions, the calibration curve is linear for the concentration range of 4.0-115 ng mL-1. The detection and quantification limits of the method were 1.20 and 3.94 ng mL-1 of Cu(II) ion. The enhancement factor of 250 was achieved for 100 mL samples containing the analyte with RSDs were ³1.25 %. The interference effect of some anions and cations was also tested. The method was applied to determine of copper in environmental, water and biological samples.