The aim of this study was to determine by static bioassay whether water hardness affects the toxicity of Zn and Cu to a fish, Gambusia holbrooki Girard, 1859. The acute toxicity of selected heavy metals to G. holbrooki was determined in soft, hard and very hard water (25, 125 and 350 mg L À 1 as CaCO 3 ). Results showed that water hardness had a significant effect on Cu and on Zn toxicity on fish. Toxicity of Cu and of Zn increased with decreasing water hardness. The 96 hours LC 50 values for G. holbrooki were higher in the hard and very hard water compared with soft water. Water hardness had a much smaller effect upon the acute toxicity of Cu than that of Zn. It was observed that the 96 hours LC 50 for Cu at the soft, hard and very hard water was found to be 0.017, 0.17 and 0.65 mg L À 1 , respectively, while the 96 hours LC 50 for Zn at the soft, hard and very hard water was found to be 0.46, 48.1 and 121.6 mg L À 1 , respectively.