This study examined the uptake and effects of copper (Cu) from flooded agricultural soils to epibenthic amphipods (Hyalella azteca) using 10-day sediment toxicity tests. Soils were collected from 10 citrus agricultural sites in South Florida. One sediment toxicity test was conducted with one flooding of the 10 soils, and based on the results of this test a second sediment toxicity test was conducted with 4 of the soils, after four 14-day flooding and four 14-day drying intervals over 4 months. Sediment toxicity tests were conducted under flow-through conditions using U.S. EPA methodology. Effects on survival, dry weight, and whole-body Cu concentrations of H. azteca were determined. Cu concentrations in overlying water and sediment of both sediment toxicity tests exceeded regulatory criteria for aquatic organisms. Although survival of H. azteca was not consistently affected from the first to the second sediment toxicity tests, dry weight was consistently reduced and related to Cu concentrations in soil, overlying water, and pore water. Furthermore, whole-body tissue Cu concentrations were significantly higher in H. azteca in all 10 soil-water treatments in the first sediment toxicity test and in all 4 soil-water treatments in the second sediment toxicity test compared to controls. Whole-body tissue concentrations and effects on dry weight were related to Cu exposures in soil, overlying water, and pore water. In these managed soil-water systems, small fish consuming H. azteca with high concentrations of Cu may be at risk.