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The objective of this study was to determine if a high fat diet having a 2:1 saturated-polyunsaturated fatty acid ratio exacerbates signs of copper deficiency. Male weanling Long-Evans rats were randomly placed into one of the following treatment groups: adequate copper low fat or deficient copper high fat. The levels of fat used were 31 or 12% of daily energy, and copper concentrations were 94.5 micromol/kg and <15.8 micromol/kg in the copper-adequate and copper-deficient diets, respectively. Cardiac hypertrophy as well as lower liver copper levels and superoxide dismutase activity were observed in both groups of copper-deficient rats. Irrespective of copper level, consumption of the high fat diet resulted in the thickening of the interventricular septum and left ventricular free wall. Electrocardiograms revealed that the copper-deficient high fat diet led to a significantly smaller QT interval compared with all other groups. Significantly greater S-wave voltage due to copper deficiency was observed. Significantly lower heart cytochrome c oxidase (CCO) activity was found in the copper-deficient groups with the copper deficient high fat group showing the lowest activity. Western blots of the cardiac non-myofibrillar fraction demonstrated lower amounts of CCO nuclear encoded peptides in the copper-deficient groups, with the least amount seen in the copper-deficient high fat treatment. These data suggest that a high level of dietary fat exacerbates some of the signs of copper deficiency.