Several disturbances concerning lipid metabolism are found in obese Zucker rats. The present study was designed to determine the effect of energy restriction on fatty acid profile of adipose tissue triglycerides from two different anatomic locations, in genetically obese rats. The impact of the addition of a high amount of olive oil to a restricted diet was also considered. Lean and obese male Zucker rats, 11 weeks old, were used. The former (n = 7) were fed a control diet ad libitum during 4 weeks, and the latter (n = 21) were randomized into 3 groups and fed either a control diet ad libitum or diets restricted to 75% of lean rat intake which provided different amounts of olive oil as fat source. Obesity induced strong modifications in fatty acid profiles from subcutaneous and mesenteric adipose tissues that were not corrected when a standard-fat 25% energy-restricted diet was used for feeding. In contrast, when a high amount of olive oil was included into the restricted diet, a new fatty acid profile in adipose tissue from obese rats, that mirrors the one of lean rats, was observed. Significant differences between two anatomic locations were found in the pattern of modifications produced by obesity, but not by dietary treatments.