Key points Dietary fish oil has been found to have protective effects against cardiovascular disease, particularly in its role as an anti‐arrhythmic agent. An additional mechanism proposed for the putative cardio‐protective effect is enhanced efficiency of metabolic energy usage by the heart. We tested whether dietary supplementation of fish oil enhances cardiac efficiency or otherwise improves mechano‐energetic performance. Experiments were performed at two distinct physiological levels (whole organ and isolated tissues) employing two independent metabolic indices (oxygen consumption and heat production), respectively. Feeding rats with either a fish oil‐enriched or a saturated fatty acid‐enriched diet did not alter the efficiency of either the isolated whole heart or its left ventricular trabeculae. Abstract Numerous epidemiological studies, supported by clinical and experimental findings, have suggested beneficial effects of dietary fish or fish oil supplementation on cardiovascular health. One such experimental study showed a profound (100%) increase in myocardial efficiency (i.e. the ratio of work output to metabolic energy input) of the isolated whole heart, achieved by a corresponding decrease in the rate of myocardial oxygen consumption. However, a number of other investigations have returned null results on the latter energetic index. Such conflicting findings have motivated us to undertake a re‐examination. To that effect, we investigated the effects of dietary fatty acid supplementation on myocardial mechano‑energetics, with our primary focus on cardiac efficiency. We used both isolated hearts and isolated left ventricular trabeculae of rats fed with one of three distinct diets: reference (REF), fish oil‐supplemented (FO) or saturated fat‐supplemented (SFA). For all three groups, and at both spatial levels, we supplied 10 mm glucose as the exogenous metabolic substrate. In the working heart experiments, we found no difference in the average mechanical efficiency among the three dietary groups: 14.8 ± 1.1% (REF), 13.9 ± 0.6% (FO) and 13.6 ± 0.7% (SFA). Likewise, we observed no difference in peak mechanical efficiency of left ventricular trabeculae among the REF, FO and SFA groups: 13.3 ± 1.4, 11.2 ± 2.2 and 12.5 ± 1.5%, respectively. We conclude that there is no effect of a period of pre‐exposure to a diet supplemented with either fish oil or saturated fatty acids on the efficiency of the myocardium at either spatial level: tissue or whole heart.