Fat oxidation during exercise is associated with cardio-metabolic benefits, but the extent of which whole-body exercise modality elicits the greatest fat oxidation remains unclear. We investigated the effects of treadmill, elliptical and rowing exercise on fat oxidation in healthy individuals. Nine healthy males participated in three, peak oxygen consumption tests, on a treadmill, elliptical and rowing ergometer. Indirect calorimetry was used to assess maximal oxygen consumption (V̇O_2peak), maximal fat oxidation (MFO) rates, and the exercise intensity MFO occurred (Fat_max). Mixed venous blood was collected to assess lactate and blood gases concentrations. While V̇O_2peak was similar between exercise modalities, MFO rates were higher on the treadmill (mean ± SD; 0.61 ± 0.06 g·min^-1) compared to both the elliptical (0.41 ± 0.08 g·min^-1, p = 0.022) and the rower (0.40 ± 0.08 g·min^-1, p = 0.017). Fat_max values were also significantly higher on the treadmill (56.0 ± 6.2 %V̇O_2peak) compared to both the elliptical (36.8 ± 5.4 %V̇O_2peak, p = 0.049) and rower (31.6 ± 5.0 %V̇O_2peak, p = 0.021). Post-exercise blood lactate concentrations were also significantly lower following treadmill exercise (p = 0.021). Exercising on a treadmill maximizes fat oxidation to a greater extent than elliptical and rowing exercises, and remains an important exercise modality to improve fat oxidation, and consequently, cardio-metabolic health.

• The ability to oxidize fat has been associated with improved oxidative enzymes activity and mitochondrial biogenesis.

• The present study examined the effects of treadmill, elliptical, and rower exercises on maximal fat oxidation rates (MFO), the intensity were MFO was observed (Fat_max) and on fat oxidation curves in healthy and young participants.

• Both MFO and Fat_max were higher during treadmill exercise. Multiple linear mixed-effects regression analyses further revealed an effect of exercise modality on fat oxidation curves.

• Adequate selection of exercise modality during training may have a meaningful impact on substrate oxidation. Treadmill exercise should be considered in training design for those looking to maintain or improve metabolic profiling.