Muscle deoxygenation responses provide information about the training impulse of an exercise session enabling adaptation to be predicted. Our aim was to investigate muscle oxygenation profiles during prolonged low-intensity eccentric and concentric cycling. Twelve healthy men performed two 45-min exercise sessions of concentric (CON) and eccentric (ECC) cycling, matched for the same heart rate at the start of each session. Mechanical power output during ECC was ∼2.5 times that of CON (210±40 W vs. 82±16 W). Oxygen uptake, blood lactate, cardiac output and systolic arterial pressure responses did not differ between exercises. Heart rate was similar at 5 min of each exercise bout but progressively increased during ECC and was higher at 15, 30 and 45 min of ECC compared to CON (+10 bpm), with a trend for a lower stroke volume. Diastolic and mean blood pressures were higher during ECC. No significant differences were observed in muscle oxygenation profiles. Muscle oxygenation responses during prolonged low-intensity exercise were not affected by the type of muscle action at the same metabolic demand and cardiac output.