An increase to above-baseline levels of electromyography (EMG) mean power spectral frequency (MPF) has been observed previously during muscle recovery following fatiguing contractions and has been explained by membrane hyperpolarization due to increased activation of the Na+-K+ pump. It is hypothesized that this membrane mechanism is impaired by muscle fiber damage following eccentric exercise. Thus, the aim of the study was to investigate surface EMG signal characteristics during recovery from fatigue after eccentric exercise. Ten healthy subjects performed sustained isometric knee extensions at 40% of the maximal torque (MVC) until task failure before, immediately after and 24 and 48 h after eccentric exercise. Bipolar surface EMG signals were recorded from six locations over the quadriceps during the sustained isometric contraction and during 3-s long contractions at 40% MVC separated by 1-min intervals for 15 min (recovery). Before the eccentric exercise, MPF of EMG signals increased to values above baseline during recovery from the fatiguing isometric contraction (P<0.001), whereas immediately after and 24 and 48 h after the eccentric task, MPF was lower than baseline during the entire recovery period (P<0.01). In conclusion, delayed-onset muscle soreness abolished the supranormal increase in EMG MPF following recovery from fatigue.