Neuromuscular electrical stimulation (NMES) has been shown to impart a number of health benefits and can be used to produce functional outcomes. However, one limitation of NMES is the onset of NMES-induced fatigue. Multi-channel asynchronous stimulation has been shown to reduce NMES-induced fatigue compared to conventional single-channel stimulation. However, in previous studies in man, the effect of stimulation frequency on the NMES-induced fatigue has not been examined for asynchronous stimulation. Low stimulation frequencies are known to reduce fatigue during conventional stimulation. Therefore, the aim of this study was to examine the fatigue characteristics of high- and low-frequency asynchronous stimulation as well as high- and low-frequency conventional stimulation. Experiments were performed in both able-bodied and spinal cord injured populations. Low frequency asynchronous stimulation is found to have significant fatigue benefits over high frequency asynchronous stimulation as well as high- and low-frequency conventional stimulation, motivating its use for rehabilitation and functional electrical stimulation (FES).