[Purpose] The objective of this study was to investigate the effect of the duration of muscle relaxation during intermittent isometric exercises (IIEs). [Subjects] Subjects were twelve healthy males.[Methods] The subjects performed 10 cycles of intermittent isometric knee extension at 60% maximum voluntary contraction for 8 seconds. One cycle of IIE consisted of a muscle contraction phase and subsequent muscle relaxation phase. To determine the durations of the muscle relaxation phase, two recovery periods (T_R1.0 and T_R0.5) of oxygenation level in the vastus lateralis muscle (VL) were measured with near-infrared spectroscopy immediately after the isometric knee extension. T_R1.0 and T_R0.5 were 30.6 ± 6.8 seconds and 15.4 ± 4.8 seconds, respectively. Therefore, IIEs were performed as two different types (IIE-T_R1.0 and IIE-T_R0.5). [Results] Deoxygenation during muscle contraction and relaxation phases in VL were significantly stronger in IIE-T_R0.5 than in IIE-T_R1.0. The reduction in median power frequency, an indirect measure of lactate accumulation in VL, was also significantly greater in IIE-T_R0.5 than in IIE-T_R1.0. [Conclusion] The results of this study suggest that the duration of muscle relaxation determines both deoxygenation and lactate accumulation in active muscles during IIE.