This study is to investigate the effect of deep sea water (DSW) on the exercise-induced fatigue of rats. The exhaustive exercise test on the treadmill and the measurement of biochemical values related to fatigue were conducted after the two-week feeding to male Wistar rats with different hardnesses DSW (DRO, D100, D600) in different dosages (6, 12, 30 mL/kg·d). It was found that the exhausting time and the ratio of lactic acid elimination to lactic acid increment from the experiment groups were significantly better than those from the control group. The BUN level of rats fed with D100 in a dosage of 30 mL/kg·d and D600 in three dosages distinctly was lower than that of the control group. The liver glycogen content in the group that the rats fed with D100 in dosages of 6 and 30 mL/kg·d and D600 in a dosage of 30 mL/kg·d showed significant difference compared with control group. However, the weight gain, kidney-body weight ratio, spleen-body weight ratio, and total plasma protein level in the rats of experimental groups showed no significant difference from those of control group. It suggested that endurance, adaptation for exercising load and accelerating elimination in fatigue of rats were improved when fed with higher hardness and dosage of DSW.