The effect of dietary iron treatment on hematological parameters, serum iron status and physical performance was assessed in 12 female distance runners during severe physical training. The subjects were instructed to take a tablet containing 60 mg iron/day (iron supplement group:n =6) or a matching placebo tablet (placebo group:n= 6) in a randomized double‐blind fashion for 8 weeks during training. Except for the iron or placebo supplement, the subjects’activities of daily life and quantity of distance running training were similar in both groups. To assess the effect of iron dosage, the lactate threshold (LT), onset of blood lactate accumulation (OBLA) and maximal oxygen uptake (VO2max) were measured by treadmill incremental exercise tests. Resting hematological parameters and 3000 m running performance were obtained pre‐and post‐training with supplementation. In the iron supplement group, a significant improvement was observed in serum ferritin, running velocity both at LT (VLT) and OBLA (VOBLA), and the 3000 m running performance. In the placebo group, however, serum ferritin decreased significantly and there were no significant improvements in blood lactate variables (e.g., VLTand VOBLA) and running performance. These results indicate that by regular rigorous training without iron supplement, female distance runners have a tendency to induce an iron deficient condition, which, in turn, might disrupt improvement in running performance. In conclusion, the present study suggests that the 60 mg of regular iron supplement a day might be useful for prevention of iron deficiency in female athletes during periods of rigorous training.