The effect of an endurance training regimen on muscle oxidative enzymes and work performance was studied in iron-deficient and -sufficient rats. Three-week-old male Sprague-Dawley rats (n = 40) were randomly assigned to diets containing either 6 mg iron/kg (iron deficient) or 50 mg iron/kg (iron sufficient). After 2 wk, each group of rats was further divided into untrained or endurance-trained subgroups. Training consisted of daily treadmill running of gradually increasing duration for a 1-mo period. After the training period, sedentary and endurance-trained iron-deficient rats were anemic (Hgb approximately 8 g/dl compared with 16 g/dl in the 2 control groups) and had significantly lower skeletal muscle cytochrome c concentration, cytochrome oxidase activity, and succinic oxidase activity compared with the iron-sufficient groups. In response to training iron-deficient rats also generally had a substantial increase in skeletal muscle oxidative enzymes (P less than 0.05), in contrast to iron-sufficient animals, in which there was little or no training effect. Work performance in response to training in the iron-deficient rats improved more than sixfold in an endurance type of exercise (P less than 0.05), but maximal oxygen consumption during a brief, intense type of exercise was not significantly affected. The results suggest that endurance training of iron-deficient rats results in a milder anemia and less drastic reduction of skeletal muscle oxidative enzymes which in turn allows better performance in an endurance type of exercise.