This study was designed to investigate the effect of short-term, submaximal training on changes in blood substrates, metabolites, and hormonal concentrations during prolonged exercise at the same power output. Cycle training was performed daily by eight male subjects ([Formula: see text], [Formula: see text]) for 10–12 days with each exercise session lasting for 2 h at an average intensity of 59% of [Formula: see text]. This training protocol resulted in reductions (p < 0.05) in blood lactate concentration (mM) at 15 min (2.96 ± 0.46 vs. 1.73 ± 0.23), 30 min (2.92 ± 0.46 vs. 1.70 ± 0.22), 60 min (2.96 ± 0.53 vs. 1.72 ± 0.29), and 90 min (2.58 ± 1.3 vs. 1.62 ± 0.23) of exercise. The reduction in blood lactate was also accompanied by lower (p < 0.05) concentrations of both ammonia and uric acid. Similarly, following training lower concentrations (p < 0.05) were observed for blood β-hydroxybutyrate (60 and 90 min) and serum free fatty acids (90 min). Blood glucose (15 and 30 min) and blood glycerol (30 and 60 min) were higher (p < 0.05) following training, whereas blood alanine and pyruvate were unaffected. For the hormones insulin, glucagon, epinephrine, and norepinephrine, only epinephrine and norepinephrine were altered with training. For both of the catecholamines, the exercise-induced increase was blunted (p < 0.05) at both 60 and 90 min. As indicated by the changes in blood lactate, ammonia, and uric acid, a depression in glycolysis and IMP formation is suggested as an early adaptive response to prolonged submaximal exercise training.Key words: exercise, training, blood metabolites, substrates, hormones.