Introduction: Evidence has shown that an increase in the skeletal and hepatic biochemical markers is one of the main factors in the exhaustion of the individual. The aim of this study was to investigate the effect of one session of exhaustive training on some biochemical markers of skeletal muscles and hepatic metabolism in men handball players. Methods: The present study was a quasi-experimental research method in which 12 handball players with at least 2 years' experience in a superior league (age= 21.42±1.56 years, height= 186 ± 5.85 cm and weight= 83.25 ± 10 kg, body mass index= 24.09 ± 2.93 kg. m-2) were randomly selected. Fasting blood samples were collected before and immediately after the maximum Bruce protocol test in order to measure the biochemical changes in skeletal muscles (LDH, CPK and lactate) and hepatic metabolism (ALT and AST). Paired t-test was used to analyze the data. The significance level was less than 0.05. Results: The results showed that the lactate dehydrogenase (P = 0.004), lactate (P = 0.001) and alanine aminotransferase (P = 0.001) levels increased significantly immediately after the exhaustive exercise. The levels of creatine phosphokinase (P = 0.20), aspartate aminotransferase (P = 0.16) increased but the increase was not statistically significant. Conclusion: According to the findings of this study, it can be concluded that progressive exhaustive exercise may increase in some of the biochemical markers of skeletal muscle and liver metabolism. Therefore, it is recommended to trainers and athletes to pay more attention to the training principles of practice. پرونده مقاله