Introduction: Nowadays, physical activities have become an important way for people to improve their physical fitness. Running is considered to be an exercise option with low investment, low consumption, and no space restrictions. It can play a role in physical fitness, entertainment, and communication. Objective: Analyze the impact of running activities on athletes’ physical fitness. Methods: Volunteer athletes were randomly divided into two groups (experimental and control groups). The experimental group performed running activities for eight weeks while the control group did not participate in running. Key physiological and biochemical indicators such as height, weight, blood pressure, blood lipids, etc. were recorded. The data were collected and processed according to the mathematical statistics method to analyze the data. Results: There is no significant difference in height after the experiment (P>0.05). The value of body weight curve was changed after the increment of running exercise (P<0.05). There was no significant difference in bust size between the two tests (P>0.05). Waist circumference, hip circumference and upper limb circumference were significantly different after the test (P<0.05). The subjects’ blood pressure and heart rate before and after the test were significantly different, proportional to the changes in mean vital capacity (P<0.01). Conclusion: Long-term running training impacts athletes’ physical and mental health. Level of evidence II; Therapeutic studies - investigation of treatment outcomes.