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Objectives. This study aims to investigate whether mento-physical exercises can decrease mental fatigue during shift work. Methods . Research subjects including the control room staff of an urban train system were chosen randomly, for whom mental fatigue was assessed before and after rest breaks for control and experimental groups. A new protocol was applied in the experimental group, including breathing exercises, isometric and isotonic exercises, and progressive relaxation during the inter-shift break. A designed questionnaire and the Stroop test were used to evaluate fatigue and the reaction time, respectively. Results . Pre and post-test results showed that mental, visual and physical fatigue and reaction time decreased by 14.48, 49.22, 26.85 and 8.35% in the control group and 36.42, 48.48, 76.37 and 20.56% in the experimental group, respectively. Conclusions . At the 5% α level, Student's t test showed that mento-physical exercises effectively decreased mental and physical fatigue as well as reaction time in the experimental group compared to the control group.