Background: Repeated sprint training in hypoxia has shown to be superior to training in normoxia. However, both natural and simulated altitude training are strategies that are not always accessible and expensive due to traveling or the need for advanced equipment. A possible way of simulating hypoxic conditions is to administer carbon monoxide (CO) prior to training as it has been shown to reduce the oxygen transport and delivery to the muscle similar to systemic hypoxia. It is therefore hypothesized that inhaling a small bolus of CO prior to repeated-sprint exercise will potentiate the acute physiological responses during the training, thus induce performance improvements associated with repeated sprint ability compared to placebo control. Methods: 23 endurance-trained individuals (18 men and 5 women), age 33.2 ± 6.9 years, body mass 78.3 ± 11.0 kg, height 180.7 ± 7.3 cm, performed a 3-week supervised repeated sprint exercise protocol on electromagnetically braked cycle ergometers. A repeated sprint ability test was conducted pre-and post-training intervention. Participants were randomly assigned to either inhaling CO (INCO group) or inhaling a sham gas (NOCO group). Analysis of covariance (ANCOVA) was conducted to determine statistical significance by controlling for pre-test values. Results: Periodic inhalation of CO prior to repeated sprint exercise led a medium, significant difference in mean power decrement (INCO -2.63%, NOCO 8%; p <0.05, ηp2 = 0.19) and fatigue index (INCO 0.28%, NOCO 8.2%; p <0.05, ηp2 = 0.24) compared to sham gas, despite that both groups increased in number of sprints (INCO 16.58%, NOCO 27.60%; p <0.05; ηp2 = 0.71) with no difference between groups. Conclusion: Our findings in this study showed a positive effect on an improved ability to sustain power output during repeated sprints when periodic inhalation of CO is administered. Therefore, it is suggested that periodic inhalation of CO prior to repeated sprint exercise might be a nuanced way to induce favorable physiological adaptations, thus improving performance associated with repeated sprint ability.