Objective To investigate the effects of single high-intensity interval exercise on positive and negative emotions and lateralization of frontal lobe α frequency band in depressed college students by analyzing the changes of the scores of positive and negative affect schedule (PANAS) and electroencephalography (EGG) α frequency band at the frontal lobe. Methods A total of 38 college students in mild and moderate depressed mood were screened by self-rating depression scale (SDS). They were randomly divided into high-intensity interval exercise group (n=19) and control group (n=19) by coin toss. The high-intensity interval exercise group performed single high-intensity interval exercise for 20 min, while the control group sat still for 20 min. After the intervention, SDS was used again to test the depression scores of the subjects 1 week later. Before and immediately after the intervention, PANAS was used to test the emotional state of the subjects, and the power values of frontal lobe α frequency band of the subjects at rest were collected by EGG topographic map. The PANAS scores, SDS scores and the changes of total power of the frontal lobe α frequency band and lateralization of the 2 groups were compared before and after the intervention. Results The SDS scores of the subjects were decreased, the positive emotion scores were increased, and the negative emotion scores were decreased in the single high-intensity interval exercise group after intervention (all P<0.05). There were no significant differences in the above indexes in the control group before or after intervention (all P>0.05). The total power values of the right frontal lobe α frequency band of the subjects were increased in the high-intensity interval exercise group after intervention (P<0.05), and the lateralization of the frontal lobe α frequency band was improved (both P<0.05); the total power values of the left and right frontal lobe α frequency band were increased in the control group after intervention (both P<0.05), but there was no significant difference in the lateralization of the frontal lobe α frequency band (P>0.05). Conclusion Single high-intensity interval exercise can effectively improve college students' mood and lateralization of frontal lobe α frequency band.