At vital moments in professional soccer matches, penalties were often missed. Psychological factors, such as anxiety and pressure, are among the critical causes of the mistakes, commonly known aschoking under pressure. Nevertheless, the factors have not been fully explored. In this study, we used functional near-infrared spectroscopy (fNIRS) to investigate the influence of the brain on this process. Anin-situstudy was set-up (N= 22), in which each participant took 15 penalties under three different pressure conditions: without a goalkeeper, with an amiable goalkeeper, and with a competitive goalkeeper. Both experienced and inexperienced soccer players were recruited, and the brain activation was compared across groups. Besides, fNIRS activation was compared between sessions that participants felt anxious against sessions without anxiety report, and between penalty-scoring and -missing sessions. The results show that the task-relevant brain region, the motor cortex, was more activated when players were not experiencing performance anxiety. The activation of task-irrelevant areas was shown to be related to players experiencing anxiety and missing penalties, especially the prefrontal cortex (PFC). More particularly, an overall higher activation of the PFC and an increase of PFC lateral asymmetry were related to anxious players and missed penalties, which can be caused by players' worries about the consequences of scoring or missing the penalty kicks. When experienced players were feeling anxious, their left temporal cortex activation increased, which could be an indication that experienced overthink the situation and neglect their automated skills. Besides, the left temporal cortex activation is higher when inexperienced players succeeded to score a penalty. Overall, the results of this study are in line with the neural efficiency theory and demonstrate the feasibility and ecological validity to detect neurological clues relevant to anxiety and performance from fNIRS recordingsin the field.