The amount of information exchange through the internet is less than face-to-face communication due to technical limitations of online communication. Previous studies have reported that online communication, compared with face-to-face communication, generated fewer turn-taking, less group satisfaction, and less likeable for members. Although behavioral and psychological studies which investigated the difference between online and face-to-face communication have been accumulated, the underlying neural correlates remain unclear. The purpose of this study was to investigate the difference between face-to-face communication and online communication by measuring inter-brain synchronization using functional near-infrared spectroscopy (NIRS) hyperscanning.The experimental protocol was approved by the ethics committee of the Tohoku University School of Medicine. 30 healthy university students participated in the experiment. Five participants with the same sex who did not know each other before the experiment made a group. We arranged face-to-face (FtoF) communication group and online communication group. Participants were asked to talk about assigned topics for five minutes with wearing the NIRS.The inter-brain synchronization difference was analyzed between the FtoF and online communication group. We found stronger inter-brain synchronization in the FtoF group than the online group at left lateral prefrontal cortex (in the range of periods 1.0–1.6 s and 21.0–23.6 s) and medial prefrontal cortex (in the range of periods 1.0–1.6 s, 2.8–4.7 s and 8.8–10.5 s).Our findings suggest that online communication cannot induce the inter-brain synchronization and quality of communication is lower than FtoF communication.