Anxiety disorders are one of the most prevalent and increasing neuropsychological disorders worldwide. Chamaecyparis obtusa (hinoki) extract has been reported to modulate emotional behaviors. However, the potential anxiolytic effect of hinokinin, a major component of hinoki extract, remains unknown.  In the present study, we examined the anxiolytic effect of the hinoki extract and hinokinin using the elevated plus maze (EPM) test in mice. Male C57BL/6J mice intranasally administered vehicle, diazepam, or hinoki extract. The results showed that administration of hinoki extract significantly increased time spent in the open arm of the EPM, similar to diazepam treatment. Hinokinin alone also significantly reduced anxiety-like behaviors, as evidenced by increased time spent in the open arm. However, acute immobilization stress elevated both anxiety-like behaviors and blood corticosterone levels. Hinokinin restored open-arm exploration and normalized blood corticosterone levels in stressed mice. These results suggest that hinokinin may contribute to the antianxiety effects of hinoki extract through the regulation of stress hormone.