Background: Nivolumab monotherapy, an immune checkpoint inhibitor, is a standard of care for patients with metastatic gastric cancer (mGC) as a later-line treatment. Several studies have suggested that the circadian rhythm is essential in immune system function, including anti-cancer immunity. This study evaluated whether the time-of-day patterns of nivolumab infusion alter the efficacy of mGC treatment. Methods: This study retrospectively analyzed 298 consecutive mGC patients who received nivolumab monotherapy at a single institution between December 2014 and December 2022. Patients were divided into early (EA)- and late (LA)-nivolumab infusion groups: EA group = ≥70% of their infusions before 14:00, and LA group = <70% of their infusions before 14:00. Treatment efficacy (overall response rate [ORR], disease control rate [DCR], progression-free survival [PFS], and overall survival [OS]) was evaluated. Multivariate analyses included variables with P values <0.1 in univariate analyses. Results: A total of 248 patients were eligible; 140 were classified as having EA, and 108 as having LA. Most baseline characteristics were similar between EA and LA. In contrast, the median infusion time was earlier in EA than in LA (11:50 vs. 14:22). The median follow-up time was 9.0 months (range, 1.2–64.7 months). ORR and DCR were significantly higher in the EA than in the LA among 149 patients with measurable lesions (ORR, 16.9% vs. 3.3%, P=0.01; DCR, 47.2% vs. 20.0%, P<0.01). Patients with EA had significantly better prognoses than those with LA in terms of PFS (median PFS, 2.3 vs. 1.6 months.; hazard ratios [HR], 0.65; P<0.01) and OS (median OS, 7.6 vs. 3.9 months.; HR, 0.64; P<0.01). EA patients received the subsequent chemotherapy more frequently than those in LA (54% vs. 32%; P<0.01). Multivariate analyses, including factors such as age, ECOG performance status, histological type, ascites, modified Glasgow Prognostic Score, and timing of nivolumab infusion, showed that EA was an independent prognostic factor for PFS (adjusted HR, 0.66; P<0.01) and OS (adjusted HR, 0.62; P<0.01). Conclusions: Our data suggest that efforts to schedule nivolumab infusions before mid-afternoon could be considered in daily practice.