Research primates may experience pain due to veterinary or experimental procedures and effective perioperative care is essential from a welfare and scientific standpoint. The aim of this work was to (1) create and validate a macaque grimace scale, and (2) assess analgesic therapeutic efficacy during the night. Video recording of cynomolgus macaques (Macaca fascicularis) (n=43) pre- and post-surgery was conducted, 1940 images were scored by observers (n=12), and detailed day (111 h) and night (224 h) behaviours were assessed. A novel grimace tool was developed and results indicated overall good inter-rater reliability (ICCaverage meanSD: 0.670.28). Construct/criterion validity was established as grimace scores increased significantly up to 17 h post-op compared to baseline (p<0.001), and behavioural trends correlated with grimace scores (rho=0.22-0.35, p<0.001). General and pain-associated night time behaviours significantly differed from baseline (p<0.05), but not social behaviour. These findings are important to refine perioperative care of research primates.