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Pediatric nurses constitute a critical workforce in Ghana due to the continuous and high-intensity demands of child care. However, their occupational health outcomes remain underexplored, particularly in northern Ghana. This study assessed the effect of night shift work on daytime sleepiness and physiological health among pediatric nurses in the Tamale Metropolis, Northern Ghana. A cross-sectional survey was conducted among 175 pediatric nurses. A stratified sampling technique was used to ensure proportional representation of pediatric nurses from each health facility, followed by simple random sampling for participant selection. Data were collected using a structured questionnaire incorporating the Epworth Sleepiness Scale (ESS) and Night Shift Physiological Health Assessment Scale (NSPHAS). Appropriate inferential statistics were used, with statistical significance set at p  < 0.05. The mean daytime sleepiness score following night shift work was 10.12 (SD = 5.55), with 20.6% of participants exhibiting severe excessive daytime sleepiness. The highest chance of dozing was reported when lying down to rest in the afternoon (31.4%). Sleep disturbances recorded the highest mean physiological health score (2.87 ± 0.98), followed by gastrointestinal disturbances and eating habit disruptions (2.29 ± 0.91), and cardiovascular and physical strain (2.28 ± 0.91). Significant differences in physiological health outcomes were also observed across sociodemographic and work-related characteristics ( p  < 0.05). Night shift work is associated with increased daytime sleepiness and physiological health disturbances among pediatric nurses. Context specific interventions, including improved shift scheduling and targeted sleep health education, may help reduce these adverse outcomes.