Purpose: Although the 24-hours urinary copper excretion is useful for the diagnosis of Wilson disease (WD), there are practical difficulties in the accurate and timed collection of urine samples. The purpose of this study was to verify if the spot morning urinary Copper/ Zinc (Cu/Zn) ratio could be used as a replacement parameter of 24-hours urinary copper excretion in the diagnosis of WD. Methods: A cross-sectional study was conducted at the Department of Pediatric Gastroenterology and Nutrition, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh, from June 2019 to May 2021 on 67 children over three years of age who presented with liver disease. Twenty-seven children who fulfilled the inclusion criteria for WD were categorized into the test group, and the remaining forty children were considered to have non-Wilsonian liver disease and were categorized into the control group. Along with other laboratory investigations, spot morning urinary samples were estimated for the urinary Cu/Zn ratio in all patients and were compared to the 24-hour urinary copper excretion. The diagnostic value of the Cu/Zn ratio was then analyzed. Results: Correlation of spot morning urinary Cu/Zn ratio with 24-hours urinary copper excretion was found to be significant (r=0.60). The area under ROC curve with 95% confidence interval of morning urinary Cu/Zn ratio measured using 24-hours urine sample was 0.84 (standard error, 0.05; p<0.001). Conclusion Spot morning urinary Cu/Zn ratio seems to be a promising parameter for the replacement of 24-hours urinary copper excretion in the diagnosis of WD.