Background: Aspartame is a low-calorie artificial non-sugar sweetener and is commonly used as a sugar substitute in foods and beverages. Pancreas has a vital role in carbohydrate metabolism. The aim of the study: was to study the effect of aspartame on the structure of pancreas. Methods: Eighteen albino Wistar rats (200-250gm) were divided into three groups; control group, aspartame-treated group, was treated with aspartame (250 mg/kg/day) for six months and withdrawal group: was treated with aspartame (250 mg/kg/day) for six months then left without treatment for another month. Serum amylase and lipase were measured, and histological study of the pancreas was conducted. Results: histopathological changes were detected in both pancreatic acini and islets of Langerhans, increased collagen deposition around the acini and blood vessels. Additionally, aspartame supplementation resulted in elevated levels of serum amylase and lipase. Upon withdrawal of aspartame, an obvious improvement was noticed in the withdrawal group in comparison with the aspartame-treated group. Conclusions: Aspartame produces degenerative changes in the pancreas of the rats and causes uprising of serum pancreatic enzymes. Withdrawal of aspartame results in partial restoration of the histological structure of the pancreas. From these results, it is recommended to spot a light on the dangerous impacts of the usage of aspartame in clinical studies.