Abstract: Objective: Exploring the laxative and intestinal protective effects of glutinous rice banana enzyme (GRBE) on constipated mice. Methods: A mouse constipation model was established by administering loperamide hydrochloride (LH). The apparent status, melena discharge time, and small intestine propulsive ratio of mice in each group were observed. The changes of serum tumor necrosis factor-alpha (TNF-α), vasoactive intestinal peptide (VIP), substance P (SP), gastrin (GAS), and motilin (MTL) in each group of mice were analyzed. In addition, the damage of jejunal villi was compared by histopathology. Results: During the administration period, the mice did not experience adverse symptoms such as abdominal distension and diarrhea. The weight increased steadily, with a good mental state and smooth and glossy hair, indicating that GRBE was safe and non-toxic. Compared with the model group, the time to pass the first black stool in the GRBE high-dose group was significantly shortened (P<0.01), and the wet weight, number of particles, and moisture content of the black stool were significantly improved (P<0.001). The ink propulsion rate of mice in the GRBE high-dose group increased significantly (P<0.001), indicating that GRBE had the effect of promoting small intestinal peristalsis in mice. GRBE could improve the intestinal motility and defecation function of constipated mice by reducing the levels of serum VIP, SP and TNF-α (P<0.001) and increase the levels of GAS and MTL (P<0.001). The histopathological results showed that the damage to the jejunal villi of constipated mice in each GRBE dose group was improved to varying degrees, suggesting that GRBE had a certain protective effect on the damage to the jejunal villi caused by LH. Conclusion: Obvious laxative and intestinal protective effects of GRBE on constipated mice are demonstrated, thus providing a theoretical basis for the further development of functional fermented beverages from glutinous rice banana.