Ameliorative Effects of Soybean Powder Fermented by Bacillus subtilis on Constipation Induced by Loperamide in Rats

Constipation is a prevalent gastrointestinal disorder that significantly impairs quality of life. While pharmacological agents such as loperamide are widely used to induce constipation in experimental models, there is increasing interest in natural alternatives for alleviating intestinal dysfunction. In this study, we investigated the laxative effects of soybean powder fermented by Bacillus subtilis DKU_09 in a loperamide-induced rat model of constipation. The probiotic strain was isolated from cheonggukjang, a traditional Korean fermented soybean paste, and its identity was confirmed through 16S rRNA sequencing. Fermented soybean powder was characterized morphologically via scanning electron microscopy and chemically via HPLC to assess its isoflavone content. Rats were administered loperamide (5 mg/kg) for four days to induce constipation and were then treated with fermented soybean powder at doses of 100, 200, or 300 mg/kg. No pharmacological laxatives (e.g., PEG) were used as a positive control; instead, values from the treatment groups were compared with those from the loperamide-only constipation group. Key outcomes of fecal output, water content, colonic fecal retention, and gastrointestinal transit ratio were measured. The fermented product significantly improved stool frequency and moisture content, reduced colonic fecal retention, and restored gastrointestinal transit in a dose-dependent manner. Notably, the 300 mg/kg group demonstrated nearly complete recovery of fecal parameters without affecting body weight. Statistical analysis was performed using one-way ANOVA followed by Tukey's post hoc test. These findings suggest that Bacillus subtilis-fermented soybean powder exerts synergistic laxative effects through the combined action of probiotic viability and fermentation-enhanced bioactive compounds such as aglycone isoflavones. This study supports the potential use of fermented soybean-based nutraceuticals as a natural and safe intervention for constipation and gastrointestinal dysregulation.