Blending Three Probiotics Alleviates Loperamide-Induced Constipation in Sprague-Dawley (SD)-Rats

Abstract

BIOVITA 3 bacterial species (BIOVITA 3), a probiotic blend powder containing Clostridium butyricum IDCC 1301, Weizmannia coagulans IDCC 1201 and Bacillus subtilis IDCC 1101, has been used as a food ingredient for gut health. However, its efficacy in improving constipation has not been reported. Therefore, we aimed to investigate the functional effects of oral administration of BIOVITA 3 as well as its component strains alone (at 1.0×10⁹ CFU/day) in Sprague-Dawley (SD) rats with loperamide-induced constipation. The study included fecal analysis, gastrointestinal transit ratio, histopathological analysis, short chain fatty acids (SCFAs), and metagenome analysis. As results, the BIOVITA 3 group showed significant improvements in fecal number, water content, gastrointestinal transit ratio, and thickening of the mucosal layer. In the SCFAs analysis, all probiotic-treated groups showed an increase in total SCFAs compared to the loperamide-constipated group. Changes in microbial abundance and the diversity index of three groups (normal, constipated, and BIOVITA 3) were also defined. Of these, the BIOVITA 3 showed a significant improvement in loperamide-constipated SD-rats. This study suggests the possibility that BIOVITA 3 can be applied as an ingredient in functional foods to relieve constipation.

Keywords: BIOVITA 3 bacterial species; constipation; gut microbiota; probiotics.

Fig. 1.. Effect of BIOVITA 3 and its component strains treatment of loperamide-induced constipated SD rats.

(a) Change in body weight, (b) number of feces per cage, (c) water content (%) in fecal, and (d) gastrointestinal charcoal transit ratio (%). Bars represent the mean±standard deviation (n=10). ^†† p<0.01 vs. G1, ^* p<0.05 and ^** p<0.01 vs. G2. G1, normal group; G2, loperamide-induced constipation group; G3, Bacillus subtilis IDCC 1101 and loperamide-induced constipation group; G4, Weizmannia coagulans IDCC 1201 and loperamide-induced constipation group; G5, Clostridium butyricum IDCC 1301 and loperamide-induced constipation group; G6, BIOVITA 3 and loperamide-induced constipation group; SD, Sprague-Dawley.

Fig. 2.. Histological analysis of colon tissue stained with H&E (Bar=100 μm).

(a) Photographs of colon sections, (b) mucosal layer length, (c) muscle layer length. ^† p<0.05 vs. G1, ^* p<0.05 and ^** p<0.01 vs. G2. G1, normal group; G2, loperamide-induced constipation group; G3, Bacillus subtilis IDCC 1101 and loperamide-induced constipation group; G4, Weizmannia coagulans IDCC 1201 and loperamide-induced constipation group; G5, Clostridium butyricum IDCC 1301 and loperamide-induced constipation group; G6, BIOVITA 3 and loperamide-induced constipation group.

Fig. 3.. Comparative analysis of the fecal microbiome in loperamide-induced constipated SD rats.

(a) Taxonomic abundance at the phylum level and (b) family level. (c) Alpha-diversity index values were statistically analyzed by the Wilcoxon rank-sum test. (d) Principal coordinate analysis (PCoA) of beta-diversity. ^* p<0.05 and ^** p<0.01, significantly different by PERMANOVA. G1, normal group; G2, loperamide-induced constipation group; G6, BIOVITA 3 and loperamide-induced constipation group; SD, Sprague-Dawley.

Fig. 4.. Core gut microbial analysis in SD rats with loperamide-induced constipation by treated probiotics.

The core microbiota identified by the linear discriminant analysis (LDA) effect size algorithm is shown with heat maps of the log-scaled relative abundance. Each plot represents biomarkers in the fecal samples in the G1, G2, and G6 groups at the genus level (a) and species level (b) (n=8). SD, Sprague-Dawley.