# S-equol producing bacteria: isolation and identification from Albino Wistar rat gut microbiota

## TL;DR

This study isolates and identifies bacteria from rat gut microbiota that convert soy compounds into health-benefiting S-equol, with potential for functional food and therapeutic applications.

## Contribution

The study isolates and identifies novel S-equol-producing bacteria from rat gut microbiota, including their phylogenetic classification and S-equol production capabilities.

## Key findings

- Four S-equol-producing bacteria were isolated from albino Wistar rats and identified as C. freundii, Escherichia fergusonii, and Enterococcus faecalis.
- S-equol concentrations ranged from 5.90 to 7.56 µg/g of fermented soybean across different bacterial strains.
- Dietary interventions, particularly fermented soy feed, increased S-equol fecal excretion compared to urine.

## Abstract

The metabolism of soy isoflavones by gut microbiota is critical for the bioactivation and bioavailability of these compounds, particularly daidzein, which is further metabolized by gut bacteria to produce S-equol. S-equol, an exclusive gut bacterial metabolite, is associated with health benefits such as reduced blood pressure, cardiovascular disease prevention, and protection against hormone-related cancers due to its estrogen-mimicking structure and antioxidant properties. However, the limited availability of S-equol-producing bacteria has hindered its production and utilization. This study investigates the isolation and characterization of S-equol-producing microbes from albino Wistar rats and explores the impact of dietary interventions on S-equol production. Preliminary tests showed that both dietary groups excreted more S-equol in feces than urine, with rats on fermented soy feed showing higher S-equol levels due to the presence of daidzein, a precursor. In this study, we isolated four anaerobic S-equol-producing bacteria — MG1 (PX459562), MG2 (PX459563), MG3 (PX459564), and MG4 (PX459565) from the intestine and feces of albino Wistar rats. High-Performance Thin-Layer Chromatography (HPTLC) and High-Performance Liquid Chromatography (HPLC) confirmed the presence of S-equol, with concentrations ranging from 5.90 to 7.56 µg/g of fermented soybean across different strains. Phylogenetic analysis revealed that the isolates belonged to the Enterobacteriaceae and Enterococcaceae families, identifying MG1 as C. freundii strain ATCC 8090, MG2 as Escherichia fergusonii strain NBRC 102419, and both MG3 and MG4 as Enterococcus faecalis strain NBRC 100480. Our findings underscore the significant role of gut microbiota in metabolizing daidzein into S-equol, highlighting the potential for utilizing these bacterial strains in functional food development and therapeutic applications. While the pathogenic nature of E. fergusonii (MG2) precludes its therapeutic use, strains MG1, MG3, and MG4, which match common commensal bacteria, show promise for commercial S-equol production and may serve as valuable resources for further investigation and utilization in promoting health and preventing associated diseases.

• Dietary intervention modulates gut microbiota in albino Wistar rats.

• Soybean fermentation enables efficient conversion of daidzin to bioactive S-equol.

• Novel S-equol–producing microbes were isolated and identified.

The online version contains supplementary material available at 10.1007/s00253-026-13759-4.