To quantify interindividual differences in the human intestinal microbial metabolism of (−)-epicatechin (EC), in vitro anaerobic incubations with fecal inocula from 24 healthy donors were conducted. EC-derived colonic microbial metabolites were qualitatively and quantitively analyzed by liquid chromatography triple quadrupole mass spectrometry (LC-TQ-MS) and liquid chromatography time-of-flight mass spectrometry (LC-TOF-MS). Quantitative microbiota characterization was achieved by 16S rRNA analysis. The results obtained show 1-(3′,4′-dihydroxyphenyl)-3-(2″,4″,6″-dihydroxyphenyl)-2-propanol (3,4-diHPP-2-ol) and 5-(3′,4′-dihydroxyphenyl)-γ-valerolactone (3,4-diHPV) to be key intermediate microbial metabolites of EC and also revealed the substantial interindividual differences in both the rate of EC conversion and the time-dependent EC metabolite pattern. Furthermore, substantial differences in microbiota composition among different individuals were detected. Correlations between specific microbial phylotypes and formation of certain metabolites were established. It is concluded that interindividual differences in the intestinal microbial metabolism of EC may contribute to interindividual differences in potential health effects of EC-abundant dietary foods or drinks.

Multiple beneficial effects have been attributed to green tea catechins (GTCs). However, the bioavailability of GTCs is generally low, with only a small portion directly absorbed in the small intestine. The majority of ingested GTCs reaches the large intestinal lumen, and are extensively degraded via biotransformation by gut microbiota, forming many low-molecular-weight metabolites such as phenyl-γ-valerolactones, phenolic acids, butyrate, and acetate. This process not only improves the overall bioavailability of GTC-derived metabolites but also enriches the biological activities of GTCs. Therefore, the intra- and inter-individual differences in human gut microbiota as well as the resulting biological contribution of microbial metabolites are crucial for the ultimate health benefits. In this review, the microbial degradation of major GTCs was characterized and an overview of the in vitro models used for GTC metabolism was summarized. The intra- and inter-individual differences of human gut microbiota composition and the resulting divergence in the metabolic patterns of GTCs were highlighted. Moreover, the potential beneficial effects of GTCs and their gut microbial metabolites were also discussed. Overall, the microbial metabolites of GTCs with higher bioavailability and bioactive potency are key factors for the observed beneficial effects of GTCs and green tea consumption.

Background: Ferroptosis is a regulated cell death process linked to various diseases. This study explored whether Epigallocatechin-3-gallate (EGCG), a tea-derived antioxidant, could regulate ferroptosis to alleviate dextran sulfate sodium (DSS)-induced colitis. Methods: A DSS-induced colitis model was used to assess EGCG’s effects. Ferroptosis markers, oxidative stress, and iron metabolism were evaluated, alongside Nrf2-GPX4 pathway activation and ferritin (FTH/L) expression. Results: Iron dysregulation and oxidative stress contributed to DSS-induced colitis by activating ferroptosis in colonic epithelial cells. EGCG supplementation inhibited ferroptosis, reducing oxidative damage. Mechanistically, EGCG activated the Nrf2-GPX4 pathway, enhancing antioxidant defense, and improved iron metabolism by upregulating ferritin expression. Conclusions: EGCG effectively suppressed DSS-induced ferroptosis and colitis, highlighting its potential as a ferroptosis inhibitor and therapeutic agent.

(-)-Epigallocatechin gallate (EGCG) has been associated with multiple beneficial effects. However, EGCG is known to be degraded by the gut microbiota. The present study investigated the hypothesis that microbial metabolism would create major catechol-moiety-containing microbial metabolites with different ability from EGCG to induce nuclear factor-erythroid 2-related factor 2 (Nrf2)-mediated gene expression. A reporter gene bioassay, label-free quantitative proteomics and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) were combined to investigate the regulation of Nrf2-related gene expression after exposure of U2OS reporter gene or Hepa1c1c7 cells in vitro to EGCG or to its major microbial catechol-moiety-containing metabolites: (-)-epigallocatechin (EGC), gallic acid (GA) and pyrogallol (PG). Results show that PG was a more potent inducer of Nrf2-mediated gene expression than EGCG, with a 5% benchmark dose (BMD₅) of 0.35 µM as compared to 2.45 µM for EGCG in the reporter gene assay. EGC and GA were unable to induce Nrf2-mediated gene expression up to the highest concentration tested (75 µM). Bioinformatical analysis of the proteomics data indicated that Nrf2 induction by PG relates to glutathione metabolism, drug and/or xenobiotics metabolism and the pentose phosphate pathway. Taken together, our findings demonstrate that the microbial metabolite PG is a more potent inducer of Nrf2-associated gene expression than its parent compound EGCG.

Scope: An in vitro faecal incubation model combined with reporter gene assay based testing strategy is developed to characterize interindividual differences in the gut microbial conversion of (‐)‐epigallocatechin‐3‐O‐gallate (EGCG) and its consequences for potential activation of Nrf2‐mediated gene expression.Method & Results: Anaerobic human faecal incubations are performed to characterize the microbial metabolism of EGCG including interindividual variability. EGCG derived intestinal microbial metabolite patterns show substantial interindividual differences that are correlated to relative microbial abundances determined by 16S rRNA sequencing. Results obtained show the time‐dependent formation of gallic acid, pyrogallol, phenylpropane‐2‐ols, phenyl‐γ‐valerolactones, and 5‐(3′,5′‐dihydroxyphenyl)valeric acid as the major metabolites, with substantial interindividual differences. The activity of the formed metabolites in the activation of EpRE‐mediated gene expression is tested by EpRE‐LUX reporter gene assay. In contrast to EGCG, at low micromolar concentrations, especially gallic acid, pyrogallol, and catechol induce significant activity in the EpRE‐LUX assay.Conclusions: Given these results and taking the level of formation into account, it is concluded that especially gallic acid and pyrogallol contribute to the EpRE‐mediated beneficial effects of EGCG. The interindividual differences in the formation may result in interindividual differences in the beneficial effects of EGCG and green tea consumption.

(-)-Epicatechin (EC) is one of the most popular polyphenols present in various food products in daily life. Upon intake, it is intensively metabolized by microbiota in the large intestine. In the present study, intra- and inter-individual variations in this gut microbial conversion of EC and the concomitant formation of its major metabolites, including 5-(3′,4′-dihydroxy phenyl)-γ-valerolactone (3,4-diHPV), were identified and quantified via liquid chromatography triple quadrupole mass spectrometry (LC-TQ-MS) in anaerobic fecal incubations. In addition, the bioactivity of EC and 3,4-diHPV in activating Nrf2-mediated gene expression was tested quantifying their effects in the U2OS Nrf2 CALUX assay (a reporter gene assay that is used to test the potency of chemicals in activation of Nrf2 signaling), and on the expression levels of Nrf2-related proteins in Hepa1c1c7 and Caco-2 cells via nanoLC-MSMS. A quantitative real-time polymerase chain reaction (RT-qPCR) was carried out to confirm selected Nrf2-regulated gene expressions at the mRNA level. Results obtained show that both intra- and inter-individual differences exist in human gut microbial EC degradation and 3,4-diHPV formation, with inter-individual differences being more distinct than intra-individual differences. The metabolite, 3,4-diHPV, showed higher potency in the U2OS Nrf2 CALUX assay than EC itself. Among the obviously altered Nrf2-related proteins, 14 and 10 Nrf2-associated proteins were upregulated to a higher extent upon 3,4-diHPV treatment than in the EC treated group for Hepa1c1c7 and Caco-2 cells, respectively. While only three and four of these Nrf2-associated proteins were induced at a higher level upon EC than upon 3,4-diHPV treatment for Hepa1c1c7 and Caco-2 cells, respectively. RT-qPCR results showed that indeed Nrf2-mediated genes (e.g., Nqo1 and Ugt1a) were only induced significantly in 3,4-diHPV treated and not in EC treated Hepa1c1c7 cells. Taken together, the results suggest that the major colonic EC metabolite, 3,4-diHPV, was more capable of inducing Nrf2-mediated gene expression than its parent compound EC. This implies that the evident inter- and intra-individual differences in the microbial conversion of EC to this major metabolite 3,4-diHPV may affect the overall health-promoting effects of EC consumption related to the Nrf2 pathway activation.