Fungi and Gut Microbes Reshape the Tea Compound EGCG in Surprisingly Different Ways

Dark tea, the post-fermented tea beloved across parts of China for its earthy depth and reputed health benefits, owes much of its character to microscopic workers. Two fungi in particular, Aspergillus cristatus and Aspergillus niger, dominate the fermentation of products such as Fu brick tea and Liubao tea, secreting extracellular enzymes that systematically remodel the chemistry of the tea leaf. A new study published in Current Research in Food Science has now traced, molecule by molecule, what these fungi do to epigallocatechin gallate, or EGCG, the most abundant catechin in fresh tea leaves, and how the same compound is transformed once it reaches the gut. The results reveal a strikingly context-dependent process: the same two fungi can dismantle EGCG at very different speeds depending on whether they are working in a simple liquid medium or on an actual tea matrix.

EGCG accounts for roughly half of the total catechins in fresh tea leaves and is prized for antioxidant, anti-inflammatory and anticancer activities. Yet it is also notoriously unstable and poorly absorbed. During fermentation it is converted into a family of derivatives, including epigallocatechin, epicatechin, catechin, theasinensins, theaflavins, fuzhuanins and teadenols, many of which show better bioavailability and distinct biological activities. The research team, led by Zexin An and Mingzhi Zhu of Hunan Agricultural University, set out to map this transformation comprehensively, combining targeted quantification with untargeted metabolomics across four experimental systems: liquid fermentation with each fungus separately, solid-state fermentation of sterile raw dark tea, an in vivo rat study, and an in vitro anaerobic culture of rat gut microbiota.

In the liquid fermentation experiments, the fungi were grown for five days before EGCG was added to a final concentration of 1 milligram per milliliter, and samples were collected over 72 hours. In a sterile medium without fungi, EGCG and gallic acid remained stable, ruling out significant auto-oxidation. Once A. cristatus was present, EGCG became undetectable within 12 hours, whereas A. niger needed a full 24 hours to finish the job. First-order kinetic modeling quantified the difference: the apparent disappearance rate constant for A. cristatus was 0.326 per hour, roughly 3.05 times that of A. niger at 0.107 per hour, with corresponding half-lives of 2.13 and 6.49 hours. Gallic acid, the primary hydrolysis product of EGCG, accumulated steadily in both systems.

Untargeted metabolomics using ultra-high-performance liquid chromatography coupled to a Q-Exactive Orbitrap mass spectrometer detected 3,402 annotated metabolic features across the fungal cultures. After filtering for features that changed more than twofold relative to controls, the team identified 235 differential metabolites in the A. cristatus system and 114 in the A. niger system. By tracking how each metabolite’s abundance rose and fell relative to EGCG and gallic acid, the researchers organized them into early-, mid- and late-stage products. Early-stage metabolites included epigallocatechin itself and complex polyflavonoids such as samarangenin A and epigallocatechin-(4β→8)-epicatechin 3,3′-digallate, showing that fungal fermentation drives not only degradation but also polymerization from the very beginning. Mid-stage products shared by both fungi included theasinensin B, oolongtheanin, theaflavin 3,3′-digallate, prodelphinidin A2 3′-gallate and isoneotheaflavin 3-O-gallate, several of which have documented antihypertensive, cholesterol-lowering, antiviral or alpha-glucosidase-inhibiting activities.

The late stages diverged sharply between the two fungi. In A. cristatus cultures, 3-hydroxyflavone emerged as a dominant late-stage EGCG-associated metabolite, accumulating at more than 100 times the level seen with A. niger, a remarkable demonstration of the species-specific conversion efficiency. A. cristatus also generated a diverse set of gallic-acid-derived products, with database-assisted annotations pointing to compounds including ellagic acid, dihydroxyphaseic acid, camonagrel and lidoflazine. A. niger, by contrast, produced fewer but distinctive derivatives, with flavonoids such as quercetin 3-(6”-caffeylgalactoside), isotheaflavin, fuzhuanins A and prodelphinidin A1 peaking at 36 hours before declining, while pelargonidin 3-sambubioside and a methylenedioxy-substituted phenylcoumarin continued rising through 72 hours.

The most surprising result came when the team moved from the simplified liquid system to real tea. Sterile raw dark tea was fermented with each fungus for 15 days, mirroring the roughly two-week “flowering” stage of Fu brick tea production. Visually, the cultures diverged dramatically: A. cristatus eventually produced the characteristic golden “golden flowers” on the leaves, while A. niger grew vigorously and turned the leaves brownish. But the kinetics of EGCG disappearance reversed. In tea fermented with A. niger, EGCG dropped to 209.5 micrograms per gram by day 5 and was nearly undetectable by day 15, whereas A. cristatus fermentation showed a slower, more gradual decline that plateaued after day 10. The authors suggest several possible explanations, including the known ability of high EGCG concentrations to inhibit A. niger by suppressing chitinase and chitin synthase expression, and the possibility that the tea matrix redirects A. cristatus toward alternative metabolic pathways. They also caution that the tea was autoclaved before inoculation, a treatment that may itself alter catechins, proteins and their interactions, and that untreated raw tea was not directly compared.

To untangle the chemistry of the tea matrix, the researchers applied weighted correlation network analysis to 5,144 detected compounds. In the A. cristatus fermentations, the data sorted into 15 color-coded modules, with the Meblue module of 651 metabolites showing a highly significant positive correlation with EGCG content. Within it, 585 core metabolites passed stringent thresholds, including 13 flavans such as catechin, epigallocatechin, taxifolin, neotheaflavin and methylated epigallocatechin gallates. A network built around EGCG linked it to 244 highly associated metabolites. Molecular networking on the GNPS platform then clustered structurally similar compounds sharing a common fragment ion at m/z 170.02, revealing a putative EGCG dimer at m/z 915.161, a potential epigallocatechin dimer at m/z 611.14, and other catechin relatives whose mass shifts hint at hydroxymethylation and related modifications. A. niger fermentations yielded their own module structure and unique core metabolites, including 4-gallocatechol, which accumulated significantly by day 15 and may represent a core late-stage product.

The study then followed EGCG into the body. Rats received a single oral dose of 500 milligrams per kilogram of body weight, and fecal samples were collected over 24 hours. EGCG first appeared in feces at 6 hours post-gavage, peaked at 440.27 micrograms per gram at 10 hours, and had fallen to just 6.14 micrograms per gram by 24 hours, pinpointing a 6-to-12-hour window as the key intestinal transit phase. Metabolomics of 106 available fecal samples yielded 14,931 annotated features, from which 683 EGCG-associated candidates were selected by chemical-class filtering and 59 core metabolites were prioritized using statistical models. Among 13 core flavans, the team found not only monomeric catechins and their derivatives but also a high-molecular-mass EGCG-associated product and putative theaflavin-related compounds, suggesting that oxidative transformations, potentially involving both non-enzymatic reactions and gut microbial interactions, occur within the intestine. A dimethylated catechin derivative also appeared, a modification that may alter polarity, stability and metabolic kinetics.

Perhaps the most instructive contrast was between the living gut and the petri dish. When rat gut microbiota were cultured anaerobically with EGCG at three concentrations, the bacteria reduced EGCG levels over time but unexpectedly failed to degrade it completely, even after 72 hours. Only nine flavans were detected among 5,942 annotated features, mostly monomeric catechins such as epigallocatechin, catechin and epicatechin, along with sulfated derivatives including epicatechin 4′-sulfate and 7′-sulfate. Seven of these nine metabolites also appeared in the fungal fermentations, and epicatechin 4′-sulfate showed similar concentration trends in vivo and in vitro. The far richer metabolic profile seen in living rats therefore cannot be explained by fecal microbiota alone; host enzymatic reactions, intestinal absorption, dynamic pH, bile, oxygen gradients and the mucus-associated microenvironment all likely contribute. 16S rDNA sequencing of the cultures showed Ligilactobacillus, Lactobacillus, Streptococcus and Escherichia-Shigella as dominant genera, with exploratory correlations between microbial abundance and EGCG levels that did not remain statistically significant after correction for multiple testing.

The authors are candid about the limits of the work: without isotope-labelled EGCG, the proposed pathways remain putative, since endogenous fungal metabolites, native tea constituents and host-derived compounds may change in parallel with EGCG. Future studies using carbon-13-labelled EGCG with high-resolution tandem mass spectrometry will be needed to confirm precursor-product relationships. Even so, the study delivers a clear message with practical implications. EGCG transformation is not a single uniform degradation pathway but a strain- and substrate-dependent process, meaning that fungal strain selection and fermentation conditions could be deliberately tuned to control how much EGCG is depleted and which catechin derivatives accumulate in dark tea. Given that many of the identified derivatives carry documented health-promoting activities, that tuning could shape not just the flavor of fermented tea but its functional value, while deepening our understanding of what happens to tea polyphenols between the cup and the colon.

Subject of Research: Microbial and gut microbiota biotransformation of the tea catechin EGCG during dark tea fermentation and intestinal metabolism

Article Title: Aspergillus Species and Gut Microbiota-Mediated Biotransformation of Epigallocatechin Gallate: From Dark Tea Fermentation to Intestinal Metabolism

Article References: An, Z., Ma, R., Huang, X., Zhang, Y., Huang, J., Liu, Z., Wang, K., & Zhu, M. (2026). Aspergillus Species and Gut Microbiota-Mediated Biotransformation of Epigallocatechin Gallate: From Dark Tea Fermentation to Intestinal Metabolism. Current Research in Food Science, Article 101587. https://doi.org/10.1016/j.crfs.2026.101587

Image Credits: AI Generated

DOI: Not provided

Keywords: EGCG, dark tea, Aspergillus cristatus, Aspergillus niger, fermentation, gut microbiota, metabolomics, catechins, gallic acid, theaflavins, bioavailability, biotransformation