The gut microbiota plays a key role in host nutrition and immune regulation, and its composition and function are strongly influenced by diet. Functional foods can positively influence microbial composition and activity, positioning the gut microbiome as a key target for nutritional interventions. Here, we evaluated the effects of a novel legume snack consisting of chickpeas and dark chocolate on short-chain fatty acid (SCFA) production and gut microbiota composition using an in vitro human colon model. Faecal samples from three healthy donors were inoculated under anaerobic conditions and exposed to chickpeas, chocolate, or the combined snack for 48 h. SCFA concentrations were quantified over time, and 16 S rRNA gene sequencing was used to assess microbial community composition. SCFA increased over fermentation time: the snack promoted higher acetate, propionate and, to a lesser extent, butyrate production compared with the control and individual ingredients, with the most consistent effects observed after 48 h. Microbiota profiling indicated that donor variability, incubation time and treatment contributed to differences in overall microbial composition. Differential abundance analysis, accounting for fermentation time, identified specific bacterial genera significantly associated with treatment after multiple testing correction. Moreover, several bacterial taxa showed positive associations with SCFA production and fermentation progression, suggesting links between microbial compositional shifts and functional metabolic activity during fermentation. Overall, these findings suggest that the chickpea-dark chocolate snack may represent a fermentable substrate capable of modulating microbial activity and SCFA production under in vitro conditions, providing preliminary evidence to support further investigation of its potential functional properties.

IntroductionPolyphenol-rich pomegranate extract has been shown to inhibit microbial trimethylamine (TMA) production from L-carnitine. Previous clinical studies have examined effects of polyphenol-rich interventions on fasting trimethylamine N-oxide (TMAO) concentrations but have not assessed pharmacokinetic TMAO responses following an oral carnitine challenge (OCC). We investigated whether a single dose of pomegranate extract attenuates the plasma TMAO response to an OCC in healthy adults.MethodsThis two-phase dietary intervention study enrolled 34 healthy, omnivorous adults. In Phase I, participants completed an OCC (1.5 g L-carnitine) to identify high TMAO producers (increase ≥ 5 μmol/L). Twenty high producers entered Phase II; an 18-day double-blind, randomized, placebo-controlled, crossover study with two 48-h pharmacokinetic interventions separated by a 10-day washout. Interventions consisted of an OCC with concurrent pomegranate extract (1.6 g) or placebo. Each OCC was preceded by a 48-h low-TMAO precursor run-in diet. Participants arrived fasted (>8 h), and all meals during the intervention periods were fully standardized to minimize dietary variability. Blood, urine, and stool samples were collected, and TMAO was quantified using LC-MS/MS. Differences in TMAO area under the curve (AUC) were analyzed using linear mixed-effects models.ResultsNinety one percent of participants meeting the Phase I inclusion criteria produced substantial TMAO quantities from L-carnitine. This proportion exceeds that of earlier reports. Pomegranate extract did not reduce TMAO AUC in the full Phase II cohort (placebo/pomegranate ratio 0.993, 95% CI 0.81–1.22; P = 0.945; n = 16). However, a post hoc subgroup analysis showed that the effect of the pomegranate extract on plasma TMAO differed by age and sex.ConclusionUnder tightly controlled dietary conditions, a single dose of pomegranate extract did not reduce post-OCC TMAO responses in the overall cohort. Post hoc analyses suggest potential sex- and age-dependent effects, warranting confirmation in larger, adequately powered studies.Clinical trial registrationhttps://clinicaltrials.gov/, identifier NCT06518343.

AIMS:This project aimed to investigate production of the inhibitory neurotransmitter γ-aminobutyric acid (GABA) from potential probiotic strains. We studied production in co-cultures and faecal fermentations and examined the effect of selected strains on the faecal microbiome composition and metabolome in vitro. METHODS AND RESULTS:Strains of intestinally derived Bifidobacterium adolescentis and Lactiplantibacillus plantarum from fermented cereals were grown singly, in co-culture and in faecal fermentations designed to simulate colonic conditions. Isolates synthesized varying amounts of GABA in vitro; GABA production could be increased by co-culture, lactic acid, or reduced pH but was decreased in the presence of high buffering. In faecal fermentations, selected strains inoculated singly or in combination persisted over 24 h and increased the GABA concentration without causing major disruptions in the microbiome or metabolome. Bifidobacterium adolescentis supplementation increased short-chain fatty acids acetate and propionate, and L. plantarum was associated with increased succinate levels, while all treatments exhibited a reduction in Escherichia compared to the controls. CONCLUSIONS:GABA production from these lactic acid bacteria is strain-specific and the combination of these two species shows potential for future next-generation probiotic development.

Polyphenol-rich tart cherry (TC) supplementation has previously been shown to enhance recovery from exercise-induced muscle damage (EIMD). However, no studies have investigated the effects of different doses of TC supplementation on such recovery. The molecular mechanism(s) of action through which TC elicits these effects is unknown. Here we show that 7 days TC supplementation prior to EIMD increases TC-derived plasma phenolic acid concentrations and profoundly alters the balance of structural and contractile muscle proteins, and macrophage cell infiltration within the exercised muscle. We demonstrate that plasma hippuric acid concentrations are positively correlated with muscle function in young active males. These molecular changes occurred in the absence of TC-accelerated recovery of muscle function. These findings suggest that TC warrants exploration as an intervention that may enhance musculoskeletal adaptations to training stimuli.Clinical Trial Registrationhttps://clinicaltrials.gov/study/NCT04725149, identifier: NCT04725149.

BACKGROUND & AIMS:The contribution of the gut-liver axis during liver regeneration remains poorly understood. Understanding the mechanisms underlying this process is critical for developing novel therapeutic strategies for liver diseases and transplantation. Here, we aimed at defining the role of intestinal sirtuin-1 during liver regeneration in mice. METHODS:We performed partial hepatectomy to intestinal-specific sirtuin-1 knockout mice (SIRTintKO) and wild-type littermates. Immunostaining and immunoblotting were performed on liver and intestinal tissues to assess proliferation and senescence, as well as farnesoid X receptor and fibroblast growth factor-15 protein expression. Bulk RNA sequencing was performed on liver tissues. RESULTS:We found that SIRTintKO mice had significantly reduced hepatocyte proliferation and increased hepatocyte senescence, accompanied by accumulation of bile acids in the liver that was associated with profuse parenchymal damage. Still, SIRTintKO restored liver mass at comparable levels to wild-type mice at 10 days after partial hepatectomy, which was accompanied by the activation of liver progenitor cells in the livers of knockout mice. Transcriptomic analysis of bulk RNA sequencing data from liver tissue samples at the priming (6 hours) and proliferative phase (24 hours) after partial hepatectomy highlighted that impaired hepatocyte proliferation in SIRTintKO mice coincided with the downregulation of the signal transducer and activator of transcription pathway and disruption of amino acid and lipid metabolism. Mechanistically, intestinal sirtuin-1 depletion was associated with reduced expression of farnesoid X receptor and fibroblast growth factor-15 in the small intestine. The intestinal-specific activation of farnesoid X receptor with fexaramine treatment successfully re-established hepatocyte proliferation and restored the liver parenchyma integrity in SIRTintKO mice. CONCLUSIONS:Intestinal sirtuin-1 is a key regulator of liver regeneration through upstream control of the farnesoid X receptor/fibroblast growth factor-15 axis following partial hepatectomy.

High circulating levels of trimethylamine N-oxide (TMAO) are linked to metabolic diseases, adverse outcomes after heart failure, and atherogenic effects in animal models and in human subjects. l-Carnitine and choline are major dietary precursors of TMAO. These are first converted to trimethylamine (TMA) by gut microbiota, which is absorbed by the host and converted into TMAO by hepatic flavin-containing monooxygenases (FMOs). The minimal absorption of pomegranate polyphenols by the host suggests that they may reach the colon for further metabolism by the gut microbiome. This study investigates the ability of a polyphenol-rich pomegranate extract to inhibit TMA production by human fecal microbiota. Batch fermentations were conducted with 1% human fecal inoculum, l-carnitine, or choline, and a pomegranate extract (anaerobic, pH 6.6-7.1, 37°C) for 24 or 48 h. Methylamines were quantified using LC-MS/MS with isotopically labeled internal standards. The pomegranate extract significantly delayed and reduced the rate of TMA production from both choline and l-carnitine. The effect was dose-dependent for l-carnitine, with the highest dose delaying the average midpoint of l-carnitine metabolism by 16 h (95% CI = 8.4-24; p = 0.001). The pomegranate extract significantly reduced TMA production from choline and l-carnitine in vitro.

It was recently reported that degradation of anthocyanins in the anaerobic human colon occurs via both microbiota-dependent and spontaneous processes. However, the existing literature only describes the spontaneous degradation of anthocyanins in aerobic conditions. We investigated the loss of cyanidin-3-O-glucoside (Cy3Glc) over time under aerobic and anaerobic conditions. In anaerobic conditions, spontaneous breakdown products were consistent with Cy3Glc undergoing a classic pH-dependent transformation to colourless intermediates (i.e., cyanidin (Cy) hemiketal-glucoside, Cy chalcone-glucoside, Cy chalcone quinone-glucoside, 2,4,6-trihydroxyethenylbenzene-glucoside), but no other breakdown products. In contrast, under aerobic conditions, in addition to the pH-dependent intermediates, multiple other breakdown products were identified, including the previously reported protocatechuic acid and phloroglucinaldehyde, and several products of oxidation reactions reported for the first time (coumarin-glucoside, 2,4,6-trihydroxyphenylacetic acid, 2,4,6-trihydroxyphenylglyoxylic acid, and 3,4-dihydroxyphenylglyoxylic acid). These observations show degradation of anthocyanins is completely different in aerobic compared to anaerobic conditions, and useful information of the spontaneous degradation of anthocyanins cannot be obtained using aerobic conditions.

Over recent decades, dietary patterns have changed significantly due to the increasing availability of convenient, ultra-processed refined foods. Refined foods are commonly depleted of key bioactive compounds, which have been associated with several deleterious health conditions. As the gut microbiome can influence the brain through a bidirectional communication system known as the 'microbiota-gut-brain axis', the consumption of refined foods has the potential to affect cognitive health. In this study, multi-omics approaches were employed to assess the effect of a refined diet on the microbiota-gut-brain axis, with a particular focus on bile acid metabolism. Mice maintained on a refined low-fat diet (rLFD), consisting of high sucrose, processed carbohydrates and low fibre content, for eight weeks displayed significant gut microbial dysbiosis, as indicated by diminished alpha diversity metrics (p < 0.05) and altered beta diversity (p < 0.05) when compared to mice receiving a chow diet. Changes in gut microbiota composition paralleled modulation of the metabolome, including a significant reduction in short-chain fatty acids (acetate, propionate and n-butyrate; p < 0.001) and alterations in bile acid concentrations. Interestingly, the rLFD led to dysregulated bile acid concentrations across both the colon (p < 0.05) and the brain (p < 0.05) which coincided with altered neuroinflammatory gene expression. In particular, the concentration of TCA, TDCA and T-alpha-MCA was inversely correlated with the expression of NF-kappa B1, a key transcription factor in neuroinflammation. Overall, our results suggest a novel link between a refined low-fat diet and detrimental neuronal processes, likely in part through modulation of the microbiota-gut-brain axis and bile acid dysmetabolism.

Emulsions are commonly used to fortify beverages with oil-soluble nutrients. Such emulsions must be physically stable, prevent the cargo nutrient from chemical degradation, and release the nutrient at the right time during digestion. Wood hemicelluloses, with their proven capability in producing nanoemulsions and preventing oil oxidation, have the potential to facilitate such a fortification method. These hemicelluloses are unique, as the presence of residual lignin was proven essential in their functionality. However, the structural interplay between the residual lignin and the polysaccharide fraction at the oil-water interface is unknown, including their effects on the storage stability and digestibility of a loaded bioactive compound. We studied the performance of emulsions stabilized by two grades of birch glucuronoxylans (GX) as vessels for vitamin D3. Both GXs showed similar protective performance to methylcellulose regardless of the lignin content during storage and the gastric phase of digestion. The unrefined GX, which contained larger lignin moieties, delayed the release of vitamin D3 during intestinal digestion. We concluded that the interface is primarily built of a polysaccharide layer with the lignin moieties facing towards the oil phase. We have thus highlighted the potential of GX-stabilized emulsions as carriers of vitamin D3. Additionally, these results advance the understanding of lignin-polysaccharide interplay towards a rationalized strategy in biorefining of wood hemicelluloses, for which the ratio of lignin-to-polysaccharide can be adjusted during extraction and post-extraction treatments to obtain a tailor-made emulsifier for various emulsion-based products.

Background & Aims: Senescence has been reported to have differential functions in cholangiocytes and hepatic stellate cells (HSCs) during human and murine cholestatic disease, being detrimental in biliary cells and anti-fibrotic in HSCs. Cholestatic liver disease is associated with loss of intestinal barrier function and changes in the microbiome, the mechanistic cause of which is undetermined. Methods: Intestinal samples were analysed from controls and patients with primary sclerosing cholangitis, as well as wild-type (WT) and p16-3MR transgenic mice. Cholestatic liver disease was induced by bile duct ligation (BDL) and DDC diet feeding. Fexaramine was used as an intestinal-restricted FXR agonist and antibiotics were given to eliminate the intestinal microbiome. Senescent cells were eliminated in p16-3MR mice with ganciclovir and in WT mice with the senolytic drug ABT-263. In vitro studies were done in intestinal CaCo-2 cells and organoids were generated from intestinal crypts isolated from mice. Results: Herein, we show increased senescence in intestinal epithelial cells (IECs) in patients with primary sclerosing cholangitis and in mice after BDL and DDC diet feeding. Intestinal senescence was increased in response to reduced exposure to bile acids and increased presence of lipopolysaccharide in vitro and in vivo during cholestatic liver disease. Senescence of IECs was associated with lower proliferation but increased intestinal stem cell activation, as supported by increased organoid growth from intestinal stem cells. Elimination of senescent cells with genetic and pharmacological approaches exacerbated liver injury and fibrosis during cholestatic liver disease, which was associated with increased IEC apoptosis and permeability. Conclusions: Senescence occurs in IECs during cholestatic disease and the elimination of senescent cells has a detrimental impact on the gut-liver axis. Our results point to cell-specific rather than systemic targeting of senescence as a therapeutic approach to treat cholestatic liver disease. (c) 2024 The Authors. Published by Elsevier B.V. on behalf of European Association for the Study of the Liver (EASL). This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

TenA thiamin-degrading enzymes are commonly found in prokaryotes, plants, fungi and algae and are involved in the thiamin salvage pathway. The gut symbiont Bacteroides thetaiotaomicron (Bt) produces a TenA protein (BtTenA) which is packaged into its extracellular vesicles. An alignment of BtTenA protein sequence with proteins from different databases using the basic local alignment search tool (BLAST) and the generation of a phylogenetic tree revealed that BtTenA is related to TenA-like proteins not only found in a small number of intestinal bacterial species but also in some aquatic bacteria, aquatic invertebrates, and freshwater fish. This is, to our knowledge, the first report describing the presence of TenA-encoding genes in the genome of members of the animal kingdom. By searching metagenomic databases of diverse host-associated microbial communities, we found that BtTenA homologues were mostly represented in biofilms present on the surface of macroalgae found in Australian coral reefs. We also confirmed the ability of a recombinant BtTenA to degrade thiamin. Our study shows that Bt tenA -like genes which encode a novel sub-class of TenA proteins are sparingly distributed across two kingdoms of life, a feature of accessory genes known for their ability to spread between species through horizontal gene transfer.

AbstractSpot urinary polyphenols have potential as a biomarker of polyphenol-rich food intakes. The aim of this study is to explore the relationship between spot urinary polyphenols and polyphenol intakes from polyphenol-rich food sources. Young adults (18–24 years old) were recruited into a sub-study of an online intervention aimed at improving diet quality. Participants’ intake of polyphenols and polyphenol-rich foods was assessed at baseline and 3 months using repeated 24-h recalls. A spot urine sample was collected at each session, with samples analysed for polyphenol metabolites using LC-MS. To assess the strength of the relationship between urinary polyphenols and dietary polyphenols, Spearman correlations were used. Linear mixed models further evaluated the relationship between polyphenol intakes and urinary excretion. Total urinary polyphenols and hippuric acid (HA) demonstrated moderate correlation with total polyphenol intakes (rs = 0·29–0·47). HA and caffeic acid were moderately correlated with polyphenols from tea/coffee (rs = 0·26–0·46). Using linear mixed models, increases in intakes of total polyphenols or polyphenols from tea/coffee or oil resulted in a greater excretion of HA, whereas a negative relationship was observed between soya polyphenols and HA, suggesting that participants with higher intakes of soya polyphenols had a lower excretion of HA. Findings suggest that total urinary polyphenols may be a promising biomarker of total polyphenol intakes foods and drinks and that HA may be a biomarker of total polyphenol intakes and polyphenols from tea/coffee. Caffeic acid warrants further investigation as a potential biomarker of polyphenols from tea/coffee.

Scope Some dietary interventions with berry fruits, berry fruit extracts, and purified anthocyanins have been reported to beneficially alter lipoprotein profiles in hyperlipidemic participants. The major anthocyanins in human diets are glycosides of cyanidin and delphinidin, and structure can influence both absorption and bioactivity. The aim of this study is to determine the effects of two major types of anthocyanins on low‐density lipoprotein cholesterol and other cardiometabolic markers for cardiovascular disease (CVD) risk in hyperlipidemic individuals. Methods and results Fifty‐two hyperlipidemic participants complete this randomized, placebo‐controlled, double‐blind, three arm crossover trial. Participants ingest capsules containing 320 mg of anthocyanins (bilberry trihydroxy‐type or black rice dihydroxy‐type) or placebo once daily for 28 days. Biomarkers of CVD risk are measured before and after the intervention period. Compared to the placebo, neither anthocyanin treatment significantly ( p < 0.05) changes circulating levels of lipoproteins (total‐/high‐density lipoprotein (HDL)‐/low‐density lipoprotein (LDL)‐cholesterol, triglycerides, Apolipoprotein B (ApoB)), biomarkers of glycemic control (fasting glucose, fructosamine), biomarkers of HDL function (ApoA1, HDL3, paraoxonase‐1 (PON1) arylesterase, and lactonase activities), or plasma bile acids. Conclusions These data do not support the notion that regular consumption of anthocyanins beneficially affects glycemic control or lipoprotein profiles or functions. It is possible the no effect observation is due to the relatively short duration of treatments.

Prediction of retention times (RTs) is increasingly considered in untargeted metabolomics to complement MS/MS matching for annotation of unidentified peaks. We tested the performance of PredRet (http://predret.org/) topredict RTs for plant food bioactive metabolites in a data sharing initiative containing entry sets of 29-103 compounds (totalling 467 compounds, >30 families) across 24 chromatographic systems (CSs). Between 27 and 667 predictions were obtained with a median prediction error of 0.03-0.76 min and interval width of 0.33-8.78 min. An external validation test of eight CSs showed high prediction accuracy. RT prediction was dependent on shape and type of LC gradient, and number of commonly measured compounds. Our study highlights PredRet's accuracy and ability to transpose RT data acquired from one CS to another CS. We recommend extensive RT data sharing in PredRet by the community interested in plant food bioactive metabolites to achieve a powerful community-driven open-access tool for metabolomics annotation.