Meiyn Wng*, Jinying Li Ting Hu*, Hui Zho*

a Tianjin Key Laboratory of Food and Biotechnology, School of Biotechnology and Food Science, Tianjin University of Commerce, Tianjin 300134, China.

b Hubei Key Laboratory for EFGIR, Huanggang Normal University, Huanggang 438000, China.

Keywords:

Tea polyphenols

Metabolism

Gut microbiota

Bioeffects

A B S T R A C T

Tea represents an abundant source of naturally occurring polyphenols.Tea polyphenols (TPs) have received growing attentions for its wide consumption in the world, and more importantly its pleiotropic bioeffects for human health.After ingestion, TPs may undergo absorption and phase II reaction in the small intestine,and most undigested proportion would be submitted to the colon to interact with gut microbiota.Interactions between gut microbiota and TPs are bidirectional, including not only bacteria-mediated TPs metabolism,e.g., removal of gallic acid moiety and ring fission to release phenolic acid catabolites, but also TPs-based modification of bacterial profiles.Crosstalk between TPs and gut microbes may benefit for gut barrier function,for example, improvement of the intestinal permeability to alleviate inflammation.Moreover, by reshaping microbial composition and associated metabolites, TPs may exert a systemic protection on host metabolism,which contributes to improve certain chronic metabolic disorders.Given that, further understanding of the metabolic fate of TPs and interplay with gut microbiota as well as potential health-promoting effects are of great significance to development and application of tea and their polyphenolic components in the future as dietary supplements and/or functional ingredients in medical foods.

1.Introduction

Tea, leaf or bud from the plantCamellia sinensis, are the most popularly consumed beverage around the world, second only to water.Based on the manufacturing process, tea is mainly categorized into unfermented green tea, semifermented oolong tea,fermented black tea, and post-fermented dark tea.Tea is extremely rich in dietary polyphenols [1].Dominant polyphenols in green tea (GTPs) are characterized by flavan-3-ol monomers, including(?)- epigallocatechin-3-gallate (EGCG), (?)-epigallocatechin (EGC),(?)-epicatechin-3-gallate (ECG), and (?)-epicatechin (EC) [2].A typical cup of brewed green tea beverage (e.g., 2.5 g of leaves steeped for 3 min in 250 mL of water) contains 240-320 mg of catechins,of which EGCG is the most abundant, accounting for almost half of the total GTPs [3,4].In fermented tea, however, these flavan-3-ol monomers are transformed via the peroxidase- and oxidase-mediated reaction into novel dimeric and polymeric derivatives, known as theaflavins ( TFs) and thearubigins (TRs).The common TFs are theaflavin, theaflavin-3-O-gallate, theaflavin-3’-O-gallate and theaflavin-3,3’-O-digallate.In brewed black tea, GTPs, TFs, and TRs respectively contribute 3%-10%, 2%-6%, and > 20% to its dry weight [4,5].Recently, tea polyphenols (TPs) have been in the spotlight as a result of their potential to interplay with gut microbiota and multiple benefits for human health [6,7].

Upon ingestion, TPs may undergo an extensivein vivometabolic process such as hydrolysis of the ester bonds for catechin gallates(e.g., EGCG and ECG ) in gut, phase II biotransformation in gut and liver, and interplay with gut microbiota, releasing free catechins,glucuronidated/sulfated/methylated conjugates, phenolic acids, and/or other catabolites [8-10].Of note, the relationship between TPs and gut microbiota is bidirectional – microbiota metabolizing TPs and in turn, TPs shaping microbial profiles.Indeed, emerging evidence in recent years has pointed to the key role of dietary phytochemicals including TPs in the modulation of the composition and function of the intestinal flora.In this direction, TPs are found to positively modify the microbial diversity and composition in the intestine,and further regulate the levels of gut microbe-derived metabolites,e.g.short-chain fatty acids and bile acids [11-13].As a result, the crosstalk between TPs and gut microbes may not only benefit for the improvement of local damage within gut, such as the intestinal inflammation and permeabilization, but also for the prevention and/or treatment of some systemically metabolic diseases, such as obesity and type 2 diabetes mellitus (T2DM) [14,15].

This review focuses on the crosstalk between TPs and gut microbiota.On one hand, it provides a summary of the intestinal fate of TPs after ingestion, with an emphasize of the key role of gut bacteria in this metabolic process.On the other hand, the modulatory effect of TPs on gut microbiome as well as associated healthpromoting effects are discussed.The aim is from the perspective of gut microbiota to explain the underlying mechanism by which tea or TPs consumption benefits human health.

2.Intestinal fates of TPs

After ingestion, TPs are absorbed to different extents in the small intestine depending on their chemical structures and undergo an intensive biotransformation in gut by phase II enzymes as well as local microbiota.Understanding the metabolic process and metabolites of TPs are important as they define the molecular targets and bioactivities of TPs.

2.1 Small intestinal fate: absorption and phase II conjugation

GTPs maintain relatively stable during gastric transit, but they are rapidly absorbed and/or biotransformed during passage through the small intestine [16,17].Gut plays a dominant role in the bioconversion of flavan-3-ols prior to a liver detoxification process [18].Typically,abundant phase II enzymes in enterocytes are capable of conjugating some TPs to generate glucuronidated, sulfatedand, and methylated(methoxy) derivatives upon disposal of uridine-5’-diphosphateglucuronosyl-transferases (UGTs), sulfotransferases (SULTs),and catechol-O-methyltransferases (COMTs), respectively [19].In this direction, a clinical trial was performed to investigate the kinetic process of GTPs among 10 healthy volunteers who consumed 500 mL of green tea that contained 257 μmol EGC, 230 μmol EGCG, 58 μmol EC and 49 μmol ECG [20].A total of 7 metabolites were identified in the plasma in the form of glucuronided, methylglucuronided and methyl-sulfated conjugates of EC and glucuronided,sulfated and methyl-sulfated EGC with 29-126 nmol peak concentrations occurring approximately 2 h post-intake, indicating a biotransformation in the upper gastrointestinal tract [20].Methylation of EC and EGC appeared to occur only after the glucuronidation or sulfation reaction.Also, unchanged EGCG and ECG were detected in the plasma, with a peak concentration of 55 and 25 nmol, respectively [20].Actually, few dietary polyphenolics are detected in plasma in their intact form after ingestion.The exceptional behavior of EGCG and ECG in this regard might be explained by the presence of the 3-O-galloyl moiety, since gallic acidper seis well absorbed with a urinary excretion of 37% of ingestion [21,22].The urinary excretion of flavan-3-ol metabolites 0–24 h after green tea administration corresponded to 27% of intake, which, summed to an 8% excretion of phase II conjugated GTPs originating from absorption in the small intestine, accounts for 35% absorption [20,23].Another human intervention study estimated the 48 h bioavailability of GTPs was close to 62% [24].In brief, these data imply that GTPs are highly bioavailable, being absorbed and excreted to a much greater extent than most other polyphenols.

Compared with GTPs, nutrikinetics of TFs and TRs principally derived from the fermented tea have received only limited attention,although black tea consumption is far more extensive in the United States and Europe.Such researches have focused upon the absorption of monomeric flavan-3-ols and flavonols (e.g., quercetin and kaempferol) from black tea beverages with or without addition of milk [25,26].For TFs, only trace amounts (< 0.001% of intake) was absorbed in the upper gastrointestinal tract and TRs are not expected to be absorbed [12,27].These TPs would be submitted to the colon and undergo locally microbial degradation, according to recent mice and human studies [9,28].

2.2 Colonic fate: microbial-media production of phenyl-γvalerolactones (PVLs) and phenylvaleric acids (PVAs)

Unabsorbed TPs in the upper gastrointestinal tract will sequentially pass into the large intestine, where harbors an enormous population of microbiota with an estimated load of up to 1010–1012CFUs/mL [29].Arising through gut microbiota catabolism,PVLs and their related PVAs, as well as the smaller phenolic acid catabolites are reported as the main metabolites of catechins as well as polymeric flavan-3-ols in human [30-32].The peak concentration of PVLs in the plasma occurred 5-12 h post-consumption of green tea,suggesting the importance of microbial activity on TPs degradation.Actually, the recognition of the key role of microbial community in thein vivometabolism of TPs leads to a resurgence of interest in the colonic degradation and metabolites, and the consequences on bioavailability and bioactivity of TPs [33-36].

After ingestion of green tea by human, it was estimated over 70% of GTPs reaching the colon and subjected to bacteriamediated catabolism [18].For galloylated flavan-3-ol monomers including EGCG and ECG, the microbial metabolism generally starts with a rapid cleavage of the gallic acid moiety by microbial esterases to release free EGC and EC [37-39].By screening 169 bacterial strains,Raoultella planticola,Enterobacter aerogenes,Bifidobacterium longumsubsp.infantis, andKlebsiella pneumoniaesubsp.pneumoniaewere found to be involved in the degalloylated reaction of EGCG [39].Further catabolism of EGC and EC in the colon includes C-ring opening, A-ring fission, PVL/PVA formation, and smaller phenolic acid production, and the proposed pathway for this bacterial catabolism is illustrated in Fig.1.When free epi(gallo)catechins are released from the bacteria-mediated degalloylation, their C-ring would undergo ring opening yielding corresponding diphenylpropan-2-ol intermediates.And then, these diphenylpropan-2-ols are converted to PVL/PVA metabolites via ring-A fission by the action of specific bacterial species,Flavonifractor plautii., for instance [36].Typical PVLs/PVAs derived from EGC and EC are 5-(3’,4’,5’-trihydroxyphenyl)-γvalerolactone (M4)/5-(3’,4’,5’-trihydroxyphenyl)-γ-hydroxyvaleric acid and 5-(3’,4’-dihydroxyphenyl)-γ-valerolactone (M6)/5-(3’,4’-dihydroxyphenyl)-γ-hydroxyvaleric acid, respectively, and/or 5-(3’,5’-dihydroxyphenyl)-γ-valerolactone (M6’)/5-(3’,5’-dihydroxyphenyl)-γ-hydroxyvaleric acid and 5-(3’-hydroxyphenyl)-γ-valerolactone/5-(3’,4’-dihydroxyphenyl)-γ-hydroxyvaleric acid generated through a further 4’-deoxidation reaction [39-42].Among which, 5-(3’,5’-dihydroxyphenyl)-γ-hydroxyvaleric acid was identified as the major metabolite of EGCG by rat intestinal flora [39], whereas M6 was the main component determined in the cecum and colon of rats after EC intake [43].Interestingly, M4, one major PVL from EGCG, can be dehydroxylated at C-5’ position to convert into the major PVL metabolite of ECG – M6, indicating a relevance between thein vivobiotransformation of EGCG and ECG.

Fig.1 Proposed metabolic pathways for EGCG and ECG in the large intestine.Based on data from literatures [39,44].

The intestinal fate of GTPs usually does not end at PVLs/PVAs, as these structures can be further catabolized by gut microbiota to result in various smaller phenolic acids, such as phenylpropionic and benzoic acid derivatives, through a successive loss of carbon atoms from the side chain by means ofβ-oxidation(Fig.1) [23,43,45].However, whether these catabolic processes occur in the colon, after absorption or both, remains uncertain.Also,whether 3-(3’,4’-dihydroxyphenyl)propionic acid is catabolized viaα-oxidation to produce 2-(3’,4’-dihydroxyphenyl)acetic acid is another unanswered question [46].

PVLs/PVAs and simple phenolic acids can be absorbed and further biotransformed by phase II enzymes in colonocytes and/or hepatocytes to give conjugated derivatives, and majority of them are ultimately excreted in urine.In this regard, the recovery of radioactivity in urine was evaluated to be 78% after ingestion of14C-EC by rats [43].In addition, faeces may contain relatively small amounts of undigested GTPs, unabsorbed catabolites, and conjugated catabolites released from the enterocytes/colonocytes or excreted in bile [47].For example, after feeding14C-EC to male volunteers,approximately 9% of the ingested radioactivity was voided in faeces over a 72 h period, principally as PVAs/PVLs as well as a smaller amount of 3-(3’-hydroxyphenyl)propionic acid [30].

With regard to theaflavin and its gallates, they are resistant to absorption in their intact form not only in the small intestine but in the colon.An evidence was from a human feeding study, in which two healthy subjects respectively consumed high up to 700 mg of TFs, equivalent to approximately 30 cups of black tea [27].Peak concentration of TFs was, however, determined as low as 0.5 and 1.0 μg/L in the plasma of these two volunteers, respectively [27].Although almost no intact theaflavin or galloylated theaflavin was present in the plasma after black tea consumption by human, a significant increase of phenolic acid derivatives were detected in their urine and feaces, which hinted that after ingestion, TFs might undergo a gut flora-mediated catabolism first, and then the postabsorption [48-50].

For thea flavin gallates, their degradationin vivogenerally begins with the hydrolysis of the ester bond by gut microbiota to remove gallic acid moiety.For example, theaflavin-3,3’-digallate could be successively degalloylated to convert to theaflavin-3 (or 3’)-gallate and then to free theaflavin byLactobacillus plantarum299v andBacillus subtilis, with a large release of gallic acid[51].In vivo, some of the gallic acid appeared to be methylated by mammalian enzymes to form 3- and 4-O-methygallic acid, while most was decarboxylated,probably by bacterial enzymes, to afford pyrogallol that might be further subjected to phase II metabolism [52].Compared with gallic acid, the theaflavin skeleton is relatively resistant to the colonic bacteria-mediated degradation.A recent study detected a total of 964 μmol of phenolic catabolites in 0–30 h urine of human who consumed 1 g of thea flavin extract that contained 177 μmol thea flavin,307 μmol theaflavin-3-O-gallate, 172 μmol theaflavin-3’-O-gallate,and 332 μmol thea flavin-3,3’-O-digallate [52].Among these phenolic catabolites, 625 μmol of sulfated pyrogallols were identified as gallic acid metabolites, whereas only 2.1 μmol of 3-hydroxyhippuric acid and 7.3 μmol of 4-hydroxyhippuric acid were considered to be associated with theaflavin skeleton degradation.Another evidence pointing to the stability of thea flavin in the distal gut was that over a 24 h period of incubation with human feaces, 67% of theaflavin skeleton still remained undisposed, which was much higher than flavan-3-ol monomers such as EGCG, ECG, EC and EGC [23,52,53].

Data from human urine metabolome suggested that only limited amount of theaflavin skeleton was degraded, and produced 3-(4’-hydroxyphenyl)propionic acid through series of ring fission reactions mediated by the combination of bacterial and mammalian enzymes [52].Potentially, 3-(4’-hydroxyphenyl)propionic acid may be further catabolized into 4-hydroxybenzoic acid throughβ-oxidation of the fatty acid side chain, and possibly glycinated in the liver to afford 4-hydroxyhippuric acid ultimately [52].A possible metabolic pathway of thea flavin gallates after ingestion is proposed in Fig.2.In addition, traces of 3’- and 4’-hydroxyphenylacetic acid and 3’-methoxy-4’-hydroxymandelic acid were identified in human urine 0-30 h post-intake of TFs [52], which was assumed to be linked to the breakdown of the basic thea flavin ring.However, exactly how much of these phenolic acid catabolites are derived from the catabolism of thea flavin skeleton remains to be ascertained in the future study.

Fig.2 Exemplified biotransformation of TFs in gut.The illustrated example is thea flavin-3-O-gallate.It assumed that thea flavin-3’-O-gallate and thea flavin-3,3’-O-digallate undergo a similar degalloylated process in the colon to release thea flavin.Red arrows represent bacteria-mediated steps, and blue arrows indicate mammalian enzyme-involved bioconversions.GlcUA, glucuronide.Based on data from literature [52].

2.3 Inter-individual variability in metabolism and metabolites of TPs

Inter-individual variation in the metabolic process and metabolites have been investigated for a variety of phytochemicals, and it is well established that multiple factors including sex, age, and dietary habits may affect thein vivokinetics of these components [54].For TPs, since most of them pass unabsorbed to the distal gastrointestinal tract after ingestion, local microbial community is arguably one key factor affording the individual discrepancy observed during their metabolic process [55].In this context, inter-individual variations in the composition and function of the intestinal flora might give rise to some specific metabolites.

Overwhelming clinical evidence suggests the differences in the profiles of the microbial-derived catabolites of flavan-3-ol monomers and oligomers [56-58].For example, after consumption of green tea for 7 consecutive days, great individual variation was found not only in the amounts but in the components of PVLs in urines among eight healthy adults, and the variation coefficient for these PVLs varied in most cases between 200% and 300% [58].Additionally,in vitrofaecal fermentation is frequently used in the primary determination of both the gut bacterial catabolism and possible individual variation derived from extremely diverse microbial profiles in gut.In this direction, several GTPs was respectively incubated with fecal slurries from 3 donors, and results showed that catabolite profiles and amounts varied 2–18 folds among volunteers [23].Nonetheless, there is limited data regarding the specific microbial strains and enzymes involved in the biotransformation of TPs to specific metabolites and factors potentially modulating their activities.

Available researches have proposed food matrix as a possible factor influencing the absorption and metabolism of flavan-3-ols when they are consumed in foods as part of a daily diet [59,60].In this regard, a nutrition-based nutrikinetic model was established to evaluate the modulatory capacity of the food matrix and the colonic microbiome to the intestinal metabolism of black tea as well as other flavan-3-ol-rich foods, such as red wine [61].It was interestingly discovered that the production of typical PVL conjugates was determined by the biotransformation capacity of individuals’gut microbiome rather than by the specific food matrix, and it displayed a positive correlation with the abundance of Clostridia and Actinobacteria, includingF.plautii,Clostridium leptum,Sporobacter termitidis,Eubacterium ramulus, andRuminococcus bromii, and the genusPropionibacterium.

In conclusion,in vivometabolism and metabolites of TPs might vary greatly depending on individual microbial profiles.Elucidation of metabotypes is of interest to further understand the healthpromoting effects of some specific intestinal metabolites such as PVLs on an individual basis, which might change the way in which the chronic bioeffects of TPs are studied.In this direction, a doubleblind randomized controlled trial demonstrated that the equol producer phenotype is critical in unlocking the vascular benefits of equol, a gut flora-derived metabolite of the isoflavone daidzein and available in 30% of Western populations after soy intake, which explained the controversial cardioprotective property of daidzein-containing foods [62].Given specific colonic metabolites of isoflavone where metabotypes have been linked to personalized health benefits, flavan-3-ols,such as TPs, or flavan-3-ols-rich food, such as tea, may benefit some individuals more than others.These newly findings suggest a dietary phytochemical-based individualized treatment may be a warranted research topic in the future.

3.Crosstalk between TPs and gut microbiota: local and systemic effects

After a sequential metabolism in the gastrointestinal tract, the bioavailable fraction of TPsin vivomight consist of the intact parent compound, free TPs without gallic acid moiety, PVLs/PVAs, sulfated/glucuronidated/methylated conjugates, and small phenolic acids.The intestinal metabolism is highly relevant to TPs’ bioactivities, and the metabolites might confer more or similar, or perhaps less activities.Actually, emerging evidence indicates while gut flora metabolizes TPs, TPs shape microbiota, and this crosstalk between TPs and bacteria might benefits each other and ultimately exert a positive effect on health [63-65].TPs and their metabolites in gut can exhibit a locally biological effects, for instance, protection of the intestinal barrier, and may also exert some systemic actions when entering into the bloodstream.TPs-induced modification of the gut bacteria and associated resultant health benefits are summarized in Fig.3.Notably,local effects of TPs within gut can in part underlie their systemic benefits, for example, positively shaping microbial profiles and further modification of microbial metabolites to ultimately rebuild glucose and lipid homeostasis [13,66].

Fig.3 Potential modification of TPs consumption on gut microbial profiles as well as associated local and systemic effects.

3.1 TPs-induced modification of gut microbiome

In the past 20 years, there has been overwhelming evidence to support the important role of gut bacteria in the digestion of dietary phytochemicals [67-69], including TPs as discussed above.Indeed, in addition to microbiota catabolizing TPs, TPs shape gut microbiome,which indicates the relationship between TPs and gut microorganism is bidirectional [70].

TPs are capable of reshaping the gut flora composition in a positive way, for example, modulating the ratio of Firmicutes to Bacteroidetes (F/B ratio) under the pathological condition such as obesity.Firmicutes and Bacteroidetes are two dominant phyla among the massive community of microbes harbored in the human gut,mainly acting on the metabolism of dietary fiber involving a complex metabolic energy-harvesting dynamic.High F/B ratio is reported to be related to obesity in human and mice, while weight loss intervention was accompanied by an increased abundance of Bacteroidetes [71,72].TPs were reported to be capable of ameliorating high-fat diet induced body weight gain or obesity through modification of gut dysbiosis,particularly by means of enriching Bacteroidetes and lowering F/B ratio [14,73].The effective forms of TPs were diverse, including EGCG3”Me [74], and polyphenol extract from green tea [75-77]and oolong tea [78].Although many studies determined an increased F/B ratio after TPs-based intervention, two exceptions were reported -tea infusions did not obviously affect the F/B ratio [79]and Pu-erh tea extract actually increased the F/B ratio as compared to the control group with high-fat diet [80].The inconsistent results of changes in bacterial composition at the phylum level in different researches are probably due to differences in bacteria at the lower taxonomic ranking or due to different dosages of TPs.Given that, evaluating the effects of TPs on the intestinal bacteria at a more specific taxonomic classification such as genus or even species, and quantitation of active components may help establish a more convincing causal relationship between TPs consumption and microbial composition.

BifidobacteriumandLactobacillusare well recognized beneficial bacteria in human gut and have been using as commercial probiotics.The abundance of these two probiotics was significantly enriched by TPs preparations from Pu-erh tea, oolong tea, and green tea [81-85].BothBifidobacteriumandLactobacillusare known to prefer to grow at relatively low pH condition.Thus, the increased proportions of these two bacterial genera might be of relevance to the decrease of pH during TPs administration [86,87].

Recently,Akkermansia muciniphilahas been proposed as a promising bacterial species with gut-protective and health-promoting potentials since it was first identified in human feaces in 2004 [88,89].A.muciniphilais relatively abundant in healthy individuals,accounting for about 4% of the bacterial population in gut.However,its abundance declines significantly in certain metabolic disorder condition, such as type 2 diabetes mellitus (T2DM) and obesity [90,91].The declined proportion ofA.muciniphilawas found to be normalized when treated with TPs-based dietary supplements such as green tea powder [82], Pu-erh tea extract [92], and EGCG [93].For example,polyphenols-rich Pu-erh tea extract was found to effectively prevented mice from high-fat diet-induced obesity by remodeling the disrupted intestinal homeostasis, mainly characterized by enriching the relative abundance ofA.muciniphila[80].Further specific bacteria administration revealed thatA.muciniphilapromoted white adipose tissue browning, improved the glucose and lipid dysmetabolism, and prevented mice from weight gain when fed with a high-fat diet [80].

In addition to promoting the growth of some beneficial bacterial species, TPs inhibit the growth of some pathogenic strains, e.g.,Clostridium difficile,C.perfringens,Bacteroidesspp.,Escherichia coliO157:H7, andHelicobacter pylori[94,95].Mechanistically, TPs may target cell membranes of pathogenic bacteria, damage their lipid bilayer, promote the production of hydrogen peroxide, and further lead to the cell membrane permeabilization to inhibit the growth of potential pathogens [96,97].

The above discussion illustrates that TPs improving the bacterial dysbiosis is usually accompanied by multiple positive modifications in host metabolic parameters, such as weight, blood lipid, blood glucose, and insulin sensitivity.In this context, we postulate that TPs might represent a group of promising dietary phytochemicals capable of positively shaping the intestinal profiles and further remodeling the metabolic homeostasis of host, which is beneficial for the treatment of some systemically metabolic syndromes.

3.2 The intestinal barrier protective property: local effect of TPs within gut

Gut barrier is a highly selective barrier, which allows a continuous influx of nutrients, ions and water, but at the same time prevents multiple luminal insults, e.g., endotoxins and dietary antigens [98].When the barrier is dysfunctional, it may enable an invasion of external factors and trigger local injuries within gut, such as inflammation, colitis, and inflammatory bowel disease (IBD) among others [99-101].As one important structural element of the intestinal barrier, gut microbiota plays a vital role in sustaining the intestinal microecosystem [102,103].Interactions between TPs and gut microbiota can lead to a metabolism of TPs on one hand, and on the other hand, TPs as well as their metabolites would reshape microbial profiles and confer protective actions onto gut barrier.

A feeding study with tea discovered a substantial increase in the overall diversity of gut microbiota in mice after infusion of oolong tea,black tea, and green tea, respectively [79].Also, the composition of gut bacteria was modified, with decreased proportions of 8 phylotypes(e.g.,Allobaculumsp.OTU383 andC.leptumOTU450) and enriched abundances of six species includingAlistipessp., Lachnospiraceae,S24-7,Akkermansiasp., andRikenella microfusus[79].Altered composition of the gut microbiota may further lead to a change in their function.For example, typical metabolites ofAlistipesandRikenella, i.e., short chain carboxylic acids including succinic acid, propionic acid, acetic acid, and alcohols, enable tight junction assembly and epithelium differentiation in gut, which altogether can contribute to strengthen of the gut barrier [104,105].EC feeding was demonstrated to positively shape the integrity and function of the tight junctions in high-fat diet feeding mice, and protect the intestinal barrier from permeabilization [106].With regard toAkkermansia,a mucin-degrading bacterium, its adequate proportion is directly associated with the health of the intestinal mucosal barrier [107].Anotherin vivostudy demonstrated that a dietary supplementation with 2% (m/m) green tea extract for 8 weeks protected high-fat diet feeding mice from inflammation and endotoxin translocation [108].The intestinal barrier protection of green tea extract may contribute,at least in part, to its ability to remodel gut microbial homeostasis,mainly manifested in enriching the overall bacterial diversity and decreasing F/B ratio [108].

In addition to improving gut microbial dysbiosis, TPs might exert their protective action on the intestinal barrier by downregulating certain inflammatory response pathways, suppressing inflammatory factors, and/or giving antioxidant activity.In dextran sodium sulfate-induced colitis model of mice, total GTPs (or EGCG)administration dramatically decreased the levels of inflammatory markers, e.g.interleukin-6, tumor necrosis factor alpha, and serum amyloid A, and significantly ameliorated the interruption of the colonic architecture [109].The anti-inflammatory activity of total GTPs (or EGCG) was estimated to be comparable with sulfasalazine,a commonly used drug for treatment and maintenance in IBD [109].In the aspect of antioxidant capacity, EGCG showed a better restorative effect on damaged glutathione, an essential intracellular element to protect enterocytes against free radicals, in colon than sulfasalazine [109].Additionally, GTPs were reported to suppress the intestinal inflammation by means of down-regulating toll-like receptor 4 expression through 67 kDa laminin receptor and inhibiting nuclear factor-kappa B pathways [110].Although as a standard care for patients with IBD, sulfasalazine has severe adverse effects,mainly lack of response, male infertility, and fibrosis that may lead to intestinal resection [111,112].In this scenario, flavan-3-ol monomers,total TPs, or tea consumption might be a promising candidate as a form of complementary and alternative medicine to relieve symptom and improve quality of life for IBD patients.

3.3 Host metabolic protection: systemic effect of TPs

During the past two decades, available observational findings suggest that gut microbiome is involved in sustaining the systemic metabolism homeostasis of the human host and, when dysfunctional,to the pathogenesis of multiple common metabolic disorders, e.g.,obesity, T2DM, hypercholesterolemia and nonalcoholic steatohepatitis among others [113,114].Metagenome-wide association studies found that despite the huge disparity in pathologies of these chronic dysmetabolism, they share certain disease-specific abnormalities in the composition and function of gut microbiota [115-117].The interplay modes between the intestinal microbiome and its host are established primarily by means of bioactive metabolites.These metabolites are literally group of gut microbiota-derived small molecules, which are produced by specific microbes as their intermediate or end metabolic products, such as bile acids (BAs), short-chain fatty acids (SCFAs),branched-chain amino acids (BCAAs), lipopolysaccharides (LPS),and trimethylamine (TMA) [118,119].These bioactive metabolites originally produced in gut might enter the enterohepatic circulation through absorption and finally reach the bloodstream of the host,and exert certain systemic effects.Given above, we assume when gut bacterial dysbiosis occurs, metabolic profiles of microbiota are modified, followed by host dysmetabolism, and then metabolic disorders in host might appear.Recently, TPs-based dietary intervention was reported to show systemic metabolism-protective potential by remodeling gut microecosystem and rebuilding a systemically metabolic homeostasis of the host.

BAs are the dominant hepatic catabolites of cholesterol, which can facilitate the digestion and absorption of lipids.Interestingly,interactions between gut microbiota and BAs are subtle.On one hand, microbes are involved in BAs metabolism, principally converting primary to secondary BAs via a series of deconjugation,dehydroxylation, and isomerization processes in gut [120].On the other hand, BAs reshape gut microbiota profiles, by means of a direct modulation or a receptor-mediated pathway [121-123].Thus, gut bacteria-BA interaction significantly affects cholesterol metabolism, lipid homeostasis and energy harvest of the host.A recent study established a diet induced mice model of weight gain and hyperlipidemia, to investigate the modulatory effect of TPs onin vivometabolism [124].Results displayed that body weight and levels of cholesterol and total triglyceride were significantly lowered by Pu-erh tea treatment.Changes of the blood and hepatic lipids in mice were confirmed in human subjects [124].Mechanistically, it was demonstrated that TBs in Pu-erh tea suppressed the bile salt hydrolase related bacteria and activity, further inhibited farnesoid X receptor(FXR)- fibroblast growth factor (FGF) 15/19 signaling pathway in gut while activated an alternative BA synthetic pathway-hepatic FXR-small heterodimer partner (SHP) signaling, which led to an elevated BA production in the liver and fecal excretion and ultimately,diminished the serum cholesterol level [124].Fecal metabolomics combined with 16S rRNA gene sequencing discovered Pu-erh teainduced microbial composition modification in mice and human showing the similar tendencies at the phylum, class and even genus levels, characterized by declined relative abundances ofLactobacillus,Bacillus,Lactococcus, andStreptococcus[124].Also, decreased proportions of two additional generaEnterococcusandLeuconostocwere detected in mice stool samples after Pu-erh tea infusion.These reduced microbial genera have been documented to be linked to the activity of bile salt hydrolase that are involved in deconjugation of primary BAs to form unconjugated BAs [125].The cholesterol- and lipid-lowering effects of Pu-erh tea have shown certain values in clinic for the treatment of obesity, fatty liver and nonalcoholic fatty liver disease [124,126].In diet-induced obese mice, EGCG administration was also found to deactivate FXR-regulated pathway in the intestine whereas active FXR mediated signaling in the liver, which may be a compensation way of BA synthesis as well [93].Furthermore, EGCG activated Takeda G protein receptor (TGR)-5 based on increased glucagon-like peptide 1 release in enteroendocrine L cells, thereby improved liver function and enhanced glucose tolerance in obese mice [93,127].This metabolism-protective function of EGCG may contribute, at least in part, to its capacity to regulate gut microbes, for instance, promoting the bloom ofA.muciniphila[93].

The anaerobic microbiome fermentation degrades nondigestible carbohydrates to produce SCFAs acetate, butyrate, and propionate in the cecum and colon.SCFAs are representative beneficial metabolites derived from gut bacteria, which contribute to functional epithelial barrier, reduced food intake, increased energy expenditure,and improved glucose homeostasis [128,129].Microbiome-wide association studies have uncovered a general down-regulation of SCFAs in faeces of patients with dysmetabolism, e.g., obesity and T2DM, and improved SCFA levels help alleviate metabolic disorders[130,131].For example, mice treated with a butyrate precursor drug (tributyrin) were protected from diet-induced obesity, hepatic steatosis and insulin resistance [132].Some TPs were reported to raise levels of SCFAs by optimizing gut microbial composition and ultimately, benefit host for their metabolic homeostasis.For example, green tea leaf powder dose-dependently reduced body weight and cholesterol level of high-fat diet fed mice, and improved systemic inflammation [133].Also, increased microbial diversity and SCFAs production were observed after green tea leaf powder administration, as well as altered bacterial composition, including increased relative abundances ofAlloprevotella,Butyrivibrio,Blautia,Oscillibacter, andRuminiclostridium, and decreased proportions ofErysipelatoclostridiumandDesulfovibrio.Among which,AlloprevotellaandButyrivibriobelongs to SCFAs producing bacterial genera [134], which might be responsible for up-regulated SCFA levels during green tea leaf powder treatment.BlautiaandOscillibactermight be involved in improving glucose and lipid homeostasis and inhibiting weight gain [135].ErysipelatoclostridiumandDesulfovibriowere considered as opportunistic pathogens in the gut [136].Therefore, mechanism underlying the modulatory effect of green tea leaf powder on host systemic metabolism is related to its ability to positively reshape gut microbiota as well as associated bacterial metabolites.

Taken together, selected TPs are capable of reshaping gut bacterial composition as well as their metabolites and as a result,may benefit for the metabolic homeostasis of the host.Thus, specific TPs may afford not only local benefits within gut, but some positive systemic effects, for instance, prevention of dyslipidemia and obesity as mentioned above.A double-blind, placebo-controlled clinical study found that regular administration of GTP (600 mg/day) for one year led to a 90% chemoprevention efficacy among 60 males with highgrade prostate intraepithelial neoplasia, and exhibited no any adverse effects [137].A cohort study performed in Japan where 80% of the population consumes green tea and over half of them daily drink 3 or more cups (100 mL/cup), confirmed a negative correlation between green tea intake and cardiovascular disease mortality [138].These evidences propose a TPs-based preventive/therapeutic strategy for a series of metabolic syndromes.

4.Conclusions and perspectives

Knowledge ofin vivokinetics is important for the application of dietary phytochemicals in the prevention/treatment of diseases.After ingestion, TPs undergo intense metabolic processes that may include remove of gallic acid moiety, phase II conjugation, and bacterial catabolism.This review summarizes the intestinal fates, metabolic pathways, and major metabolites of TPs, highlighting the important role of gut microbiota in TPs degradation.Of note, the connection between TPs and gut flora is bidirectional and, more significantly,benefits each other.As a result, TPs may give gut barrier protective effects via crosstalk with gut microbes, and based on the local benefits, might further exert systemic protection for host metabolism,such as lowering blood lipids and inhibiting body weight gain.

The metabolism of TPsin vivois rapid and extensive, which may require a fasting and repeated administration to sustain effective plasma concentrations.However, excessive TPs in blood may in turn reach a toxic level.For example, a single dose administration of EGCG (1 500 mg/kg, i.g.) increased the plasma level of alanine aminotransferase, one important indicator evaluating the liver damage,by 138 folds and lowered the survival by 85% in mice model [139].Hepatic necrosis was also observed after high oral doses of EGCG,which was relevant to the pro-oxidant effects of EGCG, inducing mitochondrial toxicity and free radicals-promoting production in the liver [140].In humans, ingestion of high levels ranging from 10-29 mg/kg/day of the green tea extract was found to induce liver toxicity [141].Nonetheless, regular or moderate consumption of green tea is considered to be safe [142].Anin vivostudy with mice found that pretreatment with dietary EGCG (3.2 mg/g diet, equivalent to 500 mg/kg/day, 2 weeks) significantly mitigated hepatotoxicity induced by the high dose (750 mg/kg, i.g., once daily for 3 days)administration [143].In this context, compared with a high dose intake, low-dose dietary supplement with TPs or tea might be an alternative strategy to exert bioeffects while mitigating and even avoiding the potential toxicity.However, available data regarding TPs applications in clinic are still limited and sporadic.Future researches are warranted to determine the safe dose of TPs as well as tea consumption and protecting actual magnitude of different health uses.

Finally, interactions among TPs, microbiome and host are complicated.Although the understanding of the complexity has gained momentum and become clear gradually with the studies of metabolomics and microbiome-wide association, it still remains a huge challenge.For example, there is great inter-individual variability in microbiota profiles, dietary habit, and host metabolic state, and how to determine the proper dosage to deal with these variations to guarantee the dietary intervention effects.Moreover, there is limited information on underlying action mechanisms of TPs including potential action targets, microbiota/TPs crosstalk, and ultimate health effects.All these puzzles wait to be answered to provide enough evidence to support tea- or its polyphenolics-based application.

Declaration of competing interest

The authors declare no conflict of interest.

Acknowledgements

We would like to acknowledge the financial projects of the National Natural Science Foundation of China (No.81803548),Natural Science Foundation of Tianjin (No.19JCQNJC12400), Hubei Province Technical Innovation Special Project (No.2019ABA100),and Tianjin Science and Technology Support Special Project (No.19YFZCSN00010).