茶儿茶素氧化机理

在制茶发酵过程中,儿茶素可发生苯骈环化反应而形成茶黄素,茶黄素还可转化成茶萘酚醒、脱氢茶黄素和高聚合物;同时,儿茶素可发生二聚合反应产生原花青素类以及双黄烷醇类等物质;儿茶素还可能与茶没食子素、杨梅甙、阿福豆素、维生素C、茶氨酸发生聚合反应,形成加合物。体外自由基清除实验表明,儿茶素具有优良的抗氧化特性。儿茶素在清除自由基时,自身氧化产生双黄烷醇,也可发生A环的断裂而产生羧酸类物质。     

Antiviral Effect of Theaflavins on Tobacco Mosaic Virus

Theaflavins, the oxidized products formed from tea leaf catechins during black tea fermentation, showed an antiviral activity on TMV. From the survey of the interactions of theaflavins with RNA and its related substances, it was presumed that theaflavins disturbed the replication cycle of TMV through binding to TMV-RNA.     

Anxiolytic effects of theaflavins via dopaminergic activation in the frontal cortex

Epidemiological investigations have reported that the habit of drinking tea reduces the risk of developing a mental disorder, including anxiety disorder and depression. Theaflavins, black tea polyphenols, show antibacterial and anti-oxidative effects, but their effects on brain function, especially mental condition, have not been elucidated. The present study demonstrated that theaflavins increased dopamine (DA) turnover in the frontal cortex and showed an anxiolytic effect in mice. Theaflavin consumption increased the time spent by mice in the open arms of an elevated plus maze test. Theaflavin administration increased the levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and the ratios of DOPAC/DA and (DOPAC+homovanillic acids)/DA indicating DA turnover, in the frontal cortex. These results suggest that the consumption of theaflavins induced anxiolytic effects via activation of the dopaminergic system in the frontal cortex, which support the findings of previous epidemiological studies. Theaflavins in black tea may be helpful to reduce anxiety in daily life. (150/150 words).     

Neuroprotective Effects of Theaflavins Against Oxidative Stress-Induced Apoptosis in PC12 Cells

Oxidative stress can induce neuronal apoptosis via the production of superoxide and hydroxyl radicals. This process is as a major pathogenic mechanism in neurodegenerative disorders. In this study, we aimed to clarify whether theaflavins protect PC12 cells from oxidative stress damage induced by H₂O₂. A cell model of PC12 cells undergoing oxidative stress was created by exposing cells to 200 μM H₂O₂ in the presence or absence of varying concentrations of theaflavins (5, 10, and 20 μM). Cell viability was monitored using the MTT assay and Hoechst 33258 staining, showing that 10 μM theaflavins enhanced cell survival following 200 μM H₂O₂ induced toxicity and increased cell viability by approximately 40?%. Additionally, we measured levels of intracellular reactive oxygen species (ROS) and antioxidant enzyme activity. This suggested that the neuroprotective effect of theaflavins against oxidative stress in PC12 cells is derived from suppression of oxidant enzyme activity. Furthermore, Western blot analyses indicated that theaflavins downregulated the ratio of pro-apoptosis/anti-apoptosis proteins Bax/Bcl-2. Theaflavins also downregulated the expression of caspase-3 compared with a H₂O₂-treated group that had not been treated with theaflavins. Interestingly, this is the first study to report that the four main components of theaflavins found in black tea can protect neural cells (PC12) from apoptosis induced by H₂O₂. These findings provide the foundations for a new field of using theaflavins or its source, black tea, in the treatment of neurodegenerative diseases caused by oxidative stress.     

Peroxidase-mediated oxidation of catechins

Theaflavins and thearubigins are major pigments in black tea, and they are produced by oxidation and polymerization of flavan-3-ols (catechins) during tea fermentation. In the course of studies on the mechanism of black tea polyphenol formation, we have systematically studied peroxidase-mediated oxidation of tea catechins. Individual catechins, appropriate pairs of catechins, and the combination of selected catechins and theaflavins have been used as substrates of peroxidase. Different dimers, trimers and a tetramer are formed. This work has led to the characterization of some of the new trimers in black tea by LC/ESI-MS/MS. The proposed mechanisms for the peroxidase-mediated reaction are discussed.     

New dibenzotropolone derivatives characterized from black tea using LC/MS/MS

Theaflavins and thearubigins are major pigments in black tea, and it is generally accepted that they are produced by oxidation of flavan-3-ols (catechins) during tea fermentation. In the course of studies on the oxidation mechanism of tea polyphenols, especially the formation of thearubigins, a method combining the enzymatic synthesis and LC/ESI-MS/MS analysis was developed to search for new higher molecular weight polymers from black tea. Three new dibenzotropolones, theadibenzotropolone A, B, and C, together with one new tribenzotropolone, theatribenzotropolone A, were formed by the reaction of theaflavins and tea catechins with horseradish peroxidase in the presence of H(2)O(2). The structures of these new benzotropolone derivatives were elucidated on the basis of MS and 2D NMR spectroscopic analyses. The existence of these compounds in black tea was characterized by LC/ESI-MS/MS. Theadibenzotropolone A and B were the first benzotropolone-type trimers of catechins found in the black tea extract. The observation that galloyl ester groups of theaflavins can be oxidized to form di- or tri-benzotropolone skeletons strongly implied that this type of oxidation is an important pathway to extend the molecular size of thearubigins.     

Effect of oral theaflavin administration on body weight,fat, and muscle in healthy subjects: a randomized pilot study

Theaflavins are reddish-colored polyphenols in black tea. To test the efficacy of theaflavin administration on body fat and muscle, we performed a randomized, double-blind, placebo-controlled study and investigated the effect of theaflavins administration on the body composition using of healthy subjects. In this study, 30 male and female Japanese were enrolled and participants were randomly allocated to receive placebo, theaflavin (50 or 100 mg/day), or catechin (400 mg/ml) for 10 weeks. The effects were evaluated using body weight, body fat percentage, subcutaneous fat percentage, and skeletal muscle percentage. Theaflavin administration significantly improved body fat percentage, subcutaneous fat percentage, and skeletal muscle percentage when compared to with the placebo. In contrast, there was no significant difference in all measured outcomes between the catechin and the placebo groups. The results indicate that oral administration of theaflavin had a beneficial effect on body fat and muscle in healthy individuals.     

A Single Oral Administration of Theaflavins Increases Energy Expenditure and the Expression of Metabolic Genes

Theaflavins are polyphenols found in black tea, whose physiological activities are not well understood. This study on mice evaluated the influence of a single oral administration of theaflavins on energy metabolism by monitoring the initial metabolic changess in skeletal muscle and brown adipose tissue (BAT). Oxygen consumption (VO₂) and energy expenditure (EE) were increased significantly in mice treated with theaflavin rich fraction (TF) compared with the group administered vehicle alone. There was no difference in locomotor activity. Fasting mice were euthanized under anesthesia before and 2 and 5, 20-hr after treatment with TF or vehicle. The mRNA levels of uncoupling protein-1 (UCP-1) and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) in BAT were increased significantly 2-hr after administration ofTF. The levels of UCP-3 and PGC-1α in the gastrocnemius muscle were increased significantly 2 and 5-hr after administration of TF. The concentration of phosphorylated AMP-activated protein kinase (AMPK) 1α was also increased significantly in the gastrocnemius 2 and 5-hr after treatment with TF. These results indicate that TF significantly enhances systemic energy expenditure, as evidenced by an increase in expression of metabolic genes.     

The Microbiota Is Essential for the Generation of Black Tea Theaflavins-Derived Metabolites

Background

Theaflavins including theaflavin (TF), theaflavin-3-gallate (TF3G), theaflavin-3′-gallate (TF3′G), and theaflavin-3,3′-digallate (TFDG), are the most important bioactive polyphenols in black tea. Because of their poor systemic bioavailability, it is still unclear how these compounds can exert their biological functions. The objective of this study is to identify the microbial metabolites of theaflavins in mice and in humans.

Methods and Findings

In the present study, we gavaged specific pathogen free (SPF) mice and germ free (GF) mice with 200 mg/kg TFDG and identified TF, TF3G, TF3′G, and gallic acid as the major fecal metabolites of TFDG in SPF mice. These metabolites were absent in TFDG- gavaged GF mice. The microbial bioconversion of TFDG, TF3G, and TF3′G was also investigated in vitro using fecal slurries collected from three healthy human subjects. Our results indicate that TFDG is metabolized to TF, TF3G, TF3′G, gallic acid, and pyrogallol by human microbiota. Moreover, both TF3G and TF3′G are metabolized to TF, gallic acid, and pyrogallol by human microbiota. Importantly, we observed interindividual differences on the metabolism rate of gallic acid to pyrogallol among the three human subjects. In addition, we demonstrated that Lactobacillus plantarum 299v and Bacillus subtilis have the capacity to metabolize TFDG.

Conclusions

The microbiota is important for the metabolism of theaflavins in both mice and humans. The in vivo functional impact of microbiota-generated theaflavins-derived metabolites is worthwhile of further study.     

Theaflavins attenuate hepatic lipid accumulation through activating AMPK in human HepG2 cells

Black tea is one of the world's most popular beverages, and its health-promoting effects have been intensively investigated. The antiobesity and hypolipidemic effects of black tea have attracted increasing interest, but the mechanisms underlying these phenomena remain unclear. In the present study, the black tea major component theaflavins were assessed for their hepatic lipid-lowering potential when administered in fatty acid overload conditions both in cell culture and in an animal experimental model. We found that theaflavins significantly reduced lipid accumulation, suppressed fatty acid synthesis, and stimulated fatty acid oxidation. Furthermore, theaflavins also inhibited acetyl-coenzyme A carboxylase activities by stimulating AMP-activated protein kinase (AMPK) through the LKB1 and reactive oxygen species pathways. These observations support the idea that AMPK is a critical component of decreased hepatic lipid accumulation by theaflavin treatments. Our results show that theaflavins are bioavailable both in vitro and in vivo and may be active in the prevention of fatty liver and obesity.     

Black tea theaflavins suppress dioxin-induced transformation of the aryl hydrocarbon receptor

Dioxins cause various adverse effects through transformation of aryl hydrocarbon receptor (AhR). In this study, we investigated whether black tea extract and its components, theaflavins, suppress AhR transformation in vitro. First, we confirmed that black tea extract strongly suppressed AhR transformation compared to green and oolong tea, although the catechin contents did not change significantly among the extracts. Then we isolated four theaflavins as active compounds from black tea leaves. They suppressed 1 nM 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced AhR transformation in a dose-dependent manner. The IC(50) values of theaflavin, theaflavin-3-gallate, theaflavin-3'-gallate, and theaflavin-3,3'-digallate (Tfdg) were 4.5, 2.3, 2.2, and 0.7 muM, respectively. The suppressive effect of Tfdg was observed not only by pre-treatment but also by post-treatment. This suggests that theaflavins inhibit the binding of TCDD to the AhR and also the binding of the transformed AhR to the specific DNA-binding site as putative mechanisms.     

Modulation of tea and tea polyphenols on benzo(a)pyrene-induced DNA damage in Chang liver cells

The protective effects of three tea extracts (green tea, GTE; oolong tea, OTE; and black tea, BTE) and five tea polyphenols (epicatechin, EC; epicatechin gallate, ECG; epigallocatechin, EGC; epigallocatechin gallate, EGCG; and theaflavins, THFs) on benzo[a]pyrene (B[a]P)-induced DNA damage in Chang liver cells were evaluated using the comet assay. B[a]P-induced DNA damage in Chang liver cells was significantly (p < 0.05) inhibited by GTE and OTE at a concentration of 10 microg/ml and by BTE at 25 microg/ml. At a concentration of 100 microg/ml, the % tail DNA was reduced from 33% (B[a]P treated only) to 10, 9, 13%, by GTE, OTE and BTE, respectively. EC and ECG did not cause DNA damage in cells according to the results of the comet assay; however, EGC, EGCG and theaflavins caused DNA damage in cells at a concentration of 100 microM. The results indicated that EC and ECG had protective effects against B[a]P-induced DNA damage in cells at a concentration of 10-100 microM. Although EGC, EGCG and the theaflavins caused DNA damage at a high concentration, but they had protective effects against B[a]P-induced DNA damage in cells at a low concentration of 10-50 microM. The results also showed that the DNA damage in cells induced by EGC, EGCG, and the theaflavins was due to the generation of superoxide during incubation with cells at a higher concentration. Therefore, tea catechins and THFs play an important role in enabling tea extracts to inhibit DNA damage in Chang liver cells.     

Optimization of theaflavin biosynthesis from tea polyphenols using an immobilized enzyme system and response surface methodology

Theaflavins were synthesized from tea polyphenols extracted from green tea using an immobilized polyphenol oxidase system. To optimize the production of theaflavins, response surface methodology was applied to determine the effects of five critical variables and their mutual interactions on theaflavin biosynthesis at five levels. A total of 52 individual experiments were performed and a statistical model predicted that the highest theaflavin concentration was 0.766mgml^–1 at optimized conditions. Using these optimal parameters under experimental conditions in three independent replicates, the average value of the biosynthesized theaflavin concentration reached 0.75±0.017mgml^–1 and matched the value predicted by the modelRevisions requested 03 November 2004; Revisions received 7 December 2004     

Theaflavins inhibit the ATP synthase and the respiratory chain without increasing superoxide production

Four dietary polyphenols, theaflavin, theaflavin-3-gallate, theaflavin-3'-gallate and theaflavin-3,3'-digallate (TF3), have been isolated from black tea, and their effects on oxidative phosphorylation and superoxide production in a model system (Escherichia coli) have been examined. The esterified theaflavins were all potent inhibitors of the membrane-bound adenosine triphosphate (ATP) synthase, inhibiting at least 90% of the activity, with IC(50) values in the range of 10-20 μM. ATP-driven proton translocation was inhibited in a similar fashion, as was the purified F(1)-ATPase, indicating that the primary site of inhibition was in the F(1) sector. Computer modeling studies supported this interpretation. All four theaflavins were also inhibitory towards the electron transport chain, whether through complex I (NDH-1) or the alternative NADH dehydrogenase (NDH-2). Inhibition of NDH-1 by TF3 appeared to be competitive with respect to NADH, and this was supported by computer modeling studies. Rates of superoxide production during NADH oxidation by each dehydrogenase were measured. Superoxide production was completely eliminated in the presence of about 15 μM TF3, suggesting that inhibition of the respiratory chain by theaflavins does not contribute to superoxide production.     

Preparative isolation and purification of theaflavins and catechins by high-speed countercurrent chromatography

High-speed countercurrent chromatography (HSCCC) has been applied for the separation of theaflavins and catechins. The HSCCC run was carried out with a two-phase solvent system composed of hexane-ethyl acetate-methanol-water-acetic acid (1:5:1:5:0.25, v/v) by eluting the lower aqueous phase at 2 ml/min at 700 rpm. The results indicated that pure theaflavin, theaflavins-3-gallate, theaflavins-3'-gallate and theaflavin-3,3'-digallate could be obtained from crude theaflavins sample and black tea. The structures of the isolated compounds were positively confirmed by (1)H NMR and (13)C NMR, MS analysis, HPLC data and TLC data. Meanwhile, catechins including epigallocatechin gallate, gallocatechin gallate, epicatechin gallate and epigallocatechin were isolated from the aqueous extract of green tea by using the same solvent system. This study developed a modified method combined with enrichment theaflavins method by using HSCCC for separation of four individual theaflavins, especially for better separation of theaflavins monogallates.     

Theaflavins retard human breast cancer cell migration by inhibiting NF-κB via p53-ROS cross-talk

The present study demonstrates that theaflavins exploit p53 to impede metastasis in human breast cancer cells. Our data suggest that p53-dependent reactive oxygen species (ROS) induce p53-phosphorylation via p38MAPK in a feedback loop to inhibit IκBα-phosphorylation and NF-κB/p65 nuclear translocation, thereby down-regulating the metastatic proteins metalloproteinase (MMP)-2 and MMP-9. When wild-type p53-expressing MCF-7 cells are transfected with p53 short-interfering RNA, or treated with a pharmacological inhibitor of ROS, theaflavins fail to inhibit NF-κB-mediated cell migration. On the other hand, NF-κB over-expression bestows MCF-7 cells with resistance to the anti-migratory effect of theaflavins. These results indicate that inhibition of NF-κB via p53-ROS crosstalk is a pre-requisite for theaflavins to accomplish the anti-migratory effect in breast cancer cells.

Structured summary

MINT-7295816: p53 (uniprotkb:P04637) physically interacts (MI:0915) with IKK beta (uniprotkb:O14920) by anti bait coimmunoprecipitation (MI:0006)     

Analysis of theaflavins in biological fluids using liquid chromatography–electrospray mass spectrometry

A HPLC–MS procedure for the sensitive and specific analysis of the black tea flavonoid theaflavin in human plasma and urine was developed. Levels were measured after enzymatic deconjugation, extraction into ethyl acetate, and separation by HPLC, using tandem mass spectrometry as a detecting system. Two healthy volunteers consumed 700 mg theaflavins, equivalent to about 30 cups of black tea. The maximum concentration detected in blood plasma was 1.0 μg l⁻¹ in a sample collected after 2 h. The concentration in urine also peaked after 2 h at 4.2 μg l⁻¹. Hence, only minute amounts of theaflavins can be detected in plasma and urine samples of healthy volunteers after ingestion.     

Evaluation of the antigenotoxic potential of monomeric and dimeric flavanols, and black tea polyphenols against heterocyclic amine-induced DNA damage in human lymphocytes using the Comet assay

The polyphenolic dimers, epicatechin-4beta-8-catechin (B1), epicatechin-4beta-8-epicatechin (B2), catechin-4beta-8-catechin (B3), catechin-4beta-8-epicatechin (B4), and the gallate ester epicatechin-4beta-8-epicatechin gallate (B'2G) were isolated from grape seeds, and theaflavins and theafulvins from black tea brews. The ability of these naturally-occurring polyphenols to afford protection against the genotoxicity of the heterocyclic amine 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) was compared with that of the monomeric tea flavanols, (+)-catechin (C), (-)-epicatechin (EC), (-)-epicatechin gallate (ECG), (-)-epigallocatechin (EGC) and (-)-epigallocatechin gallate (EGCG). Genotoxic activity was evaluated in human peripheral lymphocytes using the Comet assay. At the concentration range of 1-100 microM, neither the monomeric nor the dimeric flavanols prevented the lymphocyte DNA damage induced by Trp-P-2. In contrast, both of the black tea polyphenols, theafulvins and theaflavins, at a dose range of 0.1-0.5 mg/ml, prevented, in a concentration-dependent manner, the DNA damage elicited by Trp-P-2. Finally, neither the monomeric and dimeric polyphenols (100 microM) nor the theafulvins and theaflavins (0.5mg/ml) caused any DNA damage in the human lymphocytes. These studies illustrate that black tea theafulvins and theaflavins, if absorbed intact, may contribute to the anticarcinogenic potential associated with black tea intake.     

Black tea theaflavins inhibit formation of toxic amyloid-β and α-synuclein fibrils

Causal therapeutic approaches for amyloid diseases such as Alzheimer's and Parkinson's disease targeting toxic amyloid oligomers or fibrils are still emerging. Here, we show that theaflavins (TF1, TF2a, TF2b, and TF3), the main polyphenolic components found in fermented black tea, are potent inhibitors of amyloid-β (Aβ) and α-synuclein (αS) fibrillogenesis. Their mechanism of action was compared to that of two established inhibitors of amyloid formation, (-)-epigallocatechin gallate (EGCG) and congo red (CR). All three compounds reduce the fluorescence of the amyloid indicator dye thioflavin T. Mapping the binding regions of TF3, EGCG, and CR revealed that all three bind to two regions of the Aβ peptide, amino acids 12-23 and 24-36, albeit with different specificities. However, their mechanisms of amyloid inhibition differ. Like EGCG but unlike congo red, theaflavins stimulate the assembly of Aβ and αS into nontoxic, spherical aggregates that are incompetent in seeding amyloid formation and remodel Aβ fibrils into nontoxic aggregates. When compared to EGCG, TF3 was less susceptible to air oxidation and had an increased efficacy under oxidizing conditions. These findings suggest that theaflavins might be used to remove toxic amyloid deposits.