Penta-O-galloyl-ß-d-glucose inhibits the formation of advanced glycation end-products (AGEs): A mechanistic investigation.

Penta-O-galloyl-ß-d-glucose (PGG) was prepared from tannic acid methanolysis products based on HSCCC, and its protective effects and mechanism on the glucose-induced glycation were investigated for the first time. PGG was confirmed to exhibit strong anti-AGEs effects in bovine serum albumin (BSA)-glucose (Glu) and BSA-methylglyoxal (MGO) glycation systems. It was showed that PGG could inhibit the AGEs formation by blocking glycated intermediates (fructosamine and α-dicarbonyl compounds), eliminating radicals, and chelating metal-ions. In-depth mechanism analysis proved that PGG could prevent BSA from glycation by hindering the accumulation of amyloid fibrils, stabilizing the BSA secondary structures, and binding the partial glycation sites. Furthermore, PGG exhibited a prominent trapping capacities on the reactive intermediate MGO by generating PGG-mono-MGO adduct. This research indicated that PGG could be an effective agent to block Glu/MGO-triggered glycation and offered new insights into PGG as a functional ingredient in food materials for preventing diabetic syndrome.

Change in membrane fluidity induced by polyphenols is highly dependent on the position and number of galloyl groups.

The cell membrane fluidity was very important in adipogenesis and galloyl groups on polyphenolic structures could enhance their antiadipogenic activity. However, the effect of polyphenols on membrane fluidity and the role of galloyl groups in fluidity changes remain unclear. Therefore, the present study chose structurally different polyphenols to compare their effects on the membrane morphology and fluidity of 3T3-L1 preadipocytes, and then the reasons behind the changes of membrane fluidity induced by galloylated polyphenols were explored from structural and molecular insights using liposome model and molecular dynamic simulation technology. Our results indicated that galloylated polyphenols could significantly change 3T3-L1 cell membrane morphology and decrease membrane fluidity, while non-galloylated ones could not. The membrane interference effect of polyphenols was enhanced as the number of galloyl groups increased. Morever, the decrease in membrane fluidity induced by galloylated polyphenols was due to the disturbance of polyphenols on lipid alkyl chains in the cell membrane. Galloylated polyphenols could not only locate in the polar head, but also insert into hydrophobic center of lipid bilayer to interfere with the lipid alkyl chains arrangement, thus decreasing the membrane fluidity and showing strong affinity for the membrane. In addition, differences in position of galloyl groups in polyphenols induced distinct effect on cell membranes interactions, thus affecting the binding manner and bioactivity. The results expanded the understanding on the strong antiadipogenic activity of galloylated polyphenols through the aspect of their effects on cell membrane by both experimental and theoretically simulated ways.

The galloyl moiety enhances inhibitory activity of polyphenols against adipogenic differentiation in 3T3-L1 preadipocytes.

Previous studies have proved that the characteristic galloyl moiety in polyphenols is crucial for their biological activities. However, whether the presence of the galloyl moiety in the structure of polyphenols has a great contribution to their inhibition of adipogenic differentiation is not clear. Therefore, in this study, seven polyphenols with different galloylation degrees were chosen for exploring the contribution of the galloyl group to the lipid-lowering property of polyphenols and its molecular mechanism. Our results showed that the existence of the galloyl moiety in the structure of polyphenols was necessary for their inhibition of adipogenic differentiation, which could help to delay cells from entering the G2/M phase as well as to hinder the MCE process in the early stage of differentiation and the downstream PPARγ and C/EBPα related MAPK signaling pathway, probably via binding to IR and disturbing the α-helix in its conformation. Our finding highlighted that the existence of galloyl groups in polyphenols was crucial for their anti-adipogenic activity, and provided new insights into the mechanism by which galloylated polyphenols suppress adipocyte differentiation.

Inhibitory Effects against Alpha-Amylase of an Enriched Polyphenol Extract from Pericarp of Mangosteen (Garcinia mangostana).

The pericarp of mangosteen, a by-product of the mangosteen, is rich in polyphenols. In this study, an efficient and environmentally friendly method for preparative enrichment of polyphenols from mangosteen pericarp (MPPs) was developed, and the inhibitory effects on starch digestion were also evaluated. It was found that the optimal extract method of MPPs was at a solid to solvent ratio of 1:50 g/mL, pH of 2, and at 80 °C for 2 h. The IC50 of MPPs for α-amylase was 0.28 mg/mL. Based on the fluorescence quenching results, we presumed that MPPs could alter the natural structure of α-amylase, resulting in inhibitory activity on α-amylase. In addition, MPPs significantly reduced the blood glucose peak and AUC of glucose responses in rats after ingestion of the starch solution. Taken together, MPPs may have the potential as a functional supplement for blood glucose control and diabetes prevention.

Lipid rafts as potential mechanistic targets underlying the pleiotropic actions of polyphenols.

Polyphenols have attracted a lot of global attention due to their diverse biological actions against cancer, obesity, and cardiovascular diseases. Although extensive research has been carried out to elucidate the mechanisms of pleiotropic actions of polyphenols, this remains unclear. Lipid rafts are distinct nanodomains enriched in cholesterol and sphingolipids, present in the inner and outer leaflets of cell membranes, forming functional platforms for the regulation of cellular processes and diseases. Recent studies focusing on the interaction between polyphenols and cellular lipid rafts shed new light on the pleiotropic actions of polyphenols. Polyphenols are postulated to interact with lipid rafts in two ways: first, they interfere with the structural integrity of lipid rafts, by disrupting their structure and clustering of the ordered domains; second, they modulate the downstream signaling pathways mediated by lipid rafts, by binding to receptor proteins associated with lipid rafts, such as the 67 kDa laminin receptor (67LR), epidermal growth factor receptor (EGFR), and others. This study aims to elaborate the mechanism of interaction between polyphenols and lipid rafts, and describe pleiotropic preventive effects of polyphenols.

Simultaneous determination of the pharmacokinetics of A-type EGCG and ECG dimers in mice plasma and its metabolites by UPLC-QTOF-MS.

A-type epigallocatechin-3-gallate (EGCG) and epicatechin-3-O-gallate (ECG) dimers have multiply biological activities. In this study, the pharmacokinetics of them were investigated in mice after a single dose intravenous administration, and the metabolites in mice plasma and urine were investigated by ultra-performance liquid chromatography-Quadrupole-time of flight mass spectrometer (UPLC-QTOF-MS). Our results showed that the half-life (t1/2) of A-type EGCG and ECG dimers were 116.37 min and 33.04 min, respectively, and the maximal concentration in plasma was 32.81 µg/mL and 55.59 µg/mL, respectively. It was found that two dimers were firstly experienced by quinone methide (QM) fission to form the EGCG and ECG analogue, and the phase II metabolites were generated subsequently. The main metabolites in plasma and urine were glucuronidation and sulphation derivatives. In addition, small molecule weight of phenolic acids were detected in urine.

Anthocyanins from black peanut skin protect against UV-B induced keratinocyte cell and skin oxidative damage through activating Nrf 2 signaling.

Excessive Ultraviolet (UV) irradiation induces skin damage. In the present study, the potential protective activity of anthocyanins (cyanidin-3-O-sophoroside and cyanidin-3-O-sambubioside) from black peanut against skin damage induced by UV-B was evaluated in vitro and in vivo. Treatment with anthocyanins significantly reversed UV-B induced oxidative damage and following apoptotic death in human HaCaT cells. Nuclear-factor-E2-related factor 2 (Nrf 2) was activated by anthocyanins through Nrf 2 protein stabilization and nuclear translocation, along with the expressions of antioxidant responsive element (ARE)- related genes (HO1, GCLC and NOQ1). Nrf 2 knockdown in HaCaT cells by targeted-shRNA plasmid markedly abolished the protective activity of anthocyanins against UV-B irradiation. Additionally, topical application of anthocyanins (5 mg cm-2) inhibited UV-B induced oxidative stress and cell apoptosis in BALB/c mouse skin tissues. The protective effect of anthocyanins can be explained by the regulation of oxidative-stress and the suppression of cell apoptosis through the activation of Nrf-2 by interaction with the MAPK and NF-κB signaling pathways. Our results suggested that anthocyanins from black peanut skin might be used as a potential photochemo-protective agent against UV-B induced skin damage.

GC-(4→8)-GCG, A Proanthocyanidin Dimer from Camellia ptilophylla, Modulates Obesity and Adipose Tissue Inflammation in High-Fat Diet Induced Obese Mice.

SCOPE: Excessive fat accumulation in adipose tissue leads to obesity and related chronic inflammation. This study aims to examine the effects of gallocatechin -(4→8)-gallocatechin-3-O-gallate (GC-(4→8)-GCG), a main proanthocyanidin dimer from Camellia ptilophylla (Cocoa tea), on adipocyte- and adipose-related inflammation in vivo and in vitro. METHODS AND RESULTS: C57BL/6 mice are fed a high-fat diet (HFD) and GC-(4→8)-GCG (40 or 80 mg kg-1 d-1 ) for 8 weeks. The metabolic profiles, adipose tissue hypertrophy, macrophage infiltration, and inflammatory cytokine production are investigated. Additionally, 3T3-L1 preadipocytes are utilized to investigate the effect of GC-(4→8)-GCG on preadipocyte differentiation and the tumor necrosis factor (TNF)-α-stimulated inflammatory response in vitro. GC-(4→8)-GCG supplementation decreases HFD-induced epididymal white adipose tissue (eWAT) hypertrophy, suppresses proinflammatory cytokine production and macrophage infiltration in eWAT, and improves insulin sensitivity in HFD-induced obese mice. In vitro, GC-(4→8)-GCG shows a strong anti-adipogenic potential in 3T3-L1 preadipocyte by inhibiting the expression of key adipogenic transcription factors and decreasing the production of proinflammatory cytokines by inhibiting the activation of the nuclear factor (NF)-κB, Janus tyrosine kinase/signal transducer and activator of transcription (JAK/STAT3) and mitogen-activated protein kinase (MAPK) signaling pathways. CONCLUSION: GC-(4→8)-GCG can modulate obesity and improve obesity-related insulin resistance by inhibiting preadipocyte differentiation and the related proinflammatory responses.

Shepherd's Purse Polyphenols Exert Its Anti-Inflammatory and Antioxidative Effects Associated with Suppressing MAPK and NF-κB Pathways and Heme Oxygenase-1 Activation.

Shepherd's purse (Capsella bursa-pastoris (L.) Medik.), a wild herb as a traditional herbal medicine, has been proved with multiple healthy benefits. In this study, the chemical constituents of shepherd's purse were identified by UPLC-QTOF-MS/MS. The antioxidative and anti-inflammatory potential of shepherd's purse extract (SPE) were also investigated applying lipopolysaccharide- (LPS-) induced inflammation in RAW 264.7 macrophages and a carrageenan-induced mice paw edema model. Twenty-four chemical compounds were identified mainly including phenolic acids and flavonoids. The data also indicated SPE inhibited the productions of NO, PGE2, TNF-α, and IL-6 stimulated with LPS. In addition, SPE inhibited the increase of reactive oxygen species (ROS) and upregulated the expression of heme oxygenase-1 (HO-1). We further found that SPE inhibited the phosphorylation of P38 MAPK and activation of NF-κB. In vivo mice model also indicated that SPE showed strong antioxidative and anti-inflammatory activity.

Penta-O-galloyl-ß-d-glucose, a hydrolysable tannin from Radix Paeoniae Alba, inhibits adipogenesis and TNF-α-mediated inflammation in 3T3-L1 cells.

Penta-O-galloyl-ß-d-glucose (PGG) was purified and identified from Radix Paeoniae Alba by HSCCC and HPLC/ESI-MS, and its inhibitory effects on adipogenesis and TNF-α-induced inflammation were assessed in 3T3-L1 cell line. The results showed that PGG dose-dependently reduced intracellular lipids accumulation, and this involved decrease the expression levels of major adipogenic markers, PPARγ, C/EBP α, through MAPKs inhibition. This was accompanied by a reduction of lipogenic genes, ACC, FAS, and SCD-1, involved in fatty acid synthesis. Furthermore, PGG also inhibited TNF-α-induced expression of inflammatory cytokines including IL-6 and MCP-1 in the matured 3T3-L1 adipocytes. The inhibitions were likely mediated by blocking the MAPKs and NF-κB activation. These findings highlighted that PGG could serve as a potent therapeutic agent for controlling obesity and obesity-related chronic inflammation.

Inhibitory Effect of Persimmon Tannin on Pancreatic Lipase and the Underlying Mechanism in Vitro.

Pancreatic lipase (PL) is a critical enzyme associated with hyperlipidemia and obesity. A previous study of ours suggested that persimmon tannin (PT) was the main component accounting for the antihyperlipidemic effects of persimmon fruits, but the underlying mechanisms were unclear. In this present study, the inhibitory effect of PT on PL was studied and the possible mechanisms were evaluated by fluorescence spectroscopy, circular dichroism (CD) spectra, isothermal titration calorimetry (ITC), and molecular docking. PT had a high affinity to PL and inhibited the activity of PL with the half maximal inhibitory concertation (IC50) value of 0.44 mg/mL in a noncompetitive way. Furthermore, molecular docking revealed that the hydrogen bonding and π-π stacking was mainly responsible for the interaction. The strong inhibition of PT on PL in the gastrointestinal tract might be one mechanism for its lipid-lowering effect.

Position and orientation of gallated proanthocyanidins in lipid bilayer membranes: influence of polymerization degree and linkage type.

It is well known that the biological activity of gallated proanthocyanidins (PAs) is highly structure-dependent. Polymerization degree (DP) and linkage types affect their biological activity greatly. Positions and orientations of gallated PAs in lipid bilayer reveal their structure-function activity at the molecular level. The present work aimed at determining the locations and orientations of epigallocatechin-3-gallate (EGCG) and its derivatives: A-type and B-type EGCG dimers and trimers in 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC) and 1-palmitoyl-2-oleoyl phosphatidylethanolamine (POPE) lipid bilayer via molecular dynamic (MD) simulations. The results showed that EGCG and its derivatives localized in the lipid bilayer or on the bilayer/water interface. Their penetration depths and orientations depended on both DP and linkage types. The penetration depths decreased with the increase of DP, sequencing to be EGCG > EGCG dimers > EGCG trimers. Spatially stretched A-type PAs could form more hydrogen bonds (H-bonds) with deep oxygen atoms of lipid bilayer and have higher affinity to the lipid bilayer than B-type PAs. Our results will provide an explicit evidence for PAs' distinct biological activities.

Separation and purification of four phenolic compounds from persimmon by high-speed counter-current chromatography.

An efficient method was established by high-speed counter-current chromatography (HSCCC) for preparation of four phenolic compounds from the depolymerization products of persimmon tannin. Using the two solvent systems of n-hexane/ethyl acetate/water (3:17:20, v/v/v) and ethyl acetate/methanol/water (50:1:50, v/v/v), the preparative isolation was successfully performed by a two-step separation. The yields of one run (150mg crude sample) for gallic acid, methyl gallate, and epigallocatechin-3-gallate-(4ß→8, 2ß→O→7)-epigallocatechin-3-gallate dimer (A-type EGCG dimer) were 4.7, 44.2 and 5.9mg, respectively. In addition, 4.6mg epicatechin-3-gallate-(4ß→8, 2ß→O→7)-epicatechin-3-gallate dimer (A-type ECG dimer) was obtained by further preparative high-performance liquid chromatography (prep-HPLC). The purities of these compounds were all above 95.0% and their structures were identified by HPLC/ESI-MS. We found that HSCCC had definite advantages for the preparation of dimeric procyanidins compared with previous methods. Furthermore, it was shown that the four phenolic compounds possessed greater antioxidant activities than Trolox.

A-type ECG and EGCG dimers inhibit 3T3-L1 differentiation by binding to cholesterol in lipid rafts.

The present study aimed to explore the underlying mechanisms of epicatechin-3-gallate-(4ß→8, 2ß→O→7)-epicatechin-3-gallate (A-type ECG dimer) and epigallocatechin-3-gallate-(4ß→8, 2ß→O→7)-epigallocatechin-3-gallate (A-type EGCG dimer) involved in their strong inhibitory effects on 3T3-L1 preadipocytes differentiation. In the synthetic "lipid raft-like" liposome, A-type ECG and EGCG dimers incorporated into the liposome with high affinity and decreased the fluidity of the liposome. In 3T3-L1 preadipocytes, A-type ECG and EGCG dimers possibly bonded to lipid rafts cholesterol and disrupted the integrity of lipid rafts, thus exerting their notable inhibitory effects on 3T3-L1 preadipocytes differentiation by suppressing mitotic clonal expansion process and mRNA levels of PPARγ, C/EBPα and SREBP1C. A highly positive correlation between the cholesterol binding capacity of the two dimers and their inhibitory effect on 3T3-L1 preadipocytes differentiation (R2=0.9328) was observed. Molecular dynamics simulation further verified that A-type ECG and EGCG dimers could bond to cholesterol via hydrogen bonding. The results of this study suggested that the disruption of A-type ECG and EGCG dimers on membrane lipid rafts by targeting cholesterol in the lipid rafts was involved in the underlying mechanisms of their strong inhibitory effects on 3T3-L1 preadipocytes differentiation. This broadens the understanding of the molecular mechanisms of polyphenols on modulating and controlling of metabolic dysregulation, particularly adipocyte differentiation, which is a significant risk factor associated with the development of cardiovascular disease.

Comparison of disaggregative effect of A-type EGCG dimer and EGCG monomer on the preformed bovine insulin amyloid fibrils.

In the present study, the disruptive effects of epigallocatechin-3-gallate (EGCG) and A-type dimeric epigallocatechin-3-gallate (A-type EGCG dimer) on the preformed bovine insulin amyloid fibrils were studied by several biophysical methods including thioflavin-T (ThT) fluorescence assay, 1-anilinonaphthalene-8-sulfonic (ANS) fluorescence assay, Congo red (CR) binding assay, dynamic light scattering (DLS), transmission electron microscopy (TEM), Gel electrophoresis (SDS-PAGE) and Bradford assay. Our results demonstrated that A-type EGCG dimer showed significantly more potential disaggregative effects on the bovine insulin amyloid fibrils than EGCG. A-type EGCG dimer could not only dramatically promote the disaggregation of the preformed bovine insulin amyloid fibrils, but also restructure the amyloid fibrils into amorphous aggregates. While, EGCG could only shorten and thin the fibrils, but induce no small amorphous aggregates. Our present results provided additional evidence for the more potent disaggregation effects of dimeric polyphenols than monomeric polyphenols and suggested that A-type EGCG dimer seems to have potential application as an excellent anti-amyloidogenic agent.

Both non-covalent and covalent interactions were involved in the mechanism of detoxifying effects of persimmon tannin on Chinese cobra PLA2.

Persimmon tannin (PT) has been shown to inhibit snake venom activities and toxicities both in vitro and in vivo. To clarify the detoxifying mechanism of PT on snake venom, the interaction of characteristic structural elements of PT (EGCG, ECG, EGCG dimer and ECG dimer) and Chinese cobra phospholipase A2 (PLA2) was studied. The results revealed that except non-covalent bonds like hydrogen bonds, hydrophobic bonds and iron bonds were formed between PT and PLA2, covalent interaction was also occurred. PT could bind with the key active residues of PLA2, such as lysine, histidine, tryptophan and tyrosine, restraining their activity and disturbing the structure of PLA2, thus showing detoxifying effects on snake venom.

Molecular Insight into Affinities of Gallated and Nongallated Proanthocyanidins Dimers to Lipid Bilayers.

Experimental studies have proved the beneficial effects of proanthocyanidins (Pas) relating to interaction with the cell membrane. But the detailed mechanisms and structure-function relationship was unclear. In present study, molecular dynamics (MD) simulations were used to study the interactions of four PA dimers with a lipid bilayer composed of 1:1 mixed 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC) and 1-palmitoyl-2-oleoyl phosphatidylethanolamine (POPE). The results showed that the gallated PA dimers had much higher affinities to the bilayer with lower binding free energies compared with nongallated PA dimers. The gallated PA dimers penetrated deeper into the bilayer and formed more hydrogen bonds (H-bonds) with bilayer oxygen atoms, especially the deeper oxygen atoms of the lipids simultaneously, thus inducing stronger lateral expansion of the membrane and lipid tails disorder. The present results provided molecular insights into the interactions between PA dimers and bio-membranes and agreed with our experimental results well. These molecular interactions helped to elucidate the structure-function relationship of the PA dimers and provided a foundation for a better understanding of the underlying mechanisms of the bioactivities of PA oligomers.

Structure-Dependent Membrane-Perturbing Potency of Four Proanthocyanidin Dimers on 3T3-L1 Preadipocytes.

Proanthocyanidins (PAs) have been widely recognized for their broad spectrum of beneficial health effects, which are highly structure-dependent. It was found that PA dimers epicatechin-3-gallate-(4ß→8,2ß→O→7)-epicatechin-3-gallate (A-type ECG dimer) and epigallocatechin-3-gallate-(4ß→,2ß→O→7)-epigallocatechin-3-gallate (A-type EGCG dimer) inhibit the differentiation of 3T3-L1 cells significantly, whereas epicatechin-(4ß→8,2ß→O→7)-epicatechin (A-type EC dimer) and epicatechin-(4ß→8)-epicatechin (B-type EC dimer) showed little effect in previous work. However, the underlying mechanisms are unclear. To test whether bilayer perturbation may underlie this diversity of actions, we examined the bilayer-modifying effects of the four dimers in both 3T3-L1 cell and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine liposome models by using scanning electron microscopy, fluorescent spectroscopy, differential scanning calorimetry, and molecular dynamics methods. Our results revealed that A-type ECG and EGCG dimers had a high affinity for the lipid bilayer and could form simultaneous hydrogen bonds (H-bond) with both the surface oxygen acceptors and the deeper inside lipid oxygen atoms. However, A-type and B-type EC dimers contacted only the surface oxygen atoms with limited and significantly fewer H-bonds. A-type ECG and EGCG dimers notably distorted the membrane morphology of 3T3-L1 cells. In the present study, we found there was a high positive correlation between the membrane-disturbing abilities of the four dimers and their 3T3-L1 cell differentiation inhibitory effects as previously reported. This indicated that the strong 3T3-L1 cell differentiation inhibitory effect of A-type ECG and EGCG dimers might be due to their strong bilayer-perturbing potency.

A-type dimeric epigallocatechin-3-gallate (EGCG) is a more potent inhibitor against the formation of insulin amyloid fibril than EGCG monomer.

Because fibrillary protein aggregates is regarded to be closely associated with many diseases such as Alzheimer's disease, diabetes, and Parkinson's disease, growing interest and researches have been focused on finding potential fibrillation inhibitors. In the present study, the inhibitory effects of epigallocatechin-3-gallate (EGCG) and A-type dimeric epigallocatechin-3-gallate (A-type EGCG dimer) on the formation of insulin fibrillation were compared by multi-dimensional approaches including thioflavin-T (ThT) fluorescence assay, 1-anilinonaphthalene-8-sulfonic (ANS) fluorescence assay, dynamic light scattering (DLS), transmission electron microscopy (TEM), Fourier transform infrared (FTIR) spectroscopy and circular dichroism (CD) spectroscopy. Our results confirmed that A-type EGCG dimer is a more potent inhibitor against the formation of bovine insulin amyloid fibril than EGCG. In addition, A-type EGCG dimer could not only inhibit insulin amyloid fibril formation, but also change the aggregation pathway and induce bovine insulin into amorphous aggregates. The results of the present study may provide a new guide on finding novel anti-amyloidogenic agents.

Efficient wastewater treatment by membranes through constructing tunable antifouling membrane surfaces.

In the present study, a facile in situ approach for constructing tunable amphiphilic or hydrophilic antifouling membrane surfaces was demonstrated by exquisitely manipulating the microphase separation and surface segregation behavior of the tailor-made ternary amphiphilic block copolymers during the commonly utilized wet phase inversion membrane-formation process. Under dead-end filtration for oily wastewater treatment, the membrane with amphiphilic surface exhibited over 99.5% retention ratio of chemical oxygen demand (COD) without appreciable membrane fouling: the water permeation flux was slightly decreased during operation (total flux decline was 6.8%) and almost completely recovered to the initial value (flux recovery ratio was more than 99.0%) after simple hydraulic washing. While for the proteins-containing wastewater treatment, the membrane with hydrophilic surface exhibited about 52.6% COD retention ratio and superior antifouling performance: only 17.0% total flux decline and also more than 99.0% flux recovery ratio. Hopefully, the present approach can be developed as a competitive platform technology for the preparation of robust and versatile antifouling membrane, leading to the high process efficiency of wastewater treatments.