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[Polyphenols and endogenous antioxidant defences: effects on glutathione and glutathione related enzymes].

Among diet antioxidants, polyphenols, naturally occurring in vegetables, fruits and plant-derived beverages such as tea, red wine, and extra virgin olive oil, are the most abundant ones. In vitro cell culture experiments have shown that polyphenols possess antioxidant properties, and it is thought that these activities can contribute to the prevention of several oxidative stress-associated diseases. It has however become clear that the mechanisms of action of polyphenols go beyond their intrinsic reducing capabilities, being able to exert other additional effects that are as yet poorly understood. This article gives an overview of the most recent data on the subject and describe the additional functions that polyphenols can have in biological systems, focusing on their effects on glutathione and its related enzymes. Evidence is provided of a tight connection between exogenous and endogenous antioxidants that appear to act in a coordinated fashion. Experimental data indicate that polyphenols may offer an indirect protection by activating endogenous defense systems. It is reasonable to hypothesize that this is achieved, at least in part, through antioxidant responsive elements (ARE) present in the promoter regions of many of the genes inducible by oxidative and chemical stress. The latest studies strongly suggest that dietary polyphenols can stimulate antioxidant enzyme transcription through ARE.

Animals↗

[Evaluation of two methods to determine the polyphenol content in raw and cooked beans and its effect on protein digestibility].

The polyphenolic compounds present in raw and cooked, and dried, with and without the cooking broth of common white, black and red beans (Phaseolus vulgaris) were measured by the Folin-Denis method for total polyphenols, and by the protein precipitation method of Hagerman-Butler, which measures their biological activity. The polyphenol content was measured during 20 consecutive days on the same sample, using three different extracts of volume from each sample. Statistical analysis of the results by the Folin-Denis method indicated that variability among the three aliquots was different for each bean color. A non-parametric analysis, however, indicated that the average in the three levels of concentration for beans of all colors, was the same. A similar analysis of the results by the Hagerman-Butler method demonstrated that variability and average values for the three aliquots were equal for black and red beans but not for white beans. The coefficient of variation was lower for the higher aliquot of the extract. A significant correlation (r = 0.72, P less than 0.05, n = 60) was found between the two methods for all beans using the larger aliquot of the extract. The correlation was highly significant (r = 0.84, P less than 0.05) when white bean values were eliminated. The polyphenolic content varied with seed color and the thermic process reduced their content, as measured by the two methods. The losses in polyphenolics as measured by the Folin-Denis in the cooked beans dried with broth, varied from 31.4 to 36.3%, and from 39.8 to 51.1% for the cooked bean flour dried without broth. The losses by the Hagerman-Butler method were from 25.0 to 93.5% in the cooked bean flours dried with cooking broth, and from 33.3 to 95.7% when dried without the broth. The higher losses were recorded for red beans. In vivo digestibility for cooked bean flours, dried and without broth, were 73.2, 69.6 and 64.5%, and 71.9, 71.9 and 68.8% for white, red and black beans, respectively. A negative correlation (r = -0.39) and significant (p less than 0.05) was found between polyphenolic content in the diet and in vivo protein digestibility.

Analysis of Variance↗

Polyphenol antioxidants in citrus juices: in vitro and in vivo studies relevant to heart disease.

It is well known that eating fruits and vegetables lowers the risk of chronic diseases such as heart disease and cancer. The question of what is/are the active ingredient(s) is still unresolved. The initial hypothesis was that the antioxidant vitamins were responsible. However, recently the polyphenols have been investigated since they have been found to have beneficial properties such as being strong antioxidants. We measured the polyphenol content of citrus juices by an oxidation-reduction colorimetric method (Folin) using catechin as the standard. The order was tangerine juice > grapefruit juice > orange juice. The antioxidant contribution of ascorbic acid was measured by the difference in Folin reactive content following removal by ascorbate oxidase. Ascorbate contributed 56 to 77% of the antioxidant content of orange juice, 46% of the single tangerine juice measured, and 66 to 77% of grapefruit juices. Polyphenol quality in the juices was analyzed by using the inhibition of lower density lipoprotein oxidation promoted by cupric ion, an in vitro model of heart disease. Quality decreased in the following order: orange juice > grapefruit juice > tangerinejuice. In orange juice polyphenols accounted for 84-85% of antioxidant quality. The pure polyphenol hesperidin, which is common in juices, ascorbic acid, and the citrus juices, were not able to bind with LDL+VLDL and protect it from oxidation. In a hamster model of atherosclerosis, the juices were able to significantly inhibit atherosclerosis and lowered cholesterol and triglycerides. Ascorbic acid alone in the dose provided by the juices was found to have the same effect on atherosclerosis. However, the polyphenols in the citrus

Adult↗

The involvement of polyphenols and peroxidase activities in heavy-metal accumulation by epidermal glands of the waterlily (Nymphaeaceae).

Co-localization of polyphenols and peroxidase activity was demonstrated in epidermal glands of the waterlily (Nymphaea) by histochemistry. Total phenols, tannins and peroxidase activity were determined quantitatively in plant extracts. Polyphenols were partially identified and were found to consist mainly of hydrolyzable tannins, gallic and tannic acid derivatives. Nymphaea polyphenols were shown to chelate Cr, Hg, and Pb in vitro, and Cd-binding by polymerized polyphenols was demonstrated in leaves exposed to Cd in vivo. Both polyphenols and peroxidases were found at very high constitutive levels, which were not induced or altered by external conditions, such as light and heavy-metal stress. It is suggested that the polymerization of polyphenols by peroxidases, enhanced after heavy-metal uptake and detoxification, is responsible for the binding of heavy metals in Nymphaea epidermal glands.

Cadmium↗

Antiangiogenic mechanisms of diet-derived polyphenols.

Accumulating evidence demonstrates that polyphenols in natural products are beneficial against human lethal diseases such as cancer and metastasis. The underlying mechanisms of anti-cancer effects are complex. Recent studies show that several polyphenols, including epigallocatechin-3-gallate (EGCG) in green tea and resveratrol in red wine, inhibit angiogenesis when administrated orally. These polyphenols have direct effects on suppression of angiogenesis in several standard animal angiogenesis models. Because angiogenesis is involved in many diseases such as cancer, diabetic retinopathy and chronic inflammations, the discovery of these polyphenols as angiogenesis inhibitors has shed light on the health beneficial mechanisms of natural products, which are rich in these molecules. At the molecular level, recent studies have provided important information on how these molecules inhibit endothelial cell growth. Perhaps the greatest therapeutic advantage of these small natural molecules over large protein compounds is that they can be administrated orally without causing severe side effects. It is anticipated that more polyphenols in natural products will be discovered as angiogenesis inhibitors and that these natural polyphenols could serve as leading structures in the discovery of more potent, synthetic angiogenesis inhibitors.

Journal Article↗

Antimicrobial effects of green tea polyphenols on thermophilic spore-forming bacteria.

The inhibitory action of tea polyphenols towards the development and growth of bacterial spores was examined. Among the tested Bacillus bacteria, tea polyphenols showed antibacterial effects towards Bacillus stearothermophilus, which is a thermophilic spore-forming bacterium. The heat resistance of B. stearothermophilus spores was reduced by the addition of tea polyphenols. Clostridium thermoaceticum, an anaerobic spore-forming bacterium, also exhibited reduced heat resistance of its spores in the presence of tea polyphenols. (-)-Epigallocatechin gallate, which is the main component of tea polyphenols, showed strong activity against both B. stearothermophilus and C. thermoaceticum. The heat resistance of these bacterial spores was more rapidly decreased by the addition of tea polyphenols at high temperatures.

Journal Article↗

Nitric oxide production and endothelium-dependent vasorelaxation induced by wine polyphenols in rat aorta.

1. The aim of this work was to investigate the mechanism of vasorelaxation induced by red wine polyphenolic compounds (RWPC) and two defined polyphenols contained in wine, leucocyanidol and catechin. The role of the endothelium, especially endothelium-derived nitric oxide (NO), was also investigated. 2. Relaxation produced by polyphenols was studied in rat aortic rings with and without functional endothelium, pre-contracted to the same extent with noradrenaline (0.3 and 0.1 microM, respectively). RWPC and leucocyanidol, but not catechin, produced complete relaxation of vessels with and without endothelium. However, 1000 fold higher concentrations were needed to relax endothelium-denuded rings compared to those with functional endothelium. 3. High concentrations of catechin (in the range of 10(-1) gl-1) only produced partial relaxation (maximum 30%) and had the same potency in rings with and without endothelium. 4. The NO synthase inhibitor, N omega-nitro-L-arginine-methyl-ester (L-NAME, 300 microM) completely abolished the endothelium-dependent but not the endothelium-independent relaxations produced by all of the polyphenolic compounds. 5. In contrast to superoxide dismutase (SOD, 100 u ml-1), neither RWPC nor leucocyanidol affected the concentration-response curve for the NO donor, SIN-1 (3-morpholino-sydnonimine) which also produces superoxide anion (O2-). 6. In aortic rings with endothelium, RWPC (10(-2) gl-1) produced, a 7 fold increase in the basal production of guanosine 3':5'-cyclic monophosphate (cyclic GMP) which was prevented by L-NAME (300 microM). 7. Electron paramagnetic resonance (e.p.r.) spectroscopy studies with Fe(2+)-diethyldithiocarbamate as an NO spin trap demonstrated that RWPC and leucocyanidol increased NO levels in rat thoracic aorta about 2 fold. This NO production was entirely dependent on the presence of the endothelium and was abolished by L-NAME (300 microM). 8. These results show that RWPC and leucocyanidol, but not the structurally closely related polyphenol catechin, induced endothelium-dependent relaxation in the rat aorta. They indicate that this effect results from enhanced synthesis of NO rather than enhanced biological activity of NO or protection against breakdown by O2. It is concluded that some polyphenols, with specific structure, contained in wine possess potent endothelium-dependent vasorelaxing activity.

Animals↗

Green tea polyphenols block endotoxin-induced tumor necrosis factor-production and lethality in a murine model.

Green tea polyphenols are potent antioxidants. They have both anti-cancer and anti-inflammatory effects. However, their mechanisms of actions remain unclear. In inflammation, tumor necrosis factor-alpha(TNFalpha) plays a pivotal role. NF-KB, an oxidative stress -sensitive nuclear transcription factor, controls the expression of many genes including the TNFalpha gene. We postulated that green tea polyphenols regulate TNFalpha gene expression by modulating NF-KB activation through their antioxidant properties. In the macrophage cell line, RAW264.7, (-)epigallocatechin gallate (EGCG), the major green tea polyphenol, decreased lipopolysaccharide (LPS)-induced TNFalpha production in a dose-dependent fashion (50% inhibition at 100 mmol/L). EGCG also inhibited LPS-induced TNFalpha mRNA expression and nuclear NF-KB-binding activity in RAW264.7 cells (30-40% inhibition at 100 mmol/L). Similarly, EGCG inhibited LPS-induced TNFalpha production in elicited mouse peritoneal macrophages. In male BALB/c mice, green tea polyphenols (given by oral gavage 2 h prior to an i.p. injection of 40 mg LPS/kg body wt) decreased LPS-induced TNFalpha production in serum in a dose-responsive fashion. At a dose of 0.5 g green tea polyphenols/kg body wt, serum TNFalpha was reduced by 80% of control. Moreover, 0.5 g green tea polyphenols/kg body wt completely inhibited LPS-induced lethality in male BALB/c mice. We conclude that the anti-inflammatory mechanism of green tea polyphenols is mediated at least in part through down-regulation of TNFalpha gene expression by blocking NF-KB activation. These findings suggest that green tea polyphenols may be effective therapy for a variety of inflammatory processes.

Administration, Oral↗

Reduced progression of atherosclerosis in apolipoprotein E-deficient mice following consumption of red wine, or its polyphenols quercetin or catechin, is associated with reduced susceptibility of LDL to oxidation and aggregation.

The effect of consuming red wine, or its major polyphenol constituents catechin or quercetin, on the development of atherosclerotic lesions, in relation to the susceptibility of plasma LDL to oxidation and to aggregation, was studied in atherosclerotic apolipoprotein E deficient (E degree) mice. Forty E degree mice at the age of 4 weeks were divided into four groups, 10 mice in each group, and were supplemented for up to 6 weeks in their drinking water with placebo (1.1% alcohol); catechin or quercetin (50 micrograms/d per mouse), or red wine (0.5 mL/d per mouse). Consumption of catechin, quercetin, or red wine had no effect on plasma LDL or HDL cholesterol levels. The atherosclerotic lesion area was smaller in the treated mice by 39%, 46%, and 48%, respectively, in comparison with E degree mice that were treated with placebo. In accordance with these findings, cellular uptake of LDL derived after catechin, quercetin, or red wine consumption was found to be reduced by 31%, 40%, and 52%, respectively. These results were associated with reduced susceptibility to oxidation (induced by different modes such as copper ions, free radical generator, or macrophages) of LDL isolated after red wine or quercetin and, to a lesser extent after catechin consumption, in comparison with LDL isolated from the placebo group. Similar results were obtained when LDL was preincubated in vitro with red wine or with the polyphenols prior to its oxidation. Even in the basal oxidative state (not induced oxidation), LDL isolated from E degree mice that consumed catechin, quercetin, or red wine for 2 weeks was found to be less oxidized in comparison with LDL isolated from E degree mice that received placebo, as evidenced by 39%, 48%, and 49% reduced content of LDL-associated lipid peroxides, respectively. This effect could be related to enhanced serum paraoxonase activity in the polyphenol-treated mice. LDL oxidation was previously shown to lead to its aggregation. The present study demonstrated that the susceptibility of LDL to aggregation was reduced in comparison with placebo-treated mice, by 63%, 48%, or 50% by catechin, quercetin, and red wine consumption, respectively, and this effect could be shown also in vitro. The inhibition of LDL oxidation by polyphenols could be related, at least in part, to a direct effect of the polyphenols on the LDL, since both quercetin and catechin were found to bind to the LDL particle via the formation of an ether bond. We thus conclude that dietary consumption by E degree mice of red wine or its polyphenolic flavonoids quercetin and, to a lesser extent, catechin leads to attenuation in the development of the atherosclerotic lesion, and this effect is associated with reduced susceptibility of their LDL to oxidation and aggregation.

Animals↗

Cancer chemoprevention and apoptosis mechanisms induced by dietary polyphenolics.

This review summarises current knowledge on the various molecular chemopreventive or therapeutic mechanisms that may be involved when the administration of flavonoids or polyphenols prevented chemical carcinogenesis in animal models. These mechanisms can be subdivided into the following: 1) the molecular mechanisms involved in preventing carcinogen metabolic activation, 2) the molecular mechanisms for preventing tumour cell proliferation by inactivation or downregulation of prooxidant enzymes or signal transduction enzymes, 3) the molecular cell death mechanisms for the induction of tumour cell death (apoptosis) and the molecular mechanisms for the inhibition of isolated mitochondria functions. Many of the flavonoids and polyphenols found in diets, supplements or herbal medicine were also ranked using "accelerated cytotoxic mechanism screening" by a combinatorial approach utilising isolated rat hepatocytes. A strong correlation of an early collapse of the mitochondrial membrane potential and cell death was found for most of the cytotoxic polyphenols but did not occur with non-toxic polyphenols. This screening could prove useful for eliminating polyphenols that have the potential for adverse health effects and for selecting safe and effective polyphenolic candidates for further development as supplements for preventing cancer or cardiovascular disease. Safety concerns of flavonoid/polyphenol supplements are also reviewed.

Animals↗

Inhibition of activator protein 1 activity and cell growth by purified green tea and black tea polyphenols in H-ras-transformed cells: structure-activity relationship and mechanisms involved.

ras gene mutation, which perpetually turns on the growth signal transduction pathway, occurs frequently in many cancer types. The mouse epidermal JB6 cell line has been transfected with a mutant H-ras gene to mimic carcinogenesis in vitro. These transformed cells (30.7b Ras 12) are able to grow in soft agar, exhibiting anchorage independence and high endogenous activator protein 1 (AP-1) activity, which can be detected by a stable AP-1 luciferase reporter. The present study investigated the ability of different pure green and black tea polyphenols to inhibit this ras signaling pathway. The major green tea polyphenols (catechins), (-)-epigallocatechin-3-gallate (EGCG), (-)-epigallocatechin, (-)-epicatechin-3-gallate, (-)-epicatechin, and their epimers, and black tea polyphenols, theaflavin, theaflavin-3-gallate, theaflavin-3'-gallate, and theaflavin-3,3'-digallate (TFdiG), were compared with respect to their ability to inhibit the growth of 30.7b Ras 12 cells and AP-1 activity. All of the tea polyphenols except (-)-epicatechin showed strong inhibition of cell growth and AP-1 activity. Among the catechins, both the galloyl structure on the B ring and the gallate moiety contributed to the growth inhibition and AP-1 activity; the galloyl structure appeared to have a stronger effect on the inhibitory action than the gallate moiety. The epimers of the catechins showed similar inhibitory effects on AP-1 activity. The addition of catalase to the incubation of the cells with EGCG or TFdiG did not prevent the inhibitory effect on AP-1 activity, suggesting that H2O2 does not play a significant role in the inhibition by tea polyphenols. Both EGCG and TFdiG inhibited the phosphorylation of p44/42 (extracellular signal-regulated kinase 1 and 2) and c-jun without affecting the levels of phosphorylated-c-jun-NH2-terminal kinase. TFdiG inhibited the phosphorylation of p38, but EGCG did not. EGCG lowered the level of c-jun, whereas TFdiG decreased the level of fra-1. These results suggest that tea polyphenols inhibited AP-1 activity and the mitogen-activated protein kinase pathway, which contributed to the growth inhibition; however, different mechanisms may be involved in the inhibition by catechins and theaflavins.

Animals↗

Polyphenols purified from the Brazilian aroeira plant (Schinus terebinthifolius, Raddi) induce apoptotic and autophagic cell death of DU145 cells.

Polyphenols extracted from many plants have shown antiproliferative and antitumor activities in a wide range of carcinogenesis models. The antiproliferative effects of polyphenols purified from the Brazilian aroeira plant (Schinus terebinthifolius, Raddi) were investigated on the androgen-insensitive DU145 human prostatic carcinoma cell line. A F3 fraction purified from leaf extract inhibited the DU145 cell proliferation more than 30-fold compared to the crude extract. By flow cytometric analysis, the polyphenol fraction was demonstrated to induce G0/G1 cell growth arrest and cell apoptosis. This apoptosis was evidenced by caspase 3 stimulation in F3-treated cells as compared to crude extract treated cells. The acid phosphatase activity of lysosomes was strongly activated in the lysosomal fraction of the F3-treated DU145 cells. This lysosomal activation, together with the appearance of autophagic vacuoles, suggests that "type 2 physiological cell death" was also involved in this antiproliferative effect. HPLC analysis of this F3 fraction showed 18 different subfractions. Among these subfractions, F3-3, F3-7 and F3-13 strongly inhibited DU145 cell proliferation in a dose-dependent manner. However, the nature of these polyphenols remains unknown since only one (Isoquercitrin) of the tested pure polyphenols co-migrated with F3-13. Since lysosomotropic drugs are considered as possible regulators of lysosome activity, aroeira polyphenols could target lysosomes of prostatic cancer cells to induce autophagic cell death.

Anacardiaceae↗

Microbial degradation of phloroglucinol and other polyphenolic compounds.

Biodegradation of phloroglucinol (1,3,5-trihydroxybenzene) and other polyphenolic compounds by microbes may occur by aerobic and anaerobic metabolic pathways. Aerobic microbes may initiate the mineralization of phloroglucinol or other polyphenolics by either a reductive pathway, epoxide formation, or a specific hydroxylating mechanism. Cleavage of the various intermediates of phloroglucinol and polyphenolic degradation may occur by intradiol and extradiol mechanisms. The reductive pathway in contrast to other mechanisms utilized by aerobic microbes, seems both cumbersome and energy wasteful. The degradation of lignin and its associated phenolics follows an enzymatic combustion process which resembles a nonspecific enzyme-catalyzed burning. Anaerobic mineralization of phloroglucinol and its associated polyphenolics by several microbes seems to favour the reductive formation of a dihydrophloroglucinol (1,3-dioxo-5-hydroxycyclohexane), which is cleaved by a specific hydrolase. Mineralization of numerous other polyphenolic compounds by anaerobes seems to utilize phloroglucinol as a central metabolite.

Aerobiosis↗

Green tea polyphenol blocks h(2)o(2)-induced interleukin-8 production from human alveolar epithelial cells.

Reactive oxygen species (ROS) play crucial roles in ischemia-reperfusion (IR) injury of lung transplants. Reactive oxygen species may stimulate the production of neutrophil chemotactic factors such as interleukin-8 (IL-8), from alveolar epithelial cells, causing recruitment and activation of neutrophils in the reperfused tissue. Green tea polyphenol has potent anti-oxidative activities and anti-inflammatory effects by decreasing cytokine production. In the present study, we found that green tea polyphenol significantly inhibited IL-8 production induced by hydrogen peroxide (H(2)O(2)) in human lung alveolar epithelial cells (A549 line). It has been shown that mitogen activated protein kinases, such as Jun N-terminal kinase (JNK), p38 and p44/42, could mediate IL-8 production from a variety of cell types. We further investigated the effect of green tea polyphenol on these protein kinases, and demonstrated that H(2)O(2)-induced phosphorylation of JNK and p38 but not p44/42 was inhibited by green tea polyphenol in A549 cells. We speculate that green tea polyphenol may inhibit H(2)O(2)-induced IL-8 production from A549 cells through inactivation of JNK and p38.

Antioxidants↗

Protein quality of pigeonpea (Cajanus cajan (L.) Millsp.) as influenced by seed polyphenols and cooking process.

True protein digestibility (TD), biological value (BV), and net protein utilization (NPU) of low-polyphenol pigeonpea cultivars (Nylon, BDN 2, and ICPL 87067) were significantly higher than those of the high polyphenol cultivars (C 11, ICPL 87, and ICPL 151) when whole-seed samples were compared. Most of the polyphenols (80-90%) were concentrated in the seed-coat. Dhal (decorticated split cotyledons) samples of low (Nylon) and high (C 11) polyphenol cultivars revealed no large differences in TD, BV, and NPU values of these cultivars. This indicated an adverse affect of seed-coat polyphenols on protein quality of pigeonpea whole-seed. The cooking process significantly increased TD in both whole seed and dhal samples. BV of both whole-seed and dhal samples was reduced remarkably by cooking. However, NPU of the cooked whole-seed and dhal samples was significantly higher than in the raw samples. No noticeable differences due to cooking were observed in amino acid composition of whole-seed and dhal samples of these cultivars.

Amino Acids↗

Green and black tea consumption by humans: impact on polyphenol concentrations in feces, blood and urine.

The objective of the study was to determine the effects of green tea, black tea and decaffeinated black tea consumption on urinary and fecal excretions and whole blood and blood serum concentrations of polyphenols. The 56 day study was divided into four randomly arranged experimental periods of 14 days each during which the 10 healthy adult subjects consumed a laboratory controlled, constant, measured diet based on ordinary foods. During separate periods, subjects received no tea, green tea, regular black tea or decaffeinated black tea beverages at the three daily meals. Subjects made complete collections of urine and stools throughout the study and fasting blood samples were drawn at the beginning of the study and at the end of each experimental period. Polyphenols contained in urine, feces, whole blood, blood serums, food and tea were analyzed by the spectrophotometry method of Wah Lau et al. (1989). Green tea consumption resulted in highest intakes in greatest fecal and urinary excretions, highest retentions, and high whole blood concentrations of polyphenols followed by effects of regular black tea, decaffeinated black tea and no tea treatments. These results indicate that polyphenols from tea are at least partly absorbable. Hence, both positive and negative effects of dietary polyphenol may occur internal to the body proper and not only as effects within the intestines.

Analysis of Variance↗

Bioguided extraction of polyphenols from grape marc by using an alternative supercritical-fluid extraction method based on a liquid solvent trap.

The biological activity of polyphenols extracted from grape marc was studied with a view to finding a new use for this winery waste. Polyphenols were extracted by using an alternative supercritical-fluid extraction method based on the use of a liquid trap that allows extracted polyphenols to be retained in a saline buffer, thus avoiding the need for the organic solvent required to elute polyphenols from a solid trap. The major extraction variables influencing the performance of the liquid trap (viz. CO(2) modifier content, flow-rate, extraction time and trap volume) were optimized. The proposed method was applied to the supercritical-fluid extraction extraction of 0.3 g grape marc with CO(2) modified with 3% methanol at 350 bar at 50 degrees C (CO(2) density 0.9 g mL(-1)) for 20 min, using a liquid flow-rate of 0.9 mL min(-1). The polyphenol extracts thus obtained exhibited cytotoxic effects that induced apoptosis in tumour cells.

Apoptosis↗

Proteomic profile of the dentine pellicle modified with plant polyphenols and fluoride.

OBJECTIVES: Despite its protective role, the dentine pellicle has rarely been studied, therefore we aimed to map out the proteomic profile of in vitro dentine pellicles before and after modification. MATERIALS AND METHODS: A total of 135 human dentine specimens were prepared. After initial pellicle formation with 150 µl pooled human saliva (37 °C, 30 min), the dentine specimens were immersed in one of 9 pellicle modification solutions (2 ml/specimen): deionized water (non-modified pellicle), SnCl2/NaF/AmF (commercial solution containing 800 ppm Sn2+ and 500 ppm F-), NaF solution (500 ppm F- ), and six polyphenol solutions (2 mg / ml) with or without 500 ppm F-: blueberry extract (BBE and BBE + F-), green tea extract (GTE and GTE + F-) and grape seed extract (GSE and GSE + F-). After another aliquot of saliva (150 µl, 37 °C, 60 min), the pellicles were harvested with sodium dodecyl sulphate by rubbing with cotton balls, and taken to proteomic analyses by Liquid Chromatography-Tandem Mass Spectrometry after tryptic digestion. RESULTS: A total of 382 proteins were identified in all the proteomic analyses for all groups. Pellicle modification with fluoride, either NaF or SnCl2/NaF/AmF, led to the presence of 12 or 14 exclusive proteins, respectively, whereas modification with the solutions containing polyphenols presented less exclusive proteins (4-6 proteins). The number of exclusive proteins was even lower for when polyphenols and fluoride (GTE + F- and GSE + F-) were used, with lower abundance of proteases. CONCLUSIONS: We conclude that NaF and SnCl2/NaF/AmF significantly modify the proteome of the dentine pellicle. The combination of fluoride with polyphenols further lowers the abundance of proteins and proteases, which explains the positive effect of these solutions on the dentine pellicles. CLINICAL RELEVANCE: Plant extract solutions with fluoride can significantly modify the proteomic structure of the dentine pellicle, which clarifies the mechanism of action of polyphenols on the protection of dentine demineralization.

Humans↗