Metabolic Fate and Cardiometabolic Effects of Phenolic Compounds from Red-Fleshed Apple in Hypercholesterolemic Rats: A Comparative Study with Common White-Fleshed Apple. The AppleCOR Study.

The present study aims to investigate the metabolic fate and the cardiometabolic effects of phenolic compounds provided by a red-fleshed apple variety biofortified in anthocyanins (ACN). Wistar rats are fed with high-fat diet (HFD) to induce hypercholesterolemia and supplemented with red-fleshed apple (HFD+R), white-fleshed apple (HFD+W), or an ACN-rich infusion from aronia fruit (HFD+A) providing matched content and profile of ACN. Plasma biochemical parameters, histological analysis, and phenol biological metabolites are determined. Plasma, urine, and feces show a significant increase of ACN metabolites after HFD+R and HFD+A, while flavan-3-ols are significantly increased after HFD+W and dihydrochalcones derivatives increased after both apples supplementation. A cardioprotective effect is observed after both apples and aronia infusion supplementation in the reduction of aortic thickness. The kidney function is improved after all supplementations and a decrease in insulin plasma concentration after both apples supplementation (HFD+R and HFD+W) is also observed. The findings support that ACN without apple matrix can induce cardioprotective effects. ACN or flavan-3-ols, together with dihydrochalcones, compose a phenolic phytocomplex in red- and white-fleshed apples, respectively, which can act synergistically in the attenuation of cardiovascular outcomes in hypercholesterolemic rats.

Thermal and non-thermal processing of red-fleshed apple: how are (poly)phenol composition and bioavailability affected?

The present study evaluated the impact of different thermal (infrared-drying, hot air-drying and purée pasteurization) and non-thermal (freeze-drying) processing technologies on red-fleshed apple (poly)phenolic compounds. We further investigated the processing effect on the (poly)phenol bioavailability in a crossover postprandial study where three subjects consumed three apple products (freeze-dried snack, hot air-dried snack and pasteurized purée). (Poly)phenolic compounds present in the apple products and their biological metabolites in urine were analyzed using liquid chromatography coupled to mass spectrometry (UPLC-MS/MS). When comparing different processes, infrared-drying caused important losses in most of the apple (poly)phenolics, while hot air-drying and purée pasteurization maintained approximately 83% and 65% of total (poly)phenols compared with the freeze-dried snack, respectively. Anthocyanins in particular were degraded to a higher extent, and hot air-dried apple and pasteurized purée maintained respectively 26% and 9% compared with freeze-dried apple snack. The acute intake showed that pasteurized purée exhibited the highest (poly)phenol bioavailability, followed by hot air-drying and freeze-dried snack, highlighting the impact of processing on (poly)phenols absorption. In conclusion, for obtaining affordable new red-fleshed apple products with enhanced (poly)phenol bioavailability, purée pasteurization and hot air-drying represent viable techniques. However, to obtain a red-fleshed apple snack with high anthocyanin content, freeze-drying is the technique that best preserves them.

Consumption evaluation of one apple flesh a day in the initial phases prior to adenoma/adenocarcinoma in an azoxymethane rat colon carcinogenesis model.

Colorectal cancer (CRC) is the fourth cancer with the most new cases reported in 2018 worldwide. Consumption of fruit and vegetables is a protective factor against the risk of CRC. Beyond this, flavonoids could orchestrate these healthy effects. Apart from containing the typical apple flavonoids, red-fleshed apples also contain anthocyanins, mainly cyanidin-3-O-galactoside (Cy3Gal). Through an azoxymethane rat carcinogenesis model, a study was carried out in order to assess the possible protective effects of apple polyphenols, with special attention to anthocyanins. In addition, apart from negative and positive controls, a group with chemotherapy with 5-fluorouracil (5FU) was included to compare their performance against the output collected from the animal treatments with white-fleshed apple (WF), red-fleshed apple (RF) and Cy3Gal (AE). Although the 5FU group presented the best performance towards aberrant crypt foci (ACF) inhibition (70.1%), rats fed with white-fleshed apples ('Golden Smoothee') were able to achieve 41.3% ACF inhibition, while none of the challenged treatments (WF, RF and AE) suffered mucin depletion in their colonocytes. Expression changes of 17 genes related to CRC were assessed. In detail, the ACF inhibition phenotype detected in 5FU and WF groups could be explained through the expression changes detected in the apoptosis-related genes of Aurka, p53 and Cox2. Moreover, in the apple consumption groups (WF and RF), a reduced protein expression of matrix metalloproteinases with gelatinase activity (MMP-2 and 9) was detected. Overall, our study suggests an effect of apple polyphenols and apple anthocyanin Cy3Gal against colon carcinogenesis, retarding/diminishing the appearance of the precancerous markers studied.

Anthocyanin Tissue Bioavailability in Animals: Possible Implications for Human Health. A Systematic Review.

Anthocyanins (ACNs) are promising health-enhancing phenolic compounds. We focus on ACN animal tissue bioavailability to provide an evidentiary link between tissue ACNs and their associated health properties. We performed a systematic review of electronic libraries; 279 results were retrieved, and 13 publications met inclusion criteria. Extracted information included animal model employed, administration route, doses, analysis method, and ACN concentration values in tissues. Total ACN concentrations were detected in mice kidney (2.17 × 105 pmol/g), liver (1.73 × 105 pmol/g), heart (3.6 × 103 pmol/g), and lung (1.16 × 105 pmol/g); and in pig brain (6.08 × 103 pmol/g). ACNs showed a predominance of parent ACNs in long-term experiments versus an ACN metabolite predominance in short-term experiments. ACNs detected in animal tissues, such as cyanidin-3-glucoside, suggest it may have an important role in human health. This information could be useful to determine proper ACN-intake biomarkers in biological samples in futures studies.

Seasonal Variability of the Phytochemical Composition of New Red-Fleshed Apple Varieties Compared with Traditional and New White-Fleshed Varieties.

The main objective of this study was to evaluate the impact of the season on the apple phytochemical composition (phenolic compounds, triterpenes, and organic and ascorbic acids). For this proposal, four red-fleshed and five white-fleshed apple varieties from two consecutive seasons (2015 and 2016) were studied. A significant interaction with the season in some compounds was observed. The total phenolic content in the apple flesh from 2015 was higher than that from 2016 probably related with the lower rainfall during the harvest period in 2015 that could have favored hydric stress in the apple trees. The impact of the season on the apple skin was different. The 2016 season was characterized by higher maximum and minimum temperatures resulting in a higher content of flavonols, triterpenes, and organic acids. Anthocyanin concentration in both the flesh and skin of the red-fleshed apples showed no clear relationship to the season, and each variety showed an individual pattern.

Bioavailability of the ferulic acid-derived phenolic compounds of a rice bran enzymatic extract and their activity against superoxide production.

Rice bran is an exceptional source of such antioxidant molecules as γ-oryzanol and ferulic acid, but their bioavailability and metabolism within this matrix remain unknown. The aims of this work were to describe the oral bioavailability and metabolic pathways of the ferulic acid-derived phenolic compounds contained in a rice bran enzymatic extract (RBEE), and to determine its effect on NADPH oxidase activity. Wistar rats were administered with RBEE and sacrificed at different times over a period of 24 h to obtain plasma. An additional group was used for collection of urine and faeces over a period of 48 h. The phenolic metabolites were determined by ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS), and plasma pharmacokinetic parameters were calculated. In parallel, aortic rings were incubated in the plasma of rats sacrificed 30 min after RBEE gavage, or in the presence of RBEE, ferulic acid or γ-oryzanol. Endothelin-1-induced superoxide production was recorded by lucigenin-enhanced luminescence. Twenty-five ferulic acid metabolites showing biphasic behaviour were found in the plasma, most of which were found in the urine as well, while in the faeces, colonic metabolism led to simpler phenolic compounds. Superoxide production was abrogated by phenolic compound-enriched plasma and by RBEE and ferulic acid, thus showing the biological potential of RBEE as a nutraceutical ingredient.

Phytochemical Profiles of New Red-Fleshed Apple Varieties Compared with Traditional and New White-Fleshed Varieties.

This study is an exhaustive chemical characterization of the phenolic compounds, triterpenes, and organic and ascorbic acids in red-fleshed apple varieties obtained by different breeding programs and using five traditional and new white-fleshed apple cultivars as reference. To carry out these analyses, solid-liquid extraction (SLE) and ultraperformance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) were used. The results showed that the red-fleshed apples contained, in either the flesh or peel, higher amounts of phenolic acids (chlorogenic acid), anthocyanins (cyanidin-3-O-galactoside), dihydrochalcones (phloretin xylosyl glucoside), and organic acids (malic acid) but a lower amount of flavan-3-ols than the white-fleshed apples. These quantitative differences could be related to an up-regulation of anthocyanins, dihydrochalcones, and malic acid and a down-regulation of flavan-3-ols (anthocyanin precursors) in both the flesh and peel of the red-fleshed apple varieties. The reported results should be considered preliminary because the complete phytochemical characterization of the red-fleshed apple cultivars will be extended to consecutive harvest seasons.

Exploring the Colonic Metabolism of Grape and Strawberry Anthocyanins and Their in Vitro Apoptotic Effects in HT-29 Colon Cancer Cells.

Beneficial properties attributed to the intake of fruit and red wine have been associated with the presence of significant amounts of anthocyanins. However, their low absorption and consequent accumulation in the gut have generated the suspicion that colonic metabolites of anthocyanins are probably involved in these protective effects. Grape pomace and strawberry extracts, rich in malvidin- and pelargonidin-glucoside, respectively, were fermented in vitro using human feces as microbial inoculum. After 8 h of anaerobic incubation, the anthocyanins were almost completely degraded, whereas their microbial metabolite concentrations were highest at 24 h. Syringic acid and tyrosol were the main metabolites of grape and strawberry extracts, respectively. On the basis of the metabolites detected, metabolic pathways of malvidin- and pelargonidin-glucosides were proposed. Anthocyanin-rich grape and strawberry extracts and their generated metabolites such as hydroxyphenylacetic acid showed apoptotic effects in HT-29 colon cancer cells and may suggest their possible contribution as anticarcinogenic agents.

Understanding of human metabolic pathways of different sub-classes of phenols from Arbutus unedo fruit after an acute intake.

Arbutus unedo is a small Mediterranean fruit, commonly named strawberry tree, which is a rich source of different sub-classes of phenolic compounds, the more representative being the gallic acid derivatives, including its mono and oligomeric forms esterified with quinic and shikimic acids. In addition, galloyl derivatives, particularly gallotannins, described in A. unedo, are part of a very selective phenolic group, present in a reduced number of plant-products. The aim of the present study is to provide a better understanding of human metabolic pathways of different sub-classes of phenols from the A. unedo fruit after an acute intake by healthy adults. Therefore, the A. unedo phenolic metabolites were studied in whole blood samples (0 to 24 h), urine (24 h) and feces (12 and 24 h). Special focus was placed on the application of dried blood spot (DBS) cards for the sample collection and for the analysis of phenolic metabolites in whole blood samples. The results of the blood analysis revealed two peaks for the maximum concentrations of the main phenolic metabolites. Furthermore, it is appropriate to highlight the application of DBS cards as an efficient and accurate way to collect blood samples in post-prandial bioavailability studies. The analysis of urine (24 h) gave a wide range of phenolic metabolites showing the extensive metabolism that A. unedo phenolic compounds underwent in the human body. The results of the study provide a relevant contribution to the understanding of the in vivo human bioavailability of phenolic compounds, especially galloyl derivatives, a singular phenolic sub-group present in the A. unedo fruit.

Stability and metabolism of Arbutus unedo bioactive compounds (phenolics and antioxidants) under in vitro digestion and colonic fermentation.

The natural antioxidants of Arbutus unedo highlight the importance of this fruit as natural source of bioactive compounds. In the present study, to evaluate the stability of phenolic compounds, ascorbic acid and fat-soluble antioxidants (α-tocopherol, ß-carotene and lutein), in vitro gastrointestinal digestion was applied to A. unedo fruit. After that, the non-absorbable fraction was anaerobically incubated with human faeces and the metabolic pathway for gallotannins, ellagitannins, flavan-3-ols and anthocyanins from A. unedo fruit was proposed. The results showed that the presence of pectin from the fruit hampered the solubilization of the phenolic compounds (with exception of gallic and ellagic acids) and fat-soluble vitamins during gastric digestion. Degradation of pectin-gel during the duodenal digestion favored the release of the phenolic compounds and fat-soluble antioxidants to the media. The catabolic activity of human microbiota led to the generation of a wide range of simple phenols, such as p-hydroxybenzoic acid and catechol, derived from the catabolism of gallotannins, ellagitannins, flavan-3-ols and anthocyanins.

Study of the catabolism of thyme phenols combining in vitro fermentation and human intervention.

The gut metabolism of four thyme phenolics (monoterpenes thymol and carvacrol, rosmarinic acid, and eriodictyol) was evaluated in vitro. After the in vitro transformations of the individual phenols had been studied, the presence of their microbial metabolites was investigated in human feces collected before and after a sustained intake (3 weeks) of 25 mL/day of a thyme phenol-enriched olive oil. Results of in vitro fermentation showed low degradation of thymol and carvacrol. By contrast, large catabolism was noted when rosmarinic acid and eriodictyol were fermented, yielding hydroxyphenylpropionic acid as the main metabolite. In accordance with these results, after the in vivo intervention with thyme phenol-enriched olive oil, an increase in the concentration of hydroxyphenylpropionic and phenylpropionic acids was observed in human feces, confirming the effective in vivo microbial degradation of rosmarinic acid and eriodictyol. Carvacrol was detected in fecal samples at trace levels, suggesting that monoterpenes are well absorbed in the upper part of the gastrointestinal tract.

Faecal microbial metabolism of olive oil phenolic compounds: in vitro and in vivo approaches.

SCOPE: In the present study, the individual colonic metabolism of the main components of the virgin olive oil phenolic fraction was evaluated by an in vitro model using human faecal microbiota. To assess differences in metabolism related to the molecular structure, four phenolic standards were selected, tyrosol, hydroxytyrosol, hydroxytyrosol acetate and oleuropein. After studying the in vitro colonic metabolism pathways of the individual phenols, the presence of their colonic metabolites was investigated in human faecal samples obtained before and after the sustained intake (3 weeks) of a daily dose of 25 mL of a phenol-enriched olive oil. METHODS AND RESULTS: The in vitro colon fermentation of the four individual phenolic compounds revealed (i) an increase in phenolic acids, (ii) the stability of hydroxytyrosol and tyrosol and (iii) the high degradation of hydroxytyrosol acetate and oleuropein. Additionally, a moderate intake of a phenol-rich olive oil raised the concentration in human faeces of free hydroxytyrosol and phenylacetic and phenylpropionic acids. CONCLUSION: The products of colonic catabolism of olive oil phenolic compounds could be good candidates for novel preventive strategies and open a promising line of research into the preventive action of olive oil phenols in colon and other bowel diseases.

Optimisation and validation of analytical methods for the simultaneous extraction of antioxidants: application to the analysis of tomato sauces.

In the present study, simultaneous extraction of natural antioxidants (phenols and carotenoids) in complex matrices, such as tomato sauces, is presented. The tomato sauce antioxidant compounds studied were the phenolics hydroxytyrosol, from virgin olive oil, quercetin and its derivatives, from onions, and quercetin-rutinoside as well as the carotenoid, lycopene (cis and trans), from tomatoes. These antioxidant compounds were extracted simultaneously with n-hexane/acetone/ethanol (50/25/25, v/v/v). The phenolics were analysed by ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS), and lycopene (cis- and trans-forms) was analysed using high-performance liquid chromatography coupled to a diode array detector (HPLC-DAD). After studying the parameters of these methods, they were applied to the analysis of virgin olive oil, fresh onion, tomato concentrate and tomato powder, and commercial five tomato sauces. Subsequently, the results obtained in our laboratory were compared with those from the Gallina Blanca Star Group laboratory.

Effect of the co-occurring components from olive oil and thyme extracts on the antioxidant status and its bioavailability in an acute ingestion in rats.

The aim of this work was to examine whether bioactives in thyme could enhance the antioxidant capacity of phenolics in virgin olive oil and their bioavailability in Wistar rats. After acute oral administration of extracts from olive cake (OE), thyme (TE) or their combination (OTE), blood samples were collected from 0 to 360 min. Plasma antioxidant status was analyzed by DPPH and FRAP in plasma and by SOD, CAT and GPx activities in erythrocytes. Plasma pharmacokinetics of the main metabolites of bioactives in olive oil and thyme were characterized. Plasma non-enzymatic antioxidant capacity was significantly modulated by OE, TE, and OTE in a time-, assay, and extract-dependent manner. OE, TE, and OTE all significantly decreased superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity and catalase (CAT) activity was increased. Pharmacokinetic results showed that plasma concentration (Cmax) of the main olive phenolic metabolites in rats fed with OTE were similar to those of OE. These results indicate that an enhanced bioavailability of olive phenolic compounds could occur in the presence of thyme, although any synergistic effect was observed in the antioxidant status when both phenolic extracts were administered. Antioxidant protection by phenolics from olive and thyme against oxidative stress occurs primarily through a direct antioxidant effect and may be related to the phenolic plasmatic metabolites.

Gallic acid is an active component for the anticarcinogenic action of grape seed procyanidins in pancreatic cancer cells.

The aim of the present work was to evaluate the effects of a grape seed procyanidin extract (GSPE) on proliferation and apoptosis in the pancreatic adenocarcinoma cell line MIA PaCa-2 and identify the components of the extract with higher activity. The effects of the extract were analyzed on the proliferation and apoptosis processes in MIA PaCa-2 cells, as well as in the levels of the apoptosis markers Bcl-2 and Bax, the mitochondrial membrane potential, and reactive oxygen species levels. Finally, the components of the extract with higher effects were elucidated using enriched fractions of the extract and pure compounds. The results showed that GSPE inhibits cell proliferation and increases apoptosis in MIA PaCa-2 cells, which is primarily mediated by the downregulation of the antiapoptotic protein Bcl-2 and the depolarization of the mitochondrial membrane. GSPE also reduced the formation of reactive oxygen species. The component of the extract that possesses the highest antiproliferative and proapoptotic activity was gallic acid. In conclusion, GSPE acts as anticarcinogenic in MIA PaCa-2 cells, with gallic acid as the major single active constituent of the extract.

Effect of the co-occurring olive oil and thyme extracts on the phenolic bioaccessibility and bioavailability assessed by in vitro digestion and cell models.

Olive oils flavoured with edible herbs have grown in popularity because of their added value and potential health benefits. However, the combined presence of different phytochemicals from olive oil and herbs requires study of their possible interactions during intestinal transport and metabolism. The aim of this study was firstly to evaluate the effect on bioaccessibility of the co-occurring bioactive compounds from olive oil and thyme through an in vitro digestion model of three extracts: olive extract (OE), thyme extract (TE) and a combination of both (OTE). The bioaccessible fractions were exposed to Caco-2 and HepG-2 cell models, as well as to a co-culture of both of these. Results indicated that the bioaccessibility of hydroxytyrosol was enhanced when OTE was digested. After Caco-2 cells exposure, no significant differences were observed in hydroxytyrosol transport, whereas the main flavonoids from thyme seemed to undergo an enhanced basolateral permeation when both phenolic sources where exposed.

Application of dried spot cards as a rapid sample treatment method for determining hydroxytyrosol metabolites in human urine samples. Comparison with microelution solid-phase extraction.

Two different rapid sample pretreatment strategies, dried spot cards, and microelution solid-phase extraction plates (µSPE), with ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) have been developed and validated for the determination of hydroxytyrosol and its metabolites in spiked human urine samples. Hydroxytyrosol, hydroxytyrosol-3'-O-glucuronide, hydroxytyrosol-4'-O-glucuronide, hydroxytyrosol-3-O-sulphate, and homovanillic alcohol-4'-O-glucuronide were used as the target compounds. Using the FTA DMPK-A dried urine spot card under optimum conditions, with 5 µL of preconcentrated urine volume and 100 µL of methanol/water (50/50, v/v) as the elution solvent, the extraction recovery (%R) of the compounds studied was higher than 80%, and the matrix effect (%ME) was less than 8%. The stability of these cards and punching at the centre or side of the card were also studied, obtaining an excellent stability after 7 days of storage and complete homogeneity across the surface of the dried drop. The different µSPE parameters that affect the efficiency were also studied, and under optimum conditions, the %R and the %ME were higher than 70% and lower than 17%, respectively. The linearity range in dried urine spot cards was 2.5-20 µM for all the metabolites, with the exception of hydroxytyrosol-3-O-sulphate and hydroxytyrosol, which were 0.3-70 µM and 2.5-50 µM respectively. With regards to µSPE, the linearity range was 0.5-5 µM for all the studied compounds, except for hydroxytyrosol-3-O-sulphate, which was 0.08-5 µM. The quantification limits (LOQs) were 0.3-2.5 µM and 0.08-0.5 µM in dried spot cards and in µSPE, respectively. The two developed methods were then applied and compared for determining hydroxytyrosol and its metabolites in human 24 h-urine samples after a sustained consumption (21 days) of a phenol-enriched virgin olive oil. The metabolites identified were hydroxytyrosol in its glucuronide and sulphate forms, homovanillic alcohol in its glucuronide and sulphate forms, homovanillic acid sulphate and hydroxytyrosol acetate sulphate.

Biomarkers of food intake and metabolite differences between plasma and red blood cell matrices; a human metabolomic profile approach.

Untargeted metabolomic analyses of plasma and red blood cells (RBCs) can provide complementary information on biomarkers of food consumption. To assess blood collection differences in biomarkers, fasting blood was drawn from 10 healthy individuals using sodium citrate and lithium heparin as anticoagulants. Plasma and RBCs were separated into aqueous and lipid fractions to be analyzed using 1D and 2D (1)H NMR spectroscopy. Fatty acids were analyzed using gas chromatography-mass spectrometry (GC-MS). Polyphenols were extracted from plasma and RBCs by micro-elution solid-phase extraction and analyzed by ultra performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). (1)H NMR demonstrated higher aqueous metabolites such as glucose in plasma compared to RBCs, while RBCs contained higher ADP-ATP, creatine and acetone than plasma. Lipoproteins and their subclasses were higher in plasma than in RBCs. Percentages of saturated fatty acids (SFA) 16 : 0, 17 : 0, 20 : 0, 24 : 0 and polyunsaturated fatty acids (PUFA) 22 : 6 n-3 (docosahexaenoic acid) and 20 : 4 n-6 (arachidonic acid) were higher in RBCs than in plasma (p < 0.05), while SFA 14 : 0, monounsaturated fatty acids (MUFA) 14 : 1 n-5, 16 : 1 n-7, 17 : 1 n-7 and 18 : 1 n-9 and PUFA 18 : 3 n-3, 18 : 2 n-6, 18 : 3 n-6 and 20 : 3 n-6 were higher in plasma than in RBCs (p < 0.05). Polyphenols differed in plasma from those of RBCs. Biomarker concentrations were lower in sodium citrate compared to lithium heparin plasma. In conclusion, metabolomic profiles generated by NMR spectroscopy, GC-MS and UPLC-MS/MS analyses of RBCs versus plasma show complementary information on several specific molecular biomarkers that could be applied in nutritional assessment.

Flavanol metabolites distribute in visceral adipose depots after a long-term intake of grape seed proanthocyanidin extract in rats.

A considerable number of epidemiological investigations and intervention studies have supported an association between the intake of flavanol- and proanthocyanidin-containing foods and a decreased risk of metabolic diseases. Nonetheless, less is know about the capacity of tissues to accumulate flavanols and/or their metabolites. The main objective of the present study was to determine (n 20) plasma bioavailability and disposition in the liver, muscle, brown adipose tissue (BAT) and white adipose tissues (mesenteric and perirenal) in rats after a long-term consumption of three doses of grape seed phenolic extract (5, 25 and 50 mg/kg body weight) for 21 d in order to determine whether there is a dose-response relationship. Glucuronidated conjugates (total glucuronidated conjugates: C(5 mg/kg) 1·9; C(25 mg/kg) 6·4; C(50 mg/kg) 27·7 µmol/l plasma) followed by methyl glucuronidated conjugates (total methyl glucuronidated conjugates: C(5 mg/kg) 1·98; C(25 mg/kg) 4·48; C(50 mg/kg) 12·5 µmol/l plasma) were the main flavanol metabolites quantified in plasma, also detecting a dimer in its free form (C(25 mg/kg) 0·74; C(50 mg/kg) 0·79 µmol/l plasma). Each of the studied organs has a particular behaviour of accumulation and response to the assayed grape seed extract doses, with an exponential bioavailability-dose relationship in BAT, in which flavanols could play an important role in the reduction or prevention of obesity, modulating the functionality of that tissue.

Distribution of procyanidins and their metabolites in rat plasma and tissues in relation to ingestion of procyanidin-enriched or procyanidin-rich cocoa creams.

BACKGROUND: Procyanidins are extensively metabolized via phase-II and microbial enzymes. However, their distribution in the body is not well characterized. AIM: This study investigates the distribution of procyanidins (monomers and dimers) and their phase-II metabolites in plasma and tissues (thymus, heart, liver, testicle, lung, kidney, spleen and brain). METHODS: Wistar rats were fed with 1 g of cocoa cream (CC), 50 mg of procyanidin hazelnut skin extract (PE) and 50 mg PE in 1 g CC (PECC). The rats were killed at 0, 1, 1.5, 2, 3, 4 and 18 h after gavage, and the plasma and tissues were analyzed by UPLC-MS/MS. RESULTS: Epicatechin-glucuronide was the main metabolite in the plasma after the CC intake, with C(max) at 423 nM and t(max) at 2 h, and methyl catechin-glucuronide (301 nM, 2 h) was the main metabolite in the plasma after the PE intake. As a result of the PECC enrichment, epicatechin-glucuronide (452 nM, 1.5 h) and catechin-glucuronide (297 nM, 2 h) were the main metabolites in the plasma. Methyl catechin-glucuronide was found in the liver after PE (8 nmol/g tissue, 4 h) and PECC (8 nmol/g, 1.5 h). The kidney was found to contain a high concentration of phase-II metabolites of procyanidins and is therefore thought to be the main site of metabolism of the compounds. Methyl catechin-sulfate (6.4 nmol/g, 4 h) was only quantified in the brain and after PE intake. Catechin metabolites were not found in the spleen or heart. Phenolic acids were detected in all tissues. CONCLUSIONS: The formulation of a product enriched or fortified with procyanidins is a way to increase their bioavailability, with clear effects on the plasmatic pharmacokinetics, and a greater accumulation of phenolic metabolites in such tissues as the liver, kidney, lung and brain.