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A simple technique for the routine capillary electrophoretic analysis of phenolic compounds in red wines from different Spanish regions, is described. Separation was achieved using a fused-silica capillary column with a borate buffer at pH 9.5, and UV detection at 280 nm. The compounds identified were: (-)-epicatechin, (+)-catechin, (-)-epigallocatechin, syringic acid, vanillic acid, gallic acid, 3,4 dihydroxybenzoic acid, p-coumaric acid, caffeic acid and the hydroxycinnamic tartaric acid esterscis- andtrans-coumamyl tartaric acid andcis-caffeoyl tartaric acid. Different wines showed very similar electrophoretic profiles, but significant quantitative differences were observed.

Six anthocyanin pigments were found to be responsible for the red colour of pomegranate juice (cv ‘Mollar’). These were analysed quantitatively and qualitatively by HPLC and identified as delphinidin 3-glucoside and 3,5-diglucoside, cyanidin 3-glucoside and 3,5-diglucoside and pelargonidin 3-glucoside and 3,5-diglucoside. The fruit skin contained only the cyanidin and pelargonidin derivatives. Generally, there is an increase in juice pigmentation with fruit ripening. During early maturation stages the 3,5-diglucosides were the main pigments and the delphinidin-based derivatives were the predominant compounds. In late maturation stages the proportion of monoglucosides increased to reach values similar to or higher than those of the diglucosides, and the cyanidin-based derivatives were the predominant substances. The concentration of pigments in juice obtained from mature pomegranates ranged between 50 and 100 μg of anthocyanin per gram fresh weight of arils. The development of anthocyanins was also tested in fruits produced in two different orchards, and in fruits from different locations in the tree. Red coloured fruits, located in the outer parts of the tree, and yellow fruits, on the inner branches, were analysed. The juices obtained with the seed coats of both fruit types, and from the two orchards selected, showed the same anthocyanin profile. However, the total amount of pigments in the juice was generally smaller in those fruits with reddish skins (outer fruits), than in those with yellow skins (inner fruits). As juice pigmentation is one important quality factor in pomegranate, these results could be used in on-line determinations of the pomegranate quality by non-destructive physical methods.

Urolithins are dibenzo[b,d]pyran-6-one derivatives that are produced by the human gut microbiota from ellagitannins and ellagic acid (EA). These metabolites are much better absorbed than their precursors and have been suggested to be responsible for the health effects attributed to ellagitannins and EA that occur in food products as berries and nuts. In the present review, the role and potential of urolithins in human health are critically reviewed, and a perspective of the research approach needed to demonstrate these health effects is presented, based on the existing knowledge. The analytical methods available for urolithin analysis, their occurrence in different tissues and biological fluids, and their metabolism by human gut microbiota are considered. In addition, the interindividual variability observed for the production of urolithins (metabotypes) and its relationship with health status and dysbiosis are also reviewed. The potential mechanisms of action of urolithins are also critically discussed, paying attention to the concentration and the type of metabolites used in the in vitro and in vivo assays and the physiological significance of the results obtained. The gut microbiota metabolism of EA to urolithins and that of daidzein to equol, their individual variations, and the effects on health are also compared.

Both stingless-bee honey and Apis mellifera honey contain flavonoids that originate from nectar, pollen, and plant resins collected by bees. Such flavonoids indicate the botanical and geographical origins of honey and have associated health benefits. A survey of flavonoids in pot honey from Venezuela, Australia, Brazil, and Bolivia shows they are richer in flavonoid glycosides than Apis mellifera honey. Honey of Apis contains larger amounts of flavonoid aglycones that originate from propolis or from enzymatic hydrolysis of nectar flavonoid glycosides. The low diastase content of stingless-bee honey may explain the higher content of flavonoid glycosides in this honey.

Short-chain fatty acids are the major end products of bacterial metabolism in the large bowel. They derive mostly from the bacterial breakdown of carbohydrates and are known to have positive health benefits. Due to the biological relevance of these compounds it is important to develop efficient, cheap, fast, and sensitive analytical methods that enable the identification and quantification of the short-chain fatty acids in a large number of biological samples. In this study, a gas chromatography-mass spectrometry method was developed and validated for the analysis of short-chain fatty acids in faecal samples. These volatile compounds were extracted with ethyl acetate and 4-methyl valeric acid was used as an internal standard. No further cleanup, concentration, and derivatization steps were needed and the extract was directly injected onto the column. Recoveries ranged between 65 and 105%, and no matrix effects were observed. The proposed method has wide linear ranges, good inter- and intraday variability values (below 2.6 and 5.6%, respectively) and limits of detection between 0.49 μM (0.29 μg/g) and 4.31 μM (3.8 μg/g). The applicability of this analytical method was successfully tested in faecal samples from rats and humans.

Bacterial infections remain an important problem for human health. The control of bacterial infections has been traditionally treated by inhibiting microbial growth using different types of antibiotics. However, the ability of different bacteria to resist the inhibitory action of antibiotics has become a global problem. In fact, there is an important need for the development of new antimicrobials that act on novel bacterial targets. Many pathogenic bacteria control their population and regulate gene expression in response to their cell population density using diffusible signalling com¬pounds. This type of communication has been referred to as “quorum sensing” (QS). This phenomenon can be essential for the synchronization of the virulence production factors, which make it an attractive therapeutic target. Therefore, the search of non-toxic compounds, which inhibit QS and so, the virulence of pathogenic bacteria can bring new alternatives for the treatment of bacterial infections in humans. In this work, we made an attempt to screen the anti-QS activity of 11 bioactive compounds extracted from fruits and vegetables using the biosensor strain, Chromobacterium violaceum. The anti-QS activity was determined quantifying the violacein production of the biosensor strain at three concentrations (50, 100 and 200 µg/ml). At least five of the tested compounds (cinnamaldehyde, pomegranate extracts, ellagic acid resveratrol and rutin) showed anti-QS activity against the biosensor strain. The obtained results showed the potential of bioactive compounds extracted from fruits and vegetables to be used as a new category of anti-pathogenic compounds against bacterial infections. Fil: Truchado , P.. Consejo Superior de Investigaciones Cientificas. Center For Research In Agricultural Genomics.; España Fil: Tomás Barberán, F.. Consejo Superior de Investigaciones Cientificas. Center For Research In Agricultural Genomics.; España Fil: Allende, A.. Consejo Superior de Investigaciones Cientificas. Center For Research In Agricultural Genomics.; España Fil: Ponce, Alejandra Graciela. Universidad Nacional de Mar del Plata. Facultad de Ingeniería. Departamento de Ingeniería Química; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata; Argentina

The efficacy of resveratrol and some glucosyl, glucosylacyl, and glucuronide derivatives in inhibiting the adhesion of Salmonella Typhimurium, Escherichia coli O157:H7, and Listeria monocytogenes Scott A to Caco-2 and HT-29 colonic cells was investigated. The three bacteria strains were capable of adhering to both colonic epithelial cell lines, which responded by producing the pro-inflammatory interleukin 8 (IL-8). Adhesion inhibition of E. coli O157:H7 and S. Typhimurium to colonic cells was ≥60 and ≥40%, respectively, when resveratrol and most of the resveratrol derivatives were applied. Lower adhesion inhibition was observed for the bacteria with higher adherence potential, L. monocytogenes (≥20%). Resveratrol-3-O-(6'-O-butanoyl)-β-D-glucopyranoside (BUT) (50 and 100 μM) and resveratrol-3-O-(6'-O-octanoyl)-β-D-glucopyranoside (OCT) (50 μM) reduced IL-8 secretion by 100%. These results suggest that one mechanism for the beneficial attributes of resveratrol and especially the derivatives BUT and OCT could be the ability to reduce the adhesion and consequent pro-inflammatory cytokine production in intestinal epithelial cells in response to pathogen adhesion. The potential use of these compounds in the prevention of foodborne infections, intestinal homeostasis loss, and inflammatory bowel diseases could be another step in finding coadjuvants or alternatives to antibiotic treatments.

The cardioprotective role of resveratrol as part of the human diet is not yet clear. Our aim was to investigate the effect of a grape supplement containing 8 mg resveratrol in oxidized LDL (LDLox), apolipoprotein-B (ApoB), and serum lipids on statin-treated patients in primary cardiovascular disease prevention (PCP).A triple-blind, randomized, placebo-controlled trial was conducted. Seventy-five patients (three parallel arms) consumed one capsule (350 mg) daily for 6 months containing resveratrol-enriched grape extract (GE-RES, Stilvid®), grape extract (GE, similar polyphenolic content but no resveratrol), or placebo (maltodextrin). After 6 months, no changes were observed in the placebo group and only LDL cholesterol (LDLc) decreased by 2.9% (p = 0.013) in the GE group. In contrast, LDLc (-4.5%, p = 0.04), ApoB (-9.8%, p = 0.014), LDLox (-20%, p = 0.001), and LDLox/ApoB (-12.5%, p = 0.000) decreased in the Stilvid® group, whereas the ratio non-HDLc (total atherogenic cholesterol load)/ApoB increased (8.5%, p = 0.046). No changes were observed in hepatic, thyroid, and renal function. No adverse effects were observed in any of the patients.This GE-RES reduced atherogenic markers and might exert additional cardioprotection beyond the gold-standard medication in patients from PCP. The presence of resveratrol in the GE was necessary to achieve these effects.

Short-chain fatty acids are the major end products of bacterial metabolism in the large bowel. They derive mostly from the bacterial breakdown of carbohydrates and are known to have positive health benefits. Due to the biological relevance of these compounds it is important to develop efficient, cheap, fast, and sensitive analytical methods that enable the identification and quantification of the short-chain fatty acids in a large number of biological samples. In this study, a gas chromatography-mass spectrometry method was developed and validated for the analysis of short-chain fatty acids in faecal samples. These volatile compounds were extracted with ethyl acetate and 4-methyl valeric acid was used as an internal standard. No further cleanup, concentration, and derivatization steps were needed and the extract was directly injected onto the column. Recoveries ranged between 65 and 105%, and no matrix effects were observed. The proposed method has wide linear ranges, good inter- and intraday variability values (below 2.6 and 5.6%, respectively) and limits of detection between 0.49 μM (0.29 μg/g) and 4.31 μM (3.8 μg/g). The applicability of this analytical method was successfully tested in faecal samples from rats and humans.

[Background] Rosemary (Rosmarinus officinalis L.) extracts (REs) exhibit hepatoprotective, anti-obesity and anti-inflammatory properties and are widely used in the food industry. REs are rich in carnosic acid (CA) and carnosol which may be responsible for some of the biological activities of REs. The aim of this study was to investigate whether inhibition of lipase activity in the gut may be a mechanism by which a RE enriched in CA (40%) modulates body weight and lipids levels in a rat model of metabolic disorders and obesity., [Methods and Principal Findings] RE was administered for 64 days to lean (fa/+) and obese (fa/fa) female Zucker rats and body weight, food intake, feces weight and blood biochemical parameters were monitored throughout the study. Lipase activity (hydrolysis of p-nitrophenylbutyrate) was measured in the gastrointestinal tract at the end of the study and the contents of CA, carnosol and methyl carnosate were also determined. Sub-chronic administration of RE moderately reduced body weight gain in both lean and obese animals but did not affect food intake. Serum triglycerides, cholesterol and insulin levels were also markedly decreased in the lean animals supplemented with RE. Importantly, lipase activity was significantly inhibited in the stomach of the RE-supplemented animals where the highest content of intact CA and carnosol was detected., [Conclusions] Our results confirm that long-term administration of RE enriched in CA moderates weight gain and improves the plasma lipids profile, primarily in the lean animals. Our data also suggest that these effects may be caused, at least in part, by a significant inhibition of gastric lipase and subsequent reduction in fat absorption., This project was financed by the Spanish Centre for the Development of Industrial Technology (CDTI) as part of the project SENIFOOD, which belongs to the CENIT subvention program (Ref: CEN-20091006).

The presence of grape and wine polyphenol resveratrol (RES) in the diet is negligible. Therefore, the cardiovascular benefits of this molecule, in a dietary context, remain to be established. We aimed to investigate, through dietary intervention, the effects of a resveratrol-rich grape extract (GE-RES) on the prevention of early aortic lesions in pigs fed an atherogenic diet (AD). These effects were compared with those produced by a grape extract lacking RES (GE) or RES alone. Pigs fed the AD for 4 months showed early atherosclerotic lesions in the thoracic aorta: degeneration and fragmentation of elastic fibers, increase of intima thickness, subendothelial fibrosis, and accumulation of fatty cells and anion superoxide radicals. GE-RES was the most effective treatment and prevented the disruption of aortic elastic fibers, decreased their alteration (57%), and reduced the intima thickness (33%) and the accumulation of fatty cells (42%) and O(2)(•-) (38%) in aortic tissue. In addition, GE-RES moderately downregulated the expression of the suppressors of cytokine signaling 1 (SOCS1) and 3 (SOCS3), key regulators of vascular cell responses, in peripheral mononuclear blood cells. Our results suggest that the consumption of this GE-RES nutraceutical, in a dietary prevention context, could prevent early atherosclerotic events. The presence of RES in the grape extract strengthened these effects.

Flavanones, flavonoids abundant in Citrus , have been shown to interfere with quorum sensing (QS) and affect related physiological processes. We have investigated the QS-inhibitory effects of an orange extract enriched in O-glycosylated flavanones (mainly naringin, neohesperidin, and hesperidin). The QS-inhibitory capacity of this extract and its main flavanone components was first screened using the bacteriological monitoring system Chromobacterium violaceum . We next examined the ability of the orange extract and of some of the flavanones to (i) reduce the levels of the QS mediators produced by Y. enterocolitica using HPLC-MS/MS, (ii) inhibit biofilm formation, and (iii) inhibit swimming and swarming motility. Additionally, we evaluated changes in the expression of specific genes involved in the synthesis of the lactones (yenI, yenR) and in the flagellar regulon (flhDC, fleB, fliA) by RT-PCR. The results showed that the orange extract and its main flavanone components inhibited QS in C. violaceum, diminished the levels of lactones secreted by Y. enterocolitica to the media, and decreased QS-associated biofilm maturation without affecting bacterial growth. Among the tested compounds, naringin was found to inhibit swimming motility. Exposure to the orange extract and (or) to naringin was also found to be associated with induction of the transcription levels of yenR, flhDC, and fliA. This work shows the in vitro QS-inhibitory effects of an orange extract enriched in flavanones against a human enteropathogen at doses that can be achieved through the diet and suggests that consumption of these natural extracts may have a beneficial antipathogenic effect.

The search for complementary treatments in primary prevention of cardiovascular disease (CVD) is a high-priority challenge. Grape and wine polyphenol resveratrol confers CV benefits, in part by exerting anti-inflammatory effects. However, the evidence in human long-term clinical trials has yet to be established. We aimed to investigate the effects of a dietary resveratrol-rich grape supplement on the inflammatory and fibrinolytic status of subjects at high risk of CVD and treated according to current guidelines for primary prevention of CVD. Seventy-five patients undergoing primary prevention of CVD participated in this triple-blinded, randomized, parallel, dose‐response, placebo-controlled, 1-year follow-up trial. Patients, allocated in 3 groups, consumed placebo (maltodextrin), a resveratrol-rich grape supplement (resveratrol 8 mg), or a conventional grape supplement lacking resveratrol, for the first 6 months and a double dose for the next 6 months. In contrast to placebo and conventional grape supplement, the resveratrol-rich grape supplement significantly decreased high-sensitivity C-reactive protein (26%, p 0.03), tumor necrosis factor- (19.8%, p 0.01), plasminogen activator inhibitor type 1 (16.8%, p 0.03), and interleukin-6/interleukin-10 ratio (24%, p 0.04) and increased anti-inflammatory interleukin-10 (19.8%, p 0.00). Adiponectin (6.5%, p 0.07) and soluble intercellular adhesion molecule-1 (5.7%, p 0.06) tended to increase and decrease, respectively. No adverse effects were observed in any patient. In conclusion, 1-year consumption of a resveratrol-rich grape supplement improved the inflammatory and fibrinolytic status in patients who were on statins for primary prevention of CVD and at high CVD risk (i.e., with diabetes or hypercholesterolemia plus >1 other CV risk factor). Our results show for the first time that a dietary intervention with grape resveratrol could complement the gold standard therapy in the primary prevention of CVD. © 2012 Elsevier Inc. All rights reserved.

High-performance liquid chromatography with diode-array detection was used to characterize the phenolic profile of apricot fruits, juices and jams. Chlorogenic acid was the major phenolic compound present in all samples. Quercetin 3-rutinoside was the main flavonoid and kaemp-ferol 3-rutinoside was present in smaller amounts with traces of other quercetin and kaempferol glycosides. Quantitative differences in phenolics were observed in eleven apricot fruit varieties at three stages of maturity and also in some processed apricot products. A common phenolic profile was observed in all cases. The major portion of the flavonoids occurred in the skin of the fruit and the total phenolic content of apricot fruits had no influence over the softening behaviour observed in some varieties.

The flavonoids present in twenty samples of Portuguese heather honey were analysed by reversed-phase HPLC. The flavonoid fraction was purified by a combination of chromatography on Amberlite XAD-2 and Sephadex LH-20 columns. The total amount of flavonoids ranged between 60 and 500 μg/100g honey, values which are much smaller than those previously found for Spanish rosemary honey samples (500-2000 μg/100 g honey). The main flavonoids present in Portuguese heather honey were the flavanones pinocembrin and pinobanksin and the flavones chrysin and galangin. These analyses showed that all the samples contained a similar flavonoid pattern composed of at least 22 compounds. The most characteristic substances were myricetin, myricetin 3-methyl ether, myricetin 3'-methyl ether and tricetin. These four flavonoids have in common the structural feature of having a trioxygenated ring B (3',4',5'-trioxygenation), this being an attribute not found in the flavonoids detected in any other honey sample of different plant origin analysed so far, suggesting that these substances could probably be used as markers for the botanical origin of heather honey.

The separation of thirteen flavonoids from honey by micellar electrokinetic capillary chromatography (MECC) is described. All the flavonoids were separated on a fused-silica column (75 cm × 75 μm I.D.) with 0.2 M sodium borate buffer (pH 8.0)-50 mM sodium dodecyl sulphate-10% methanol. These conditions were applied to the separation of flavonoids from lavender, rosemary, citrus and heather honey samples, to establish correlations between the flavonoid profiles and the botanical origin of honey. Citrus honey was characterized by the accumulation of hesperetin, lavender by luteolin, rosemary by 8-methoxykaempferol and heather by some unidentified flavonoids. The influence of the geographical origin on the honey flavonoid pattern was also studied by MECC. Honey samples from Spain, Mexico and Canada were analysed and no significant differences were found.

The general objective is to provide an alternative methodology based on capillary electrophoresis (CE) to characterize flavonoids from honey and hence determine its botanical origin. The specific objective is to compare the separation of flavonoids by CE with those achieved by HPLC to assess CE as an alternative technique for the determination of honey flavonoids. Fourteen different flavonoids isolated from honey were analysed by MECC and compared to the HPLC separations. It was difficult to find specific experimental conditions to separate all the flavonoids from honey in a single MEKC run. Three chromatographic conditions are optimized and, depending on the flavonoid markers sought in honey, the appropriate detection method should be chosen. Compared to the HPLC results, it is clear that CE could be an alternative technique in honey flavonoids analysis and particularly in the study of its geographical and floral origin.

This work was afforded from 2 points of view, phytochemical evaluation and relation to antioxidant activity and dietary burden of phenolics of a cup of "Mountain tea", a drink obtained by domestic infusion of Sideritis. Phytochemically, two extraction protocols using water and methanol as solvent were used for comparison. Methanol and boiling water extracts (by domestic infusion procedure) showed that extracts were rich in bound forms of phenolics such as hydroxycinnamic acids, phenylethanoid glycosides and flavonoid glycosides. The total phenolic content for Sideritis species ranged around 190 mg per serving (2 g infusion bag) for methanol extracts and around 72 mg per serving in water extracts. Among the two different Macedonian Sideritis species, Sideritis raeseri (wild growing) showed the highest phenolics content in both extracts (212 mg and 89 mg per serving, respectively). Concerning the phenolic content in the different aerial parts, leaf was the richest plant organ in phenolics followed by flower and stem with the lowest amount. The methanol extract from Sideritis raeseri (wild growing) showed the highest antioxidant capacity as shown by DPPH, ABTS and FRAP assays. The antioxidant capacity was linearly correlated with phenolic content. Nutritionally, the dietary burden of phenolics of a "Mountain tea" bag for domestic infusion (serving size) was established at 89 mg for an homogeneous and equal distribution of the different aerial parts (leaf, flower and stem). However, and according to our results a rate of 60% leaf and 40% flower would increase the content of bioavailable phenolics and also the total phenolics content of a serving bag of "Mountain tea".

Metabolic and cardiovascular diseases (CVDs) have risen to alarming proportions, and there is a need for therapeutic and preventive measures. The polyphenol resveratrol (RES) protects against CVDs, but in vivo molecular mechanisms responsible for protection are not yet understood. Peripheral blood mononuclear cells (PBMNCs) are involved in the development of atherosclerosis and metabolic disorders. The identification of PBMNCs genes responding to dietary compounds might help to understand the mechanisms underlying the effects of polyphenols. We determined gene expression differences between PBMNCs from pigs fed a high-fat diet manifesting a mild increase of cholesterol and pigs fed a high-fat diet containing low doses of RES. Although the consumption of RES did not modify the levels of cholesterol, microarray analyses indicated that some of the differentially expressed genes, collagens (COL1A, COL3A), lipoprotein lipase (LPL) and fatty-acid binding proteins (FABPs) involved in CVDs and lipid metabolism were up-regulated by the high-fat diet and down-regulated by RES. Reverse transcriptase polymerase chain reaction confirmed that RES and RES-containing grape extract prevented the induction of FABP4 in PBMNCs in female pigs fed a high-fat diet. Low micromolar concentrations of RES and its metabolite dihydroresveratrol exerted a minor but significant reducing effect on the induction of FABP4 expression in human macrophages treated with oxidized low-density lipoprotein. Our results show that the consumption of low doses of RES modulates the expression of genes related to lipid metabolism and metabolic disorders that are affected by a high-fat diet and suggest that some of the circulating RES metabolites may contribute to these effects.

Propolis and propolis balsam from Canada was analyzed by gas chromatography-mass spectrometry and high performance liquid chromatography and the results compared. The compounds identified indicated that the main plant source was Populus from Section Aigeiros

An easy technique that enables the analysis of flavonoids from citrus jams is described. The flavonoids from commercial sour orange, sweet orange, mandarin, lemon, bitter lemon, grapefruit and lime jams were analysed by HPLC. Each citrus jam has a characteristic flavonoid pattern that can be used in genuineness determinations. Commercial jams produced with mixtures of orange, lemon and grapefruit can easily be characterized by HPLC. No differences between the flavonoid profiles of natural fruits and those of the corresponding jams were observed, showing that the manufacturing process does not affect the flavonoid profile and, therefore, that this profile can be used in genuineness studies.

Two distinct flavonoid patterns have been found in plants of Gratiola officinalis which are not correlated with morphological differences. One of the chemotypes is characterized by the production of 8-hydroxylated flavone O -glycosides, among which are the new plant substances 8-hydroxychrysoeriol 8- O -glucuronide, hypolaetin 7- O -sophoroside, 8-hydroxychrysoeriol 7- O -sophoroside and isoscutellarein 8- O -sophoroside. The flavonoids of the other chemotype are lacking 8-hydroxylation, but some are hydroxylated in the 6-position. Both chemotypes are similar to each other in that they also contain flavone C -glycosides, a class of flavonoid which is rare in the Scrophulariaceae.

The phenolic compounds present in 38 propolis samples produced in tropical Venezuela by imported Apis mellifera and five indigenous species of stingless bees, and seven propolis samples from different temperate regions were analysed by HPLC. The different tropical propolis generally showed a single phenolic profile characterized by the occurrence of polyprenylated benzophenones as major components. Flavonoids were present in only a few samples, and in each case they were found to be lipophilic methylated 6-oxygenated flavones (eupatorin, hispidulin, 5-hydroxy-6,7,3′,4′-tetramethoxyflavone, etc.). Generally, no correlations between the composition of tropical propolis and the place of collection or the bee species were found. However, propolis from temperate areas contained flavonoids (with an unsubstituted ring B) as the main phenolic components (pinobanksin, pinobanksin 3-acetate, pinocembrin, chrysin, galangin and tectochrysin). These analyses confirmed that the source of the phenolics found in propolis produced in temperate regions was different Populus species of section Aigeiros (Salicaceae). The resins exuded by the flowers of Clusia minor and C. major (Guttiferae) contained the polyprenylated benzophenones found in the majority of the propolis studied, suggesting that this is the main source for propolis in tropical Venezuela.

The occurrence of the anti-inflammatory flavonoids hypolaetin 8-glucoside, cirsiliol and sideritoflavone, and other flavonoids has been investigated in taxonomically related taxa of the genus Sideritis by HPLC. All the analysed taxa showed very similar flavonoid aglycone patterns, while they had clear differences in the flavonoid glycoside patterns. The analyses of flavonoid glycosides allowed the differentiation of five different groups of Sideritis taxa. There is a correlation between the chemical groups and the geographical location of the analysed taxa. In S. mugronensis, S. tragoriganum and S. reverchonii , the flavonoids hypolaetin 8-glucuronide, hypolaetin 4′-methyl ether 8-glucoside and 8-glucuronide have also been identified, this being the first time that these substances are reported in the Labiatae.

Twenty flavonoid aglycones from honey were analysed by HPLC on reversed-phase columns. Different solvents were used in order to optimize the detection of those flavonoids which could be considered as markers for the floral origin of honey. None of the solvent systems used allowed the resolution of all the flavonoids from honey included in this analysis. The different solvent systems were then applied to the analysis of flavonoids from citrus and rosemary honeys. The methanol-water system permitted the separation of hesperetin, the marker of citrus honey, whereas the acetonitrile-water system was the best for the separation of all the flavanones and the detection of apigenin, the marker of rosemary honey. The presence of the flavone techtochrysin was also demonstrated in both honeys. The use of a diode-array detector proved very useful for studies of the floral origin of honey by HPLC flavonoid analysis.

The flavonoids present in beeswax produced in “La Alcarria” region were analyzed by HPLC. Pinocembrin, pinobanksin, pinobanksin 3-acetate, chrysin, galangin and techtochrysin were detected as the main flavonoid constituents. This is the first detailed report on the flavonoids of beeswax. These substances are already present when wax scales are secreted by bees. The same flavonoid compounds were generally present in honey, propolis and Populus nigra bud exudates collected in the same geographical region. These results indicate that beeswax flavonoids originate from those of honey and/or propolis, and suggest that analysis of beeswax flavonoids could be used as an adjunct in the detection of beeswax adulterations.

The flavonoids present in honey samples from different geographical areas (Europe, North America, Equatorial regions, South America, China and Australia) have been analysed by HPLC. These flavonoids are incorporated into honey from propolis, nectar or pollen. As a general rule, honey samples from the Northern Hemisphere (where poplars, the source of propolis, are native) show flavonoid profiles characterized by the presence of propolis flavonoids. In contrast, honey samples from most Equatorial regions and Australia are generally devoid of propolis-derived flavonoids, showing only flavonoids from other plant sources. However, several honey samples from Central and South America and from New Zealand do contain the flavonoids characteristic of propolis. This means that importedApis mellifera colonies may locate poplar trees, occasionally finding an imported specimen in gardens or agro-industrial exploitations, and incorporate propolis flavonoids into honey. These preliminary results show that flavonoid analysis could be used as an adjunct to geographical origin studies of honey.

The occurrence of two dihydrochalcone glycosides has been demonstrated in Golden Delicious apple juices and jams by HPLC analysis. These substances have been isolated by a combination of Amberlite XAD-2 and reversed-phase liquid chromatography and by semipreparative HPLC. They have been completely identified as 2′,4′,6′,4- tetrahydroxydihydrochalcone -2′-O-β- d -glucopyranoside (phloridzin) and 2′,4′,6′,4- tetrahydroxy-dihydrochalcone -2′-O-(6″-β- d -xylopyranosyl )-β- d - glucopyranoside by spectroscopic methods.

8 páginas, 5 figuras, 4 tablas.-- et al., To select and establish floral biomarkers of the botanical origin of Diplotaxis tenuifolia honeys, the flavonoids and glucosinolates present in bee-deposited nectar collected from hive combs (unripe honey) and mature honey from the same hives fron which the unripe honey samples were collected were analyzed by LC-UV-PAD-ESI-MSn. Glycosidic conjugates of the flavonols quercetin, kaempferol, and isorhamnetin were detected and characterized in unripe honey. D. tenuifolia mature honeys contained the aglycones kaempferol, quercetin, and isorhamnetin. The differences between the phenolic profiles of mature honey and freshly deposited honey could be due to hydrolytic enzymatic activities. Aliphatic and indole glucososinolates were analyzed in unripe and mature honeys, this being the first report of the detection and characterization of glucosinolates as honey constituents. Moreover, these honey samples contained different amounts of propolis-derived flavonoid aglycones (1765−3171 μg/100 g) and hydroxycinnamic acid derivatives (29−1514 μg/100 g). Propolis flavonoids were already present in the freshly deposited nectar, showing that the incorporation of these compounds to honey occurs at the early steps of honey production. The flavonoids quercetin, kaempferol, and isorhamnetin and the glucosinolates detected in the samples could be used as complementary biomarkers for the determination of the floral origin of Argentinean Diplotaxis honeys., Support of the EUfunded Project BEE-DOC, ref: FP7-KBBE-2009-3 (244956), is gratefully acknowledged. P.T. holds a Ph.D. grant from the Seneca Foundation (Murcia, Spain).

Orange juice is a very rich source of dietary flavanones. The effect of flavanone concentration and solubility of orange beverages on their bioavailability has been studied in a crossover study with 10 healthy volunteers. Five different beverages with different flavanone concentrations were evaluated. Commercial orange juices (29.2-70.3 mg of flavanones/100 mL) were compared with experimental orange beverages in which the flavanone concentration was enhanced (110.2 mg/100 mL). Hesperetin and naringenin glucuronides and sulfates were detected and quantified in plasma and urine. The study shows that the solubility of the flavanones, and particularly that of hesperidin, in the juice is a key factor for the bioavailability as flavanone excretion and the C(max) in plasma correlate well with the soluble flavanone concentration in the juice, whereas it has no correlation with the total flavanone intake. In addition, a large interindividual variation was observed, this being consistent for each individual after the intake of the different beverages, suggesting that flavanone bioavailability is also dependent on the occurrence of specific microbiota that is able to remove the rutinosides from the juice glycosides, which results in aglycones that are then absorbed from the gut.

The voltammetric behaviour of ellagic acid (EA) is investigated by cyclic, differential pulse and square-wave voltammetry (CV, DPV and SWV, respectively). Based on the anodic oxidation peak at approximately 0.42 V in acetic/acetate buffer (pH 5.5) a robust and a highly reliable square-wave voltammetric method is presented for the determination of EA. The oxidation peak current was linearly dependent on the concentration of EA in the range of 1.0 × 10−7–1.5 × 10−6 mol/L (r = 0.9997), with a detection limit of 1.0 × 10−8 mol/L (S/N = 3) and a quantification limit of 3.4 × 10−8 mol/L (S/N = 10), good reproducibility and a satisfactory level of selectivity towards others polyphenols. The proposed method was applied to the determination of free and total EA in fruits, nuts and juices with good analytical results being obtained.

12 páginas, 7 figuras, 2 esquemas., There is no pharmaceutical or definitive surgical cure for inflammatory bowel diseases (IBDs). The naturally occurring polyphenol resveratrol exerts anti-inflammatory properties. However, its rapid metabolism diminishes its effectiveness in the colon. The design of prodrugs to targeting active molecules to the colon provides an opportunity for therapy of IBDs. Herein we explore the efficacy of different resveratrol prodrugs and pro-prodrugs to ameliorate colon inflammation in the murine dextran sulfate sodium (DSS) model. Mice fed with a very low dose (equivalent to 10 mg for a 70 kg-person) of either resveratrol-3-O-(6′-O-butanoyl)-β-d-glucopyranoside (6) or resveratrol-3-O-(6′-O-octanoyl)-β-d-glucopyranoside (7) did not develop colitis symptoms and improved 6-fold the disease activity index (DAI) compared to resveratrol. Our results indicate that these pro-prodrugs exerted a dual effect: (1) they prevented the rapid metabolism of resveratrol and delivered higher quantities of resveratrol to the colon and (2) they reduced mucosal barrier imbalance and prevented diarrhea, which consequently facilitated the action of the delivered resveratrol in the colon mucosa., This work was supported by the projects 200670F0131 (CSIC), CICYT-BFU2007-60576, and CSD2007-00063 (Fun-C-Food; Consolider Ingenio 2010). M.L. and R.L. are holders of a JAE-DOC grant from CSIC and M.V.S of a “Ramón y Cajal” grant from MCINN. J.T.C. is holder of a predoctoral grant from MCINN.

The occurrence of flavonoid p -coumaroylglucosides and 8-hydroxyflavone 7-allosylglucosides is restricted, in the Labiatae, to some genera of subfamily Lamioideae (tribe Stachydeae). Generally both types of compounds do not co-occur in the same species. Thus, flavonoid p -coumaroylglucosides are present in all the species studied of the genera Ballota , Phlomis and Marrubium , and are universally present in the subgenus Marrubiastrum of Sideritis , in subgenus Galeopsis and in section Betonica of genus Stachys , while 8-hydroxyflavone 7-allosylglucosides accumulate in the other Sideritis , Stachys and Galeopsis species. Amongst the flavonoid p -coumaroylglucosides, the apigenin derivatives occur most frequently but also the corresponding luteolin, chrysoeriol, kaempforol and isorhamnetin glycosides have been detected. The 8-hydroxyflavone allosylglucosides occur as such and also as the monoacetylated and diacetylated derivatives. The presence of p -coumaroylglucosides and/or 8-hydroxyflavone allosylglucosides shows the affinities of Anisomeles , Pogostemon , and Leonurus species with Ballota , Marrubium , Phlomis , Sideritis , Stachys and Galeopsis species. These results suggest that flavonoid p -coumaroylglucosides are valuable markers for chemotaxonomic studies.

A comparative study of the recovery of flavonoids (both aglycones and glycosides) by different Amberlite XAD resins has been achieved. No significant differences in the percentages of recovery were found with the chemical nature and physical characteristics of the different resins. A flavonoid recovery higher than 90% is possible in a number of cases. The possibility of fractionating the adsorbed flavonoids by desorption with methanol: water mixtures has also been studied. The polystyrene resins (XAD-2, XAD-4 and XAD-16) were found to be more useful for fractionation purposes than the polyacrylic resins (XAD-7, XAD-8). The results suggest that Amberlite XAD-2 is the most suitable for the recovery and fractionation of flavonoids from plant extracts and foods rich in sugars and other polar compounds such as honey and fruit jams.

The flavonoids present in ten selected samples of “La Alcarria” honey with different pollen compositions have been HPLC analysed in order to establish if correlations between botanical origin and flavonoid profiles are possible. A common flavonoid pattern is observed in the different samples showing that pollen is not the main source of honey flavonoids. A close correlation between the flavonoid patterns of honey flavonoids and propolis flavonoids has been found suggesting that flavonoid analysis could be more useful in geographical origin determinations than in botanical origin studies.

The phenolic compounds of 15 strawberry cultivars grown in Spain were analyzed and quantified: anthocyanins (20.2-47.4 mg/100 g of fw) (cyanidin 3-glucoside and pelargonidin 3-glucoside, 3-rutinoside, and 3-malonyl glucoside), flavonols (1.5-3.4 mg/100 g of fw) (quercetin 3-glucuronide and kaempferol 3-glucoside and 3-p-coumaroyl-glucoside), proanthocyanidins (53.9-163.2 mg/100 g), p-coumaroyl-glucose (0.84-6.70 mg/100 g), ellagitannins (9.67-22.86 mg/100 g) (sanguiin H-6, lambertianin C, and galloyl bis-HHDP-glucose), and ellagic acid glycosides (0.88-2.06 mg/100 g of fw) (two ellagic acid deoxyhexosides). Proanthocyanidins, the main phenolic compounds, were characterized by phloroglucinol degradation. Their mean degree of polymerization ranged from 3.4 for cv. Chiflon to 5.8 for cv. Ventana, the average value being 4.3. The terminal unit of proanthocyanidin oligomers was always (epi)catechin (17.36-29.93%) and (epi)catechin (61.66-75.39%) or (epi)afzelechin (4.50-10.54%) as extension units. Different combinations of (epi)catechin and (epi)afzelechin were detected, and their sequence of linkage was characterized by HPLC-MS-MS. Relative percentages of dimers, trimers, tetramers, and pentamers were evaluated by the extracted ion chromatogram (EIC) analysis.

8 páginas, 5 figuras, 3 tablas., Bacteria are able to communicate and coordinate certain processes using small secreted signaling molecules called autoinducers. This phenomenon, known as “quorum sensing” (QS), may be essential for the synchronization of virulence factors as well as biofilm development. The interruption of bacterial QS is acknowledged to attenuate virulence and considered to be a potential new therapy to treat infections caused by pathogenic bacteria. N-Acyl-l-homoserine lactones (AHLs) have been identified as the main bacterial signaling molecules in Gram-negative bacteria. This study evaluates the capacity of chestnut honey and its aqueous and methanolic extracts to inhibit bacterial AHL-controlled processes in Erwinia carotovora, Yersinia enterocolitica, and Aeromonas hydrophila. This study is the first in applying liquid chromatography coupled with tandem mass spectrometry to determine the QS inhibitory activity of honey against pathogenic bacteria. The tandem mass spectrometry analysis of culture supernatants confirmed the presence of three main AHLs: N-(3-oxohexanoyl)-l-homoserine lactone (3-oxo-C6-HSL) and N-hexanoyl-l-homoserine lactone (C6-HSL) in E. carotovora and Y. enterocolitica and N-butanoyl-l-homoserine lactone (C4-HSL) in A. hydrophila. The effect of chestnut honey and its aqueous and methanolic extracts (0.2 g/mL) on AHL concentration and biofilm formation in bacterial cultures was determined. The obtained results revealed their potential use as QS inhibitors or regulators of the degradation of QS signals, with the methanolic extract showing less inhibitory capacity. Thus, the QS inhibitory activity of chestnut honey seems to be related to the aqueous phase, suggesting that the carbohydrate fraction contains an antipathogenic substance responsible for the inhibitory activity., Support of the EUFunded Project “Bees in Europe and Sustainable Honey Production” (“BEE SHOP”, ref: FOOD 2006-022568-BEESHOP) is gratefully acknowledged. P.T. holds a Ph.D. grant from the Seneca Foundation (Murcia, Spain).

Epidemiology supports the important role of nutrition in prostate cancer (PCa) prevention. Pomegranate juice (PJ) exerts protective effects against PCa, mainly attributed to PJ ellagitannins (ETs). Our aim was to assess whether ETs or their metabolites ellagic acid and urolithins reach the human prostate upon consumption of ET-rich foods and to evaluate the effect on the expression of three proliferation biomarkers. Sixty-three patients with BPH or PCa were divided into controls and consumers of walnuts (35 g walnuts/day) or pomegranate (200 mL PJ/day) for 3 days before surgery. Independently of the ETs source, the main metabolite detected was urolithin A glucuronide, (3,8-dihydroxy-6H-dibenzo[b,d]pyran-6-one glucuronide) (up to 2 ng/g) together with the traces of urolithin B glucuronide, (3-hydroxy-6H-dibenzo[b,d]pyran-6-one glucuronide) and dimethyl ellagic acid. The small number of prostates containing metabolites was likely caused by clearance of the compounds during the fasting. This was corroborated in a parallel rat study and thus the presence of higher quantities of metabolites at earlier time points cannot be discarded. No apparent changes in the expression of CDKN1A, MKi-67 or c-Myc were found after consumption of the walnuts or PJ. Our results suggest that urolithin glucuronides and dimethyl ellagic acid may be the molecules responsible for the beneficial effects of PJ against PCa.

The consumption of flavonoid-rich Citrus extracts has been associated with multiple beneficial effects including anti-inflammatory properties, but the potential effects on the inflammatory responses in the gut have not been thoroughly investigated. We used microarrays to search for molecular changes induced in human colon fibroblasts in response to the exposure to a flavanone-rich bitter orange extract under physiologically representative conditions. Dietary nontoxic levels of the predigested extract induced moderate but significant changes in the expression of genes associated with tissue repair and inflammation. Among the top regulated genes, plasminogen activator inhibitor 1 (PAI-1) was downregulated, and the matrix metallopeptidase 12 (MMP-12) was upregulated (mRNA and protein levels). Both proteins are involved in extracellular matrix (ECM) remodeling and fibroblast migration. The extract also affected the fibroblast migration and reduced monocyte adhesion, but the response was different in unstimulated cells and in cells pretreated with TNF-alpha. Collectively, these results were indicative of a moderate activation of the colon fibroblast inflammation-related function after exposure to the extract. Further investigations are required to identify the in vivo role of this Citrus derived extract in the maintenance of the normal balance in the intestine and in the pathogenesis of inflammatory diseases.

Novel gene expression profiles and cellular functions modulated in Caco-2 cells in response to the dietary polyphenol, ellagic acid (EA), and its colonic metabolites, urolithin-A (3,8-dihydroxy-6H-dibenzo[b,d] pyran-6-one) and urolithin-B (3-hydroxy-6H-dibenzo[b,d] pyran-6-one) have been identified. Exposure of cells to EA and urolithins arrested cell growth at the S- and G(2)/M-phases. Transcriptional profiling using microarray and functional analysis revealed changes in the expression levels of MAPK signalling genes such as, growth factor receptors (FGFR2, EGFR), oncogenes (K-Ras, c-Myc), and tumour suppressors (DUSP6, Fos) and of genes involved in cell cycle (CCNB1, CCNB1IP1). Results suggest that EA and urolithin-A and -B, at concentrations achievable in the lumen from the diet, might contribute to colon cancer prevention by modulating the expression of multiple genes in epithelial cells lining the colon. Some of these genes are involved in key cellular processes associated with cancer development and are currently being investigated as potential chemopreventive targets.

The biological activity of dietary polyphenols and glucosinolates is mediated by their bioavailability and metabolism. In the present review, the scientific knowledge regarding the absorption and metabolism of these bioactive compounds is presented. The comparative absorption kinetics of different phenolics and the key role of colonic microbiota on phenolic metabolism are also reviewed. In addition, the physiological changes of glucosinolates after intake and the effect of food matrix on their bioavailability are discussed.

9 páginas, 4 figuras, 5 tablas., HPLC-MS–MS analysis of unifloral honey extracts has shown the occurrence of flavonoid glycosides in most of the analyzed samples. These compounds are not present in large amounts, but can reach up to 600 μg/100 g honey in canola and rapeseed honeys. Rhamnosyl-hexosides (tentatively rutinosides and neohesperidosides) and dihexosides (hexosyl(1→2)hexosides and hexosyl(1→6)hexosides) of flavonols such as quercetin, kaempferol, isorhamnetin and 8-methoxykaempferol, are the main flavonoid glycosides found in honey. However, flavonoid triglycosides and monoglycosides are also detected in some floral origins. Eucalyptus and orange blossom nectars were collected and analyzed showing that nectar flavonoid glucosides, as is the case of eucalyptus flavonoids, can be readily hydrolyzed by the bee saliva enzymes, while flavonoid rhamnosyl-glucosides, as is the case of citrus nectar flavonoids, are not hydrolyzed, and because of these reasons the flavonoid glycoside content of citrus honey is higher than that of eucalyptus honey that contains mainly aglycones. The flavonoid glycoside profiles detected in honeys suggest that this could be related to their floral origin and the results show that the HPLC-MSn ion trap analysis of flavonoid glycosides in honey is a promising analytical method to help in the objective determination of the floral origin of unifloral honeys., This work has been funded by the European Commission; project Beeshop, FOODCT-2006-022568.

7 páginas, 4 figuras, 3 tablas., To identify potential floral markers of chestnut honey, the phytochemicals present in chestnut floral nectar collected by bees were analyzed. Two nitrogen-containing compounds were detected, isolated, and identified as 4-hydroxyquinaldic acid (kynurenic acid) and 4-quinolone-2-carboxylic acid by 1H NMR and 13C NMR. In addition, chestnut nectar contained the monoterpene 4-(1-hydroxy-1-methylethyl)cyclohexa-1,3-diene-1-carboxylic acid, its gentiobioside ester, and the flavonol quercetin 3-pentosylhexoside. These nectar markers were found in different chestnut unifloral honey samples, although the flavonol was not detected in all samples analyzed. The terpenoid derivatives had previously been found in linden and tilia honeys. These results show that quinoline alkaloids are potentially good markers of chestnut honey, as they were not detected in any other unifloral honey analyzed so far. They are present at concentrations ranging from 34 to 65 mg/100 g of honey in the samples analyzed. In addition, the terpenoid and flavonoid derivatives present in nectar, although not exclusively characteristic of this floral origin, are good complementary markers for the determination of the floral origin of chestnut honey., This work has been funded by the European Commission, Project Beeshop, FOOD CT-2006-022568.

8 páginas, 3 figuras, 6 tablas., Quorum sensing (QS) inhibitory activity of 29 unifloral honeys was evaluated using the bacterial model Cromobacterium violaceum. The tested honeys were able to inhibit the production of acyl-homoserine lactones (AHLs) produced by C. violaceum at 0.1 g/ml. However, chestnut and linden honey samples showed the highest inhibitory activity, while orange and rosemary were less effective in inhibiting QS. When honey samples from the same floral origin obtained from different geographical regions were compared, they showed similar QS inhibitory activity. Thus, one of the factors which influence the inhibitory activity could be derived from the floral origin, independently of the geographic location. It was observed that unifloral honey samples showed “non-peroxide” anti-QS activity, which was not linearly correlated with total and individual phenolic compounds. The obtained results showed that the preservative properties of honey could be due to both the antimicrobial properties and the QS inhibitory capacity., Support of the EU-Funded Project ‘‘Bees in Europe and Sustainable Honey Production” (‘‘BEE SHOP”, Ref: FOOD 2006-022568- BEESHOP) is gratefully acknowledged. A. Allende and F. López-Gálvez hold grants founded by CSIC (JaeDoc- 2007 and I3P contracts) and P. Truchado holds a PhD grant from the Seneca Fundation (Murcia, Spain).

5 flavonoids aglycons and 8 flavonol glycosides were isolated from broad bean pods. By chemical, chromatographic and spectral methods the aglycons were identified as 7,3',4'-trihydroxyflavone, 7,4'-dihydroxyflavone, geraldone, butein and keamferol. The concentration changes of the flavonoids in pod tissues with progressive stages of maturity were evaluated by HPLC analysis.

A phytochemical study of the flavonoid glycosides present in the aerial parts of Galeopsis angustifolia has shown the existence of hypolaetin, isoscutellarein and hypolaetin 4′-methyl ether 7- O -β- d -(2″- O -β- d -allosyl)glucosides as well as their mono- O -acetylated and di- O -acetylated derivatives in the sugar moiety. The distribution of these compounds in different Galeopsis species has been studied by reversed-phase HPLC. This survey shows that the 8-hydroxyflavone glycosides are confined to subgenus Ladanum while subgenus Galeopsis accumulates luteolin, apigenin and scutellarein 7- O -β- d -glucuronides and p -coumaroyl glucosides of apigenin and luteolin instead.

Recently, it was shown that the exposure to the potent hop phytoestrogen 8-prenylnaringenin (8-PN) depends on intestinal bacterial activation of isoxanthohumol (IX), but this occurs in only one-third of tested individuals. As the butyrate-producing Eubacterium limosum can produce 8-PN from IX, a probiotic strategy was applied to investigate whether 8-PN production could be increased in low 8-PN producers, thus balancing phytoestrogen exposure. Using fecal samples from high (Hop +) and low (Hop -) 8-PN-producing individuals, a Hop + and Hop - dynamic intestinal model was developed. In parallel, Hop + and Hop - human microbiota-associated rats were developed, germ-free (GF) rats acting as negative controls. IX and then IX + E. limosum were administered in the intestinal model and to the rats, and changes in 8-PN production and exposure were assessed. After dosing IX, 80% was converted into 8-PN in the Hop + model and highest 8-PN production, plasma concentrations, and urinary and fecal excretion occurred in the Hop + rats. Administration of the bacterium triggered 8-PN production in the GF rats and increased 8-PN production in the Hop - model and Hop - rats. 8-PN excretion was similar in the feces (294.1 +/- 132.2 nmol/d) and urine (8.5 +/- 1.1 nmol/d ) of all rats (n = 18). In addition, butyrate production increased in all rats. In conclusion, intestinal microbiota determined 8-PN production and exposure after IX intake. Moreover, E. limosum administration increased 8-PN production in low producers, resulting in similar 8-PN production in all rats.