The matrix of fruit & vegetables modulates the gastrointestinal bioaccessibility of polyphenols and their impact on dietary protein digestibility.

Fruit and vegetables (F&V) polyphenols have numerous positive health effects, ascribed either to their antioxidant activity within the gastrointestinal tract (GIT) or to bioactivity of their absorbed metabolites. The effect of the F&V matrix on the gastrointestinal bioaccessibility of polyphenols was investigated along with its possible interaction with protein digestion. Minipigs were fed a complete meal with either cubed F&V (apple, plum, artichoke) added, or the corresponding phenolic extract (PE). Gastric and ileal chymes were kinetically collected over the postprandial period. The overall polyphenol bioaccessibility in the stomach was found to be 1.5% and 3.1% after F&V and PE consumption, respectively. The lower release rate from artichoke than from apple showed evidence of a plant effect. Flavanol monomers and glucoside conjugates were not recovered in the ileum in agreement with their absorption in the upper GIT. Interestingly, PE, but not F&V, significantly decreased the speed and efficiency of dietary protein digestion.

Characterization and quantification of fruit phenolic compounds of European and Tunisian pear cultivars.

The flesh and peel of 19 pear cultivars (8 Tunisian dessert cultivars, 8 European dessert cultivars and 3 French perry pear cultivars) were studied for their phenolic composition. Phenolic compounds were identified by HPLC/ESI-MS2 and individually quantified by HPLC-DAD. Five classes of polyphenols were present: flavan-3-ols, phenolic acids, flavonols, anthocyanins and simple phenolics (hydroquinones). The total phenolic content ranged between 0.1g/kg Fresh Weight (FW) ('Conference' cultivar) and 8.6g/kg FW ('Plant De Blanc' cultivar) in the flesh and between 1.6g/kg FW ('William vert' cultivar) and 40.4g/kg FW ('Arbi Chiheb' cultivar) in the peel. Procyanidins, analyzed after thioacidolysis, were the main phenolic compounds in all pear cultivars either in the pulp or the peel, their constitutive units being essentially (-)-epicatechin. Tunisian dessert pears and French perry pears are richer in procyanidins with very high degree of polymerization (>100) for Tunisian pears. Peel procyanidins were less polymerized (from 4 to 20). Pear peel phenolic profile was more complex especially for Tunisian cultivars, with flavonols and in some cultivars anthocyanins.

p-Hydroxyphenyl-pyranoanthocyanins: An Experimental and Theoretical Investigation of Their Acid-Base Properties and Molecular Interactions.

The physicochemical properties of the wine pigments catechyl-pyranomalvidin-3-O-glucoside (PA1) and guaiacyl-pyranomalvidin-3-O-glucoside (PA2) are extensively revisited using ultraviolet (UV)-visible spectroscopy, dynamic light scattering (DLS) and quantum chemistry density functional theory (DFT) calculations. In mildly acidic aqueous solution, each cationic pigment undergoes regioselective deprotonation to form a single neutral quinonoid base and water addition appears negligible. Above pH = 4, both PA1 and PA2 become prone to aggregation, which is manifested by the slow build-up of broad absorption bands at longer wavelengths (λ ≥ 600 nm), followed in the case of PA2 by precipitation. Some phenolic copigments are able to inhibit aggregation of pyranoanthocyanins (PAs), although at large copigment/PA molar ratios. Thus, chlorogenic acid can dissociate PA1 aggregates while catechin is inactive. With PA2, both chlorogenic acid and catechin are able to prevent precipitation but not self-association. Calculations confirmed that the noncovalent dimerization of PAs is stronger with the neutral base than with the cation and also stronger than π-π stacking of PAs to chlorogenic acid (copigmentation). For each type of complex, the most stable conformation could be obtained. Finally, PA1 can also bind hard metal ions such as Al3+ and Fe3+ and the corresponding chelates are less prone to self-association.

Chemically synthesized glycosides of hydroxylated flavylium ions as suitable models of anthocyanins: binding to iron ions and human serum albumin, antioxidant activity in model gastric conditions.

Polyhydroxylated flavylium ions, such as 3',4',7-trihydroxyflavylium chloride (P1) and its more water-soluble 7-O-ß-d-glucopyranoside (P2), are readily accessible by chemical synthesis and suitable models of natural anthocyanins in terms of color and species distribution in aqueous solution. Owing to their catechol B-ring, they rapidly bind FeIII, weakly interact with FeII and promote its autoxidation to FeIII. Both pigments inhibit heme-induced lipid peroxidation in mildly acidic conditions (a model of postprandial oxidative stress in the stomach), the colorless (chalcone) forms being more potent than the colored forms. Finally, P1 and P2 are moderate ligands of human serum albumin (HSA), their likely carrier in the blood circulation, with chalcones having a higher affinity for HSA than the corresponding colored forms.

Fruits, vegetables and their polyphenols protect dietary lipids from oxidation during gastric digestion.

Previous studies indicate that the ingestion of oxidized vegetable oils leads to the incorporation of chemically reactive molecules issued from the decomposition of the initial lipid hydroperoxides into lipoproteins. The aim of the present study is to investigate the oxidation of dietary lipids in the gastric compartment and their inhibition by plant polyphenols provided either as fruit and vegetables (F&V) or an extract. Six minipigs received a standard Western diet containing primarily sunflower oil, ground beef meat, and starch. Polyphenols in different matrix forms were ingested either as cubed F&V or as the corresponding hydroacetonic extract. Sampling of the gastric digesta allowed the kinetic investigation of pH, heme and non-heme iron forms, total lipids, lipid-derived conjugated dienes (CD) and TBARS. F&V and the corresponding polyphenol extract delayed the gastric digestion process as shown for total lipid and heme iron contents. This study also demonstrated the occurrence of in vivo oxidation of dietary lipids in the presence of meat iron. Interestingly, F&V played a protective role by totally inhibiting the accumulation of CD while largely decreasing the formation of TBARS. The polyphenol extract similarly slowed down the TBARS formation although it had no effect on the CD accumulation.

Reactivity of food phenols with iron and copper ions: binding, dioxygen activation and oxidation mechanisms.

In this work, the affinity of common dietary phenols (gallic acid, caffeic acid, catechin, and rutin) for iron and copper ions was quantitatively investigated in neutral phosphate buffer as well as the reactivity of the complexes toward dioxygen. Contrasting behaviors were observed: because of the competing phosphate ions, Fe(III) binding is much slower than Fe(II) binding, which is rapidly followed by autoxidation of Fe(II) into Fe(III). With both ions, O2 consumption and H2O2 production are modest and the phenolic ligands are only slowly oxidized. By contrast, metal-phenol binding is fast with both Cu(I) and Cu(II). With Cu(I), O2 consumption and H2O2 production are very significant and the phenolic ligands are rapidly oxidized into a complex mixture of oligomers. The corresponding mechanism with Cu(II) is hampered by the preliminary rate-determining step of Cu(II) reduction by the phenols. The consequences of these findings for the stability and antioxidant activity of plant phenols are discussed.

Pink discoloration of canned pears: role of procyanidin chemical depolymerization and procyanidin/cell wall interactions.

After canning, pear pieces turn occasionally from whitish-beige to pink. Conditions were set up to obtain this discoloration systematically and investigate its mechanism. Canned pears showed a significantly lower L* coordinate compared with fresh pears, and the L* coordinate of canned pears decreased with decreasing pH. The values of the a* and b* coordinates increased significantly after processing, the increase being greater for the more acidic pH values, with corresponding redder colors. After canning, polyphenol concentrations decreased significantly, mainly due to loss of procyanidins. This supported the hypothesis of conversion of procyanidins to anthocyanin-like compounds. However, no soluble product was detected at 520 nm, the characteristic wavelength of anthocyanins. When purified procyanidins were treated at 95 °C at three different pH values (2.7, 3.3, and 4.0), procyanidin concentrations decreased after treatment, the more so as the pH was lower, and a pinkish color also appeared, attributed to tannin-anthocyanidin pigment. The pink color was bound to cell walls. Extraction of the neoformed pink entities was attempted by successive solvent extractions followed by cell wall degrading enzymes. The pink color persisted in the residues, and canned pears gave significantly higher amounts of residues after solvent and enzyme treatments than fresh pears. Procyanidins were the entities responsible for the appearance of pink discoloration. However, it seems that this pink discoloration also involved the formation of strong, probably covalent, bonds to the cell wall.

Dietary iron-initiated lipid oxidation and its inhibition by polyphenols in gastric conditions.

The gastric tract may be the first site where food is exposed to postprandial oxidative stress and antioxidant activity by plant micronutrients. After food intake, dietary iron, dioxygen, and emulsified lipids come into close contact and lipid oxidation may take place. This study investigated lipid oxidation and its inhibition by dietary polyphenols in gastric-like conditions. Lipid oxidation induced by heme and nonheme iron was studied in acidic sunflower oil-in-water emulsions. The emulsifier type (bovine serum albumin, phospholipids), pH, and iron form were found to be factors governing the oxidation rates. Quercetin, rutin, and chlorogenic acid highly inhibited the metmyoglobin-initiated lipid oxidation in both emulsified systems at pH 5.8. Additionally, quercetin inhibited nonheme iron-initiated processes, while it was inefficient with hematin as an initiator. The presence of human gastric juice did not influence lipid oxidation, although it diminished the antioxidant activity of phenolics. Model emulsions may thus be valuable tools to study the gastric stability of polyunsaturated lipids.

The impact of industrial processing on health-beneficial tomato microconstituents.

The effect of industrial processing was investigated on the stability of tomato carotenoids, phenolic compounds and ascorbic acid. A deep insight in the processed products allowed the quantification of caffeic acid hexosides, which are far more important contributors than the well-known chlorogenic acid, dicaffeoylquinic acids and quercetin oligosaccharides (new feruloyl, sinapoyl and syringoyl derivatives of quercetin apiosylrhamnosylglucoside). (E)-ß-Carotene and (E)-lycopene were also quantified along with different mono- and di-(Z)-isomers of lycopene which were tentatively assigned. Processing of fresh tomato into paste had an overall positive effect on the contents in phenolic compounds, no effect on lycopene and a slight and high detrimental effect on ß-carotene and ascorbic acid, respectively. The balance between the increase in tomato matrix extractability and microconstituent catabolism was further observed in two contrasted transformations of paste into sauce. Overall, the nutritional quality of tomato-processed products, except for ascorbic acid, is mainly preserved through manufacture.

Food grade lingonberry extract: polyphenolic composition and in vivo protective effect against oxidative stress.

Fractionation of the polyphenols constituting a food grade lingonberry extract (Vaccinium vitis-idaea) highlighted a composition more complex than described until now in the berry. Procyanidins B1, B2, and A2 were identified by UPLC/ESI-MS(2) along with the presence of other flavanol oligomers. Processing induced the release of large amounts of aglycones for ferulic acid, p-coumaric acid, and quercetin. The described anthocyanic composition of lingonberry was completed with hexoside derivatives of peonidin, petunidin, malvidin, and delphinidin. Besides confirmation of in vitro antioxidant activity, in vivo study was performed on rats fed a diet inducing oxidative stress. Supplementation with lingonberry extract significantly decreased the total oxidant status and favorably affected antioxidant defense enzymes in red blood cells and liver. A drop in the serum reduced glutathione level was also prevented, and uric acid was maintained at low level, confirming the antioxidant activity of the extract (5% proanthocyanidins) from a dosage of 23 mg/kg of body weight.

Chemical synthesis of citrus flavanone glucuronides.

Flavanone glucuronides are the major phenolic metabolites detected in human plasma after consumption of citrus fruits. As such, they might display significant cardioprotective effects. In this work, glucuronides of naringenin (4'- and 7-O-beta-d-glucuronides) and hesperetin (3'- and 7-O-beta-d-glucuronides), the major flavanone aglycones in grapefruit and orange, respectively, have been chemically synthesized. On the one hand, the most reactive hydroxyl group, C7-OH, was protected by selective benzoylation to allow subsequent glucuronidation of C4'-OH (naringenin) or C3'-OH (hesperetin) (B-ring). On the other hand, the selective debenzoylation at C7-OH of the perbenzoylated flavanone aglycones allowed glucuronidation at the same position (A-ring). After careful deprotection, the target compounds were purified and characterized by nuclear magnetic resonance and mass spectrometry.

Olive phenols efficiently inhibit the oxidation of serum albumin-bound linoleic acid and butyrylcholine esterase.

BACKGROUND: Olive phenols are widely consumed in the Mediterranean diet and can be detected in human plasma. Here, the capacity of olive phenols and plasma metabolites to inhibit lipid and protein oxidations is investigated in two plasma models. METHODS: The accumulation of lipid oxidation products issued from the oxidation of linoleic acid bound to human serum albumin (HSA) by AAPH-derived peroxyl radicals is evaluated in the presence and absence of phenolic antioxidants. Phenol binding to HSA is addressed by quenching of the Trp214 fluorescence and displacement of probes (quercetin, dansylsarcosine and dansylamide). Next, the esterase activity of HSA-bound butyrylcholine esterase (BChE) is used as a marker of protein oxidative degradation. RESULTS: Hydroxytyrosol, oleuropein, caffeic and chlorogenic acids inhibit lipid peroxidation as well as HSA-bound BChE as efficiently as the potent flavonol quercetin. Hydroxycinnamic derivatives bind noncompetitively HSA subdomain IIA whereas no clear site could be identified for hydroxytyrosol derivatives. GENERAL SIGNIFICANCE: In both models, olive phenols and their metabolites are much more efficient inhibitors of lipid and protein oxidations compared to vitamins C and E. Low postprandial concentrations of olive phenols may help to preserve the integrity of functional proteins and delay the appearance of toxic lipid oxidation products.

Inhibition of the peroxidation of linoleic acid by the flavonoid quercetin within their complex with human serum albumin.

This work provides a quantitative kinetic analysis of oxidative pathways involving linoleic acid and the common dietary antioxidant quercetin (flavonoid), both bound to human serum albumin (HSA). In particular, it is shown that quercetin, although embedded in drug site I, is oxidized as quickly as free quercetin under a flux of hydrophilic peroxyl radicals. This observation suggests that efficient charge relays are established between the periphery of HSA and bound quercetin. Moreover, the peroxidation of HSA-bound linoleic acid is shown to take place at some specific fatty acid binding sites once one to two critical HSA residues are themselves oxidized. Quercetin efficiently delays the onset of lipid peroxidation. The inhibition persists long after the total consumption of quercetin, in agreement with some quercetin oxidation products exerting a residual antioxidant activity. Consistently, HSA markedly increases the maximal concentration of a two-electron oxidation product of quercetin that is accumulated and then consumed in the course of the peroxidation. The additional observation of the faster consumption of the single Trp residue in the presence of quercetin suggests that HSA enhances the antioxidant activity of quercetin by regenerating some of its oxidation products retaining a H-donating activity.

Flavonoids and their oxidation products protect efficiently albumin-bound linoleic acid in a model of plasma oxidation.

Although LDL esterified polyunsaturated fatty acids (PUFA) contribute largely to the pool of oxidizable lipids in plasma, they coexist with a non-negligible content of free PUFA. In some pathological conditions, the free PUFA/albumin ratio becomes abnormally elevated. Modeling was performed in a system constituted of linoleic acid bound to human serum albumin (HSA) in which oxidation was initiated by hydrophilic AAPH. Inhibition of lipid peroxidation was evaluated for various flavonoids. The accumulations of hydroperoxyoctadecadienoic acids (HPODE), hydroxyoctadecadienoic acids (HODE) and ketooctadecadienoic acids (KODE) were similarly inhibited: isoquercitrin>quercetin>catechin=isorhamnetin>>kaempferol>quercetin-4'-beta-D-glucoside=quercetin-3,4'-di-beta-D-glucoside. Surprisingly, quercetin and isorhamnetin afforded a protection to linoleic acid long after their consumption. Elucidation by mass spectrometry and NMR of the quercetin oxidation products and assessment of their antioxidant capacity pointed out that 3,4-dihydroxybenzoic acid and 2-(3,4-dihydroxybenzoyl)-2,4,6-trihydroxybenzofuran-3(2H)-one are major contributors to the apparent quercetin antioxidant capacity.

Regio- and stereoselective oxidation of linoleic acid bound to serum albumin: identification by ESI-mass spectrometry and NMR of the oxidation products.

An efficient RP-HPLC method was developed for the detection of the oxidation products derived from the AAPH-initiated peroxidation of linoleic acid bound to human serum albumin. Diode array UV-detection allowed the quantification at 234 nm of four regioisomeric hydroperoxyoctadecadienoic acids (HPODE) and four hydroxyoctadecadienoic acids (HODE) while at 280 nm four oxooctadecadienoic acid isomers (KODE) were detected. Full identification of the different underivatized HODE, HPODE and KODE isomers was achieved by negative ESI-mass spectrometry outlining common fragmentation pathways for 9- and 13-regioisomers. Chemical synthesis of 9-(E,Z)-, 9-(E,E)-, 13-(Z,E)- and 13-(E,E)-KODE helped to their structural characterization by 1H NMR. Lipid peroxidation in the presence of albumin proved to be regioselective with a larger accumulation of 13-HPODE and 9-KODE isomers. Thermodynamically more stable E,E-stereoisomers were also favored by albumin for both HPODE and KODE.

Quantitative kinetic analysis of hydrogen transfer reactions from dietary polyphenols to the DPPH radical.

Diphenylpicrylhydrazyl (DPPH) is widely used for quickly assessing the ability of polyphenols to transfer labile H atoms to radicals, a likely mechanism of antioxidant protection. This popular test generally pays no attention to the kinetics of H atom transfer, which however could be even more important than the total H-atom-donating capacities (stoichiometry, EC50) typically evaluated. In the present work, a series of dietary polyphenols belonging to the most representative families (flavonols from onion, flavanol monomers and oligomers from barley, and caffeic acid and caffeoyl esters from artichoke and endive) are characterized not only by their total stoichiometries (n(tot)) but also by their rate constants of first H atom abstraction by DPPH (k(1)), deduced from the kinetic analysis of the decay of the DPPH visible band following addition of the antioxidant. The mildly reactive DPPH radical allows a good discrimation between polyphenols, as demonstrated by the relatively large ranges of k(1) (ca. 400-5000 M(-)(1) s(-)(1)) and n(tot) (ca. 1-5) values typically measured with antioxidants having a single polyphenolic nucleus. With antioxidants displaying more than one polyphenolic nucleus (procyanidin oligomers, dicaffeoyl esters), the kinetic analysis makes it possible to demonstrate significant differences in reactivity between the subunits (two distinct k(1) values whose ratio lies in the range 3-10) and nonadditive stoichiometries.