Monomeric flavanols or catechins (flavan-3-ols), biosynthetic precursors of proanthocyanidins, are characterized for having a C₆-C₃-C₆ skeleton with a hydroxyl group in position three of the C-ring (Figure 1). Flavan-3-ols represent the largest class of monomeric C₆-C₃-C₆ flavanols. The two compounds catechin and epicatechin are among the commonest flavonoids known, sharing a distribution almost as widespread as the related flavonol, quercetin [1,2]. Catechin (C), epicatechin (EC), gallocatechin (EC), epigallocatechin (EGC) and their galloyl substituted derivatives (ECG, and EGCG) are usually found in plant-derived foods and food products. Catechins are rarely found in nature in their glycosylated form, unlike anthocyanidins which are commonly found as anthocyanins. On the other hand, flavanols are commonly found in plant-derived food products in their polymerized forms as oligomers (dimers to pentamers) or polymers (six or more units). The most common oligomers are the B series, B1 to B8, formed by two flavanol units, either catechin or epicatechin, joined by a C4–C8 linkage (B1 to B4) [4→8 bond (epicatechin-(4β→8)-catechin); Figure 2], or C4–C6 linkage (B5 to B8). The least frequent dimers are the A series, characterized by the presence of double linkages between the two “catechin” units, one C4–C8 or C4–C6 and an additional one between C2 and C5 or C7 [1,2].

Type-A proanthocyanidins (Figure 3) are less common in food plants, however their presence in peanuts and almond skins and in some berries has been described [9–12]. In general, flavanols are widely distributed in plant foods.

Flavanols are mainly present in fruits and derived products like fruit juices or jams; also in tea, cocoa, and cereals (Table 1). They are however almost non-existent in vegetables and legumes, with the notable exception of lentils and broad beans [12,13]. In many cases flavanols are present in the peels or seeds of fruits and vegetables, being discarded when eaten or during processing, and therefore their dietary intake is limited.

Consumption of total catechins and dimeric procyanidins has been estimated to be between 18 and 50 mg per day in the Spanish population [26] and 50 mg (s.d. 56 mg/day) catechins per day in a nationwide dietary survey among 6,200 Dutch men and women aged 1–97 years [27]. Catechin intake increased with age, and the intake was higher in women (60 mg/day) than in men (40 mg/day) [27]. However it has to be taken into consideration that the bioavailability of flavanols varies depending on the degree of polymerization and the existence of galloyl residues in the molecule.

The food sources of catechins (tea, chocolate, apples, pears, grapes and red wine) are very popular and highly consumed. The usual sources of dietary catechins in the studies of bioavailability of flavanols with human volunteers are cocoa or tea [28]. After ingestion of chocolate, representing doses of 46, 92, and 138 mg of unmetabolized catechin, the maximal plasma concentrations (C_max) of 0.13, 0.26, and 0.36 μmol/L, respectively, were reached in 2 h (T_max) [26]. Similarly, 220 mg of epicatechin (EC) from chocolate ingestion reached C_max of 4.77 μmol/L also in 2 hours, with urinary excretion (as a % of intake) of 29.8% [28]. Tea catechins (mainly constituted by epigallocatechin and epigallocatechin-3-gallate) at a total dose of 240 mg gave a 0.5 μmol/L C_max with a T_max that, depending on the specific metabolites, ranged from 1.6 to 2.3 hours, with a mean level of excretion of 8.1 (% of intake), but with clear differences between catechins (28.5%) and gallocatechins (11.4%) [29].

Bioavailability differs markedly among catechins, and galloylation of catechins also reduces their absorption. The wide nature of procyanidins and flavan-3-ols affects their bioavailability since they can be detected at different degrees of polymerisation and acylation (with gallic acid). The galloylation of flavan-3-ols reduces their absorption; polymerised procyanidins are not absorbed and small amounts of procyanidin dimers B1 and B2 have been detected in plasma [30–32]. Galloylated catechins are rapidly eliminated, probably not by degalloylation, which has been shown to be a minor process in humans, but rather by preferential excretion of these compounds in bile [27,28]. In all cases, catechin and gallocatechin were found in urine excreted as methylated and/or sulphated or glucuronidated forms. We should note that, as stated by Stalmach et al. [29], there is still a controversy regarding the bioavailability of catechins, mainly due to a lack of a standardised methodology for both the processing of plasma samples and the analytical determination of the metabolites in the processed extracts.

The maximum concentration of metabolites of flavan-3-ols in plasma varied between 0.03 and 2.7 μmol/L (after a 50 mg aglycone equivalent supply) and the urinary excretion was 0.1–55%. The high interval in the absorption and urinary excretion was due to the wide variability in the nature of flavan-3-ols [28].

Epigallocatechin gallate is the only known polyphenol present in plasma in large proportions in a free from. The flavan-3-ols are highly methylated and glucuronidated in the 3′ and 4′ positions (catechin and epicatechin metabolites mainly) [33].

Procyanidin intake in the United States, excluding monomers, has been estimated to be 53.6 mg per day [34]. Very few studies have shown absorption of procyanidin dimers in humans [32–37] and none has shown, so far, absorption of higher polymerized compounds in humans. However certain studies have reported the absorption of procyanidins, up to trimers, in rats [38,39], showing that in the case of flavanol dimers and trimers, no glucuronidation or sulphatation take place.

The flavanols is a very complex group of polyphenols ranging from the monomeric flavan-3-ols (catechin and epicatechin; gallocatechin; and epigallocatechin and the corresponding gallate esters) to polymeric procyanidins known as condensed tannins including a whole range of oligomeric intermediates going from dimers up to undecamers and dodecamers and then polymers, mainly provided by fruits, tea and wine. The main products or metabolites detected in urine as a result of the bacterial metabolism after the intake of catechin and epicatechin are 3-hydroxyphenylpropionic acid, δ-(3,4,-dihydroxyphenyl)-γ-valerolactone; δ-(3-hydroxyphenyl)-γ–valerolactone; 3-hydroxyhippuric acid (from both bacterial and human metabolism) [40–43].

After in vitro incubation of epicatechin with human intestinal bacteria the metabolites pyrogallol, 3′,4′-dihydroxyphenylvaleric acid, 3′-hydroxyphenylvaleric acid, 3,4-dihydroxyphenylpropionic-, 3′-hydroxyphenylpropionic-, 3′-methoxyphenylvaleric acid and 2″,3″-dihydroxyphenoxyl-3′,4′-dihydroxyphenylpropionic acid were detected. However, epicatechin gallate and epigallocatechin gallate were not degraded by the microbiota [44].

The quantitative importance and biological activities of the microbial metabolites have seldom been examined in vivo by measuring the microbial metabolites formed. In four groups of rats fed for 8 days a diet supplemented with 0.12 g/100 g catechin, 0.25 or 0.5 g/100 g red wine powder containing proanthocyanidins, phenolic acids, flavanols, anthocyanins and flavonols or an unsupplemented diet the main metabolites from the catechin diet were 3-hydroxyphenylpropionic acid, 3-hydroxybenzoic acid and 3-hydroxyhippuric acid. Their total urinary excretion accounted for 4.7 g/100 g of the catechins ingested and that of intact catechins for 45.3 g/100 g. For wine polyphenols, the same microbial metabolites as observed for the catechin diet were identified in urine, along with hippuric, p-coumaric, vanillic, 4-hydroxybenzoic and 3-hydroxyphenylacetic acid. All together, these aromatic acids accounted for 9.2 g/100 g of the total wine polyphenols ingested and intact catechins for only 1.2 g/100 g. The higher excretion of aromatic acids by rats fed wine polyphenols is likely due to their poor absorption in the proximal part of the gut. Since microbial metabolites still bear a reducing phenolic group and should also prevent oxidative stress in inner tissues, these microbial metabolites and their biological properties help explain why the health effects of polyphenols deserve more attention [45].

Studies on the microbial metabolism of dietary condensed tannins (oligomeric and polymeric procyanidins) showed that benzoic, phenylacetic, phenylpropionic and phenyllactic acid derivatives, as well as phloroglucinol, δ-(3-hydroxyphenyl)-γ-valerolactone, and 1-(3-hydroxyphenyl)-3-(2,4,6-trihydroxyphenyl) propan-2-ol were produced in rat cecal models [46]. In another study, condensed tannins produced 4-hydroxy-phenylacetic, 3-phenylpropionic, 3-hydroxyphenylacetic, 4-hydroxyphenylpropionic, 3-hydroxyphenylvaleric and 3-hydroxyphenyl propionic [41], while valerolactones were not detected.

As instestinal metabolism strongly influences the bioavailability of flavonoids, the investigated microbial deconjugation and degradation of the most common flavan-3-ols using the pig cecum in vitro model system developed by Van’t Slot and Humpf [47], showed that the microbial degradation of (+)-/(−)-catechin, (−)-epicatechin, (−)-gallocatechin, (−)-epigallocatechin, (−)-gallocatechin gallate, (−)-epigallocatechin gallate, procyanidin B2, and gallic acid, under anaerobic physiological conditions in single incubation experiments and as a mixture, was complete within 4–8 h. No difference was observed for catechin enantiomers. In addition to monomeric flavonoids, procyanidins were also metabolized by the intestinal microbiota, as shown for procyanidin B2. The hydroxylated phenolcarboxylic acids produced were similar for all the tested substances [47].

Anthocyanins are natural pigments responsible for the blue, purple, red and orange colors of many fruits and vegetables. More than 500 different anthocyanins have been described in the literature. Anthocyanins are food bioactive compounds with a double interest, one technological, due to their impact on the sensorial characteristics of food products, and the other for their health related properties throw different biological activities, one of them being their implication on cardiovascular disease risk protection [48].

Anthocyanins are mainly present in nature in the form of heterosides. The aglycone form of the anthocyanins, so-called anthocyanidin, is structurally based on the flavilium or 2-phenyl-benzopyrilium cation, with hydroxyl and methoxyl groups present at different positions of the basic structure (Figure 4). Depending on the number and position of the hydroxyl and methoxyl groups as substituents, different anthocyanins have been described, and six of them are commonly found in fruits and vegetables: pelargonidin, cyanidin, delphinidin, petunidin, peonidin and malvidin [2,3,13].

Anthocyanidins are unstable to light and are water-insoluble so that they do not occur usually in their free state. Instead, they are present in the cell vacuole linked to sugars, which provide stability and water solubility. These glycosides are called anthocyanins. Some time ago, the known glycosidic variation among these pigments was restricted to four main types: 3-monoglycosides, 3-diglycosides, 3,5-diglycosides and 3-diglycoside-5-monoglycosides. The most common sugar present was glucose, but rhamnose, xylose and galactose were also encountered. More recent research has revealed many more complex structures. There are many anthocyanins with acyl substituents linked to sugars, aliphatic acids (such as malonic, succinic, malic and acetic acid), cinnamic acids (such as p-coumaric, ferulic, or sinapic acid), and pigments with both aliphatic and aromatic substituents. A further complexity in some anthocyanins results from the presence of several acylated sugars in the structure. These anthocyanins have sometimes been designated as polyglycosides [2,3,13,49].

The main dietary sources of anthocyanins are red fruits (Table 2), like berries and red grapes, red wine, cereals and purple corn, as well as some vegetables such as red cabbage [48–73]. Daily consumption of total anthocyanins has been estimated to be between 3 and 215 mg/day [55,71,76,77]. Since dietary anthocyanins are restricted to berries, red fruits and red wine it should be assumed that there are variations between individuals due to differences in the intake of these anthocyanin-rich products. However, the influence of methodological differences in the assessment, as well as nutritional, social and cultural differences of the investigated populations may also explain the wide range of anthocyanin consumption estimated by different authors [28,48,49].

Until very recently, anthocyanins were believed to have a very low bioavailability. Clinical studies carried out with humans consuming different types of fruit containing anthocyanins have shown that these phenolic pigments are poorly absorbed [78,79]. The concentration of anthocyanins in plasma ranged 10–50 nmol/L (after a 50 mg of aglycone equivalent supply) and the T_max was 1.5 h. At intakes of 188–3,570 mg total cyanidin glycosides, the C_max were in the range of 2.3–96 nmol/L [71,81]. The urinary excretions were found to be in the range between 0.018% and 0.37 % of the intake. The time of appearance in the plasma was consistent with absorption at the stomach (24%) but also at the small intestine (23%) [74,82–84]. Maximal urinary excretion is usually achieved in less than 4 h. Nevertheless, some studies revealed that the bioavailability of anthocyanins was underestimated since all of their metabolites might not have been yet identified [28].

Anthocyanins are some of the few polyphenols that can be detected in plasma in the native form found in plant foods (glycosides). However, they can also suffer glucuronidation and sulphatation reactions as glucuronide and sulphate conjugates were detected in urine in several studies. Kay and co-workers [85] have also identified an oxidized derivative of anthocyanins. Methylation at the 3′-hydroxyl is also very common in anthocyanins [82,86]. The ratio of metabolites present in plasma or urine varied depending on the experiment, type of food administered, sample conservation, extraction procedures as well as the sensitivity and resolution of the HPLC methods used (6% [87], 25% [88], 68% [85], or 85% [82]).

Besides the usual organs where the metabolization and elimination of the anthocyanins takes place, like liver or kidney, these polyphenols have also been detected, in the course of animal studies, in brain regions that mediate cognitive behaviour (cortex, hippocampus), and eye [74,90,91].

Only a small part of the dietary anthocyanins are absorbed as such or as hydrolysis products in which the sugar moiety is removed. Thus, large amounts of the ingested compounds are likely to enter the colon. The studies looking at the colonic microbiota metabolism of anthocyanins are scarce. Using in vitro models, the microbial deglycosylation and degradation of six anthocyanins exhibiting three different aglycones with mono- or di-β-d-glycosidic bonds were investigated using HPLC-DAD and GC-MS detection [92]. All the anthocyanidin glycosides detected were hydrolysed by the microflora within 20 min and 2 h of incubation, depending on the sugar moiety. Due to the high instability of the liberated aglycones at neutral pH, primary phenolic degradation products were already detected after 20 min of incubation. Further metabolism of the phenolic acids was accompanied by demethylation. Because of their higher chemical and microbial stability, phenolic acids and/or other anthocyanin metabolites might be mainly responsible for the observed antioxidant activities and other physiological effects in vivo [92]. Cyanidin 3-rutinoside is first transformed into the corresponding glucoside and then to the aglycone. This is reported to be achieved by gut microbiota, and the aglycones, which are chemically unstable, are converted to phenolic acids that then can be further metabolized by the gut microbiota. The anthocyanin nucleus is broken down and protocatechuic (3,4-dihydroxybenzoic acid) acid is detected as a product of human colonic microbiota [92,93]. Other metabolites produced in vitro by the pig gut microbiota include syringic acid (3,5-dimethoxy-4-hydroxybenzoic acid), vanillic acid (3-methoxy-4-hydroxybenzoic acid), phloroglucinol aldehyde (2,4,6-trihydroxybenzaldehyde), phloroglucinol acid (2,4,6-trihydroxybenzoic acid), and gallic acid (3,4,5-trihydroxybenzoic acid), depending on the chemical structure of the anthocyanins [92]. In the case of methylated anthocyanins, the ring-cleavage degradation products can be de-methylated by the pig gut microbiota. This shows that anthocyanins as being flavonoid compounds, also suffer the ring cleavage, to release the B-ring and A-ring derived products [74].

More recently, it has been shown that anthocyanins may be metabolised by intestinal microflora [94,95], or simply being chemically degraded [85], producing a set of new products that have not yet been totally identified, but that include the phenolic acids: gallic acid, protocatechuic acid, syringic acid, vanillic acid and phloroglucinol. Looking only to the anthocyanin glucuronides and sulphates, the existent studies on anthocyanin bioavailability in humans have shown that for a total anthocyanin intake of 0.05–1.9 g, the corresponding C_max of 0.001–0.2 μM was reached (T_max) in a time range of 0.5–4 hours after ingestion of the respective dose, with a urinary excretion that in no case exceeded 5% of the ingested dose [96].

An investigation on the promotion of an optimal platelet function via the reduction of platelet hyper-reactivity using dietary solutions, assayed the anti-platelet activity of physiologically relevant concentrations of the anthocyanins delphinidin-3-O-rutinoside, cyanidin-3-O-glucoside, cyanidin-3-O-rutinoside, and malvidin-3-O-glucoside, and their putative colonic metabolites, dihydroferulic acid, 3-(3-hydroxyphenyl)propionic acid, 3-hydroxyphenylacetic acid and 3-methoxy-4-hydroxypheylacetic acid, both separately and in combination. Anti-thrombotic properties were exhibited by 10 μmol/L dihydroferulic acid, and 3-(3-hydroxyphenyl)propionic acid, 1 μmol/L delphinidin-3-O-rutinoside, and a mixture of all the tested compounds [124]. Anthocyanins also inhibit the thrombin receptor activating peptide (TRAP)-induced platelet aggregation but did not influence platelet reactivity when faced with strong agonists such as collagen and ADP [124].

Anthocyanins are capable of acting on different cells involved in the development of atherosclerosis. The chemokine monocyte chemotactic protein 1 (MCP-1) is known to mediate in the recruitment of macrophages to sites of infection or inflammation, and direct involvement of MCP-1 on atherogenesis has been established. Anthocyanins have been shown to have a protective effect against TNF-α induced MCP-1 secretion in primary human endothelial cells [125].

Vascular endothelial growth factor (VEGF) is a major pro-angiogenic and pro-atherosclerotic factor. Anthocyanins, specifically delphinidin and cyanidin, have been shown to prevent the expression of VEGF stimulated by platelet derived growth factor(AB) (PDGF(AB) in vascular smooth muscle cells by preventing activation of p38 mitogen-activated protein kinases (p38 MAPK) and c-Jun N-terminal kinase (JNK) [126].

Anthocyanin extracts from chokeberry, bilberry, and elderberry have shown endothelium-dependent relaxation capacity in porcine coronary arteries [127]. Moreover, chronic ingestion of anthocyanins increased cardiac glutathione concentrations in rats [128]. In a recent study, rats treated with isoproterenol to induce postinfarction remodeling were fed with an alcohol-free red wine showing a protective effect on hearts by repressing hypertrophy-associated increased phosphorylation of protein kinase C (PKC) α/β II and by activating Akt/protein kinase B (Akt) [129].

Anthocyanins have an effect on cholesterol distribution, protecting endothelial cells from CD40-induced proinflammatory signalling [130].

Pelargonidin inhibits iNOS protein and mRNA expression as well as the NO production in a dose-dependent manner in macrophages exposed to an inflammatory stimulus (lipopolysaccharide, LPS). Pelargonidin also inhibits the activation of NF-κB, which is a significant transcription factor for iNOS [131]. In macrophages, blackberry anthocyanins inhibit LPS induced nitric oxide biosynthesis [132].

Ischemia/reperfusion injury leads to irreplaceable myocyte cell necrosis and apoptosis with concomitant activation of signal transducers and activators of transcription 1 (STAT1) and STAT3. It has been shown that the anthocyanin delphinidin decreases the extent of both necrotic and apoptotic cell death in cultured cardiomyocytes and reduces infarct size after ischemia in rats and that both effects are mediated by inhibition of STAT1 activation [133]. In rats, a long-term dietary intake of corn-derived anthocyanins, mainly cyanidin and pelargonidin based, made the myocardium less susceptible to ischemia-reperfusion injury ex vivo as well as in vivo when comparing it with a control consisting of anthocyanin-free corn. However results from human trials are controversial. Recently Curtis et al. [134], showed no effect on biomarkers of cardiovascular disease (CVD), including inflammatory biomarkers, platelet reactivity, lipids, and glucose; and liver and kidney function, as well as anthropometric measures, blood pressure, and pulse after a 12-week intervention with 500 mg cyanidin in postmenopausal women.

Many phenolic compunds are potent effectors of biologic processes and have the capacity to influence disease risk via several complementary and overlapping mechanisms. The current knowledge on mechanisms by which dietary phenolic compounds play a role in preventing degenerative pathologies was summarized recently [8]. In particular, the complex interactions between these dietary molecules and their molecular targets including the cell signaling pathways and response: NF-κB signaling pathway, Activator protein 1(AP-1), the Phase II enzyme activation and Nrf, and the mitogen-activated protein kinase (MAPK) signaling pathway. Future studies should focus on commonly consumed anthocyanins, in order to examine dose-response effects, and be of long enough duration to allow assessment of clinically relevant endpoints [111].