Functional foods for health promotion: state-of-the-science ondietary flavonoidsExtended abstracts from the 12th Annual Conference onFunctional Foods for Health Promotion, April 2009

Gary Williamson, Helmut Sies, David Heber, Carl L Keen, Ian A Macdonald, Lucas Actis-Gorreta,Tony Y Momma, Javier I Ottaviani, Roberta R Holt, Hagen Schroeter, and Christian Heiss

The extended abstracts in this report are based on presentations from the 12th

Special Conference on Functional Foods for Health Promotion, cosponsored by theNorth American branch of the International Life Sciences Institute (ILSI NorthAmerica) Project Committee on Flavonoids and the American Society for Nutrition atthe Experimental Biology meeting in April 2009. The theme of this year's specialconference was “State-of-the-Science on Dietary Flavonoids.” The conference beganwith a general introduction and overview of flavonoids and their presence in the dietas well as the estimated intake levels in the US population. Subsequentpresentations addressed issues pertaining to study design and interpretation,mechanisms of action, and the potential health impacts related to inflammation,the vasculature, and the brain. The present summary of the current science indicatesthat dietary flavonoids, particularly flavanols, show promising potential forreducing cardiovascular disease risk via reduction of inflammation andimprovement in vascular function. However, the existing data must be interpretedcautiously, with consideration given to the compound tested (i.e., parent ormetabolite), the use of controls, and the practicality of the concentrations used.While more data are needed on the long-term health impacts of dietary flavonoidsin humans, including the efficacious dose, current data indicate it may soon bepossible to develop public health messages about flavonoid-rich foods.nure_257 736..743

© 2009 International Life Sciences Institute

METHODOLOGICAL ISSUES IN DESIGN ANDINTERPRETATION OF STUDIES ON DIETARY

FLAVONOIDS AND HEALTH

Gary WilliamsonThe literature on flavonoids and health has expandedconsiderably over the last 2 decades, with a quarter of thepapers on vitamin/flavonoids and health published in

2008 being on flavonoids, compared to only 5% in 1988.This is coupled with increasing consumer interest in foodand in government-led campaigns encouraging the con-sumption of five portions of fruits or vegetables per day.Unfortunately, there is also a considerable amount of falseor misleading information in the press and on the Inter-net, which threatens to undermine the serious scientificadvances that have been made in this area. Painstaking

Affiliations: G Williamson is with the University of Leeds, Leeds, UK. H Sies is with the Heinrich-Heine University, Düsseldorf, Germany.D Heber is with the University of California, Los Angeles, California, USA. CL Keen, L Actis-Gorreta, TY Momma, Ji Ottaviani, and RR Holt arewith the Department of Nutrition at the University of California, Davis, California, USA. H Schroeter is with Mars Inc., McLean, VA, USA.C Heiss is with the Division of Cardiology, Vascular Medicine, and Pulmonology at Heinrich-Heine University, Düsseldorf, Germany.IA Macdonald is with the University of Nottingham Medical School, Nottingham, UK.

Correspondence: S Weiss, ILSI North America, 1156 Fifteenth Street, NW, Suite 200, Washington, DC 20005, USA. E-mail: sweiss@ilsi.org,Phone: +1-202-659-0074, Ext. 119, Fax: +1-202-659-3859.

Key words: cognition, diet, flavonoids, flavanols, functional foods, inflammation, mechanisms, methodology, vascular health

Meeting Report

doi:10.1111/j.1753-4887.2009.00257.xNutrition Reviews® Vol. 67(12):736–743736

resource-intensive long-term human studies are notcarried out overnight or cheaply. In the quest for fundingand for publication of high-impact papers, it is temptingto perform quick experiments, over-interpret them, andpublish the results as a health effect. Examples include invitro cellular systems treated with very high concentra-tions of flavonoids that far exceed the concentrationsfound in vivo. This leads to erroneous conclusions onhealth (and, indeed, in some cases, on toxicity). Under-standing the absorption and metabolism of dietary fla-vonoids is necessary before in vitro experiments can beinterpreted correctly and in order to design appropriateand successful in vivo intervention studies on biologicaleffects.

Knowledge of the mechanism of flavonoid absorp-tion and the amount of flavonoid absorbed has increasedtremendously in the last 2 decades. Although manyfactors affect bioavailability, such as the complexity of thefood matrix, the chemical form of the compound of inter-est, and the structure and amount of coingested com-pounds,1 it is possible to compare and contrast therelative absorption of different classes of flavonoids.2 Theprocess of absorption follows several steps, dependingon the chemical structure and on any other moietiesattached to the parent flavonoid molecule. Many fla-vonoids are glycosylated in food through a b-linkage toone or more residue(s) of glucose, arabinose, galactose, orrhamnose.3 The glucose and, to a certain extent, the ara-binose and galactose residues, are hydrolyzed by endog-enous enzymes, such as lactase, present in the brushborder of the small intestine enterocytes, and theflavonoid molecule (aglycone) is then absorbed by theintestine. However, when polyphenols are bound to arhamnose sugar, such as in rutin (quercetin-gluco-rhamnoside) or in hesperidin (hesperetin-gluco-rhamnoside), the brush border enzymes are not efficientat hydrolyzing the flavonoid-sugar bond and these mol-ecules continue into the gastrointestinal tract and reachthe colon, where microflora possess the enzymes neces-sary to hydrolyze the rhamnose moiety.4 At this stage, thepolyphenol is either absorbed as such, or is furthermetabolized by bacteria into lower-molecular-weightmetabolites, which are then absorbed by the colon. Somepolyphenols, especially catechins and procyanidins, arenot glycosylated, and so do not require deglycosylationbefore absorption. Within the enterocyte, most polyphe-nols are again conjugated but with glucuronide andsulfate moieties. The enterocytes have apical and basolat-eral transporters; the former are responsible for efflux(e.g., MRP2, P-glycoprotein) and the latter for absorption(e.g., MRP3, MRP1, and others as yet unknown).5 It isclear that the ratio between these two types of transport-ers would be predicted to be an important determinant ofbioavailability. A proportion of some flavonoids, e.g.,

galloylated catechins, phenolic acids, and isoflavones(partially) escape conjugation in the intestine, and conse-quently are found in both the conjugated and unconju-gated forms in the plasma, so that blood contains amixture of flavonoid forms resulting from intestinalmetabolism and from subsequent hepatic metabolism.6 Itshould be noted that the exact nature of flavonoid conju-gates in blood is only known for a limited number ofcompounds. Flavonoids are excreted either in the urine orthrough the bile, but the half-life varies considerably,between 2 and 20 h depending on the flavonoid. Under-standing of these processes has helped improve thedesign of in vitro experiments on cultured cells. Manyauthors now take the approach of using physiologicallyrelevant concentrations of flavonoids in molecular orcell-culture experiments, and some are using the conju-gated forms actually found in vivo.7,8 These approachesshould help advance the field, since the number of effectsobserved will decrease and only those which are physi-ologically relevant will be reported.

Acknowledgment

Declaration of interest. G Williamson is a part-timeemployee of the Nestle Research Center, Lausanne,Switzerland.

REFERENCES

1. Scholz S, Williamson G. Interactions affecting the bioavailabil-ity of dietary polyphenols in vivo. Int J Vitam Nutr Res.2007;77:224–235.

2. Manach C, Williamson G, Morand C, Scalbert A, Remesy C.Bioavailability and bioefficacy of polyphenols in humans. I.Review of 97 bioavailability studies. Am J Clin Nutr.2005;81(Suppl):S230–S242.

3. Rhodes MJ. Physiologically-active compounds in plant foods:an overview. Proc Nutr Soc. 1996;55:371–384.

4. Nielsen IL, Chee WS, Poulsen L, et al. Bioavailability is improvedby enzymatic modification of the citrus flavonoid hesperidin inhumans: a randomized, double-blind, crossover trial. J Nutr.2006;136:404–408.

5. Hayeshi R, Hilgendorf C, Artursson P, et al. Comparison of drugtransporter gene expression and functionality in Caco-2 cellsfrom 10 different laboratories. Eur J Pharm Sci. 2008;35:383–396.

6. Williamson G, Barron D, Shimoi K, Terao J. In vitro biologicalproperties of flavonoid conjugates found in vivo. Free RadicRes. 2005;39:457–469.

7. Tribolo S, Lodi F, Connor C, et al. Comparative effects of quer-cetin and its predominant human metabolites on adhesionmolecule expression in activated human vascular endothelialcells. Atherosclerosis. 2008;197:50–56.

8. Shirai M, Kawai Y, Yamanishi R, Kinoshita T, Chuman H, Terao J.Effect of a conjugated quercetin metabolite, quercetin3-glucuronide, on lipid hydroperoxide-dependent formationof reactive oxygen species in differentiated PC-12 cells. FreeRadic Res. 2006;40:1047–1053.

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HOW DO DIETARY FLAVONOIDS EXERT THEIRBIOLOGICAL EFFECTS?

Helmut SiesThe biological effects of dietary flavonoids are manifoldand short-term effects can be distinguished from long-term effects; their health implications are for inflamma-tion, heart disease, and cancer.1 Flavonoid actions havebeen grouped as follows: radical scavengers, metal chela-tors, antioxidants, and enzyme inhibitors. One crucialquestion considers the concentrations achieved at bio-logical target sites and another research area addressesmetabolism and transport: Are the parent compoundsthe biologically active compounds, or are they to be con-sidered “prodrugs,” giving rise to biologically activemetabolites?

Recent interest has focused on the flavonoid effectson the vascular endothelium.2 The following processesmodulate the in vivo efficacy of flavonoids: 1) rate andextent of absorption in the intestine as well as of entero-hepatic circulation; 2) metabolic conversions in intestinalcells, liver, and other cells or tissues; 3) binding toalbumin or cellular proteins; 4) plasma levels of aglyconesand conjugates; 5) accumulation in target cells, such asvascular endothelium; 6) urinary elimination rate; and 7)the molecular modes of action.

The dietary flavan-3-ol (-)-epicatechin, has beenidentified as a bioactive component in flavanol-richcocoa, which modulates endothelial vascular functionthrough elevation of bioactive nitric oxide in humans.The response of flow-mediated dilation (FMD) of thebrachial artery closely follows the time-course of the con-centration of (-)-epicatechin and its metabolites in bloodplasma.3,4 The peak response occurs at about 2 h andlevels off to return to baseline at 4–6 h. This may be calleda short-term effect. It has been proposed that endothelialNO metabolism rather than general antioxidant activity isthe major target of dietary flavanols and that NADPHoxidase activity is a crucial site of action.2

When flavanol-rich product is ingested repetitively,there is a change in the baseline FMD in the form of asustained increase, reaching a plateau at 5–8 days.5 Thismay be called a long-term effect, although longer termeffects at the level of months should be examined as well.Interestingly, the short-term response is maintained, i.e.,the FMD response subsequent to ingestion of an acutedose is unaltered but commences from a higher baselinefollowing several days of flavanol ingestion.5 The mecha-nism for the long-term effect involves changes in geneexpression and in protein patterns, with one examplebeing a decrease in arginase activity, which results in aug-mented substrate supply for NO synthase.

Similar studies were performed with otherflavonoid-containing foods and beverages. An early study

was conducted using black tea in patients with coronaryartery disease.6 A recent meta-analysis put the resultsobtained from several groups into perspective.7 Thus, vas-cular responses of conduit arteries, as well as in themicrocirculation in skin,8 have been identified as targetparameters of potential use as noninvasive functionalbiomarkers for vascular disease and its prevention. Fla-vonoids affect endothelial function not solely as antioxi-dants but also, and more importantly, as modulatorysignaling molecules; postprandial oxidative stress isattenuated when dietary flavonoids are supplied togetherwith a meal rich in oxidized lipids.9

Acknowledgment

Declaration of interest. H Sies has conducted research oncocoa flavanols that was supported by grants from MarsInc.

REFERENCES

1. Middleton E, Kandaswami C, Theoharides TC. The effects ofplant flavonoids on mammalian cells: implications for inflam-mation, heart disease and cancer. Pharmacol Rev.2000;52:673–751.

2. Schewe T, Steffen Y, Sies H. How do dietary flavanols improvevascular function? A position paper. Arch Biochem Biophys.2008;476:102–106.

3. Heiss C, Dejam A, Kleinbongard P, Schewe T, Sies H, Kelm M.Vascular effects of cocoa rich in flavan-3-ols. JAMA.2003;290:1030–1031.

4. Schroeter H, Heiss C, Balzer J, et al. (-)-Epicatechin mediatesbeneficial effects of flavanol-rich cocoa on vascular function inhumans. Proc Natl Acad Sci USA. 2006;103:1024–1029.

5. Heiss C, Finis D, Kleinbongard P, et al. Sustained increase inflow-mediated dilation after daily intake of high-flavanolcocoa drink over 1 week. J Cardiovasc Pharmacol. 2007;49:74–80.

6. Duffy SJ, Keaney JF Jr, Holbrook M, et al. Short- and long-termblack tea consumption reverses endothelial dysfunction inpatients with coronary artery disease. Circulation.2001;104:151–156.

7. Hooper L, Kroon PA, Rimm EB, et al. Flavonoids, flavonoid-richfoods, and cardiovascular risk: a meta-analysis of randomizedcotrolled trials. Am J Clin Nutr. 2008;88:38–50.

8. Heinrich U, Neukam K, Tronnier H, Sies H, Stahl W. Long-termingestion of high flavanol cocoa provides photoprotectionagainst UV-induced erythema and improves skin condition inwomen. J Nutr. 2006;136:1565–1569.

9. Sies H, Stahl W, Sevanian A. Nutritional, dietary and postpran-dial oxidative stress. J Nutr. 2005;135:969–972.

FLAVONOIDS AND INFLAMMATION

David HeberFlavonoids are widely accepted to be the predominantpolyphenols in our diet and they typically have an average

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molecular weight of approximately 300 Daltons.1

However, larger polyphenolic compounds (>1000molecular weight) are also ubiquitous in our diet yet theiramounts have been underestimated, in part due to thelack of authentic chemical standards. These compoundsinclude tannins, which are divided into two groups: 1)proanthocyanidins, or condensed tannins, found in greentea, cacao/chocolate, blueberries, cranberries, grape seed,etc.; and 2) hydrolyzable tannins found in pomegranate,strawberries, raspberries, some nuts (peels of walnuts,almonds, pecans), and oak-aged beverages. Flavonoidspossess antioxidative and radical scavenging activities.They also regulate cellular activities of the inflammation-related cells including mast cells, macrophages, lympho-cytes, and neutrophils.

Flavonoids can modulate the activities of arachidonicacid metabolizing enzymes such as phospholipase A2,cyclooxygenase, lipoxygenase, and inducible nitric oxidefound in macrophages.2–5 Inhibition of these enzymes byflavonoids reduces the production of arachidonic acid,prostaglandins, leukotrienes, and peroxynitrates, whichare crucial mediators of inflammation.

Inflammation is classically defined by calor (heat),rubor (redness), and tumor (swelling), but the role oflow-grade chronic inflammation signaled by increasedlevels of C-reactive protein has been identified as anindependent risk factor for heart disease. The associa-tion of low-grade inflammation with abdominal obesityand reduced intake of flavonoids in the diet has spurredinterest in dietary modification for the prevention ofchronic diseases. The association between the moderndiet and low-grade inflammation is multifactorial andincludes the observation that omega-3 fatty acid intakeis reduced along with the pro-inflammatory and pro-oxidant activities of refined carbohydrates, such as fruc-tose. Although not fully understood, several mechanismsof action are proposed to explain the in vivo anti-inflammatory effects of flavonoids, including inhibitionof eicosanoid-generating enzymes, such as phospholi-pase A2, cyclooxygenases, and lipoxygenases, therebyreducing the concentrations of prostanoids and leuko-trienes. Recent studies have also shown that certainflavonoids, especially flavone derivatives, express theiranti-inflammatory activity, at least in part, by the modu-lation of proinflammatory gene expression, such ascyclooxygenase-2, inducible nitric oxide synthase, andnumerous cytokines and chemokines.2 The inhibition ofenzyme activities, which are increased by these variousdietary factors, by flavonoids is an important cellularmechanism balancing excess inflammation. However,flavonoids also modulate proinflammatory gene expres-sion through inhibition of the activation and action ofnuclear factor-kappa B (NF-kB), a protein that binds tospecific promoter regions on the genome stimulating

the production of numerous mediators of inflammation.This factor remains inactive in the cytoplasm untilphosphorylation of inhibitory kappaB-alpha (IkB-alpha)leads to its release of the p65/p50 dimer of NF-kBwhich binds to nuclear DNA leading to gene acti-vation of multiple mediators of inflammation. Normally,this response is inactivated rapidly, but in chronicinflammatory conditions, including in some cancer cells,this response continues to be activated. Several labora-tories have demonstrated inhibition of IkB-alphaphosphorylation and inhibition of binding of the p65/p50 dimer to its promoter region of the genome asmechanisms by which flavonoids can inhibit excessiveinflammation.

The flavonoids are metabolized by the liver and byintestinal flora to yield smaller compounds such as thephenolic acids from green tea.6–8 Using a bacterial prepa-ration imitating the normal human colonic flora, it ispossible to demonstrate the conversion of larger polyphe-nols to phenolic acids. These phenolic acids, in turn, dem-onstrate similar anti-inflammatory effects as the parentcompounds. Therefore, the impact of flavonoids oninflammation occurs via direct effects and those of fla-vonoid metabolites.

Chronic inflammation is increasingly shown to beinvolved in the onset and development of several patho-logical disturbances such as arteriosclerosis, obesity, dia-betes, neurodegenerative diseases, and even cancer.Given that many drugs with anti-inflammatory actionshave side-effects that make them unsuitable for primaryprevention, there is an urgent need to develop infor-mation on safe dietary modifications that can takeadvantage of naturally occurring anti-inflammatorycompounds. Flavonoids belong to a group of naturalsubstances occurring normally in the diet that exhibita variety of beneficial effects on health. The anti-inflammatory properties of flavonoids suggest they havepotential utility for reducing the risk of inflammatorydiseases related to the modern diet. Several mechanismsof action have been proposed to explain the in vivo anti-inflammatory actions of flavonoids, such as anti-oxidantactivity, inhibition of eicosanoid-generating enzymes, ormodulation of the production of proinflammatorymolecules. Recent studies have also shown that someflavonoids are modulators of proinflammatory geneexpression, thus leading to attenuation of the inflamma-tory response. However, much work remains to be donein order to achieve definitive conclusions about thepotential of flavonoids in disease prevention. This briefsummary has outlined the known mechanisms involvedin the anti-inflammatory activity of flavonoids and theimplications of these effects for protecting against age-related chronic diseases including cancer and cardiovas-cular disease.

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Acknowledgment

Declaration of interest. D Heber has received researchfunding from the California Strawberry Commission andfrom POM Wonderful Inc.

REFERENCES

1. Beecher GR. Overview of dietary flavonoids: nomenclature,occurrence and intake. J Nutr. 2003;133:3248S–3254S.

2. Kim HP, Park H, Son KH, Chang HW, Kang SS. Biochemicalpharmacology of biflavonoids: implications for anti-inflammatory action. Arch Pharm Res. 2008;31:265–273.

3. O’Leary KA, de Pascual-Tereasa S, Needs PW, Bao YP,O’Brien NM, Williamson G. Effect of flavonoids and vitamin Eon cyclooxygenase-2 (COX-2) transcription. Mutat Res.2004;551:245–254.

4. Sakata K, Hirose Y, Qiao Z, Tanaka T, Mori H. Inhibition of induc-ible isoforms of cyclooxygenase and nitric oxide synthase byflavonoid hesperidin in mouse macrophage cell line. CancerLett. 2003;199:139–145.

5. Cho SY, Park SJ, Kwon MJ, et al. Quercetin suppresses proin-flammatory cytokines production through MAP kinases andNF-kappaB pathway in lipopolysaccharide-stimulated mac-rophage. Mol Cell Biochem. 2003;243:153–160.

6. Henning SM, Choo JJ, Heber D. Nongallated compared withgallated flavan-3-ols in green and black tea are more bioavail-able. J Nutr. 2008;138:1529S–1534S.

7. Gao K, Xu A, Krul C, et al. Of the major phenolic acids formedduring human microbial fermentation of tea, citrus, and soyflavonoid supplements, only 3,4-dihydroxyphenylacetic acidhas antiproliferative activity. J Nutr. 2006;136:52–57.

8. Henning SM, Niu Y, Lee NH, et al. Bioavailability and antioxi-dant activity of tea flavanols after consumption of green tea,black tea, or a green tea extract supplement. Am J Clin Nutr.2004;80:1558–1564.

IMPACT OF FLAVONOIDS ON VASCULAR REACTIVITYAND VASCULAR HEALTH

Carl L Keen, Lucas Actis-Gorreta, Tony Y Momma, Javier IOttaviani, Roberta R Holt, Hagen Schroeter, ChristianHeissCurrent dietary guidelines recommend a diet rich inplant foods. Fruits, vegetables, and nuts have beenstrongly associated with a decreased risk of cardiovascu-lar disease (CVD); however, causal mechanisms under-lying the putative positive effects of fruits and vegetablesfor CVD risk have yet to be firmly elucidated.1 Thesefoods provide not only a number of essential vitaminsand minerals, but also diverse phytonutrients, such asflavonoids, which have been of increasing interest due totheir chemical diversity and range of potential biologicaleffects. Indeed, we are just beginning to determine whichcompounds, and potential foods, drive the putative posi-tive cardiovascular health effects of plant foods. It mustbe noted that the search for the biologically active com-

ponents of plant foods is complicated by the fact thatagricultural practices and food processing methods canaffect the type, concentration, and bioactivities ofnumerous plant constituents.

With respect to flavonoids, epidemiological studiessupport the concept that regular consumption of dietsrich in these nutrients is associated with a reduced risk forvascular disease.2 During the last decade, there has beenincreasing interest in the concept that flavanols, whichrepresent a class of the flavonoid family, may be particu-larly important in this relationship. Individuals and cul-tural groups who consume high amounts of flavanol-richfoods have been reported to have reduced risk for CVDmortality.3,4 It has been speculated that the putative posi-tive health effects of flavanols can be attributed tomultiple mechanisms, including flavanol-inducedchanges in oxidant defense, platelet reactivity, and vascu-lar function.2

Cardiovascular disease is a leading cause of morbid-ity and mortality in most countries. The initiation andprogression of atherosclerotic vascular disease is multi-factorial in nature and is currently viewed as a chronicinflammatory disease that is initiated, and promoted, by anumber of cell types, including leukocytes and platelets,which lead to a damaged and dysfunctional endothelium.Consumption of flavanol-rich foods has been reportedto influence platelet reactivity as well as fibrinolysis. Inhuman subjects, platelet-primary hemostasis (as mea-sured by the PFA-100) was found to be reduced as soon as2 h following consumption of a cocoa drink containing900 mg of total flavanol, including the flavanols epicat-echin and catechin, and their oligomers (procyanidins).At the same time, there was reduced expression on theplatelet surface of glycoprotein IIb/IIIa and P-selectin,which mediate platelet-platelet and platelet-leukocyteinteractions, respectively.5 Isolated specific compoundsfound in flavanol-rich foods have been reported to beeffective in reducing platelet reactivity and the overallthrombotic state. For example, dimer B1 was reported toreduce platelet reactivity 2 h post consumption in humanvolunteers. In these individuals, there was a significantreduction in PAI-1 activity and an increase in fibrin-D-dimer activity, indicating an increase in overall fibrin-olytic activity (Actis-Goretta et al., unpublished data).

A number of studies have shown that flavanol-richfoods can provide acute, as well as chronic, improvementsin vascular reactivity in both healthy and at-riskpopulations.6–8 Illustrative of this, within 2 h of consump-tion of a flavanol-rich cocoa that provided 917 mg offlavonoids, there was a significant improvement in flow-mediated dilation (FMD) in healthy subjects; this effectwas not seen in subjects who were given a cocoa drinkthat was similar in composition to the high-flavanolcocoa but which contained only 37 mg of flavanols.

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Blockade of nitric oxide (NO) synthase eliminated thisputative flavanol-mediated response.8 High flavanolintakes can provide a significant increase in baselineFMD that can be observed after 3 days of continuouscocoa consumption, an effect that can be sustained for aperiod of 1 week.7 The above studies were performed inrelatively healthy individuals; an important question is ifthese types of vascular effects also occur in individualswith established CVD. To address this issue, work hasbeen conducted with fully medicated diabetic patients.Results to date suggest there are similar positive flavanoleffects in these individuals.5 Importantly, the vascularimprovements observed in diabetic subjects were main-tained for up to 1 month when the subjects consumed thehigh-flavanol beverages on a daily basis.5 It is importantto note that the studies above use FMD as a prognosticallyvalidated measure of vascular reactivity. FMD measuresthe dilation of the brachial artery as an assessment ofoverall CVD risk, but it does not directly measure theresponse of vessels that cause disease, i.e., the coronaryarteries. Recently, high-flavanol chocolate was reported toimprove coronary vasodilatation in heart-transplantrecipients,3 suggesting that flavanols may be effective inthe coronary circulation.

Assuming that high-flavanol cocoa has been estab-lished as a food that is effective for improving acute vas-cular function, significant questions remain concerningthe identity of this complex food’s bioactive compo-nent(s) that contribute to the improved FMD response, aswell as the mechanism(s) of action of these compo-nent(s). While cocoa can be high in numerous flavanolsand procyanidins, it is particularly rich in the flavanolepicatechin. Isolated epicatechin provided to human sub-jects at a dose similar to that provided in the 900-mgcocoa drinks described above resulted in an FMDresponse that was similar in magnitude to that obtainedwith the complete cocoa beverage.8 However, severalresearchers have shown that following its absorption, epi-catechin is rapidly metabolized to a number of differentmetabolite products.5,8 The identity of the metabolite(s)responsible for the above FMD effect is an issue ofongoing debate. In addition to instigating an improvedFMD response, an increase in NO availability has beenobserved that can be correlated to circulating flavanolmetabolites.8 It has been reported that the flavanolmetabolites, as well as cocoa consumption, can reduce,through unknown mechanisms, arginase activity, therebyproviding another possible mechanism for enhanced NOavailability.9 In summary, there is extensive evidence forthe concept that certain flavanols can alter plasma NOpools and influence endothelial health; the identity of thespecific metabolite(s) that drive these effects, and themechanisms underlying them, are areas of activeresearch.

The above clinical work is consistent with epidemio-logical findings of associations between the consumptionof high-flavanol diets and a reduced risk for vasculardisease. While additional studies in this area are clearlyneeded, it could be argued that for healthy individuals,incorporating into the diet fruits and vegetables that arehigh in flavanols would help in reducing vascular diseaserisk. In addition, health professionals should begin toconsider the potential value of flavanol-rich foods andbeverages for patients who are either at risk for, or alreadyhave, vascular disease. Importantly, the utilization ofmethods, such as FMD, could be applied as a means todemonstrate the efficacy of diet in the treatment and pre-vention of vascular disease in high-risk individuals. Wesuggest this could represent a breakthrough in the dietarymanagement of many patients, as it could provide ameans of rapid feedback as to the positive effects ofdietary modification.

Acknowledgments

Funding. This body of work was supported, in part, bygrants from Mars Inc.

Declaration of interest. The research group has receivedfunding from Mars Inc. CL Keen has worked as a con-sultant for Mars Inc. H Schroeter is employed by Mars,Inc. The remaining authors have no relevant interests todeclare.

REFERENCES

1. Dauchet L, Amouyel P, Hercberg S, Dallongeville J. Fruit andvegetable consumption and risk of coronary heart disease: ameta-analysis of cohort studies. J Nutr. 2006;136:2588–2593.

2. Erdman Jr. JW, Balentine D, Arab L, et al. Flavonoids and hearthealth: Proceeding of the ILSI North America Flavonoids Work-shop, May 31–June 1, 2005, Washington, DC. J Nutr.2007;137(Suppl):S1–S20.

3. Corti R, Flammer AJ, Hollenberg NK, Luscher TF. Cocoa andcardiovascular health. Circulation. 2009;119:1433–1441.

4. Mink PJ, Scrafford CG, Barraj LM, et al. Flavonoid intake andcardiovascular disease mortality: a prospective study in post-menopausal women. Am J Clin Nutr. 2007;85:895–909.

5. Holt RR, Actis-Goretta L, Momma T, Keen CL. Dietary flavanolsand platelet reactivity. J Cardiovasc Pharmacol.2006;47(Suppl):S187–S196.

6. Balzer J, Rassaf T, Heiss C, et al. Sustained benefits in vascularfunction through flavanol-containing cocoa in medicated dia-betic patients a double-masked, randomized, controlled trial. JAm Coll Cardiol. 2008;51:2141–2149.

7. Heiss C, Finis D, Kleinbongard P, et al. Sustained increase inflow-mediated dilation after daily intake of high-flavanolcocoa drink over 1 week. J Cardio Pharmacol. 2007;49:74–80.

8. Schroeter H, Heiss C, Balzer J, et al. (-)-Epicatechin mediatesbeneficial effects of flavanol-rich cocoa on vascular function inhumans. Proc Nat Acad Sci USA. 2006;103:1024–1029.

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9. Schnorr O, Brossette T, Momma TY, et al. Cocoa flavanols lowervascular arginase activity in human endothelial cells in vitroand in erythrocytes in vivo. Arch Biochem Biophys.2008;476:211–215.

FLAVONOIDS AND THE BRAIN

Ian A MacdonaldThere is now a substantial body of epidemiological evi-dence showing that diets rich in fruits and vegetablesconfer health benefits. It appears that the flavonoidcontent of fruits and vegetables (including green tea,cocoa, and berry fruits) convey at least some of thisbenefit, both for promoting cardiovascular health and forpotentially sustaining cognitive function during ageing.Increased neuronal activity requires an increase in bloodflow to support the metabolic requirements, whilsthuman ageing is associated with a decline in brain bloodflow. Thus, there is substantial interest in compounds,including components of food, which might increasebrain blood flow and thus improve brain (particularlycognitive) function in normal ageing as well as in clinicalconditions such as dementia. This presentation examinedthe experimental evidence for potential beneficial effectsof flavonoids, both in animal models of cognitive decline,and in human studies of brain blood flow and brainfunction.

Hartman et al.1 showed in a transgenic mouse modelof Alzheimer’s disease that providing pomegranate juice(which is rich in phenolics and flavonoids) for 6.5 monthsimproved the animals’ abilities to learn a complex taskcompared to the control mice. This improved learningand retention of memory was at least partly due toincreased swimming speed in the mice treated withpomegranate juice. Similar benefits were observed whenepigallocatechin gallate (EGCG- one of the major fla-vonoids in green tea) was provided to the same type oftransgenic mice in a different study by Rezai-Zade et al.2

The beneficial effects were more noticeable when theEGCG was administered via intraperitoneal (i.p.) injec-tion, rather than in the drinking water, which may be dueto poor bioavailability of the EGCG when given orally tothese mice. However, it was also noticeable that the degreeof cognitive impairment seen in the transgenic mice wasrather variable between groups, being much moremarked in the mice which served as the comparatorgroup for the i.p. injection than in the mice receivingEGCG in the drinking water.

Feeding a purified plant-derived flavanol(-epicatechin) increases the angiogenic response in thehippocampus and enhances memory retention in micesubjected to a learning and memory task, as shown in astudy by van Praag et al.3 That study assessed the effects of

both epicatechin and regular exercise on learning andmemory retention and the most marked positive effectwas noticed for the combination of exercise and epicat-echin. A study of aged rats, performed by Williams et al.,4

showed that provision of freeze-dried blueberries in thediet improved the performance of a number of memorytasks. Whilst this effect could be due to a number ofcompounds found in the blueberries, it was found thatbrain levels of flavanols were increased to a greater extentthan the levels of anthocyanins.

The demonstration that -epicatechin and blueberrieshave effects on learning and memory has led to interest inthe possibility that flavanols and other flavonoids couldincrease brain blood flow and improve memory inhuman subjects. It has previously been shown that con-sumption of cocoa flavanols for 5 days enhances the func-tional magnetic resonance imaging (fMRI) response to acomplex cognitive task in healthy young women.5 Theseyoung subjects had good initial cognitive function, whichwas not altered by the cocoa flavanols. However, theenhanced fMRI response is indicative of an increasedbrain blood flow response to the task, and this occurred inareas of the brain associated with attention and switchinginto a response suppression mode (right dorsolateral pre-frontal cortex), attention allocation (parietal cortex), andwith processing response conflict and overcoming priorresponse suppression (anterior cingulated cortex).However, such studies have not yet been performed inconditions of cognitive impairment, so it is not knownwhether flavanols will confer benefits in such individuals.We did show in a pilot study of healthy young womenthat total brain grey matter blood flow (measured by arte-rial spin-labeling and MRI) was increased by a single,large dose of cocoa flavanols5 and Sorond et al.6 showedthat consumption of a drink with a high cocoa flavanolcontent for 14 days was associated with an increase inbrain blood flow in older individuals, as assessed withtrans-cranial Doppler ultrasound. Clearly, it will be ofmajor interest to confirm these observations in the elderlyand to assess whether they are accompanied by benefitssuch as improved cognitive function.

Acknowledgment

Declaration of interest. IA Macdonald serves on the advi-sory boards of Mars Europe and Coca-Cola Europe. Hehas received funding from Mars Inc. and Unilever.

REFERENCES

1. Hartman RE, Shah A, Fagan AM, et al. Pomegranate juicedecreases amyloid load and improves behavior in a mousemodel of Alzheimer’s disease. Neurobiol Dis. 2006;24:506–515.

2. Rezai-Zadeh K, Arendash GW, Hou H, et al. Green teaepigallocatechin-3-gallate (EGCG) reduces beta-amyloid

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mediated cognitive impairment and modulates tau pathologyin Alzheimer transgenic mice. Brain Res. 2008;1214:177–187.

3. van Praag H, Lucero MJ, Yeo GW, et al. Plant-derived flavanol(–)epicatechin enhances angiogenesis and retention of spatialmemory in mice. J Neurosci. 2007;27:5869–5878.

4. Williams CM, El Mohsen MA, Vauzour D, et al. Blueberry-induced changes in spatial working memory correlate withchanges in hippocampal CREB phosphorylation and brain-derived neurotrophic factor (BDNF) levels. Free Radic Biol Med.2008;45:295–305.

5. Francis ST, Head K, Morris PG, Macdonald IA. The effect offlavanol-rich cocoa on the fMRI response to a cognitive task inhealthy young people. J Cardiovasc Pharmacol. 2006;47:S215–S220.

6. Sorond FA, Lipsitz LA, Hollenberg NK, Fisher NDL. Cerebralblood flow response to flavanol-rich cocoa in healthy elderlyhumans. Neuropsychiatr Dis Treat. 2008;4:433–440.

Acknowledgment

ILSI North America is a public, not-for-profit, scientificfoundation that advances the understanding and applica-

tion of scientific issues related to the nutritional qualityand safety of the food supply. It carries out its mission bysponsoring relevant research programs, professional edu-cation programs and workshops, seminars and publi-cations, as well as providing a neutral forum forgovernment, academic, and industry scientists to discussand resolve scientific issues of common concern for thewell-being of the general public. ILSI North America’sprograms are supported primarily by its industry mem-bership. For more information about ILSI NorthAmerica, visit the website at www.ilsina.org, or send ane-mail to ilsina@ilsi.org.

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