The Impact of Cocoa Processing on Flavanol Content Absorption and Health Effects
Lead Research Organisation:
University of Reading
Department Name: Food and Nutritional Sciences
Abstract
Representing one of the most important lifestyle factors, diet can strongly influence the incidence of cardiovascular disease and thus a healthy diet is crucial for healthy ageing. Recent dietary intervention studies, in particular those using wine, tea and cocoa, have demonstrated beneficial effects on reducing cardiovascular disease risk, including an ability to lower blood pressure and to prevent blood vessel ageing. While such foods and beverages differ greatly in chemical composition, nutrient content and calories per serving, they have in common that they are amongst the major dietary sources of a group of plant compounds known as flavanols. The beneficial effects of flavanols have been attributed to their potential to be absorbed into the blood and to exert direct actions on blood vessels. Whilst flavanol-rich foods can be regarded as being protective against cardiovascular disease, the content of active flavanols is significantly reduced during industrial processing. This is because during heating and storage, the flavanols react with to varying degrees with sugars to form new products known as flavanol-Maillard conjugates. At present, information regarding the absorption of these new conjugates and whether they possess similar beneficial properties to native flavanols is lacking. This proposal is designed to address these questions and to unravel the significance of industrial food processing on their inherent health properties. On completion of the proposal, we will be in an excellent position to advise manufacturers of flavanol-rich foods on the best processing conditions required to produce foods with optimum beneficial cardiovascular effects. Building on existing human work conducted in our laboratory, this multidisciplinary study is designed to: 1) investigate the formation of flavanol-Maillard products during the processing of a common flavanol-rich food, namely cocoa; 2) determine the fate of these compounds in the human stomach, small intestine and large intestine; and 3) assess their ability to exert beneficial effects human cells. The first objective will inform us of the major flavanol-Maillard conjugates formed in the flavanol-rich foods during the processing, in this case heating of the cocoa beans during roasting. Although we will study cocoa, heat processing is relevant to many other flavanol rich-foods and therefore will have wider relevance. Secondly, we will examine the absorption of these conjugates, along with native flavanols by feeding processed cocoa to human and measuring them in blood. This will tell us whether heat processing reduces the absorption of native flavanols and also whether the flavanol-Maillard conjugates are absorbed by humans. We will also investigate their metabolism in the large intestine and whether they have a beneficial effect on the balance of the gut microflora, akin to changes seen with pre-biotic functional foods. Lastly, we will examine the beneficial effects of the conjugates in cellular models of human colon cancer and cardiovascular disease using state-of-the-art molecular techniques. We predict that this proposal will help determine the optimum industrial processing conditions required to generate flavanol-rich foods capable of exerting the strongest cardiovascular protection. It will inform both industry and the consumer and will help us develop future strategies to maximise flavanols in our foods. The proposal will broaden understanding of the role that diet plays in the prevention of cardiovascular disease and will help provide evidence for new and promising dietary strategies for tackling cardiovascular disease. The results of this study therefore have important implications for an ageing population where an improvement in healthy ageing is greatly desired. Moreover, the potential benefits in terms of quality of life are relevant to the population as a whole, as are the potential savings in health care costs.
Technical Summary
Dietary derived flavanols have been shown to reduce cardiovascular risk. These beneficial effects have been attributed to their absorption and actions on the vascular endothelium. However, food processing alters the flavanol content of foods due to interactions with Maillard reaction products, formed primarily during heating. This proposal is designed to unravel the significance of these reactions for the beneficial vascular effects of flavanol-rich foods. We will determine which flavanol-Maillard conjugates are generated from native flavanols and carbohydrate precursors under heat processing [150C; 30min or 120C; 5min], using state-of-the-art analytical analysis to characterise products, including specially developed HPLC protocols and LC-MS/MS. Major flavanol-Maillard derivatives identified will be synthesised. To assess human absorption and vascular function we will perform randomised, double blind, triple cross-over human intervention studies. These will use analytically well characterised foods: 1) standard high-flavanol, 2) heat processed high-flavanol; and 3) a low-flavanol. Blood samples will be collected at 0, 0.5, 1, 2 and 4 hr and flavanols and/or flavanol-Maillard conjugates will be measured by HPLC. Vascular function will be measured using state-of-the-art Laser Doppler imaging with iontophoresis. We will also investigate the absorption of individual flavanol-Maillard conjugates in cellular models of the human small intestine. The extent to which flavanol-Maillard conjugates are metabolised in the large intestine and their effects on the microflora will be performed using HPLC and fluorescent in situ hybridisation (FISH) with molecular probes targeting 16S rRNA, respectively. Finally the cellular activity of flavanol-Maillard conjugates will be assessed (using Western immunoblotting), in terms of their ability to: 1) inhibit the growth of colon adenocarcinoma cells and 2) to increase endothelial nitric oxide production by eNOS activation.
Publications
Mills CE
(2017)
Mediation of coffee-induced improvements in human vascular function by chlorogenic acids and its metabolites: Two randomized, controlled, crossover intervention trials.
in Clinical nutrition (Edinburgh, Scotland)
Lamport DJ
(2015)
The effect of flavanol-rich cocoa on cerebral perfusion in healthy older adults during conscious resting state: a placebo controlled, crossover, acute trial.
in Psychopharmacology
Rendeiro C
(2015)
The mechanisms of action of flavonoids in the brain: Direct versus indirect effects.
in Neurochemistry international
Rodriguez-Mateos A
(2015)
Influence of age on the absorption, metabolism, and excretion of cocoa flavanols in healthy subjects.
in Molecular nutrition & food research
Kean RJ
(2015)
Chronic consumption of flavanone-rich orange juice is associated with cognitive benefits: an 8-wk, randomized, double-blind, placebo-controlled trial in healthy older adults.
in The American journal of clinical nutrition
Mills CE
(2013)
The effect of processing on chlorogenic acid content of commercially available coffee.
in Food chemistry
Rodriguez-Mateos A
(2013)
Blueberry intervention improves vascular reactivity and lowers blood pressure in high-fat-, high-cholesterol-fed rats.
in The British journal of nutrition
Del Rio D
(2013)
Dietary (poly)phenolics in human health: structures, bioavailability, and evidence of protective effects against chronic diseases.
in Antioxidants & redox signaling
Spencer JP
(2012)
Neuroinflammation: modulation by flavonoids and mechanisms of action.
in Molecular aspects of medicine
Williams R
(2012)
Flavonoids, cognition, and dementia: Actions, mechanisms, and potential therapeutic utility for Alzheimer disease
in Free Radical Biology and Medicine
Gómez-Alonso S
(2012)
Inhibition of colon adenocarcinoma cell proliferation by flavonols is linked to a G2/M cell cycle block and reduction in cyclin D1 expression
in Food Chemistry
Rodriguez-Mateos A
(2012)
Procyanidin, anthocyanin, and chlorogenic acid contents of highbush and lowbush blueberries.
in Journal of agricultural and food chemistry
Vauzour D
(2011)
Potential Health Effects of Champagne Wine Consumption
in Journal of Wine Research
Spencer JP
(2010)
The impact of fruit flavonoids on memory and cognition.
in The British journal of nutrition
Vauzour D
(2010)
Sulforaphane protects cortical neurons against 5-S-cysteinyl-dopamine-induced toxicity through the activation of ERK1/2, Nrf-2 and the upregulation of detoxification enzymes.
in Molecular nutrition & food research
Vauzour D
(2010)
Polyphenols and human health: prevention of disease and mechanisms of action.
in Nutrients
Vauzour D
(2010)
Moderate Champagne consumption promotes an acute improvement in acute endothelial-independent vascular function in healthy human volunteers.
in The British journal of nutrition
Macready AL
(2010)
Cognitive tests used in chronic adult human randomised controlled trial micronutrient and phytochemical intervention studies.
in Nutrition research reviews
Spencer JP
(2010)
Beyond antioxidants: the cellular and molecular interactions of flavonoids and how these underpin their actions on the brain.
in The Proceedings of the Nutrition Society
Vauzour D
(2010)
Caffeic acid, tyrosol and p-coumaric acid are potent inhibitors of 5-S-cysteinyl-dopamine induced neurotoxicity.
in Archives of biochemistry and biophysics
Schroeter H
(2010)
Recommending flavanols and procyanidins for cardiovascular health: current knowledge and future needs.
in Molecular aspects of medicine
Spencer JP
(2009)
Flavonoids and cognition: the molecular mechanisms underlying their behavioural effects.
in Archives of biochemistry and biophysics
Spencer JP
(2009)
Flavonoids and brain health: multiple effects underpinned by common mechanisms.
in Genes & nutrition
Vafeiadou K
(2009)
The citrus flavanone naringenin inhibits inflammatory signalling in glial cells and protects against neuroinflammatory injury.
in Archives of biochemistry and biophysics
Description | Dietary flavanols are well reported to induce beneficial effects on the human vascular system and thus reduce the risk of cardiovascular disease. As such, there is interest in developing food products that are rich in these natural food components. Cocoa is an excellent source of flavanols and improves blood perfusion and lowers blood pressure on consumption. However, during industrial processing the levels of flavanols in cocoa are reduced, in part due to their reaction with carbohydrates present in the cocoa to form flavanol-Maillard conjugates. At present information regarding these processes and their effects on the beneficial properties of cocoa are unknown. Our proposal was designed to unravel the significance of industrial food processing on the inherent health properties of cocoa, in particular the roasting and alkalisation steps where the greatest degree of flavanols are thought to be lost. The major findings of our work were as follows: 1. Initially we studied the reactions of flavanols with cocoa carbohydrates under typical roasting and alkalizing conditions. Here, cocoa flavanols were found to react with sugar derivatives generated as a result of the Maillard reaction to form flavanol-Maillard complexes. In addition, flavanols were observed to yield novel flavanol-C-glycosides. High levels of flavanol-C-glycosides and lower amounts of flavanol-Maillard complexes were identified in commercial cocoa powders. 2. A randomized, controlled, human intervention trial was run in order to assess the impact of cocoa alkalisation on its inherent beneficial cardiovascular effects. Three cocoa powders were used which had been subjected to varying degrees of alkalization: 1) natural or non-alkalized cocoa (1740 mg flavanols); 2) medium alkalised cocoa (403 mg flavanols); and 3) heavily alkalized cocoa (1.3 mg flavanols). The impact of these cocoa powders on the vascular system was assessed in 10 healthy men by measuring physiologically significant increases in blood flow/perfusion (using Flow-Mediated Dilatation, a technique used by cardiologists to assess the global health of the vascular system). Our data indicated for the first time that both non-alkalised and medium alkalised cocoas induce a similar extent of vascular benefits at 2 and 4 h post ingestion. In contrast the heavily alkalized cocoa had no beneficial effects. 3. The 3 alkalised cocoas were also found to exert a positive effect on the growth of beneficial bacterial species, Bifidobacterium spp and Eubacterium rectal-C. coccoides. Furthermore, they inhibited of the growth of the pathogenic C. histolyticum group, which has been shown to contribute to the progression of colonic cancer and the onset of inflammatory bowel disease. The degree of cocoa processing did not alter these beneficial effects. These findings were supported by observations that cocoa roasting also had not effect on its positive influence on the gut microbiota. 4. Alkalised cocoa was observed to reduce the growth of human large intestinal adenocarcinoma cells, although to a lesser extent then that of the non-alkalised equivalent. These differences were found to be dependent on the ability of native flavanols to inhibit the phosphatidylinositol 3-kinase (PI3K)-AKT pathway, which plays critical role in cancer cell proliferation. Our data provide information that will allow industry to optimise levels of these beneficial micronutrients in foods. Overall, the data broaden our understanding of the role that food processing plays in inherent health benefits of foods containing flavanols and other phytochemicals. Indeed, although our data was collected using cocoa as a model food, the data have relevance to other food and beverages, including tea, coffee, fruit juices/smoothies and a variety of foods containing fruit and vegetables. Our data will help to deliver healthier foods to the consumer thus reducing disease risk and health care costs at the population level. |
Exploitation Route | This work has already led to updated and improved cocoa processing protocols in order to retain the maximum level of flavanols in products. |
Sectors | Agriculture Food and Drink Education Healthcare Pharmaceuticals and Medical Biotechnology |
Description | BBSRC research grant 2014 |
Amount | £534,820 (GBP) |
Funding ID | BB/M002802/1 |
Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
Sector | Public |
Country | United Kingdom |
Start | 01/2015 |
End | 12/2017 |
Description | EUropean Union 7th Framework research grant |
Amount | £320,000 (GBP) |
Organisation | European Research Council (ERC) |
Sector | Public |
Country | Belgium |
Start | 05/2010 |
End | 05/2012 |
Description | Mars Nutrition Research Council funding |
Amount | £86,000 (GBP) |
Organisation | Mars Nutrition Research Council, Mars Incorporated |
Sector | Private |
Country | United States |
Start | 07/2008 |
End | 06/2011 |
Description | Mars Symboscience |
Organisation | Mars Incorporated UK |
Department | Mars Symboscience |
Country | United States |
Sector | Private |
PI Contribution | R&D of new products based on the research |
Collaborator Contribution | Help with the synthesis of physiologically relevant metabolites of flavanols, through the use of their facilities. |
Impact | Manuscript and addtional funding |
Description | The effects of flavanols on brain and vascular health |
Organisation | Mars Incorporated UK |
Country | United Kingdom |
Sector | Private |
PI Contribution | Wrote successful grant application and hosted PhD student |
Collaborator Contribution | Supply of research intervention diets for human clinical trial (£60,000). Fully funded PhD studenship (stipend + fees + runing costs). Enhances my groups work in the flavonoid vascular area. |
Impact | Greatly expands the flavonoid brain and vascular research programme. |
Start Year | 2007 |
Description | 3rd International Conference on Polyphenols and Health |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Health professionals |
Results and Impact | International conference; 40 min oral presentation Press release and round table discussions. |
Year(s) Of Engagement Activity | 2009 |
Description | 3rd International Congress on Wine and Health |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Participants in your research and patient groups |
Results and Impact | International conference; 30 min oral presentation Press release |
Year(s) Of Engagement Activity | 2008 |
Description | 4th Internation Conference on Polyphenols and Health |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | 45 min Plenary lecture with questions and discussion afterwards, around 20 min Requests for publications and further work with Industry stimulated. |
Year(s) Of Engagement Activity | 2011 |
Description | 5th Internation Conference on Polyphenols and Health |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Professional Practitioners |
Results and Impact | 1h Plenary session with round table discussion (1h). Discussion was wide ranging and set targets for research over the next 2-5 years requests for Industry projects and manuscript requests |
Year(s) Of Engagement Activity | 2013 |
Description | Diet and Brain Health Film |
Form Of Engagement Activity | A magazine, newsletter or online publication |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | Recorded and film for the University of Reading regarding the research area. ~15 min and published on the Universities website Increased recruitment. Increased Industrial funding. |
Year(s) Of Engagement Activity | 2010 |
Description | Dissemination conference |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Public/other audiences |
Results and Impact | Public engagement event run by the Department of Food and Nutritional Sciences targeted primarily at the public but with literature reaching a wider audience. After session I received 18 emails requesting further information, publications and advice |
Year(s) Of Engagement Activity | 2013,2014 |
Description | Fourth European Nutrigenomics Conference |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Participants in your research and patient groups |
Results and Impact | European Conference; 20 min oral presentation Press release for conference and media interest |
Year(s) Of Engagement Activity | 2008 |
Description | Mars Nutrition Research Council |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Public/other audiences |
Results and Impact | Industrial Conferenece; 40 min oral presentation Funding directed towards this research in 2007, 2008 and 2009. |
Year(s) Of Engagement Activity | 2007 |
Description | Oxygen Club California |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Health professionals |
Results and Impact | International conference; 45 min oral presentation Press conference and discussion regarding future research focus in the area. |
Year(s) Of Engagement Activity | 2009 |
Description | Oxygen Club of California |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Participants in your research and patient groups |
Results and Impact | International conference; 30 min oral presentation. media interest in the USA |
Year(s) Of Engagement Activity | 2007 |
Description | Royal Pharmaceutical Society |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Health professionals |
Results and Impact | International conference; 45 min oral presentation Press release and media interviews |
Year(s) Of Engagement Activity | 2009 |
Description | Sixth Form Summer School |
Form Of Engagement Activity | Participation in an activity, workshop or similar |
Part Of Official Scheme? | No |
Geographic Reach | National |
Primary Audience | Schools |
Results and Impact | organisation and running of the laboratory sessions as part of the School of Food Biosciences 'Sixth Form Open Day'. This course gives A-level students a chance to experience the academic research environment. Experiments related to this MRC grant were used in order to show how certain nutrients may be beneficial to human brain health. June 2005, 2006, 2007 and 2008 Increased recruitment of students. |
Year(s) Of Engagement Activity |
Description | Society for Free Radical Research-International |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Health professionals |
Results and Impact | International conference; 30 min oral presentation Media interest |
Year(s) Of Engagement Activity | 2007 |
Description | TV Programme |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Media (as a channel to the public) |
Results and Impact | Part of a TV programme 'The Food Hospital". Filmed a piece relating to the MRC funded research. A number of schools have requested lay talks to pupils. |
Year(s) Of Engagement Activity | 2014 |
Description | The Nutrition Society |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Health professionals |
Results and Impact | International conference; 30 min oral presentation Press release and significant media interest. Collaborations made and grants applied for. |
Year(s) Of Engagement Activity | 2008 |
Description | The Times, The Observer, The Daily Mail and The Wine Spectator |
Form Of Engagement Activity | A magazine, newsletter or online publication |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Media (as a channel to the public) |
Results and Impact | Edited Interviews for publication, mainly 2007 and 2008 Interest from the general public and NHS |
Year(s) Of Engagement Activity | 2009 |
Description | The Wye Valley School |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | Local |
Primary Audience | Schools |
Results and Impact | Talk to 6th-form students about University level science and research. Increased recruitment and interest in science in general. |
Year(s) Of Engagement Activity | 2012 |
Description | Third European Nutrigenomics Conference |
Form Of Engagement Activity | A talk or presentation |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Health professionals |
Results and Impact | European conference; 20 min oral presentation Conference press release and media interest |
Year(s) Of Engagement Activity | 2007 |
Description | Website articles - Various |
Form Of Engagement Activity | A magazine, newsletter or online publication |
Part Of Official Scheme? | No |
Geographic Reach | International |
Primary Audience | Public/other audiences |
Results and Impact | Usually telephone interviews followed by publication of edited interview on public websites. Interest from the general public usually by email |
Year(s) Of Engagement Activity |