1. #1 Whole wheat potential for health Unlocking the bioactive ingredients in whole wheat with novel technologies developed in HEALTHGRAIN Jan A. Delcour Katholieke Universiteit Leuven, Leuven Belgium Porto, September 24 2010 EU integrated project: Exploiting bioactivity of European cereal grains for improved nutrition and health benefits

2. #2 Take Home Messages Wheat grain contains interesting nutrients such as fibre and numerous phytonutrients which can provide health benefits. The HEALTHGRAIN diversity screen shows large variation in bioactive components which can be exploited by wheat breeders. Novel technologies (innovative milling, dry fractionation and enzyme technologies) can increase safety and ‘unlock’ physiologically active fractions from wheat grains. These include soluble arabinoxylan oligosaccharides (AXOS), included in the definition of dietary fibre. A major feature of AXOS is colonic fermentation to short chain fatty acids, butyric acid in particular (fuel for colon mucosa cells).

3. #3 Background State of the matter:  Strong evidence of whole grain health benefits is mainly based on epidemiology (food consumption of many persons (e.g. 100.000) is correlated with illness and causes of death, during many years)  Little insight in the mechanisms of health benefits, and the importance of individual components and/or combinations of compounds/ synergistic effects  There should and can be more major possibilities for healthy bread, pasta and other cereal grain based products than just whole grain ….. using grain based raw materials, fractions, components, and production processes

4. #4 Testa (1%)  Alkylresorcinols Aleurone layer (6-9%) Starchy endosperm (80-85%) Starch & Protein Lipids Antioxidants Vitamin E B vitamins Minerals Plant sterols Enzymes Soluble & insoluble dietary fibre (arabinoxylans) Proteins Antioxidants (phenolic acids) Vitamin E B vitamins Minerals Phytic acid Enzymes Germ (3%) Insoluble dietary fibre (arabinoxylans, cellulose, lignin) Antioxidants (phenolic acids) Crease Brush Inner - and Outer pericarp (4-5%) Bran WHEAT KERNEL – Important components are present in minor fractions

5. #5 WHOLE WHEAT FOOD BIOACTIVITY Whole meal vs. white flour: 2.5 – 5 x higher levels Phytosterols Cholesterol-lowering Alkylresorcinols Enzyme inhibitors Phytate Antioxidant Vitamin E antioxidant Folate Choline, betaine Methyl donors Magnesium, chromium Improved insulin sensitivity Ferulic acid Antioxidant Oligo- Saccharides Prebiotic Tocopherols & - trienols Antioxidants, reduce LDL oxidation Lignans Phytoestrogens Beneficial physiological effects in humans Synergistic effects Dietary fibre Improved intestinal function, and glucose and lipid metabolism

6. #6 HEALTHGRAIN - STRATEGIC OBJECTIVE HEALTHGRAIN, an Integrated Project of EU’s 6 th Framework Programme aimed at providing the scientific basis for improving the well-being, and reducing the risk of metabolic syndrome diseases in Europe by increasing the intake of protective grain components. The aim was to exploit European grain, and new ingredients thereof, in development of tailored products with improved health benefits.

7. #7 CONSUMER RESEARCH  Consumer expectations DISSEMINATION AND TECHNOLOGY TRANSFER Breeders, food industry, trade, consumer organisations, authorities  Web-pages, e-learning, courses, newsletters, workshops GRAIN IMPROVEMENT AND BIOTECHNOLOGY TOOLKIT  Tools for molecular breeding TECHNOLOGY AND PROCESSING  Technology for new products and ingredients NUTRITION AND METABOLISM  Physiological responses and health benefits HEALTHGRAIN PROJECT STRUCTURE Integrated project: ’from fork to farm’- 5 Modules INDUSTRIAL PLATFORM NUTRITION INFORMATION NETWORK CONSUMER COMMUNICATION PANEL

8. #8 CONSUMER RESEARCH DISSEMINATION AND TECHNOLOGY TRANSFER Breeders, food industry, trade, consumer organisations, authorities GRAIN IMPROVEMENT AND BIOTECHNOLOGY TOOLKIT (module leader Peter Shewry) Identification of sources of natural variation in European wheat cultivars (in comparison with other grains); dietary fibre, starch, lignans, alkylresorcinols, phenolic acids, folate, tocopherols and tocotrienols and sterols. THE HEALTHGRAIN DIVERSITY SCREEN Generation of new variation especially in starch and arabinoxylan Development of ”OMICS” technologies, and a ”Biotechnology Toolkit” to assist breeders NUTRITION AND METABOLISM  INDUSTRIAL PLATFORM NUTRITION INFORMATION NETWORK CONSUMER COMMUNICATION PANEL Grain improvement = higher levels of fibre and other health promoting compounds

9. #9 IMPROVED VARIETIES DEVELOP NOVEL TOOLS FOR BREEDERS PLANT BREEDING DETERMINE NATURAL VARIATION IN BIOACTIVE COMPONENTS GENERATE NEW VARIATION- mutants & transgenics How to increase bioactive compounds in wheat

10. #10 Trials Martonvasar, Hungary 150 wheat varieties + 50 other cereal grains (rye, oat, barley, spelt) Follow-up: 26 selected wheat and 5 rye varieties grown in England, France, Hungary, Poland THE HEALTHGRAIN DIVERSITY SCREEN Determining natural variation in levels of bioactive compounds

11. #11 Phenolic Acids Alkylresorcinols Tocols Folate Sterols Flour Total Arabinoxylan Flour Soluble Arabinoxylan 670-959 μg/g dm 241-677 μg/g dm 325-1171 μg/g dm 325-774 ng/g dm 0.29-1.38% 1.36-2.74%27-80 μg/g dm Concentration ranges of bioactive components across 150 bread wheats grown on one site The HEALTHGRAIN diversity screen shows large variation in bioactive components in wheat which can be exploited by breeders

12. #12 DIVERSITY SCREEN – CONCLUSIONS 1. There is no relationship between the contents of bioactive components and the age or geographical origin of the varieties 2. It is possible to combine high levels of phytochemicals and dietary fibre components with good yield and processing properties 3. Fibre levels (Arabinoxylans, β-glucans): genotype most important Environmental effects (2007) : UK: fibre ↑, Hungary: fibre ↓ 4. Environmental factors important for levels of choline and betaine 5.Near infrared spectroscopic methods sufficiently sensitive to predict Total arabinoxylan, Soluble arabinoxylan, Sterols (cholesterol lowering), Alkylresorcinols (markers of whole grain content).

13. #13 DIVERSITY SCREEN - FOLLOW-UP 1.Findings can be applied now – buy wheat varieties with high levels of desired components *e.g. fibres, anti-oxidants 2.Generation of new varieties: 7 to 10 years 3.Results of 200 varieties/ (grown in Martonvasar): 11 papers) in special issue of J. Agr. Food Chem. Volume 56, 2008. 4.Results of 26 wheat + 5 rye lines 4 locations: 10 papers in special issue J. Agr. Food Chem. 2010 ________________________________________________________ HEALTHGRAIN Methods Handbook : Analysis of Bioactive Components in Small Grain Cereals. Edited by Peter R. Shewry and Jane L. Ward (2009). Published by AACC International.

14. #14 Take Home Messages Wheat grain contains interesting nutrients such as fibre and numerous phytonutrients which can provide health benefits The HEALTHGRAIN diversity screen shows large variation in bioactive components which can be exploited by wheat breeders Novel technologies (innovative milling, dry fractionation and enzyme technologies) can increase safety and ‘unlock’ physiologically active fractions from wheat grains These include soluble arabinoxylan oligosaccharides (AXOS), included in the definition of dietary fibre. A major feature of AXOS is colonic fermentation to short chain fatty acids, butyric acid in particular (fuel for colon mucosa cells).

15. #15 CONSUMER RESEARCH DISSEMINATION AND TECHNOLOGY TRANSFER Breeders, food industry, trade, consumer organisations, authorities GRAIN IMPROVEMENT AND BIOTECHNOLOGY TOOLKIT TECHNOLOGY AND PROCESSING NUTRITION AND METABOLISM INDUSTRIAL PLATFORM NUTRITION INFORMATION NETWORK CONSUMER COMMUNICATION PANEL TECHNOLOGY AND PROCESSING (Module leader Jan Delcour) New dry and wet fractionation of wheat and rye to produce specific well characterised fractions containing high levels of dietary fibre and phytochemicals Production of functional non- digestible cereal carbohydrates Technology and processing methods to deliver cereal foods with high levels of outer grain layers and good sensory quality

16. #16 Testa (1%)  Alkylresorcinols Aleurone layer (6-9%) Starchy endosperm (80-85%) Starch & Protein Lipids Antioxidants Vitamin E B vitamins Minerals Plant sterols Enzymes Soluble & insoluble dietary fibre (xylans,  -glucans) Proteins Antioxidants (phenolic acids) Vitamin E B vitamins Minerals Phytic acid Enzymes Asparagine Germ (3%) Insoluble dietary fibre (xylans, cellulose, lignin) Antioxidants (phenolic acids) Contaminants pesti-/ herbicides mycotoxins, ‘microbials’ Crease Brush Inner - and Outer pericarp (4-5%) Bran THE WHEAT GRAIN KERNEL

17. Wheat Bran’ Bran Abraded Wheat Flour Bran Fractions Bran Extracts (AXOS) Milling Fermented Bran (New) Products Wheatgrain based raw materials, fractions, components, and production processes provide major possibilities for healthy bread, pasta and other cereal grain based products

18. Pericarp Seed coat Aleurone Starchy endosperm = Flour Technical Bran = Pericarp, Seed coat, and Aleurone Courtesy Karin Autio, VTT

19. #19 AX rich materials Flour Whole meal AXOS with specific structural features Bread containing AXOS with specific structural features AXOS with documented health effects/claims AXOS containing bread with documented health effects/claims Xylanase technology Functional carbohydrates: the AXOS concept. prebiotics originating from wheat / grain  From arabinoxylan (AX) rich source to AX prebiotic oligosaccharides (AXOS) - promoting health promoting intestinal bacterial flora  Use the compounds present in grains and xylanase enzymes instead of adding prebiotics

20. #20 From AX to prebiotic AXOS 5 O O O OH OH 5 4 3 2 1 A O OH OH OH O O O OH O 5 4 3 2 1 1 2 3 4 B D 5 4 3 2 1 5 1 2 3 4 5 O OH OH OH O O OH OH OH O O O O 1 2 3 4 5 4 3 2 1 5 O OH OH O OH O O O O O OH H 3 CO 1 2 3 4 C

21. #21 From AX to prebiotic AXOS Wheat bran AXOS Wheat bran Destarched, deproteinised wheat bran Wheat bran AXOS extract Amylase Protease Filtration Xylanase Filtration Concentration Spray drying J. Sci. Food Agric. 2006, 86, 1722

22. #22 From AX to prebiotic AXOS 2. colon transversum Data from U. Gent, Labmet Simulator of the Human Intestinal Microbial Ecosystem 1. colon ascendens

23. #23 AXOS can stimulate intestinal microflora in tranverse colon to produce benefical short chain fatty acids (SCFA) Data from U. Gent, Labmet colon Transversum (AXOS main effect) ‘In vitro’ Simulator of the Human Intestinal Microbial Ecosystem colon Ascendens (inulin main effect)

24. #24  WP 4.1 IN VITRO MODELS Product characteristics, release features and bioavailability of components associated with ”whole grain” - studies using in-vitro models simulating the human gastrointestinal tract, and cell-based bioassays WP 4.2 ANIMAL MODELS Bioavailability and metabolic effects of bioactive components associated with ”whole grain” - studies using animal models WP 4.3 MEAL STUDIES IN HUMANS Bioavailability and acute metabolic effects of components associated with ”whole grain” – acute and semi-acute meal studies in healthy and ”at risk” subjects WP 4.4 DIETARY INTERVENTIONS Longer-term effects of whole-grain and associated product features (GI) on metabolic risk factors and weight regulation - studies in “at risk” subjects and in type 2 diabetics Module 4 NUTRITION AND METABOLISM

25. #25 Testa (1%)  Alkylresorcinols Aleurone layer (6-9%) Starchy endosperm (80-85%) Starch & Protein Lipids Antioxidants Vitamin E B vitamins Minerals Plant sterols Enzymes Soluble & insoluble dietary fibre (arabinoxylans,  -glucans) Proteins Antioxidants (phenolic acids) Vitamin E B vitamins Minerals Phytic acid Enzymes Germ (3%) Insoluble dietary fibre (arabinoxylans, cellulose, lignin) Antioxidants (phenolic acids) Crease Brush Inner - and Outer pericarp (4-5%) Bran THE WHEAT GRAIN KERNEL -UNEVEN DISTRIBUTION OF TARGET COMPONENTS

26. #26 Rats fed with wheat and rye fractions Aleurone / similar fractions: highest butyrate production (hind gut). wholemeal flour < bran < aleurone

27. #27 In Vitro ANTIOXIDANT CAPACITY: main active component: ferulic acid from aleurone and bran fractions

28. #28 HEALTHGRAIN nutrition interventions

29. #29 Effect of refined vs. whole grain products on body weight and composition and risk markers of cardiovascular disease (Copenhagen University) Randomisation Δ weight loss : 0.81 kg from randomisation Will inclusion of wheat based whole grain foods in hypo-caloric diet enhance weight loss and improve CVD risk factors? Design: open-labelled randomised trial, 79 overweight women metabolic syndrome features Foods: whole (WG) or refined wheat flour (RW) basis (2000 kJ/d, 12 wk after a 2 week run-in period on refined grains). Results: Body weight, waist circumference, % body fat: significant decrease (P<0.01), both groups, WG Group: significantly (P=0.03) greater decrease in % body fat (-3%) than RG group (- 2.1%). Total and LDL cholesterol changes differed significantly (P<0.05) (RG +5%; WG +0%). Insulin, glucose, and C-reactive protein levels did not change in either group. Weight measured at -2 wks is non-fasting weight. Repeated measures ANCOVA (adjusted for age and body weight at randomisation): P-value <0.001 for time, p=0.15 for diet Conclusions: Both diets effective in inducing weight loss, but whole grain foods had a more beneficial effect on body fat percentage and cholesterol levels than refined grains.

30. #30 Nutrition studies – conclusions Positive impact of whole grain vs refined grain products (human study, less body fat) Higher impact of aleurone rich fractions compared to whole grain butyrate production ↑ (studies in rats and pigs) anti-oxidant capacity and anti-inflammatory response ↑ (vitro, human) lower homocysteine levels → reduced risk for heart disease (human) Stronger glycemic effects of barley kernel and rye products and specially treated starch vs. Wholemeal wheat products ______________________________________________ 1)Composition: Benefits of whole grain can be related both to grain fibre and other bio-active substances. 2)Processing effects

31. #31 Some suggested mechanisms for metabolic benefits of whole grain Insulin resistance Dietary fibre Whole grain (WG) intake Blood lipids Bioactive components Type 2 diabetes (Betaine, choline, folate) Antioxidant/anti- inflammatory status (e.g. Mg) Homocysteine GI Colonic fermentation (Viscosity/ food structure) Oligo-saccharides, RS (vitamins and phenolic anti- oxidants) CVD ( e.g SCFA, phenolic metabolites) (3) (1) (2)

32. #32 Take Home Messages Wheat grain contains interesting nutrients such as fibre and numerous phytonutrients which can provide health benefits The HEALTHGRAIN diversity screen shows large variation in bioactive components which can be exploited by wheat breeders Novel technologies (innovative milling, dry fractionation and enzyme technologies) can increase safety and ‘unlock’ physiologically active fractions from wheat grains. These include soluble arabinoxylan oligosaccharides (AXOS), included in the definition of dietary fibre. A major feature of AXOS is colonic fermentation to short chain fatty acids, butyric acid in particular (fuel for colon mucosa cells).

33. #33 MORE OF GRAIN - FROM SCIENCE TO ACTION