Structural diversity of polyphenols and main dietary sources

Polyphenols are secondary plant metabolites and although they must contain a characteristic phenolic ring, they have very diverse structures⁽ Reference Tsao ^{4 –} Reference Harborne ^{7 )} (Fig. 1). There are relatively simple structures with recognisable phenolic cores such as the phenolic acids (e.g. hydroxybenzoic acids and hydroxycinnamic acid derivatives) and stilbenes and lignans⁽ Reference Tsao ^{4 )}. The flavonoids are a major sub-group based on a C₆–C₃–C₆ core structure with variations that lead to the most common groups, e.g. flavan-3-ols or catechins, flavonols and anthocyanidins. Other flavonoid groups such as flavanones, etc. are important in citrus fruits⁽ Reference Crozier, Jaganath and Clifford ^{5 )}.

Fig. 1. Structures of common polyphenol groups.

There are also complex large molecular weight tannin derivatives⁽ Reference Harborne ^{7 )}, which are split into condensed tannins (e.g. proanthocyanidins composed of flavan-3-ol groups) and hydrolysable tannins (e.g. the ellagitannins which are composed of esters of ellagic acid derivatives). However, the enormous complexity of (poly)phenols lies in the potential for multiple decorations with other groups, including sugars, alcohols and acids⁽ Reference Harborne ^{7 )}. In fact, most common flavonoids are rarely found in foods without attached sugars, acids or alcohols. For example, the hydroxycinnamic acid, caffeic acid, is most often found as chlorogenic acid, an ester with quinic acid, and anthocyanidins and flavonols are rarely found without attached sugars (i.e. as anthocyanins and flavonol glycosides)⁽ Reference Tsao ^{4 –} Reference Harborne ^{7 )}.

Plant-based foods are the primary source of polyphenols and excellent analytical work by various groups, including data on Finnish intake⁽ Reference Ovaskainen, Törrönen and Koponen ^{8 )} has outlined the polyphenol contents of common foods (Fig. 2) Although the highest sources are clearly fruits, coffee and tea make a larger overall contribution to intake due to higher volume of intake. In fact, the major dietary sources of polyphenols vary with habitual diet with tea intake more important in the UK and wine and fruit intake in Mediterranean diets⁽ Reference Ovaskainen, Törrönen and Koponen ^{8 )}. It is clear that berries can contribute high polyphenol content and there is evidence that increased berry intake has had beneficial effects on health in Scandinavian diets⁽ Reference Puska, Pietinen and Uusitalo ^{9 )}. As berries are generally universally regarded as palatable, greater inclusion in the diet represents a plausible means to increase overall polyphenol intake. In addition, although coffee and tea are rich in specific polyphenols⁽ Reference Zamora-Ros, Rothwell and Scalbert ^{10 )} (flavan-3-ols in teas and hydroxycinnamic acid derivatives in coffee), berry intake brings higher levels of specific components such as anthocyanins and ellagitannins⁽ Reference Tsao ^{4 ,} Reference Manach, Scalbert and Morand ^{6 ,} Reference Harborne ^{7 )}. Although other fruits and vegetables, nuts and certain breads can be reasonably good sources of polyphenols, the focus of the present paper is on berries given the long history of the Hutton Institute in berry cultivation and breeding.

Fig. 2. Total phenol contents of common foods.

Enhancing polyphenol content and composition in berries: breeding

There is substantial variation in polyphenol composition between different species of berries. Therefore as anthocyanins have been implicated as influencing human disease⁽ Reference Cassidy, Rogers and Peterson ^{1 ,} Reference He and Giusti ^{11 )} then selecting anthocyanin-rich berry species such as black currant, blueberries and blackberries could be important (Fig. 3) for intervention trials. Within species, there is also great variation in polyphenol content but also in composition. Some of the most different material may arise from wild, unimproved varieties and provides another source of polyphenol variation. For example, anthocyanin composition in raspberry varieties can be markedly different (Fig. 4(a)), which may influence potential bioactivity. There are substantial differences in flavonol content and composition in black currant genotypes (Fig. 4(b)), which can be of importance as quercetin has been implicated in various health benefits⁽ Reference Boots, Haenen and Bast ^{12 )}. The differences between genotypes are under genetic control and can be selected for in breeding programmes. The variations can be even more subtle. In blueberry, certain genotypes accumulate a higher proportion of acetylated anthocyanins which set them aside from other genotypes (Fig. 4(c)). This can be of biological relevance because acylated anthocyanins improve colour tone and stability in juices⁽ Reference Kalt, McDonald and Ricker ^{13 )} but have been suggested to be less bioavailable than non-acylated anthocyanins⁽ Reference Prior and Wu ^{14 )}.

Fig. 3. Representative polyphenol composition of berry species.

Fig. 4. Polyphenol compositional diversity in different varieties of berry species. (a) anthocyanin diversity in raspberries; (b) flavonol diversity in black currant varieties; (c) anthocyanin diversity in blueberries; (d) anthocyanins in a raspberry variety grown in different locations in Europe.

Polyphenol content and composition can also be influenced by environmental or climatic conditions. For example, identical varieties of raspberries grown under identical agronomic conditions at different sites across Europe have different compositions but these compositions appear to be consistent at those locations across seasons (Fig. 4(d)). Given that predicted climatic changes envisaged under models suggest Europe will become generally warmer, varieties that are currently favoured in more southerly European locations may shift northward in future years. What is clear is that inter-genotype variation can be compounded by variation due to environmental and agronomic conditions and berries (or berry products) used in intervention studies must have their polyphenol content and composition closely defined.

Classical breeding usually involves selecting parents with useful suitable genotypic variation in polyphenol content and composition (and other favourable traits such as pest resistance, good yield, low fertiliser inputs, etc.) then screening for progeny with consistently elevated levels of specific components⁽ Reference Finn, Hancock and Hancock ^{15 ,} Reference Brennan and Graham ^{16 )}. This essentially serendipitous gene-shuffling approach is often slow as many berry species require years before they reach sexual maturity and begin to set fruit. However, modern exploitation of increasing genetic and genomic resources (genetic maps, genome sequences, etc.) can provide gene-based markers for polyphenol traits that accelerate breeding by selecting for genotypes likely to have beneficially altered polyphenol composition well before sexual maturity and fruit set. Genetic maps and genome sequences are already available for some berry species⁽ Reference Brennan, Jorgensen and Hackett ^{17 ,} Reference McCallum, Woodhead and Hackett ^{18 )} and our understanding of the biosynthesis of these polyphenol components is increasing rapidly.

Another means to obtain polyphenol-enriched berries would be to use a synthetic biology approach to modulate the biosynthesis of individual components by targeted alteration in the expression of specific genes. Techniques such as CRISPR (clustered regularly interspaced short palindromic repeats) are being developed that allow the surgical elevation of expression of endogenous genes that may modulate polyphenol content/composition⁽ Reference Belhaj, Chaparro-Garcia and Kamoun ^{19 )}. However, in addition to concerns about genetic manipulation, various polyphenols are sensorially important; ellagitannins provide the astringency essential for the recognisable flavour profile of raspberries and flavonoids can influence bitterness⁽ Reference Drewnowski and Gomez-Carneros ^{20 )}. Therefore, increasing the levels of certain polyphenols may lead to unpalatable fruits especially as domestication and concerted breeding programmes have consistently selected for fruits with reduced bitterness and astringency, as can be noted if wild berry varieties are tasted.

Another possibility would be to ‘lift’ biosynthetic pathways from plants into suitable microbial vectors such as yeasts and to produce polyphenols under large-scale in vitro conditions as envisaged in the European Union funded BACHBERRY grant. This approach has already been achieved for high-value plant metabolites (such as artemisinin)⁽ Reference Ro, Paradise and Ouellet ^{21 )} or alkaloid precursors⁽ Reference Brown, Clastre and Courdavault ^{22 )} and is probably better suited for the production of specific polyphenols as pharmaceuticals.

Processing methods for polyphenol-enriched products

Human subjects have been processing foods and altering their polyphenol composition for centuries. Fermentation of teas, cocoa beans and grapes, or roasting of cocoa and coffee beans has long been used and can massively influence polyphenol composition and potential bioactivity. For example, oxidation during black tea processing leads to the formation of characteristic orange-brown theaflavin products from flavan-3-ols⁽ Reference Subramanian, Venkatesh and Ganguli ^{23 )}. Fermentation during wine making often results in substantial degradation of grape anthocyanins, but can produce unusual anthocyanin derivatives. For example, Vitisin A derivatives are formed through reaction with pyruvate during red wine production⁽ Reference Fulcrand, Benabdeljalil and Rigaud ^{24 )}. These process-derived pigments alter the colour and tone of the wines and although they are more stable under gastrointestinal conditions⁽ Reference McDougall, Fyffe and Dobson ^{25 )}, their uptake is less well understood⁽ Reference Oliveira, Wu and Zhang ^{26 )}. Fruit wines made from Salal berries retain red colouration due to their high content of stable anthocyanin diglycosides containing pentoses⁽ Reference McDougall, Austin and Van Schayk ^{27 )}.

The simplest way to reformulate to obtain polyphenol-enriched products may be to add berry juices to staple juices such as orange and apple juices. Berry juices, purees and ‘wastes’ or co-products (from juicing or pre-prepared fruit products) added to apple/citrus juices can easily double the polyphenol content and introduce specific novel polyphenol components (e.g. anthocyanins or ellagitannins). Studies on (e.g.) black currant-enriched apple juices and raspberry-enriched orange juices have illustrated that palatability is not reduced and the added colouration can enhance acceptability⁽ Reference Shahidi, Alasalvar, Shahidi and Alasalvar ^{28 )}.

Another method to increase polyphenol intake is to use berry purees in products rather than clarified juices. The purees have the added benefits of the non-extractable polyphenols, which are either covalently attached to cell walls or embedded in the tissue matrix. These components have been called the ‘neglected and underrecorded polyphenol content of foods, the missing polyphenols’⁽ Reference Saura-Calixto ^{29 )}. As they are essentially antioxidant-rich dietary fibre, they can have positive effects through the gut such as enhancing lipid excretion, promoting chemopreventive effects throughout the gastrointestinal tract, encouraging growth of colonic microbiota and the release of systemic polyphenol metabolites through colonic degradation.

It is well known that techniques used to aid juicing such as addition of pectinolytic enzymes can have various effects on polyphenol composition and result in a balance of release of polyphenol from non-extractable polyphenols and degradation of flavonoid glycosides⁽ Reference Rommel, Heatherbell and Wrolstad ^{30 )}. As novel extraction and processing methods (e.g. supercritical CO₂, pressurised water, microwave-enabled systems, pulsed electric fields, etc.) become available, their effects on polyphenol content and composition need to be defined.

Berry wastes or juicing residues could be used to improve nutrient stability in bakery⁽ Reference Ranawana, Raikos and Campbell ^{31 )} or meat products perhaps through inhibition of lipid oxidation. These could substitute for synthetic antioxidants currently used and have the added bonus of providing higher polyphenol content and dietary fibre.