Polyphenols (noun, pronunciation of the singular /pɑli'finəl/ or /pɑli'fɛnəl/) are a structural class of natural, synthetic, and semisynthetic organic chemicals characterized by the presence of large multiples of phenol structural units (right). The number and characteristics of these phenol structures underlie the unique physical, chemical, and biological (metabolic, toxic, therapeutic, etc.) properties of particular members of the polyphenol class. The name derives from poly-, from the ancient Greek word πολύς (polus, meaning “many, much”) and the word phenol which refers to a chemical structure formed by attaching to an aromatic benzenoid (phenyl) ring, an hydroxyl (-OH) group akin to that found in alcohols (hence the "-ol" suffix). The term polyphenol appears to have been in use since 1894.

Definition of the term polyphenol

Basis in the organic chemical and natural products literature

The most research-informed and chemistry-aware definition of the polyphenol term describes the class as (i) generally moderately water-soluble compounds, (ii) with molecular weight of 500–4000 Da, (iii) >12 phenolic hydroxyl groups, and (iv) 5–7 aromatic rings per 1000 Da, where the limits to these ranges are necessarily somewhat flexible.

Examples of the compound class include the black tea antioxidant theaflavin-3-gallate shown below, and the hydrolyzable tannin, tannic acid, shown above. Notably, the historically important chemical class of the tannins is a subset of the polyphenols. These examples highlight the high density of phenolic substructures that characterize the class and underlie their properties, and introduces their origin as plant-derived substances (phytochemicals).

Individual polyphenols engage in reactions related to their core structure—standard phenolic reactions (e.g., ionization, oxidations to ortho- and para-quinones, and other underlying aromatic transformations related to the presence of the phenolic hydroxyl, etc.; see phenol image above)—as well as reactions related to their peripheral structures (e.g., nucleophilic additions, oxidative and hydrolytic bond cleavages, etc.). As critically, per the definition, the polyphenols display behaviors more explicitly limited to the polyphenol class—for instance, formation of particular metal complexes (e.g., intense blue-black iron(III) complexes), and precipitation of proteins and particular amine-containing organics (e.g., particular alkaloid natural products).

Underlying importance of class definition to structure-activity understanding

Less than rigorous use of the polyphenol term toward the lower molecular weight end of the range is understandable; it is associated, for instance, with the limited understanding of chemical structure in play when a plant extract is first studied (see, for instance, pharmacognosy to understand the process from plant extract to determined chemical structures), and with a desire to associate newly discovered phenolic substances with particularly well-studied, pharmacologically interesting "true" polyphenols (e.g., for pattern, see ,).

The need to distinguish between structure classes, even in rough terms, relates to the critical concept of structure-activity relationships (SAR)—the fact that the biological activities of chemical agents are structure-derived and structure-dependent., Hence, understanding of the way in which phenolic chemical structures vary (disambiguating types) is important to understanding of how and why biological activities vary with those structures (see, e.g., ,). This in turn underlies good correlative research and consumer decision-making about the use of phenolic substances, including relevant simple, oligo-, and polyphenols.

Catalog of conflicting definitions based on common and web-usage

In the Merriam Webster and Babylon on-line dictionaries, rather than differentiation based on the presence of many (poly) phenolic substructures, polyphenols are instead defined as molecules with more than one hydroxyl group on one or more phenol unit per molecule. At wiktionary, they are alcohols [sic] containing two or more benzene rings that each have at least one hydroxyl group attached, a definition that likewise confounds simple natural phenols and polyphenols, and further confounds the very different properties of alcoholic and phenolic hydroxyl groups. Likewise, the term polyphenol is increasingly confounded in some parts of the bioactivity and analytical research literature with simple and mid-molecular weight plant phenolic natural products (see next paragraph), though less so in literatures of modern organic and natural products chemistry where more precise understandings of naming apply.

In the parts of literature where polyphenol is used less rigorously, the term is used almost interchangeably with other terms/structures such as simple natural phenols, intermediate weight phenolics, and the newer and less precedented term phenoloids.

Finally, some web sources give definitions focusing on particular antioxidant properties or other health benefits, or on polyphenol plant origins. The latter distinction is rejected on chemical terms, because identical chemical agents can be isolated from various natural sources other than plants, or can be prepared synthetically.

Different aims of lexicography vs. chemical nomenclature in polyphenol definition

It is generally understood that the aims of normal English lexicography versus chemical nomenclature vary and are to an extent at odds. Dictionaries of English words on the web or otherwise collect and report the meanings of words as their uses appear and change over time. Chemical nomenclature on the other hand (with IUPAC nomenclature as the best example) is necessarily more restrictive: it aims to standardize communication and practice so that when a chemical term is used, it has a fixed meaning relating to chemical structure, and thereby, can give insights into chemical properties and derived molecular functions. This tension between scientific vs. other definitions is evident in the phenol and polyphenol literature, and correlates with commercial interest in promoting particular nutraceutical and supplement products (e.g., ), and with broad scientific interest in new phenolic substances—where new reports of properties often precede structure assignment, and practice dictates that new discoveries be tied to earlier ones, As such, first assignments of structure type may be inaccurate. For instance, when the term polyphenol appeared in the Medical Subject Headings (MeSH) database of the National Library of Medicine in relation to a 1980 report on sclerotization induced by extracts from the Surinam cockroach. The NLM entry's"note" on use of the term polyphenol states: "GEN only; [One should] use the precise structure header, most commonly in the FLAVONOIDS group; this [polyphenol] term only refers vaguely to phenolic (aromatic) hydroxyls…".

Hence, while the polyphenol definition provides only rough boundaries, they are structurally and functionally relevant and useful; despite web writings to the contrary, all chemicals presenting a phenolic group are not polyphenols, and they do not provide the health benefits associated with, for instance, tea and wine polyphenols (see below). As noted, the problematic bisphenol A can fit a loose definition. Phenol itself, C₆H₅OH, is not a therapeutic agent; it is caustic, and toxic if taken internally. As structural complexity increases from phenol to di-,tri- and oligophenols, and on through to polyphenols, one encounters hepatotoxins, drugs, endocrine disruptors, industrial dyes, plant pigments (i.e., all manner of functionally active agents) before arriving at commercially and medically important true polyphenols.

In short, discussion of phenolic substances and their general and specific biologic and therapeutic properties is necessarily connected to chemical structure, with chemical classifications such as "polyphenol" substituting in literature and common use for detailed chemical knowledge. The ability to correlate beneficial properties—value in commercial tanning, anti-proliferative pharmacology, etc.—of the polyphenol chemical class will only be as strong as the definitions applied to this and other phenol classes.

Origin of the rigorous polyphenol definition

Building off of earlier natural products research efforts of E.C. Bate-Smith, A. Swain and T. White that characterized specific structural characteristics common to plant phenolics used in tanning, research chemist Edwin Haslam and colleagues developed the four-point definition based on solubility, molecular weight, numbers of phenolic substructures, and aromatic ring density presented above. This definition is now know as the White–Bate-Smith–Swain–Haslam (WBSSH) definition. It is the definition applied in the highest level research and practical discussions of the structure class.

Polyphenols, versus phenolic polymers

The use of the term polyphenol does not indicate that the class are a type of polymer. Generally, there are no repeating monomeric structures inherent to polyphenols, though in some molecules such as tannic acid—shown above—and in the condensed tannins, particular phenolic units appear repeatedly. An example which highlights the distinction is the complex, random polymeric phenolic polymer lignin that constitutes the woody tissue of gymnosperms, the most abundant phenolic and, in general, polymeric substance. Lignin is formed from radical-mediated cross-linking of simple phenolics derived from the plant phenylpropanoid pathway. Lignin is a phenolic polymer, but not a polyphenol. Note, there is theoretically no reason to exclude the possibility of polymerization of a phenolic monomer, such that it would result in formation of a structure that would be both a polyphenol by the WBSSH definition, and have the repeating characteristics of a true polymer. The arecatannin-type natural products from Ceylonese cassia bark and Areca seed may be considered as an example of such.

Chemical Structure and Properties

Structural features

In common with simple and mid-molecular weight phenolic dimers and trimers (natural phenols), the phenol substructures of polyphenols have various further nomenclatures depending on the number of phenolic hydroxyl groups. A phenol, per se, is the term for a substructure with one phenolic hydroxyl group, catechol- and resorcinol-types (benzenediols) have two, and pyrogallol- and phloroglucinol-types (benzenetriols) have three. Polyphenols always have heteroatom substituents other than hydroxyl groups; as might be expected, ether and ester linkages are common, as are various carboxylic acid derivatives (see theaflavin gallate image).

Image:Phenol2.svg	[[Image:Pyrocatechol.svg	[[Image:pyrogallol2.svg	[[Image:Resorcine.svg	[[Image:Phloroglucin.svg
[[Phenol	Pyrocatechol	Pyrogallol	Resorcinol	Phloroglucinol

As the earlier images suggest, polyphenol compositions are normally limited to carbon, hydrogen and oxygen in undefined proportion. Carbon frameworks can be complex, arising from various biosynthetic pathways aimed at plant and related secondary metabolites; e.g., the 7-atom ring (7-membered ring) appearing in theaflavin structure above is an example of a "carbocycle" that is of a nonbenzenoid aromatic tropolone type. In addition, various biaryls and triaryls occur (e.g., biphenyls), see figure at right, and benzopyrans and normal and C-glucoside derivatives frequently appear (further figure at right) —e.g. in condensed, complex and hydrolyzable tannins such as in stenophyllanin A (1), acutissimin B (2), mongolicain A (3), stenophynin A (4), mongolicanin (5), and mongolicin B (6). Spiro-type structures as illustrated at right appear as in preceding compound (3); furanoid, pyrone, and other heterocycles appear as in compounds (4) and (6); (diaryl)methyl structures as in (1), (2), and (6); as do pyrans and dioxins, etc. Because of the preponderance of saccharide-derived core structures (e.g., see tannic acid image above), as well as spiro- and other structure types, natural chiral (stereo) centers abound.

Chemical properties

Polyphenols are molecules producing autofluorescence, especially lignin and the phenolic part of suberin.

Polyphenols are reactive species toward oxidation. The Fenton's reagent, a mixture producing reactive oxygen species, used in association with a photo-oxidation system may be used to treat oil mill waste water. A complex mixture of polyphenols, found in food for example, can undergo autoxidation during the ageing process. Simple natural phenols can lead to the formation of B-type procyanidins in wines or in model solutions. This is correlated to the non enzymatic browning color change characteristic of this process. This phenomenon can be observed in foods like carrot purees. ABTS may be used to characterise polyphenol oxidation products.

Polyphenols can interact with proteins (case of tannins) and other food matrices.

Chemical synthesis

Polyphenols can be synthetised chemically from phenol to produce a mixture of phenylene and oxyphenylene.

Natural phenols can be enzymatically polymerised. Laccase and peroxidase induced the polymerization of syringic acid to give a poly(1,4-phenylene oxide) bearing a carboxylic acid at one end and a phenolic hydroxyl group at the other.

Classification and nomenclature

In terms of chemical classification, polyphenols are generally divided into hydrolyzable tannins (gallic acid esters of glucose and other sugars or cyclitols) and phenylpropanoids, such as lignins, flavonoids, and condensed tannins. This division is derived from the variety of simple polyphenolic units derived from secondary plant metabolism as well as classical divisions based upon the relative importance of each base component to different fields of study. Tannin chemistry originated in the importance of tannic acid to the tanning industry; lignins to the chemistry of soil and plant structure; and flavonoids to the chemistry of plant secondary metabolites for plant defense, and flower color (e.g. from anthocyanins).

Chemical uses

Some polyphenols are traditionally used as dyes. For instance, in the Indian subcontinent, the pomegranate peel, high in tannins and other polyphenols, or its juice, is employed in the dyeing of non-synthetic fabrics.

Polyphenols, especially, tannins, can be used as precursors in green chemistry notably to produce plastics or resins by polymerisation with or without the use of formaldehyde or adhesives for particleboards. The aims are generally to make use of plant residues from grape, olive (called pomaces) or pecan shells left after processing.

A special form of polyphenols-derived resins are the EUV resists.

Polyphenols are also used for the production of creosote to treat wood.

Biology

Occurrence in nature

The most abundant polyphenols are the condensed tannins, found in virtually all families of plants, and comprising up to 50% of the dry weight of leaves. Plant polyphenols have antioxidant action and may help reduce tooth decay.

Some polyphenols produced by plants in case of pathogens attacks are called phytoalexins. Such compounds can be implied in the hypersensitive response of plants. High levels of polyphenols in some woods can explain their natural preservation against rot.

Cucumbers grown on vermicompost may be less susceptible to striped cucumber beetle attack due to a higher induced level of polyphenols in the plant.

Polyphenols can be involved in allelopathic interactions in soil or in water. The fruit pulp of the plant Liriope muscari contains phenolic compounds which inhibit its own seeds germination.

Polyphenols can be a source of pollution in sites near processing plants producing olive oil, coffee (see coffee wastewater) or paper. Laccases (found for instance in the fungal species Panellus stipticus) can be used in bioremediation.

Polyphenol content can be an element of chemotaxonomy.

Polyphenols can also be found in animals. In arthropods like insects and crustaceans polyphenols play a role in epicuticle hardening (sclerotization). The hardening of the cuticle is due to the presence of a polyphenol oxidase. In crustaceans, there is a second oxidase activity leading to cuticle pigmentation. There is apparently no polyphenol tanning occurring in arachnids cuticle.

Metabolism

Biosynthesis

Polyphenol oxidase (PPO) is an enzyme that catalyses the oxidation of o-diphenols to produce o-quinones. It is the rapid polymerisation of o-quinones to produce black, brown or red polyphenolic pigments that is the cause of fruit browning. In insects, PPO serves for the cuticle hardening.

Laccase is a major enzyme that initiates the cleavage of hydrocarbon rings, which catalyzes the addition of a hydroxyl group to phenolic compounds. This enzyme can be found in fungi like Panellus stipticus, organisms able to break down lignin, a complex aromatic polymer in wood that is highly resistant to degradation by conventional enzyme systems.

Anthracyclines or hypericin are derived from polyketides cyclisation.

The glycosylated form increases the solubility of polyphenols.

Content in food

Polyphenols are considered antinutrients, compounds that interfere with the absorption of nutrients as a mechanism of plant defense against herbivores. Leaf protein concentrates used as animal food can also be rich in polyphenols. Animals produce salivary proteins, like protein IB5, as herbivore adaptations to plant defense, to counter the tannins astringent effect. Some animals like the greater glider or the koala can digest low nutrient foliage, specifically eucalypt leaf matter, which contains a variety of phenolic and terpenoid compounds and a high concentration of lignified fiber. Some frugivores like birds are reported to not sense astringency.

In human food

Polyphenols from algae (phlorotannins) may be used as additives to prevent lipid oxidation during fish preservation.

Role of processing in phenolic composition

Phenolic and carotenoid compounds with antioxidant properties in vegetables have been found to be retained significantly better through steaming than through frying.

Polyphenols in wine, beer and various nonalcoholic juice beverages can be removed using finings, substances that are usually added at or near the completion of the processing of brewing.

Marketing argument

The presence of polyphenols is a marketing argument to sell functional foods, dietary supplements or anti-aging creams.

Functional foods may contain polyphenols. For superfruit beverages, which may include extracts from fruits like açai or pomegranate, the detailed composition of polyphenols is usually not revealed on the nutrition label. Instead, there may be an ORAC value given for the in vitro antioxidant capacity of the product. Polyphenol-enriched drinks may actually deliver the intended blend of bioavailable polyphenols, which would normally require consumption of several different plant-derived foods.

As a matter of pharmacovigilance, health benefits from using these products have not been scientifically confirmed or approved by regulatory authorities and may only be supported by preliminary research. Accordingly, there are no recommended Dietary Reference Intake levels established for polyphenols as exist for essential nutrients.

Potential health effects

Research on polyphenols

Bioavailability

Questions on the relationship between health benefits and polyphenols generally revolve around bioavailability.

Compared with the effects of polyphenols in vitro, the effects in vivo, although the subject of ongoing research, are limited and vague. The reasons for this are 1) the absence of validated in vivo biomarkers, especially for inflammation or carcinogenesis; 2) long-term studies failing to demonstrate effects with a mechanism of action, specificity or efficacy; and 3) invalid applications of high, unphysiological test concentrations in the in vitro studies, which are subsequently irrelevant for the design of in vivo experiments. In rats, polyphenols absorbed in the small intestine may be bound in protein-polyphenol complexes modified by intestinal microflora enzymes, allowing derivative compounds formed by ring-fission to be better absorbed.

Matrix effect

The poor bioavailability of polyphenols in vivo may be due to a matrix effect. Casein found in milk added to a polyphenol-rich food like tea may reduce the absorbed polyphenols content by the formation of a complex as demonstrated in vitro. Moreover other substances such as caffeine can form insoluble complexes with polyphenols.

Polyphenols may also interact with fibers like pectins and have a positive effect in large intestine accessibility.

Analysis

The analysis techniques are those of phytochemistry: extraction, isolation, structural elucidation, then quantification.

Extraction

Extraction of polyphenols can be performed using a solvent like water, hot water, methanol, methanol/formic acid, methanol/water/acetic or formic acid etc. Liquid liquid extraction can be also performed or countercurrent chromatography. Solid phase extraction can also be made on C18 sorbent cartridges. Other techniques are ultrasonic extraction, heat reflux extraction, microwave-assisted extraction, critical carbon dioxide, pressurized liquid extraction or use of ethanol in an immersion extractor. The extraction conditions (temperature, extraction time, ratio of solvent to raw material, solvent and concentrations) have to be optimized.

Mainly found in the fruit skins and seeds, high levels of polyphenols may reflect only the measured extractable polyphenol (EPP) content of a fruit which may also contain non-extractable polyphenols.

Concentration can be made by ultrafiltration. Purification can be achieved by preparative chromatography.

Analysis techniques

Phosphomolybdic acid is used as a reagent for staining phenolics in thin layer chromatography. Polyphenols can be studied by the mean of spectroscopy, especially in the ultra violet domain, by fractionation or paper chromatography. They can also be analysed by chemical characterisation.

Instrumental chemistry analyses include separation by high performance liquid chromatography (HPLC), and especially by reversed-phase liquid chromatography (RPLC), can be coupled to mass spectrometry. Purified compounds can be identified by the mean of nuclear magnetic resonance.

Microscopy analysis

The DMACA reagent is an histological dye specific to polyphenols used in microscopy analyses. The autofluorescence of polyphenols can also be used, especially for localisation of lignin and suberin.

Quantification

A method for polyphenolic content quantification is volumetric titration. An oxidizing agent, permanganate, is used to oxidize known concentrations of a standard tannin solution, producing a standard curve. The tannin content of the unknown is then expressed as equivalents of the appropriate hydrolyzable or condensed tannin.

Some methods for quantification of total polyphenol content are based on colorimetric measurements. Some tests are relatively specific to polyphenols (for instance the Porter's assay). Total phenols (or antioxidant effect) can be measured using the Folin-Ciocalteu reaction. Results are typically expressed as gallic acid equivalents. Polyphenols are seldom evaluated by antibodies technologies.

Other tests measure the antioxidant capacity of a fraction. Some make use of the ABTS radical cation which is reactive towards most antioxidants including phenolics, thiols and vitamin C. During this reaction, the blue ABTS radical cation is converted back to its colorless neutral form. The reaction may be monitored spectrophotometrically. This assay is often referred to as the Trolox equivalent antioxidant capacity (TEAC) assay. The reactivity of the various antioxidants tested are compared to that of Trolox, which is a vitamin E analog.

Other antioxidant capacity assays which use Trolox as a standard include the diphenylpicrylhydrazyl (DPPH), oxygen radical absorbance capacity (ORAC), ferric reducing ability of plasma (FRAP) assays or inhibition of copper-catalyzed in vitro human low-density lipoprotein oxidation.

New methods including the use of biosensors can help monitor the content of polyphenols in food.

Quantitation results produced by the mean of diode array detector-coupled HPLC are generally given as relative rather than absolute values as there is a lack of commercially available standards for every polyphenolic molecules.

Other techniques

Chemometrics analyses on acquired data can be performed to compare samples from different origins.

Genetic analysis

In Vitis vinifera cells suspensions, polyphenol production is regulated by sugars levels.

See also

Polyphenolic protein

References

External links

groupe polyphenols website

; Other tools

Phenol-Explorer, the first comprehensive and freely available electronic database on polyphenol content in foods.

KNApSACK

massbank.jp, a high resolution Mass Spectral Database

PubChem, NCBI

liberherbarum.com, the incomplete reference-guide to Herbal medicine, Copyright © Erik Gotfredsen.

metabolomics.jp (English, Japanese)

KEGG: Kyoto Encyclopedia of Genes and Genomes

ChEBI

Comparative Toxicogenomics Database for toxicity

Category:Phytochemicals Category:Dietary antioxidants

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