WO2012150370A1 - Synergic polyphenol combination - Google Patents

Abstract

The present invention relates to a combination of the polyphenols resveratrol, quercetin and catechin in a molar ratio of approximately 1:1:2 or, alternatively, approximately 1:1:5, which has synergic antioxidant power. This combination of polyphenols is suitable for incorporation into any type of pharmaceutical, cosmetic, foodstuff or veterinary formulation and may be used in the prevention and treatment of pathological or physiological conditions that can be treated with antioxidant and free-radical scavenger substances.

Description

 D E S C R I P C I O N

"SYNERGIC POLYPHENOL COMBINATION" Field of technique

 The present invention relates to polyphenol compositions that have a synergistic antioxidant power. Prior art

 Polyphenolic compounds of natural origin are recognized antioxidant agents that have been shown to be effective as free radical scavengers. Free radicals are highly reactive molecules that are naturally generated in the tissues, and that can also be formed in an increased way due to environmental stress situations such as, for example, cigarette smoke, the presence of ionizing radiation, ultraviolet radiation, or environmental pollution, as well as other causes such as stress situations or the practice of a very intense exercise. Free radicals are capable of inducing the oxidation of nucleic acids, proteins and lipids of the organism, which can contribute to the acceleration of the aging process and trigger diseases associated with aging such as degenerative diseases, cancer, cardiovascular diseases and dermatological diseases.

 In the state of the art it is described that the antioxidant properties of naturally occurring polyphenols can be applied in different therapeutic areas, and can also contribute to delaying the signs of aging thanks to their ability to reduce cellular and tissue oxidation.

Resveratrol is a polyphenolic compound present in the seed and in the skin of the grape (Vitis vinifera) and in other plant products. Its therapeutic interest has been reinforced since its possible involvement in the cardioprotective effects of red wine was postulated, as described in the article Siemann et al., Concentration of the phytoalexin resveratrol in wine, Am. J. Eno. Vitic, Free. Rad. Res., 1992, 43, 49-52, and its possible participation in the so-called French paradox: The French population, which has a diet rich in saturated fats, has a lower disease mortality rate cardiovascular than the rest of developed countries. This reduction was associated with the increased consumption of red wine and the presence of resveratrol in red wine.

 Numerous studies, conducted in experimental models and published over the last few years confirm that resveratrol:

 - it can be effective in preventing or slowing the progression of various pathologies such as cancer, diabetes, and cardiovascular diseases,

 - it has antiviral and anti-inflammatory properties, showing itself as a good alternative for the treatment of chronic inflammation,

 - it has protective properties in different organs, especially in ischemic heart disease associated with myocardial infarction, thanks to its ability to prevent the oxidation of low density lipoproteins (LDL), and

 - slows the symptoms of aging in yeasts and some simple vertebrates.

 Also noteworthy is the study by Baur et al., Resveratrol improves health and survival of mice on a high-calorie diet, Nature, 2006, 444, 337-342, in which it was shown that rats with a high-fat diet had higher pathologies and less survival than others undergoing the same diet, but those that were also given resveratrol.

 Although pharmacological effects have been observed for resveratrol even at the relatively low concentrations present in the usual sources of the diet, given its low bioavailability (Walle et al., High absorption but very low bioavailability of oral resveratrol in humans, Drug Metab. Disp., 2004, 32 (12), 1377-1382), it is desirable to administer higher doses to achieve higher plasma and tissue concentrations and to enhance their therapeutic effects. However, the possible adverse effects of the continued administration of high doses of resveratrol are unknown and, in any case, the economic cost would be very high.

Other polyphenols of the flavonoid family known for their antioxidant properties are quercetin and catechins as described, for example, in articles Zhang et al., Antioxidant properties of quercetin, Adv. Exp. Med. Biol., 201 1, 915, 283-9, and Plumb et al., Antioxidant properties of catechins and proanthocyanidins: effect of polymerisation, galloylation and glycosylation, 1998, 29 (4), 351-8.

 In the state of the art combinations of resveratrol with other polyphenols have been described to increase their bioavailability.

 For example, the combination with quercetin may contribute to increasing the bioavailability of resveratrol, as described in the article De Santi et al., Sulphation of resveratrol, a natural compound present in wine, and its inhibition by natural flavonoids, Xenobiotica, 2000, 30 (9), 857-866.

 Some combinations of resveratrol with other polyphenols, in particular with quercetin and catechins, in different proportions have been described in the state of the art.

 International patent application WO-A-2010/135589 is specifically directed to drinkable dietary supplements prepared based on a suspension of resveratrol in water to which, optionally, other ingredients are added, for example, quercetin and / or catechins from an extract of green tea, and which are mainly in the form of epigallocatechin gallate, as well as other antioxidants in the form of extracts or concentrates of plant origin, and also other agents called "energy", also of plant origin. No preferred ratio between resveratrol and the rest of the additional components is described. In particular, a specific example of a beverage is described, in which the weight ratio between resveratrol, quercetin and catechins was found to be 17.2: 3.4: 1, equivalent to a molar ratio of 21, 8: 3.3 :one .

International patent application WO-A-03/015738 describes a cosmetic composition that can be used for the fight against skin aging and that includes a combination of grape seed extracts, grape leaves and skins. grape. Such grape seed extracts contain polyphenols, especially catechins in amounts up to 15% by weight, grape leaf extracts contain quercetins in amounts up to 5% by weight, and grape skin extracts contain resveratrols in amounts. up to 5% by weight. An example is also described with a mixture of skin, leaf and grape seed extracts, which represents a combination of resveratrol: quercetin: catechins in an approximate ratio of 1: 1: 3 by weight, equivalent to an approximate molar ratio of 1, 3: 1: 3.1. Be describes that said combination has a synergistic effect as regards the inhibition of matrix metalloproteinases (MMP).

 In international patent application WO-A-02/081651 compositions with resveratrol, catechin, epicatechin and quercetin are described, alone or in combination, for use as cardioprotectors, and their fibrinolytic activity is analyzed in an in vitro model with endothelial cells human derived from the umbilical vein (HUVEC) and in an in vivo model in rat. The examples describe compositions in the form of capsules or orange juice in which resveratrol, quercetin and catechins are combined, in a weight ratio of 1: 2.5: 7.5, equivalent to a 1: 1 molar ratio , 9: 5.9.

 On the other hand, in US patent application US-A-2010/0247670 a dry extract obtained from black grape of the Vitis labrusca variety is described, which contains resveratrol, quercetin, catechin and tannins. It also describes the extraction with hot water of the dry extract to obtain infusions, which were effective for the treatment of various conditions, such as common cold, hypercholesterolemia or hyperglycemia, or for dermatological treatments of cosmetic type. The analysis of two of the prepared infusions is also described, according to which the weight ratio between resveratrol, quercetin and catechin is 1: 5: 15 and 1: 4: 63, equivalent respectively to molar ratios of 1: 3, 8: 1 1, 8 and 1: 3: 49.5.

 Similarly, in the Spanish patent application ES-A-2217966 pharmaceutical, dietary and cosmetic compositions are described with antioxidant properties comprising a skin extract, seeds and black grape rasp of the species Vitis vinifera containing polyphenols and anthocyanins. Among the polyphenols are catechin, gallic acid, quercetin, and resveratrol. It is described that said combination of polyphenols and anthocyanins has antioxidant properties, in the proportions in which it is obtained directly from the plant extract. The result of the analysis of a black grape extract is also described, in which resveratrol, quercetin and catechin are in an approximate ratio of 1: 3: 21, equivalent to an approximate molar ratio of 1: 2.3: 16.5 .

The need therefore remains to be able to have a resveratrol composition with improved antioxidant properties, which has a high antioxidant power even at relatively low doses. Object of the invention

 The object of the invention is a synergistic combination of polyphenols.

A composition comprising said combination is also part of the object of the invention.

 The use of the combination for the preparation of a composition with antioxidant power is also part of the invention.

 The combination for use as an antioxidant is also part of the invention.

Detailed description of the invention

 The object of the present invention is a synergistic combination of polyphenols comprising resveratrol, quercetin and catechin, with a molar ratio between resveratrol, quercetin and catechin of about 1: 1: 2 or about 1: 1: 5.

 The authors of this invention have developed a combination of the resveratrol, quercetin and catechin polyphenols in such proportions that, surprisingly, it has a synergistic antioxidant power.

 In a preferred embodiment, the combination of antioxidants comprises resveratrol, quercetin and catechin in a 1: 1: 2 or 1: 1: 5 molar ratio.

 In a preferred embodiment, the molar ratio is 1: 1: 2.

 In another preferred embodiment, the molar ratio is 1: 1: 5.

 In the context of the invention, the term "approximately" means, for the value "1" of the molar ratio, a variation between 0.9 and 1.1, preferably between 0.95 and 1.05; for the value "2" a variation between 1, 8 and 2.2, preferably between 1, 9 and 2.1; and for the value "5" a variation between 4.5 and 5.5, preferably between 4.75 and 5.25, and more preferably between 4.9 and 5.1.

 Throughout the description the percentages refer to percentages by weight and the relationships are molar ratios.

 Within the framework of this invention, "dry extract" means the resulting product after subjecting the plant extract to a drying process whereby substantially all of the water it contains is removed.

In the context of this invention, "purified extract" means an extract having a polyphenol content equal to or greater than 90% by weight On the dry extract. In the case of resveratrol preferably equal to or greater than 95%, more preferably equal to or greater than 98%; in the case of quercetin preferably equal to or greater than 95% expressed as quercetin dihydrate, more preferably equal to or greater than 98%; and in the case of the catechin a catechin content expressed as catechin preferably equal to or greater than 80%, more preferably equal to or greater than 85%.

Antioxidants

 Antioxidants are substances that can protect cells against oxidation and the effects of free radicals, because they eliminate these radicals from the medium and because they inhibit oxidation by oxidizing themselves.

 Among the antioxidants are polyphenols, which are compounds widely distributed in the plant kingdom, and have an appropriate chemical structure to capture free radicals.

 In its molecule, polyphenols incorporate more than one phenol group. Its property as an antioxidant comes from its great reactivity as electron donors and the ability of the radical formed to stabilize and delocalize the missing electron.

 The main sources of polyphenols are red fruits, tea leaves, beer, grapes, wine, olive oil, chocolate, cocoa, nuts, and pomegranates, among others.

Resveratrol

 In the context of the invention the term resveratrol refers to trans-resveratrol and cis-resveratrol interchangeably, and also includes the active ingredient and its possible hydrates and / or solvates.

 Trans-resveratrol is the common name that designates 5 - [(1 £) -2- (4- hydroxyphenyl) ethenyl] -1,3-benzenediol, which can also be called 3,5,4'-trihydroxystilbene, and that It has the following structure:

El cis-resveratrol es el nombre común que designa al 5-[(1 Z)-2-(4- hidroxifenil)etenil]-1 ,3-bencenodiol, y tiene el doble enlace de la figura anterior en posición cis.

Cis-resveratrol is the common name that designates 5 - [(1 Z) -2- (4- hydroxyphenyl) ethenyl] -1, 3-benzenediol, and has the double bond of the previous figure in cis position.

 Trans-resveratrol is a phenolic compound found in nature, produced by various plants. For example, it is one of the constituents of the skin and the grape seed and, consequently, one of the components of wine, especially red wine.

 Trans-resveratrol was first isolated in 1940 from the root of hellebore, as described in the article MJ Takaoka, Of the phenolic substances of white hellebore (Veratrum grandiflorum Loes, fil), J. Faculty Sci Hokkaido Imperial University, 1949, 3, 1-16.

 Trans-resveratrol can be obtained as an extract from various plant species such as, for example, skin and grape seed {Vitis), blueberries (Vaccinium), Polygonum cuspidatum root, hops (Humulus lupulus), Cassia garrettiana , Cassia Quinquangulata, Gnetum klossii, Pterolobium hexapetallum, peanut (Arachis hypogaea), pistachio (Pistacia vera), rhubarb leaves and root (Rheum rhabarbarum and Rheum Rhaponticum), Bauhinia racemosa bark, Veratrum grandiflorum bark Veratrum formosanum root, eucalyptus, spruce and spruce, among others.

 Commercially, plant extracts with various trans-resveratrol contents can be found, reaching values greater than 95% on the dry extract.

 For example, Polygonum cuspidatum root extract of the company Select Botanical, is a purified extract that has a maximum water content of 8% by weight, and whose dry extract has a trans-resveratrol content greater than 98%, determined by HPLC.

Likewise, there are extracts from grape seed and Polygonum cuspidatum root generally, with very variable resveratrol contents that can be between 0.05% and 98% determined by HPLC. The determination of resveratrol content by HPLC can be done, for example, as described in the article Kolouchová-Hanzlíková et al., Rapid method for resveratrol determination by HPLC with electrochemical and UV detections in wines, Food Chem., 2004, 87 ( 1), 151-158. Likewise, trans-resveratrol can be obtained in substantially pure form through suppliers such as, for example, the Sigma-Aldrich company.

 Trans-resveratrol can also be synthesized, for example as described in the article Fariña et al., An improved synthesis of resveratrol, in Nat. Prod. Res., 2006, 20 (3), 247-52.

 On the other hand, cis-resveratrol can be obtained, for example, from trans-resveratrol by exposure to ultraviolet radiation.

 Transrosveratrol is preferably used in the combination of the invention.

 In the combination of the invention, resveratrol may be of synthetic or natural origin. When it is naturally occurring, preferably the source of resveratrol is a plant extract. More preferably as a source of resveratrol, an extract of plant origin with a high trans-resveratrol content is used, preferably greater than 95% on the dried extract, determined by HPLC, more preferably a Polygonum cuspidatum root extract, and even more preferably a purified extract of Polygonum cuspidatum root with a trans-resveratrol content greater than 98% on the dry extract, determined by HPLC. Quercetin

 Quercetin or 2- (3,4-dihydroxyphenyl) -3,5,7-trihydroxy-4H-1-benzopyran-4- one, is a polyphenol that structurally belongs to the flavonoid group and responds to the following formula:

Quercetin is widely distributed in nature, and is present in a variety of plants and fruits. Because of this, quercetin can be obtained as an extract, from various vegetables and fruits such as, for example, green tea and black tea (Camellia sinensis), capers, mountain celery (Levisticum officinale), apples, onion, black grape, citrus, papaya, tomato, broccoli, cauliflower, chili (Capsicum annum), blueberries, or Sophora japan, among others.

 Quercetin can be found commercially as an extract of plant origin with various contents, which may be greater than 98% on the dry extract, determined by HPLC, expressed as quercetin dihydrate. For example, the purified extract of acacia flowers from Japan or tree of pagodas (Sophora Japónica), from the Select Botanical company, is presented in the form of a yellow powder containing quercetin in the form of quercetin dihydrate, with a content of water that is a maximum of 12.5%, and that has a minimum content of quercetin dihydrate of 98% on the dry extract, determined by HPLC, for example as described in the article Beecher et al., Analysis of tea polyphenols , Proc. Soc. Exp. Biol. Med., 1999, 220 (4), 267-70.

 Quercetin is also found as an onion extract (Allium cepa) with varying concentrations between 0.5% and 40%, and as apple, asparagus and grape extracts.

 Quercetin can also be obtained commercially in substantially pure form through suppliers such as Sigma-Aldrich.

 On the other hand, quercetin can be synthesized, for example, as described in the article Shakhova et al., Zh. Obshch Khim., 1962, 32, 390.

 In the context of the present invention, the term quercetin is broadly understood and includes the active ingredient and its possible hydrates and / or solvates. In a preferred embodiment of this invention, quercetin is in the form of dihydrate.

In the combination of the invention, quercetin may be of synthetic or natural origin. When it is naturally occurring, preferably the source of quercetin is a plant extract. More preferably, an extract of plant origin with a high quercetin content is used, such as for example greater than 95% on the dry extract expressed as quercetin dihydrate, and more preferably a japanic Sophora extract, and even more preferably a purified extract of Japan sophora with a content greater than 98% quercetin dihydrate on the dry extract. Catechin

 Catechin is the common name by which it knows the product (2R, 3S) -2- (3,4-dihydroxyphenyl) -3,4-dihydro-2 / - / - chromen-3,5,7-triol, a polyphenol also belonging to the flavonoid group, and which has the following structure:

The catechin molecule has two asymmetric carbons or chiral centers in positions 2 and 3, as indicated in the previous structure and, therefore, there are actually four different stereoisomers: two of them in the cis configuration and two in the configuration trans. The two trans forms correspond to the enantiomer (2R, 3S), represented in the above formula, and to the enantiomer (2S, 3R), in which the configuration of the two asymmetric carbons 2 and 3 is reversed.

 The two enantiomers that are in cis configuration are usually referred to as epicatechin. Thus, the name epicatechin corresponds to the product (2R, 3R) -2- (3,4-dihydroxyphenyl) -3,4-dihydro-2H-chromen-3,5,7-triol, which has the following structure:

As with the trans isomer, epicatechin is understood as the set formed by the two cis isomers, both the enantiomer (2R, 3R) represented above, and the corresponding enantiomer (2S, 3S), in which the configuration of the two asymmetric carbons 2 and 3. Catechins are also found in nature in the form of dimers and trimers, which are known by the name of proanthocyanidins or procyanidins or oligomeric proanthocyanidins.

 In the context of the present invention, any of the 4 stereoisomers mentioned, that is, both the catechin and the epicatechin and their respective enantiomers, is called catechin, any of its possible dimers or trimers in the form of type B proanthocyanidins (dimers formed from catechin and / or epicatechin) or proanthocyanidins type C (trimer formed from epicatechin), dimer in the form of gallate, and any mixture of the above.

 Commercially the source of catechin can be a plant extract with different contents in catechins, which can be equal to or greater than 90% on the dry extract expressed as catechin. For example, the purified extract of Vitis vinifera from the company Select Botanical, which is presented in the form of an orange powder with a water content of at most 8% and with a catechin content of approximately 90% on the dry extract expressed as catechin, determined by spectrophotometry, for example as described in the article Peri et al., An Assay of Different Phenolic Fractions in Wines, Am. J. Enol. Vitic., 1971, 22 (2), 55-58. Said extract contains a mixture of trans-catechin, epicatechin, dimer in the form of gallate, and proanthocyanidins. Other sources of catechins are extracts from green tea, cocoa, green coffee and blueberries.

 Commercially, pure enantiomeric forms of catechin can also be obtained, for example, (2S, 3R) from suppliers such as Sigma-Aldrich.

 In a preferred embodiment of the present invention, catechin (2S, 3R) -2- (3,4-dihydroxyphenyl) -3,4-dihydro-2H-chromen-3,5,7-triol is used in substantially pure form.

In a more preferred embodiment of the present invention, the catechin is the trans isomer, in any of its enantiomeric (2S, 3R) or (2R, 3S) forms, or a mixture of the two in any proportion, and optionally further containing epicatechin in any of its two enantiomeric forms and / or proanthocyanidins, or any mixture of the above in any proportion, preferably obtained from plant extracts. In an even more preferred embodiment, the source of catechin is a grape extract, that is, an extract of Vitis vinifera preferably with a catechin content equal to or greater than 80% on the dry extract expressed as catechin, and even more preferably employs a purified extract of Vitis vinifera with a catechin content equal to or greater than 90% on the dry extract expressed as catechin.

 In the present invention, plant extracts can be used as sources of antioxidants containing different resveratrol, quercetin, catechin, or extracts containing mixtures of two or more of these polyphenols.

 Preferably a plant extract containing substantially pure resveratrol is used, that is, with a resveratrol content greater than 90% over the dry extract, preferably greater than 92%, more preferably greater than 95%, and even more preferably greater than 98% .

 Preferably a plant extract containing substantially pure quercetin is used, that is, with a content greater than 90% on the dry extract, preferably greater than 92%, more preferably greater than 95%, and even more preferably greater than 98% expressed as Quercetin dihydrate

 Preferably, a plant extract containing substantially pure catechin is used, that is, with a catechin content greater than 80% over the dry extract, preferably greater than 85%, and more preferably greater than 90% expressed as catechin.

In a particularly preferred embodiment, the combination of antioxidants comprises resveratrol, quercetin and catechin in a molar ratio resveratrol: quercetin: catechin of approximately 1: 1: 2, the source of resveratrol is a root extract of Polygonum cuspidatum, preferably with a content of trans-resveratrol greater than 95% on the dry extract, the source of quercetin is an extract of Sophora japan, preferably with a content of quercetin dihydrate greater than 98% on the dry extract, and the source of catechin is an extract of Vitis vinifera, preferably with a catechin content equal to or greater than 90% on the dry extract expressed as catechin. In an especially preferred embodiment, the combination of antioxidants comprises resveratrol, quercetin and catechin in a molar ratio resveratrol: quercetin: catechin of approximately 1: 1: 5, the source of resveratrol is a root extract of Polygonum cuspidatum, preferably with a content of trans-resveratrol greater than 95% on the dry extract, the source of quercetin is an extract of Sophora japan, preferably with a content of quercetin dihydrate greater than 98% on the dry extract, and the source of catechin is an extract of Vitis vinifera, preferably with a catechin content equal to or greater than 90% on the dry extract expressed as catechin.

 In another especially preferred embodiment, the combination of antioxidants consists essentially of resveratrol, quercetin and catechin in a molar ratio resveratrol: quercetin: catechin of approximately 1: 1: 2, the source of resveratrol is a root extract of Polygonum cuspidatum, preferably with a content of trans-resveratrol greater than 95% on the dry extract, the source of quercetin is an extract of Sophora japan, preferably with a content of quercetin dihydrate greater than 98% on the dry extract, and the source of catechin is a Vitis vinífera extract, preferably with a catechin content equal to or greater than 90% on the dry extract, expressed as catechin.

 In another especially preferred embodiment, the combination of antioxidants consists essentially of resveratrol, quercetin and catechin in a molar ratio resveratrol: quercetin: catechin of approximately 1: 1: 5, the source of resveratrol is a root extract of Polygonum cuspidatum, preferably with a content of trans-resveratrol greater than 95% on the dry extract, the source of quercetin is an extract of Sophora japan, preferably with a content of quercetin dihydrate greater than 98% on the dry extract, and the source of catechin is a Vitis vinifera extract, preferably with a catechin content equal to or greater than 90% on the dry extract expressed as catechin.

 In a preferred embodiment, the antioxidant combination does not contain routine.

The combination of polyphenols consisting of trans-resveratrol, catechin ((2R, 3S) -2- (3,4-dihydroxyphenyl) -3,4-dihydro-2H-chromen-3,5,7-triol, its enantiomer or mixtures of both), epicatechin ((2R, 3R) -2- (3,4-dihydroxyphenyl) -3,4-dihydro-2H-chromen-3,5,7-triol, its enantiomer, or mixtures of both), Quercetin and rutin in a 1: 1: 1: 1: 1 molar ratio, where each of the polyphenols have a chromatographic purity equal to or greater than 90%, such as those supplied by the Sigma-Aldrich company, is not part of the invention.

Antioxidant power tests

 The antioxidant power of the compositions of the present invention can be determined using an in vitro method designed to assess the free radical scavenging capacity of the tested substances. For example, the method called TOSC (Total Oxidant Scavenging Capacity), which allows the determination of the antioxidant power of compounds at low concentrations.

 With the TOSC method, the antioxidant power of a compound can be determined at a specific concentration, and it can be applied to both solutions with pure antioxidants and complex mixtures.

In the TOSC method, the ability of the substance to be tested to inhibit the oxidation of α-keto-Y-methylthiobutyric acid (KMBA) is determined by the action of free radicals such as, for example, hydroxyl (HO), peroxyl (ROO) or peroxynitrite (ONOO ^" ). As described in Winston et al., A rapid gas chromatographic assay for determining oxyradical scavenging capacity of antioxidants and biological fluids, Free Rad. Biol. Med ., 1998, 24 (3), 480-493, the reaction that takes place is as follows:

H ₃ CS-CH ₂ -CH ₂ -CO-COOH ► H ₂ C = CH ₂ + 1/2 (CH ₃ -S) + C0 ₂

_Udder «leno

Preferably, the TOSC test against peroxyl radicals (ROO) was performed since they are more stable and have a longer half-life than hydroxyl radicals or peroxynitrite radicals. Peroxyl radicals can be obtained by thermal decomposition and symmetric hydrolysis of the 2,2'-azobis (2-methylpropionamidine) compound (ABAP). Thus, the antioxidant power of the tested substances can be measured by the ability to inhibit the formation of ethylene in the presence of peroxyl radicals compared to a control reaction, in which an antioxidant substance is not used as an inhibitor.

 TOSC values are determined according to the methodology described in the articles Regoli et al., Quantification of total oxidant scavenging capacity of antioxidants for peroxynitrite, peroxyl radicáis, and hydroxyl radicáis, Toxicol. Appl. Pharm., 1999, 156, 96-105 and Regoli et al., Total oxidant scavenging capacity (TOSC) of microsomal and cytosolic fractions from Antarctic, Arctic and Mediterranean scallops: differentiation between three potent oxidants, Aquat. Toxicol., 2000, 49 (1-2), 13-25.

Essentially, TOSC values are quantified by measuring ethylene formation, by comparison of the integrated areas for each tested substance and the control substance, according to the following equation:

Control substances, without the ability to eliminate free radicals, have a TOSC value of 0%, since they have the same area under the curve as the control. Compounds that completely prevent the formation of ethylene have an area under the curve equal to 0 and, therefore, a TOSC value of 100%.

 Surprisingly, the authors of the present invention observed that the combination of the resveratrol, quercetin and catechin polyphenols in the molar proportions approximately 1: 1: 2 and approximately 1: 1: 5 show a synergistic anti-radical activity, since values of TOSC for combinations significantly greater than the sum of the TOSC values of its components, as described in Example 3.

 As a result of this synergistic effect, it is possible to obtain a potent antioxidant effect without the need to ingest large amounts of resveratrol, or of the other individual antioxidants, which is advantageous in order to minimize possible risks of side effects.

Compositions

Also part of the object of the invention is a composition comprising the combination of polyphenols of the invention and an acceptable vehicle. The composition is selected from a pharmaceutical composition, a cosmetic composition, a food composition, a dietary supplement, and a veterinary composition, and the vehicle is respectively pharmaceutical, cosmetic, food, dietary, or veterinarily acceptable.

 Pharmaceutical forms suitable for incorporating the combination of polyphenols of the present invention can be formulated for oral, nasal, rectal, parenteral or topical administration. Both solid and semi-solid pharmaceutical forms such as tablets, creams, hard capsules, soft capsules, dragees, powder presentations are suitable; such as liquid or semi-liquid, such as syrups or any type of aqueous, or non-aqueous solution, suspension or emulsion.

 Pharmaceutical compositions are formulated according to the procedures that are known to the expert in pharmaceutical technology, as described, for example, in the book M. E. Aulton, Pharmacy. The science of the design of pharmaceutical forms, second edition, Elsevier, Madrid, 2004. For example, by mixing the antioxidant combination of the invention with at least one pharmaceutically acceptable carrier or excipient and, optionally, with one or more products with therapeutic or dietary functionalities, such as vitamins, minerals, trace elements and other micronutrients.

 The excipients suitable for use in the pharmaceutical compositions object of the present invention are well known to the expert in pharmaceutical technology and are described, for example, in the book R. C. Rowe, P. J. Sheskey and P.J. Weller, Handbook of Pharmaceutical Excipients, fourth edition, Pharmaceutical Press, 2003.

Thus, for example, among the types of excipients and vehicles suitable for use in liquid formulations, in the form of solutions, suspensions or emulsions, are, for example, solvents, buffers, viscosity modifiers, density modifiers, surfactants and emulsifiers, flocculating agents, humectants, preservatives, sweeteners and flavorings, and mixtures of the foregoing. Similarly, among the types of excipients and / or vehicles suitable for use in solid pharmaceutical compositions are, for example, fillers, binders, non-stick agents, lubricants, anti-caking agents, materials for the coating of tablets, gelatin for hard and soft capsules, plasticizers, stabilizers, preservatives, dyes, essences, and mixtures thereof.

 The combination of polyphenols of the invention can also be incorporated into cosmetic compositions for external application, typically for hair or skin care. Cosmetic forms suitable for incorporating the antioxidant combinations of the present invention are, for example, creams, gels, milks, lotions, sprayable emulsions, and solid bars, among others.

 The cosmetic compositions are formulated according to the procedures that are known to the person skilled in the art, by mixing the combination of polyphenols of the invention with at least one cosmetically acceptable carrier in which it is dissolved, emulsified, dispersed, or it is suspended. The vehicle is selected from water, a water miscible non-aqueous vehicle, such as ethanol or isopropanol, and a water non-water miscible vehicle, such as paraffin oil. Optionally, the cosmetic compositions may contain at least one additional cosmetic ingredient, which may be chosen, for example, from the group consisting of surfactants and emulsifiers, lipid compounds and emollients, consistency factors and thickeners, stabilizers, hydrotropes, preservatives, essences, colorants, silicone compounds, fats, waxes, lecithins, phospholipids, UV sun protection factors, film-forming agents, self tanners, or tyrosine inhibitors (depigmentation agents), or mixtures of the above.

 Likewise, the combination of polyphenols of the present invention can be incorporated into food products and dietary supplements, for example in the form of nutraceutical products or functional foods or food supplements for sports nutrition.

 The elaboration of this type of products, incorporating the antioxidant combination of the invention, is carried out by methods known to the expert in food technology.

Any type of food composition or dietary supplement is suitable for incorporating the combination of polyphenols object of the invention, for example, infusions, coffees, drinks in general such as juices or other Alcoholic or non-alcoholic beverages, pastries or bakery products, jams, energy bars, cereals, flours, dairy products such as cheese, milk, butter, yogurts or cream, chocolate-based products and sweets such as candy, chewing gum or ice cream , oils, margarine, preserves.

 Optionally, the food product incorporating the combination of polyphenols of the invention may additionally contain other products with therapeutic or dietary functionalities, preferably, vitamins, minerals, trace elements or other micronutrients, or other plant extracts of polyphenols source.

 Likewise, the food product containing the combination of polyphenols of the invention may additionally contain one or more ingredients, which are chosen from among the usual authorized food additives, well known to those skilled in the art, for example of the type dyes, preservatives , condiments and spices, flavor enhancers, thickeners and gelling agents.

 The combination of polyphenols of the invention can also be incorporated into veterinary compositions intended for animal feed, in particular for the feeding of domestic animals. Such combinations may be incorporated into pet food compositions, such as those described in patent applications EP-A-1514480 and WO-A-2005/1 10037, by methods well known to those skilled in the art. Applications

 The use of the combination for the preparation of a composition with antioxidant power is also part of the invention.

 The composition is selected from the group consisting of a pharmaceutical composition, a cosmetic composition, a food composition, a dietary supplement, and a veterinary composition.

 The combination of the invention for use as an antioxidant is also part of the invention.

The combination of polyphenols object of the present invention can be used therapeutically to prevent or decrease the progression of all those pathological conditions that can be treated with antioxidants and / or free radical scavengers, such as cancer, cardiovascular diseases, inflammatory diseases, coronary diseases, diabetes, Alzheimer's disease, multiple sclerosis or dermatological conditions such as psoriasis or the appearance of eczema , among other.

 The combination of the invention is particularly useful for delaying the symptoms of aging, for its ability to protect organs, tissues and cells against the harmful effects of oxidation. It is also appropriate, for example, to treat premature aging of the skin by preventing the breakdown of collagen fibers and, in general, all tissues.

 The combination of the invention is also particularly suitable for those subjects most likely to generate free radicals in an increased manner in their organism, for example, smokers, people especially exposed to adverse conditions of environmental pollution, ionizing radiation, excessive sun exposure, subject people to stressful situations, or people who practice intense physical exercise.

 The following examples serve to illustrate the invention but should not be construed as limiting the same.

Example 1: Preparation of an antioxidant composition of resveratrol,

 Quercetin and catechin in 1: 1: 2 molar ratio based on resveratrol, quercetin and catechin from plant extracts

In a container equipped with a mixer for solid products, 253.16 g of purified extract of Polygonum cuspidatum root were weighed with a water content of 8% and a content of 98% of resveratrol on the dry extract, equivalent to 1 mol of resveratrol; 386.59 g of purified extract of Sophora japan with a water content of 12.5% and a content of 100% quercetin dihydrate on the dry extract, equivalent to 1 mol of quercetin; and 701, 1 1 g of purified Vitis vinífera extract with a water content of 8% and a catechin content of 90% on the dry extract expressed as catechin, including 12.5% of monomeric catechins, the rest being dimeric procyanidins of type B and trimeric of type C, equivalent to 2 moles of catechin, and the whole was mixed until obtaining a complete homogenization. Example 2: Preparation of an antioxidant composition of resveratrol,

 Quercetin and catechin in 1: 1: 2 molar ratio based on resveratrol and quercetin from vegetable extracts and catechin (2S.3R) In a container equipped with a mixer for solid products 253.16 g of purified extract of root of Polygonum cuspidatum with a water content of 8% and a 98% content of resveratrol on the dry extract, equivalent to 1 mol of resveratrol; 386.59 g of purified extract of Sophora japan with a water content of 12.5% and a content of 100% quercetin dihydrate on the dry extract, equivalent to 1 mol of quercetin; and 580.52 g of catechin (2S, 3R) substantially pure (Sigma), equivalent to 2 moles of catechin, and the whole was mixed until complete homogenization. Example 3: Antioxidant Power Test

 The antioxidant power of the compositions of the present invention was measured by the TOSC method, as described above. Solutions of concentration 1 μ concentración of the substances or mixtures to be tested were used, and distilled water as a control, whose TOSC value is 0%.

 The following solutions were prepared, using distilled water as solvent:

1) Resveratrol (R): 253.16 mg of purified extract of Polygonum cuspidatum root was weighed, as used in Example 1, and dissolved in 1 I of distilled water. 1 ml of the resulting solution was taken and diluted with distilled water to a volume of 1 I. The concentration of resveratrol was 1 μΜ. 2) Quercetin (Q): 386.57 mg of purified Sophora japan extract were weighed, as used in Example 1, and dissolved in 1 I of distilled water. 1 ml of the resulting solution was taken and diluted with distilled water to a volume of 1 I. The concentration of quercetin was 1 μΜ. 3) Catechin (2S.3R) (C): 290.26 mg of substantially pure catechin (2S.3R) (Sigma) was weighed and dissolved in 1 I distilled water. 1 ml of the resulting solution was taken and diluted with distilled water to a volume of 1 I. The catechin concentration was 1 μΜ. 4) Catechin (C _ex tr): 350.56 mg of purified grape seed extract Vitis vinifera was weighed, as used in Example 1, and dissolved in 1 I distilled water. 1 ml of the resulting solution was taken and diluted with distilled water to a volume of 1 I. The catechin concentration was 1 μΜ.

5) Resveratrol-quercetin-catechin R + Q + C _ex tr (1: 1: 2): 0.5 ml of solution R, 0.5 ml of solution Q and 1 ml of solution C _ex tr were taken , and mixed. This solution contained a concentration of 0.25 μΜ of resveratrol, 0.25 μΜ of quercetin and 0.5 μΜ of catechin. The total concentration of polyphenols was 1 μΜ.

6) Resveratrol-quercetin-catechin R + Q + C _ex tr (1: 1: 5): 0.5 ml of solution R, 0.5 ml of solution Q and 2.5 ml of solution C were taken _ex tr, and mixed. This solution contained a concentration of 0.143 μΜ of resveratrol, 0.143 μΜ of quercetin and 0.714 μΜ of catechin. The total concentration of polyphenols was 1 μΜ.

7) Resveratrol-quercetin-catechin (2S, 3R) R + Q + C (1: 1: 2): 0.5 ml of solution R, 0.5 ml of solution Q and 1 ml of solution were taken C, and mixed. This solution contained a concentration of 0.25 μΜ of resveratrol, 0.25 μΜ of quercetin and 0.5 μΜ of catechin. The total concentration of polyphenols was 1 μΜ. 8) Resveratrol-quercetin-catechin (2S, 3R) R + Q + C (1: 1: 5): 0.5 ml of solution R, 0.5 ml of solution Q and 2.5 ml of solution were taken solution C, and mixed. This solution contained a concentration of 0.143 μΜ of resveratrol, 0.143 μΜ of quercetin and 0.714 μΜ of catechin. The total concentration of polyphenols was 1 μΜ.

Ethylene formation during the TOSC test was monitored by gas chromatography, with HP 5890-series II equipment, HP Chemstation 5890 Software, and calculated by determining the area under the curve, using the SRI PeakSimple ^{® program} . Linear regression was calculated using the GraphPad Prism ^{® program} . The slope of the regression was calculated in the linear margin of the TOSC control curve versus the concentration of the assets tested.

The results shown in Table I were obtained.

TABLE I

The column "TOSC calculated" represents the TOSC value that theoretically corresponds to the samples of the different combinations of antioxidants taking into account only an additive effect of each of them. Thus, the calculated value is obtained by multiplying the concentration of each of the antioxidants in the sample by the experimental TOSC of each of the individual antioxidants and dividing by the concentration of the antioxidant in the individual sample. For example in the case of sample 5:

 TOSC calculated = (0.25 μΜ R x 22.00 / 1 μΜ) + (0.25 μΜ Q x 1 1.00 / 1 μΜ) + + (0.5 μΜ C x 8.00 / 1 μΜ) = 12.25%

 The "Increment" column corresponds to the difference between the calculated and experimental TOSC value, and allows quantifying the synergistic effect observed in the antioxidant power of the combinations. It can be seen that the combination of resveratrol, quercetin and catechin of the invention, both in the 1: 1: 2 molar ratio, and in the 1: 1: 5 molar ratio, shows a remarkable synergistic effect. Likewise, it can be observed that, although the synergistic effect with pure catechin (2S, 3R) is slightly higher than that of catechin from grape extract, a comparable synergistic effect can be appreciated in both cases, independent of the source of Catechins used.

 The "Potency" column reflects the number of times the antioxidant power increases with the combinations object of the invention compared to the antioxidant power obtained from the separate addition of the antioxidant powers of resveratrol, quercetin and catechin.

Example 4: Preparation of a food supplement for the prevention of skin aging

Soft gelatin capsules (8/10 oval) of red color were prepared with the following filling per capsule:

Component Quantity

 (mg)

Purified extract of Polygonum cuspidatum root from Example 1 33.53

Purified extract of Japan Sophora from Example 1 52,18

Purified grape seed extract of Example 1 94.65

D acetate, L-a-tocopherol 10.00

Borage oil (> 20% γ-linoleic acid) 150.00

Hydrogenated soybean fat 10.00 Beeswax 5.00

The molar ratio resveratrol: quercetin: catechin in this preparation is 1: 1: 2.04. Example 5: Preparation of a food supplement for the improvement of the prevention of age-related macular degeneration

Soft gelatin capsules (14/16 oblong) of black color were prepared with the following filling per capsule:

The molar ratio resveratrol: quercetin: catechin in this preparation is 1: 0.97: 1, 98. Example 6: Preparation of a food supplement for repair of expression wrinkles Envelopes were prepared with the following filling per envelope:

The molar ratio resveratrol: quercetin: catechin in this preparation

1: 1: 2.

Example 7: Preparation of a functional coffee-based food for the prevention of cardiovascular disease

Single-dose sachets were prepared with the following filling per envelope: Component Quantity

Decaffeinated coffee 30.00 g

Purified extract of Polygonum cuspidatum root from Example 1 37.61 mg

Purified extract of Japan Sophora from Example 1 58.62 mg

Purified grape seed extract of Example 1 261, 16 mg

Aroma of moka 50.00 mg

Citric acid 100.00 mg

The molar ratio resveratrol: quercetin: catechin in this preparation is

1: 1: 5.

Example 8: Preparation of a food supplement based on natural substances as an antioxidant in sports nutrition

Soft gelatin soft capsules (20 oblong) of purple color were prepared with the following filling per capsule:

Component Quantity

 (mg)

Coenzyme Q10 100.00

Purified Polygonum cuspidatum root extract of Example 1 37.61

Purified extract of Japan Sophora from Example 1 58.62

Purified grape seed extract of Example 1 261, 16

RRR-alpha-tocopherol (67%) 15.00

Vitamin C 60.00

Beta-carotene dunaliella (95% retinol) 30.00

Wheat Germ Oil 500.00 Hydrogenated soybean fat 20.00

Beeswax 10.00

The molar ratio resveratrol: quercetin: catechin in this preparation is

1: 1: 5.

Example 9: Preparation of a nutritional supplement for the improvement of the appearance of the skin and brightness of the fur in pets (mammals)

Soft gelatin capsules (18/20 oblong) of red color were prepared with the following filling per capsule:

The molar ratio resveratrol: quercetin: catechin in this preparation is 1: 1: 5.09.

Example 10: Preparation of a nutritional supplement to improve the fertility of stallions (mammals) Soft gelatin capsules (22 oblong) of red color were prepared with the following filling per capsule:

The molar ratio resveratrol: quercetin: catechin in this preparation is

1: 1: 5.

Claims

1. - Synergistic combination of polyphenols comprising resveratrol, quercetin and catechin, characterized in that the molar ratio between resveratrol, quercetin and catechin is 0.9-1, 1: 0, 9-1, 1: 1, 8-2.2 or 0.9-1, 1: 0, 9-1, 1: 4, 5-5, 5, with the proviso that the combination of polyphenols consisting of trans-resveratrol, catechin, epicatechin, quercetin and routine in one 1: 1: 1: 1: 1 molar ratio, where each of the polyphenols have a chromatographic purity equal to or greater than 90% is expressly excluded. 2. - Combination according to claim 1, characterized in that the molar ratio is 1: 1: 2 or 1: 1: 5. 3. - Combination according to claim 2, characterized in that the molar ratio is 1: 1: 2. 4. - Combination according to claim 2, characterized in that the molar ratio is 1: 1: 5. 5. Combination according to any one of claims 1 to 4, characterized in that the resveratrol is trans-resveratrol. 6. - Combination according to any one of claims 1 to 5, characterized in that the resveratrol is of natural origin. 7. - Combination according to claim 6, characterized in that the source of resveratrol is a plant extract. 8. - Combination according to claim 7, characterized in that the plant extract is a root extract of Polygonum cuspidatum. 9. - Combination according to any one of claims 1 to 8, characterized in that the quercetin is of natural origin. 10. - Combination according to claim 9, characterized in that the source of quercetin is a plant extract. eleven . - Combination according to claim 10, characterized in that the plant extract is an extract of Sophora Japan. 12. - Combination according to any one of claims 1 to 1 1, characterized in that the source of catechin is a plant extract. 13. Combination according to claim 12, characterized in that the plant extract is an extract of Vitis vinífera. 14. - Combination according to claim 1, characterized in that the resveratrol: quercetin: catechin molar ratio is 0.9-1, 1: 0.9-1, 1: 1, 8-2.2, the source of resveratrol is a Polygonum cuspidatum root extract, the source of quercetin is an extract of Sophora japan, and the source of catechin is an extract of Vitis vinifera. 15. - Combination according to claim 1, characterized in that the resveratrol: quercetin: catechin molar ratio is 0.9-1, 1: 0.9-1, 1: 4.5-5.5, the source of resveratrol is a Polygonum cuspidatum root extract, the source of quercetin is an extract of Sophora japan, and the source of catechin is an extract of Vitis vinifera. 16. Composition comprising the combination according to any one of claims 1 to 15 and an acceptable vehicle. 17. - Composition according to claim 16, characterized in that the composition is pharmaceutical and the vehicle is pharmaceutically acceptable. 18. - Composition according to claim 16, characterized in that the composition is cosmetic and the vehicle is cosmetically acceptable. 19. Composition according to claim 16, characterized in that the composition is a dietary supplement and the vehicle is dietically acceptable. 20. Composition according to claim 16, characterized in that the composition is food and the vehicle is food acceptable. twenty-one . - Composition according to claim 16, characterized in that the composition is veterinary and the vehicle is veterinarily acceptable. 22. - Use of the combination according to any one of claims 1 to 15 for the preparation of a composition with antioxidant power. 23. Combination according to any one of claims 1 to 15 for use as an antioxidant.