EP2496206A2

EP2496206A2 - Composition comprising an extract from polygonum cuspidatum and a polyphenolic extract from red vitis vinifera

Info

Publication number: EP2496206A2
Application number: EP10798587A
Authority: EP
Inventors: Ettore Bergamini; Gabriella Cavallini; Sara Straniero
Original assignee: Functional Point Srl; Gori Zina
Current assignee: Functional Point Srl; Gori Zina
Priority date: 2009-11-05
Filing date: 2010-11-04
Publication date: 2012-09-12
Also published as: ITMI20091940A1; WO2011055222A3; WO2011055222A2

Abstract

Composition comprising an extract of polygonum cuspidatum and a polyphenolic extract of red vitis vinif era.

Description

COMPOSITION COMPRISING A WATER-SOLUBLE EXTRACT OF POLYGONUM CUSPIDATUM AND A POLYPHENOLIC EXTRACT OF RED VITIS VINIFERA AND USES THEREOF.

DESCRIPTION

The present invention relates to a composition comprising an extract of polygonum cuspidatum and a polyphenolic extract of red vitis vinifera.

It is commonly known that human cells and tissues are continuously exposed to the damaging effects of reactive substances such as free radicals, which may be endogenous (products of cellular metabolism or of external origin (high concentrations of oxygen, generators of radicals present in the environment, including smoke, sunlight and the like) and which are responsible for acute or chronic pathological conditions, such as, in particular, ageing, of an intrinsic nature (caused by the free radicals generated by the normal cellular oxidative metabolism, and of an extrinsic nature (produced by free radicals generated by environmental factors outside the organism) .

The ageing process is in fact a natural phenomenon caused by the accumulation, in old cells, of altered molecules and organelles, said accumulation producing a consequent reduction of the metabolism and the functions of all the cells and all the tissues .

This reduction is determined by the speed of said accumulation, which varies from person to person depending on the respective cellular defences, and increases with age, in all individuals, since the generation of free radicals increases with the number of altered mitochondria and peroxisomes which, in turn, increase with age, while the cellular defences decline.

Slowing down of the functions of most of the cells, which has a very important role in maintaining the integrity of the structure of the tissues, starts to occur generally at around the age of 30 years and may depend on many factors, such as the genetic constitution, general state of health, life style and environment (e.g. Masoro E (ed) Handbook of Physiology 11: Aging. 1995) .

It is also known that nutrition and physical exercise may also improve (or negatively affect) the functionality of autolysis which forms the main defence mechanism of cells and that the antioxidants of so-called functional foods may protect the cellular membranes from the effects of a reduction in the antioxidant defences at an advanced age .

The definition of "functional foods" may be applied, among other things, to plant derivatives which naturally contain products which are useful for human health and are capable of preventing illnesses (Boue SM, Cleveland TE, Carter-Wientjes C, Shih BY, Bhatnagar D, McLachlan JM, Burow ME. Phytoalexin-enriched functional foods. J Agric Food Chem. 2009 Apr 8;57(7): 2614-22).

These substances are known to include stilbenes which have been recently been shown to be effective in stopping the cancerogenous process, modifying initiation, development and progression thereof (Pervaiz S, Holme AL. Resveratrol : Its Biological Targets and Functional Activity. Antioxid Redox Signal. 2009 May 11. [Epub ahead of print]) .

Stilbenes include resveratrol (trans-3 , 4 ' , 5- trihydroxystilbene) , a natural, non-flavonoid, polyphenolic antioxidant derived from many plants, including grapes, berries, peanuts and contained in wine, in particular red wine.

The plants which contain resveratrol in high concentrations include also Polygonum Cuspidatum. {Rocha-Gonzalez HI, Ambriz-Tututi M, Granados-Soto V. Resveratrol: a natural compound with pharmacological potential in neurodegenerative diseases. CNS Neurosci Ther. 2008 Fall ; 14 (3): 234- 47. Epub 2008 Jul 29)

Resveratrol is considered to be a promising natural bioactive molecule with phytotherapeutic and pharmacological potential (Shils ME, Shike M, Ross AC, Caballero B, Cousins RJ Modern Nutrition in Health and Disease, 10^th ed, p. 1796, Lippincott Williams & Wilkins, Baltimore, 2006) .

During the last few years resveratrol has been the subject of many in vitro and in vivo investigations aimed at understanding biological activities, including anti-oxidant and anti-inflammatory, anti-aggregating, anti -atherogenous , phyto- estrogenous, cell-proliferation-inhibiting and immunomodulating activities (Pervaiz S, Holme AL. Resveratrol: Its Biological Targets and Functional Activity. Antioxid Redox Signal. 2009 May 11. [Epub ahead of print]). Resveratrol provides protection against diabetes (Subramanian S Resveratrol, a natural phytoalexin, normalizes hyperglycemia in streptozotocin-nicotinamide induced experimental diabetic rats. Bio ed Pharmacother. 2008 Nov; 62 (9) : 598 -605) and neurodegeneration {Rocha- Gonzalez HI, Ambriz-Tututi M, Granados-Soto V. Resveratrol : a natural compound with pharmacological potential in neurodegenerative diseases. CNS Neurosci Ther. 2008 Fall ; 14 (3) :234- 47. Epub 2008 Jul 29) .

Resveratrol may also have cardioprotective and chemopreventive properties.

During the last two decades resveratrol and similar polyphenols have been very much at the centre of attention, resulting in the publication of a large number of articles which have highlighted the beneficial effects in many human illness models and in cardio and neuroprotection, immune regulation and chemoprevention of neoplasia (Pervaiz S, Holme AL. Resveratrol: Its Biological Targets and Functional Activity. Antioxid Redox Signal. 2009 May 11) .

In 2003, David Sinclair and his team at Harvard have added the increase in longevity to the list of potential benefits of the substance, publishing in Nature the fact that resveratrol increases the survival of yeast cells. Since then studies carried out on worms, fruit flies, fish and rats have established a link between resveratrol and longevity. Other studies have demonstrated the anti -tumoral and anti - inflammatory effects. Other very recent studies have shown that resveratrol lessens in a reproducible manner the oxidative damage of tissues with a high age-related oxidative damage and reproduces the effects of calorific restriction of mammals (Wong YT, Gruber J, Jenner AM, Ng MP, Ruan R, Tay FE. Elevation of oxidative- damage biomarkers during aging in F2 hybrid mice: protection by chronic oral intake of resveratrol. Free Radic Biol Med. 2009 Mar 15; 46 (6) : 799-809) .

The attribution of all the beneficial effects to resveratrol alone is, however, not possible and the evaluations as to the amounts of pure resveratrol which are necessary for obtaining the advantages described above appear to cast doubt on such an association .

It must also be underlined that the beneficial effects of pure resveratrol were observed in vitro using amounts which were far higher (tens of milligrams per Kg of body weight) than those which proved to be effective when taken together with functional foods.

Considering that a glass of wine contains micrograms of resveratrol and that daily consumption of wine together with all kinds of food results in an amount ranging between 5 and 10 mg/day, it is clear that what happens in real life is very different from that which occurs in most of the studies on animals where hundreds of milligrams of pure resveratrol are used.

The use of pure resveratrol in high concentrations increases, however, the risk of toxicity of the resveratrol itself since in high concentrations it may have pro-oxidant properties depending on the doses (de la Lastra CA, Villegas I. Resveratrol as an antioxidant and pro-oxidant agent: mechanisms and clinical implications . Biochem Soc Trans. 2007 Nov;35(Pt 5).-1156-60) .

In high doses resveratrol may in fact strengthen the death signals, depressing the redox proteins and strengthening the apoptotic proteins {Dudley J, Das S, Mukherjee S, Das DK. Resveratrol , a unique phytoalexin present in red wine, delivers either survival signal or death signal to the ischemic myocardium depending on dose. J Nutr Biochem. 2009 Jun;20 (6) : 443 -52) .

The studies carried out have also shown that resveratrol is a phytoalexin, i.e. a defensive substance produced by plants in response to environmental stresses and pathogenic microorganism infections; in basic conditions resveratrol is absent or present in very low concentrations which may increase in a significantly rapid manner in the case of a more general defensive response (Zamboni A, Gatto P, Cestaro A, Pilati S, Viola R, Mattivi F, Moser C, Velasco R. Grapevine cell early activation of specific responses to DIMEB, a resveratrol elicitor BMC Genomics . 2009 Aug 6_/10(1) :363. [Epub ahead of print])} it follows that resveratrol is an additional defence which operates always in the presence of - and most probably in co-operation with - other defences which are always present in the cells.

Also known are the following:

FR2867977: cosmetic, pharmaceutical and derrao- pharmaceutical compositions (not in the form of tablets) for combating wrinkles and skin lines, containing resveratrol (0.001-40%) in water- insoluble form or vegetable derivatives and extracts or mixtures thereof, which are active owing to the relaxing action of these compositions on the skin muscles.

FR2923717: compositions containing derivatives obtained by means of alkylation of at least one stabilized phenolic function by means of esterification with mainly unsaturated fatty acids of polyphenols and stilbenes extracted from vitis vinifera or polygonum cuspidatum which are used for combating illnesses and ageing in human beings and in particular for cosmetic, dietary and therapeutic applications.

WO02/081651: compositions which comprise cardioprotective fibrinolytic agents in the form of tablets, beverages or alcohol -free integrators containing catechin (5-1000 mg) , epicatechin (2- 1000 mg) , quercetin (2-1000 mg) , resveratrol (1-50 mg) or their complexes or metabolic or synthetic derivatives, singly or in combination, capable of increasing the fibrinolytic activity.

WO2004/006881 : cosmetic preparations in the form of a cream, ointment, emulsions, liposomes, gel, lotions and sprays which are able to clarify the skin and eliminate blemishes and which contain epsilon-viniferin optionally in combination with resveratrol and polyphenols in oligo or polymer form.

WO2007/146318 : describes fluid compositions containing trans-resveratrol and polyphenols able to provide users who cannot tolerate alcohol with beverages which are alcohol -free or have a low alcohol content and which contain trans -resveratirol and polyphenols in concentrations comparable to or greater than those of red wine in packaging able to protect the trans-resveratrol from oxidation by the oxygen in the atmosphere .

The document also describes beverages containing trans-resveratrol in concentrations ranging from 5 to 300 mg/1 with or without stabilizers, vitamin A,. E and C, substances which absorb UV rays, one or more polyphenols, . proteins, minerals, tea, coffee and aromas .

WO2008/114141 : compositions capable of inhibiting 5 -alpha-reductase , containing a combination of omega- 6 fatty acids and rutin in concentrations ranging between 0.1 and 15%, in a suitable vehicle, useful for regulating nutrition of the hair bulbs and the production of sebum, formulated as a pharmaceutical, dietary or cosmetic product for treating hair loss.

WO2009/53802 : alimentary alcohol-free beverages for anti-ageing use, containing trans-resveratrol, citric acid, fructose, polyphenols extracted from black grape, extract of green tea, vitamin E, vitamin C and niacin, associated with edible-fruit juices ; US2004/02412261 (EP 1284133), products which exploit the effects of combinations in synergic proportions of the following natural anti -oxidants : a) polyphenols, in particular catechols; b) quercetin; c) resveratrol, able to develop a particularly active action as inhibitors of metalloproteinases . The preparations, in addition to the combination of pure substances, may be derived from the combination of extracts - of an aqueous or hydroalcoholic nature obtained via conventional techniques - of the following:

a) grape seeds (rich in catechols - max. 15%) ;

b) vine leaves (rich in quercetin - max. 5%) ;

c) grape skin (rich in resveratrol, max. 5%), mixed together in the ratios 1 : (0.1-10) : (0.1-10) so as to form each of the three components from 0.1 to 25% by weight of the preparation.

The products may therefore contain up to 75% by weight of the mixture of the three extracts and contain quantities of resveratrol less than 5% by weight .

XP-002593863 : cosmeceutical products derived from grass and also extracts of grape seeds (vitis vinifera) and active principles contained therein, including resveratrol; XP-002593864 : this deals with the protective action of many vegetable phenolic compounds against photo- ageing and UV-ray skin cancer, summarising the results achieved in- the sector up to 2003. More generally it describes the beneficial dermatological effects of grape seed extracts.

All of the prior art, although referring to the use of resveratrol and polyphenols for various pharmacological, dermatological and anti -ageing cosmetic applications, therefore describes compositions mainly obtained from the preliminary selection of natural active principles considered to be specifically active, extracted from grapes, using various methods, so as to provide special formulations of the said active principles with the aim of obtaining specialized ad hoc effects.

The technical problem which is posed, therefore, is that of developing a composition which comprises resveratrol in combination with other components, able to produce a defensive action in the human body against cell and tissue ageing, without the negative consequences of an increase in the degenerative effects of ageing. These results are achieved according to the present invention by a composition according to the characteristic features of Claim 1.

Further details may be obtained from the following description of non-limiting examples of embodiment of the composition according to the present invention.

The composition according to the present invention comprises an extract of polygonum cuspidatum containing trans-resveratrol

and a polyphenolic extract of red vitis vinifera in relative proportions ranging between 1% and 25% by weight .

For the purposes of the present patent the expression "extract" is understood as meaning a phyto-composite comprising all the components present in red vitis vinifera with their natural balance .

Preferably extraction is performed using a hydroalcoholic method which does not introduce chemical molecules which are damaging for human health .

This definition therefore includes polyphenols which are produced naturally (for example by means of extraction from red wine or red grape) , but not the polyphenols obtained synthetically.

Preferably, the hydrosoluble extract of polygonum cuspidatum has a content of trans-resveratrol of not less than 20%.

It is envisaged, moreover, that the composition comprises trans-resveratrol added in an amount ranging between 0.1 and 5% by weight of the content of polyphenolic extract of red vitis vinifera.

The composition according to the present invention may also be associated with ubiquinol and/or ubiquinone and/or liposoluble vitamins with an anti -radical action such as vitamen E and vitamin F (Omega 3) which improve the absorption and stabilize and prolong the protective effects of the composition .

In applications for cosmetic and/or dermatological use the compositions may be associated with dolichol and/or with vegetable polyprenols from which dolichol may be synthesized, in order to improve protection of the skin against the acute and chronic damage produced by exposure to sunlight .

The composition according to the invention may be administered both systemically, preferably orally, and topically.

Preferably said oral form may be formulated in the form of a capsule, coated or uncoated tablet, granular product or emulsion; the topical form may in turn be preferably formulated as a cream, gel or ointment .

Preferred embodiments of the composition are described below in non-limiting examples thereof with reference to different methods of administration :

CAPSULES

Polyphenolic extract from red

vitis vinifera g 0.90

Extract from polygonum cuspidatum g 0.15

Resveratrol added g 0.002

Vitamin E g 0.04

Vitamin C g 0.05

CREAM

Glyceryl monostearate g 16

Cetyl alcohol g 1

Mineral oil g 10

Sesame oil g 9

Glycerin g 7

Vitamin E g Polyphenolic extract from red

vitis vinifera g 2

Extract from polygonum cuspidatum g 0.4

Resveratrol added g 0.05

Water g 54.95

EMULSION FOR CONDIMENTS

Extra virgin olive oil g 5

Fish and/or walnut and/or linseed oil g 3

Vitamin E g 0.05

Red wine vinegar g 0.5

Ethyl alcohol g 0.5

Extract from polygonum cuspidatum g 0.1

Polyphenolic extract from red

vitis vinifera g 0.4

Vitamin C g 0.05

NaCl g 0.05

KC1 g 0.05

Water g 0.3

The results obtained from the experimental tests carried out are described below in the examples of application of the composition according to the invention . EXAMPLE 1

Protection from photo- lipoperoxidat ion with preloading of polyphenols extracted from red wine The rat liver cells isolated according to Seglen (Seglen PO, Preparation of isolated liver cells. Methods Cell Biol 1976; 13: 29-83) were incubated and irradiated (for 20 or 40 minutes) as described in Parentini I, Bergamini E, Cecchi L, Cavallini G, Donati A, Maccheroni M,Tamburini I, Gori Z. The effect of carbon tetrachloride and ultraviolet radiation on dolichol levels in liver cells isolated from 3- and 24 -month-old male Sprague- Dawley rats. Biogerontology. 2003; 4:365-70).

The preloading was carried out incubating the isolated liver cells (20 mg/ml) in 24 ml of solutions containing the polyphenols (RWP) in the final concentration of 0 , 50, 250 and 500 mg/100 ml for 10 minutes.

At the end, the cells were centrifuged, separated from the supernatant and re-suspended in the final concentration of 6 mg/ml.

50 ml of the suspension were poured onto 12x12 cm Petri dishes so as to obtain by means of sedimentation cellular monolayers which were photo- irradiated for 20 or 40 minutes as described in Parentini I, Bergamini E, Cecchi L, Cavallini G, Donati A, Maccheroni M,Tamburini I, Gori Z. The effect of carbon tetrachloride and ultraviolet radiation on dolichol levels in liver cells isolated from 3- and 24 -month-old male Sprague- Dawley rats. Biogerontology. 2003; 4:365-70). The cells were then separated by means of centrifuging and 1 ml amounts of the incubation means were sampled for the Malondialdehyde (MDA) content as described in Guarini M, Stabile A, Cavallini G, Donati A, Bergamini E. Effects of oxidative stress on the Dolichol content of isolated rat liver cells. Free Radic Res. 2007; 41:1283-8 and shown in Figure 1.

The results are shown in the bar chart of Fig. 1, expressed as nmols of MDA/mg cells against the irradiation time.

The standard error (±) of the mean of 5 samples is shown. The results show that, in our experimental conditions, the production of MDA is linearly dependent upon the duration of irradiation and that preloading of the cells with RWP (Red Wine Polyphenols) has a protective effect which, in turn, is dependent upon the concentration of the RWPs used during preloading.

EXAMPLE 2

Effects of preloading with pure resveratrol isolated rat liver cells incubated and irradiated for 40 minutes.

The methods of dosage of the MDA production are similar to those described in Example 1. In the graph of Fig. 2 which shows the standard error (±) of the mean of 8 samples, the MDA production values obtained, illustrated as described for Figure 1, show that preloading with pure resveratrol in a low concentration (0.7xlO^~6M) surprisingly increases by a highly significant amount the production of MDA (P<0.001) caused by photoirradiation, instead of inhibiting it as one might have expected.

The increase in the photo- lipoperoxidation disappears only with preloading using higher concentrations of resveratrol (20x10 ^~6 M) , which are difficult to achieve in vivo.

EXAMPLE 3

Effects of preloading with RWP (at the concentration of 250 mg/100 ml) and resveratrol (at the concentrations of 0.7; 2; 6 and 20 xlO^"6 M) for

10 minutes on the production of MDA caused by photoirradiation for 40 minutes.

The techniques for isolation, preloading and irradiation of the cells and dosage of the MDA production are again those described in connection with Example 1.

As illustrated in Fig. 3 which shows the standard error (±) of the mean of 8 samples, the MDA production values obtained, expressed in nmols/mg cells, show that, in the presence of polyphenols of red wine, the resveratrol always has (both in low concentrations and in high concentrations) a significant capacity for protection against photo- lipoperoxidation .

It is therefore clear how the composition according to the invention is determined by the combination of complex natural extracts, naturally balanced both individually and in combination, without preliminary separation and recombination of the active components as indicated by the prior art.

In addition to this, the choice of polygonum cuspidatum and vitis vinifera for obtaining the extracts according to the present invention provides a selection thereof such as to determine the special relative weight ratios of resveratrol , and complex polyphenols considered to be effective for the predefined aims, avoiding pretreatment and recombination which may give rise to alteration of the active components with a consequent reduction in their effectiveness.

In addition interaction between the components which could alter in an unexpected manner the action thereof is avoided.

Although described in connection with certain constructional forms and certain preferred examples of embodiment of the invention, it is understood that the scope of protection of the present patent is defined solely by the following claims.

Claims

1. Composition comprising an extract of polygonum cuspidatum and a polyphenolic extract of red vitis vinifera in a relative proportion of between 1% and 25% by weight.

2. Composition according to Claim 1, characterized in that the extract of polygonum cuspidatum has a content of trans-resveratrol of not less than 20%.

3. Composition according to Claim 1, characterized in that it comprises trans-resveratrol added in proportion to the content of extract of red vitis vinifera ranging between 0.1 and 5% by weight.

4. Composition according to Claim 1, characterized in that said vitis vinifera is picked at a latitude lower than 45°N/S.

5. Composition according to Claim 1, characterized in that it comprises ubiquinol and/or ubiquinone.

6. Composition according to Claim 1, characterized in that it comprises liposoluble vitamins with anti -radical action such as vitamin E and/or vitamin F (Omega 3) .

7. Composition according to Claim 1, characterized in that it comprises dolichol and/or vegetable polyprenols from which dolichol may be synthesized.

8. Composition according to one of the preceding claims, characterized in that it may be administered orally.

9. Composition according to Claim 8, characterized in that it is formulated in the form of a capsule or tablet, which may be coated or uncoated, a granular product or emulsion.

10. Composition according to one of the Claims 1 to 7, characterized in that it may be administered topically.

11. Composition according to Claim 1, characterized in that it is formulated in the form of a cream, gel or ointment .

12. Composition according to Claim 1 for dietary use .

13. Composition according to Claim 1 for anti- ageing dermatological use.