WO2012063198A1

WO2012063198A1 - Compositions for the prevention and treatment of erectile dysfunction and impotence and for improving sport performance

Info

Publication number: WO2012063198A1
Application number: PCT/IB2011/054975
Authority: WO
Inventors: Francesco Di Pierro; Giulia Federica Merizzi
Original assignee: Medestea Biotech S.P.A.
Priority date: 2010-11-08
Filing date: 2011-11-08
Publication date: 2012-05-18
Also published as: IT1402672B1; ITTO20100888A1

Abstract

The present invention relates to compositions comprising active principles of vegetable origin whether or not combined with nitric oxide promoters and/or protectors for the prevention and treatment of erectile dysfunction and impotence and for improving sports performance.

Description

Compositions for the prevention and treatment of erectile dysfunction and impotence and for improving sports performance

Technical background

The physiology of male sexual function can be summarized schematically in 5 basic phases: libido, erection, ejaculation, orgasm, detumescence. Among these, no phase guarantees the next: libido may not be followed by erection (impotence), erection may not be followed by ejaculation (retrograde ejaculation), ejaculation may not be followed by orgasm (anorgasmy), orgasm may not be followed by detumescence (priapism). Moreover, cases have been described of ejaculation and/or orgasm in the absence of erection, demonstrating that some phases may be autonomous and not necessarily connected with the preceding phases. Among the various disorders that can affect the sphere of male sexuality, erectile dysfunction, so-called impotence, is the commonest problem (affecting between 10 and 35% of the male population) as well as the most feared (for obvious reasons of a psychological nature). According to recent estimates, impotence is due to an endocrine disorder in 5% of cases, to a psychogenic disorder in a percentage of cases from 10 to 20% and to an organic problem, essentially of a circulatory character, in all other cases (75- 85%). Whereas impotence associated with endocrine disorders can be tackled exclusively with hormone therapy based on testosterone and psychogenic impotence is the preserve of the psychologist, it is nevertheless true that organic impotence with a vascular basis can be tackled with therapy that restores the potentialities of the damaged circulatory function.

The implications of a circulatory nature connected with impotence become clear both on examining the detailed structure of the penis and the physiology of erection, and considering data showing that subjects with atherosclerosis or chronic diabetes (and therefore characterized by severe vascular impairment) are particularly exposed to the risk of suffering from erectile dysfunction. The erectile tissue of the penis consists of two "corpora cavernosa" located on the back of the penis and of a "corpus spongiosum" that surrounds the urethra and forms the gland. These structures consist essentially of spongy tissue that is bounded by a thin fibrous sheet called the "tunica albuginea". The phenomenon of erection results from vasodilatation of the arteries of the corpora cavernosa following relaxation of the smooth muscles. This relaxation is due to the action of cyclic guanosine monophosphate (cGMP) on the parietal myocytes of the arteries of the corpora cavernosa. This substance, released in its turn by the tissue guanylate cyclases stimulated by nitric oxide, is then responsible for opening of the vessels and, consequently, for the considerable flow of blood which, entering the corpora cavernosa and the trabecular spaces, causes the penis to become rigid (tumescence). Finally, stiffening of the penis is ensured by the expansion of the trabecular walls against the tunica albuginea which, compressing the subtunical venules, prevents venous return which, at the appropriate time, will cause detumescence.

It is therefore clear from this analysis that any disorder or alteration that can affect the functionality of the vascular system (e.g. reducing the contractility of the parietal myocytes, damaging the endothelial structures or inducing fibrosis of the corpora cavernosa) is in actual fact able to reduce the potentialities of the erectile mechanism.

Accordingly, impotence of vascular aetiology requires a therapeutic approach that tackles the changes of the circulatory phenomena.

A closely related area, at least from the circulatory standpoint, is that of an athlete's sports efficiency, which can be designated more simply as performance. This is influenced, in its physiology, by the general circulatory capacity with particular reference to the circulatory capacity at the level of the striated skeletal musculature. In this connection, functional ingredients that are able to protect the tissues and the endothelia from production of excesses of oxygen free radicals (ROS), to promote arteriolar pulsatility, and therefore the supply of nutrients and oxygen, to favour vasodilatation and blood supply and to protect the connective tissues from fibrosis, in its turn able to limit the reactivity and quickness of the muscles as well as the capacity for resistance to stress, produce, as has been observed and is described below, an increase in sports performance. Accordingly, formulations that have been shown to be capable of this dual function, i.e. of having a positive influence on problems of erectile dysfunction and on sports performance, contain the active principles stated below.

Leuco-anthocyanins of Vitis vinifera

The so-called "French paradox" is a phenomenon that is well known to the scientific community throughout the world. According to this paradox, although having a high-fat diet, the French appear to be extraordinarily protected against cardiovascular diseases. Epidemiological studies in this connection have emphasized that this paradox can be explained by the chemistry of red wine: the latter, which is well present in the French diet and is extremely rich in polyphenols with extremely high antioxidant power, is said to be responsible for targeted anti-atherosclerotic action.

According to the most recent studies, the presence of high blood levels of low-density lipoproteins (LDL), caused for example by a high-fat diet, would be a necessary but not sufficient condition for generating an atheromatous lesion. The latter instead would only be generated following oxidation of the LDLs which, thus modified, would engulf the circulating monocytes, transforming them into so-called "foam cells", a keystone in the aetiology of atherosclerosis. The role of antioxidants in this connection would then be clear: limiting and/or opposing the oxidation of the LDLs would block any incipient process which, beginning with formation of an atheromatous plaque, leads to various forms of pathology (infarct, stroke etc.).

A long series of studies, still in progress, has made it possible to characterize, from the chemical and pharmacological standpoint, these polyphenols, chemically leuco- anthocyanins, i.e. procyanidolic oligomers derived from the condensation of monomer units of flavan-3-ols and flavane-3,4-diols, free or esterified with gallic acid, having the following properties:

- extremely high antioxidant activity (they act at molarity 50 times lower compared with vitamin E);

- non-competitive inhibitory activity of enzymes such as xanthine oxidase (which generates the free radicals implicated in damage of the endothelial walls) and of enzymes such as elastase, collagenase, hyaluronidase and beta-glucuronidase (which regulate the turnover of the components of the extracellular matrix that surrounds the capillary walls);

- capillary protective properties that are able to bind non-specifically the fibres that make up the walls of the vessels;

- good bioavailability following oral administration and tropism for the cardiovascular bed and for all those tissues, such as the walls of arteries, that are particularly rich in glycosaminoglycans.

It can be concluded that the leuco-anthocyanins of Vitis vinifera - just like the procyanidolic oligomers from maritime pine bark, proanthocyanidins and anthocyanins from species of the genus "vaccinium", which are closely related chemically - constitute a valid tool for the prevention and treatment of vascular disorders and of all those situations normally associated with problems that affect the vasal structures and the extravasal matrix.

Standardized extract of Ginkgo biloba One of the aspects that most characterizes the aetiology of erectile dysfunction is reduced capacity for blood perfusion at the level of the arterioles of the corpora cavernosa, apparently due to phenomena of fibrosis or to the contraction of the smooth muscles of the arterial compartment. Ginkgo biloba extract is characterized by the presence of some active principles that have potent vasomotor action, which act by increasing blood perfusion, both central and peripheral: these are ginkgo-flavone glycosides (24%) and terpenoids (6%), in particular ginkgolides and bilobalides, which have been investigated in particular for their anti-PAF (Platelet Aggregating Factor) capacity. Experiments conducted on the isolated aorta, submitted to perfusion, have identified how the extract exerts its action on the tunica media of the arterial wall owing to direct interaction with parietal myocytes, improving both the phasic and the tonic component of myocyte contraction that normally develops the sphygmic wave.

The strong antioxidant properties of Ginkgo biloba extract are equally well known. Recently, moreover, the role of the extract has been highlighted in prolonging the action of EDRF (Endothelium-Derived Relaxing Factor), better identified as "nitric oxide", a substance that is known for its relaxing action on vessels. This capacity would be the result of interaction between nitric oxide and guanylate cyclase which, leading to a rise in cGMP levels, would cause relaxation of the parietal myocytes leading to decontraction of those arterial branches with outlet at the level of the trabeculae of the corpora cavernosa.

Flavonoid dimers from Ginkgo biloba

As mentioned above, the leaves of Ginkgo biloba are rich in valuable active principles. The three fractions that are most important pharmacologically, identified and characterized to date, are the flavonoid glycosides, the terpene lactones and the biflavones. The flavonoid glycosides and the terpene lactones constitute the active fractions of the standardized extract of Ginkgo biloba, present at 24 and 6%, respectively.

The biflavone component, however, cannot be extracted by the normal extraction processes, but requires special extraction procedures and constitutes a product per se. So far 5 biflavones have been identified, namely amentoflavone, bilobetin, isoginkgetin, ginkgetin and sciadopitysin. These biflavones, as well as being potent antioxidants, have anti-inflammatory microvasculokinetic, anti-phosphodiesterase and antiallergic properties. In contrast to the terpene lactones, they do not display any capacity for interaction with PAF.

With regard to microvasculokinetic activity, it is important to emphasize that the biflavones improve, after acute treatment, the amplitude of the arteriolar sphygmic wave and, after chronic treatment, the capillary density in tissues with trophic-connective disorders of a varying degree.

The anti-phosphodiesterase activity of the biflavones is important since the phosphodiesterases (PDE) are cellular enzymes that interact with the cyclic nucleotides, which in turn are involved as second messengers in intracellular signal transduction and so are responsible, among other things, for vasomotor activity at the level of the capillary arterioles. The anti-inflammatory properties of the biflavones, in particular those of amentoflavone, have been demonstrated both in vitro, by assessing the interaction of said biflavones with cyclooxygenase, lipoxygenase and phospholipase A2, and in vivo, in various animal models of inflammation (oedemas induced by carrageenan, by Croton oil, etc.). Purified triterpenes oi Centella asiatica

Centella asiatica is characterized by the presence of a triterpene fraction which has, among other things, lymph-draining action and stimulation of collagen production. Using an extremely complex purification process it is possible to obtain, starting from the above-ground part of Centella asiatica, a mixture of this fraction composed of madecassic acid (30%), asiatic acid (30%), asiaticoside (40%>). These substances are triterpenes that recognize the fibroblast as main target and, by interacting with this, perform their functions, accelerating the uptake and metabolism of lysine and proline (two amino acids that are fundamental for the final structure of collagen), increasing the synthesis and release of tropocollagen and stimulating the turnover of the acid mucopolysaccharides in connective tissue.

L-Arsinine

In the treatment and in the prevention of erectile dysfunction, phosphodiesterase inhibitors, such as sildenafil and/or its analogues, are widely used, or the derivatives of Ginkgo biloba already mentioned, which prolong the half-life of the cyclic nucleotides and consequently improve the local circulatory capacity, thus permitting a prolonged erection. Another approach for potentiating the action of phosphodiesterase inhibitors consists of potentiating the nitric oxide (NO) pathway by combining a producer of nitric oxide: L-arginine, which exerts a direct action, by reaction with molecular oxygen, causing production of citrulline and NO. The latter, by activating PDE-cGMP, increases the local level of cyclic nucleotides causing vasodilatation. Citrulline in its turn permits rapid feedback in the production of arginine. D(+)-Limonene (from essential oil)

Limonene is a cyclo-olefin classified as a cyclic monoterpene. Colourless at room temperature, it possesses a strong odour of oranges, lemons or turpentine depending on the chiral composition. It takes its name from lemon, and lemon peel, like the peel of other citrus fruits, contains large amounts of it. Being a chiral molecule, it can be in dextrorotatory and laevorotatory enantiomeric forms and in the racemic form, also called the dipentene form. The R enantiomer smells of orange, the S enantiomer of lemon. The chemical compound occurring in nature, and therefore obtainable by extraction, is D- limonene, i.e. (4R)-(+)-4-isoprenyl-l-methylcyclohexene. In the plant cell it is synthesized starting from geranyl pyrophosphate, by cyclization of a neryl carbocation or an equivalent thereof. The final step consists of loss of a proton with formation of the alkene.

Data in our possession and not yet published reveal a clear potentiating effect on induction of inducible NO synthase, when this is combined with active principles known to have this action, for example Ginkgo biloba extracts. This activity is at least partly responsible for the clinical findings reported below in the treatment of erectile dysfunction.

Description of the invention The first object of the present invention is to provide a composition that allows to tackle the organic causes connected with those erectile dysfunctions that have already led, or are on the point of leading, to true impotence, and that displays the optimum requirements such as:

- vasokinetic action on the small arteries and on the precapillary arterioles, able to cause relaxation of the smooth muscles of the arterioles and to increase the volume and flow rate of the local blood flow, but without acting on important parameters such as pressure (systolic and diastolic) and heart beat;

- vasokinetic action both on the phasic component and on the tonic component of the myocyte contraction that normally develops the sphygmic wave;

- antioxidant action, to promote the effects of nitric oxide, which involve relaxation of the parietal myocytes and then decontraction of the arterial branches that open into the trabeculae of the corpora cavernosa;

- activity of improvement of the replacement of collagen fibres, with consequent reduction of sclerosis of the affected tissues.

Another object of the invention is the use of the combinations for influencing sports efficiency and performance. Clinical data in fact show that in athletes these formulations reduce the production of ROS for an average value of 40%, with peaks of 60% in smokers who engage in sports. The parameters of blood perfusion measured with echo-doppler also show a 25% improvement in terms of blood circulation. This is followed by an increase in sports performance which can be assessed, for example in aerobic endurance sports, at about 15% after 3 months of treatment.

Thus, it has been found that a composition comprising leuco-anthocyanins of Vitis vinifera or molecules that are closely related chemically, obtainable from other genera (Pinus, Vaccinium, etc.) such as procyanidolic oligomers, proanthocyanidins, anthocyanins, standardized extract of Ginkgo biloba, flavonoid dimers from Ginkgo biloba, purified triterpenes of Centella asiatica, L-arginine and limonene, satisfies the requirements stated above.

Accordingly, the present invention relates to compositions comprising:

a) leuco-anthocyanins of Vitis vinifera (or analogues from other genera) b) standardized extract of Ginkgo biloba;

c) flavonoid dimers from Ginkgo biloba;

d) purified triterpenes of Centella asiatica;

e) L-arginine, and

f) D-limonene (essential oil)

The compositions of the present invention comprise the various components within the following dosages expressed as mg/day:

a) leuco-anthocyanins (or analogues) of Vitis vinifera: from 10 to 500; b) standardized extract of Ginkgo biloba: from 10 to 400; c) flavonoid dimers from Ginkgo biloba: from 10 to 200; d) purified triterpenes of Centella asiatica: from 10 to 200; e) L-arginine: from 100 to 3500, and

f) D-limonene: from 5 to 1000.

According to a preferred aspect, the compositions of the present invention will the various components within the following dosages expressed as mg/day:

a) leuco-anthocyanins of Vitis vinifera: from 100 to 140; b) standardized extract of Ginkgo biloba: from 80 to 120; c) flavonoid dimers from Ginkgo biloba: from 20 to 60; d) purified triteipenes of Centella asiatica: from 15 to 45; e) L-arginine: from 100 to 400, and

f) limonene from 10 to 700.

A suitable formulation advantageously contains the following ingredients in the following unit doses:

Leucocyanidins 150 mg

Extract of Ginkgo biloba 120 mg

Flavonoid dimers of Ginkgo biloba 50 mg

Extract of Centella asiatica 30 mg

L-Arginine 50 mg

Limonene 10 mg According to a preferred aspect of the invention, the components from a) and d) can be present in the form of extract or as active ingredients contained therein.

According to a preferred aspect of the invention, the components from a) and d) will be present in the form of complexes with a soya phospholipid, such as lecithins, phosphatidylcholine, phosphatidylethanolamines, phosphatidylserine and similar, in particular soya distearoylphosphatidylcholine. These complexes, known as phytosomes, are described, for example, in EP 441 279 and are available commercially under the trade name Fitosoma®.

The lipophilic moiety of this complex, soya phospholipid, allows better interaction of the active ingredient with the colic acids supplied for emulsion and capture of the substances to be conveyed, by absorption, to the systemic circulation. Toxicological studies on animals and pharmacokinetic studies on humans have in fact demonstrated absence of toxicity and greater absorption of the ingredient administered as phytosome compared with the ingredient administered in free form.

According to a preferred aspect the limonene can be contained in the form of essential oil from Citrus spp. with titre above 85% or pure in the form of oil either obtained by extraction or obtained synthetically.

According to a preferred aspect of the present invention, the compositions in question will be able to contain additional components, with therapeutic action, or complementary, or otherwise useful for the purposes of the invention. Examples of said additional components are plant extracts, vitamins, amino acids, trace elements, and the like. These additional components will be present in amounts corresponding to their recommended daily dose (or to submultiples or multiples thereof). Examples of these additional components are extracts of valerian, melissa, kava kava, passionflower, hypericum, thyme, Eschscholtzia or eurycoma; amino acids such as citrulline, ornithine or lysine; group B vitamins, vitamin D, vitamin C, vitamin and vitamin E; extract and/or hydrolysate of fish cartilage, preferably from shark; and trace elements such as Mg. Mn, K in various salifications with inorganic and organic salts and in amino chelate forms. The compositions of the invention will be able to be formulated in any form suitable for oral administration, for example as capsules of hard or soft gelatin, tablets, effervescent or chewable tablets, granules or powders in sachets, solid forms for controlled release, chewing gums and the like.

The compositions of the present invention will be able to be formulated suitably for administration by the oral route and will be prepared according to conventional methods that are well known in pharmaceutical technology, such as those described in "Remington's Pharmaceutical Handbook", Mack Publishing Co., N.Y., USA, using excipients, diluents, fillers, and anti-caking agents that are acceptable for their intended use.

The compositions of the invention, in relation to the first object of the invention, are useful, as well as for the prevention and treatment of erectile dysfunction and impotence, also for improving and prolonging an erection, for improving sperm production and the vitality of the spermatozoa and for delaying ejaculation.

The clinical data supporting the validity of the proposed formulations are presented below.

ACTIVITY TESTING

Objectives and Methods

To verify the efficacy of the product, an activity test was conducted, against placebo, on 36 male subjects, healthy and sexually active, aged between 22 and 60 years. The subjects were included in the study on a voluntary basis after establishing absence of pathological problems of erectile dysfunction and absence of current severe systemic pathologies. The volunteers, after they had signed an informed consent form, were divided into three groups with 12 subjects in each: the subjects belonging to group A received 14 soft gelatin capsules formulated as in example 7 (see formulation examples) but without the presence of limonene, the subjects in group B received 14 soft gelatin capsules containing inert material (wheat germ oil); while the subjects in group C received 14 capsules of gelatin formulated as in example 7. All the capsules used in the study were of identical colour and were supplied in dark glass bottles. All the subjects (group A, group B and group C) took one capsule a day for 14 days and maintained normal sexual activity during this period.

The efficacy of the treatment was evaluated by a self-assessment questionnaire that the study participants completed at the end of the observation period ( 14 days).

The possible activity of limonene alone was tested on an additional group of 12 subjects called group D. They received 14 soft gelatin capsules, of identical appearance to the others, containing wheat germ oil (inert) and 10 mg of limonene (the same amount of limonene present in the capsules of group C ).

Results

The study was carried out between March 1 , 2010 and July 30, 2010. One subject in group A stopped the treatment following the onset of mild and 'transient side-effects (hot flushes, decrease in pressure and feeling of restlessness) and therefore has not been included in the analysis of the results, while all the subjects in groups B and C completed the study. The results of the self-assessment questionnaires are presented in Table 1 (group A), in Table 2 (group B) and in Table 3 (group C). The results for group D are shown in Table 4.

Parameter Results (group A)

within 45 minutes: 1/1 1 (9.1 % of subjects)

Beginning (average) of

within 60 minutes: 3/1 1 (27.3% of subjects) perception of the effect of the

within 90 minutes: 4/1 1 (36.4% of subjects) treatment

no perception: 3/1 1 (27.3% of subjects)

Presence of unusual nocturnal 5/1 1 (45.5% of subjects)_, erections

Presence of unusually intense

6/1 1 (54.5% of subjects)

morning erections

Greater intensity of erection as

8/1 1 (72.7% of subjects)

a result of sexual stimuli

Greater ease of maintaining

7/1 1 (63.6% of subjects)

erection during intercourse

Quicker recovery following

6/1 1 (54.5% of subjects)

intercourse

Up to 4 hours: 3/1 1 (27.3% of subjects)

Duration of the effect of the Up to 8 hours: 6/1 1 (54.5% of subjects) treatment More than 8 hours: 0/1 1

no perception: 2/1 1 ( 18.2% of subjects)

Table 1 - results of self-assessment questionnaire, group A

Parameter Results (group B)

within 45 minutes: 2/12 ( 16.7% of subjects)

Beginning (average) of

within 60 minutes: 0/12 (0% of subjects) perception of the effect of the

within 90 minutes: 1/12 (8.3% of subjects) treatment

no perception: 9/12 (75% of subjects)

Presence of unusual nocturnal

2/12 ( 16.7% of subjects)

erections

Presence of unusually intense

1/12 (8.3% of subjects)

morning erections

Greater intensity of erection as

2/12 ( 16.7% of subjects)

a result of sexual stimuli

Greater ease of maintaining

1 /12 (8.3% of subjects)

erection during intercourse

Quicker recovery following

1 /12 (8.3% of subjects)

intercourse

Duration (average) of the Up to 4 hours: 3/12 (25% of subjects) effect of the treatment Up to 8 hours: 0/12 (0% of subjects) More than 8 hours: 0/12 (0% of subjects) no perception: 9/12 (75% of subjects)

Table 2 - results of self-assessment questionnaire, group B

Parameter Results (group C)

within 45 minutes: 2/12 ( 16.6% of subjects)

Beginning (average) of

within 60 minutes: 5/12 (41 .6% of subjects) perception of the effect of the

within 90 minutes: 3/12 (25.0% of subjects) treatment

no perception: 1/12 (8.3% of subjects)

Presence of unusual nocturnal

10/12 (83.3% of subjects)

erections

Presence of unusually intense

8/12 (66.6% of subjects)

morning erections

Greater intensity of erection as

Ί 1/12 (91 .6% of subjects)

a result of sexual stimuli

Greater ease of maintaining

10/12 (83.3% of subjects)

erection during intercourse

Quicker recovery following

9/12 (75.0% of subjects)

intercourse

Up to 4 hours: 3/12 (25.0% of subjects)

Duration of the effect of the Up to 8 hours: 7/12 (58.3% of subjects) treatment More than 8 hours: 2/12 ( 16.6% of subjects) no perception: 0/12

Table 3 - results of self-assessment questionnaire, group C Some subjects belonging to group C also reported a considerable improvement in athletic performance.

7 subjects out of 12 in group C (formulation example 7) in fact engaged in sports in the study period (including 4 jogging and 3 gymnasium); all 7 subjects reported reduced fatigue and increased stamina. An improvement, but to a lesser extent, was reported in 4 out of 6 subjects in group A (formulation example 7 without limonene) who engaged in sports activities.

In group B (placebo), in which 5 subjects out of 12 engaged in sports during the study period (jogging), only one subject reported a slight, but perceptible improvement on an exercise bike.

Table 4 - results of self-assessment questionnaire, group D

No subject noticed any improvement in athletic, training, although 8 of them engaged in sports activities regularly (3 jogging, 2 cycling, 3 gymnasium).

Study conclusions The formulation as in formulation example 7 (group C) showed significantly greater efficacy compared with the same formulation without the compound limonene (group A). Both showed significant differences relative to the placebo group (group B). These considerations apply to all the parameters included in the self-assessment questionnaire. It can be seen, however, that the two parameters most influenced by the formulation containing limonene are, following sexual stimulation, the intensity of erection and maintenance thereof during intercourse.

Moreover, the formulation containing limonene improved the athletic performance of the treated subjects more clearly compared with the group treated with the formulation without limonene and, even more so, compared with the group treated with the placebo formulation.

It can clearly be seen that the presence of limonene is fundamental for greater success, in terms of efficacy, of the formulation. Even if the mechanism still seems to be far from being fully elucidated, it seems clear that the presence of limonene produces a valid enhancer effect. Probably its presence generates synergy with the proanthocyanidin fraction from Vitis vinifera, through its capacity for endothelial protection and protection of the half-life of vascular NO. Moreover, regarding the real presence of NO, there is dual activation of the NO-producing systems; in fact these systems are supplemented with raw material from the quota of L-arginine present in the formulation and are protected from the anti-phosphodiesterase inhibitory action exerted by the dimer fraction and by the ginkgoflavone-glycoside fraction (both from G. biloba), which are able to prolong the half- life of NO.

The test for checking whether limonene is able to produce, when taken on its own, an improvement in erectile or sports activity had a completely negative outcome. The synergistic result from combining limonene with the plant extracts can neither be explained, nor interpreted, as the summation of the effects of the individual components.

CLOSER EXAMINATION OF SPORTS ACTIVITY A double-blind, placebo-controlled clinical study was recently completed on the efficacy of formulation 7 on 20 subjects engaging in amateur cycling for 30 days. 10 subjects were treated with the formulation as in example 7 and 10 were treated with a placebo, of identical appearance.

The results indicate a significant difference between the two groups in the parameters assessed, described below.

In particular, it was noted that in the treated subjects the heart rate (HR) on exertion both at 50% and at 90% of maximum power was more uniform compared with the subjects who took the placebo. Moreover, the values of V02 (volume of oxygen consumed) recorded during the recovery period after exertion were lower in the treated subjects. The correlation between these two parameters allows us to state that the subjects were less fatigued on exertion and that also the recovery period was consequently shorter.

A number of haematochemical parameters were measured to evaluate the level of oxidative stress in the subject. The following were evaluated:

• Carbonylate proteins (CP. markers of oxidative stress, connected with the oxidizing action of the free radicals on said proteins).

• Determination of thiobarbituric acid (TBARS. standard marker of oxidative stress, connected with the oxidizing action of free radicals on lipids).

• 1L6 (cytokine with pro-inflammatory activity)

All the parameters proved to be statistically significantly lower compared with the value at time zero in the treated group. This decrease in the oxidative parameters of the organism is an indicator that fatigue during athletic activity was lower owing to the action of the supplement. This explains the improvement in athletic performance recorded in the previous study.

FORMULATION EXAMPLES (dose/unit or treatment) Example 1 : CAPSULES TYPE 1

Leucocyanidin 120 mg

Extract of Ginkgo biloba 100 mg

Flavonoid dimers of Ginkgo biloba 40 mg

Extract of Centella asiatica 5 mg

L-Arginine 200 mg

Limonene e.o. 13 mg

Microcel ® 40 mg

Magnesium stearate 5 mg

Aerosil ® 10 mg

Example 2: FILM-COATED TABLETS

Leucocyanidin 120 mg

Extract of Ginkgo biloba 100 mg

Flavonoid dimers of Ginkgo biloba 40 mg

Extract of Centella asiatica 10 mg

L-Arginine 200 mg

Limonene, supported 50 mg

Microcel ® 340 mg

Dicaphos ® 333 mg

Magnesium stearate 10 mg

Aerosil ® 8 mg

Explocel ® 18 mg

Sepifilm ® 22.8 mg

Shellac 7 mg

Colorant 0.2 mg

Example 3: FILM-COATED TABLETS

Leucocyanidin phytosome 60 mg Extract of Ginkgo biloba phytosome 50 mg

Flavonoid dimers of Ginkgo biloba phytosome 20 mg

Extract of Centella asiatica phytosome 10 mg

L-Arginine 400 mg

Limonene, supported 25 mg

Microcel ® 340 mg

Dicaphos ® 333 mg

Magnesium stearate 10 mg

Aerosil ® 8 mg

Explocel ® 18 mg

Sepifilm ® 22.8 mg

Shellac 7 mg

Colorant 0.2 mg Example 4: DOUBLE-LAYER PROGRAMMED-RELEASE TABLETS Layer with normal release

Leucocyanidin 60 mg

Extract of Ginkgo biloba 50 mg

Flavonoid dimers of Ginkgo biloba 40 mg

Extract of Centella asiatica 15 mg

L-Arginine 200 mg

Limonene, supported 10 mg

Dicaphos ® 304 mg

Explocel ® 15 mg

Aerosil ® 3 mg

Magnesium stearate 6 mg

Colorant 1 mg

Layer with delayed release Leucocyanidin 80 mg

Extract of Ginkgo biloba 75 mg

Flavonoid dimers of Ginkgo biloba 30 mg

Extract of Cenlella asiatica 10 mg L-Arginine 250 mg

Limonene, supported 30 mg

Metholose ® 80 mg

Aerosil ® 2 mg

Magnesium stearate 3 mg Microcel ® 80 mg

Dicaphos ® 164 mg

Example 5: SACHETS Leucocyanidin 120 mg Extract of Ginkgo biloba 100 mg

Flavonoid dimers of Ginkgo biloba 40 mg

Extract of Centella asiatica 30 mg

L-Arginine 1600 mg Limonene microencapsulated for coacervation 75 mg

Fructose 1873 mg

Aerosil ® 15 mg

Orange flavour 180 mg

Citric acid 220 mg Acesulfame K 25 mg

E102 2 mg

Example 6: ORO-DISPERSIBLE FORMULATION Leucocyanidin 120 mg

Extract of Ginkgo biloba 100 mg

Flavonoid dimers of Ginkgo biloba 40 mg Extract oi Centella asiatica 30 mg

L-Arginine 600 mg

Limonene e.o. 5 mg

Sorbitol 160 mg

Orange flavour 20 mg

Mandarin flavour 5 mg

Acesulfame K 2 mg

Aerosil ® 5 mg

Fructose 1530 mg

Example 7: SOFT GELATIN CAPSULES

Leucocyanidin phytosome 150 mg

Extract of Ginkgo biloba phytosome 120 mg

Flavonoid dimers of Ginkgo biloba phytosome 50 mg

Extract of Cen fella asiatica phytosome 30 mg

L-Arginine 50 mg

Limonene e.o. 10 mg

Food-grade oil q.s.

Claims

1. Composition comprising:

a) leuco-anthocyanins of Vitis vinifera or analogues from other genera; b) standardized extract of Ginkgo biloba;

c) dimeric flavones from Ginkgo biloba;

d) purified triterpenes of Centella asiatica;

e) L-arginine, and

f) D-Limonene (essential oil).

2. Composition according to claim 1 , wherein the components a) to d) are present in extract form or as active ingredients contained therein.

3. Composition according to claim 1 , wherein the components a) to d) are present in a complex with phospholipids.

4. Composition according to claim 3, wherein the components a) to d) are present in a complex with distearoylphosphatidylcholine soy.

5. Composition according to the preceding claims, in which the components, expressed as mg/day, are in the following amounts:

a) leuco-anthocyanins (or analogues) of Vitis vinifera: 10 to 500;

b) standardized extract of Ginkgo biloba: 10 to 400;

c) Ginkgo biloba dimeric flavones from: 10 to 200;

d) purified triterpenes oi Centella asiatica: 10 to 200;

e) L-arginine: 100 to 3500, and

0 Limonene from 5 to 1000.

6 Composition according to claim 5, wherein the components are present in the following amounts

a) leuco-anthocyanins (or analogues) of Vitis vinifera: 100 to 140; b) standardized extract of Ginkgo biloba: from 80 to 120; c) Ginkgo biloba dimeric flavones: 20 to 60;

d) purified triterpenes of Centella asiatica: 15 to 45;

e) L-arginine: 100 to 400, and

f) Limonene from 5 to 350.

7. Composition according to the preceding claims, further comprising additional components, with curative or supplementary activity, selected from extracts of valerian, melissa, kava kava, passionflower, hypericum, thyme, Eschscholtzia or eurycoma; amino acids such as citrulline, ornithine or lysine; group B vitamins, vitamin D, vitamin C, vitamin K and vitamin E; extract and/or hydrolyssate fish cartilage, preferably of sharks; and oligoelements such as Mg, Mn, K in the form of inorganic, organic salts and aminochelate forms.

8. Composition according to the preceding claims, in a form suitable for oral administration.

9. Composition according to claim 8, in the form of hard or soft gelatin capsules, tablets, effervescent tablets or chewable tablets, sachets of powder or granular, controlled-release solid dosage forms, chewing gum and the like.

10. Composition according to any of claims 1 to 9 for the treatment and prevention of erectile dysfunction and impotence, to improve and prolong an erection, to improve sperm production and viability of sperm and/or to delay ejaculation.

1 1 . The use of a composition according to any of claims 1 to 9 to improve sports performance of a healthy subject.