ITRM20120591A1 - COMPOSITIONS TO BE USED IN STAGE AND INFLAMMATORY FLEBOPATHOLOGIES - Google Patents

Description

DESCRIPTION OF THE INVENTION

FIELD OF APPLICATION

The present invention in the chemical-pharmaceutical field relates to compositions for the preparation of drugs, food supplements and cosmetics based mainly on medicinal plants. The compositions are all characterized by the presence of rutin and luteolin, pure or as titrated dry extracts, in a mixture that negatively modulates the expression of the IL1-β and iNOS genes and by other molecules or dry extracts all useful for subjects suffering from venous insufficiency .

STATE OF THE ART

Chronic Venous Insufficiency (CVI) appears to be a very relevant clinical condition that manifests itself with the appearance of varicose veins; these appear as abnormal and saccular dilations of the veins, especially of the lower limbs, which often take on a tortuous course. Pathological dilation of the veins is associated with a modification of the venous walls that can be localized, with nodular or diffuse areas. Varicose disease generally depends on various haemodynamic factors that influence venous return. Important factors such as venous musculature, muscle pump during phasic contractions and venous tone under the control of the vegetative nervous system are important. Among the factors that can cause varicose veins to develop are recognized:

• Constipation, which hinders the return of venous blood upwards;

• Obesity:

Muscle pump deficiency

»Work in orthostatic position (standing position) in a warm environment (which favors a dilation of the vessels);

«Alterations of the veins present at birth (congenital);

o Age (between 20 and 50 years old);

° Sex (predominantly female).

The factor that most affects venous blood pressure is posture; during the standing position, especially if prolonged, the venous pressure increases considerably, up to ten times the normal pressure. Consequently, the upright posture often determines the onset of a marked venous stasis and the appearance of edema affecting the feet. Varicose veins are much more frequent after 50 years of age, in obese people and especially women, due to the high venous pressure that pregnancy causes in the lower limbs. Furthermore, the high levels of estrogen present in the female sex facilitate the relaxation of the muscles of the venous wall. The familial tendency seen in varicose vein disease is believed to be due to impaired development of the venous wall.

The real cause of the dilation of the veins of the lower limbs, however, has not yet been defined the attention of the researchers has turned to the structure of the varicose vein wall, in which there seems to be less collagen and elastin than in the normal venous wall. . Beyond the pharmacological treatment, it is evident that physical exercise and diet are important for improving the quality of life of subjects suffering from stasis and inflammatory phlebopathies.

A diet followed by subjects with phlebopathologies should include functional nutrients aimed at improving remodynamics, maintaining the tone of the smooth muscle of the veins. In addition, a diet should promote the control of body weight; promote normal digestion and liver function; affect the regularity of intestinal transit; promote the synthesis of collagen and elastin in the epithelial cells of the veins. The present invention exploits the idea of having looked for substances such as rutin and luteolin present in plants useful for drip pathologies. In particular, a mixture of rutin and luteolin has been identified which inhibits the synthesis of the gene coding for interleukin 1-beta found in turn to repress the synthesis of elastin. Rutin and Luteolin are associated with plant substances in new compositions for use in phlebopathology.

There are numerous patented compositions both for pharmacological use and for nutritional use based on the two flavonoids Diosmin and Hesperidin which are indicated for the treatment of varicoses. Various associations between molecules of plant origin have been shown to be useful for the well-being of those suffering from varicosis. In the pharmaceutical compositions, hesperidin and diosmin alone or in combination are contained in 500 mg per total dose. It should be borne in mind that for functional nutritional preparations only concentrations of 300mg / day are currently permitted as the maximum daily intake. Patent EP2022345A1 describes a food composition comprising between 25% and 80% by weight of Hesperidin; between 1% and 10% of Isonaringina; between 0.5% and 1% by weight of Eriocitrine; and between 0.1% and 1% by weight of Diosmin. This formulation is patented for the preparation of a food supplement capable of preventing, reducing or treating cardiovascular problems such as varicose veins, heavy or tired legs, hemorrhoids. Diosmin and Hesperidin must be present as all published studies confirm the efficacy of these molecules in the treatment of phlebopathologies. However, the low dosage allowed for supplements is not sufficient as the works on the subject show: T. Silambarasan, B. Raja: Diosmin, a bioflavonoid reverses alterations in blood pressure, nitric oxide, lipid peroxides and antioxidant status in DOCA-sait induced hypertensive rats . Eur. J. Pharmacol. 679 (2012) 81-89; Nicolaides AN .: From symptoms to leg edema: efficacy of Daflon 500 mg. Angiology. 2003 Jul-Aug; 54 Suppl 1: S33-44; and the work of A-W. N. Meshikhes: Daflon for haemorrhoids: a prospective, multi-center observational study 004Surg J R Coll Surg Edinb (rei 2: 6; 335-338.

Doses and time of intake are particularly important, for example prolonged use of Diosmin alone at high concentrations has been associated with bleeding (R. M. Kumar, J. J. Van Gompel, R. Bower; A. A. Rabinstein: Spontaneous Intraventricular Hemorrhage Associated with Prolonged Diosmin Therapy. Neurocrit Care (2011) 14: 438-440).

In a pharmaceutical formulation under patent EP1707192A3, Diosmin is associated with one or more flavonoids in microcrystalline form, such that the specific surface area (measured with the BET technique) is between 4 and 18m <2> / g and such that the weight ratio / weight between flavonoids and Diosmin must be between 0.03 and 0.1.

Studies conducted on compounds associated with Diosmin and Hesperidin have suggested new applications in the field of venous insufficiency as in patent EP2208498A1 Hesperidin and Diosmin are associated with calcium dobesilate (calcium 2,5-dihydroxybenzenosulfonate) which is only allowed for pharmacological use. This patent protects the compositions in which Diosmin can vary between 250mg and 450mg, Hesperidin is between 25mg and 50mg and calcium dobesilate between 250mg and Ig.

Hesperidin and Diosmin have been associated with various molecules also with general applications, as in patent IT1249041B. There are also compositions indicated for the symptomatic treatment of venous insufficiency as described in the patent FR2967064 in which an extract from Punica granatum is used to prevent heavy legs. The same patent shows the studies of the association of hesperidin with other substances such as vitamins and antioxidants. In patent EP1637134, eicosapentaenoic acid is used for the reduction of some symptoms of varicose veins. In the patent W02006089317 to obtain a preventive mixture for capillary and microcirculatory problems many components are used such as Escin, rutoside, troxerutin, Diosmin, Hesperidin, gingko, Vitis viniferae (red vine leaves) and / or Ruscus aculeatus (Rusco) and optionally a vitamin, flavorings, trace elements, minerals and / or an additional addition. Other compositions containing Diosmin and / or Hesperidin have applications in pathologies such as diabetes or obesity.

The present invention is new thanks to the demonstration of the effects of rutin and luteolin on the metabolism of Saos-2 cells and while considering the state of the art it proposes new compositions useful for their effectiveness in the treatment of stasis and inflammatory phlebopathies as they reduce the expression of some inflammatory genes. The association of rutin and luteolin in a given concentration reduces the expression of the IL1-beta gene responsible for the decrease in elastin as shown in the work of S. Lin, K. Mequanint: The role of Ras-ERK-IL-lb signaling pathway in upregulation of elastin expression by human coronary artery smooth muscle cells cultured in 3D scaffo! ds. Biomaterials 33 (2012) 7047e7056. The reduction of elastin is one of the possible causes of some venous insufficiency.

EXAMPLE 1

The present invention makes use of the association of Rutin and luteolin with Diosmin and Hesperidin, the use of which for the treatment of venous insufficiency is consolidated. In a preferred composition Diosmin and Hesperidin are added as extracts from fruits of the genus Citrus. The maximum quantity that can be added in the case of food supplements is established by the competent control body. -

According to the present invention, the onesie and the luteolin are useful for improving the state of well-being and health of individuals suffering from venous insufficiency. Rutin and Luteolin are present in various drugs but in this formulation Hieracium pilosella have been used due to the presence of luteolin (Haag-Berrurier M, Duquenois P .: Extraction and physicochemical characterization of the luteolin in the flowers of the hawkweed Hieracium pilosella L .. C. R. Hebd Seances Acad Sci. 1962 May 7; 254: 3419-21) and Taraxacum officinolis.

In order to establish a composition useful for the purposes of the invention, studies were carried out on the effect of rutin and luteolin added, individually and as mixtures, to the culture medium of Saos-2 human cells (HTB-85 line in the ATCC catalog ) and analyzing the dose-response effect on the modulation of the expression of some genes encoding factors involved in inflammatory processes: interleukin 1-beta (IL1-beta) and tumor necrosis factor-alpha (TNF-alpha); in the defense processes against antioxidants: superoxide dismutase (SOD-2) and the inducible enzyme responsible for the synthesis of nitric oxide (iNOS). In the experiments, Saos-2 cells were grown as described in the work A. Scotto d'Abusco, C. Ciclone, V. Calamia, R. Negri, C. Giordano, B. Grigolo, L. Politi, R. Scandurra: Glucosamine and its N-acetyl-phenylalanine derivative prevent TNF-α-induced transcriptional activation in human chondrocytes. Clin. Exp. Rheumatol. 2007; 25: 847-852. Once a minimal cell concentration was reached, the Saos-2s were treated for 24 hours, all at various molecular concentrations. The measurement was performed using Reai Time PCR; total messenger RNA- {RNA) was extracted from each sample of treated cells according to the method described elsewhere (Scotto d'Abusco et al, 2007). untreated cells (CTL) are used to calibrate the results, the variation of the amount of RNA in the CTL is assumed to be 1.0. The values of the target gene Ct were normalized with respect to the value of the gene coding for GAPDH. The data in the treated cells were using the method and expressed as the change with respect to the CTL according to the following expressions:

where is it

ΔCt 1 = Ctgene control - Ct (GAPDH) control;

ΔCt 2 = treated Ctgene - treated Ct (GAPDH)

Figure 1 shows the most interesting results derived from the study. The mixture of rutin and luteolin at a concentration of 50 μΜοΙ is active on Saos-2 cells and is able to statistically significantly lower the expression of the gene encoding IL1-beta while it does not induce changes in the expression of the genes encoding TNF -alpha and SOD-2 but strongly decreases that of the gene encoding iNOS.

FIGURE 1

Considering the symptomatology of varicose veins it is useful to add rutosides to the composition which, for example, can be found in Ruscus aculeatus; the quantities can also be evaluated considering the study by W. Vanscheidt et al. Efficacy and Safety of a Butcher's broom preparation (ruscus aculeatus L.extract) compared to Placebo in patients suffering from chronic venous insufficiency. Azneim.-Forsch./Drug.Res. 52.4, 243-250 (2002).

To the composition can be added Aesculus hyppocastanus which contains escin, a saponin with vasoprotective and vasoconstrictive properties as suggested in the Raffetto JD study, Khalil RA .: Ca (2 +) - dependent contraction by the saponoside escin in rat vena cava: implications in venotonic treatment of varicose veins. J Vasc Surg. 2011; 54 (2): 489-96. Epub 2011 Apr 16. The addition of saponins is also useful because to improve the bioavailability, both at the intestinal and piasmatic level, of the other substances used in the compositions as it lowers the surface tension of the dispersed systems containing the composition.

To enhance the anti-inflammatory action Curcuma Ionga was used which contains curcuminoids probably responsible for the anti-inflammatory action as suggested in the work of Julie S. Jurenka: Antiinflammatory Properties of Curcumin, a Major Constituent of Curcuma Ionga: A Review of Preclinical and Clinical Research- Alt. Med. Review 14, 2, 2009, 141-153.

According to the present invention, a typical composition contains 92% (W / W) Diosmin and 93% (W / W) Hesperidin preferably in the form of extracts from the fruits of Citrus aurantium -L and- each substance is present up to a maximum of 300 mg / day. A 60 ° ethanol extract from Ruscus aculeatus up to a maximum of 50 mg with saponins (such as ruscogenin) between 3% (W / W) and 10% (W / W). A 60 ° ethanol extract from Hieracium pilosella with a content greater than 1% of flavonoids including luteolin and its derivatives such as luteolin-7-O-glucoside. An ethyl acetate extract of Curcuma Ionga containing between 90% (W / W) and 95% (W / W) of curcuminoids. An extract in ethanol: water (70:30) from Aesculus hyppocastanus seeds containing between 15% (W / W) and 25% (W / W) of triterpene glycosides including aescin- An aqueous extract from Taraxacum officinale Web. Containing between 15% (W / W) and 25% (W / W) of inulin.

EXAMPLE 2

In a preferred composition, the dry extracts of the drugs Citrus aurantium, Aesculus hyppocastonus, Ruscus aculeatus, Hieracium pilosella, Curcuma Ionga, Taraxacum officinale Web are used to improve fluid drainage in subjects suffering from venous insufficiency. The composition differs from the previous ones for the addition of quantities of drugs such as Hieracium pilosella and / or Taraxacum officinole able to enhance the drainage of body fluids. In venous insufficiency the local drainage of the vessels is particularly important to eliminate the transudate liquids both from the most sloping veins and from the lymphatic vessels. The hydrostatic pressure also prevents the lymphatic system from draining these extravasal fluids which over time, being also inflammatory in nature, organize themselves creating fibrosis. Hieracium pilosella from ethnobotanical studies is known for its diuretic properties and can be useful for counteracting skin redness as suggested by L. Tognetti, S. Giorgini, and T. Lotti. Erythema multiforme-like from a slimming drug preparation cutaneous adverse drug reaction. Indian Dermatol Online J. 2011 Jul-Dec; 2 (2): 78-81. Taraxacum officinale Web can also be useful for draining liquids as noted by B.A. Clare, R.S. Conroy, and K. Spelman: The Diuretic Effect In Human Subjects Of An Extract Of Taraxacum Officinale Folium Over A Single Day. The Jour. Altern. He accomplished. Med. Vol. 15, 8, 2009, 929-934.

A typical composition would be in capsule, tablet, sachet and could also be spary while attempts to obtain effervescent forms with salts induced an increase in blood pressure. Since some of the substances used are unpleasant to the taste, the tablet composition requires the use of flavoring and since some powdered drugs are bulky (eg turmeric) glidants and / or lubricants must be added. Vitamin C is added as an antioxidant which is suggested useful for the trophism of veins by F. Luo, Q. Lv, Y. Zhao, G. Hu, G. Huang, j.Zhang, C. Sun, X. Li and K. Chen: Quantification and Purification of Mangiferin from Chinese Mango (Mangifera indica L.) Cultivars and Its Protective Effect on Human Umbilìcal Vein Endotheiial Cells under H2O2-induced Stress; int J. Mol. Ski. 2012, 13, 11260-11274.

In the preferred composition, type 0 capsules were prepared containing a dry extract of Citrus aurantium titrated with 100 ÷ 150 mg Hesperidin 90% (W / W); 10Q ÷ 150 mg of Diosmin 92% (W / W); 25 ÷ 60 mg of a dry extract from Ruscus aculeatus with 5% saponins (W / W); 25 ÷ 100 mg of a dry extract from Aesculus hyppocastanus with Escin 20% (W / W), 80 ÷ 200 mg of ascorbic acid; 50 ÷ 120 mg of a dry extract from Hieracium pilosella containing 1% (W / W) of flavonoids and 0 ÷ 120 mg of a dry extract from Taraxacum officinale Web with inulin 22% (W / W).

EXAMPLE S

To identify an alternative composition to the previous ones, it was based on the comparison of cohort data of subjects suffering from venous insufficiency, accustomed to taking nutraceuticals as well as drugs. On the basis of the analysis of the various products taken (drugs, food supplements, galenic and cosmetic preparations) the following composition was established: Hesperidin and Diosmin titrated at least to 90% must be present in an overall ratio of 1: 0.9 (w / w) and such that in each single dose, regardless of the daily dose, the ratio between the total weight of Hesperidin plus Diosmin with respect to the total composition is between 0.4 and 0.6. As for the weight of the other ingredients, intended as titrated dry extracts (E.S.), each must be in the following proportional ratios with respect to the total dosage: E.S. with Escin between 0.1 and 0.2; E.S. with ruscogenins between 0.05 and 0.1; E.S. with curcuminoids between 0.05 and 0.2; coenzyme Q10 between 0.05 and 0.16. When the composition has been defined, the maximum possible amount of ascorbic acid is added, considering the pharmaceutical technique of the formulation used and the maximum amount allowed by law.

Claims (1)

CLAIMS 1 Compositions containing rutin and luteolin in a mixture such that it reduces the expression of the gene coding for IL1-beta, useful for people suffering from phlebopatoiogie. In the compositions, rutin and luteolin are added pure or preferably as titrated dry extract of Hieracium pilosella and / or Taraxacum officinale Web. The compositions associate rutin and luteolin with one or more of ascorbic acid, diosmin, hesperidin from a dry extract of Citrus aurantium, escin from a dry extract of Aesculus hyppocastanus, ruscogenine from a dry extract of Ruscus aculeatus; curcuminoids from Curcuma lónga; coenzyme Q10, so that the compositions have pharmaceutical, food and nutritional, cosmetic applications, capable of increasing the well-being of a person suffering from venous insufficiency. 2 compositions according to claim 1 with an anti-inflammatory physiological effect useful in stasis and inflammatory phlebopathies. 3 compositions according to claims 1 and 2 containing diosmin 92% (W / W) and hesperidin 93% (W / W) dry extract from the fruits of Citrus aurantium L. and each molecule present up to the maximum contribution allowed by current legislation. A 60 ° ethanol extract from Ruscus aculeatus up to a maximum of 50 mg containing saponins (ruscogenins) between 3% (W / W) and 10% (W / W). A 60 ° ethanol extract from Hieracium pilosella with a content greater than 1% of flavonoids including luteolin and its derivatives such as luteolin-7-Ο-glucoside. An ethyl acetate extract of Curcuma longa containing between 90% (W / W) and 95% (W / W) of curcuminoids. An extract in ethanol: water (70:30) from Aesculus hyppocastanus seeds containing between 15% (W / W) and 25% (W / W) of triterpene glycosides including aescin. An aqueous extract from Taraxacum officinale Web. Containing between 15% (W / W) and 25% (W / W) of inulin. 4 compositions according to claims 1 and 2 where Hesperidin and Diosmin, pure or as dry extracts having a minimum titer 90%, must be present in a ratio of 1: 0.9 (w / w) and such that the amount of Hesperidin plus Diosmin compared to the total weight of the dose is between 0.4 and 0.6. As for the weight of the other ingredients, added pure or as dry extracts (E.S.), they must each be in the following proportional ratios with respect to the total dose: containing Escin (20% titer) between 0.1 and 0.2; E.S. with ruscogenins (5% titer) between 0.05 and 0.1; E.S, with curcuminoids (95% titre) between 0.05 and 0.2; coenzyme Q10 between 0.05 and 0.16. When the composition is defined, the maximum possible quantity of ascorbic acid is added considering the pharmaceutical technique of the formulation used and the maximum contribution allowed by current legislation. 5 compositions according to claims 1 and 2 containing dry fruit extract from Citrus aurantium with 100 ÷ 150 mg Hesperidin 90% (W / W); 100 ÷ 150 mg of Diosmin 92% (W / W); 25 ÷ 60 mg of a dry extract of the rhizome from Ruscus aculeatus with 5% saponins (W / W); 25 ÷ 100 mg of a dry extract of the seeds from Aesculus hyppocastanus with Escin 20% (W / W), 80 ÷ 200 mg of ascorbic acid; 50 ÷ 120 mg of a dry extract of the Hieracium pilosella plant containing 1% (W / W) of flavonoids and 0 ÷ 120 mg of a dry extract of the plant from Taraxacum officinale Web with 22% (W / W) inulin. 6 compositions according to claims 1 and 2 in the form of capsules, tablets, sachets, sprays.