HRP20090650A2 - Formulation based on resveratrol and extracts from pomegranate, olive leaf and cinnamon used as an antioxidant agent - Google Patents

Abstract

The invention relates to a therapeutic agent containing variable proportions of: (i) resveratrol (1), in a 0.01% by weight, preferably 0.1-3%; (ii) pomegranate extract (Garnet L. pellet), in a content of 0.1-50%, preferably 0.5-25%; (iii) olive (Olea europea L.) leaf extract, in a percentage by weight of 0.1-50%, preferably 0.5-25%; (iv) cinnamon extract (Cinnamomum zeylanicum L.), by weight of 0.1-50%, preferably 0.5-25%; (v) additional plant extracts that support the antioxidant action of the main active ingredients of the formulation: garlic extract (Allium sativum L.), grape seed extract (Vitis vinifera L.), coriander extract (Coriandrum sativum L.), saffron extract (Crocus sativus L. ) and barley extract (Hordeum vulgare L.), by weight of 0.1-50% by weight, preferably 0.5-25%; and (vi) the excipients necessary for the administration of said active substances as a finished therapeutic form: tablet, capsule, tincture, syrup, instant beverage powder, or instant beverage powder, in the weight range of 5-99; The present invention solves the technical problem of effective treatment of oxidative stress, which is at the molecular level the major or one of the main causes of a number of diseases and conditions of modern humans such as: (i) cardiovascular disease (hypertension, hyperlipidemia, atherosclerosis); (ii) diabetes (and in general metabolic syndrome); (iii) immune system diseases (low level of immunity, frequent infections); (iv) brain and nervous system diseases (neurodegenerative diseases); (v) tumor diseases; and (vi) an accelerated aging process.

Description

Field of invention

The subject invention relates to a therapeutic agent that contains:

(i) resveratrol (1);

[image]

(ii) pomegranate extract (Punica granatom L);

(iii) olive (Olea europea L.) leaf extract;

(iv) cinnamon extract (Cinnamomum zeylanicum L.);

(v) additional plant extracts that support the antioxidant action of the main active ingredients of the formulation: garlic extract (Allium sativum L.), grape seed extract (Vitis vinifera L), coriander extract (Coriandrum sativum L.), saffron extract (Crocus sativus L.) and barley extract (Hordeum vulgare L.); you

(vi) auxiliary substances that are necessary so that the listed active substances can be used as a finished therapeutic form: tablets, capsules, tinctures, syrups, instant powder for a drink, or instant powder for an effervescent drink.

The formulation of the subject invention has an unexpectedly strong antioxidant effect, which enables its use for the prevention and treatment of oxidative stress conditions as the main cause:

(i) cardiovascular diseases (hypertension, hyperlipidemia, atherosclerosis);

(ii) diabetes (and metabolic syndrome in general);

(iii) diseases of the immune system (low level of immunity, frequent infections);

(iv) diseases of the brain and nervous system (neurodegenerative diseases);

(v) tumor diseases; you

(vi) accelerated aging process.

Summary of the invention

The subject invention solves the technical problem of effective therapy of the state of oxidative stress, which at the molecular level is the main or one of the main causes of a number of diseases of modern man, such as: cardiovascular diseases (hypertension, hyperlipidemia, atherosclerosis), diabetes (and metabolic syndrome in general), diseases of the immune system (low level of immunity, frequent infections), diseases of the brain and nervous system (neurodegenerative diseases), tumor diseases, and the aging process.

The subject invention relates to a therapeutic agent containing variable proportions:

(i) resveratrol (1), in a mass proportion of 0.01-10%, preferably 0.1-3%;

(ii) pomegranate extract (Punica granatum L), in a mass proportion of 0.1-50%, preferably 0.5-25%;

(iii) olive (Olea europea L.) leaf extract, in a mass proportion of 0.1-50%, preferably 0.5-25%;

(iv) cinnamon extract (Cinnamomum zeylanicum L.) in a mass proportion of 0.1-50%, preferably 0.5-25%;

(v) additional plant extracts that support the antioxidant action of the main active ingredients of the formulation: garlic extract (Allium sativum L), grape seed extract (Vitis vinifera L.), coriander extract (Coriandrum sativum L.), saffron extract (Crocus sativus L.) and barley extract (Hordeum vulgare L.), in mass fractions of 0.1-50%, preferably 0.5-25%; you

(vi) auxiliary substances that are necessary so that the listed active substances can be used as a finished therapeutic form: tablets, capsules, tinctures, syrups, instant powder for a drink, or instant powder for an effervescent drink, in a mass fraction of 5-99.19 %.

State of the art

Oxidative stress at the molecular level is the main or one of the main causes of a number of diseases and conditions of modern man such as: cardiovascular diseases (hypertension, hyperlipidemia, atherosclerosis), diabetes (and metabolic syndrome in general), diseases of the immune system (low level of immunity, frequent infections ), brain and nervous system diseases (neurodegenerative diseases), tumor diseases, and (accelerated) aging process.

The state of oxidative stress is caused by the constant exposure of the organism to elevated concentrations of harmful free radicals. The latter are also called reactive oxygen species (ROS). The superoxide anion radical (•O2-) is the most significant reactive oxygen species (ROS) that occurs as a result of the "leakage" of electrons during the complex process of oxidative phosphorylation in the mitochondria of human cells. Namely, in the process of oxidative phosphorylation (OF), which is one of the fundamental metabolic processes in the body, natural electron transporters from the Krebs cycle (citric acid cycle), reduced forms of nicotinamide adenine dinucleotide (NAOH) and flavin adenine dinucleotide (FADH2), enable the biosynthesis of adenosine triphosphate (ATP), the basic molecule for short-term cellular energy storage. so that NADH, via the enzyme NADH dehydrogenase within the complex, donates electrons to ubiquinone (Q) with the formation of ubiquinol (QH2) [F.L. Crane, Biochemical Function of Coenzyme Q10, J. Am. Coll Nutr. 20 (2001) 591]. the oxidized form of NAD+ that returns to the Krebs cycle.

Ubiquinol (QH2) is a lipophilic molecule that can travel through the mitochondrial membrane and through further stages of oxidative phosphorylation [complex n (succinate dehydrogenase), complex III (cytochrome bci complex) and complex IV (cytochrome c oxidase)] enables the biosynthesis of ATP [H. Schagger, K. Pfeiffer: The Ratio of Oxidative Phosphorylation Complexes I-IV in Bovine Heart Mitochondria and the Composition of Respiratory Chain Supercomplexes, J. Biol. Chem. 276 (2001) 37861], The terminal electron acceptor is oxygen, which with hydrogen ions, which are also formed in the process of oxidative phosphorylation (OF), gives water.

It is believed that the most sensitive sites where electron "leakage" can occur in the OF process are complex I and complex III [A.J. Lambert, M.D. Brand: Inhibitors of the Quinone-binding Site Allow Rapid Superoxide Production from Mitochondrial NADH:Ubiquinone Oxidoreductase (Complex I ), J. Biol, Chem. 279 (2004) 39414]. The acceptor of those electrons that "uncontrollably" leak out of the OF process is also molecular oxygen. However, this results in a one-electron reduction of molecular oxygen, in which the mentioned superoxide anion radical (•O2-) is formed, which is very harmful to the cell.

Superoxide anion radical (•O2-) as a highly reactive species in the cell can cause a series of harmful reactions. Here are just a few of them as an example:

(i) •O2- causes the formation of other harmful radicals such as hydroxyl radicals (HO•), hydroperoxides of higher fatty acids at allylic positions (ROOH, where R= the residue of a higher fatty acid), and corresponding hydroperoxides (ROO•) or alkoxide radicals (RO•), etc.;

(ii) •O2- causes the formation of extremely harmful peroxynitrite anion (OONO-), which further undergoes homolytic decomposition and produces harmful hydroxyl radicals (HO•) and nitrogen dioxide (NO2);

(iii) causes numerous secondary harmful effects due to a drop in the equilibrium concentration of nitrogen monoxide (NO), which disrupts the natural processes it regulates: regulation of circulation (arterial, venous, lymphatic), vasodilation, signal transmission in the nervous system, collagen biosynthesis, proliferation and cell growth, pain physiology, regulation of the immune system, etc.

Ultimately, •O2- contributes to the progression of the process of damage to various structural and functional molecules of the organism, which causes various diseases such as tumor diseases or aging.

Due to the fact that it is not only a radical but a radical anion, the superoxide anion radical (•O2-) is relatively stable in an aqueous medium such as the cytosol. The half-life of •O2- in relatively low concentrations, comparable to those in the cell, eg 0.1 nM, is 14 hours. It follows that in the cell it is necessary to effectively remove •O2-, which is always formed as a by-product during the OF process, that is, during the catabolism of nutrients (food metabolism).

The natural way cells fight against superoxide anion radicals (•O2-) is the existence and action of the natural enzyme superoxide dismutase (SOD). The latter effectively converts •O2- into relatively less harmful hydrogen peroxide (H2O2). A series of experimental findings showed that a decrease in the concentration or lack of SOD causes serious damage to mitochondria, which ultimately leads to cell damage, acceleration of the aging process of the organism, cell death, or the appearance of tumor diseases [P.L. Larsen: Aging and resistance to oxidative damage in Caenorhabditis elegans, Proc. Natl. Acad Sci USA 90 (1993) 8905; K. Aquilano, P. Vigilanza, G. Rotilio, M.R. Ciriolo: Mitochondrial damage due to SODI deficiency in SH-SY5Y neuroblastoma cells: a rationale for the redundancy of SODI, FASEB J. 20 (2006) E930; M.S. Wolin: Extracellular Superoxide Dismutase Depeletion in Hypertension Unmasks a New Role for Angiotensin II in Regulating Cu,Zn-Superoxide Dismutase Activity, Hypertension 48 (2006) 368].

The slightly less harmful hydrogen peroxide (H2O2) thus created is further broken down by disproportionation using the enzyme catalase or glutathione peroxidase-1 to harmless molecules: water and molecular oxygen, which returns to the oxidative phosphorylation (OF) process.

Complementary enzymes are other types of glutathione peroxidase (so-called lipid peroxidase) whose natural function is to split harmful lipid peroxides [created by the reaction of superoxide anion radical (•O2-) with lipids at the allylic positions of unsaturated higher fatty acids] into harmless alcohol derivatives.

The actions of the protective enzymes superoxide dismutase, catalase and glutathione peroxidase, which remove harmful free-radical species or their precursors (e.g. peroxides), are schematically shown in Scheme 1,

I. Action of superoxide dismutase (SOD):

[image]

II. Action of catalase (CAT):

[image]

III. Action of glutathione peroxidase (GPX1) and glutathione reductase (GR):

a) Removal of RA'.

[image]

b) Regeneration of glutatone (GSH):

[image]

Scheme 1. Mechanisms of action of natural enzymes that protect the body from harmful free radicals.

The most important mechanism of action of antioxidants is the elimination of the harmful superoxide anion radical (•O2-). This prevents the above-mentioned reactions in which other harmful free radicals are formed or prevents its binding to nitrogen monoxide (NO).

The reaction of the superoxide anion radical (•O2-) with nitrogen monoxide (NO) produces peroxynitrite anion (O=N-O-O")- Peroxynitrite undergoes homolytic decomposition, resulting in:

(i) peroxonitritic acid, which further decomposes with the formation of extremely harmful free hydroxide radicals (HO•) and nitrogen dioxide (NO2); or

(ii) nitrosoperoxycarbonate (ONOOCO2-; formed by the reaction of peroxynitrite with carbon dioxide) which gives the carbonate radical (•CO2-) and nitrogen dioxide (NO2).

Due to its extremely strong oxidative effect, peroxonitrite can cause a number of harmful effects at the molecular level due to the reaction, for example nitration of tyrosine branches, with a number of functional and building proteins of the cell, nucleic acids, enzymes, etc. [J.S. Beckman: Oxidative damage and tyrosine nitration from peroxynitrite, Chem. Crisp. Toxicol 9 (1996) 836-844].

For the above reasons, and for the purpose of preventing the occurrence of the most common above-mentioned diseases of modern man, a key prerequisite at the molecular level is the maintenance of a low concentration of free radicals, first of all the formation of the superoxide anion radical (•O2-), from which a number of other more or less harmful reactive oxygen species (ROS).

This can be achieved in two ways:

(i) caloric restriction; proper and measured nutrition, which is adjusted in quantity and composition and distributed throughout the day in several meals, which prevents excessive intake of nutrients, and thus a significant reduction in the state of oxidative stress; and/or

(ii) constant intake of harmless antioxidants; which in states of oxidative stress destroy (directly or indirectly) harmful superoxide anion radicals (•O2-) and ine radicals, reducing the level of oxidative stress itself.

The modern pharmaceutical and dietary industry offers a large number of different products that are based on vitamins, different antioxidants, or their combinations, which are claimed to have certain positive effects in the prevention of the above-mentioned diseases or alleviate already existing conditions and diseases.

In a large number of the mentioned antioxidants available on the market with a purpose similar to the formulation of the subject invention, the following most well-known active substances are represented: ecsveratrol (1), green tea extract (Camellia sinensis L.), rooibos extract (Aspalathus linearis L.), sycamore fruit extract ( Silybum marianum L.), ascorbic acid (vitamin C), rutin, quercetin, cyanidin or blueberry extract (Vaccinium myrtillus L.), lycopene or extracts with a higher lycopene content, tetrahydrocurcumin, rosmarinic acid or rosemary extract (Rosmarinus officinalis L.), ellagic acid, chlorogenic acid, oleuropein or olive leaf extract (Olea europea L.), grape seed extract (Vitis vinifera L.), glutathione, noni plant extract, aloe vera extract (Aloe vera L.), etc.

The formulation of the subject invention consists of the following active substances: resveratrol (1), pomegranate extract (Punica granatom L.), olive (Olea europea L.) leaf extract, cinnamon extract (Cinnamomum zeylanicum L.), garlic extract (Allium sativum L. ), grape seed extract (Vitis vinifera L.), coriander extract (Coriandrum sativum L.), saffron extract (Crocus sativus L.) and barley extract (Hardeum vulgare L.). Resveratrol and all the mentioned plant extracts are well described in the literature.

Resveratrol (1) or 3,5-dihydroxy-[2-(4-hydroxyphenyl)ethenyl]benzene (3,5,4'-trihydroxystilbene) is a natural polyphenol with the strongest antioxidant activity.

[image]

Numerous works from the literature prove that resveratrol (1) acts as effective:

(i) cardioprotective [J.F. Lin, S.M. Lin, C.L. Chih, M.W. Nien, H.H. Su, B.R. Hu, S.S. Huang, S.K. Tsai: Resveratrol reduces infarct size and improves ventricular function after myocardial ischemia in rats, Life Sci. 83 (2008) 313-317; A. Y. Chan, V.W. Dolinsky, C.L. Soltys, B. Viollet, S. Baksh, P.E. Light, J.R. Dyck: Resveratrol inhibits cardiac hypertrophy via AMP-activated protein kinase and Akt, J. Biol Chem. 29 (2008) 24194-24201; GM Do, E.Y. Kwon, H.J. Kim, S.M. Jeon, T.Y. Ha, T. Parit, M.S. Choi: Long-term effects of resveratrol supplementation on suppression of atherogenic lesion formation and cholesterol synthesis in apo E-deficient mice, Biochem. Biophys. Crisp. Commun. 374 (2008) 55-59; J.-T. Hwang, D.Y. Kwon, O.J. Parit, M.S. Kim: Resveratrol protects ROS-induced cell death by activating AMPK in H9c2 cardiac muscle cells, Gene Nutr. (2008) 323-326];

(ii) neuroprotective [S. Yousuf, F. Atif, M. Ahmad, N. Hoda, T. Ishrat, B. Khan, F. Islam: Resveratrol exerts its neuroprotective effect by modulating mitochondrial dysfunction and associated cell death during cerebral ischemia, Brain Res. 1250 (2009) 242-253; K.-T. Lu, M.-C. Ko, B.-Y. Chen, J.-C. Huang, C.-W. Hsieh, M.-C. Lee, R.Y.Y. Chiou, B.-S. Wung, C.-H. Peng, Y.L. Yang: Neuroprotective Effects of Resveratrol on MPTP-Induced Neuron Loss Mediated by Free Radical Scavenging, J. Agric. Food Chem. 56 (2008) 6910-6913]; and

(iii) as a potential antiaging agent [J.L. Barger, T. Kayo, T.D. Pugh, T.A. Prolla, R. Weindruch: Short-term consumption of a resveratrol-containing nutraceutical mixture mimics gene expression of long-term caloric restriction in mouse heart, Exp. Gerontol. 43 (2008) 859-866].

The cardioprotective action of resveratrol is based on its antioxidant effect, which improves ventricular function in myocardial ischemia, prevents the formation of atherogenic lesions, and blocks cholesterol biosynthesis.

The neuroprotective effect of resveratrol is based on the binding of harmful free radicals, the regulation of mitochondrial dysfunction and the associated cell death during cerebral ischemia.

Taking resveratrol slows down the aging process according to the caloric restriction mechanism.

Pomegranate (Punica granatom L.) has been used as an edible plant since biblical times [J.A. Duke, P.-A.K. Duke, J.L. Du Cellier: Pomegranate (Punica granatum L.), Duke's Handbook of Medicinal Plants of the Bible, Taylor&Francis Corp. (2008) 362-367]. Pomegranate juice contains ingredients for which the following pharmacological effects have been proven;

(i) antioxidant [C. Guo, J. Wei, J. Yang, J. Xu, W. Pang, Y. Jiang: Pomegranate juice is potentially better than apple juice in improving antioxidant function in elderly subjects, Nutr. Crisp. 28 (2008) 72-77];

(ii) antihypertensive (angiotensin-converting enzyme (ACE) inhibitor [M. Aviram, L. Domfeld: Pomegranate juice consumption inhibits serum angiotensin converting enzyme activity and reduces systolic blood pressure, Atherosclerosis 158 (2001) 195-198]; and

(iii) preventively against atherosclerosis [M. Aviram, L. Domfeld, M. Rosenblat, N. Volkova, M. Kaplan, R. Coleman, T. Havek, D. Presser, B. Fuhnnan: Pomegranate juice consumption reduces oxidative stress, atherogenic modifications to LDL, and platelet aggregation: studies in humans and in atherosclerotic apolipoprotein E-deficient mice, Am. J. Clin. Nutr. 71 (2000) 1062-1076].

Olive (Olea europea L.) leaf extract has been used as a medicinal drug since ancient times [J.A. Duke, P.-A.K. Duke, J.L. Du Cellier: Olive (Olea europea L.), Duke's Handbook of Medicinal Plants of the Bible, Taylor&Francis Corp. (2008) 306-309]. It contains oleuropein and hydroxytyrosol as the main active ingredients. Modern pharmacology of olive leaf extracts proves the following actions:

(i) antioxidant and antihyperlipidemic [H. Jemai, I. Fki, M. Bouziz, Z. Bouallagui, A. EI Feki, H. Isoda, S. Savadi: Lipid-Lowering and Antioxidant Effects of Hydroxytyrosol and Its Triacetylated Derivative Recovered from Olive Tree Leaves in Cholesterol-Fed Rats, J. Agric. Food Chem. 56 (2008) 2630-2636]; and

(ii) antimicrobial [A.N. Sudjana, C. D'Orazio, V. Ryan, N. Rasool, J. Ng, N. Islam, T.V. Riley, K.A. Hammer: Antimicrobial activity of commercial Olea europea (olive) leaf extract, Int. J. Antimicrobial Agents (2009)].

Cinnamon (Cinnamomum zeylanicum L.) is a plant whose ground bark is widely used as a spice. Hydrophilic extracts of cinnamon bark contain more polyphenols and polyhydroxyterpenoids. The latter show the following pharmacological effects:

(i) antioxidant and immunomodulating [H. Hello, J.F. Urban, R.A. Anderson: Cinnamon Polyphenol Extract Affects Immune Responses by Regulating Anti- and Proinflammatory and Glucose Transporter Gene Expression in Mouse Macrophages, J. Nutr. (2008) 833-840]; you

(ii) antiviral [Y. Orihara, H. Hamamoto, H. Kasuga, T. Shimada, Y. Kawaguchi, K. Sekimizu: A silkworm-baculovirus model for assessing the therapeutic effects of antiviral compounds: characterization and application to the isolation of antivirals from traditional medicines, J. Gene. Virol. 89 (2008) 188-194].

Garlic (Allium sativum L.) is a well-known plant that has been used for thousands of years for various medicinal purposes, including for the treatment of cardiovascular diseases, skin diseases, infections, etc. [J.A. Duke, P.-A.K, Duke, J.L. Du Cellier: Garlic (Allium sativum L.), Duke's Handbook of Medicinal Plants of the Bible, Taylor&Francis Corp. (2008) 23-30].

The most well-known active ingredient of garlic is alliin, which breaks down catalyzed by the enzyme allinase to give aiicin (diallylthiosulfinate). Allicin gives garlic its characteristic taste and smell, as well as a number of valuable pharmacological effects [J.A. Milner: Garlic (Allium sativum), Encyclopedia of Dietary Supplements (2005) 229-236; S. Gorinstein, Z. Jastrzebski, J. Namiesnik, Hanna Leontowicz, M. Leontowicz, S. Trakhtenberg: The atherosclerotic heart disease and protective properties of garlic: contemporary data, Mol. Nutr. Food Res. 51 (2007) 1365-1381].

Allicin further undergoes decomposition with the formation of other active ingredients of garlic: diallyldisulfide, diallyltrisulfide, diallyltetrasulfide and aoyene. All the listed sulfur compounds are easily further metabolized with the formation of allylmethylsulfide (which can be felt in the breath exhaled by a person who has consumed garlic).

Due to its strong antioxidant effect, garlic and its extracts prevent diseases that are caused by a prolonged state of oxidative stress, so that they act:

(i) cardioprotective;

(ii) antihyperlipidemic; you

(iii) prevent atherosclerosis (due to the antihyperlipidemic effect and by increasing the concentration of anticoagulants in the blood plasma);

Grape seeds (Vitis vinifera L.) and their extracts are a known natural source of strong antioxidants (+)-catechin, (-)-epicatechin and their oligomers called procyanidins (eg procyanidin C) [D.L. Clouatre, C. Kandaswami: Grape Seed Extract, Encyclopedia of Dietary Supplements (2005) 309-325]. The mentioned compounds are partially esterified with gallic acid, which is why a certain amount of them is present in grape seeds in the form of corresponding gallic esters.

Grape seeds and their extracts show pharmacological effects:

(i) antioxidants;

(ii) cardioprotective;

(iii) hepatoprotective;

(iv) an anti-inflammatory agent; you

(v) skin and wound treatments.

Coriander (Coriandrum sativum L.) or its extracts were used in the treatment of hyperlipidemia, anxiety, inflammatory changes, infections, etc. [J.A. Duke, P.-A.K. Duke, J.L. Du Cellier: Coriander (Coriandrum sativum L.), Duke's Handbook of Medicinal Plants of the Bible, Taylor&Francis Corp. (2008) 138-143].

In modern fartnacognosy, coriander and its extracts are used as:

(i) antioxidants; and

(ii) antihyperlipidemics [P. Dhanapakiam, J.M. Joseph, V.K. Ramaswamy, M. Moorthi, A.S. Kumar: The cholesterol lowering property of coriander seeds (Coriandrum sativum): mechanism of action, J. Environ. Biol. 29 (2008) 53-56; V. Chithra, S. Leelamma: Hypolipidemic effect of coriander seeds (Coriandrum sativum): mechanism of action, Plant Foods Human Nutr. 51 (1997) 167-172].

Saffron (Crocus sativus L.) is a well-known spice and medicinal plant [J.A. Duke, P.-A.K. Duke, J.L. Du Cellier: Saffron (Crocus sativus L.), Duke's Handbook of Medicinal Plants of the Bible, Taylor&Francis Corp. (2008) 143-148].

Modern studies prove that saffron and its extracts act as strong:

(i) antioxidants and preventive agents for tumor diseases [M.Venkatraman, D. Konga, R. Peramaivan, E. Ganapathy, S. Dhanapal: Reduction of Mitochondrial Oxidative Damage and Improved Mitochondrial Efficiency by Administration of Crocetin against Benzo[a]pyrene Induced Experimental Animals, Biol. Pharm. Bull. 31 (2008) 1639-1645];

(ii) antidiabetics [L. Sheng, Z. Qian, Y. Shi, L. Yang, L. Xi, B. Zhao, X. Xu, H. Ji: Crocetin improves the insulin resistance induced by high-fat diet in rats, Brit. J. Pharmacol 154 (2008) 1016-1024];

(iii) cardioprotective [X.-C. Snen, Y. Lu, Z.-Y. Qian: Effects of crocetin on the matrix metalloproteinases in cardiac hypertrophy induced by norepinephrine in rats, J. Asian Nat. Prod. Crisp. 8 (2006) 201-208].

Barley (Hordeum vulgare L.) has been cultivated as a cultivated plant and used for food and medicinal purposes for thousands of years [J.A. Duke, P.-A.K. Duke, J.L. Du Cellier: Barley (Hordeum vulgare L.) Duke's Handbook of Medicinal Plants of the Bible, Taylor&Francis Corp. (2008) 205-209].

Modern medicine proves the strong pharmacological action of barley extracts in the regulation of the immune system due to its p-glucan polysaccharide content [J.J. Volman, J.D. Ramakers, J. Plat: Dietary modulation of immune system function by p-glucans, Physiol & Behavior 94 (2008) 276-284; O. Kanauchi, T. Oshima, A. Andoh, M. Shioya, K. Mitsuvama: Germinated barley foodstuff ameliorates inflammation in mice with colitis through modulation of mucosal immune system, Scand. J. Gastroenterol. 43 (2008) 1346-1352].

The technical problem of more effective treatment of the state of oxidative stress of the organism, and consequently the therapeutic prevention and treatment of various diseases (cardiovascular, nervous and immune system) caused by the permanent state of the same, is solved by this invention in a completely new and significantly more effective way, as will be demonstrated in the detailed description of the invention .

Detailed description of the invention

The subject invention solves the technical problem of effective therapy of the state of oxidative stress, which at the molecular level is the main or one of the main causes of a number of diseases of modern man, such as:

(i) cardiovascular diseases (hypertension, hyperlipidemia, atherosclerosis);

(ii) diabetes (and metabolic syndrome in general);

(iii) diseases of the immune system (low level of immunity, frequent infections);

(iv) diseases of the brain and nervous system (neurodegenerative diseases);

(v) tumor diseases; you

(vi) accelerated aging process.

The subject invention relates to a therapeutic agent containing variable proportions:

(i) resveratrol (1), in a mass proportion of 0.01-10%, preferably 0.1-3%;

(ii) pomegranate extract (Punica granatum L.), in a mass proportion of 0.1-50%, preferably 0.5-25%;

(iii) olive (Olea europea L.) leaf extract, in a mass proportion of 0.1-50%, preferably 0.5-25%;

(iv) cinnamon extract (Cinnamomum zeylanicum L.), in a mass fraction of 0.1-50%, preferably 0.5-25%;

(v) additional plant extracts that support the antioxidant action of the main active ingredients of the formulation: garlic extract (Allium sativum L.), grape seed extract (Vitis vinifera L.), coriander extract (Coriandrum sativum L.), saffron extract (Crocus sativus L. ) and barley extract (Hordeum vulgare L.), in mass fractions of 0.1-50%, preferably 0.5-25%; you

(vi) auxiliary substances that are necessary so that the listed active substances can be used as a finished therapeutic form: tablets, capsules, tinctures, syrups, instant powder for a drink, or instant powder for an effervescent drink, in a mass fraction of 5-99.19 %;

Preparation of the formulation of the subject invention

The formulation of the subject invention includes the following finished forms suitable for oral administration:

(i) tablets;

(ii) capsules;

(iii) tincture (solution);

(iv) syrup;

(vi) instant potion burst; you

(vii) instant powder for effervescent drink.

Mass fractions of the main active substances [resveratrol (1), pomegranate extract (Punica granatom L.), olive (Olea europea L.) leaf extract and cinnamon extract (Cinnamomum zeylanicum L.)], and auxiliary active substances [garlic extract (Allium sativum L.) grape seed extract (Vitis vinifera L.), coriander extract (Coriandrum sativum L.), saffron extract (Crocus sativus L.) and barley extract (Hordeum vulgare L.)] are listed at the beginning of the chapter "Detailed description of the invention" .

The active substances used for the preparation of the formulation of the subject invention are characterized by the following level of purity/content of active substances:

(i) resveratrol (1), content >98%;

(ii) pomegranate extract (Punica granatum L.), ellagic acid content >90%;

(iii) olive (Olea europea L.) leaf extract, oleuropein content >20%;

(iv) cinnamon extract (Cinnamomum zeylanicum L.), polyphenol content >50%;

(v) garlic extract (Allium sativum L.), allicin content>10%;

(vi) grape seed extract (Vitis vinifera L.), proanthocyanidin content >95%;

(vii) coriander extract (Coriandrum sativum L.), a standard commercially available dry extract;

(viii) saffron extract (Crocus sativus L.), a standard commercially available dry extract;

(ix) and barley extract (Hordeum vulgare L.), p-glucan content >70%.

Alternatively, instead of pure resveratrol, extracts of plants rich in resveratrol can be used to prepare the formulation of the subject invention, such as extracts of Polygonum cuspidatum L. The average expert in the field knows that in this case the amount of such raw material should be converted to resveratrol content and at the same time the content of excipients should be corrected for the proportion of ballast substances from extracts.

The excipients required to prepare the above-mentioned finished therapeutic forms are common substances known to the average person skilled in the art. They are selected from the group consisting of various technological additives that are necessary to incorporate the listed active substances in powder form into the final pharmaceutical or dietary form suitable for oral use: fillers, binders, disintegrators, lubricants, emulsifiers, diluents, humectants, thickeners, preservatives , antioxidants, stabilizers, dyes, fragrances, pH control agents, and other functional additives.

In the case of solid oral forms such as tablets or capsules, substances well known to a person skilled in the art such as microcrystalline cellulose, lactose monohydrate, calcium hydrogen phosphate, sucrose, glucose, fructose, silicon dioxide, talc, sorbitol, mannitol, starch, modified starches can be used as fillers. , inulin, or a mixture of these substances.

In the case of solid oral forms such as powdered instant drinks, substances such as sucrose, glucose, fructose, xylitol, citric acid, sorbitol, mannitol, starch, modified starches, sodium bicarbonate, or mixtures of these substances can be used as fillers.

Polyvinylpyrrolidone, lactose monohydrate, glucose, mannitol, sorbitol, starch, modified starches, carrageenans, alginates, sodium carboxymethylcellulose, methylcellulose, 2-hydroxyethylcellulose, sucrose, gelatin, or mixtures of these substances can be used as binders in solid oral forms.

As disintegrators in solid oral forms, the following can be used: polyvinylpyrrolidone, copolymers of polyvinylpyrrolidone, agar-agar, calcium carbonate, starch, alginic acid, calcium silicate, sodium starch glycolate, or mixtures of the aforementioned substances.

The following can be used as lubricants in solid oral forms such as tablets: talc, stearic acid, magnesium stearate, calcium stearate, zinc stearate, solid polyethylene glycols, solid polypropylene glycols, or mixtures of the aforementioned substances.

As emulsifiers in liquid preparations such as tincture or syrup can be used: ethoxylates of higher fatty alcohols such as polyoxyethylene(2) lauryl ether, polyoxyethylene(10) lauryl ether, polyoxyethylene(23) lauryl ether, polyoxyethylene(2) stearyl ether, polyoxyethylene(10) stearyl ether, polyoxyethylene( 23) stearyl ether, polyoxyethylene (2) oleyl ether, polyoxyethylene (10) oleyl ether, polyoxyethylene (23) oleyl ether, where the numbers 2, 10 and 23 indicate the average number of ethylene glycol units attached to a higher fatty alcohol; higher fatty acid ethoxylates such as polyoxyethylene(2) laurate, polyoxyethylene(10) stearate, polyoxyethylene(23) oleate, where the numbers 2, 10 and 23 indicate the average number of ethylene glycol units attached to the higher fatty acid; sorbitan esters such as polyoxyethylene sorbitan monolaurate (Tween 20), polyoxyethylene sorbitan monopahnitate (Tween 40), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan tristearate (Tween 65), polyoxyethylene sorbitan sesquioleate (Tween 85) ; castor oil or its ethoxylates; hydrogenated castor oil or its ethoxylates; or mixtures of these substances.

As diluents or fillers in the liquid forms of the formulation of the subject invention, such as tinctures or syrups, other substances selected from the group consisting of: ethanol, glycerol, 1,2-propylene glycol, polyethylene glycol 400, polyethylene glycol 600, polyethylene glycol can be used in addition to purified water. 1000, polyethylene glycol 2000, polyethylene glycol 4000, polyethylene glycol 6000, polypropylene glycol 425, polypropylene glycol 100, other polypropylene glycols, polyglycerols, sorbitol, sucrose, glucose, glucose syrup, fructose, honey, mullin, or mixtures of the above substances.

As humectants in the formulation of the present invention, substances selected from the group consisting of: glycerol, 1,2-propylene glycol, polyethylene glycol 400, polyethylene glycol 600, polyethylene glycol 1000, polyethylene glycol 2000, polyethylene glycol 4000, polyethylene glycol 6000, polypropylene glycol 425, polypropylene glycol 1000, others can be used. polypropylene glycols, polyglycerols, sorbitol, or mixtures of these substances.

In the formulation of the subject invention in liquid forms such as tinctures, syrups or drinks, for thickening and/or suspending, auxiliary substances selected from the group consisting of:

polyacrylic acid or its salts with sodium hydroxide, potassium hydroxide, or ammonium hydroxide, methylcellulose, sodium carboxymethylcellulose, 2-hydroxyethylcellulose, 2-hydroxypropylcellulose, bentonite, starch, modified starches, gelatin, polyglycerols, polyethylene glycol 400, polyethylene glycol 600, polyethylene glycol 1000, polyethylene glycol 2000, polyethylene glycol 4000, polyethylene glycol 6000, polypropylene glycol 425, polypropylene glycol 2000, agar-agar, carrageenan, gum arabic, tragacanth, alginic acid, sodium alginate, pectin, or mixtures of the above substances.

The following substances are used as preservatives for the long-term preservation of the microbiological purity of the formulation of the invention in all cases where the content of the aqueous phase is relatively high, as is the case with syrups: methyl 4-hydroxybenzoate, ethyl 4-hydroxybenzoate, propyl 4-hydroxybenzoate, butyl 4-hydroxybenzoate, sorbic and/or benzoic acid or their salts with pharmaceutically acceptable bases, or mixtures of the aforementioned substances.

One or more compounds from the group consisting of: 2,6-di-tert-butyl-4-hydroxytoluene (BHT), tert-butylhydroxyanisole (BHA), tocopherol, tocopheryl acetate, other tocopherol esters can be used as antioxidants in the formulation of the present invention. , ascorbic acid, ascorbyl palmitate, lecithin, sodium sulfite, sodium metabisulfite, sodium thiosulfate, sulfur(IV) oxide, cysteine hydrochloride, N-acetylcysteine, propyl gallate, or mixtures of the aforementioned substances.

As stabilizers in the formulation of the present invention, one or more additives can be used that act as ligands for transition metals, which, converting them into complexes, stabilize the formulation in the long term against possible metal-catalyzed decomposition of one or more components of the formulation. Stabilizers are selected from the group consisting of disodium citrate [Na2(C(OH)(COOH)(CH2COO)2] or other salts of citric acid, disodium tartrate or other salts of tartaric acid, or mixtures of the aforementioned substances.

The formulation of the subject invention can be produced according to the procedures for the preparation of the specified forms as is customary in the art [S.C. Gad (Ed.): Pharmaceutical Manufacturing Handbook: Production and Processes, Wiley (2008)].

Thus, tablets are prepared by direct compression of a homogenized tablet mixture or wet granulation with subsequent tableting of the granulate. In any case, in addition to the listed active substances, the tablet also contains one or more auxiliary substances that are necessary for production: filler, binder, disintegrant, lubricant, and possibly coating agents, etc.

Capsules are produced by encapsulating a homogenized mixture of the listed active substances and one or more auxiliary substances or their granules in standard gelatin or vegetable capsules.

The tincture (solution) is produced by mixing the listed active substances in a mixture of suitable diluents (solvents; usually a water-alcohol medium) with the addition of one or more auxiliary substances. The procedure is carried out at room or slightly elevated temperature.

The syrup is produced by mixing the above-mentioned active substances in the basic syrup (base), which, according to the rules of the profession, may contain an aqueous solution of sucrose, honey, glucose, glucose syrup, fructose, etc. In the case of using sweeteners, the aqueous suspension is thickened using standard (above listed) food/pharmaceutical thickeners. Sweeteners are selected from the group consisting of: sodium saccharin, potassium acesulfame, sucralose, sodium or calcium cyclamate, xylitol, sorbitol, stevia plant extract, or mixtures of the aforementioned substances.

Instant drink powder is produced by dry homogenization (mixing) of the listed active substances with auxiliary substances selected from the following groups: diluents (usually sabarose, glucose or mixtures of sucrose and other sweet sugars and citric acid), emulsifiers (usually polyethylene glycol, sorbitan esters of higher fatty acids with H.L.B>10) with the addition of other technological additives such as colors, flavors, preservatives, etc.

In addition to the above, sodium hydrogencarbonate (NaHCO3) is added to powders for effervescent instant drinks, which reacts with citric acid with the release of gaseous carbon dioxide (CO2), which creates foaming when the instant powder is dissolved in water and gives a refreshing taste.

The formulation of the subject invention can be the basis for other finished pharmaceutical forms for oral administration, such as suspensions or drops (which are a variant of tincture/solution), etc., which are not described here, but such application does not change the essence of the subject invention.

Pharmacological action of the formulation of the subject invention

During the examination of the antioxidant activity of various natural compounds and plant extracts, we found that the formulation of the subject invention unexpectedly shows a significantly stronger antioxidant activity than resveratrol itself (1), which is generally known as "the strongest natural antioxidant".

To test the strength of the antioxidant effect of the formulation of the subject invention, resveratrol as a standard, and the combination of resveratrol with the mentioned extracts, a well-known method of in vitro determination of antioxidant activity based on the inhibition (reduction) of free 1,1-diphenyl-2-picrylhydrazinyl (DPPH•) was used radicals [F. Nikhat, D. Satynarayana, E.V.S. Subhramanvam: Isolation, Characterization, and Screening of Antioxidant Activity of the Roots of Syzygiumcuminii (L) Skeel, Asian J. Res. Chem. 2 (2009) 218-221; J. Muselík, M. Garcia-Alonso, M.P. Martin-López, M. Žemlička, J.C. Rivas-Gonzalo: Measurement of Antioxidant Activity of Wine Catechins, Procyanidins, Anthocyanins, and Pyranoanthocyanins, Int. J. Mol. Sci. 8 (2007) 797-809; T.K. Panovska, S. Kulevanova, M. Stefova: In vitro antioxidant activity of some Teucrium species (Lamiaceae), Acta Pharm. 55 (2005) 207-214].

The DPPH• inhibition method is based on the reaction of the examined antioxidant with the mentioned free radicals. If the tested substance has a reduction potential (if it is an antioxidant), then it reduces DPPH radicals to 1,1-diphenyl-2-picrylhydrazine (DPPH-H). As the DPPH-radical solution is purple and the final hydrazine DPPH-H is yellow, the quantity of the reaction is determined spectrophotometrically, by measuring the absorbance at 517 nm.

By applying the aforementioned analytical in vitro method, it was determined that the mixture of resveratrol (1) and extracts of pomegranate (Punica granatum L.), olive (Olea europea L.) leaves and cinnamon (Cinnamomum zeylanicum L.) acts as a much stronger antioxidant than resveratrol itself and and from other possible combinations:

(i) resveratrol and pomegranate extract;

(ii) resveratrol and olive leaf extract;

(iii) resveratrol and cinnamon extract;

(iii) resveratrol and pomegranate and olive leaf extracts;

(iv) resveratrol and pomegranate and cinnamon extracts;

(v) resveratrol and olive leaf and cinnamon extracts.

The test results are presented in Table 1.

Table 1. Results of determination of antioxidant activity of resveratrol (1), two- and three-component mixtures of resveratrol with plant extracts of pomegranate (Punica granatum L.), olive leaf (Olea europea L.) and cinnamon (Cinnamomum zeylanicum L.), and formulations of the subject invention by the method of inhibition of the 1,1-diphenyl-2-picriIhydrazinyl (DPPH) radical.1

[image]

1 A detailed description of the analysis procedure is described in Example 7 of the embodiment of the invention.

2 In order to compare the results, an optimal mass concentration (y) of 10 µg/mL was determined for the resveratrol standard, while all other experiments were recalculated to the same total concentration. The share of extracts with a lower composition of the active substance increased at the expense of ballast substances.

3 Obtained from spectrophotometric measurement of absorbance at 517 nm. It indicates the percentage of DPPH that reacted (reduced) under the action of the tested antioxidant sample.

4 IC50 is a value that indicates the mass concentration (γ/µg/mL) at which the tested antioxidant inhibits (reduces) 50% of the starting amount (quantity) of DPPH.

5 Mass concentrations of active substances: resveratrol + pomegranate extracts + olive leaf extract + cinnamon extract = 2+2.2+10+4 µg/mL with the addition of garlic, grape seed, coriander, saffron and barley extracts, each in a concentration of 1.0 µg/mL (ready-made mixture equivalent to 10 ng/mL resveratrol).

From the obtained results, it was concluded that the mixture of resveratrol (1) with extracts of pomegranate, olive leaf and cinnamon really acts as a significantly stronger antioxidant than the same amount of resveratrol itself. As the latter is generally known as the "strongest natural antioxidant", the results undoubtedly point to some kind of synergistic action. Quantitatively expressed (comparison of results in experiments 12 and 1), the increase in the degree of inhibition of DPPH-radicals amounts to approx. 7% (increase with 87 to 93%), with a significant decrease in the IC50 value from 2.72 to 2.16, which shows an increase in antioxidant activity by 20.6%.

At the same time, all tested two- and three-component mixtures of resveratrol and extracts of pomegranate, olive leaf and cinnamon also have high antioxidant activity, but significantly weaker than resveratrol itself (standard) (Experiments 2-7).

Pomegranate extract in combination with resveratrol also seems to show extremely high antioxidant activity probably due to the fact that it is almost pure ellagic acid (the pomegranate extract used contained >90% ellagic acid) (Experiment 2). However, there is no indication of a synergistic effect.

Two- and three-component mixtures of pomegranate, olive leaf and cinnamon extracts also act as strong antioxidants, but the level of this activity is significantly lower than the experiments in which resveratrol was also present (Experiments 8-11).

Finally, the formulation of the subject invention (Experiment 13), which, in addition to resveratrol (1) and extracts of pomegranate, olive leaf and cinnamon, also contained extracts of garlic, grape seeds, coriander, saffron and barley, showed a slightly higher antioxidant effect (comparison of the results of Experiment 12 and 13). It cannot be said with certainty that the last extracts act additionally synergistically, but they definitely enhance the antioxidant effect to some extent. As it is known from pharmacognosy that these extracts, each in their own way, help with conditions and diseases such as hypertension, hyperlipidemia, diabetes, obesity, weak immunity, infections, etc., they are included in the definition of the formulation of the subject invention as "additional active substances" because they really support the basic antioxidant action of the "main active substances": resveratrol and extracts of pomegranate, olive leaf and cinnamon.

The formulation of the subject invention in the form of capsules (the preparation is described in Example 2) was tested on several people who suffered from or suffered from the above-mentioned common diseases whose origin and development is based on constant exposure to a state of oxidative stress, mainly due to excessive food intake. Often, the clinical history of such patients includes smoking, poor-quality nutrition, genetic predisposition to hypertension and/or hyperlipidemia, bad lifestyle habits, and often alcoholism.

The composition of the formulation of the subject invention in the form of capsules that was used to test the effectiveness in patients was as follows:

Content of active substances in one capsule:

(i) 5 mg resveratrol;

(ii) 75 mg of pomegranate extract (Punica granatum L.);

(iii) 75 mg of olive leaf extract (Olea europea L.);

(iv) 25 mg of cinnamon extract (Cinnamomum zeylanicum L.);

(v) 30 mg of garlic extract (AIlium sativum L.);

(vi) 80 mg of grape seed extract (Vitis vinifera L.);

(vii) 25 mg of coriander extract (Coriandrum sativum L.);

(viii) 20 mg of saffron extract (Crocus sativus L.);

(ix) 25 mg of barley extract (Hordeum vulgare L.);

The study regimen was as follows: Duration: 30 days. Dosage: 2×2 capsules per day (after breakfast and after lunch).

Case 1: Male, 60 years old, end-stage renal disease - dialysis, type II diabetes, atherosclerosis. Values of key parameters before applying the formulation of the subject invention: creatinine 823, blood glucose (guk) 11.2, blood pressure 190/110, LDL cholesterol 6.3. Values after treatment: creatinine 523, glucose 7.2, blood pressure 160/110, LDL cholesterol 4.1.

Case 2: Male, 54 years old, elevated blood cholesterol. Values before applying the formulation of the subject invention: LDL cholesterol 5.8. Value after application: LDL cholesterol 3.2.

Case 3: Male, 48 years old, type II diabetes and elevated cholesterol. Values of key parameters before applying the formulation of the subject invention: guk 8.5; LDL cholesterol 6.1. Values after application: guk 6.6; LDL cholesterol 2.8.

Case 4: Male, 34 years old, hypertension. Value before applying the formulation of the subject invention: pressure 160/90. Value after application: 125/90

Case 5. Woman, 48 years old, Alzheimer's disease, early stage of dementia, apathy and inability to remember new information, elevated cholesterol. Key values before applying the formulation of the subject invention: cholesterol LDL cholesterol 5.2. Value after application: LDL cholesterol 2.6. At the end of the Study, the disappearance of apathy and subjective improvement (feeling of better energy), improvement in the memory of new information was noted.

Conclusion

From the in vitro test of the antioxidant effect by the method of inhibition (reduction) of DPPH-free radicals, it was determined that the formulation of the subject invention unexpectedly shows a significantly stronger antioxidant effect than the equivalent amount (amount) of resveratrol itself.

In tests on patients, the positive effect of the formulation of the subject invention on diseases caused by a prolonged state of oxidative stress was confirmed: hypertension, hyperlipidemia, atherosclerosis, neurodegenerative diseases (Alzheimer's disease), etc.

The possible reason for the unexpected synergistic effect of resveratrol with the active ingredients of other extracts lies in the fact that all antioxidants do not have the same mechanism of action. According to the literature, the possible mechanisms of action of antioxidants are:

(i) reductive action;

(ii) the ability to bind (addition; e.g. to the C=C bond) free radicals; or

(iii) binding (in a complex) of transition metal cations that can participate in redox reactions (primarily Fe2+/Fe3+, Cu2+, Mn2+);

In this way, antioxidants prevent the formation (initiation) or remove already formed (blockade of chain reaction propagation) harmful free radicals.

It is likely that the antioxidants of the formulation of the subject invention act by several mechanisms at the same time, which leads to a quantitatively extremely strong antioxidant activity (synergism), which on a macro-level has a positive effect on the mentioned diseases caused by a state of oxidative stress.

Examples of embodiments of the invention

Example 1. Preparation of the formulation of the subject invention in the form of 500 mg tablets

Composition (100 g tablet mixture): (a) Resveratrol (1.00 g; 1%), (b) pomegranate extract (Punica granatom L.; 5.00 g; 5%), (c) olive leaf extract (Olea europea L.; 5.00 g; 5%), (d) cinnamon extract (Cinnamomum zeylanicum L.; 5.00 g; 5%), (e) garlic extract (Allium sativum L.; 5.00 g; 5 %), (f) grape seed extract (Vitis vinifera L.; 5.00 g; 5%), (g) coriander extract (Coriandrum sativum L.; 5.00 g; 5%), (h) saffron extract ( Crocus sativus L.; 5.00 g; 5%), (i) barley extract (Hordeum vulgare L.; 5.00 g; 5%), (j) lactose monohydrate (55.20 g; 55.2%) , (k) sodium starch glycolate (1.50 g; 1.5%), (l) polyvinylpyrrolidone (1.50 g; 1.5%), (m) vegetable magnesium stearate (0.80 g; 0.8 %).

Preparation procedure: Weighed ingredients (a)-(l) were mixed and homogenized by mixing at room temperature for 15 minutes with a little purified water. Then (m) was added, and the resulting mixture was stirred at room temperature for 15 minutes. Then, the homogenized granulate was dried at 40 °C in a high vacuum for 1 h, ground, sieved and pressed into tablets. Approximately 195 tablets were obtained. Average tablet weight 512 mg.

Example 2. Preparation of the formulation of the subject invention in the form of 600 mg capsules

Composition (100 g of encapsulation mixture): (a) Resveratrol (1.00 g; 1%), (b) pomegranate extract (Punica granatum L; 15.00 g; 15%), (c) olive leaf extract (Olea europea L; 15.00 g; 15%), (d) cinnamon extract (Cinnamomum zeylanicum L; 5.00 g; 5%), (e) garlic extract (Allium sativum L; 6.00 g; 6%), (f) grape seed extract (Vitis vinifera L; 16.00 g; 16%), (g) coriander extract (Coriandrum sativum L; 5.00 g; 5%), (h) saffron extract (Crocus sativus L; 4 .00 g; 4%), (i) barley extract (Hordeum vulgare L; 5.00 g; 5%), (j) inulin (17.20 g; 17.2%), (k) talc (10, 00 g; 10%), (1) magnesium stearate (0.80 g, 0.8%).

Preparation procedure: Weighed ingredients (a)-(l) were mixed, ground and homogenized by mixing at room temperature for 30 minutes. The homogenous mixture thus prepared was filled into standard "0" type gelatin capsules using a manual laboratory capsule maker. Approximately 200 capsules were obtained. The obtained capsules are characterized by a net mass of approximately 500 mg and a gross mass of approximately 600 mg (the mass of the empty capsule is 95-100 mg).

Composition of active substances in one capsule:

(i) 5 mg resveratrol;

(ii) 75 mg of pomegranate extract (Punica granatum L);

(iii) 75 mg of olive leaf extract (Olea europea L);

(iv) 25 mg of cinnamon extract (Cinnamomum zeylanicum L);

(v) 30 mg of garlic extract (Allium sativum L);

(vi) 80 mg of grape seed extract (Vitis vinifera L);

(vii) 25 mg of coriander extract (Coriandrum sativum L);

(viii) 20 mg of saffron extract (Crocus sativus L.);

(ix) 25 mg of barley extract (Hordeum vulgare L.);

Example 3. Preparation of the formulation of the subject invention in the form of a tincture

Composition (1 L tincture): (a) Resveratrol (2.50 g; 0.25%), (b) pomegranate extract (Punica granatom L; 10.00 g; 1%), (c) olive leaf extract (Olea europea L; 10.00 g; 1%), (d) cinnamon extract (Cinnamomum zeylanicum L.; 5.00 g; 0.5%), (e) garlic extract (Allium sativum L.; 5.00 g; 0.5%), (f) grape seed extract (Vitis vinifera L; 5.00 g; 0.5%), (g) coriander extract (Coriandrum sativum L; 5.00 g; 0.5%), ( h) saffron extract (Crocus sativus L; 5.00 g; 0.5%), (i) barley extract (Hordeum vulgare L.; 5.00 g; 0.5%), (j) 96% ethanol (400 .00 g; 40%), (k) purified water (547.50 g; 54.75%).

Preparation procedure: Components (a)-(i) were added to the mixture of solvents (j) and (k) respectively. The suspension was intensively stirred at room temperature for 1 h. It was then filtered. A tincture was obtained in the form of a reddish-brown clear liquid. The finished tincture is packed in dark glass bottles of 50 mL with a dropper. Alcohol content approx. 38-40%.

Example 4. Preparation of the formulation of the subject invention in the form of syrup

Composition (1 L syrup): (a) Resveratrol (2.50 g; 0.25%), (b) pomegranate extract (Punica granatim L; 10.00 g; 1%), (c) olive leaf extract (Olea europea L.; 10.00 g; 1%), (d) cinnamon extract (Cimamomum zeykmicum L.; 5.00 g; 0.5%), (e) garlic extract (Allium sativum L; 5.00 g; 0.5%), (f) grape seed extract (Vitis vinifera L; 5.00 g; 0.5%), (g) coriander extract (Coriandrum sativum L; 5.00 g; 0.5%), ( h) saffron extract (Crocus sativus L; 5.00 g; 0.5%), (i) barley extract (Hordeum vulgare L; 5.00 g; 0.5%), (j) sucrose (550.00 g ; 55%), (k) methyl 4-hydroxybenzoate (2.00 g; 0.2%), (l) propyl 4-hydroxybenzoate (1.00 g; 0.1%), (m) 96% ethanol ( 10.00 g; 1%), (n) polyoxyethylene sorbitan monooleate (Tween 80; 10.00 g; 1%), (o) purified water (374.50 g; 37.45%).

Preparation procedure: Ingredient (o) is heated to 60 °C. Then (j) and (n) were added, and the mixture was intensively stirred at that temperature until a thick (syrupy) liquid was formed. Ingredients (a)-(i) were added to the resulting thick liquid. The mixture was stirred at temperatures from 60°C to room temperature for 2 h. Then a previously prepared solution of ingredients (k) and (1) in solvent (m) was added. The product thus prepared was filtered through a thin filter paper. The syrup of the formulation of the subject invention was obtained in the form of a thick orange-brown to red-brown thick liquid.

Example 5. Preparation of the formulation of the subject invention in the form of a powder for an instant drink

Composition (1000 g powder for instant drink): (a) Resveratrol (2.50 g; 0.25%), (b) pomegranate extract (Punica granatum L; 10.00 g; 1%), (c) olive extract leaf (Olea europea L; 10.00 g; 1%), (d) cinnamon extract (Cinnamomum zeylanicum L; 5.00 g; 0.5%), (e) garlic extract (Allium salivum L.; 5.00) g; 0.5%), (f) grape seed extract (Vitis vinifera L; 5.00 g; 0.5%), (g) coriander extract (Coriandrum sativum L; 5.00 g; 0.5%) , (h) saffron extract (Crocus sativus L; 5.00 g; 0.5%), (i) barley extract (Hordeum vulgare L.; 5.00 g; 0.5%), (j) sucrose (890 .50 g; 89.05%), (k) anhydrous citric acid (50.00 g; 5%), (l) methylcellulose (2.00 g; 0.2%), (m) blackberry flavor (5, 00 g; 0.5%).

Preparation procedure: Ingredients (j) and (k) were ground and homogenized by mixing at room temperature for 15 minutes. Finely ground ingredients (a)-(i), (l) and (m) were added to the resulting mixture, and the mixture was homogenized by stirring at room temperature for 15 minutes. The product thus prepared is in the form of a pale reddish to pale brown powder, packaged in small paper bags covered with aluminum foil, 7.00 g each.

Recommended dosage: Dissolve one sachet (7.00 g) in 1 dcL (100 mL) of water and stir. The result is a dull red to red-brown, slightly cloudy drink with a refreshing blackberry taste.

Example 6. Preparation of the formulation of the subject invention in the form of a powder for an effervescent instant drink

Composition (1000 g powder for effervescent instant drink): (a) Resveratrol (2.50 g; 0.25%), (b) pomegranate extract (Punica granatum L.; 10.00 g; 1%), (c) olive leaf extract (Olea europea L; 10.00 g; 1%), (d) cinnamon extract (Cinnamomum zeylanicum L; 5.00 g; 0.5%), (e) garlic extract (Allium sativum L; 5, 00 g; 0.5%), (f) grape seed extract (Vitis vinifera L; 5.00 g; 0.5%), (g) coriander extract (Coriandrum sativum L; 5.00 g; 0.5% ), (h) saffron extract (Crocus sativus L; 5.00 g; 0.5%), (i) barley extract (Hordeum vulgare L.; 5.00 g; 0.5%), (j) sucrose ( 857.50 g; 85.75%), (k) anhydrous citric acid (60.00 g; 6%), (l) sodium bicarbonate (25.00 g; 2.5%), (m) cinnamon flavor ( 5.00 g; 0.5%).

Preparation procedure: Ingredients (j) and (k) were dried in a vacuum oven at 50°C for 1 hour. They were then ground and homogenized by mixing at room temperature for 15 minutes. Component (1) was dried separately in a vacuum oven at 80°C for 1 h. Finely ground ingredients (a)-(i) and (m) were added to the resulting mixture, and the mixture was homogenized by stirring at room temperature for 15 minutes. The thus prepared product in the form of orange-reddish to pale brown powder is packed in small paper bags covered with aluminum foil of 7.00 g each.

Recommended dosage: Dissolve one sachet (7.00 g) in 1 dcL (100 mL) of water and stir. The result is an orange-brown to red-brown, slightly cloudy effervescent drink with a refreshing cinnamon taste.

Example 7. In vitro determination of the antioxidative effect of the formulation of the subject invention

In vitro determination of the antioxidant effect of the formulation of the subject invention was carried out using the method of inhibition (reduction) of 1,1-diphenyl-2-picrylhydrazyl (DPPH•) free radicals. The tested antioxidant reacts with DPPH•, which is accompanied by a change in color from purple to yellow. This change is determined spectrophotometrically by measuring absorbance at 517 nm.

Reagents: (a) 0.1 mM methanolic solution of DPPH- was prepared by dissolving 39.4 mg of the purchased substance (Sigma-Aldrich) in methanol (analytical grade; 1 L); (b) 1% aqueous solution of ascorbic acid;

Procedure: 1.0 mL of DPPH• solution was added to 3.0 mL of a sample solution of the tested antioxidant in methanol at a concentration of 10 µg/mL (or equivalent for sample mixtures; see Table 1). After 30 minutes, the absorbance was measured at 517 nm. A blank test is prepared without the addition of the antioxidant sample. As standards, resveratrol and ascorbic acid were used (for preliminary verification of the method).

Calculation: A lower absorbance of the reaction mixture means a higher degree of DPPH• inhibition, consequently a higher antioxidant activity. The ability to reduce DPPH-free radicals was calculated by including the obtained results in the formula:

[image]

Acontrol = absorbance of the blank

Ausample = absorbance of the sample with the addition of the tested sample

Determination of IC50: The described experiments were repeated while varying the total concentration of tested antioxidant samples (or mixtures as shown in Table 1) of 1, 2, 4, 8 and 16 µg/mL. From the obtained results, the IC50 values were graphically determined as the concentration (y) at which the given antioxidant inhibits (reduces) 50% of the present quantity (quantity) of DPPH• radicals. The results are presented in Table 1.

Example 8. Pharmacological effects of the formulation of the subject invention

To test the effectiveness, a study was conducted on five patients who suffered from one or more diseases or conditions: hypertension, hyperlipidemia, Alzheimer's disease. The formulation of the subject invention was used in the form of capsules, the preparation of which is described in Example 2. Composition of active substances in one capsule:

(i) 5 mg resveratrol;

(ii) 75 mg of pomegranate extract (Punica granatum L.);

(iii) 75 mg of olive leaf extract (Olea europea L.);

(iv) 25 mg of cinnamon extract (Cinnamomum zeylanicum L.);

(v) 30 mg of garlic extract (Allium sativum L);

(vi) 80 mg of grape seed extract (Vitis vinifera L.);

(vii) 25 mg of coriander extract (Coriandrum sativum L.);

(viii) 20 mg of saffron extract (Crocus sativus L);

(ix) 25 mg of barley extract (Hordeum vulgare L.);

The study regimen was as follows: Duration: 30 days. Dosage: 2×2 capsules per day (after breakfast and after lunch).

Case 1: Male, 60 years old, end-stage renal disease - dialysis, type II diabetes, atherosclerosis. Values of key parameters before applying the formulation of the subject invention: creatinine 823, blood glucose (guk) 11.2, blood pressure 190/110, LDL cholesterol 6.3. Values after treatment: creatinine 523, glucose 7.2, blood pressure 160/110, LDL cholesterol 4.1.

Case 2: Male, 54 years old, elevated blood cholesterol. Values before applying the formulation of the subject invention: LDL cholesterol 5.8. Value after application: LDL cholesterol 3.2.

Case 3: Male, 48 years old, type II diabetes and elevated cholesterol. Values of key parameters before applying the formulation of the subject invention: guk 8.5; LDL cholesterol 6.1. Values after application: guk 6.6; LDL cholesterol 2.8.

Case 4: Male, 34 years old, hypertension. Value before applying the formulation of the subject invention: pressure 160/90. Value after application: 125/90

Case 5. Woman, 48 years old, Alzheimer's disease, early stage of dementia, apathy and inability to remember new information, elevated cholesterol. Key values before applying the formulation of the subject invention: cholesterol LDL cholesterol 5.2. Value after application: LDL cholesterol 2.6. At the end of the Study, the disappearance of apathy and subjective improvement (feeling of better energy), improvement in the memory of new information was noted.

Claims (20)

1. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, characterized by the fact that it consists of variable proportions: (i) resveratrol (1), in a mass proportion of 0.01-10%, preferably 0.1-3%; [image] (ii) pomegranate extract (Punica granatum L), in a mass proportion of 0.1-50%, preferably 0.5-25%; (iii) olive (Olea europea L.) leaf extract, in a mass proportion of 0.1-50%, preferably 0.5-25%; (iv) cinnamon extract (Cinnamomum zeylanicum L), in a mass proportion of 0.1-50%, preferably 0.5-25%; (v) additional plant extracts that support the antioxidant action of the main active ingredients of the formulation: garlic extract (Allium sativum L.), grape seed extract (Vitis vinifera L.), coriander extract (Coriandrum sativum L), saffron extract (Crocus sativus L) and barley extract (Hordeum vulgare L.), in mass fractions of 0.1-50%, preferably 0.5-25%; you (vi) auxiliary substances that are necessary so that the listed active substances can be used as a finished therapeutic form: tablets, capsules, tinctures, syrups, instant powder for a drink or instant powder for an effervescent drink, in a mass proportion of 5-99.19% ; 2. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claim 1, indicated that the auxiliary substance is selected from the group consisting of fillers, binders, disintegrators, lubricants, emulsifiers, diluents, humectants, thickeners , preservatives and antioxidants. 3. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is a filler selected from the group consisting of microcrystalline cellulose, silicon dioxide, talc, lactose monohydrate, calcium hydrogen phosphate, sucrose, glucose, fructose, sorbitol, mannitol, starch, modified starches, inulin, xylitol, citric acid, sodium bicarbonate, or mixtures of the aforementioned substances. 4. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is a binder selected from the group consisting of polyvinylpyrrolidone, lactose monohydrate, glucose, mannitol, sorbitol, starch , modified starches, carrageenans, alginates, sodium carboxymethylcellulose, methylcellulose, 2-hydroxyethylcellulose, sucrose, gelatin, or mixtures of the aforementioned substances. 5. Formulation based on resveratrol and nam, olive leaf and cinnamon extracts as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is a disintegrant selected from the group consisting of polyvinylpyrrolidone, copolymers of polyvinylpyrrolidone, agar-agar, calcium carbonate, starch, alginic acid, calcium silicate, sodium starch glycolate, or mixtures of these substances. 6. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is a lubricant selected from the group consisting of talc, stearic acid, magnesium stearate, calcium stearate, zinc stearate, solid polyethylene glycols, solid polypropylene glycols, or mixtures of these substances. 7. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is an emulsifier selected from the group consisting of: ethoxylates of higher fatty alcohols such as polyoxyethylene (2) lauryl ether, polyoxyethylene(10) lauryl ether, polyoxyethylene(23) lauryl ether, polyoxyethylene(2) stearyl ether, polyoxyethylene(10) stearyl ether, polyoxyethylene(23) stearyl ether, polyoxyethylene(2) oleyl ether, polyoxyethylene(10) oleyl ether, polyoxyethylene(23) oleyl ether, where numbers 2, 10 and 23 indicate the average number of ethylene glycol units attached to the higher fatty alcohol; higher fatty acid ethoxylates such as polyoxyethylene(2) laurate, polyoxyethylene(10) stearate, polyoxyethylene(23) oleate, where the numbers 2, 10 and 23 indicate the average number of ethylene glycol units attached to the higher fatty acid; sorbitan esters such as polyoxyethylene sorbitan monolaurate (Tween 20), polyoxyethylene sorbitan monopahnitate (Tween 40), polyoxyethylene sorbitan monostearate (Tween 60), polyoxyethylene sorbitan monooleate (Tween 80), polyoxyethylene sorbitan tristearate (Tween 65), polyoxyethylene sorbitan sesquioleate (Tween 85) ; castor oil or its ethoxylates; hydrogenated castor oil or its ethoxylates; or mixtures of these substances. 8. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is a diluent selected from the group consisting of purified water, ethanol, glycerol, 1,2-propylene glycol , polyethylene glycol 400, polyethylene glycol 600, polyethylene glycol 1000, polyethylene glycol 2000, polyethylene glycol 4000, polyethylene glycol 6000, polypropylene glycols, polyglycerols, sorbitol, sucrose, glucose, glucose syrup, fructose, honey, inulin, or mixtures of the aforementioned substances. 9. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is a humectant selected from the group consisting of glycerol, 1,2-propylene glycol, polyethylene glycols, polypropylene glycols, polyglycerols, sorbitol, or mixtures of these substances. 10. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, characterized in that the auxiliary substance is a thickener selected from the group consisting of polyacrylic acid or its salts with sodium hydroxide, potassium hydroxide, or ammonium hydroxide; methylcellulose; sodium carboxymethylcellulose; 2-hydroxyethyl cellulose; 2-hydroxypropyl cellulose; bentonite; starch; modified starches; gelatin; polyglycerols; polyethylene glycols; polypropylene glycols; agar-agar; carrageenans; gum arabic; tragacanth; alginic acid; sodium alginate; pectin; or mixtures of these substances. 11. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, characterized in that the auxiliary substance is a preservative selected from the group consisting of methyl 4-hydroxybenzoate, ethyl 4-hydroxybenzoate, propyl 4 -hydroxybenzoate, butyl 4-hydroxybenzoate, sorbic and/or benzoic acid or their salts with pharmaceutically acceptable bases, or mixtures of the aforementioned substances. 12. Formulation based on resveratrol and extracts of pomegranate, olive leaf and cinnamon as a therapeutic agent of antioxidant action, according to claims 1 and 2, indicated that the auxiliary substance is an antioxidant selected from the group consisting of 2,6-di-tert-butyl-4- hydroxytoluene (BHT), tert-butylhydroxyanisole (BHA), tocopherol, tocopheryl acetate, other tocopherol esters, ascorbic acid, ascorbyl palmitate, lecithin, sodium sulfite, sodium metabisulfite, sodium thiosulfate, sulfur(IV) oxide, cysteine hydrochloride, N-acetylcysteine , propyl gallate, or mixtures of these substances. 13. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic antioxidant for the treatment of oxidative stress conditions. 14. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for the treatment and prevention of cardiovascular diseases: hypertension, hyperlipidemia and atherosclerosis. 15. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for the treatment and prevention of diabetes. 16. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for the treatment and prevention of metabolic syndrome. 17. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for preventive and curative strengthening of the immune system. 18. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for the treatment and prevention of neurodegenerative diseases. 19. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for the prevention of the formation and as an auxiliary agent for the treatment of tumor diseases. 20. Use of the formulation described in claims 1-12, characterized in that it is used as a therapeutic agent for slowing down aging.