RU2681853C1 - Composition for correction of dyslipidemia and maintenance of functional state of human body in metabolic syndrome - Google Patents

Abstract

FIELD: medicine; pharmaceuticals.SUBSTANCE: invention relates to the pharmaceutical industry, in particular to a composition for correcting dyslipidemia and maintaining the functional state of the human body in the metabolic syndrome. Composition for the correction of dyslipidemia and the maintenance of the functional state of the human body in metabolic syndrome, which included: sterilized lyophilized culture fluid obtained by cultivating the VKPM №B-2335 Bacillus subtilis probiotic bacterial strain and containing metabolites of bacilli, zeolite, calcium stearate, resveratrol, coenzyme Q, α-tocopherol acetate, lactose, a complex of enzymes containing alpha-amylase, neutral and alkaline proteinases, beta-glucanase, cellulase, xylanase, and silicon dioxide at a certain ratio of components, the composition is made in solid form in the form of granules, capsules or sachets.EFFECT: above invention allows to expand the range of tools for the correction of dyslipidemia and maintain the functional state of the human body in the metabolic syndrome.1 cl, 26 tbl, 2 dwg, 6 ex

Description

The invention relates to medicine, relates to a composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome (MS) to improve the quality and longevity.

Metabolic syndrome combines a group of diseases, conditions or disorders manifested by certain metabolic, hormonal and clinical disorders: weight gain due to visceral fat in abdominal obesity, decreased peripheral tissue sensitivity to insulin, hyperinsulinemia, impaired carbohydrate, lipid, purine metabolism, high blood pressure (AD), elevated fasting blood glucose, high triglycerides (TG) in blood and low cholesterol a (cholesterol) high density lipoprotein (HDL). Disorders of lipid metabolism include an increase in TG (more than 1.7 mmol / L), a decrease in HDL cholesterol (less than 1.0 mmol / L in men and less than 1.2 mmol / L in women), and an increase in the level of cholesterol of low density lipoproteins (LDL) above 3.0 mmol / L. Moreover, the appearance of one of these factors indicates an increased risk of development of all the others. Patients with MS have an increased risk of developing dyslipidemia, hypertension, insulin resistance, cardiovascular disease (CVD) in the form of coronary heart disease (CHD) and heart failure, type 2 diabetes mellitus (DM), which makes it extremely necessary not to only the identification, but also the timely correction of these conditions.

A meta-analysis of large-scale studies showed that in the adult population, MS is detected in 10-30% of the population, depending on the characteristics of MS and the criteria used in its diagnosis. In Russia, the prevalence of MS varies from 20% to 35%, and in women it occurs 2.5 times more often. With age, the risk of developing MS increases [3].

Timely diagnosis and correction of individual manifestations of MS will prevent the development and progression of severe cardiovascular diseases and their complications. Thus, a decrease in body weight in patients with MS contributes to a decrease in blood pressure, and sometimes even its normalization. Normalization of disorders of carbohydrate metabolism can lead to an improvement in the lipid spectrum and also to a decrease in blood pressure. Hypolipidemic therapy, in turn, can help increase tissue sensitivity to insulin and improve carbohydrate metabolism. Properly selected antihypertensive therapy can be accompanied by an improvement in carbohydrate and lipid metabolism and increase tissue sensitivity to insulin.

Existing methods for the correction of dyslipidemia are the use of non-drug and drug methods. Non-drug methods include lifestyle correction, correction of eating behavior by reducing the intake of saturated fats, cholesterol and total calorie intake, reducing alcohol consumption and increasing physical activity. Drug treatment is carried out using four main groups of lipid-lowering drugs: GMK-CoA reductase inhibitors (statins), bile acid sequestrants, nicotinic acid (NK) (niacin) and its derivatives, fibrates (fibroic acid derivatives).

The most effective group of cholesterol lowering drugs that have radically changed the approach to the prevention of manifestations of MS and its complications is the group of statins. However, when using drugs from the group of statins, a high profile of their safety is not provided [4], which indicates the limited possibilities of their use. Statins are characterized by such side effects as dyspeptic disorders (nausea, vomiting, flatulence, stool disorders), complications of the musculoskeletal system, manifested in the form of signs of muscle weakness and pain (myopathy), which can transform into rhabdomyolysis (destruction of muscle cells tissue) if you do not cancel the drug. Uncontrolled use of statins negatively affects the liver, which is manifested by an increase in the level of hepatic enzymes - alanine aminotransferase (ALT), aspartate aminotransferase (ACT), and others. A negative effect of statins on the kidneys was discovered relatively recently. Even with a healthy urogenital system, the use of statin drugs and especially long courses can lead to severe kidney damage (tubulopathy, renal failure).

Also, statin drugs are characterized by a decrease in the effectiveness of treatment with prolonged use (the phenomenon of receptor evasion). Some patients may experience cognitive impairment in the form of reduced levels of thinking, memory impairment, reminiscent of senile dementia. What is especially important for people with MS, drugs from the group of statins can provoke the development of hyperglycemia, characterized by an increase in blood glucose and the development of insulin sensitivity to body tissues. As you can see, by correcting one of the manifestations of MS, drugs from the group of statins provoke or aggravate other pathological processes underlying it.

To date, there are prerequisites for the search for alternative methods for correcting the noted violations. A new approach to the treatment of conditions associated with MS and intestinal dysbiosis has been developed, which consists in the use of such agents as part of complex therapy that would, along with improving the clinical course and condition of intestinal microbiocenosis, achieve a positive change in the spectrum of blood lipoproteins and restore the antioxidant defense system.

As is known, in the regulation of lipid metabolism, maintaining the qualitative and quantitative composition of the intestinal microflora is essential [5, 6]. Violation of intestinal microflora occurs in 90% of patients with cardiovascular (IHD) and metabolic (obesity, dyslipoproteinemia) diseases [7]. At the same time, dysbiotic changes in the intestine, as they progress, worsen the clinical course of the underlying disease, contributing to an increase in the atherogenic potential of blood serum. Under these conditions, the severity of clinical symptoms is increasing, the duration of their clinical manifestation is lengthening, the results of treatment and the quality of life of patients are deteriorating.

These circumstances determine the need for improving approaches to the correction of lipid metabolism disorders in individuals with MS. One of them is currently considered the use of agents that contribute to the restoration of normal intestinal microflora. The prospect of using means of correction of intestinal microbiocenosis and, above all, to restore its normal microflora is theoretically justified, experimentally confirmed and has clinical experience in the treatment of, for example, coronary heart disease [8].

The range of immunobiological preparations used for the formation, preservation and correction of human microbiota is quite wide and includes [9]:

- probiotics, including autoprobiotics;

- prebiotics;

- synbiotics and metabiotics;

- metabolites of symbiotic microorganisms;

- structural components of cells of symbiotic bacteria;

- microecological engineering.

In clinical practice, drugs from the group of probiotics are of great interest, including certain types of microorganisms non-pathogenic for humans and substances of microbial origin, which, with the natural method of administration, have beneficial effects on physiological functions, biochemical and behavioral reactions of the body through optimization and stabilization of its microecological status [10]. These include, first of all, bifidobacteria and lactobacilli to regulate the balance of intestinal microflora, since bifidobacteria and lactobacilli play a crucial role in ensuring the normal functioning of the intestine. When dysbiotic conditions occur, the qualitative and quantitative composition of anaerobic bifidobacteria and lactobacilli undergoes disturbances in the first place. Therefore, drugs based on them are widely used in the practice of doctors of various specialties.

One of the promising directions for the prevention of intestinal microbiota disorders is the use of drugs whose active principle is non-living microorganisms (for example, Bifidobacterin, Lactobacterin, Sporobacterin, Laminolact, Linex, Bifiform, etc.), and metabolites of probiotic strains of microorganisms [11, 12].

Currently, the pharmaceutical market is quite well-known among such agents and is used in clinical practice to maintain optimal microecological parameters in the intestine of the probiotic complex Bactistatin [13], which includes (wt.%): Sterilized culture fluid (SCF) obtained during the cultivation of bacteria of the species Bacillus subtilis and containing metabolites of bacilli (1.0), soy flour hydrolyzate (GSM) (20.0), the natural mineral zeolite (78.0), calcium stearate (SC) or aerosil (1.0). The specified probiotic complex in composition, mechanism of action on the body and the achieved beneficial effect, expressed in the correction of intestinal microbiota disorders, is closest to the claimed composition and adopted as a prototype composition.

The basis of the prototype composition is constituted by biologically active substances (BAS) immobilized on zeolite, synthesized by Bacillus subtilis bacteria during deep growth, which determine its beneficial effect. Their list is quite extensive and includes substances that are heterogeneous in chemical composition and biological properties: proteolytic and amylolytic enzymes and coenzymes, amino acids, polypeptides, prebiotic components, polysaccharides (hexosamine, glucosamine), B vitamins (pyridoxine hydrochloride, riboflavin), nitrogen bases and their derivatives (adenine, guanine, thymine, uracil, cytosine) [14-16].

The listed biologically active substances synthesized by Bacillus subtilis bacteria have a positive effect on the macroorganism as a whole, contributing to the improvement of microecological conditions in the intestine, affecting metabolic processes and exerting an immunomodulating effect. Among them, natural antibacterial substances (bacteriocins, lysozyme, etc.), which selectively inhibit the growth and reproduction of pathogenic and conditionally pathogenic microorganisms in the intestine, without affecting the symbiotic microflora, are of particular importance.

Zeolite, which is part of the composition of the prototype, acts as an adsorbent of the metabolites of bacilli Bacillus subtilis and their carrier. Its main purpose is to immobilize the metabolites synthesized by the bacilli during cultivation and concentrated in the culture fluid, and deliver them as a carrier to the intestine. In this regard, the zeolite in the composition of the prototype is contained in the optimal amount sufficient to ensure the sorption of all metabolites. A significant decrease in the amount of zeolite will lead to the loss of some metabolites and reduce the effectiveness of the composition of the prototype. An increase in the amount of zeolite (over 85%) is also unacceptable, as it will lead to a decrease in the effectiveness of the prototype composition due to dilution with a low-active ingredient.

An important component of the prototype composition is fuels and lubricants, which is part of the protective environment of metabolites and ensures the strength of their sorption on the surface of the zeolite, as well as a source of amino acids and provides the nutritional needs of normal intestinal microflora and macroorganism cells.

SC or Aerosil are included in the composition of the prototype as auxiliary technological additives.

The main purpose of the prototype composition is to restore and maintain optimal microecological parameters in the intestine, treat diseases of the gastrointestinal tract (GIT) by introducing substances into the body that inhibit the development of pathogenic microflora. Its action is based on the fact that during transit through the gastrointestinal tract in a given area, the protective capsule is destroyed and the biologically active substances immobilized on zeolite particles are released into the intestinal cavity. At the same time, micellar structures are formed around zeolite particles, which are gradually released from the porous surface of the zeolite during movement along the gastrointestinal tract. On the one hand, this allows maintaining the activity of biologically active components of the probiotic in the gastrointestinal tract during the day, which is sufficient to restore and stimulate the functional activity of normal intestinal microflora. On the other hand, the effect of the gradual release of biologically active substances immobilized on it from the surface of the zeolite leads to the appearance of open surfaces of its porous structure, which ensures the inclusion of ion exchange mechanisms and selective sorption of toxic compounds and the cleansing of the gastrointestinal tract from toxins and decomposition products. This is especially important for general detoxification of the body.

Thus, due to the complex composition, the prototype composition contributes to the general normalization of microecological conditions in the intestine through various mechanisms, such as suppression of opportunistic flora, sorption and elimination of toxins, improvement of digestion processes, and improvement of the trophic base for normal microflora and gastrointestinal epithelium.

The expediency of prescribing the prototype composition when implementing fundamentally new treatment regimens for coronary heart disease, a disease that currently has extremely high clinical and social significance due to the high incidence rate among people of working age, as well as the severity of complications that have arisen, has also been established [17]. Along with the main reasons for the development of this pathology, there are a number of additional factors that indirectly contribute to its occurrence and development. First of all, they include intestinal dysbiosis, since an opinion has been formed about the relationship of IHD with quantitative and qualitative violations of the composition of the microbiota, as well as the key role of the intestinal microbiota with the participation of enterohepatic circulation mechanisms in the regulation of lipid metabolism. In this case, the positive effect of the prototype composition is associated with the correction of intestinal microbiocenosis.

However, along with the indicated beneficial effects for the composition of the prototype is characterized by a number of disadvantages:

- limited biological activity in relation to the effect on lipid metabolism, which is associated with the predominant effect of the drug at the level of the gastrointestinal tract;

- the inability to concentrate the culture fluid obtained on soybean medium using fuels and lubricants, due to its high viscosity;

- the use in the technological process of obtaining the composition of the prototype as a nutrient medium of native soy flour does not allow the full use of the nutritional properties of the feedstock. Additional chemical or enzymatic hydrolysis of soy flour is required, which significantly complicates the technology for obtaining the prototype composition;

- use for the nutritional needs of the normal intestinal microflora and cells of a macroorganism of an inaccessible and at the same time defective amino acid composition of an expensive food product - fuels and lubricants containing only two essential amino acids (lysine and methionine), and in very small quantities (0.2% and 0.096%, respectively).

The noted disadvantages significantly limit the possibility of widespread use of the prototype composition for the effective correction of dyslipidemia and maintaining the functional state of the human body in MS.

At the same time, MS represents a multifactorial pathological state of the organism, and the correction of its manifestations, in particular dyslipidemia, requires the development and practical use of multicomponent formulations, which, along with metabolites of the probiotic strain of microorganisms, would include safe active compounds that can have a complex effect on other non less important aspects of MS.

Unfortunately, official medicine, judging by the materials of the last conference on MS (November 2014), uses an extremely limited range of natural components to combat this multifactorial pathological condition of the body. Evidence of the effectiveness of dyslipidemia correction drugs consistent with evidence-based medicine is extremely scarce. Therefore, the search for formulations, including active natural compounds, their comprehensive study and development of recommendations for use in clinical practice are very relevant.

In this regard, the aim of the invention was to expand the range of means for correcting the manifestations of MS by creating a balanced complex that provides an effective comprehensive effect on the body, which consists in correcting the state of intestinal microbiocenosis, lipid metabolism, regulating free radical oxidation processes and maintaining the functional state of the body, indicators which is reduced during the development of MS, which does not contain substances hazardous to health, non-toxic, and is not capable of Tew to cumulation sensitizing action and long-term negative consequences of the shelf stable and well-tolerated.

Achieving this goal is possible by creating a balanced multicomponent complex, the composition of which is represented by metabolites of a probiotic strain of bacteria, safe active natural compounds, a mitochondrial metabolite, and corresponds to the main pathogenetic mechanisms of the formation of MS and its complications, which allows to improve the functional state of the gastrointestinal tract, to control the level of lipids in the blood and their normalization due to the complex effect on the mechanisms of development of lipid disorders of sharing, improve the antioxidant defense system and maintain the functional state of the organism.

In the formation of the qualitative and quantitative composition of the claimed composition, intended for the complex effect on the correction mechanisms of such manifestations of MS as dyslipidemia, it was assumed that its main components should:

- have an effective beneficial effect on the normal intestinal microflora, in particular, on lactobacilli, contributing to their growth and reproduction, and thereby improve the functional state of the gastrointestinal tract;

- normalize lipid metabolism (significantly increase the level of anti-atherogenic HDL);

- improve the condition of the antioxidant defense system;

- maintain the functional state of the body.

To implement the specified complex effects in the multicomponent composition of the claimed composition includes:

- metabolites of the probiotic strain of bacteria Bacillus subtilis VKPM No. B-2335 contained in a sterilized lyophilized culture fluid;

- lactose;

- zeolite;

- trans-resveratrol;

- α-tocopherol acetate;

- coenzyme Q ₁₀ ;

- a complex of enzymes (alpha-amylase, neutral and alkaline proteinases, beta-glucanase, cellulase, xylanase);

- auxiliary technological additive (calcium stearate and / or silicon dioxide).

In the claimed composition, as well as in the prototype composition, metabolites of the probiotic bacterial strain Bacillus subtilis VKPM are involved in providing a complex effect on the state of intestinal microbiocenosis due to the beneficial effect on the indigenous component of the microflora of the biotope and the provision of a specific antagonistic effect with respect to a wide range of pathogens No. B-2335.

The composition of the claimed composition metabolites of the probiotic strain of bacteria Bacillus subtilis VKPM No. B-2335 introduced in the lyophilized SCZ. It is in the SCZ that a unique set of metabolites is synthesized that is synthesized by bacilli during their cultivation. Of the number of metabolites, natural antibacterial substances (bacteriocins, lysozyme, etc.) are valuable, which not only selectively inhibit the growth and reproduction of pathogenic and conditionally pathogenic microorganisms in the intestine, without affecting the symbiotic microflora, but also make a significant contribution to the ability of the claimed compositions to influence factors of nonspecific resistance of the body and stimulate the functional activity of immunocompetent cells.

It is especially significant that in the composition of the claimed composition, the metabolites of Bacillus subtilis VKPM No. B-2335 represent a lactogenic factor and exhibit a lactogenic effect, which is expressed in stimulating the growth and reproduction of lactobacilli of the indigenous intestinal microflora, as a result of which its microflora are normalized.

To ensure the nutritional needs of representatives of indigenous intestinal microflora, as well as macroorganism cells, lactose is introduced into the composition of the claimed composition as a nutrient medium.

The zeolite, which is part of the claimed composition, as in the prototype composition, is intended primarily for the immobilization of metabolites produced by Bacillus subtilis during their cultivation and concentrated in the SCR. The feasibility of using zeolite as a component of the claimed composition is determined by the unique chemical structure of its molecules, as a result of which the zeolite has a high sorbing activity with respect to the metabolites of Bacillus subtilis bacteria. Optimum conditions arise for the zeolite to fulfill its main functions - the adsorbent and the carrier of metabolites Bacillus subtilis. Without being absorbed into the gastrointestinal tract and passing in transit, the zeolite transports the metabolites of Bacillus subtilis bacteria immobilized on it throughout the intestine. The release of metabolites occurs gradually and allows for a sufficiently long period of time (up to 1 day) to maintain the necessary level of effectiveness of the claimed composition.

Due to the unique chemical structure of the molecules, the zeolite has a pronounced sorption effect with respect to compounds with a low molecular weight, such as methane, hydrogen sulfide, ammonia and other toxic substances, without directly interacting with vitamins, amino acids, proteins, due to which these substances remain in the digestive tract .

In addition, the zeolite has ion-exchange properties, since the main skeleton of its crystal lattice consists primarily of a tetrahedron and has cavities in which ions (sodium, potassium, calcium) are located, which easily exchange with each other and with the surrounding substrate. Zeolite, passing through the entire length of the gastrointestinal tract, participates in selective ion exchange and serves as an additional source of a number of trace elements necessary for the body [18]. The particles of the zeolite used are no more than 500 microns in size and have an oval crystal shape, which excludes the formation of microtraumas in the intestine and is completely safe [19-21].

As an active component in the composition of the claimed composition introduced resveratrol - a natural biologically active compound from the group of polyphenols, including flavonoids, flavonols, catechins and stilbenes, a natural component of black and red grapes and other fruits, cocoa and nuts, red wine, Japanese plant root Highlander (Polygonum cuspidatum). Moreover, using its active isomeric form - trans-resveratrol (3,5,4'-trihydroxy-trans-stilbene).

Trans-resveratrol has a multifactorial mechanism of action on human organs and tissues. In this case, the fundamental properties of trans-resveratrol, determining its positive effect on the human body, are antioxidant activity as polyphenol and the induction of mitochondriogenesis. Due to the antioxidant properties, trans-resveratrol has not only a protective effect on various membrane structures, but also a regulatory effect on proliferative, differentiating processes, apoptosis, etc. A specific function of trans-resveratrol is the induction of the formation of new mitochondria through activation of sirtuins, including SIRT1. An important aspect of its action is also a pronounced anti-inflammatory effect, manifested in the suppression of the expression of inflammatory markers TNF-α, IL-6, IL-1β, inducible NO synthase (i-NOS) of cyclooxygenase COX-2 and an increase in the level of anti-inflammatory cytokine IL-10. as a result, trans-resveratrol manifests itself as an immunomodulating factor.

Trans-resveratrol is used as part of the claimed composition, first of all, to reduce tissue sensitivity to insulin, improve glucose metabolism, normalize lipid and blood sugar levels as a result of providing a preventive metabolic effect, which consists in modulating (activating) effect on the enzyme activated by adenosine-monophosphate - Cellular 5'AMP-activated protein kinase (AMPK).

AMPK protein kinase determines the energy balance in the cell, and therefore plays a key role in metabolic processes. It is contained in each cell and serves as the main regulator of the body, determining the amount of subcutaneous fat due to the ability to block the synthesis of fatty acids and accelerate their oxidation. As a result of the activation of AMPK protein kinase in the cells of the body, metabolic processes are launched and the cells lose in volume. In fact, the activation of AMPK protein kinase mimics the effects of calorie restriction in the body (effects of a low-calorie diet without changing the diet). In this way, trans-resveratrol counteracts the negative effects of eating high-fat foods.

In addition, the anti-atherogenic effect of trans-resveratrol is realized by inhibiting the oxidation of LDL, reducing platelet aggregation, vasodilating properties, anti-inflammatory effects against vascular endothelium.

The lower permissible level of trans-resveratrol intake for the prevention of cardiovascular diseases and improve insulin sensitivity for healthy people is 5-10 mg per day.

Fat-soluble vitamin E, the main effect of which is the antioxidant effect, is used in the composition of the claimed composition in the form of active α-tocopherol acetate. The specified natural compound inhibits and limits free radical reactions. The ability of α-tocopherol acetate to stimulate the synthesis of heme and heme-containing enzymes: hemoglobin, myoglobin, cytochromes (including cytochrome P-450), catalase, peroxidase is extremely important. The consequence of this is the improvement of tissue respiration, synthetic processes in them, the elimination of peroxides of unsaturated fatty acids and lipids that damage cell and subcellular membranes. As a result of the action of α-tocopherol acetate, cell and subcellular membranes (erythrocytes, cells of various tissues, lysosomes) are stabilized, their permeability decreases and integrity is preserved, protein synthesis of various tissues (collagen in subcutaneous tissue and bones, contractile proteins in skeletal, smooth muscles and myocardium, proteins of the mucous membranes, etc.), as well as the formation of a number of enzymes (creatine phosphokinase, vasopressinase), hormones (gonadotropins), etc.

α-tocopherol acetate acts as a universal protector of cell membranes from oxidative damage. It occupies a position in the membrane that prevents the contact of oxygen with unsaturated lipids of the membranes (the formation of hydrophobic complexes). This protects the biomembranes from their peroxide degradation. The antioxidant properties of α-tocopherol acetate are also due to the ability of the mobile hydroxyl of the chroman nucleus of its molecule to interact directly with free oxygen radicals (O ₂ , HO, HO ₂ ), free radicals of unsaturated fatty acids (RO, RO ₂ ) and fatty acid peroxides. The membrane-stabilizing effect of α-tocopherol acetate is manifested and in its ability to protect from the oxidation of the SH-group of membrane proteins.

α-tocopherol acetate is not only an antioxidant, but also an antihypoxant, due to its ability to stabilize the mitochondrial membrane and reduce oxygen consumption by cells. It should be noted that of all cellular organelles, mitochondria are most sensitive to damage, since they contain the most easily oxidized unsaturated lipids. Due to the membrane-stabilizing effect of α-tocopherol acetate in mitochondria, the conjugation of oxidative phosphorylation, synthesis of adenosine triphosphate (ATP) and coenzyme Q ₁₀ (KoQ ₁₀ ), a component of the respiratory chain and the main mitochondrial antioxidant, increases [22].

To maintain the vital functions of the human body, including the additional activation of energy metabolism at the cellular level and the burning of fats and carbohydrates in cells, one of the indispensable participants in the most important processes in the body is used - the mitochondrial metabolite coenzyme Q ₁₀ . The indicated active component of the claimed composition is localized in mitochondria (cell structures that are responsible for the conversion of energy into ATP molecules) and is a direct participant in the respiratory chain of electron transfer. KoQ ₁₀ provides an energy-efficient aerobic pathway for glucose utilization, since the mitochondrial electron transport chains are a biochemical mechanism for the formation of ATP along this pathway, and KoQ ₁₀ plays an important role in their functioning.

With its broad antioxidant properties, KoQ ₁₀ prevents the deposition of cholesterol on the walls of blood vessels. KoQ _{10 also} has the ability to reduce swelling of the extremities and eliminate cyanosis, so it is used in congestive forms of chronic heart failure. According to many clinical studies, KoQ ₁₀ has the ability to normalize blood sugar and reduce high blood pressure, so it is prescribed for diabetes [23].

The combined use of KoQ ₁₀ and trans-resveratrol as an inducer of mitochondriogenesis allows one to ensure mutual enhancement of the effects of these compounds, since they have a common target of action - mitochondria, but realize their effects due to different mechanisms.

The use of proteolytic and amylolytic enzymes in the composition of the claimed composition, which are introduced as a complex of enzymes (alpha-amylase, neutral and alkaline proteinases, beta-glucanase, cellulase, xylanase), promotes complete digestion of food and assimilation of nutrients, as well as the formation of an optimal endoecological environment in intestines. The latter is important for maintaining normal intestinal microflora in optimal condition.

In the composition of the claimed composition using a complex of enzymes obtained biotechnologically. Unlike the most widely used enzymes of animal origin, the effect of the complex used is not aimed at replacing the functions of the human digestive enzymes, but at expanding the spectrum of their activity. This is achieved by splitting chemical bonds in the substrates entering the gastrointestinal tract and not accessible to their own digestive enzymes using the indicated enzymes.

To improve the manufacturability of the process of obtaining the claimed composition in solid form (granules, capsules or sachets), a sliding auxiliary technological additive is introduced into its composition, which is used as calcium stearate and / or silicon dioxide in an amount determined by pharmaceutical and therapeutic expediency. The use of a sliding auxiliary technological additive provides the creation on the surface of the granules obtained by the production of the claimed composition, a thin film that gives them a sliding effect, which increases the flowability of the encapsulated mass and facilitates its encapsulation or filling sachets.

The action of the claimed composition, made in capsule form, begins already in the stomach, where the dissolution of the protective capsule occurs under the influence of gastric juice. The metabolites of the probiotic strain of bacteria included in the composition of the claimed composition, the degradation of which is virtually eliminated during transit through the gastrointestinal tract, as well as the active natural compounds enter the intestine, where their beneficial effect is manifested.

Thus, the claimed composition is a balanced complex, has a complex multicomponent composition, obtained by qualitative and quantitative selection of ingredients in the following ratio, wt. %:

sterilized lyophilized culture fluid, containing metabolites of the probiotic strain of bacteria Bacillus subtilis VKPM No. B-2335 - 1.4-1.9; trans resveratrol - 3.5-4.8;

coenzyme Q ₁₀ - 7.1-9.6; α-tocopherol acetate - 0.4-0.5; lactose - 13.0-17.5; enzyme complex (alpha-amylase, neutral and alkaline proteinases, beta-glucanase, cellulase, xylanase) - 0.2-0.3; zeolite - 64.2-73.5; calcium stearate and / or silicon dioxide - 0.9-1.2.

The ability to achieve the objectives of the invention is confirmed by the results of the studies presented in the following examples.

Example 1. Obtaining the inventive composition in solid form for oral administration.

The inventive composition can be made in solid form (granules, capsules, sachets), which greatly simplifies its use when taken orally, conveniently in use, storage and transportation.

The claimed composition is first obtained in the form of granules, for which the following algorithm of actions is performed. The microorganisms Bacillus subtilis VKPM No. B-2335 are grown by the method of deep cultivation in a biological reactor (fermenter). At the end of the cultivation process, QOL with microorganisms is centrifuged to separate and subsequently remove living cells of microorganisms, and then sterilized. Before sterilization, QOL is mixed with zeolite, previously crushed to particles no larger than 500 microns in size. The resulting mixture is subjected to lyophilization, in which the biologically active metabolites of the producer are immobilized on zeolite particles. Then, lactose, coenzyme Q ₁₀ , trans-resveratrol, α-tocopherol acetate, an enzyme complex and an auxiliary technological additive are added sequentially in the required amounts to the mass, placed in the tank of the granulation unit for mixing and drying. After unloading, the mass is sieved on a vibrating screen and transferred to the encapsulation or filling stage of the sachet.

A natural zeolite — a natural microporous silicate mineral of the Kholinsky deposit (Buryatia) (KRAFT LLC, St. Petersburg) is used as a carrier sorbent [24, 25].

As you can see, the process of obtaining the inventive composition in solid form for oral administration (granules, capsules, sachets) is quite simple, and the ingredients used for this are produced under industrial conditions, and inexpensive are available.

Example 2. Safety assessment of the claimed composition.

The safety of the claimed composition was evaluated by sanitary-chemical analysis, microbiological studies, as well as the determination of toxicological

characteristics, including acute toxicity with intragastric administration, chronic toxicity and the ability to cumulate, irritating and sensitizing effects, possible long-term effects. Sanitary chemical analysis.

Sample preparation was carried out according to GOST 26929-94 “Raw materials and food products. Sample preparation. Mineralization to identify toxic elements. Interstate standard. " Lead and cadmium were determined according to GOST 30178-96 “Raw materials and food products. Atomic absorption method for the determination of toxic elements. Interstate standard. " Arsenic was determined according to GOST 26930-86 “Raw materials and food products. Method for the determination of arsenic. " Mercury was determined according to the "Guidelines for the detection and determination of total mercury in food products by flameless atomic absorption" (No. 5178-90).

The content of organochlorine pesticides (toxicants) was determined according to the "Methodological guidelines for the determination of residues of organochlorine pesticides" (No. 1766-77) and official methods of analysis of AOAC ("Official methods of analysis of the AOAC", 1984, 14th ed., Chapter 29, P . 537-538).

The studies were carried out using laboratory scales VLR-500, gas-liquid chromatograph "Carlo Erba" (model HRGC 5700), exchange cells of the Italian company "Texnoplast".

The measurement results are presented in table 1 and indicate that in the claimed composition the content of toxicants (organochlorine pesticides HCTs and DCT) and hazardous substances (lead, cadmium, mercury, arsenic) does not exceed the allowable levels established by the Technical Regulation of the Customs Union TP ТС 021 / 2011 “On Food Safety” (Appendix 3, Section 10) [26]. The same pesticides as aldrin and heptachlor in the claimed composition were not found.

Microbiological studies.

Microbiological studies were carried out in accordance with the requirements of the guidelines MUK 4.2.577-96 "Methods of microbiological control of baby food, therapeutic nutrition and their components" and MUK 2.3.2.721-98 "Determining the safety and effectiveness of biologically active food additives", GOST R 52814 -2007 “Food products. Method for the detection of bacteria of the genus Salmonella)), GOST R 52816-2007 “Food products. A method for identifying and determining the number of bacteria of the group of Escherichia coli (coliform bacteria) ", GOST 10444.12-88" Food. Method for determination of yeast and mold fungi ”, GOST 30726-2001“ Food products. Methods for the identification and quantification of bacteria of the species Escherichia coli. "

It has been experimentally established that, according to microbiological safety indicators, the claimed composition meets the requirements established by the Technical Regulation of the Customs Union TP TS 021/2011 "On Food Safety" (Annexes 1 and 2, Section 1.9) (table 2)].

Acute toxicity study.

To determine indicators of acute toxicity, the claimed composition was administered to healthy outbred white rats of both sexes weighing 130-150 g and 180-200 g intragastrically in the form of aqueous starch suspensions. The introduction was carried out through a metal atraumatic probe in increasing doses using Litchfield-Wilcoxon probital analysis in modification 3. Rota [27].

During the experiment, to study the effects of the claimed composition, experimental groups of 6 animals of the same sex in each were formed. In addition, there were similar in number groups of control animals of each sex, to which equivalent volumes of solvent were introduced along the same route — 1% starch suspension. Animals were randomly assigned to groups using randomization. The criteria for the acceptability of randomization were considered to be the absence of external signs of diseases and the homogeneity of groups by body weight (± 10%).

Experimental animals were monitored for 14 days with registration of indicators: mortality, time of death of animals, symptoms of poisoning, results of daily monitoring of general condition and behavior, weighing, results of monitoring feed and water consumption, results of autopsy and macroscopic description of dead and surviving animals at the end studies (euthanasia was carried out by an overdose of ether), the results of determining the mass coefficients (MK) of internal organs.

The introduction of the claimed composition in the maximum tolerated doses (20 g / kg for rats and 25 g / kg for mice) did not cause the death of experimental animals. Not noted any other signs of negative effects of the claimed composition. On the first day, lethargy, lethargy, decreased feed and water intake, and diarrhea were noted. On the remaining days, the general condition and behavior of the animals of the experimental groups did not differ from the animals of the control group. In particular, no dyspeptic symptoms were noted. At autopsy - without features. No gender differences were observed during intoxication.

At autopsy one day after acute administration, it was revealed that pleura and chest organs were not changed. Light pale pink, airy, without seals to the touch. The size of the heart is within normal limits. In the cavities of the heart contains a small amount of liquid blood. The heart muscle is dense, brown in color. The stomach is filled with a small amount of solid food. The mucous membrane is shiny, folded, slightly pinkish in color.

The mucosa of the small intestine is shiny, smooth, pinkish in color. The size and shape of the liver do not differ from the control. The surface of the liver is smooth. The capsule is thin, transparent. The liver pattern in the section is not changed. Liver tissue is moderately full-blooded. The kidneys are of normal size and shape, brownish in color, dense, with a distinct cortical and medullary substance in the section. The thyroid gland, adrenal glands and pancreas do not differ in appearance from the control.

The results of studying the dynamics of changes in body weight, water and feed consumption of experimental animals, as well as the dynamics of changes in body weight of surviving white mice receiving the inventive composition within 14 days of observation, they practically did not differ (tables 3-6).

Analysis of the MK values of the organs of surviving white rats and mice revealed that the acute administration of the claimed composition in the maximum tolerated dose did not cause any significant differences compared with the control group of animals receiving 1% starch suspension (tables 7 and 8).

The condition of the animals that survived acute intoxication indicates good tolerance and harmlessness of the claimed composition when it is administered in the maximum tolerated doses.

Thus, the results of toxicometry, necropsy data and observation of experimental animals within 14 days after acute administration allow us to classify the claimed composition as non-toxic (relatively harmless according to S.D. Zagolnikov) and low-hazard (hazard class IV according to GOST 12.1.007) substances .

The study of the ability to cumulate.

Studies of the ability to cumulate were carried out on white non-linear male rats weighing 180-200 g with the introduction of the inventive composition intragastrically through an atraumatic metal probe for 30 days at a daily dose of 1.5 g / kg. Animals of the control group received 1% starch suspension in the same volume.

Monitoring of basic physiological parameters was carried out after 30 days of chronic daily administration of the claimed composition.

No lethal effects were observed throughout the observation period. The general condition and behavior of the experimental animals remained normal and did not differ from the parameters of the control group throughout the experiment.

The autopsy of rats euthanized the day after the last injection of the claimed composition showed that the size, shape and color of their internal organs did not have macroscopic changes compared with the control group. The mucous membranes of the stomach and small intestine were shiny, pale pink, with no signs of irritation or inflammation.

During histological examination of preparations of the lungs, myocardium, liver, kidneys and gastric mucosa of experimental animals receiving the claimed composition, dystrophic, inflammatory or necrobiotic changes in organs were not detected.

Therefore, chronic intragastric administration of the claimed composition into experimental animals did not cause dystrophic or destructive changes in parenchymal organs and was not accompanied by irritation of the mucous membranes of the gastrointestinal tract (GIT). Thus, according to the integral indicator of cumulation, the claimed composition does not have the ability to cumulate.

Sensitizing study.

The study of the sensitizing effect of the claimed composition was carried out on guinea pigs. To do this, 0.02 ml of aqueous dilution of the claimed composition was administered once in a dose of 10 mg / kg into the skin of the outer surface of the animal’s ear with a tuberculin syringe, and after 10 days, the claimed composition was applied to the pre-trimmed sections of the skin of the lateral back surface of 2 × 2 cm and fixed powder form at the rate of 20 mg / cm ² . Blood sampling in experimental animals was carried out 3 hours after the staging of skin samples.

The results of evaluating the effect of the claimed composition on the sensitizing effect are presented in table 9 and indicate the absence of signs of sensitization in the peripheral blood - eosinophilia or an increase in the level of lysis in the reaction of specific leukocyte lysis (RSLL) compared with the control. Examination of the back skin of experimental animals showed no signs of irritation. Therefore, the claimed composition upon epicutaneous contact does not have a sensitizing effect, that is, it does not provoke the development of allergies.

A study of the development of possible long-term effects.

The study of a possible gonadotoxic effect was carried out in accordance with the guidelines for indicators required in the initial assessment of chemicals [28, 29]. Mutagenic activity was studied by the frequency of micronuclei formation in polychromatophilic bone marrow erythrocytes (from the femur) of white mice in accordance with the methodological recommendations [30, 31].

The results of a study of the possible gonadotoxic and mutagenic effects of the claimed composition when administered for 10 days at a dose of 1 g / kg, carried out 15 days after the end of its administration, indicate the absence of significantly significant changes in gonadotoxicity and mutagenicity indicators (table 10). Inhibition of spermatogenesis or an increase in the frequency of mutations was also not observed.

Therefore, the claimed composition does not have a toxic effect on reproductive function and does not have a mutagenic effect. This indicates its safety in terms of the development of long-term negative consequences in humans.

Example 3. Evaluation of the stability of the claimed composition during storage.

We investigated the possibility of maintaining the activity and safety of the claimed composition, made in solid form in the form of granules, and established a guaranteed shelf life in case of storage at different temperatures and without the use of special equipment.

During the experiment, the granules were placed in 120 consumer packages in the form of double plastic bags and stored for 3.0 years at temperatures of 5 ± 2 ° С, 10 ± 3 ° С, 25 ± 5 ° С, 35 ± 6 ° С (according to 25 packs for each temperature storage mode). Every 6 months of storage, 5 packages were selected from the corresponding temperature regime and the characteristics of the granules were determined: organoleptic characteristics (appearance, color, smell), physicochemical parameters (content of coenzyme Q ₁₀ and trans-resveratrol, amylolytic and proteolytic activity, mass fraction moisture, disintegration, average weight of granules in a capsule), microbiological purity. To assess possible changes in these characteristics during storage, their values were preliminarily determined at the beginning of the study.

The data presented in table 11 indicate that at normal storage temperature (25 ± 5 ° C) organoleptic and physico-chemical parameters, as well as microbiological purity of the claimed composition, made in solid form in the form of granules, practically do not change during 3.0 years.

Thus, the guaranteed shelf life of the claimed composition is at least 3.0 years, subject to the safety of packaging. Special temperature storage conditions or the use of special equipment is not required.

Example 4. Assessment of the biological activity of the claimed composition.

The biological activity of the claimed composition was evaluated on the basis of studying its ability to normalize fat metabolism and, due to this, cause body weight correction in experimental animals, as well as maintain the functional state of the body and, above all, adaptive capabilities, reduced, as a rule, in the presence of excess body weight and obesity .

The experiments to assess these characteristics were performed on white non-linear male rats with a standard weight of 150-160 g, received from the Rappolovo RAMS nursery in the Leningrad Region, in which experimental obesity was modeled.

In modeling experimental obesity in rats, a diet saturated with fats was used as a model for inducing hyperlipidemia and obesity [32]. For this, experimental animals consumed a homogeneous feed containing (%):

-casein - 5; - sunflower oil mixed with pork fat - 40; - glucose - 43; - agar - 2; - salts, vitamins - four; - cholic acid - one; - thiouracil - one; - cholesterol - four.

In addition, the rats were additionally intragastrically injected with a 10% oil emulsion of pork fat at a dose of 100 mg / kg.

The inventive composition was administered at a dose of 150 mg / kg (approximate "therapeutic and prophylactic" dose for rats, taking into account the interspecific dose transfer ratio of 5.9 for humans with an average weight of 70 kg, based on 4 capsules per day).

The claimed composition was administered intragastrically through an atraumatic probe one hour before the administration of pork fat for 1.5 months against the background of the above diet.

The intact group of rats received the usual vivar diet.

All animals received ad libitum water.

All tests and subsequent blood sampling was carried out at the end of the study 18 hours after the last injection of the claimed composition. During this period, experimental animals were deprived of food.

Statistical processing of the experimental results was performed according to Student-Fisher.

The study of the ability to correct body weight of experimental animals due to the normalization of fat metabolism.

During the experiment, we studied the effect of the claimed composition on indicators of fat metabolism and body weight. All laboratory tests were performed according to standard methods [33, 34]. Serum β-lipoproteins (β-LPs) were determined by the turbodimetric method, free (non-esterified) fatty acids (NEFA) - titrometrically, total bilirubin (OB) - spectrophotometrically, phospholipids (PL) - by the content of total phosphorus.

The dynamics of rat body weight was studied using VLR-500 weights.

Also, at the end of the experiment, experimental animals were individually planted in metabolic cages of the Italian company Tecniplast Gazzada for 24 hours to collect excrement (feces). Then, in the feces, the amount of neutral fat, the presence of fatty acids and soap were determined by conventional methods [35].

The experimental results presented in tables 12 and 13 confirmed the adequacy of the obesity model used: in the control group (without prophylaxis), the content of total lipids, low-density lipoproteins, triglycerides, cholesterol in the blood significantly increased, and the serum phospholipids, on the contrary, decreased, increased atherogenic coefficient. The body weight of the experimental animals increased significantly.

In the experimental group receiving the claimed composition, normalization of lipid metabolism was observed. Animals in the experimental group were more active, without signs of obesity.

Thus, it was experimentally established that the claimed composition exhibits a lipid-lowering effect and effectively reduces the body weight of experimental animals.

This is also evidenced by the normalization of the coprogram for the prophylactic use of the claimed composition.

Effect on the resistance of rats to high temperatures.

The adaptogenic properties of the claimed composition were evaluated by the effect of the temperature factor on experimental animals, which allows a comprehensive assessment of the effect of the claimed composition on metabolic systems, the stability of which in conditions of hyperthermia determines the survival of individuals.

To assess the stability of rats under hyperthermia, the experimental and control groups of 6 animals in a cage were placed in a ventilated thermostat and kept at a temperature of 60 ° C. The device of the heat chamber made it possible to provide adequate gas exchange, sufficient to prevent oxygen deficiency, and made it possible to observe the animals through a glazed inspection hole. During the observation, the development of symptoms of the lesion was recorded and the time of manifestation of the convulsive syndrome and death of individuals was noted.

Experimental data indicate that a four-week administration of the claimed composition contributes to a positive effect on the resistance of rats to hyperthermia (table 14). The rate of development of the convulsive syndrome in rats receiving the claimed composition has a distinct tendency to decrease, and their lifespan in conditions of hyperthermia is significantly higher. Thus, the claimed composition has an adaptogenic effect and, when applied on a course, can increase resistance to extreme factors.

Impact on cognitive function.

Animal experiments were performed in accordance with the “Methodological recommendations for preclinical study of drugs with nootropic type of action” [36].

The basic model for assessing the effect of substances on the formation and reproduction of a commemorative trace in normal conditions and under conditions of its experimental violation is the technique of the conditioned passive avoidance reflex (passive avoidance reaction) [37]. The main advantages of this method are the speed of reflex production (learning from one sample) and the possibility of differentiated effects on different phases of memory. The method is based on the inhibition of congenital activity or acquired skills, leading to negative consequences for the body. In conditions of passive avoidance, the animal learns to avoid harmful factors by suppressing certain forms of behavior, in this study - to avoid the preferred dark space. There are several variants of the passive avoidance reaction method developed in one combination. The most commonly used form of passive avoidance reaction was used in this study.

The experimental animals were divided into 3 groups: control and two experimental animals with 10 animals each. Before developing passive avoidance reaction, experimental animals of the first experimental group were administered intragastrically the claimed composition at a dose of 100 mg / kg for 10 days. The animals of the second experimental group were also injected intragastrically for 10 days with the comparison drug Noopept ^® 10 mg tablets (CJSC Master-Clone, Moscow) in the form of an aqueous suspension at a dose of 5 mg / kg (the maximum therapeutic dose adapted for rodents in humans was per active component). Animals of the control group were injected with 1% starch suspension in a volume of 0.2 ml.

The installation for generating a passive avoidance reflex is a plywood box - a compartment that consists of an illuminated part with a 40 W incandescent lamp mounted and darkened with 2 mm diameter copper rods on the floor. To the rods through the lather and the closing key, current is supplied from the network. Between themselves, the parts are separated by a partition with an opening that overlaps the shutter.

On the 11th day of the study, the passive avoidance reaction was performed in rats in an installation, the triggering stimulus of which was the animal in a brightly lit space. The rat was placed in the bright part of the compartment. After the rat transitioned to the dark part, fatal pain was immediately applied with 5 consecutive stimuli of alternating current (1 mA, each lasting 1 s with an interval of 2 s).

During testing (repeated placement in the illuminated part of the compartment after 24 h in order to determine the preservation of passive avoidance reaction), each animal was observed for 180 s. In this case, the latent period of the rat’s transition to the dark part of the compartment, the time spent in the light and dark parts of the compartment, and the number of animals that did not enter the dark part of the compartment (number of trained animals) were recorded.

The results presented in table 15 indicate that the claimed composition and the comparison drug “Noopept ^® tablets 10 mg”, if used prophylactically, improve the mnemonic functions of the central nervous system of experimental animals: the time spent in the light chamber is increased, the latent period is extended, and the time spent is shortened in a dark chamber where animals experienced pain. Consequently, experimental animals improved memory and learning ability. Moreover, it was experimentally established that the claimed composition is not inferior in effectiveness to the well-known and widely used nootropic drug “Noopept ^® tablets 10 mg”. Effect on the resistance of rats to the effects of hypoxia.

When assessing the adaptogenic and cytoprotective effects of the claimed composition, hypoxia was simulated in a closed volume ("can hypoxia") [38].

Respiration from a confined space - re-respiration - is a fairly adequate and simple model of acute hypoxia [39]. The animal, absorbing oxygen from an enclosed space due to breathing, causes the development of its deficiency - hypoxic hypoxia, which makes it possible to evaluate the studied drug by the integrated mortality rates for a certain observation time and resistance to oxygen deficiency (maximum life expectancy).

The animals were placed in a 2 L jar tightly closed with a glass lid lubricated with sealant. The maximum lifespan and symptoms of thanatogenesis were recorded using a stopwatch. Cans with animals during the study were in the air conditioner, ensuring the constancy of the experimental conditions (temperature + 20 ° C, humidity - 65-70%, atmospheric pressure).

Before the experiment, the animals of the experimental group for four weeks were administered intragastrically the claimed composition at a dose of 150 mg / kg Animals of the control group were injected with 1% starch suspension in a volume of 0.2 ml.

It was experimentally established that the claimed composition contributes to a significant increase in the resistance of animals to the effects of adverse factors modeling the conditions of enclosed spaces, a smoky environment and underground transport, that is, it exhibits a cytoprotective effect (table 16). Thus, the claimed composition significantly increases the life time of rats.

Impact on motor and research activity.

Overweight helps reduce physical activity and emotionality. In this regard, the effect on the indicated indicators of the claimed composition in the case of its prophylactic use was studied.

Movement activity was studied using the automatic activity registration system “Activity Cell” (Ugo Basile, Italy). The experimental animals were divided into two groups: control and experimental 10 animals each.

During the experiment, to assess spontaneous motor activity (SDA), the rat was placed for 3 min in the chamber.

The results of evaluating the effect of the claimed composition on rats SDA are presented in table 17 and indicate that the prophylactic use of the claimed composition helps to increase the overall motor activity of rats.

Research activity was studied using a round plastic arena with a diameter of 100 cm with sides along the perimeter. The arena floor is raised and divided into 9 squares with holes for recording research activity. A light source is placed above the camera. The nature and number of movements are noted by the registrar. Six components of the behavior of experimental animals were recorded during 5 min of observation: the latent period of exit, vertical and horizontal activity, the number of peeps in minks, grooming and boluses. To determine the severity of the effect, the ratio of the number of movements in the experiment and control was used.

From the data presented in table 18, a decrease in the activity of experimental animals of the control group is seen. Obese rats showed a decrease in research activity (the total number of uprights, peeping in the mink), as well as an increase in anxiety (the number of bowel movements and grooming).

The prophylactic use of the claimed composition contributed to an increase in the research activity of experimental animals.

Effect on physical performance and muscle strength.

Physical performance and muscle strength are important indicators of animal activity. The biological activity of the claimed composition was evaluated by the duration of swimming and muscle strength of experimental animals.

Effect on the duration of swimming of mice.

Swimming is a heavy physical dynamic load and allows you to evaluate the effectiveness of adaptogens and the activity of redox processes in the muscles of mice. During the experiment, swimming was carried out at a water temperature of 37 ° C and was carried out with a load, which is a lead tube on a rubber ring attached to the root of the tail of the mouse and comprising 10% of body weight. At the same time, 5 animals from the control and experimental groups swam in a large bath. The animals of the experimental group were daily administered intragastrically by the claimed composition at a dose of 150 mg / kg each. Mice in the control group received daily intragastric 1% starch suspension in a volume of 0.2 ml.

The criterion of fatigue and cessation of swimming was considered the first "diving" of the mouse with the immersion of the nasal passages in water. The duration of the swimming of mice was recorded from the moment they were placed in the bath until the moment of the first “diving”. The results of testing conducted 14 days after daily administration of the claimed composition indicate that daily prophylactic administration to rats of the claimed composition has a protective effect and reduces fatigue (table 19), increases their endurance in the swimming test by approximately 93%, that is, significantly increases adaptive abilities organism.

Effect on the muscle strength of rats.

Muscle strength was evaluated in white rats using a rat dynamometer. The results presented in table 20 indicate that according to the results of the test for muscle strength in the case of daily prophylactic use of the claimed composition, an increase of this indicator by 52% is observed compared to the control.

Example 5. The study of the influence of metabolites of Bacillus subtilis VKPM No. B-2335 on the growth and reproduction of the probiotic lactobacilli Lactobacillus plantarum 8R-A3.

During the experiment, the influence of Bacillus subtilis metabolites VKPM No. B-2335 on the growth of Lactobacillus plantarum 8R-A3 lactobacilli, close to the normal component of the indigenous microflora of the human intestine, was studied in vitro.

Studies were performed using:

- direct antagonism method by direct application of sterile metabolites of Bacillus subtilis bacteria VKPM No. B-2335 or live Lactobacillus plantarum 8R-A3 lactobacilli to wells or to the surface of a solid nutrient medium (MPC-4 agar) immediately after inoculation of Lactobacillus plantarum 8R- lactobacilli into it A3 or indicator test cultures, followed by incubation for 24 hours at 37 ° C;

method of two-layer agar [40], in which a suspension of Bacillus subtilis metabolites VKPM No. B-2335 or live lactobacillus Lactobacillus plantarum 8R-A3 was introduced into the lower layer of solid nutrient medium (MPC-4 agar), and Lactobacillus plant lactobacillus planted in the upper layer -A3 or indicator test cultures.

The metabolites of Bacillus subtilis VKPM No. B-2335 were administered at concentrations of 1 g or 0.1 g in 10 ml of agar.

Lactobacilli were seeded in various amounts of 7 lg CFU / ml-2 lg CFU / ml.

After applying sterile metabolites of Bacillus subtilis VKPM No. B-2335 to wells or to the surface of an agar seeded with Lactobacillus plantarum 8R-A3 lactobacilli, experimentally, using the direct antagonism method, it was found that Bacillus subtilis metabolites VKPM No. B-2335 do not exhibit antagonistic activity against the specified strain of lactobacilli, zones of inhibition of growth of lactobacilli are absent.

Using the double-layer agar method, it was confirmed that metabolites of Bacillus subtilis VKPM No. B-2335, when administered at the indicated concentration, do not inhibit the growth of Lactobacillus plantarum 8R-A3 lactobacilli. It was experimentally established that the metabolites of Bacillus subtilis VKPM No. B-2335 stimulate the growth of probiotic lactobacilli Lactobacillus plantarum 8R-A3 (table 21). In the presence of these metabolites in the accepted concentrations, a larger number of Lactobacillus plantarum 8R-A3 lactobacilli grew (four or more times).

Thus, the lactogenic effect of Bacillus subtilis metabolites VKPM No. B-2335, which is expressed in stimulating the growth and reproduction of the probiotic lactobacilli Lactobacillus plantarum 8R-A3, was experimentally established. This suggests that the metabolites of Bacillus subtilis VKPM No. B-2335 in the composition of the claimed composition are lactogenic factor, which is due to their positive effect in the restoration of intestinal flora.

Example 6. Evaluation of the effectiveness of the claimed composition, tolerability and safety of use.

The effectiveness of the claimed composition was evaluated by the dynamics of the following indicators: clinical signs (the time of the disappearance of subjective clinical symptoms), the state of intestinal microbiocenosis and blood biochemical parameters (lipid patterns, lipid peroxidation indicators, antiradical defense markers).

We observed 40 patients with metabolic syndrome, dyslipidemia, impaired carbohydrate metabolism (increased fasting glucose and impaired carbohydrate tolerance). The average age was 55.7 ± 9 years.

Patients were randomized into 2 groups of 20 people. All patients underwent standard treatment for coronary artery disease, including the administration of nitrates, antiplatelet agents, β-blockers or calcium channel blockers, angiotensin converting enzyme (ACE) inhibitors, and diuretics. Patients of the main group additionally received the claimed composition in a dose of 360 mg 1 time per day with meals. Patients of the comparison group in addition to basic therapy for 1 month. received a drug from the group of statins (simvastatin) at a dose of 20 mg per day.

Effect on the clinical picture.

In a standardized questioning, it was noted that before treatment in 100% of cases, patients complained of pain behind the sternum or in the area of the heart, constricting and pressing, aching in nature, of moderate intensity, occurring when walking more than 500 m, shortness of breath of inspiratory nature when climbing to the 2nd floor, palpitations and interruptions in the work of the heart.

On the background of therapy, positive clinical dynamics was observed by the 30th day regarding regression of the clinical manifestations of IHD in both groups of patients. There was a significant reduction in episodes of pain, a decrease in shortness of breath and sensation of palpitations in all groups (table 22).

Against the background of treatment, positive clinical dynamics were observed in the main group: a decrease in severity up to a complete relief of abdominal pain syndrome and dyspepsia, a significant decrease in taste sensations in the mouth. In the comparison group, the dynamics of complaints from the gastrointestinal tract were not observed. In addition, in the examined patients receiving standard coronary artery disease therapy with an additional prescription of the claimed composition, caprogram normalization was noted (table 23).

Effect on intestinal microbiocenosis

When conducting a dynamic assessment of the state of intestinal microbiocenosis in individuals receiving the claimed composition, a decrease in the severity of dysbiosis, an increase in the presence of obligate intestinal flora, a decrease in the level of conditionally pathogenic microorganisms and staphylococci was found. At the same time, in the comparison group, there was a tendency to increase the severity of dysbiotic changes in the microflora of the colon with a significant decrease in the number of bifidobacteria and lactobacilli and preservation of the previous staphylococcus population (table 24).

Effect on lipid metabolism.

In the indicators of the biochemical analysis of blood against the background of the appointment in the treatment of the claimed composition, there was a positive dynamics of lipidograms, comparable in terms of the main group and the comparison group: all patients showed a tendency to decrease in total cholesterol, triglycerides, and LDL cholesterol. However, a reliable and significant increase in HDL cholesterol was registered only in those patients who were prescribed the claimed composition during treatment. The data on the dynamics of the values of lipidogram indices in patients of the main group and the comparison group are presented, respectively, in FIG. 1 and FIG. 2. Thus, the use of the claimed composition as part of complex therapy has a pronounced lipid-lowering effect, comparable with the action of simvastatin, but it also helps to increase the level of antiatherogenic high density lipoproteins.

Effect on the antioxidant defense system.

The mechanisms of changing the ratio of lipoproteins in the conditions of differentiated treatment greatly clarify the data obtained related to the assessment of the indicators of antioxidant protection and lipid peroxidation (LPO) in patients. The dynamics of lipid peroxidation and the state of the antioxidant system are presented in table 25. From the data presented it can be seen that the functional activity of antiradical enzymes - superoxide dismutase and catalase - increased only in the main group receiving the claimed composition. Activation of the substrate and enzymatic units of antioxidant defense contributed to a significant decrease in the level of LPO reactions in patients of the main group, in contrast to the comparison group, where this indicator, on the contrary, increased.

Assessment of tolerance and safety of the claimed composition.

During the observation period for a month, none of the patients stopped taking the claimed composition. High adherence to the administration of the claimed composition was demonstrated by 100% of patients.

When assessing the safety of the course administration of the claimed composition, there were no significant changes in the indicators of vital functions and the appearance of significant clinical symptoms, no clinically significant changes in the clinical and biochemical parameters of blood, changes in the general analysis of urine, which could indicate a negative effect of the claimed composition on function of relevant organs and systems. As a positive effect, one can note a tendency to a decrease in glucose level in the main group, while in the comparison group the trend was opposite.

The dynamics of the main laboratory parameters in patients included in the safety assessment group are presented in table 26. It has been experimentally established that the claimed composition has a positive effect on carbohydrate metabolism, which is manifested by a significant decrease in blood glucose.

Thus, the studies showed that the impact of the claimed composition, which is a balanced multicomponent complex, is:

- correction of the state of intestinal microbiocenosis, namely: a decrease in the severity of dysbiosis, an increase in the presence of obligate intestinal flora due to the lactogenic effect of Bacillus subtilis metabolites VKPM No. B-2335, a decrease in the level of conditionally pathogenic microorganisms and staphylococci;

- correction of lipid metabolism, which consists in normalizing lipid metabolism, namely: lowering the levels of total cholesterol, triglycerides, low density lipoprotein cholesterol, significantly increasing the level of cholesterol of antiatherogenic high density lipoproteins;

- a positive effect on carbohydrate metabolism, which is manifested by a significant decrease in blood glucose levels;

- regulation of the processes of free radical oxidation due to the activation of substrate and enzymatic (superoxide dismutase and catalase) units of antioxidant protection.

Reception of the claimed composition also provides additional positive effects, which is especially important for people with MS who are trying to maintain the functional state of their body. It significantly increases the adaptive abilities of the body: physical performance and endurance, resistance to extreme factors (including under conditions of hypoxia), mental performance by improving the mnemonic functions of the central nervous system (memory and learning ability).

In addition, the claimed composition is well tolerated by patients with MS, is safe for this category of patients, since it does not contain toxic and hazardous components in quantities exceeding acceptable levels, is non-toxic, does not have the ability to cumulate, does not have undesirable side effects, including toxic effects on reproductive function, mutagenic effect and long-term negative effects.

It should also be noted that when taking the claimed composition does not require a change in the usual way of life. The inventive composition can be taken in addition to food, without changing the diet and / or physical activity during the day, which is important for people with MS.

The inventive composition has a complex multicomponent composition, and its effectiveness with respect to the correction of lipid metabolism disorders in individuals with MS exceeds the efficiency of its constituent components, which is achieved due to the effective integrated recovery of metabolic processes in the body, which are usually disturbed in MS.

The claimed invention meets the criterion of "novelty", since for the first time to correct the manifestations of the metabolic syndrome, it is proposed to use a composition that is a balanced complex, the multicomponent composition of which is selected in such a way that correction of the intestinal microbiocenosis, lipid and carbohydrate metabolism, activation of the substrate and enzymatic units of the antioxidant protection, as well as maintaining the functional state of the body, made in solid form for oral administration nnogo introduction.

The claimed invention meets the criterion of "inventive step", since from the available information it was not obvious the feasibility of combining the probiotic strain of microorganisms, the active natural compound trans-resveratrol and the mitochondrial metabolite coenzyme Q ₁₀ in the composition of the claimed composition to ensure their effective complex effect, as well as the possibility while maintaining the functional state of the body.

The compliance of the claimed invention with the criterion of "suitability for industrial use" is confirmed by numerous data that showed a simple technology for the production of the claimed composition in solid dosage form for oral administration, the ingredients of which are manufactured under industrial conditions and are available, and the claimed composition has high storage stability, is safe, has a high level of biological activity and provides effective correction of the state of intestinal microbiocenosis, lipid and carbohydrate metabolism, activation of the antioxidant defense units and thus allows to maintain the functional condition of the organism.

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Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome

Claims (2)

Composition for correcting dyslipidemia and maintaining the functional state of the human body with metabolic syndrome, which includes: sterilized lyophilized culture fluid obtained by culturing a probiotic strain of bacteria Bacillus subtilis VKPM No. B-2335 and containing metabolites of bacilli, zeolite, calcium stearate, characterized in which is a balanced complex, which additionally includes trans-resveratrol, coenzyme Q ₁₀ , α-tocopherol acetate, lactose, a complex of farms enta containing alpha-amylase, neutral and alkaline proteinases, beta-glucanase, cellulase, xylanase, and silicon dioxide, the composition is made in solid form in the form of granules, capsules or sachets and includes ingredients in the following ratio, wt.%: sterilized lyophilized culture liquid containing probiotic metabolites bacterial strain Bacillus subtilis VKPM No. B-2335                      1.4-1.9 trans resveratrol                                                                                  3.5-4.8 coenzyme Q ₁₀                                                                                              7.1-9.6 α-tocopherol acetate                                                                                  0.4-0.5 lactose                                                                                          13.0-17.5 enzyme complex (alpha-amylase, neutral and alkaline proteinases, beta-glucanase, cellulase, xylanase)                                                                                               0.2-0.3 zeolite                                                                                                       64.2-73.5 calcium stearate and / or silicon dioxide                                               0.9-1.2

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