RU2456016C2 - Wound-healing agent for wound preventing and repair in diabetic patients - Google Patents

Abstract

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to chemical-pharmaceutical industry and concerns treating diabetic patients. The wound-healing agents for wound preventing and repair in diabetic patients contains alcohol-water extracts of yellow melilot, laurel, purple Echinacea, and also a solid extract of common licorice, aloe vera juice, taurine and the base PEO-400 and PEO-1500 alloy in the following proportions, wt %: the alcohol-water extract of yellow melilot 2.0-2.5, the alcohol-water extract of laurel 3.0-3.5, the alcohol-water extract of purple Echinacea 1.0-1.5, the solid extract of common licorice 1.0-1.5, aloe vera juice 1.0-1.5, taurine 2.0-2.5, the base - to 100.0.

EFFECT: agent promotes line skin wound healing with underlying experimental alloxan diabetes at all three stages of the wound process (exudation, proliferation, epithelisation).

1 ex, 3 tbl, 3 dwg

Description

The invention relates to medicine and for the treatment of patients with diabetes mellitus. It can be used for the prevention and treatment of complications of diabetes mellitus - diabetic foot syndrome (SDS).

Damage to the skin of the feet for a healthy person will be practically invisible, but for a patient with diabetes mellitus, against the background of a weakened immune system, the slightest cut, wear on the legs is enough for infection to occur. This condition is usually accompanied by hyperthermia, leukocytosis. In such cases, urgent surgical intervention with necrectomy is necessary. With extensive purulent-necrotic lesions, amputation is performed. In the United States, more than 50,000 amputations are performed annually, each costing about $ 10,000, with total costs reaching $ 500 million (Komelyagina E.Yu., 1998). In Russia, about 11-12 thousand high amputations are performed annually, which is relatively more than in other countries (Dedov I.I., 2000). According to Russian researchers, after amputation of the lower extremities, postoperative complications are observed in 64% of patients, the average length of hospital stay is 58.2-65.7 days, and in every second life expectancy after surgery does not exceed two years (Briskin B.S. et al. ., 1999).

Over the past 20 years, the attitude towards amputations has changed in the direction of preserving the limbs as much as possible and preventing them.

The question of the most rational composition of the comprehensive treatment of CDS and the appropriateness of including some new methods in it remains relevant. However, the use of local therapy is still an indispensable method of treatment, and, in particular, ointments of various compositions are used. The selection of optimal agents for topical treatment of trophic disorders of the skin of the lower extremities in diabetes is an urgent problem. Doctors from all over the world face it.

Local treatment should correspond to the stage of the wound process:

1. exudation;

2. proliferation;

3. epithelization.

In recent years, there have been reports of the treatment of flaccid granulating ulcers in CDS with the Kuriozin preparation, which is an associate of hyaluronic acid and zinc. The positive effect of this drug in patients with SDS was noted in the 1st phase of the wound healing process, and since patients with the 2-3rd phase of the wound healing process come to the hospital, Curiosin can only be used in the complex treatment of SDS.

The Levomekol ointment has proven itself well (V. Datsenko, T. Tamm. Study of a multicomponent water-soluble ointment with dioxidine / V. M. Dedov, T. I. Tram // Klin. Khir. - 1981, No. 1 , p. 43-45). This is a combined drug (chloramphenicol and methyluracil) for external use, with an antibacterial, anti-inflammatory effect. But it is also indicated for use mainly in the 1st phase of the wound process and often causes an allergic reaction - a skin rash.

It should be noted the use of ointment "Levosin" containing chloramphenicol, sulfadimethoxine, trimecaine, methyluracil. It has established itself as a tool used to treat infected wounds and burns (Pertsev I.M. et al., 1999). The antimicrobial effect of the ointment against pyogenic cocci, some protea strains, dysenteric bacteria and blue-green pus bacillus is significantly enhanced due to the presence of prolonged sulfanilamide - sulfadimethoxin. At the same time, it has an undesirable sharp and uncontrolled increase in the pharmacological action of substances, and also dehydrates the mucous membranes, causing irritation and a burning sensation.

There are positive reviews about the local treatment of diabetic ulcers with Ximedon (RF patent No. 2063752, IPC A61K 31/505, 9/06, 1996). This is a new pyrimidine derivative comprising xymedon and boric acid. The drug inhibits the growth of blue-green pus bacillus, but representatives of anaerobic non-clostridial microflora of the bacteroid class belong to the main role in the spread of infection through the tendons and synovial sheaths. In addition, the bacterial landscape of the wound in a patient with SDS is very wide, therefore, Xymedon can only be used as a drug for the complex treatment of SDS. It has some toxicity; in patients with diabetes mellitus, glucose tolerance should be more often determined.

All this speaks in favor of the search for new drugs for the prevention, treatment of diabetes and its complications, including those based on natural compounds. Advantages of the latter: a wide range of pharmacological effects with low toxicity, rare side effects and minimal contraindications. A complex combination of phyto compounds with accompanying substances can have a beneficial effect on the structure of the skin and the body as a whole: accelerate the regeneration of epidermal cells, penetrate deep into the subcutaneous tissue, while regulating the water-salt balance and tone, activate enzyme systems, normalize hormonal and general metabolism. The combination in the phytocomposition of certain medicinal plant materials suggests the presence of various types of activity and to obtain medicinal and therapeutic and prophylactic agents with desired properties (RF patent No. 2323275, IPC A61K 36/00, 35/64, A61P 17/02, 2009; 2328301, A61K 36/48, 2007; 2294209, A61K 36/48, 2007; 2377975, A61K 36/835, 36/54; 2290943, A61K 36/48, 2007; 2191589, A61K 36 / 28.2002).

For several years, the authors have been researching and developing gels with phytocompositions for correcting and preventing the consequences of diabetes mellitus, namely, diabetic foot syndrome (SDS) (Ogay MA / Development and research of gels with phytocompositions for preventing the consequences of diabetes mellitus / M .A.Ogai, E.F. Stepanova // Izv. VUZov. North-Caucasian region, special issue. - 2007. - P.99-100; Ogay MA. Pharmacological studies and technology of phytogels for correction of the consequences of diabetes mellitus / M .A.Ogai, E.F. Stepanova, L.P. Lariono , A.Yu. Petrov // Bulletin of the Voronezh State University, series: Chemistry. Biology. Pharmacy. - No. 2, 2009. - P.171-173; Ohay MA Pharmacotechnological studies of external medicinal products - phytogels based on medicinal plant materials for for prevention of the consequences of diabetes mellitus / M.A. Ogai, E.F. Stepanova, L.P. Larionov, A.Yu. Petrov // Kursk Scientific and Practical Bulletin "Man and His Health", No. 1. - 2010. - P .123-126 (prototype)). In these works, the wound healing activity of gels of various compositions based on herbal raw materials was studied, which included alcohol-water extracts from various medicinal plants, including sweet clover, noble laurel, purple coneflower, as well as dense licorice extract, aloe vera and individual preparation taurine. As a result of these studies, the wound healing activity of the proposed gels in animals with experimental pathology was verified and it was concluded that it is possible to study the latter in the future for the prevention and treatment of some consequences of diabetes mellitus, including “diabetic foot”. However, the optimal qualitative and quantitative ratios of the components of drugs, including alcohol-water extracts of medicinal sweet clover, noble laurel, echinacea purpurea, as well as a thick extract of licorice, aloe vera juice and taurine, have not been investigated and proposed.

The objective of the invention is to develop a new soft dosage form - phytogel for the prevention and treatment of SDS.

The technical result consists in effective tissue regeneration due to the optimal qualitative and quantitative ratio of the components of the drug prepared according to the indicated technology at all three stages of the wound healing process (exudation, proliferation, epithelization).

The technical result is achieved by the fact that the wound healing agent for the prevention and treatment of wounds in patients with diabetes mellitus (phytogel) contains alcohol-water extracts from the sweet clover, noble laurel, echinacea purpurea, as well as a dense licorice extract, aloe tree-like juice and the individual preparation taurine, as basics alloy PEO-400 and PEO-1500 with the following components, wt.%:

Alcohol extracts from clover herb 2.0-2.5 from the leaves of laurel noble 3.0-3.5 Echinacea purpurea herb 1.0-1.5 thick licorice extract 1.0-1.5 aloe juice 1.0-1.5 taurine 2.0-2.5 the basis the rest is up to 100.0

The dosage form was tested at the Pyatigorsk State Pharmaceutical Academy at the Department of Drug Technology.

The selection of qualitative and quantitative composition in this phytogel was carried out on the basis of a biological test on unicellular organisms - Parametium caudatum. This test allows you to determine the optimal ratio of components, their membrane-stabilizing and antioxidant effect. The developed phytogel significantly lengthened the paramecium stop time under the influence of cellular poisons - ethyl alcohol and hydrogen peroxide. The lengthening of the time to stop the movement of paramecium under the influence of ethyl alcohol characterizes the membrane-stabilizing activity of the developed phytogel, the components of which prevent damage to the protein part of the biomembrane. Antioxidant activity was verified by lengthening the movement time of paramecium under the influence of a 1% hydrogen peroxide solution. Apparently, this is due to the ability of the components of the developed phytogel to inhibit the lipid peroxidation of the membrane (Kudrin A.N. The system of express methods for the integrated assessment of the biological activity of individual and complex preparations at biological objects / A.N. Y. Balabyan // Russian Chemical Chem. Journal - 1997. - T.41. - No. 5. - S.114-123.

Table 1, reflects the composition of the proposed funds; table 2, contains indicators of glucose and lipid spectrum in alloxan diabetes in rats; table 3 contains the average linear wound size in rats; figure 1 - picture of a histological section in the control group; figure 2 - picture of a histological section in the group treated with the drug levomekol; figure 3 is a picture of a histological section in the group treated with the proposed tool.

In experimental studies, the wound healing effect of the developed composition on linear skin wound models against the background of experimental alloxan diabetes was first revealed.

Melilotus officinalis is a grassy aromatic biennial (GOST 14101-69). Contains coumarin, coumaric acid, mellotine, purine derivatives, essential oil, vitamin C, E, carotene. Fresh grass of Melilotus officinalis contains the coumarin glycoside, from which coumarin is formed upon drying. Coumarin contained in the herb of Melilotus officinalis has anti-inflammatory, analgesic and softening effects in boils, carbuncles, and purulent wounds. There are known studies on the analgesic and wound healing effect of ointment with sweet clover (RF patent No. 2329822), the maximum effectiveness of which is manifested when its content is 1.5-2.5 wt.%.

The leaves of the noble laurel (GOST 17594-81) contain essential oil, in its composition - myrcene, limonene, fellandren, pinene, camphene, cimol, sabinen, cineol, eugenol, methyleugenol, linalool, terpinolene, carvone. Sesquiterpene lactones: desacetyl laurenobiolide, laurenobiolide, costunolide. Flavonoids are represented by rutin, in the kempferol hydrolyzate, quercetin (Plant resources of the USSR (additions): Flowering plants, their chemical composition and use. - L .: Nauka, 1996. - P.160). It is known to use paste from dry leaves in sunflower oil for arthritis, myositis and neuralgia, decoction for dermatomycosis. The fruits were included in the I-VII edition of domestic pharmacopoeias. Fatty oil (bean ointment) was used for rubbing as a locally irritating skin product for rheumatism, paralysis, colds, scabies (Damirov, I. A. Medicinal plants of Azerbaijan / I. A. Damirov [et al.]. Baku: "Maarif", 1988 - 319 p.).

Echinacea purpurea grass (TU 9379-004-46897343-05 - grass) contains polysaccharides (heteroxylans and arabinoramnogalactans), essential oil, flavonoids, hydroxycinnamic acids (chicorye, ferulic, coumaric, coffee), tannins, saponins, aminequin acids), echinacoside (a glycoside containing coffee acid and pyrocatechol), resins, phytosterols, enzymes, and macro- and microelements. Echinacea Hexal and Echinacea Ratiopharm enter the domestic market from Germany. The drugs act as biogenic stimulants, activate cellular immunity, stimulate bone marrow hematopoiesis (Kurkin V.A. Anatomical and morphological study of Echinacea purpurea using digital microscopy / V.A. Kurkin, A.S. Chepurnova, V.M. Ryzhov, L.V. Tarasenko, G.N. Suvorova, E.V. Avdeeva // Medical almanac. - No. 2 (7). - 2009. - June. - P.204-206).

A wide range of biological activity of licorice root preparations is due to the presence of triterpene saponin - glycyrrhizic acid, its glycerretic acid aglycone, flavonoids - liquvirithin, lucurazide, etc. The results of studies on the chemistry, technology, pharmacology, biosynthesis of the main groups of biologically active substances of licorice root and rhizome, medical use and The food industry is reflected in a number of reviews and monographs. Licorice products and preparations are used as painkillers, anti-inflammatory, choleretic, hepatoprotective, laxative, diuretic and antispasmodic drugs. Preparations containing licorice extracts are intended for the treatment of erysipelas and eczema (Ammosov A.S. Use of licorice in world practice (Review of patent sources) / A.S. Ammosov, V.I. Litvinenko, T.P. Popova // Chemical and Pharmaceutical Manufacturing: Obzor. Inform. - M .: 1992. - Issue 1: - 68 p.). The finished product was used extract of licorice thick dense (VFS 42-2614-89).

The juice of aloe vera leaves contains anthracene derivatives (aloe-emodin), tarry and bitter substances, polysaccharides, succinic acid, traces of essential oil, vitamins, enzymes. Aloe is an old remedy. Currently widely used in eye practice, with diseases of the gastrointestinal tract, with anemia, radiation injuries in dentistry. The therapeutic effect of drugs is based on increasing the protective properties of the body. Juice is used for gastritis, gastroenteritis, enterocolitis; externally in the treatment of purulent wounds, burns, inflammatory skin diseases (Petkov, Veselin. Modern herbal medicine / Veselin, Petkov. - Sofia: Medicine and physical education, 1988 - 504 p.). The finished product was used aloe juice (PH LSR-000083, Moscow Endocrine Plant).

Taurine aminoethanesulfonic acid. Taurine can increase glucose utilization by animal cells. It removes the “oxidative burst” caused by homocysteine and reduces its negative effect on blood vessels. It has antioxidant properties. It has a positive effect on the metabolism of people with diabetes (Nedosugova L.V. et al., 2005). Taurine was used as a substance.

The base used is water-soluble and contains a hydrophilic non-aqueous solvent PEO-400 and a water-soluble polymer PEO-1500, related to polyethylene oxides, which are physiologically indifferent compounds. When mixing the PEO-400 solvent with the PEO-1500 polymer, the latter acts as a thickener, which, depending on the amount in the total mass, provides the required consistency of the created dosage form. Polyethylene oxides provide the interstitial action of the dosage form, which provides suppression of deep microflora. PEO-400 absorbs more water and has a long period of osmotic action, and PEO-1500, which is also an osmotically active substance, absorbs purulent exudate due to the binding of water molecules to hydroxyl and ether groups. This basis is recommended for ointment dosage forms used in the local treatment of wounds in the 1st phase of the wound healing process (RF patent No. 2367469, IPC A61K 38/43, 9/06, 2008).

Despite the fact that individual components and their combinations are found in well-known therapeutic ointments for this purpose, the proposed combination and their ratio were not found. The claimed combination of components, taken in experimentally selected, optimal amount, provides the claimed technical result. Other ratios of the components taken do not allow to achieve the necessary therapeutic effect.

A wound healing agent was prepared as follows.

Comprehensive alcohol-water extraction from medicinal sweet clover grass, noble laurel leaves and Echinacea purpurea grass was prepared as follows.

Raw materials, crushed to a size of 3-5 mm, were loaded into a maceration tank with a mechanical stirrer, poured with five times the volume of extractant (40% ethyl alcohol), taking into account the absorption coefficient of raw materials obtained experimentally (absorption coefficient of raw materials - 2.0). The use of 40% ethyl alcohol is due to the fact that it was this concentration that made it possible to extract flavonoids and related substances as efficiently as possible. The extraction was carried out for 2 days with periodic stirring at room temperature. After which the raw material was squeezed out, filtered through a double layer of gauze (Extract 1). Next, the meal was re-filled with three times the amount of extractant, also taking into account the absorption coefficient of the raw material. The extraction was carried out during the day with periodic stirring at room temperature. After that, the cake was re-squeezed and filtered through a double layer of gauze (Extract 2). Periodic change of the extractant made it possible, with less time spent on extraction, to more completely exhaust the raw materials, to reduce losses during diffusion, since the difference in concentrations is maintained and, as a consequence, the diffusion rate. Both extracts were combined and evaporated in a vacuum evaporator to a thick consistency.

To the obtained complex extraction, finished products were added dense licorice extract, aloe juice and taurine substance in the ratio (2: 3: 1): 1: 1: 2.

Next, a phytogel was prepared. The extraction was introduced at the rate of 10.0 wt.% To 90.0 wt.% Of the base, which was an alloy of PEO-400 and PEO-1500, taken in a ratio of 1: 1.

In the reactor with a steam jacket, the refractory component PEO - 1500 was melted, then PEO - 400 was introduced and mixed.

When the mixer was rotating at a speed of not more than 30 rpm, the necessary amount of molten base was fed into which the extracts and taurine were added in parts. The homogenization of the gel was carried out using a stirrer until a mass was uniform throughout the volume for 30-40 minutes.

The finished phytogel was subjected to analysis for homogeneity, appearance, organoleptic properties, microbiological purity, qualitative and quantitative content of biologically active substances.

Example

In the experiment, white outbred rats of both sexes weighing 150-175.0 g were used. Animals were obtained from the nursery, were quarantined for two weeks and kept in standard vivarium conditions. In each experimental group there were six animals (n = 6). A model of diabetes mellitus was created by intraperitoneal administration of alloxan hydrate at a dose of 170 mg / kg. Diabetes developed on the 4th day after administration of the drug.

On the 4th day after intraperitoneal administration of alloxan hydrate, 1 drop of blood was taken from the tail vein and blood glucose, total cholesterol, triglycerides, high density lipoproteins (HDL) were measured (see Table 2). All of these indicators were measured using a CardioChek portable biochemical blood analyzer using appropriate test strips. Then calculated the content of low density lipoproteins (LDL) mmol / l according to the formula 1:

The change in the above indicators was recorded upward (except for HDL, the level of which decreased from 1.03 ± 0.01 to 0.52 ± 0.04).

Linear skin wounds were applied under anesthesia - chloral hydrate (300 mg / kg). The skin of the back was cut to its own fascia. The cut length averaged 25 mm. Then, at an equal distance from the edges of the wound 1 suture was applied, bringing together the edges of the wound. For the convenience of subsequent measurement of the size of the wounds, the sutures were imposed so that the epithelium of the lateral edges of the wound did not come into contact, and in this case the epithelization occurred from the final edges of the wound.

Evaluation of the wound healing effect was carried out according to the nature of the clinical course (the presence of suppuration, the time of complete rejection of the scab, the time and dynamics of the complete fusion of the edges of the wound).

The studied objects were applied to the wounds of animals - the first group of animals was not treated (control), the second group of animals was daily applied to the wound surface with levomekol (comparison drug), the third group of animals - phytogel. All preparations were applied throughout the entire period of the experiment until the wounds healed completely.

The effect of the developed phytogel on the healing time of a linear skin wound in rats with experimental alloxan diabetes (see table 3).

By the fifteenth day of the experiment, the studied phytogel noticeably exceeds the comparison drug levomekol by 48.68%, the differences are significant (p ₁ ≤0.05 *). By the twentieth day of the experiment, the studied phytogel is 25.48% more active than the reference drug.

Complete healing of linear skin wounds was observed in two experimental groups by the twenty-fifth day of the experiment, with the exception of the control group, where M averaged 1.30 mm even by the 25th day of the experiment.

For histological examination, sections of the formed scars of three groups of experimental animals with alloxan diabetes and a linear skin wound are presented. The first group - did not receive treatment (control); the second group is treated with a comparison drug (levomecol); the third group is treated with phytogel.

When a histological examination of the slice in the control group, wound healing occurred by secondary intention (see figure 4). The wound defect is filled with granulation tissue, in which cellular elements of connective tissue predominate. The resulting vessels, inside covered with endothelium, are viewed. Incomplete epithelization is observed from the edges of the wound. Under the fused epithelium, an incomplete granulation cavity is visible, partially filled with the remains of necrotic tissue. Layers of horizontally located fibroblasts and collagen fibers are not formed, which indicates a sluggish current healing process.

When histological examination in the group treated with the comparison drug levomekol (see figure 2), epithelization of the wound surface is completely completed. The scab on the surface of the scar is absent. There is no superficial leukocyte-necrotic layer. The formed scar is wide; cellular elements of connective tissue prevail in it. A small amount of the vascular component of the granulation tissue, which is in the involution stage, is visible. Germination from below of collagen fibers is weakly expressed.

On histological sections of the third group treated with phytogel, the surface of the scar is completely covered with the epidermis (see figure 3). Under the epidermis, longitudinal layers formed by strands of fibroblasts formed. The deep layer of the scar consists mainly of fibrous elements - collagen and elastic fibers, gradually displacing the cellular component.

As a result of a histological study of sections of the formed scars after a linear skin wound on the background of experimental alloxan diabetes, it was shown that the developed phytogel in terms of indicators of the degree of epithelization and keratinization of the epidermis, the ratio of cellular and fibrous elements of connective tissue, the severity and involution of the vascular component of granulation tissue is not inferior in activity comparison drug levomekol.

The benefits of using the proposed composition:

1. The developed phytogel promotes the healing of linear skin wounds on the background of experimental alloxan diabetes mellitus at all three stages of the wound healing process (exudation, proliferation, epithelization).

2. The histological characteristics of sections of a linear skin wound characterize the maturity of the formed scar.

Table 1 The composition of the components The ratio of components to 100.0 finished phytogel Alcoholic extracts from the herb of Melilotus officinalis 2.0-2.5 from the leaves of laurel noble 3.0-3.5 Echinacea purpurea herb 1.0-1.5 thick licorice extract 1.0-1.5 aloe juice 1.0-1.5 taurine 2.0-2.5 Basis: alloy PEO-400 and PEO-1500 (1: 1) up to 100.0

table 2 Indicators of glucose and lipid spectrum in alloxan diabetes in rats, (n = 6) Before the experiment (mmol / l) Against the background of alloxan diabetes (mol / l) Glucose 3.85 ± 0.30 26.03 ± 0.67 Triglycerides 1,02 ± 0,15 1.59 ± 0.01 Total cholesterol 1.30 ± 0.05 2.56 ± 0.17 HDL 1.03 ± 0.01 0.52 ± 0.04 LDL 0,07 1.72

Claims (1)

Wound healing agent for the prevention and treatment of wounds in patients with diabetes mellitus, characterized in that it contains alcohol-water extracts: medicinal sweet clover, noble laurel, echinacea purpurea, as well as a thick extract of licorice, tree aloe juice, taurine and the base - PEO-400 and PEO alloy -1500 with the following components, wt.%:
Alcoholic extracts: Melilotus officinalis herbs 2.0-2.5 noble laurel leaves 3.0-3.5 Echinacea purpurea herb 1.0-1.5 thick licorice extract 1.0-1.5 aloe juice 1.0-1.5 taurine 2.0-2.5 the basis rest

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