RU2166950C1 - Bacterial collagenase-base pharmaceutical composition - Google Patents

Abstract

FIELD: medicine, pharmacy. SUBSTANCE: invention relates to development of the novel pharmaceutical composition containing bacterial collagenase. Pharmaceutical composition has collagenase from Streptomycins lavendulae taken in the amount 3-10 U/100 g of product and also antiseptic myramistin, proxanol and polyethylene glycol. New bacterial collagen-base pharmaceutical composition shows nontoxic nonallergenic properties that can be used for healing and cleaning the wound, burn and other damages of skin. EFFECT: enhanced effectiveness of composition. 2 cl, 4 tbl, 2 ex

Description

 The invention relates to medicine, in particular to the creation of a new pharmaceutical composition containing proteolytic enzymes for cleansing wounds and other skin lesions, namely microbial collagenase.

 Known pharmaceutical compositions that promote healing of wounds and other injuries, reduce the number of complications of the pathological process and optimize skin regeneration, containing antioxidants, immunomodulators, peptidases (Kuzina MI, Kostyuchenok BM Wounds and wound infection. - M., 1981; Methodological recommendations of the Pharmacological Committee for the experimental study of drugs for local treatment of purulent wounds, M., 1989; RU 2005488, 01/15/94, RU 2106877, 03/20/98; Enzymatic cleansing and treatment of wounds. // Symposium M. 1977 g.) .

 For the treatment of skin lesions (wounds, burns, frostbite, etc.), pharmaceutical compositions containing collagen are used. Known drugs such as Meturakol, Kolotsil, collagen sponges.

 "Meturakol" is approved by the Pharmacological Committee of the Ministry of Health of the USSR (Reg. N 87/637/3) - a combined preparation consisting of collagen obtained from a bovine split split of cattle and methyluracil is a finely porous dry white plate with a specific odor. "Meturakol" is used as a local remedy for the complex treatment of trophic ulcers, pressure sores, deep and non-healing wounds, superficial skin burns. The drug has an anti-inflammatory effect, stimulates reparative processes in wounds, accelerating the growth and maturation of granulation tissue and epithelization. When applied to the wound, “Meturakol” fits snugly to its surface, absorbs the wound, swells and gradually lyses, releasing its methyluracil.

 Sponge "Kolotsil", Order of the Ministry of Health of the USSR N 1353 dated 11/30/1984 (Registration N 8/353/9), is a combined preparation, contains collagen obtained from the bovine split hair split, furatsilin, boric acid, novocaine, honsurid, represents a three-layer finely porous plate of yellow and white with a slight specific smell. "Kolotsil" is used as a local remedy for the treatment of thermal burns of the II-III degree in the initial stage of their epithelization and for the temporary closure of granulating wounds for burns of IIIb-IV degree. The drug is also prescribed in the treatment of donor skin, trophic ulcers, pressure sores, deep wounds. "Kolotsil" has a stimulating effect on reparative processes in wounds, accelerating the growth of granulation tissue and epithelization, has anti-inflammatory, hemostatic and anesthetic properties, and also prevents plasma loss through the wound surface.

 A hemostatic collagen sponge (Registration N 78/860/7) is prepared from a collagen solution obtained from a bovine cleft split. The composition of the sponge includes boric acid and furatsilin. It is a dry porous mass of yellow color with a slight smell of acetic acid, absorbs liquid well, and slightly swells. The sponge has hemostatic and antiseptic effects, stimulates tissue regeneration.

 Collagen sponge with sanguine retrin (Registration N 96/399/3) contains 0.005 g of sanguine and a collagen solution of 2% (50 g) and is a yellowish-orange plate, it has an antimicrobial and anti-inflammatory effect, stimulates recovery processes, heals wounds, well absorbs wound discharge. A collagen sponge with sanguirytrin is used to treat infected wounds, including festering postoperative purulent wounds in phases 1 and 2 of the wound process, open infected fractures, burn and non-healing wounds and trophic ulcers.

 "Digispon" (Moscow VNIIMP and A.V. Vishnevsky Institute of Surgery) - a dressing gel antimicrobial collagen based on a collagen solution, contains gelatin, dioxidine, glutaraldehyde, is a soft porous plate. Digispon is intended for the treatment and preparation for autodermoplasty of IIIa, IIIb and IV degree burns with a complicated course, after removal of the scab, treatment of flat infected, sluggish, long-healing wounds, trophic ulcers, bedsores. The coating absorbs 40 + 3 g / g of biological fluids, vapor-permeable, plastic, provides a prolonged dosed output of dioxidine into the wound, stimulates reparative processes in conditions of infected wounds, growth of granulation tissue, accelerates marginal and islet epithelization, and helps reduce the risk of formation of coarse scar tissue.

 Collagen-containing preparations, having a number of advantages, have some drawbacks, in particular, each of these drugs can be used only at a certain stage of the wound healing process, sometimes only for severe wounds and other severe injuries of the skin and nearby tissues, and it does not cleanse wounds from necrotic tissues.

 The most well-known general-use preparation Solcoseryl (Instructions for use. Special information Solko, 1990) for healing wounds arising from cuts, burns and frostbite does not destroy collagen necrotic tissues and requires a complex individual therapeutic approach when used.

 The disadvantage of the majority of currently used preparations that heal skin lesions is the narrow focus of the action, their inability to function at different stages of the pathological process, as well as the reproduction of injured growing epithelium and stimulation of regeneration of the skin and nearby tissues. In addition, with their help, it is not possible to achieve optimal wound cleansing, since they do not contain collagenase and are not capable of decomposing collagen in the native state.

 Collagenase is an enzyme with proteolytic activity with a narrowly specific focus on the breakdown of collagen with the release of a free amino acid - hydroxyproline. According to the international classification of collagenase enzymes, it has number 3.4.24.3 (3-hydrolase, 3.4.-acts on peptide bonds, 3.4.24. - metalloproteinase (contains calcium).

Two types of collagenases are known:
1. Collagenases synthesized by certain microorganisms. So. Clostridium histolyticum (a causative agent of gas gangrene) produces collagenase, which breaks down the collagen polypeptide chain in more than 200 sites. This collagenase, which destroys the connective tissue barriers of the host organism, facilitates the penetration (invasion) of highly pathogenic clostridia into the body. The bacterium itself does not contain collagen and therefore is not exposed to collagenase. (Proteolytic enzymes / Eds. Colowick SP, Kaplan NO / Methods in enzymology. 1970. V. 14. Part B, P. 861-872; Mashkovsky M.D. // Medicines. M: Medicine, 1988, p. 11. p. 55
2. Collagenases that are found in amphibians and mammals in tissues growing or undergoing metamorphosis. (Adv. Enzymol. Collagenases and elastases // Ed. Nord FFNY Acad. Press. 1961. V. 23, P. 163-264; Nordwing A. collagenolytic enzymes. NY Acad. Press. 1971. P. 155-206). Collagenolytic activity was first detected in the caudal fin of the tadpoles during metamorphosis, when large amounts of collagen are resorbed in the tissue within a few days.

As a starting material for the isolation of collagenase, a medium was used in which explants of skin tissue and tadpole caudal fin were cultured. After purification, an enzyme preparation was obtained containing in a small amount (5%) impurities of other proteinases with caseinolytic activity. Collagenase of animal tissues has a lower molecular weight than bacterial and differs from the latter in a number of other properties. It was found that the enzyme is inactivated by cysteine and after heating at 50-60 ^o for 10 minutes has an optimum effect at pH 7.0-8.0.

 Collagenase is a unique enzyme that can selectively hydrolyze an insoluble protein - collagen, which is part of the connective tissue of a living organism. By breaking down collagen fibers extremely resistant to the action of other proteases with the formation of high molecular weight fragments, collagenase performs the most important functions in the human and animal body, is involved in the transformation of connective tissue during growth and morphogenesis, as well as in the elimination of certain pathological processes in arthritis, tumor metastases, etc. .

 Collagenases acting on different types of collagen are isolated from tissues of vertebrates, crabs, larvae of flies, insect excreta, as well as from microscopic bacteria, fungi and actinomycetes.

 The currently known group of microbial collagenases, characterized by a wide variety of properties, can be used in basic scientific research, as well as in biotechnology, when effective, specific and rapid hydrolysis of native and denatured collagen fibers of various types is required. An important area of application is in the treatment of a hernia of the intervertebral disc of the spine, burn wounds, frostbite to accelerate the rejection of scabs and necrotic ulcers, to cleanse purulent-necrotic plaques. In addition, they are necessary for the production and cultivation of cell cultures of human and animal tissues, which are a source of plasminogenesis activators in thrombolytic therapy. Viable cells of islets of Langerhans isolated using microbial collagenases are used in a new method for the treatment of diabetes (V.I. Mazurov Biochemistry of collagen proteins. - M .: Medicine, 1974).

 Among collagenases of microbial origin, the most studied and well-known are collagenases synthesized by the bacterium Clostridium histolyticum. Clostridial collagenases are characterized by high specificity of action on collagen fibers and can be referred to as “true” collagenases. They are widely used in biotechnology and for laboratory purposes. However, for the cultivation of Cl. histolyticum and mass production of collagenase require complex precautions, as this bacterium is an anaerobic pathogen that causes gas gangrene and produces toxins. For optimal biosynthesis of collagenases by Cl. Histolyticum used a complex nutrient medium.

 Highly purified clostridial collagenases are active against native and denatured collagen and do not affect other proteins, while crude preparations containing other proteases. able to break down not only collagen, but also other extracellular material, and therefore are widely used to isolate cells in various types of animal tissues. Like amphibian collagenases, purified clostridial collagenases can hydrolyze collagen to completely dissolve it, acting on both fibrous native collagen and three-helical tropocollagen particles.

Purified collagenase from Achromobacter iophagus exhibited a high specificity of action, hydrolyzing only collagen and not acting on other substrates. Its activity was 6 times higher than the activity of clostridyl peptidase. Properties of collagenases from A. iophagus and Cl. histolyticum were similar. Their activity was not affected by parachloremercurium benzoate. It was established that both drugs are metalloenzymes containing Zn in the active center and retaining activity in the presence of Ca ^{2+ ions} .

 In some strains of microorganisms, the ability to biosynthesize extracellular low molecular weight collagenolytic proteases of a new serine type, which differ from the “true” collagenases primarily in the structure of the active center, was found. The same collagenases were isolated from the insects Hypoderma lineatum and crab Uca pugilator.

 In recent years, wound healing compositions containing collagenase have appeared.

 In particular, patent RU 2033805, 04/30/95 describes a method for producing a porous wound healing material containing a collagenase solution. The microbial polysaccharide mannan is dissolved in water to a concentration of 6-8 wt.%, And then mixed with an equal volume of collagenase. This drug performs only one function - cleaning the wound surface from necrotic tissue, does not have antiseptic properties, does not relieve wounds from pus and exudate.

 Closest to the proposed composition is the drug "Iruksol" (Germany, the company Knoll AG D-67008 Ludwigskafen) (Mashkovsky MD Medicines. - M .: Medicine, 1988), which contains collagenase mixed with proteases isolated from the bacterial culture of Clostridium histolyticum (Sigma Handbook, 2000, p. 274).

 1 g of ointment contains clostridiopeptidase A 0.6 U, 10 mg chloramphenicol, concomitant enzymes in a lipophilic anhydrous ointment base.

 "Iruksol" is a combined drug with proteolytic and antimicrobial effects for external use. Collagenase, a proteolytic enzyme isolated from Clostridium histolyticum, causes lysis of necrotic tissues, thereby contributing to the enzymatic, painless cleansing of the wound. In addition, collagenase stimulates the granulation process and does not inhibit epithelization. Collagenase has no effect on intact epithelium, granulation, adipose and muscle tissues. Chloramphenicol is a broad-spectrum antibiotic, active against gram-negative and gram-positive bacteria. The cleansing effect of the Iruxol ointment on the wound in 97% of patients begins from 1 day or no later than the 14th day, while in more than 50% of patients the effect of the ointment is already apparent within the first 6 days.

 Iruksol is used for enzymatic cleansing of wounds, including varicose ulcers, bedsores, gangrene of the extremities, especially in patients with diabetes mellitus, as well as frostbite, long-term non-healing postoperative wounds, after radiation therapy, and in preparation for skin transplantation.

 The disadvantage of the analogue "Iruksol" is the content in its composition of the antibiotic chloramphenicol, which can cause an allergic reaction and currently has little effect on the pathogenic microflora adapted to it. In addition, the basis of "Iruksol" is a fabric-drying vaseline. The collagenase contained in Iruksol was obtained from a pathogenic anaerobic strain of the bacterium, and its isolation is carried out using complex and expensive technology (Susagawara R., Harper E. Publication and characterization of three forms of collagenase from Clostridium histolyticum // Biochemistry. 1984. V. 23 P. 5175-5182).

 The technical task of this invention is the creation of a new more effective and cost-effective wound healing composition, which is characterized by versatile action, versatility, the ability to remove necrotic tissue, pus and exudate from wounds, to carry out their rapid healing, not to overdry the tissue, actively remove scabs, improve the quality of the composition scar tissue, optimize repair processes, suppress pathogenic microflora, without causing allergic reactions, and thereby create amb the use of one drug for the multi-faceted impact on wound process in the different stages of its course.

 This goal is achieved in that the new pharmaceutical composition contains collagenase from Streptomyces lavendulae at the rate of 3-10 Units per 100 g of product.

The composition may contain excipients, such as antiseptics, various ointment excipients and pharmaceutically acceptable additives. In particular, we studied a composition consisting of the following components:
Collagenase from Streptomyces lavendulae - 3-10 units
Miramistin - 0.1-0.5 g
Proxanol - 22.0 - 30.0 g
Polyethylene Glycol - Remaining up to 100 g
Collagenase was chosen as the active principle of the composition, which acts selectively only on collagen fibers, provides gentle and bloodless optimal cleansing of wounds from necrotic tissues. This enzyme is isolated from a non-pathogenic strain of Streptomyces lavendulae (patent RU N 2075219, 03/10/97). The patent describes collagenase isolated from the culture fluid of the strain Streptomyces lavendulae, which has collagenolytic and general proteolytic activity, cleaves the beta chain of native collagen to 84-86 kDa + 26-28kDa, has M.m. 110-120 kDa. In addition, miramistin (FS 42-3498-98) is introduced into the pharmaceutical composition, which is a broad-spectrum antiseptic that does not cause allergies. Further, the composition contains proxanol (NTU-6-14-164-87), which has the ability to delay exudate and pus from the wounds.

 As a basis, the composition includes pharmaceutically acceptable polyethylene glycol.

 A comprehensive study of the wound healing effect of the claimed composition was carried out: a study of the effect of the composition on the rate of regeneration of skin damage under aseptic conditions, the effect on this rate during wound infection, the effect of the composition on the quality of scar formation. To conduct the above studies, the following experimental models were selected that were adequate for the set goals.

 Model of planar aseptic skin wounds.

 Linear skin wound model.

 Model of an infected (purulent) wound.

 Model of a burn wound.

Known methods were used to obtain the model and evaluate the results. (Turishchev S.N., Pharmacology 4/5, 25-36, 1996)
A model of planar aseptic skin wounds was obtained as follows.

 The experiments were carried out on male non-inbred white mice weighing 20-30 g. In animals under sterile conditions under anesthesia (sodium etaminal 50 mg / g) in the interscapular region of the back after preliminary hair removal, a full-layer skin flap was cut so that an oval wound with an area of 80-100 mm. For a comparative study of the dynamics and healing of wounds, visual observation and measurement of the area of wounds was carried out for 2-3 weeks daily, assessing the condition of the edges of the wound, its discharge, and the formation of a scar.

где d₁ - больший диаметр овала;
d₂ - меньший диаметр овала;
S - площадь раны.Treatment of animals began immediately after surgery: the test substance was applied to the wound in the form of an ointment. Control animals received a placebo. As a comparison drug used the well-known wound healing agent - "Iruksol" (Mashkovsky). The rate of wound healing was evaluated by reducing the area of the wound surface. The oval area was calculated by the mathematical formula:

where d ₁ is the larger diameter of the oval;
d ₂ is the smaller diameter of the oval;
S is the area of the wound.

 A model of a linear skin wound was obtained in the experiment as follows.

 The experiments were carried out on white non-inbred male rats weighing 300 g. In animals under anesthesia (sodium ethaminol 40 mg / kg) in the interscapular region of the back after preliminary hair removal, 2 linear incisions were made through all skin layers 1 cm long, one to the right and left of the spine (approximately 2 cm from it). One nodular suture (sterile silk) was applied to the wound. For a comparative study of the dynamics and healing of wounds, visual observation was performed for 2-3 weeks daily, assessing the condition of the edges of the wound, its detachable, and the formation of a scar. The quality of healing was evaluated by microscopic examination of the regenerates. For this, tissues from the area of the defect and the skin adjacent to it were fixed in Carnoy fluid, embedded in paraffin and sections were prepared (8-10 μm thick. The sections were stained with hematoxylin and eosin.

 Treatment of animals began immediately after surgery. A test composition in the form of an ointment was applied to the right wound. The left wound of the same animal served as a control - the ointment base was applied to it. The comparison drug is Iruxol.

A model of an infected (purulent) wound was obtained experimentally
White non-inbred rats weighing 300-320 g were used in the experiments. Under general anesthesia, the hair was cut off on the back of the cervical region of the rats and, under aseptic conditions, a skin site with subcutaneous tissue and superficial fascia was removed. A teflon ring (diameter 8 mm), coated on top with a perforated film, was inserted into the wound. The edges and bottom of the wound were additionally injured with a toothed forceps. 0.5 ml of a suspension of daily culture of pathogenic staphylococcus, strain 209 (1.5 billion microbial cells in 1 ml of physical solution) was injected into the wound surface inside the ring. The rings were removed after 2 days and the standard primary surgical treatment of the wound was performed, after which treatment with the test substance was started. Control animals received a placebo (ointment base).

 The following methods were used as methods for assessing the speed and completeness of wound healing.

Cytological examination of the wound process
A smear imprint from the surface of the wound allows us to judge the nature of the wound discharge and the cellular composition of the granulations. Such a smear is obtained by touching a specially treated glass slide to the surface of the wound, previously purified from pus. The preparations are dried in air, fixed with methyl alcohol and stained according to Romanovsky-Giemsa. At least 400 cells are counted in each smear: neutrophilic leukocytes, mononuclear cells, macrophages, fibroblasts, as well as their percentage.

Microbiological examination of an infected wound
Microbiological monitoring of the effectiveness of the treatment of wounds is carried out in dynamics (every 3 days) compared with baseline (before treatment). The qualitative composition of the microflora of the wound and the quantitative dynamics of microbial bodies in the wound are studied.

Histological studies in the dynamics of wound healing give an idea of the qualitative changes in the wound under the influence of the drug. Used staining with hematoxylin and eosin
Morphometric examination of scar tissue
Pieces of skin taken on the 13th day after surgery for histological examination were fixed in Carnoy fluid, stepwise paraffin sections were made, stained with hematoxylin and eosin and examined under a light microscope. A morphometric study of scar tissue was carried out: in serial preparations using a grid of Avtandilova G.G. the method of point counting was used to determine the bulk density of connective tissue and cellular elements. The percentage of fibroblasts per 500 cells was counted.

 The results were processed using statistical methods: the arithmetic mean value was determined, the arithmetic mean error, the significance of differences between the groups was determined using Student's criterion.

 For comparative studies used 3 groups of animals.

 The first group of animals received ointment treatment based on the claimed composition with an active ingredient content of 3 units, the second group of animals received ointment treatment based on the claimed composition with an active component content of 7 units. The third group of animals received treatment with ointment "Iruksol". Placebo was used as a control in all three groups.

 The results of the study are shown in tables 1 and 2.

Table 1 shows the dynamics of healing of wounds of the skin of the back in mice (area of planar wounds (ellipse) in mm ² under the influence of the claimed composition.

 Table 2 shows the morphometric indicators characterizing the healing of linear skin wounds in rats in the control and when exposed to the claimed composition.

 The results of studies conducted on models of linear and planar wounds showed that the claimed composition has wound healing properties and, in addition, has a positive effect on the qualitative composition of the scar tissue formed during the healing of aseptic wounds, as evidenced by the ratio of fibroblasts and fibrocytes in the connective tissue of the wound scar treated with ointment.

 The invention is illustrated by the following examples of formulations.

Example 1. Collagenase from Streptomyces lavendulae - 3 PIECES
Miramistin - 0.1 g
Proxanol - 22.0 g
Polyethylene glycol - the rest is up to 100 g
Example 2

Collagenase from Streptomyces lavendulae - 10 PIECES
Miramistin - 0.5 g
Proxanol - 30.0 g
Polyethylene glycol - the rest is up to 100 g
For toxicological studies, the active ingredient of the composition, i.e. containing enzymatic components, this ingredient has a brown color, a specific odor, pH = 5.0-7.0.

 The study of acute toxicity of the substance was carried out on white mice (males and females with a body weight of 19-21 g).

 The experiments were carried out in accordance with the "Methodological recommendations for the study of general toxic effects (pharmacological agents" (1997).

 The substance of the active ingredient was administered to animals (mice) intravenously, once, in a volume of 0.2-0.5 ml, at a rate of 0.1 ml / s.

 The duration of observation of mice was 10-15 days. During the experiment, we monitored the behavior, appearance, motor activity, and the reaction of animals to external stimuli. In cases of death, the clinical picture was recorded and an autopsy was performed on dead mice.

With the intravenous administration of the drug to mice in doses up to 3500-4000 U / kg, no signs of intoxication were observed. Animals remained motile, reacted normally to the environment. With a further increase in the dose of the administered substance, the death of individual animals was noted. Medium lethal doses (LD ₅₀ ) were established for mice with a single intravenous administration of a substance: LD ₅₀ = 12400 ± 2500 U / kg.

 The maximum tolerated dose of the substance upon intravenous administration to mice was ≈4500 U / kg. In these cases, when observing animals for 2 weeks, death was not observed.

 Studies have also been conducted on the microbiological purity and pyrogenicity of the claimed composition.

 Studies have been conducted for the presence of pyrogenic impurities in the drug. The test was carried out on chinchilla rabbits, kept on a standard diet of vivarium.

 Since this drug is used only externally as an ointment, it is difficult to establish a clear therapeutic dose. Therefore, to select a test dose for the pyrogenicity test, we used a dose for a collagenase preparation - 5 U / kg.

 It was found that this drug in a test dose of 5 PIECES per kg of animal weight is pyrogenic.

 A tendency toward a decrease in the pyrogenic reaction with a decrease in the input volume was noted.

 The establishment of a clear therapeutic dose will make it possible to more reliably select a test dose for pyrogenicity. Conducting additional tests with new samples of the drug, in a minimum volume of diluent, in a reasonable test dose, may allow you to get results indicating the absence of pyrogenicity.

 A comparative study of the antimicrobial activity of our composition and Iruksol ointment was carried out using the "wells" method (Methodological recommendations for experimental - preclinical study of drugs for local treatment of wounds. - M., 1989), for which medium N 3 and N 6 were used (formulations named media are given in the indicated recommendations). Clinical freshly isolated strains of microorganisms (S. aureus, S. epidermidis, E. Coli., Proteus sp., P. aeruginosa) from patients with purulent soft tissue wounds were used as test cultures.

 Isolation and identification of pure culture was carried out by the method described in the methodological recommendations "Bacteriological diagnosis of wound infection" (M., 1984). Microorganisms were grown on solid nutrient medium - on meat-peptone agar.

Microbial suspension of test cultures was prepared in accordance with the turbidity standard of the State Institute of Sanitary Control (GISK) them. L.A. Tarasevich. The concentration of the experimental suspension was 10 ⁸ microbial cells in 1 ml

The experimental and control microorganisms were cultured in a thermostat at a temperature of -37 ^o C for 24 to 48 hours. For the zone of growth inhibition of microorganisms, the results were taken into account.

The antimicrobial activity was studied:
- the basics of the composition (a combination of proxanol and polyethylene glycol);
- ointment composition without miramistin (collagenase + proxanol + polyethylene glycol);
- the claimed composition, in which the above basis, miramistin and collgenase.

 The control drug was Iruksol, a combination drug containing fat base, collagenase and chloramphenicol.

 The results of the study of antimicrobial activity are presented in table 4.

 The results indicate that the developed ointment composition is an active drug in relation to gram-positive and gram-negative flora. Given that the Staphylococcus group included coagulase-negative and methicillin-resistant strains (25%), the results can be considered positive.

 Positive results were also obtained for P. aeruginosa, a growth inhibition zone of 15.8 mm indicates the susceptibility of Pseudomonas aeruginosa to the test ointment, although P. aeruginosa is often known to be multiresistant to antibacterial agents.

 The claimed composition has less activity against bacteria of the Enterobacteriaceae family - E. coli, Proteus, their sensitivity can be compared as average, but it must be borne in mind that the components that make up the ointment base are classified as semipermeable.

 In accordance with modern concepts, the combination of semi-penetrating and non-penetrating components inside the cells protects healthy wound tissues from osmotic shock and makes their use harmless.

 As can be seen from the table, all studied strains of microorganisms were resistant to ointment "Iruksol".

 Conducted studies of ointments for sterility showed the absence of microorganisms in the claimed composition.

 Thus, studies on a new composition based on collagenase isolated from Streptomyces lavendulae showed that the proposed new drug is a highly effective, non-toxic, non-allergenic agent for healing wound, burn and other skin lesions.

Claims (2)

 1. A pharmaceutical composition based on collagenase of microbial origin, containing the named enzyme, antiseptic and ointment excipient, characterized in that it contains collagenase from Streptomyces lavendulae at the rate of 3-10 UNITS per 100 g of product. 2. The pharmaceutical composition according to claim 1, characterized in that it contains miramistin as an antiseptic, additionally contains proxanol, and a pharmaceutically acceptable polyethylene glycol (PEG) as an ointment excipient in the following ratio of components:
Collagenase - 3 - 10 units
Miramistin - 0.1 - 0.5 g
Proxanol - 22.0 - 30.0 g
PEG - The rest is up to 100 g

Images