RU2341282C2 - Pharmaceutical composition of pancreatropic activity and method of pharmaceutical composition production - Google Patents

Abstract

FIELD: pharmaceutical industry.

SUBSTANCE: composition contains Octreotide and Dalargin in therapeutically effective amounts as active components. Method of composition production includes dissolution of components in purified water followed by lyophilisation of solution.

EFFECT: provides high pancreatropic activity and can be applied for treatment of diseases associated with pancreatic pathology.

5 cl, 3 ex, 5 tbl

Description

The invention relates to medicine, specifically to a pharmaceutical composition having pancreatotropic activity and pronounced therapeutic effect.

One of the urgent problems of modern gastroenterology is the treatment of patients with acute and chronic forms of pancreatitis and their exacerbations [1].

Recently, the importance of endogenous opioid peptides in the formation of stress reactions in inflammatory diseases of the abdominal cavity, including lesions of the pancreatic parenchyma, has been noted.

Known drug octreotide, which is widely used for the prevention of complications after pancreatic surgery and the treatment of acute pancreatitis. Octreotide has a pronounced ability to inhibit the exocrine function of the pancreas, activates the healing processes of its parenchyma [2].

Known drug dalargin, created on the basis of endogenous opioid peptides, which also has pancreatotropic activity and is used in the conservative treatment of patients with acute and chronic forms of pancreatitis [3]. In addition, dalargin has a pronounced stress-protective and analgesic activity [4].

The objective of the invention is to create a new

a combined drug with pronounced pancreatotropic activity and therapeutic effect.

The problem is achieved by the described pharmaceutical composition, which contains dalargin and octreotide in effective amounts as active components.

The invention is illustrated by the following examples.

Example 1

8 l of water for injection is poured into a reactor equipped with an anchor stirrer. Then the valve is opened to supply sterile nitrogen to the reactor and water is bubbled for 5 minutes. After that, 10 g of dalargin substance and 1 g of octreotide substance are successively loaded into the reactor with constant stirring and bubbling of the solution with nitrogen. Mixing lead to complete dissolution. Then, the pH of the solution was adjusted to a value of 4.5-5.5 by the addition of a 1 M solution of acetic acid. The resulting solution was adjusted to a volume of 10 l with water for injection and passed through a sterilizing filter. Under aseptic conditions, bottling in ampoules with a capacity of 2 ml, 1 ml per ampoule, is carried out. Cartridges with ampoules are placed in a sublimation chamber, where they are frozen to a temperature of minus 40 ° С with a freezing rate of 10 ° С / h, and then subjected to drying by vacuum. Ampoules with the obtained dry lyophilized form of the drug are sealed under nitrogen.

Example 2

1 g of the substance of dalargin and 0.1 g of the substance of octreotide are dissolved with stirring and bubbling with nitrogen in 900 ml of water for injection, in which 0.71 g of ammonium acetate is previously dissolved, the pH of the resulting solution is adjusted to 4.5-5.0 with 1 M solution of acetic acid and add water for injection to a total volume of 1.0 l. Sterilized filtration of the resulting solution is carried out and under aseptic conditions, it is bottled under nitrogen in ampoules with a capacity of 2 ml of 1 ml in an ampoule. Filled ampoules are sealed under nitrogen.

Example 3. The study of pancreatotropic activity of the described pharmaceutical composition. The study was conducted in experiments on rats. Dalargin and octreotide were used as reference preparations. Experimental acute pancreatitis (EOP) was modeled using a well-known technique by irrigating the pancreas (pancreas) with chloroethyl (cold model of EOP) [5].

In the experiment, the pharmaceutical composition of example 1 was used.

The cold model of the experimental image intensifier was reproduced in animals as follows. In rats under anesthesia, the abdominal cavity was opened under aseptic conditions, the stomach, duodenum (duodenum) and spleen with the pancreas located under them were removed to the wound. If you place the stomach and duodenum on the right, and the spleen to the left of the surgical incision, the main part of the pancreas will be in the wound. Within 1 minute, both surfaces of the pancreas were cooled with a stream of chloroethyl until a frost deposit appeared. The temperature of this segment of the gland decreased to -30 ° C. The temperature was measured in preliminary experiments using a special thermometer, which was introduced into the thickness of the pancreas before freezing. After subsequent rapid thawing of the pancreas, together with the stomach, duodenum and spleen, they were immersed in the abdominal cavity and the wound of the abdominal wall was sutured tightly through all layers with separate nodal nylon sutures. Aseptic dressings were not applied. Modeled image intensifier tubes of moderate severity.

The maximum severity of the disease develops after 24 hours from the start of the experiments. Therefore, the substances were administered 1 day (s) after the reproduction of the image intensifier, i.e. at the height of the pathological process.

To evaluate the therapeutic effect of the selected substances, 4 series of experiments were performed on white outbred male rats weighing 250-300 g: control group I (EOP without treatment), group II - EOP (treatment with dalargin), group III - EOP (treatment with octreotide), Group IV - EOP (treatment with a pharmaceutical composition). Pharmacological substances were administered intraperitoneally (ip). The volume of the injected / b solution of substances was 1 ml, the dose was 50 μg / kg for each. The drugs were administered 2 times / day for 3 days.

The criteria for evaluating the effectiveness of treatment were: macroscopic changes in the abdominal cavity, morphological changes in pancreatic tissue, biochemical parameters (activity of pancreatic enzymes and the content of lipid peroxidation products / lipid peroxidation in the blood). The listed studies in experimental animals were carried out after decapitation 1, 5, 10 and 15 days after the start of the experiment.

Materials for morphological studies were taken from three areas of the pancreas: I - zones of damage (cooling); II - zone directly bordering the lesion (perifocal); III - zone remote from the lesion (intact). For a light-optical study, pancreatic slices were fixed with 10% neutral formalin according to Lilly and embedded in paraffin. The preparations were stained with hematoxylin-eosin, according to Van Gieson, according to Romanovsky-Giemsa; fibrin and zymogen granules were detected by Gram-Weigert. In each observation, semi-thin sections with a thickness of 1 μm were made from electronic blocks, which were stained with a mixture of azur-P and methylene blue and studied under a light microscope.

In heparinized blood plasma of rats, the activity of pancreatic amylase (substrate EPS-G7) and pancreatic lipase was determined by colorimetric method using standard commercial kits manufactured by DiaSys (Germany). As is known, they are highly specific markers of pancreatic damage [6].

In EOP, an uncompensated increase in lipid peroxidation occurs, leading to a violation of membrane structures as the most sensitive to the action of reactive oxygen species, and an increase in the concentration of lipid peroxidation products is directly dependent on the severity of inflammatory changes in pancreatic tissue. The study of the content of lipid peroxidation products (concentration of malondialdehyde / MDA /) was determined by K. Yagi [7].

The data obtained were processed by methods of variation statistics.

Morphological study

Intact animals

Histologically, the pancreatic tissue of intact animals is represented by multiscale lobules, separated by loose small-cell tissue. Intralobular ducts contain a secret. There is moderate plethora of inter- and intralobular veins. The acini in the lobules have the usual structure with clear boundaries of the cell membranes of acinar cells (AK). Mature secretory granules are localized around the center of the acinous ducts. Asynchrony of accumulation of secretory granules from small to significant in the periostroval zone is noted. Gleams of centroacinous ducts are filled with a secret. The pancreas of intact animals preserves the usual lobular structure of the gland, a clear structure of acini and an asynchronous type of secretion accumulation.

Experimental acute pancreatitis

Histologically, pancreatic tissue 1 day after reproduction of the experimental OP is represented by dissected lobules of different sizes due to pronounced interlobular edema. Individual lobules or part of them with signs of necrosis and a violation of the usual structure of the acini. AK with indistinguishable boundaries, diffuse eosinophilia of the cytoplasm. In individual AKs, the nuclei of centroacinous duct cells and pyknotic nuclei of AK are distinguishable.

On semi-thin sections in the foci of acinar lesions, the contours of necrotic acini with indistinguishable AK borders and the absence of secretory granules in them, but diffuse microvesicular cytoplasmic transformation are visible. Among the necrotic acini, preserved inserted ducts with cystic extensions of their gaps are visible. Dystrophic changes in the form of vacuolization of the cytoplasm and dissociation of secretory granules to the basement membranes with possible release of enzymes in the interstitium are observed in areas bordering on necrosis in the AK cytoplasm. In the interacinar spaces there is inflammatory leukocyte infiltration with the migration of segmented white blood cells into the cytoplasm of damaged cells and focal hemorrhages. In separate segments, the contours of necrotic acini are completely erased, and segmented white blood cells diffusely infiltrate the damaged tissue of the lobules. Interlobular inflammatory infiltration is prevalent mainly around lobules with necrotic acini.

In the preserved acinar parenchyma in the interlobular adipose and connective tissue, there are signs of edema and minimal inflammatory infiltration. Interlobular arteries and veins are dilated and full-blooded. Interlobular ducts with an expanded lumen and without contents. The acini outside the lesion are characterized by a high degree of accumulation of secretory granules.

5 days after the reproduction of the image intensifier, pancreatic tissue is represented by large lobules, most of which have the usual structure of acini. In the preserved acinar parenchyma, small and larger pancreatic lobules are found, where along with the preserved acini there are tubular duct regenerates, and the interacinar spaces are made of loose multicellular connective tissue with the presence of inflammatory infiltrate cells in the form of lymphocytes and segmented white blood cells. The existing lobules or fragments of them are completely represented by tubular ductal regenerates with cystic expansion of their lumen. On semi-thin sections, an accumulation of small ducts separated by a layer of connective tissue is more clearly visible. The epithelial cells of the ducts do not contain secretory granules. Some of the newly formed ducts are included in the system of excretory ducts of the acini, while others exist in isolation.

In lobules with tubular regenerates, signs of interlobular and intralobular sclerosis are more pronounced in places with complete replacement of ductal regenerates with inflammatory tissue with inflammatory infiltration mainly from lymphocytes.

Outside of damaged pancreatic tissue, AK shows a high degree of accumulation of secretory granules with a typical apical orientation; in individual AKs, their dissociation to the basement membranes is visible.

Histologically, after 10 days, most of the pancreatic parenchyma is represented by large lobules, separated by narrow layers of small cell connective tissue. Along with this, small and larger atrophic lobes, represented by cystically altered tubular formations, disconnected layers of connective tissue, or completely replaced by connective tissue with barely distinguishable single ducts and islets of Langengars, are visible in the marginal sections of individual lobules and subcapsularly.

In semi-thin sections, the intact acinar parenchyma is represented by large acini with a high degree of accumulation of secretory granules in the apical sections of the AK cytoplasm.

On the 15th day of the EOP, histologically, pancreatic tissue is represented by small and large lobules, separated by loose small-cell connective tissue without signs of inflammation in it. In semi-thin sections, intact pancreatic lobules are represented by different sizes of acini with an apical orientation of secretory granules. Only subcapsularly and in perivascular and periductal spaces there are wide layers of connective tissue with signs of inflammation in the form of diffuse lymphoid and leukocyte infiltration. In the underlying omentum there is an infiltrate delimited from the pancreatic tissue by a fibrous capsule. The strands of fibrous tissue extending from the capsule dissociate the infiltrate and microabscesses in it. A feature of the fibrous capsule is the predominance of mature weakly vascularized connective tissue.

Thus, with EOP without treatment in the pancreas, necrosis of acinar tissue of lobules with reactive intralobular interstitial inflammation and suppuration developed in the early stages of observation, followed by later defective tissue regeneration. Features of incomplete coagulation necrosis of acinar tissue with necrobiotic and dystrophic changes in individual ductal and AK, on the one hand, determine the migration of leukocytes with suppuration of necrotic tissue, and on the other hand, defective replacement regeneration in the form of isolated tubular (ductal) structures with subsequent replacement sclerosis and complete delimitation from a viable acinar parenchyma. It should be noted that during all observation periods in AK outside the necrosis zones, a high degree of accumulation of secretory granules remained with a slowing down of secretion secretion into the duct system.

The continued high secretory activity in the AK with a slowdown in secretion excretion into the duct system outside the damage zones is an unfavorable background for pancreatic regeneration, creating a threat of pathological secretion in the interstitium.

Treatment of experimental acute pancreatitis with dalargin

On the 5th day, pancreatic tissue is represented by large lobules separated by loose multicellular connective tissue. Most of the pancreatic lobules preserves the usual structure of acini, others are represented by tubular regenerates, disconnected by connective tissue with marked inflammatory infiltration by macrophages, leukocytes and lymphocytes. Some tubular formations retain a tendency to form excretory ducts, while others undergo cystic expansion. The most pronounced inflammatory infiltration was noted in areas of atrophied lobules. In an unchanged acinar parenchyma, there are no signs of interlobular sclerosis and inflammatory infiltration.

On semithin sections of the preserved acinar parenchyma, the lobules are separated by a loose small-cell connective tissue with single fibroblasts and macrophages in it. Acini in lobules of different sizes with underlined contours of the basement membranes and with apical orientation of secretory granules around centroacinous ducts. A more pronounced degree of accumulation of secretory granules in the AK cytoplasm is observed only around the islets of Langerhans.

On the 10th day histologically, pancreatic tissue is represented by segments of different sizes, separated by fat or loose connective tissue. Only in certain sections of the lobules can one see layers of connective tissue in the form of small foci or cords with cystic-expanded tubular structures that are clearly separated from the intact acinar parenchyma and their scale is small.

When studying semi-thin sections of zones of atrophic lobules, single ducts with loose connective tissue without signs of inflammation are traced in them. In a viable acinar parenchyma, acini of various sizes with a small degree of accumulation of secretory granules.

On the 15th day, the pancreas has the usual lobular structure of small and large-scale lobules, separated by adipose tissue or small cell connective tissue. In the marginal departments of single lobules or in interlobular spaces, focal lymphoid infiltrates with cystic dilated ducts enclosed in them can be found.

When studying semi-thin sections in segments, there are no signs of interacinar edema and sclerosis. AK are characterized by a small degree of accumulation of secretory granules and are localized only around the centroacinous ducts.

Thus, treatment with dalargin gave a positive effect, which manifested itself in the repair of necrosis zones without suppuration in the form of ductal tubular regenerates with a clear delineation of them from the saved acinar parenchyma. Only at 10 days did single atrophic lobules with signs of intralobular sclerosis still persist.

For 15 days there were no signs of sclerosis and inflammation. At all observation periods, a low degree of AK secretory activity and the preservation of cell membrane contours were observed, which should be associated with the inhibitory effect of dalargin on pancreatic secretion activity and stabilization of cell membranes.

Treatment of experimental acute pancreatitis with octreotide

On day 5 of the image intensifier tube in the pancreatic tissue there were no signs of intralobular necrosis and interstitial inflammation. In the acinar tissue of the lobules, there were large-scale changes in AK in the form of large-vacuole dystrophy mainly in the basal parts of the cytoplasm. Moreover, dystrophic changes in AK were not accompanied by inflammatory infiltration. Along with this, dystrophic changes were absent in other animals and large lobules prevailed with an emphasized structure of acini in them and a low content of secretory granules in the AK cytoplasm. Single lobules or marginal sections of large lobules were represented by duct regenerates with proliferation of connective tissue between the tubular structures. Only in the connective tissue of these lobules and in the perivascular spaces was a mild lymphoid inflammatory infiltration observed.

Acinuses with apical orientation of secretory granules in the cytoplasm of AK predominated in semi-thin sections. In individual cells, secretory granules occupied all sections of the cytoplasm. Centroacinous ducts did not contain a secret.

Histologically, on day 10, pancreatic tissue is represented by segments of different sizes, separated by layers of loose small-cell connective tissue, with the presence of small white blood cells and lymphocytes in them. In the thickness of individual lobules, there were small-sized ductal regenerates with lymphoid inflammatory tissue infiltration. There were single lobules represented not only by duct regenerates with cystic expansion of tubular structures, but also with flattening of the epithelium in them. Such atrophic lobules were surrounded by loose connective tissue containing lymphocytes and macrophages.

The study of semi-thin sections of acinar tissue outside the lesion zones revealed a low degree of accumulation of secretory granules only in the apical sections of the AK cytoplasm and the abrasion of the interacinar borders.

On the 15th day of the disease, pancreatic acinar tissue is represented by large and small lobules, separated by loose connective tissue without signs of inflammation in it. The emphasized structure of the pancreatic capsule due to its thickening and moderate sclerosis is noted. Around individual interlobular ducts, signs of periductal sclerosis and chronic inflammation in the form of diffuse lymphoid infiltration are visible.

On semi-thin sections, a clear structure of acini in lobules with apical localization of secretory granules around centroacinous ducts and the presence of secretion in their lumen is visible. The degree of accumulation of secretory granules is higher than in previous periods of observation.

Thus, the treatment of EOP with octreotide according to morphological data confirms the positive effect on the course of the disease. At the same time, signs of necrosis and interstitial inflammation of acinar tissue did not progress. Necrotic areas of pancreatic tissue were regenerated by ductal transformation followed by sclerosis and delineation from a viable parenchyma. On the 15th day of the disease, the usual structure of the pancreas was completely restored while maintaining focal chronic interstitial inflammation. It should be noted a decrease in AK secretory activity up to the 10th day of the disease with a complete restoration of the asynchronous rhythm of activity by the 15th day.

Treatment of experimental acute pancreatitis with a pharmaceutical composition

On the 5th day of treatment with EOP, pancreatic tissue is represented by different-sized lobules, separated by a loose small-cell connective tissue, sometimes multicellular due to diffuse, moderately severe inflammatory infiltration by leukocytes, macrophages and lymphocytes. The degree of inflammatory infiltration is expressed around lobules, completely represented by duct proliferates or partially in the subcapsular regions, with the separation of viable lobules into small fragments. Beyond these changes, large and small pancreas lobes have clearly formed large acini, dilated and devoid of secretion intralobular ducts and full-blooded blood vessels. Individual ductal regenerates underwent cystic expansion and are surrounded by dense connective tissue with varying degrees of vascularization and chronic inflammation.

On semi-thin sections, acini with clear contours of cell membranes, decreased basophilia of the cytoplasm and small single mature secretory granules around centroacinous ducts, which indirectly indicates inhibition of the synthesis of pancreatic enzymes.

By 10 days, the pancreas histologically preserves the usual lobular structure with the presence of large and small lobules, separated by loose, sometimes multicellular connective tissue due to diffuse and large-focal inflammatory infiltration, represented by eosinophilic segmented leukocytes, macrophages and lymphocytes. The pancreatic capsule in some areas is sclerosed and infiltrated by white blood cells.

On semi-thin sections, the acini in the lobules have a clear structure, and the cytoplasm of the apical sections contains a few secretory granules. In some acini, the degree of accumulation of secretory granules is small, and in the periostroval zones it is higher and occupies all sections of the cytoplasm. Asynchronous accumulation of secretory granules in the cytoplasm of AK is characteristic of acinar tissue under normal functioning conditions.

On the 15th day of the tube, the pancreatic tissue retains its usual lobular structure without signs of interlobular and intralobular sclerosis. The lobules retain a clear structure of the structure of the acini with a pronounced basophilia of the cytoplasm and eosinophilia of the apical sections of the AK cytoplasm.

On the semi-thin sections, the sharp contours of the acini are preserved with the accumulation of mature secretory granules around the centroacinous ducts. Acinuses are found both with a small accumulation of granules, and higher in the periostrovial zone; gaps of individual centroacinous and intralobular ducts filled with secretion. In the pancreas, there are signs of asynchrony of the secretory cycle and secretion in the lumen of the ducts.

Based on the morphological study of the pancreas during EOP after treatment with the described pharmaceutical composition, signs of a positive effect of drug therapy on the course of the disease were revealed. At the same time, necrosis did not progress, there were no signs of suppuration of necrotic tissues, early regeneration of damaged tissues occurred with a tendency to distinguish them from an unchanged viable parenchyma. By the 15th day of the disease, there were no signs of sclerosis and interstitial inflammation in the pancreas with a complete restoration of the asynchronous rhythm of pancreatic enzyme secretion.

As follows from the above data, the prognosis of the disease in each treatment group is favorable with the full restoration of the acinar tissue of the pancreas without residual effects of sclerosis and inflammation by the 15th day after the reproduction of the image intensifier.

All drugs selected for treatment (dalargin, octreotide and a pharmaceutical composition based on them) have common morphological criteria for a positive effect on the course of the image intensifier tube: prevention of an increase in the incidence of necrosis and interstitial inflammation, defective regeneration of damaged acinar tissue through ductal transformation, followed by sclerosis and delimitation from intact tissue lobules with the restoration of its secretory activity.

A pronounced therapeutic effect despite the large-scale initial damage was exerted by dalargin. The intensive migration of leukocytes into necrotic tissues while maintaining blood circulation in the microcirculation system led to repair without suppuration and severe sclerosis. Under the influence of dalargin treatment, stabilization of AK cell membranes was already observed on the 10th day, and by the 15th day the asynchronous rhythm of acinar parenchyma activity was completely restored.

Octreotide in the treatment of EOP led to a decrease in the extent of necrosis, timely repair, but for a long time (up to 10 days) signs of focal chronic interstitial inflammation and intracellular edema persisted with a sharp decrease in secretory activity and only by the 15th day the function of the acinar parenchyma was completely restored.

Treatment of EOP with the claimed pharmaceutical composition revealed a more favorable effect on the course of EOP with minimal damage by 5-10 days and complete restoration of acinar parenchyma function by the 15th day without residual effects of sclerosis and inflammation. Therefore, it is possible to recommend the use of a pharmaceutical composition based on dalargin and octreotide to confirm the positive therapeutic effect in further studies.

Biochemical research

1 day after the reproduction of the image intensifier, the activity of pancreatic amylase increased 1.7 times (p <0.05) compared with that of intact animals (table 1).

After 5, 10, and 15 days after reproduction of the image intensifier, the activity of pancreatic amylase exceeded the indicated parameter by 1.5, 1.3, and 1.2 times, respectively (p <0.05). Dalargin, octreotide and their pharmaceutical composition approximately equally reduced amylase activity during the entire observation period, i.e. were about equally effective.

Table 1. INFLUENCE OF PREPARATIONS ON THE ACTIVITY OF PANCREATIC AMILASE (ME / L) IN RATS IN ACUTE EXPERIMENTAL PANCREATITIS (EOP) (M ± m) Rat group The term after the playback of the image intensifier, days 0 one 5 10 fifteen Intact animals 1148 ± 37 The control 1953 ± 85 ^* 1689 ± 28 ^* 1520 ± 17 ^* 1421 + 15 ^* Dalargin 1307 ± 13 ^# 1228 + 14 ^# 1027 ± 96 ^# Octreotide 1440 ± 21 ^# 1317 ± 24 ^# 1137 + 19 ^# Pharmaceutical composition dalargin + octreotide 1490 ± 19 ^# 1343 + 15 ^# 1245 + 32 ^# Note. Hereinafter in the tables: ^* or ^# - p <0.05 - the differences are significant compared with intact animals or with control, respectively.

Analysis of the dynamics of activity of another pancreatic enzyme - lipase - also showed significant changes in its activity during EOP (table 2). So, for example, 1 day after the reproduction of the image intensifier, the lipase activity exceeded that of intact animals 1.6 times (p <0.05), after 5, 10 and 15 days - 1.3, 1.2 and 1.1 times respectively (p <0.05). Dalargin and octreotide equally normalized this indicator. The pharmaceutical composition of dalargin with octreotide acted similarly. Moreover, after 15 days it was significantly (p <0.05) more effective than octreotide.

Table 2. INFLUENCE OF PREPARATIONS ON THE ACTIVITY OF PANCREATIC LIPASE (ME / L) IN RATS IN ACUTE EXPERIMENTAL PANCREATITIS (EOP) (M ± m) Rat group The term after the playback of the image intensifier, days 0 one 5 10 fifteen Intact animals 3.79 ± 0.06 The control 5.97 + 0.28 ^* 4.88 + 0.05 ^* 4.42 + 0.04 ^# 4.16 + 0.03 ^* Dalargin 4.16 + 0.06 ^# 3.98 + 0.03 ^# 3.72 + 0.02 ^# Octreotide 4.14 + 0.03 ^# 3.90 + 0.03 ^# 3.82 ± 0.04 ^# Pharmaceutical composition dalargin + octreotide 4.13 + 0.02 ^# 3.87 + 0.03 ^# 3.66 ± 0.03 ^# ° ° - p <0.05 - the differences are significant compared with octreotide.

A study of the content of lipid peroxidation products (level of malondialdehyde / MDA /) in peripheral blood (plasma) showed that after 1 day. after reproduction of the image intensifier, the concentration of MDA increased sharply and exceeded that of intact animals by 2.4 times (p <0.05) (table 3). After 5, 10, and 15 days after the reproduction of the image intensifier, the MDA content exceeded the indicated indicator by 1.9, 1.7, and 1.5 times, respectively (p <0.05). Dalargin and octreotide equally normalized this indicator. The pharmaceutical composition of dalargin with octreotide was more effective in this regard. So, for example, 10 days after the reproduction of OD, the MDA level after treatment with the pharmaceutical composition was significantly (p <0.05) lower than after a course of dalargin.

Table 3. INFLUENCE OF DRUGS ON THE CONTENT OF MALON DIALDEHYDE (nmol / ml) IN RATS IN ACUTE EXPERIMENTAL PANCREATITIS (EOP) (M ± m) Rat group The term after the playback of the image intensifier, days 0 one 5 10 fifteen Intact animals 5.91 ± 0.13 The control 14.1 ± 1.3 ^* 11.3 ± 0.4 ^* 9.83 ± 0.25 ^* 8.80 ± 0.06 ^* Dalargin 9.31 ± 0.47 ^# 8.60 ± 0.15 ^# 7.37 ± 0.18 ^# Octreotide 8.65 ± 0.16 ^# 7.41 ± 0.35 ^# 5.65 ± 0.22 ^# Pharmaceutical composition dalargin + octreotide 8.16 ± 0.47 ^# 7.13 ± 0.35 ^# ° 6.47 ± 0.44 ^# ° - p <0.05 - the differences are significant compared with dalargin.

Example 4. The therapeutic effect of the composition of octreotide and dalargin in rats with acute pancreatitis (OP) of moderate and severe severity.

The experiments were carried out on white non-linear male rats weighing 220-300 g, in which they reproduced the OP as described above in example 2. Modeled OP moderate and severe severity. Animals of the control groups did not receive treatment (they were injected systemically / ip / i / b / - isotonic sodium chloride solution / NaCl / for 3 days / 2 times / day /). Experimental animals, 24 hours after the creation of the OP, were administered ip for 3 days (2 times / day) octreotide or dalargin in different doses or their composition.

The results are presented in tables 4 and 5.

Table 4 Change in mortality in rats with severe acute pancreatitis under the influence of octreotide, dalargin and their combinations The drug (dose, mg / kg) The number of rats Mortality,% Control (isotonic NaCl solution) 60 48 Octreotide (0.1) 71 32 ^x Octreotide (0.2) twenty thirty Octreotide (0.3) twenty 35 Octreotide (0.4) 40 23 ^x Octreotide (0.5) 36 25 ^x Octreotide (0.6) 32 22 ^x Octreotide (0.7) 36 17 ^x Octreotide (0.8) 40 20 ^x Octreotide (0.9) 32 19 ^x Dalargin (0.1) 40 40 Dalargin (0.2) 32 41 Dalargin (0.3) 53 34 ^x Dalargin (0.4) 16 38 Dalargin (0.5) fifteen 40 Dalargin (0.6) 48 33 ^x Dalargin (0.7) 16 31 Dalargin (0.8) 24 25 ^x Dalargin (0.9) 26 23 ^x Dalargin (1.0) twenty 20 ^x Octreotide (0.1) + dalargin (0.1) 10 thirty Octreotide (0.2) + dalargin (0.2) 10 thirty Octreotide (0.3) + dalargin (0.3) 10 twenty Octreotide (0.4) + dalargin (0.4) 10 twenty Octreotide (0.5) + dalargin (0.5) 10 twenty Octreotide (0.6) + dalargin (0.6) 10 twenty Octreotide (0.7) + dalargin (0.7) 10 twenty Octreotide (0.8) + dalargin (0.8) 10 twenty Octreotide (0.2) + dalargin (0.1) 10 twenty Octreotide (0.3) + dalargin (0.1) 10 thirty Octreotide (0.4) + dalargin (0.1) 10 thirty Octreotide (0.5) + dalargin (0.1) 10 thirty Octreotide (0.6) + dalargin (0.1) 10 thirty Octreotide (0.7) + dalargin (0.1) 10 thirty Octreotide (0.8) + dalargin (0.1) 10 thirty Octreotide (0.9) + dalargin (0.1) 10 thirty Octreotide (0.3) + dalargin (0.2) 8 25 Octreotide (0.4) + dalargin (0.2) 8 37 Octreotide (0.5) + dalargin (0.2) 10 thirty Octreotide (0.6) + dalargin (0.2) 10 thirty Octreotide (0.7) + dalargin (0.2) 10 thirty Octreotide (0.8) + dalargin (0.2) 10 thirty Octreotide (0.9) + dalargin (0.2) 10 thirty Octreotide (0.1) + dalargin (0.3) 10 twenty Octreotide (0.2) + dalargin (0.3) 8 25 Octreotide (0.4) + dalargin (0.3) 8 25 Octreotide (0.5) + dalargin (0.3) 8 37 Octreotide (0.6) + dalargin (0.3) 10 thirty Octreotide (0.7) + dalargin (0.3) 10 thirty Octreotide (0.8) + dalargin (0.3) 10 thirty Octreotide (0.9) + dalargin (0.3) 10 thirty Octreotide (0.1) + dalargin (0.6) 10 10 ^x Octreotide (0.2) + dalargin (0.6) 10 10 ^x Octreotide (0.3) + dalargin (0.6) 10 10 ^x Octreotide (0.4) + dalargin (0.6) 10 twenty Octreotide (0.5) + dalargin (0.6) 10 twenty Octreotide (0.7) + dalargin (0.6) 10 thirty Octreotide (0.8) + dalargin (0.6) 10 thirty Octreotide (0.9) + dalargin (0.6) 10 thirty Octreotide (0.1) + dalargin (0.2) 10 twenty Octreotide (0.1) + dalargin (0.4) 10 twenty Octreotide (0.1) + dalargin (0.5) 10 twenty Octreotide (0.1) + dalargin (0.7) 10 10 ^x Octreotide (0.1) + dalargin (0.8) 24 4 ^{x *} ° Octreotide (0.1) + dalargin (0.9) twenty 0 ^x * ° Octreotide (0.1) + dalargin (1.0) 10 10 ^x Note. The differences are statistically significant (Fisher's exact method) - p <0.05: ^x - compared to control; ^* - compared with octreotide at a dose of 0.1 mg / kg; ° - compared with dalargin in doses of 0.8 and 0.9 mg / kg, respectively.

Table 5 The change in mortality in rats with moderate acute pancreatitis under the influence of octreotide, dalargin and their combinations (use of drugs for therapeutic purposes) The drug (dose, mg / kg) The number of rats Mortality,% Control (isotonic NaCl solution) 68 29th Octreotide (0.1) 60 17 ^x Dalargin (0.8) 60 15 ^x Dalargin (0.9) 60 13 ^x Octreotide (0.1) + dalargin (0.8) thirty 0 ^{x *} ° Octreotide (0.1) + dalargin (0.9) thirty 0 ^{x *} ° Note. The differences are statistically significant (Fisher's exact method) - p <0.05: ^x - compared to control; ^* - compared with octreotide at a dose of 0.1 mg / kg; ° - compared with dalargin in doses of 0.8 and 0.9 mg / kg, respectively.

From table 4 it can be seen that the mortality of control rats with severe severity of acute obstruction was 48%. Separately, octreotide in doses of 0.1, 0.4, 0.5, 0.6, 0.7, 0.8 and 0.9 mg / kg and dalargin in doses of 0.3, 0.6, 0.8 , 0.9 and 1.0 mg / kg significantly (p <0.05) reduced the lethality of animals to 17-34%. The composition of octreotide and dalargin significantly (p <0.05) reduced the mortality of rats (up to 0-10%) compared with the control in the following cases: octreotide (0.1, 0.2 and 0.3 mg / kg) + dalargin ( 0.6 mg / kg); octreotide (0.1 mg / kg) + dalargin (0.7, 0.8, 0.9 and 1.0 mg / kg). The combination of octreotide (0.1 mg / kg) + dalargin (0.8 mg / kg) (lethality was 4%) and octreotide (0.1 mg / kg) + dalargin (0.9 mg / kg) (mortality was 0), which significantly (p <0.05) gave a distinct, summarized effect (i.e., octreotide at a dose of 0.1 mg / kg and dalargin at doses of 0.8 and 0.9 mg / kg respectively exceeded the lethality of the composition). Therefore, there is a composition of octreotide (0.1 mg / kg) and dalargin (0.8 and 0.9 mg / kg), which in rats with severe severity of acute severity can cause a clear summation of effects and have a pronounced therapeutic effect, reducing the lethality of animals up to 0-4%.

Similar results were obtained in experiments on rats with OP of moderate severity (Table 5). Indeed, it was found that mortality in groups of rats treated separately with octreotide (0.1 mg / kg) or dalargin (0.8 and 0.9 mg / kg) was 13-17% (p <0.05; compared with control equal to 29%).

The combination of octreotide (0.1 mg / kg) + dalargin (0.8 mg / kg) and octreotide (0.1 mg / kg) + dalargin (0.9 mg / kg) significantly (p <0.05) gave a distinct the cumulative effect (in both cases, the mortality rate was 0), inferior in mortality to octreotide at a dose of 0.1 mg / kg and dalargin at doses of 0.8 and 0.9 mg / kg, respectively.

Thus, in rats with moderate to severe OP, the composition octreotide (0.1 mg / kg) and dalargin (0.8 and 0.9 mg / kg) gives a clear summation of the effects and has a pronounced therapeutic effect, reducing the lethality of animals to 0 -four%. Thus, as a result of the above studies, it was found that the described pharmaceutical composition has a high pancreatotropic activity and has a pronounced therapeutic effect in EOP.

LITERATURE

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Claims (5)

1. A pharmaceutical composition having pancreatotropic activity, characterized in that it contains octreotide and dalargin in therapeutically effective amounts as active components. 2. The pharmaceutical composition according to claim 1, characterized in that it is made in lyophilized form. 3. The pharmaceutical composition according to claim 1, characterized in that it further comprises a pH regulating substance and water. 4. The pharmaceutical composition according to claim 3, characterized in that as the pH of the regulatory substance, ammonium acetate is used in the following ratio of components, g: dalargin 0.5-5.0 octreotide 0.05-0.5 ammonium acetate 0.71 1 M acetic acid to pH 4.5-5.5 water for injection up to 1 5. The method of obtaining the pharmaceutical composition according to claim 1, characterized in that the active ingredients are dissolved in purified water and the resulting solution is lyophilized.