RU2410113C2 - Medication possessing antioxidant and pancreoprotecting activity - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: as medication applied is dry extract from vegetable raw material. Vegetable raw material for extraction contains herb of three-lobe beggarticks; herb of marsh cudweed; marigold flowers; rhizome of horse-heal; shoots of shrubby cinquefoil; herb of St. John's wort; shoots of common bilberry. Vegetable raw material is obtained by triple extraction at temperature 18-20°C, with further combination of extracts, filtration, evaporation, purification by separation and drying in vacuum drier with further fining, with final product containing not less than 12% of tanning substances.

EFFECT: medication extends arsenal of medications, which possess anti-oxidant and pancreoprotecting activity.

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Description

The invention relates to the field of pharmacy and relates to the production of agents with antioxidant and pancreatic activity.

In the general structure of diseases of the digestive system, one of the leading places is occupied by diseases of the pancreas, in particular, acute and chronic pancreatitis. In Russia, the prevalence of chronic pancreatitis in the population is from 0.20 to 0.68%. In Buryatia, the incidence of chronic pancreatitis in 2008 was: among the adult population, 223.4 cases per 100 thousand people; in adolescents, 140.9 cases per 100 thousand people; in children, 75.0 cases per 100 thousand people.

Pancreatitis, acute and chronic, are among the most common diseases of the gastrointestinal tract (GIT). At the same time, 55.5-70.0% of patients with acute pancreatitis are people of working age. Often, the inflammatory process in the pancreas leads to the development of diabetes mellitus, the formation of pancreatic cysts, and upon transition to the chronic process, secondary development of pancreatic cancer is noted in 12.5% of cases [2, 4, 5].

To date, in the treatment of this pathology, along with basic therapy, herbal medicines are widely used both as supplementary therapy during an exacerbation of the disease, and as therapeutic and prophylactic agents at the initial stages of the disease and at the stage of anti-relapse therapy [10] . In this regard, the search for promising plants and the development on their basis of effective multicomponent herbal remedies for the prevention and treatment of pancreatic diseases are an urgent problem of modern pharmacology and gastroenterology.

Known plant collection No. 120 recommended by S. Ya. Sokolov for the treatment of pancreatitis [10]. The drug is used in the form of a water decoction prepared according to the State Pharmacopoeia of the USSR XI edition (1990) [1]. The collection contains five medicinal plants (wt. Parts): alder buckthorn (bark) - 30.0; peppermint (leaf) 20.0; dioica nettle (leaf) 30.0, marsh calf (rhizome) 10.0; Valerian officinalis (rhizome) 10.0.

Decoction from medicinal plant materials does not provide the maximum yield of biologically active substances (BAS) in the extracts and usually amounts to 20-40% of the contained BAS in the raw materials. When preparing a decoction, it is difficult to assess the degree of release of active substances with each receipt of the decoction. The disadvantage of this method is also the lack of pharmacological activity.

The authors have developed a complex herbal remedy in the form of a dry extract with a pronounced antioxidant and pancreatic effect, consisting of seven medicinal plants.

The technical result of the invention is the expansion of the assortment of herbal medicines with pancreas and antioxidant activity through the use of widespread plant materials that have a reliable and secure raw material base, increasing the pharmacological activity due to the multicomponent nature of the drug.

The technical result is achieved by the fact that the claimed medicinal product includes a dried shredded vegetable mixture of the following composition (wt. Parts):

three-part series (grass) 10 marshmallows (grass) 10 calendula officinalis (flowers) twenty elecampane high (rhizome) fifteen shrubbery five leaves (shoots) 10 Hypericum perforatum (grass) 5 blueberries (shoots) thirty

The composition obtained by the authors is extracted three times in the ratio of raw materials: extractant 1: (10). The extracts are combined, filtered and evaporated to 1/15 of the original volume. The concentrated extract is purified by separation, dried in a vacuum dryer and ground. The yield of the finished product is 28% by weight of plant materials. The proposed method allows to obtain a product, which is a finely divided amorphous hygroscopic powder with a specific odor, with a content of tannins of at least 12%. The product obtained by the authors is characterized as a tool with a pronounced antioxidant, pancreatic activity. Using a herbal remedy in the form of a dry extract will allow you to get a drug with a certain content of active substances.

The authors studied the dynamics of the extraction of tannins and extractive substances from a plant composition, depending on technological factors. A quantitative assessment was conducted on the yield of tannins and extractive substances [1].

To select the concentration of the extractant, the processes of extraction of the total collection were studied. In the study of the type of extractant tested water, 20; thirty; fifty; 70; 96.6% ethyl alcohol. The results are presented in table 1.

Table 1 The yield of tannins and extractives from the plant composition, depending on the type of extractant N Type of extractant The content of tannins,% The content of extractives,% one Purified water 5.40 25.8 Ethanol, % 2 twenty 8.32 28.54 3 thirty 9.05 28.12 four fifty 10.34 27.03 5 70 10.22 26.75 6 96.6 3.86 9.41

Tabular data show that the most optimal extractant, providing the maximum yield of tannins, is 50% and 70% ethyl alcohol. The highest yield of extractives is provided by low concentrations of ethyl alcohol (20%).

The optimal ratio of raw materials and extractant, which corresponds to 1:10. A further increase in the ratio is impractical due to a slight increase in the yield of active substances, as well as in connection with an increase in the consumption of extractant. The results are presented in table 2.

table 2 The yield of tannins and extractives, depending on the ratio of raw materials: extractant Raw Material: Extractant Ratio The output of the amount of tannins,% The output of the sum of extractive substances,% 1: 5 7.8 23.67 1: 8 8.3 25.11 1:10 8.9 26.55 1:12 9.0 26.58 1:15 8.8 26.60

The most optimal grinding of the plant composition is the degree of grinding from 1-3 mm (table 3).

Table 3 The effect of the degree of grinding of the components of the drug on the yield of tannins and extractives The degree of grinding fractions of the collection Tannins, % Extractive substances,% 0.5-1 mm 10.03 37.06 1-3 mm 9.50 37.03 3-6 mm 9.21 35.45 6-10 mm 8.50 35.09

For a more complete depletion of raw materials, the authors propose using triple extraction. It is proposed to use 70% ethanol as the extractant in the first extraction, it is proposed to use hot water in the second and third extraction, this will eliminate the stage of alcohol regeneration, and also low concentrations of ethyl alcohol allow extraction of more extractive substances (Table 1).

The proposed method allows to obtain a dry extract containing at least 12% tannins. In pharmacological tests on experimental models in laboratory animals, pronounced pancreatic and antioxidant activity of the obtained dry extract was established.

An example of a specific implementation:

Shredded and sifted grass of a tripartite series - 10.0, marshmallow dried - 10.0, St. John's wort perforated - 5.0; Calendula officinalis flowers - 20.0; rhizome of elecampane high - 15.0; shoots of shrubs 10.0 shrubs and blueberries 30.0 are loaded into the extractor, 1000.0 ml of 70% ethyl alcohol are fed there, extraction is carried out at room temperature. The ratio of raw material and extractant is 1:10. Extraction is carried out during the day, periodically mixing.

At the end of the extraction, the first water-ethanol extract is obtained in an amount of 560.0 ml (density 0.905 g / cm ³ ). The meal is extracted two more times, each time supplying hot water to the extractor in an amount equal to the volume of the drained extract. 2nd discharge - 470.0 ml (density 0.982 g / cm ³ ), 3rd discharge - 490.0 ml (density 0.987 g / cm ³ ). The first, second and third extracts are sequentially, as they are obtained, filtered through sulfuric cloth into a collector and evaporated to 1/15 of the original volume. The concentrated extract is purified by separation, dried in a vacuum dryer for 12 hours and ground in a mill. Obtain 28.0 g of the finished product, which is 28% of the weight of the original plant harvest. The dry extract is a fine brown powder with a specific odor. Hygroscopic, slightly crumpled. The moisture content is 0.95%.

Determination of antioxidant and pancreoprotective activity of the complex herbal remedy "Pancreofit"

The experiments were conducted on 156 white Wistar rats of both sexes with an initial weight of 170-200 g contained in standard vivarium conditions. Acute pancreatitis in laboratory animals was caused by the method of P.S.Simavoryan et al. by irrigation of rat pancreas with chloroethyl under thiopental anesthesia (50 mg / kg) [9]. The investigated dry extract at a dose of 300 mg / kg was administered intragastrically to the operated animals from the first day of the experiment for 21 days. As a comparison drug, plant collection No. 120, recommended by S. Ya. Sokolov for the treatment of pancreatitis, was used [10]. Comparison preparation in the form of a decoction prepared according to the GF USSR XI edition (1990), was administered to animals in a volume of 1 ml / 100 g of mass according to a similar scheme. Animals of the control group received purified water in an equivalent amount according to a similar scheme. In animals of the intact group, experimental pancreatitis was not reproduced. Studies were performed on the 3rd, 7th, 14th and 21st day of the experiment.

To evaluate the antioxidant activity of the extract in the blood serum of animals, the concentration of malondialdehyde (MDA) [11] and the level of catalase activity [3] were determined.

The activity of pancreatic enzymes - α-amylase and lipase in blood serum was determined by the enzymatic colorimetric method using standard reagent kits. Peripheral blood counts were determined on an Abacus automated hematology analyzer with a standard set of reagents (Austria). For pathomorphological studies, the rat pancreas was fixed in 10% neutral formalin, sections of the organ were stained with hematoxylin-eosin [6]. On histological preparations using the computer program "Motic Images 2000" determined the area of necrotic sites, areas of inflammatory reaction and hemorrhage. The significance of differences between these parameters among animals of the control and experimental groups was evaluated using the non-parametric Mann-Whitney test [8]. Differences were considered significant at P≤0.05.

The research results showed that cooling of the pancreas of rats with chloroethyl caused a dynamic increase in the content of MDA in blood serum in animals of the control group. On the 14th day of observation, the concentration of MDA in serum in control rats reached a maximum value (Table 4). LPO activation was correlated with a decrease in the activity of a key antioxidant enzyme, catalase. So, on the 14th day of the experiment, the activity of catalase in animals of the control group decreased by 3 times compared with that in animals of the intact group.

The course administration of the studied extract to white rats against the background of chloroethyl pancreatitis inhibited LPO, causing a decrease in the concentration of MDA in blood serum by an average of 28% over the entire duration of the experiment compared with indices in animals of the control group. The most pronounced decrease in the content of MDA in blood serum in rats of the experimental group was observed on day 21 and amounted to 38% of that in animals of the control group. In rats treated with the studied extract and collection No. 120, against the background of a decrease in serum lipid peroxidation, catalase activation was observed: on day 3 - by 18 and 15%, on 7 - by 37 and 32%, on 14 - by 40 and 23 % and on day 21 - by 55 and 39%, respectively, compared with that in animals of the control group.

Thus, the antioxidant and pancreatic protection in acute chloroethyl pancreatitis has a more pronounced antioxidant activity compared to the decoction of collection No. 120.

The results of the studies showed that pancreatic irrigation with chloroethyl causes the development of acute pancreatitis, as evidenced by an increase in serum α-amylase activity in animals of the control group on day 3 of the experiment compared with data in animals of the intact group (Table 5). In the subsequent periods of observations, the activity of this enzyme in animals of the control group continued to increase dynamically and reached its maximum value by 14 days, exceeding that in animals of the intact group by 2.5 times. The serum lipase activity in animals of the control group increased simultaneously with hyperamylasemia and exceeded that in animals of the intact group on the 14th day of observation by 2.4 times (Table 5), which is also an important diagnostic sign of the development of acute pancreatitis. The course administration of the studied extract had a pronounced pancreoprotective effect, as evidenced by less pronounced hyperamylasemia and hyperlipasemia (Table 5) at all stages of the study compared with data in animals of the control group.

In animals of the control group, anemia was observed at all times of the experiment (Table 6). The severity of the inflammatory process was evidenced by an increase in ESR and the number of leukocytes compared with those in animals of the intact group (Table 6). Against the background of the introduction of antioxidant and pancreoprotective agents in animals, less pronounced anemia is observed compared with data in rats of the control group. A decrease in ESR indicated less pronounced inflammatory processes when using the studied drugs. In animals treated with the studied extract, less pronounced leukocytosis and restoration of the total number of leukocytes on day 21 were observed in the dynamics of the study compared with the data in animals of the control group. Analysis of the leukogram indicates a normalization on the 21st day of the distribution of the formed elements of white blood in the animals of the experimental groups (Table 6).

The results of a pathomorphological study showed that on the 3rd and 7th day of the experiment in the pancreas of the animals of the control group, pronounced interlobular and intralobular edema of the interstitium was observed. Alternative processes were detected on the periphery of the organ - in the areas of irrigation of the gland with chloroethyl and spread deep into the parenchyma, accompanied by intense leukocyte infiltration. On the 14th day of the study, significant areas of hemorrhage and parenchyma area underwent necrosis were observed in animals of the control group. Structural changes in the pancreas of animals receiving the studied extract and collection No. 120 on the 3rd and 7th day of the experiment did not significantly differ from those in the animals of the control group: intralobular and interlobular edema of interstitium, inflammatory infiltration, subcapsular necrosis. On the 14th day of the experiment, in animals receiving the studied extract and collection broth No. 120, the areas of necrosis decreased by 33 and 29%, inflammatory infiltration - by 27 and 24%, and hemorrhages - by 28 and 40%, respectively, compared with those in animals of the control group (table 7). Outside the zone of damage to the pancreatic tissue, correctly formulated acini with a large number of secretory granules in the apical and subnuclear sections of the cytoplasm were observed in the lobules. On the 21st day of observation, against the background of the course administration of an antioxidant, pancreatic protection and comparison drug, the areas of necrosis decreased by 30 and 24%, respectively, the areas of inflammatory infiltration - by 1.7 times and the hemorrhage - by 2.2 and 1.7 times, respectively, compared with those in animals of the control group (table 7).

Table 7 The effect of antioxidant, pancreatic protection and comparison drug on the morphometric parameters of the pancreas in acute pancreatitis in white rats Groups of animals Indicators S necrosis, um ² S inflammatory infiltration, um ² S hemorrhage, um ² one 2 3 four 3 days The control 273.1 ± 11.5 310.1 ± 9.9 67.3 ± 2.4 Antioxidant, pancreatic protection 215.4 ± 14.1 * 264.5 ± 10.3 * 64.6 ± 4.2 Etc. comparisons 223.1 ± 13.7 * 286.7 ± 12.6 * 63.2 ± 3.5 7 days The control 311.2 ± 21.3 298.3 ± 16.3 98.3 ± 7.6 antioxidant, pancreatic protection 232.9 ± 14.7 * 246.4 ± 14.7 * 76.4 ± 3.5 * Etc. comparisons 239.4 ± 12.6 * 256.5 ± 19.1 78.3 ± 4.9 * 14 days The control 231.6 ± 18.9 201.5 ± 16.7 72.3 ± 6.3 Antioxidant, pancreatic protection 154.3 ± 10.8 * 147.7 ± 9.7 * 51.7 ± 2.2 * Etc. comparisons 163.2 ± 15.7 * 152.9 ± 10.4 * 43.4 ± 4.1 * 21 days The control 147.0 ± 9.8 132.6 ± 14.8 32.7 ± 2.9 Antioxidant, pancreatic protection 103.3 ± 4.1 * 69.2 ± 4.9 * 14.9 ± 1.1 * Etc. comparisons 112.1 ± 6.3 * 78.1 ± 6.7 * 19.8 ± 1.6 *

Thus, the claimed complex herbal remedy has a pronounced pancreatic protective effect, reducing the activity of pancreatic enzymes, limiting the area of acinar tissue necrosis, the intensity of the inflammatory reaction and reducing the hemorrhage zone against the background of acute chloroethyl pancreatitis. In general, the claimed agent is not inferior, but surpasses in certain indicators (less pronounced hyperamylasemia, hyperlipasemia, leukocytosis and neutrophilia, restoration of the number of lymphocytes) to collection No. 120. Based on the data obtained, it is possible to recommend the use of the claimed antioxidant and pancreatic protection in the complex treatment and prevention of acute pancreatitis.

Information sources

1. The State Pharmacopoeia of the USSR: issue 1. General methods of analysis. 11th ed. - M.:, 1987. - 336 p.

2. Hubergrits N.B., Khristich T.N. Clinical pancreatology. - Donetsk, 2000 .-- 416 p.

3. Korolyuk M.A., Ivanova L.I., Mayrova I.G., Tokarev V.E. Methods for determining the activity of catalase / Korolyuk M.A., Ivanova L.I., Mayrova I.G., Tokarev V.E. // Lab. A business. - 1988. - No. 1. - S.16-19.

4. Mayev I.V. Hereditary pancreatitis // Ross. journal gastroenterol., hepatol., coloproctol. - 2004. - No. 1. - S.20-25.

5. Mayev I.V., Kucheryavy Yu.A. Acute and chronic pancreatitis, pancreatic cancer - a chain of sequential events or an independent disease? // Clinical medicine. - 2005. - No. 2. - S.12-16.

6. Microscopic technology: Manual / Ed. D.S. Sarkisova and Yu.L. Perova. - M.: Medicine, 1996 .-- 544 p.

7. Morozov S.V., Dolgikh V.T., Poluektov V.L. Markers of endotoxemia and the antioxidant defense system in experimental pancreatic necrosis // Vestnik VolGMU. - 2006. - No. 1. S.74-77.

8. Sergienko V.I., Bondarenko I.B., Gaydyshev I.I. Mathematical statistics in clinical studies. - M., 2001 .-- 256 s.

9. Simavoryan P.S., Tishenina R.S. Fat carbohydrate metabolism in experimental pancreatitis // Pathological physiology and experimental therapy. - 1973. - No. 2. - S. 59-62.

10. Sokolov S.Ya. Herbal Medicine and Phytopharmacology: A Guide for Physicians. - M., 2000 .-- 976 p.

11. Osakava T., Matshushita S. Coloring conditions of thiobarbituric acid test foe detection lipid peroxides. / Osakava T., Matshushita S. // Lipids. - 1980. - Vol.15. - Number 3. - P.137-140.

Claims (1)

A medicine with antioxidant and pancreoprotective activity, which is a dry extract from plant materials, characterized in that the plant materials for extraction contain a three-part sequence (grass); marshmallow cinnamon (grass); Calendula officinalis (flowers); elecampane high (rhizome); five-leaved shrubbery (shoots); St. John's wort perforated (grass); common blueberries (shoots) with a degree of grinding of 1-3 mm and the following ratio of components, parts by weight:
three-part sequence (grass) 10 marshmallows (grass) 10 Calendula officinalis (flowers) twenty elecampane high (rhizome) fifteen shrub quintuple (shoots) 10 St. John's wort perforated (grass) 5 common blueberries (shoots) thirty
the extract was obtained at a temperature of 18-20 ° C with three extraction, the first extraction with 70% ethyl alcohol, the second and third extraction with hot water in the ratio of raw material and extractant 1:10 for 24 hours each, then the extracts are combined, filtered , evaporated, purified by separation and dried in a vacuum dryer, followed by grinding, while the final product contains at least 12% tannins.