Application of alcohol extract of dry marsh celandine grass as hepatoprotective agent

Abstract

FIELD: pharmaceuticals.SUBSTANCE: use of an alcoholic extract of dried marsh stachys herb as a hepatoprotective agent. The use of an alcoholic extract of dried marsh celandine herb, obtained by extraction with 60% ethyl alcohol 1:10 at room temperature, dried using a vacuum rotary evaporator and a drying cabinet, as a hepatoprotective agent that reduces inflammation, the risk of fibrosis, used in conditions accompanied by metabolic disorders, such as fatty hepatosis.EFFECT: above-described application expands the spectrum of hepatoprotective agents, is effective as a hepatoprotective agent that reduces inflammation, the risk of fibrosis, and is used to treat fatty hepatosis.1 cl, 4 dwg

Description

[0001] The invention relates to the field of pharmaceuticals and concerns the use of an alcoholic extract of dry marsh stachys herb as a hepatoprotective agent.

[0002] The closest technical solution to the proposed one is the use of the drug Karsil©, which contains silymarin, a lipophilic compound, the active substance of the hepatoprotective plant milk thistle Silibium marianum L. (Asteraceae family) [1].

[0003] The disadvantage of Karsil, as a prototype, is the adverse reaction that occurs during use: the phenomenon of cholestasis.

[0004] The technical objective of the proposed solution is to expand the range of hepatoprotective agents and use an alcohol-water extract of dried marsh stachys herb in order to reduce the risk of developing cholestasis, improve metabolic processes in the liver, reduce inflammation and the risk of developing fibrosis.

[0005] For this purpose, it is proposed to use an alcoholic extract of dried marsh celandine herb as a hepatoprotective agent that reduces the risk of developing cholestasis, inflammation and fibrosis, used in conditions accompanied by metabolic disorders such as hypercholesterolemia, hyperbilirubinemia, and fatty hepatosis.

[0006] The extract was obtained from the herb of the marsh stachys (Stickys palustris L.) (family Lamiaceae), collected in the second half of July 2024 on the banks of the Babka River near the village of Zaikovo in Perm Krai. The raw material was dried in a well-ventilated area at room temperature, protected from direct sunlight, and stored separately from other raw materials.

[0007] To extract from the herb of marsh stachys the sum of phenolic compounds potentially possessing hepatoprotective activity in the form of aglycones and glycosides, ethyl alcohol of medium concentration, namely 60%, was used [2].

[0008] 30.0 grams of raw material, crushed to a particle size passing through a 1 mm sieve, was placed in a 500 ml flask and filled with 300 ml (1:10) of 60% ethyl alcohol. The flask was then sealed with a rubber stopper and left to infuse for 24 hours at room temperature (an alcohol extract was obtained using the "cold" method). The raw material was then filtered through a paper filter using a Bunsen flask, a Buchner funnel, and a vacuum water-jet pump. The extractant was distilled from the resulting filtrate using a vacuum rotary evaporator until a thick consistency was achieved. The mixture was placed in an evaporating bowl and dried in a drying oven at 60°C. The dry extract was removed from the evaporating bowl with a glass rod, and the extract was ground to a powder.

[0009] The extract is a dark-green amorphous powder with a characteristic faint odor. It is moderately soluble in distilled water and readily soluble in 70% ethyl alcohol. It is very slightly soluble in chloroform, diethyl ether, and ethyl acetate.

[0010] Most liver diseases are associated with alterations in the liver's detoxifying and synthesizing functions. As an organ closely linked to the concentration of neurotransmitters in the blood, the liver alters the rate of metabolic processes. Thus, against the background of the development of hepatic encephalopathy, which occurs with fatty hepatosis, disturbances in orientation and exploratory reactions and behavior occur. In addition to clinical manifestations, a process of fatty transformation of hepatocytes, cytolysis, and subsequent inflammation occurs, which then leads to the development of replacement connective tissue – fibrosis [3].

[0011] The studies were conducted on a model of fatty hepatosis induced by the action of carbon tetrachloride (hereinafter referred to as CTC), which is a strong hepatotropic toxin [4]. Modeling of fatty hepatosis was carried out on white Wistar rats, which were kept on a standard complete feeding of dry food and had free access to water. Animals (a total of 120) were divided according to the pair-analogue method into 4 groups: Group 1 - intact (30 rats), Group 2 - received Carsil© (30 rats) and carbon tetrachloride (CTC) at a dose of 40 mg / kg of animal weight, Group 3 - received an alcoholic extract of dry marsh celandine herb at a dose of 15 mg / kg (30 rats) and CTC at a dose of 40 mg / kg of animal weight, Group 4 (30 rats) - received only CTC at a dose of 40 mg / kg. Rats in each group, except for intact rats, were administered carbon tetrachloride at a dose of 40 mg / kg in a 1:1 ratio of Vaseline oil. Karsil and an alcoholic extract of dried marsh celandine were administered for 14 days.For two weeks, the animals received Karsil at a dose of 3.15 mg / kg in terms of silibinin (the daily dose was calculated in accordance with the instructions for use) [5].

[0012] The technical result obtained by implementing the present invention consists of expanding the arsenal of plant-based hepatoprotectors. Unlike the prototype (Carsil), the ethanol-water extract of dried marsh celandine herb more effectively restores metabolic disorders induced by carbon tetrachloride, including cholestasis, inflammation, and fibrosis. It is known that the development of cholestasis significantly exacerbates metabolic disorders, and the detergent action of bile components increases hepatocyte necrosis, induces inflammation, and, consequently, promotes the development of fibrosis. The technical result of the proposed solution is an expansion of the spectrum of hepatotropic agents, increased effectiveness in the prevention of conditions accompanied by metabolic disorders (fatty hepatosis, hepatitis, liver fibrosis), and expansion of the functional capabilities of existing hepatoprotectors.

[0013] Advantages of the alcoholic extract of dried marsh celandine herb: it has a broad spectrum of hepatoprotective action without increasing the volume of secreted bile, which reduces the risk of cholestasis, has anti-inflammatory activity, helps reduce the processes of cytolysis caused by pro-inflammatory cytokines, prevents the development of an inflammatory reaction, and, thereby, prevents the growth of connective tissue - fibrosis.

[0014] Liver samples from rats treated with TCM (Group 4) revealed multiple changes consistent with the development of fatty hepatosis. The lobular and trabecular structure of the liver is disrupted. Hepatic trabeculae are 2-3 hepatocytes wide. Sinusoids are poorly perfused, containing isolated mononuclear cells. Portal tracts are equally spaced, fibrotic, and contain arterioles, venules, and bile capillaries of typical structure. Disse spaces are significantly dilated, and sinusoidal capillaries are unevenly plethoraic. Central venules are plethoraic. The liver exhibits a histological picture of chronic steatohepatitis with pronounced infiltration of the portal tracts and lobules by lymphocytes and leukocytes, hepatocyte necrosis, and disruption of the lobular histoarchitecture of the organs due to incomplete bridging portoportal fibrosis. The degree of leukocyte-infiltrating inflammatory reaction is pronounced.Hepatocytes with signs of pronounced large-droplet fatty degeneration (lesion volume ~90%, steatosis index - 4). Histological section of rat liver (x10) treated with carbon tetrachloride, stained with hematoxylin and eosin, is shown in Fig. 1.

[0015] Histological examination of the liver of rats treated with TCM and an ethanol-water extract of dried marsh stachys (Group 3) revealed preserved lobular and trabecular structures. Hepatic trabeculae are 1-2 hepatocytes wide. Hepatocytes are normal-sized with euchromic nuclei, with microvesicular fatty degeneration in the central regions. Portal tracts are equally spaced, non-fibrotic, and contain arterioles, venules, and bile capillaries of typical structure. Single mononuclear cells are present in the portal tracts. Disse spaces are dilated, and sinusoidal capillaries are unevenly plethoraic. Central venules are plethoraic. There is no pronounced leukocyte-infiltrating inflammatory reaction. There are no signs of connective tissue proliferation. Individual hepatocytes on the periphery with signs of large-drop fatty degeneration (lesion volume ~ 50%), steatosis index - 2).Histological section of the liver of rats (x10) treated with carbon tetrachloride + alcohol-water extract of dry marsh stachys herb, stained with hematoxylin and eosin, is shown in Fig. 2.

[0016] The histological picture of the liver of rats treated with Carsil and carbon tetrachloride (group 2) shows changes characteristic of the development of steatosis, inflammation and fibrosis. The lobular and trabecular structure of the liver is partially preserved. Hepatic trabeculae are 1-2 hepatocytes wide. Sinusoids are weakly filled with blood, containing single mononuclear cells. Blood vessels of the portal tracts are moderately filled with blood. Central venules are full-blooded. A pronounced leukocyte-infiltrating inflammatory reaction is noted. Portal tracts are contiguous, partially fibrotic: with focal portal and periportal fibrosis, with the formation of incomplete portoportal and portocentral septa. Hepatocytes have signs of severe large-droplet fatty degeneration (lesion volume ~ 80%, steatosis index - 3). Histological section of liver of rats (x10) treated with carbon tetrachloride + Carsil, stained with hematoxylin and eosin, is shown in Fig. 3.

[0017] Histological examination of the organs of rats from the intact group revealed no changes. Liver - architecture is preserved. Portal tracts are unevenly filled with blood, with weak lymphocytic infiltration. Bile ducts are unchanged. Border plate is preserved. Lobules have a typical beam structure. Sinusoidal capillaries with weak activation of Kupffer cells. Disse spaces are dilated. Hepatocytes are not enlarged, in a state of microvesicular steatosis mainly in the central zone of the lobules. Signs of fibrosis are not detected. Steatosis index - 0 (less than 5% of hepatocytes in the histological section are in a state of fatty degeneration). Histological section (x10) of the liver of intact rats, stained with hematoxylin and eosin, is shown in Fig. 4.

[0018] As a result of its use, it was discovered that Karsil is capable of exerting a hepatoprotective effect. However, it is unable to protect against hepatocyte degeneration and causes cholestasis and inflammation: the presence of infiltrating-inflammatory changes in sections, as well as signs of fibrosis, are observed. In some cases of drug-induced liver injury, cholestasis, inflammation, and the development of fibrosis significantly increase the risk of death in patients [6].

[0019] An alcoholic extract of dried marsh stachys herb exhibits a hepatoprotective effect, which is associated with the restoration of liver metabolism and a reduced risk of inflammation and fibrosis. Cholestasis has not been observed with this extract. An alcoholic extract of dried marsh stachys exhibits an anti-inflammatory effect, which also helps restore the structural integrity of hepatocytes. Use of the extract prevents the development of portal fibrosis of hepatocytes.

[0020] An alcoholic extract of dried marsh celandine herb stabilizes the hepatocyte cell membrane, prevents cytolysis, and has anti-inflammatory and antifibrotic effects. A drawback of the Karsil prototype: a deficiency in metabolic factors preventing the development of hepatosis.

[0021] Literature:

[0022] 1. Kurkin V.A. Milk thistle - a source of medicines (review) / / Chemical and pharmaceutical journal. - 2003. - V. 37. - No. 4. - P. 27-41.

[0023] 2. Kurkina A.V. Current aspects of standardization of medicinal plant materials containing flavonoids / / Bulletin of Siberian Medicine. - 2011. - Issue. 10(5). - P. 150-154. https: / / doi.org / 10.20538 / 1682-0363-2011-5-150-154.

[0024] 3. Bueverov, A. O. "Fatty liver": causes and consequences / A. O. Bueverov / / Consilium Medicum. - 2007. - V. 9, No. 9. - P. 74-78.

[0025] 4. Saratikov, A.S. Efficiency of enzyme-inducing agents in experimental liver damage with carbon tetrachloride / A.S. Saratikov, T.P. Novozheeva, A.I. Vengerovsky / / Experimental and clinical pharmacology. - 2003. - V. 66, No. 4. - P. 47-49.

[0026] 5. Instructions for the drug Carsil reg. No.: P N014839 / 01 dated 04 / 09 / 08 / Electronic resource: https: / / www.vidal.ru / drugs / carsil_2997 (date of access 07 / 11 / 2025).

[0027] 6. Pimanov S.I., Makarenko E.V. Idiosyncratic drug-induced liver damage: diagnosis and treatment / / Medical Council. 2017. No. 5. pp. 100-107. URL: https: / / cyberleninka.ru / article / n / idiosinkrazicheskie-lekarstvennye-porazheniya-pecheni-diagnostika-i-lechenie (date of access: 11.07.2025).

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