Traditional Chinese medicine compound composition for preventing and treating liver injury as well as preparation method and application of traditional Chinese medicine compound composition

Abstract

The invention discloses a traditional Chinese medicine compound composition for preventing and treating liver injury as well as a preparation method and application thereof. The compound composition comprises verbena, alfalfa, Chinese ladiestresses root, hairyvein agrimony, fingered citron, malt and hyperforin perforatum. The traditional Chinese medicine composition is simple in composition and reasonable in proportion, and animal experiments prove that the traditional Chinese medicine composition can improve the liver function, relieve liver inflammation injury, inhibit liver cell necrosis and fatty degeneration, inhibit oxidative stress and resist lipid peroxidation injury and has good prevention and treatment effects on liver injury, cholestatic liver diseases and the like caused by drugs. And the traditional Chinese medicine composition can obviously improve the symptoms such as weakness, nausea, skin and sclera yellow staining, yellow urine, itching, stomachache, hypochondrium lumps, body emaciation and the like in the liver injury period. The medicine is simple in preparation method, convenient to take, low in price, small in side effect and good in curative effect, and provides a good choice for vast patients with hepatopathy.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicines, and specifically relates to a traditional Chinese medicine compound composition for preventing and treating liver damage, a preparation method and an application thereof. Background Art

[0002] The liver is the main site for the metabolism and detoxification of drugs and xenobiotics in the human body. Drug-induced liver injury (DILI) caused by adverse drug reactions is an adverse reaction to drugs and certain compounds, and has become the leading cause of acute liver failure worldwide. Clinically, the main symptoms of DILI in the early stage are fatigue, nausea, yellow skin and sclera, yellow urine, etc. There are also asymptomatic patients, and only abnormal liver function is detected. The pathogenesis of drug-induced liver injury is complex, mainly involving drug factors and their metabolic mechanisms. The most common drugs that induce DILI include antibiotics, isoniazid and non-steroidal anti-inflammatory drugs. Initial hepatocyte damage is induced by drugs and / or their reactive metabolites through covalent binding or direct damage to mitochondria, leading to activation of oxidative stress and stress-sensing signaling pathways, impaired mitochondrial function and endoplasmic reticulum stress.

[0003] Cholestatic liver disease is a clinical syndrome characterized by abnormal bile secretion or excretion. Early clinical manifestations may be asymptomatic, with only elevated levels of alkaline phosphatase (ALP) and γ-glutamyl transpeptidase (GGT). However, as the disease progresses, itching, fatigue, and hyperbilirubinemia may occur, and in severe cases, it may lead to cirrhosis, liver failure, and even death. The causes of cholestatic liver disease are complex, including liver fibrosis, inflammatory damage, bile acid intrahepatic cytotoxicity, acid-induced oxidative stress, and other liver damage mechanisms, and there are many complications. At present, the treatment of cholestatic liver disease mostly uses ursodeoxycholic acid, adenosine methionine, etc., which have good efficacy but many side effects.

[0004] In view of the serious impact of drug-induced liver injury and cholestatic liver disease on human health, effective intervention measures to control the development of liver disease as early as possible and the development of corresponding therapeutic drugs are particularly important. In the intervention of therapeutic drugs, traditional Chinese medicine has significant effects in increasing efficacy and reducing toxicity, delaying the disease, protecting the liver and reducing enzymes. It can be used for overall regulation, reducing the inflammatory response of the liver and improving clinical symptoms. The holistic concept, syndrome differentiation and treatment, and the concept of preventing disease before it occurs in traditional Chinese medicine have always been the advantages of traditional Chinese medicine in preventing and treating diseases. Traditional Chinese medicine often uses multi-target and multi-component methods to treat liver disease. Early intervention with traditional Chinese medicine compound can reduce or avoid the further development of liver disease. In traditional Chinese medicine, liver disease is complex. According to different clinical symptoms, it is divided into the categories of "flank pain", "jaundice", "accumulation", "swelling" and other disease syndromes. The common pathogenesis of liver disease is mainly damp-heat, qi stagnation, and blood stasis, which are mostly syndromes of deficiency in the root and excess in the superficial. According to the common pathogenesis of liver disease, the treatment principles of clearing away heat and detoxification, removing dampness and jaundice, soothing the liver and strengthening the spleen, and promoting blood circulation and removing blood stasis are often used, but the existing prescriptions are relatively complex and the efficacy needs to be further improved. Therefore, it is necessary to develop a Chinese medicine compound composition with good liver protection effect as a treatment for patients with liver damage under the guidance of traditional Chinese medicine theory. Summary of the invention

[0005] In view of the deficiencies in the prior art, the purpose of the present invention is to provide a Chinese herbal compound composition for preventing and treating liver damage, a preparation method and application thereof, which can effectively solve the problems of complex formulations and unsatisfactory efficacy of Chinese herbal medicines for preventing and treating liver damage.

[0006] In order to achieve the above-mentioned purpose, the technical solution of the present invention is, on the one hand, to provide a Chinese medicine compound composition for preventing and treating liver damage, comprising the following raw materials in parts by weight: 20-30 parts of verbena, 15-20 parts of alfalfa, 10-15 parts of Panlong ginseng, 10-15 parts of agrimony, 6-9 parts of bergamot, 5-10 parts of malt, and 10-15 parts of Hypericum perforatum.

[0007] Preferably, the Chinese herbal compound composition comprises the following raw materials in parts by weight: 30 parts of verbena, 20 parts of alfalfa, 15 parts of Panlongshen, 15 parts of Agrimoniae, 9 parts of bergamot, 10 parts of malt, and 15 parts of Hypericum perforatum.

[0008] On the other hand, the technical solution of the present invention is to provide a method for preparing the Chinese medicine compound composition, comprising the following steps:

[0009] Step 1, weigh each raw material drug according to weight and set aside;

[0010] Step 2, crushing the verbena weighed in step 1 to obtain verbena coarse powder, adding ethanol solution to the verbena coarse powder for reflux extraction several times, filtering, and concentrating the obtained filtrate under reduced pressure to remove ethanol to obtain verbena extract solution; purifying the solution to obtain verbena extract;

[0011] Step 3, mixing the alfalfa, Panlong ginseng, Agrimony vine, bergamot, malt, and Hypericum perforatum weighed in step 1, soaking them in water, boiling them for several times, filtering them; combining the filtrate, concentrating them, and obtaining a mixed extract;

[0012] Step 4, mixing the verbena extract in step 2 and the mixed extract in step 3 evenly to obtain the Chinese medicine compound composition for treating liver injury.

[0013] Furthermore, in step 2, the concentration of the ethanol solution is 70%, v / v, and the amount used is 8-10 times the amount of verbena crude powder; the number of reflux extractions is 2 times, each time for 1-2 hours.

[0014] Further, in step 2, the purification method is: the drug solution is subjected to AB-8 macroporous adsorption resin, firstly eluted with 2-4 times the volume of the resin with a 30% ethanol solution to remove impurities, and then eluted with 6-8 times the volume of the resin with a 75% ethanol solution, the second alcohol eluate is collected, concentrated under reduced pressure, and dried to obtain the verbena extract.

[0015] Furthermore, in step 3, the amount of water added is 2-4 times the amount of the raw material medicine; the soaking time is 30-40 minutes; the decoction number is 2 times, the time is 0.5-1 hour; and the concentration is to a relative density of 1.2-1.4.

[0016] On the other hand, the technical solution of the present invention is to provide an application of the Chinese medicine compound composition in the preparation of a pharmaceutical preparation for preventing and treating liver damage.

[0017] Preferably, the Chinese herbal compound composition for preventing and treating liver damage is prepared into a pharmaceutically acceptable drug preparation according to conventional methods. The pharmaceutical dosage form of the drug preparation can be oral liquid, granules, capsules, pills, powders, microcapsules or dripping pills.

[0018] The verbena described in the technical scheme of the present invention is the dry aerial part of the verbena plant Verbena officinalis L. of the Verbenaceae family. It is bitter in taste, cool in nature, and enters the liver and spleen meridians. It has the effects of promoting blood circulation and dispersing blood stasis, promoting diuresis, relieving jaundice, and stopping malaria. It is mainly used to treat the symptoms such as accumulation of masses, dysmenorrhea, amenorrhea, laryngeal paralysis, carbuncle, edema, jaundice, and malaria. Verbena contains rich chemical components, in addition to iridoid ether terpenoid glycosides, such as verbena glycoside, there are also flavonoid components, such as kaempferol, luteolin and apigenin, and specific flavonoids such as baicalin and peony glycoside, which have the effects of anti-oxidation, anti-inflammatory, liver protection and anti-cancer. Verbena is more common in clinical research and treatment of diseases, can promote diuresis and eliminate dampness, clear away heat and detoxify, and is more common and has broad prospects in protecting liver function and relieving cholestasis.

[0019] The alfalfa described in the technical solution of the present invention is the whole herb of the leguminous plant purple alfalfa or southern alfalfa Medicago sativa L. The quercetin and luteolin in alfalfa can improve the antioxidant capacity of cells and regulate the body's immunity.

[0020] The Panlong ginseng described in the technical solution of the present invention is the root and whole herb of Spiranthes sinensis (Pers.) Ames of the orchid family. The ferulic acid esters in Panlong ginseng have strong antioxidant and anti-liver damage activities.

[0021] The agrimonia described in the technical solution of the present invention is the dried aerial part of Agrimonia pilosa Ledeb., a plant of the Rosaceae family. The main active ingredients of the agrimonia, such as flavonoids and triterpenes, have pharmacological activities such as anti-tumor, analgesic, anti-inflammatory and antioxidant.

[0022] The bergamot described in the technical solution of the present invention is the dried fruit of Citrus medica L. var. sarcodactylis Swingle, a plant of the Rutaceae family.

[0023] The malt described in the technical solution of the present invention is a processed product obtained by germination and drying of mature fruits of Hordeum vulgare L., a gramineous plant.

[0024] The Hypericumperforatum described in the technical solution of the present invention is the whole herb or the whole herb with roots of the plant Hypericumperforatum L. of the family Cucurbitaceae. The components such as rutin, hyperoside and naphthodianthrone hypericin in Hypericumperforatum have the effects of anti-oxidative stress, anti-tumor and liver protection.

[0025] Among the raw materials used to prepare the Chinese medicine composition of the present invention, verbena tastes bitter, cool in nature, has the functions of promoting blood circulation and dispersing blood stasis, detoxifying, promoting diuresis, and relieving jaundice, and is the main medicine; alfalfa tastes sweet and light, is slightly cold in nature, can clear away heat and dampness, promote diuresis and reduce swelling, Panlongshen tastes sweet and bitter, is flat in nature, has the effects of invigorating qi and nourishing yin, clearing away heat and detoxifying, and herb of agrimony tastes bitter and astringent, is flat in nature, has the functions of detoxifying and replenishing deficiency, and can also astringe and stop bleeding, stop malaria, and stop dysentery, and all three are ministerial medicines; with bergamot, malt, and Hypericum perforatum, bergamot tastes bitter and sour, is warm in nature, can soothe the liver and regulate qi, harmonize the stomach and relieve pain, dry dampness and resolve phlegm, malt is sweet and flat, can promote qi and soothe the liver, invigorate the spleen and stimulate appetite, Hypericum perforatum tastes pungent and slightly bitter, is flat in nature, can soothe the liver and relieve depression, can clear away heat and detoxify, astringe and stop bleeding, promote dampness and reduce swelling. All the medicines are used together to play the functions of clearing away heat and detoxifying, promoting dampness and relieving jaundice, promoting blood circulation and dispersing blood stasis, soothing the liver and strengthening the spleen.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] The Chinese medicine compound composition of the present invention is used for treating liver injury, can improve liver function, and regulate body immunity. According to the common pathogenesis of liver disease, which is mainly damp-heat, qi stagnation, and blood stasis, most of them are syndromes of deficiency in the root and excess in the superficial part, the use of medicine is relatively focused on the use of Chinese medicine with the effects of clearing away heat and detoxification, promoting dampness and relieving jaundice, soothing the liver and strengthening the spleen, and promoting blood circulation and removing blood stasis. After many years of clinical practice, this compound composition for preventing and treating liver injury has been developed.

[0028] The Chinese medicine compound composition of the present invention has a simple structure and a reasonable ratio. It has been verified by animal experiments that it can reduce liver inflammation damage, inhibit liver cell necrosis and fatty degeneration, inhibit oxidative stress and resist lipid peroxidation damage, and has good preventive and therapeutic effects on drug-induced liver damage, cholestatic liver disease, etc. The Chinese medicine composition is relatively safe and has obvious improvements on symptoms such as fatigue, nausea, yellowing of the skin and sclera, yellow urine, itching, abdominal pain, lumps in the ribs, and weight loss during liver damage. The Chinese medicine composition of the present invention has a simple preparation method, is easy to take, is low in price, has few side effects, and has a significant therapeutic effect, providing a good choice for a large number of patients with liver disease. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 HE staining (×200) of liver tissues of mice in each group in Experiment 1. In the figure, A: normal control group; B: model group; C: bifendate group; D: low-dose group of Chinese medicine compound composition; E: medium-dose group of Chinese medicine compound composition; F: high-dose group of Chinese medicine compound composition.

[0030] Figure 2 HE staining (×200) of liver tissues of mice in each group in Experiment 2. In the figure, A: normal control group; B: model group; C: ursodeoxycholic acid group; D: low-dose group of Chinese herbal compound composition; E: medium-dose group of Chinese herbal compound composition; F: high-dose group of Chinese herbal compound composition. DETAILED DESCRIPTION

[0031] In order to enable those skilled in the art to better understand and implement the technical solution of the present invention, the present invention is further described below in conjunction with specific embodiments, but the embodiments are not intended to limit the present invention.

[0032] Verbena, Agrimonia, alfalfa, Panlongshen, Citron, malt, and Hypericum perforatum used in the embodiments of the present invention are all commercially available, and the test methods are all conventional methods unless otherwise specified.

[0033] Example 1

[0034] A Chinese medicinal compound composition for preventing and treating liver damage comprises the following raw materials in parts by weight: 30 parts of verbena, 20 parts of alfalfa, 15 parts of Panlongshen, 15 parts of Agrimoniae, 9 parts of bergamot, 10 parts of malt and 15 parts of Hypericum perforatum.

[0035] Its preparation method is:

[0036] Step 1, weigh each raw material drug according to weight and set aside;

[0037] Step 2, crushing the verbena weighed in step 1 and passing it through a 40-mesh sieve to obtain verbena coarse powder, adding 8 times the amount of 70% (v / v) ethanol solution to the verbena coarse powder, reflux extraction, extracting twice, each extraction time for 1.5 hours, filtering, and decompressing and concentrating the obtained filtrate to remove ethanol to obtain verbena extract liquid;

[0038] The drug solution was placed on AB-8 macroporous adsorption resin, and the impurities were first eluted with 30% (v / v) ethanol solution of 2 times the volume of the resin, and then eluted with 75% (v / v) ethanol solution of 8 times the volume of the resin, and the second alcohol eluate was collected, concentrated under reduced pressure, and dried to obtain verbena extract;

[0039] Step 3, mixing the alfalfa, Panlong ginseng, Agrimony vine, bergamot, malt, and Hypericum perforatum weighed in step 1, and then adding 4 times the amount of water as the mixed raw materials to soak for 30 minutes, decocting and extracting for 1 hour, and filtering; adding water equivalent to 2 times the amount of the residue to the residue, decocting and extracting for 0.5 hours, and filtering; combining the two filtrates, concentrating to a relative density of 1.2-1.4, and obtaining a mixed extract;

[0040] Step 4, mixing the verbena extract in step 2 and the mixed extract in step 3 evenly to obtain the Chinese medicine compound composition for treating liver injury.

[0041] The above Chinese medicine compound composition is crushed, and then a proper amount of auxiliary materials are added, mixed, granulated, sieved, and dried to obtain a granular Chinese medicine compound composition for preventing and treating liver damage.

[0042] Example 2

[0043] A Chinese medicinal compound composition for preventing and treating liver damage comprises the following raw materials in parts by weight: 20 parts of verbena, 15 parts of alfalfa, 10 parts of Panlongshen, 10 parts of Agrimoniae, 6 parts of bergamot, 5 parts of malt, and 10 parts of Hypericum perforatum.

[0044] Its preparation method is:

[0045] Step 1, weigh each raw material drug according to weight and set aside;

[0046] Step 2, crushing the verbena weighed in step 1 and passing it through a 40-mesh sieve to obtain verbena coarse powder, adding 8 times the amount of 70% (v / v) ethanol solution to the verbena coarse powder, reflux extraction, extracting twice, each extraction for 1 hour, filtering, and decompressing and concentrating the obtained filtrate to remove ethanol to obtain verbena extract liquid;

[0047] The drug solution was placed on AB-8 macroporous adsorption resin, and the impurities were first eluted with 30% (v / v) ethanol solution of 2 times the volume of the resin, and then eluted with 75% (v / v) ethanol solution of 6 times the volume of the resin, and the second alcohol eluate was collected, concentrated under reduced pressure, and dried to obtain verbena extract;

[0048] Step 3, mixing the alfalfa, Panlong ginseng, Agrimony, bergamot, malt, and Hypericum perforatum weighed in step 1, and then adding 2 times the amount of water as the mixed raw materials to soak for 30 minutes, decocting and extracting for 1 hour, and filtering; adding 2 times the amount of water as the residue to the residue, decocting and extracting for 0.5 hours, and filtering; combining the two filtrates, concentrating to a relative density of 1.2-1.4, and obtaining a mixed extract;

[0049] Step 4, mixing the verbena extract in step 2 and the mixed extract in step 3 evenly to obtain the Chinese medicine compound composition for treating liver injury.

[0050] The Chinese medicine compound composition prepared in Example 2 is crushed, and then an appropriate amount of auxiliary materials are added, mixed, granulated, sieved, and dried. The obtained granules are filled into capsule shells to make capsules, thereby obtaining a capsule-type Chinese medicine compound composition for preventing and treating liver injury.

[0051] Example 3

[0052] A Chinese medicinal compound composition for preventing and treating liver damage comprises the following raw materials in parts by weight: 25 parts of verbena, 18 parts of alfalfa, 12 parts of Panlongshen, 12 parts of Agrimoniae, 8 parts of bergamot, 8 parts of malt and 12 parts of Hypericum perforatum.

[0053] Its preparation method is:

[0054] Step 1, weigh each raw material drug according to weight and set aside;

[0055] Step 2, crushing the verbena weighed in step 1 and passing it through a 40-mesh sieve to obtain verbena coarse powder, adding 10 times the amount of 70% (v / v) ethanol solution to the verbena coarse powder, reflux extraction, extracting twice, each extraction for 2 hours, filtering, and concentrating the obtained filtrate under reduced pressure to remove ethanol to obtain verbena extract liquid;

[0056] The drug solution was placed on AB-8 macroporous adsorption resin, and the impurities were first eluted with 30% (v / v) ethanol solution of 4 times the volume of the resin, and then eluted with 75% (v / v) ethanol solution of 6 times the volume of the resin, and the second alcohol eluate was collected, concentrated under reduced pressure, and dried to obtain verbena extract;

[0057] Step 3, mixing the alfalfa, Panlong ginseng, Agrimony, bergamot, malt, and Hypericum perforatum weighed in step 1, and then adding 4 times the amount of water as the mixed raw materials to soak for 40 minutes, decocting and extracting for 1 hour, and filtering; adding water equivalent to 2 times the amount of the residue to the residue, decocting and extracting for 0.5 hours, and filtering; combining the two filtrates, concentrating to a relative density of 1.2-1.4, and obtaining a mixed extract;

[0058] Step 4, mixing the verbena extract in step 2 and the mixed extract in step 3 evenly to obtain the Chinese medicine compound composition for treating liver injury.

[0059] The Chinese herbal compound composition prepared in Example 3 is crushed, and then appropriate amounts of auxiliary materials are added, mixed, sieved, granulated, and dried to obtain a pill-type Chinese herbal compound composition for preventing and treating liver damage.

[0060] Examples 1-3 all prepared Chinese herbal compound compositions for preventing and treating liver damage with good efficacy. Since the efficacy of the Chinese herbal compound compositions for preventing and treating liver damage prepared in Examples 1-3 is basically the same, only the Chinese herbal compound composition prepared in Example 1 is used to illustrate the effect of the present invention.

[0061] Validation experiment 1: Effects on the acetaminophen-induced drug-induced liver injury model in mice

[0062] Acetaminophen (APAP) is a common antipyretic and analgesic drug in clinical practice. Intake of high doses of APAP is the main cause of DILI, especially taking APAP at 2-3 times the recommended dose may cause liver damage. In this experiment, an acetaminophen-induced drug-induced liver injury model was established in mice by administering APAP to verify the effect of the Chinese medicine compound composition on the prevention and treatment of drug-induced liver injury.

[0063] 1 Experimental methods

[0064] 1.1 Preparation and administration of drug-induced liver injury model in mice

[0065] 60 SPF male KM mice were randomly and evenly divided into 6 groups, namely normal control group, model group, low, medium and high dose groups of Chinese medicine compound composition (administered 14.25, 28.5, 57g / kg) and bifendate group (administered 7.5mg / kg), with 10 mice in each group. The Chinese medicine compound composition administration group and bifendate group were gavaged with different doses of Chinese medicine compound composition and bifendate at a volume of 20ml / kg, and the normal control group and model group were given the same volume of distilled water every day. The drug was administered once a day for 7 consecutive days. One hour after the last administration, APAP 500mg / kg was gavaged to prepare drug-induced liver injury models in all groups except the normal control group.

[0066] 1.2 Specimen collection and processing

[0067] The mice were processed 24 hours after modeling, fasted but not watered for 12 hours, blood was collected after anesthesia, serum was collected by centrifugation, and the liver was removed.

[0068] 1.3 Index detection

[0069] The microplate method was used to detect the levels of serum liver function indicators alanine aminotransferase (ALT) and aspartate aminotransferase (AST). The enzyme-linked immunosorbent assay (ELISA) method was used to detect the levels of serum inflammatory factors tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). After collecting serum, the liver was removed, and the left lobe of the liver was fixed in 10% (v / v) formaldehyde solution, routinely paraffin-embedded and sectioned, and stained with hematoxylin-eosin, and histopathological changes were observed under an optical microscope.

[0070] 1.4 Statistical analysis

[0071] The experimental results were measured using The data were analyzed using SPSS 19.0 statistical software, and one-way analysis of variance was used for comparison among the groups. The difference was considered statistically significant when P < 0.05.

[0072] 2 Experimental results

[0073] 2.1 The effects of the Chinese herbal compound on the liver function of mice with drug-induced liver injury are shown in Table 1.

[0074] Table 1 Effects on liver function of mice with drug-induced liver injury ( n=10)

[0075]

[0076] Note: Compared with the model group * P<0.05, ** P<0.01.

[0077] As shown in Table 1, compared with the normal control group, the ALT and AST levels in the serum of the model group mice were significantly increased (P < 0.01), indicating that the mouse drug-induced liver injury model was successfully established. Compared with the model group, the ALT and AST levels in the low-, medium-, and high-dose groups of the Chinese medicine compound composition and the bifendate group were significantly reduced (P < 0.01 or P < 0.05), and the effect of the high-dose group of the Chinese medicine compound composition was better than that of the bifendate group, indicating that the Chinese medicine compound composition has a good improvement effect on the liver function of mice with drug-induced liver injury.

[0078] 2.2 The effects of the Chinese herbal compound composition on inflammatory indicators in mice with drug-induced liver injury are shown in Table 2.

[0079] Table 2 Effects on serum TNF-α and IL-6 in mice with drug-induced liver injury ( n=10)

[0080]

[0081]

[0082] Note: Compared with the model group * P<0.05, ** P<0.01.

[0083] As shown in Table 2, compared with the normal control group, the levels of TNF-α and IL-6 in the serum of the model group mice were significantly increased (P < 0.01). Compared with the model group, the levels of TNF-α and IL-6 in the serum of the mice in each dose group of the Chinese medicine compound composition were significantly reduced (P < 0.05 or P < 0.01), and the effect of the high dose group of the Chinese medicine compound composition was better than that of the bifendate group, indicating that the Chinese medicine compound composition can effectively reduce the inflammatory response of the liver of mice with drug-induced liver injury.

[0084] 2.3 Effects of Chinese herbal medicine compound on liver histopathology in mice Figure 1 .

[0085] Under the optical microscope, it can be seen that the liver lobule structure of the mice in the normal control group is normal, the size of the hepatocytes is uniform, and the hepatic cords are arranged neatly in a radial pattern. The hepatic cords of the mice in the model group are disordered, the hepatocytes are swollen, and obvious necrosis, fatty degeneration and inflammatory cell infiltration can be seen. Compared with the model group, the hepatic cords of the mice in the bifendate group and the Chinese medicine compound composition groups tend to be neatly arranged, and the degree of hepatic cell necrosis, fatty degeneration and inflammatory cell infiltration is significantly reduced. Among them, the high-dose group of the Chinese medicine compound composition has a more obvious therapeutic effect, which indicates that the Chinese medicine compound composition can inhibit hepatic cell necrosis, fatty degeneration and inflammatory cell infiltration to varying degrees. The higher the dose, the better the effect.

[0086] Verification experiment 2: Effects on the mouse cholestatic liver injury model induced by α-naphthalene isothiocyanate (ANIT)

[0087] 1 Experimental methods

[0088] 1.1 Preparation and administration of cholestatic liver injury model in mice

[0089] 60 SPF male KM mice were randomly and evenly divided into 6 groups, namely normal control group, model group, low, medium and high dose groups of Chinese herbal compound composition (administered 14.25, 28.5, 57g / kg) and ursodeoxycholic acid group (administered 75mg / kg), with 10 mice in each group. The Chinese herbal compound composition administration group and ursodeoxycholic acid group were gavaged with different doses of Chinese herbal compound composition and ursodeoxycholic acid at a volume of 20ml / kg, and the normal control group and model group were given the same volume of distilled water every day. The drug was administered once a day for 7 consecutive days. After 6 hours of administration on the 5th day, except for the normal control group, the other groups were gavaged with 60mg / kg ANIT olive oil solution to prepare an acute cholestatic liver injury model.

[0090] 1.2 Specimen collection and processing

[0091] After ANIT was given for 48 hours, mice were fasted but not watered for 12 hours, blood was collected after anesthesia, serum was separated, and liver was removed. The right lobe of the liver was added with 0.9% sodium chloride solution to prepare 10% liver tissue homogenate, centrifuged at 3500rpm, and the supernatant was taken for later use.

[0092] 1.3 Index detection

[0093] The levels of serum liver function indicators alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), and total bile acid (TBA) were detected by microplate method. The level of superoxide dismutase (SOD) in liver tissue homogenate was detected by microplate reader, and the level of malondialdehyde (MDA) was detected by colorimetry. The left lobe of the liver was fixed in 10% (v / v) formaldehyde solution, routinely paraffin-embedded and sectioned, stained with hematoxylin-eosin, and the pathological changes of the tissue were observed under an optical microscope.

[0094] 1.4 Statistical analysis

[0095] The experimental results were measured using The data were analyzed using SPSS 19.0 statistical software, and one-way analysis of variance was used for comparison among the groups. The difference was considered statistically significant when P < 0.05.

[0096] 2 Experimental results

[0097] 2.1 Effect of the Chinese herbal compound composition on liver function in mice with cholestatic liver injury.

[0098] Table 3 Effects on liver function of mice with cholestatic liver injury ( n=10)

[0099]

[0100] Note: Compared with the model group ** P<0.05, **P<0.01.

[0101] As shown in Table 3, compared with the normal control group, the levels of ALT, AST, TBIL, and TBA in the serum of the model group mice were significantly increased (P < 0.01), indicating that the cholestatic liver injury model of mice was successfully established. Compared with the model group, the levels of ALT, AST, TBIL, and TBA in the low-, medium-, and high-dose groups of the Chinese medicine compound composition and the ursodeoxycholic acid group were significantly reduced (P < 0.05 or P < 0.01), and the effect of the high-dose group of the Chinese medicine compound composition was significantly better than that of the ursodeoxycholic acid group, indicating that the Chinese medicine compound composition has a significant improvement on the liver function of mice with cholestatic liver injury.

[0102] 2.2 Effects of the Chinese herbal compound composition on oxidative indices of liver tissue in mice with cholestatic liver injury.

[0103] Table 4 Effects on SOD and MDA in liver tissue of mice with cholestatic liver injury ( n=10)

[0104]

[0105] Note: Compared with the model group * P<0.05, ** P<0.01.

[0106] As shown in Table 4, compared with the normal control group, the SOD level in the liver tissue of the mice in the model group was significantly decreased (P < 0.01), and the MDA level was significantly increased (P < 0.01). Compared with the model group, the SOD level in the liver tissue of the mice in each dose group of the Chinese medicine compound composition was significantly increased, and the MDA level was significantly decreased (P < 0.05 or P < 0.01), indicating that the Chinese medicine compound composition can inhibit oxidative stress in liver tissue and has a good protective effect on lipid peroxidation damage.

[0107] 2.3 Effects of Chinese herbal medicine compound on liver histopathology in mice Figure 2 .

[0108] Under the optical microscope, the normal control group mice had neatly arranged liver cells, normal liver lobule structure, no bile duct hyperplasia and fibrous tissue, normal liver cell morphology without degeneration and necrosis, and no inflammatory cell infiltration around the bile duct. The model group mice had punctate and focal necrosis of liver cells, severe bile duct hyperplasia, severe hyperplasia of surrounding fibrous tissue, and a large number of inflammatory cells infiltrating around the bile duct. Compared with the model group, the ursodeoxycholic acid group and the Chinese medicine compound composition group had intact liver lobule structure, no necrosis and degeneration of liver cells, a small amount of bile duct hyperplasia and fibrous tissue, and only a small amount of inflammatory cells infiltrating around the bile duct. Among them, the high-dose group of the Chinese medicine compound composition had more obvious efficacy, which showed that each dose group of the Chinese medicine compound composition could inhibit liver cell necrosis and inflammatory cell infiltration to varying degrees, and the higher the dose, the better the effect.

[0109] The above-mentioned acetaminophen-induced mouse drug-induced liver injury model and α-naphthalene isothiocyanate-induced mouse cholestatic liver injury model both confirmed that the Chinese herbal compound composition of the present invention can significantly improve liver function, reduce liver inflammatory response, and resist oxidative stress, and has a definite preventive and therapeutic effect on drug-induced liver injury, cholestatic liver disease, etc., so it can be used to prepare drugs for preventing and treating liver injury.

Claims

1. A Chinese medicinal compound composition for preventing and treating liver damage, characterized in that: The invention comprises the following raw materials in parts by weight: 20-30 parts of verbena, 15-20 parts of alfalfa, 10-15 parts of Panlong ginseng, 10-15 parts of agrimony, 6-9 parts of bergamot, 5-10 parts of malt and 10-15 parts of Hypericum perforatum.

2. The Chinese medicine compound composition according to claim 1, characterized in that: The invention comprises the following raw materials in parts by weight: 30 parts of verbena, 20 parts of alfalfa, 15 parts of Panlong ginseng, 15 parts of agrimony, 9 parts of bergamot, 10 parts of malt and 15 parts of Hypericum perforatum.

3. A method for preparing the Chinese medicine compound composition according to claim 1 or 2, characterized in that: The following steps are involved: Step 1, weigh each raw material drug according to weight and set aside; Step 2, crushing the verbena weighed in step 1 to obtain verbena coarse powder, adding ethanol solution to the verbena coarse powder for reflux extraction several times, filtering, and concentrating the obtained filtrate under reduced pressure to remove ethanol, thereby obtaining a verbena extract solution; Purifying the medicinal solution to obtain verbena extract; Step 3, mixing the alfalfa, Panlong ginseng, Agrimony vine, bergamot, malt, and Hypericum perforatum weighed in step 1, soaking them in water, boiling them for several times, filtering them; combining the filtrate, concentrating them, and obtaining a mixed extract; Step 4, mixing the verbena extract in step 2 and the mixed extract in step 3 evenly to obtain the Chinese medicine compound composition for treating liver injury.

4. The method for preparing the Chinese medicine compound composition according to claim 3, characterized in that: In step 2, the concentration of the ethanol solution is 70%, v / v, and the amount used is 8-10 times the amount of verbena crude powder; the number of reflux extractions is 2 times, each time for 1-2 hours.

5. The method for preparing the Chinese medicine compound composition according to claim 3, characterized in that: In step 2, the purification method is as follows: the drug solution is subjected to AB-8 macroporous adsorption resin, firstly eluted with 30% ethanol solution of 2-4 times the volume of the resin to remove impurities, then eluted with 75% ethanol solution of 6-8 times the volume of the resin, the second alcohol eluate is collected, concentrated under reduced pressure, and dried to obtain verbena extract.

6. The method for preparing the Chinese medicine compound composition according to claim 3, characterized in that: In step 3, the amount of water added is 2-4 times the amount of the raw material medicine; the soaking time is 30-40 minutes; the decoction number is 2 times, the time is 0.5-1 hour; and the concentration is performed to a relative density of 1.2-1.

4.

7. Use of the Chinese medicine compound composition as claimed in claim 1 or 2 in the preparation of a pharmaceutical preparation for preventing and treating liver damage.

8. The use according to claim 7, characterized in that: The traditional Chinese medicine compound composition for preventing and treating liver damage is prepared into a pharmaceutically acceptable drug preparation according to a conventional method.

9. The use according to claim 8, characterized in that: The pharmaceutical dosage form of the drug preparation is oral liquid, granules, capsules, pills, powders, microcapsules or dripping pills.

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