Preparation combined by two traditional Chinese medicine components and used for treating non-alcoholic fatty liver disease

Abstract

The invention discloses a traditional Chinese medicine component composition for treating non-alcoholic fatty liver disease, which is prepared from two traditional Chinese medicine components. The traditional Chinese medicine component composition is prepared from ferulic acid and chlorogenic acid. Animal experiments show that the preparation for treating the non-alcoholic fatty liver disease, which is formed by combining the two traditional Chinese medicine components, can remarkably reduce the content of triglyceride in hepatic tissue of a model mouse and relieve the degree of hepatic fatty degeneration and the degree of hepatic injury, the effect is better than that of single use of the two traditional Chinese medicine components, the effect of resisting the non-alcoholic fatty liver disease can be improved, and the preparation can be used for treating the non-alcoholic fatty liver disease. The traditional Chinese medicine composition can effectively prevent development of the non-alcoholic fatty liver disease, and can be used for treating and preventing diseases such as the non-alcoholic fatty liver disease and chronic liver injury.

Description

Technical Field

[0001] The present invention belongs to the technical field of the preparation of traditional Chinese medicine preparations. Specifically, it relates to a preparation for treating non-alcoholic fatty liver disease composed of a combination of two traditional Chinese medicine components. Background Art

[0002] Non-alcoholic fatty liver disease (NAFLD) is a progressive disease mainly characterized by excessive accumulation of fat in liver cells. Among them, about 40% of patients will evolve into non-alcoholic steatohepatitis (NASH), and then develop into progressive fibrosis. At the same time, about 10% of NAFLD patients will develop liver cirrhosis and hepatocellular carcinoma within 10 - 20 years after diagnosis. At the same time, more and more evidence shows that NAFLD is a multi-system disease, which not only increases the risk of liver-related complications, but also increases the risks of type 2 diabetes, cardiovascular diseases, kidney diseases and extrahepatic cancers, and the risks increase with the severity of NAFLD.

[0003] The high prevalence rate has attracted much attention and expectation for the R & D of NAFLD drugs. However, in the face of a huge patient group, the R & D of anti-NAFLD drugs is still extremely urgent.

[0004] In the treatment of NAFLD, traditional Chinese medicine combines the overall concept and syndrome differentiation and treatment, and the traditional Chinese medicine acts on multiple pathways, multiple targets and multiple links, which is in line with the pathological mechanisms of the multi-systematic and metabolic complex diseases of NAFLD. At present, Qiaozhi Capsule, Zhibitai Capsule, Qianggan Capsule, Dangfei Liganing Capsule and other traditional Chinese medicine preparations for treating NAFLD are widely used in clinical practice and have achieved good curative effects, indicating that traditional Chinese medicine has certain advantages and prospects in the treatment of systemic multi-systematic diseases. Therefore, it is of great significance to deeply carry out the research on the treatment of NAFLD with traditional Chinese medicine.

[0005] The components of traditional Chinese medicine compound are complex, and it is difficult to correspond the pharmacological mechanism to a single effective component. Moreover, in the specific clinical application, the standardization system of traditional Chinese medicine compound is not yet perfect, which affects the improvement of the controllability and stability of the actual curative effect and is not conducive to international connection. In recent years, on the basis of continuously discovering effective and ideal traditional Chinese medicine active ingredients for treating NAFLD, the development of traditional Chinese medicine active ingredient compounds with multi-pathway pharmacological effects and "synergistic compatibility" is one of the future development trends of traditional Chinese medicine in the treatment of NAFLD. Summary of the Invention

[0006] The purpose of the present invention is to provide a traditional Chinese medicine component composition for treating non-alcoholic fatty liver disease composed of a combination of two traditional Chinese medicine components.

[0007] In order to achieve the above purpose, the technical scheme adopted by the present invention is as follows:

[0008] In the first aspect of the present invention, there is provided a traditional Chinese medicine component composition for treating non-alcoholic fatty liver disease composed of a combination of two traditional Chinese medicine components, which is made from ferulic acid and chlorogenic acid.

[0009] The mass ratio of ferulic acid to chlorogenic acid is (1-10):3, preferably 4:3.

[0010] The traditional Chinese medicine ingredient composition for treating non-alcoholic fatty liver disease composed of 2 kinds of traditional Chinese medicine ingredients can play the roles of anti-non-alcoholic fatty liver disease and anti-liver injury.

[0011] The ferulic acid comes from traditional Chinese medicines such as Polygonum cuspidatum and can be obtained by conventional extraction methods or commercial procurement.

[0012] The chlorogenic acid comes from traditional Chinese medicines such as Artemisia capillaris and Gardenia jasminoides and can be obtained by conventional extraction methods or commercial procurement.

[0013] Tests show that the combined use of ferulic acid and chlorogenic acid is significantly superior to the single use of ferulic acid or chlorogenic acid in reducing liver triglycerides, alleviating the degree of hepatic steatosis and liver injury.

[0014] In the second aspect of the present invention, there is provided an application of the traditional Chinese medicine ingredient composition in the preparation of a drug for treating non-alcoholic fatty liver disease.

[0015] In the third aspect of the present invention, there is provided an application of the traditional Chinese medicine ingredient composition in the preparation of a drug for preventing and treating liver injury.

[0016] In the fourth aspect of the present invention, there is provided a pharmaceutical preparation which is a pharmaceutical preparation made from the traditional Chinese medicine ingredient composition and pharmaceutically acceptable excipients.

[0017] The dosage form of the pharmaceutical preparation is an oral preparation.

[0018] The dosage form is selected from granules, tablets or capsules.

[0019] In the fifth aspect of the present invention, there is provided an application of the traditional Chinese medicine ingredient composition in the preparation of a drug for treating non-alcoholic fatty liver disease.

[0020] In the traditional Chinese medicine composition of the present invention, a non-alcoholic fatty liver model of C57 mice induced by high-fat diet and high-sugar drinking water is screened by using "uniform design" and verified by a non-alcoholic fatty liver model of C57 mice induced by high-fat diet, high-sugar drinking water, and methionine- and choline-deficient diet (therapeutic administration after model establishment). The results show that the preparation for treating non-alcoholic fatty liver disease composed of two traditional Chinese medicine components of the present invention can significantly reduce the content of triglyceride (TG) in the liver tissue of model mice, alleviate the degree of hepatic steatosis and liver injury, and the above effects are better than those of single use respectively. The combination application of the two components in the preparation for treating non-alcoholic fatty liver disease of the present invention can improve the anti-NAFLD effect, effectively prevent the development of non-alcoholic fatty liver and promote the reversal of fatty liver, and can be used for treating and preventing non-alcoholic fatty liver disease, various chronic liver injuries and other diseases.

[0021] Due to the adoption of the above technical solution, the present invention has the following advantages and beneficial effects:

[0022] Based on research practice, the present application provides a compound preparation of traditional Chinese medicine components with significant effects of inhibiting liver fat deposition and anti-liver injury, especially a compound preparation of traditional Chinese medicine components for treating non-alcoholic fatty liver disease.

[0023] The preparation for treating non-alcoholic fatty liver disease composed of two traditional Chinese medicine components provided by the present invention can be further used for preparing drugs for treating chronic liver diseases, drugs for treating non-alcoholic fatty liver disease, and drugs for preventing and treating liver injuries.

[0024] Animal experiments show that the preparation for treating non-alcoholic fatty liver disease composed of two traditional Chinese medicine components provided by the present invention can significantly reduce the content of triglyceride in the liver tissue of model mice, alleviate the degree of hepatic steatosis and liver injury. The above effects are better than those of single use respectively, can improve the anti-non-alcoholic fatty liver effect, effectively prevent the development of non-alcoholic fatty liver, and can be used for treating and preventing non-alcoholic fatty liver disease, chronic liver injury and other diseases. Description of the Drawings

[0025] Figure 1 It is a schematic diagram of the pathological observation result of the liver tissue of C57 mice in Example 2 (HE staining × 200).

[0026] Figure 2 It is a schematic diagram of the pathological observation result of the liver tissue of C57 mice in Example 2 (oil red O staining × 200).

[0027] Figure 3 It is a schematic diagram of the pathological observation result of the liver tissue of C57 mice in Example 3 (HE staining × 200).

[0028] Figure 4Schematic diagram of the pathological observation results of the liver tissue of C57 mice in Example 3 (Oil Red O staining × 200). Detailed implementation manners

[0029] To illustrate the present invention more clearly, the present invention will be further described below in conjunction with preferred embodiments. Those skilled in the art should understand that the content specifically described below is illustrative rather than restrictive, and should not be used to limit the protection scope of the present invention.

[0030] Example 1

[0031] A non-alcoholic fatty liver model of mice induced by high-fat diet and high-sugar drinking water and the uniform design method were used for regression analysis to obtain the corresponding optimal formula for reducing the content of liver triglycerides

[0032] 1. Materials

[0033] (1) Animals: 60 male C57BL / 6J mice, clean grade, body weight (21 - 25) g, purchased from Shanghai Jihui Experimental Animal Co., Ltd.

[0034] (2) Main reagents and drugs: Ferulic acid, CAS number: 537 - 98 - 4, batch number: 230812; Chlorogenic acid, CAS number: 327 - 97 - 9, batch number: 231208; Crocetin, CAS number: 27876 - 94 - 4, batch number: 231023; Caffeic acid, CAS number: 331 - 39 - 5, batch number: 230927; Gallic acid, CAS number: 149 - 91 - 7, batch number: 230916; Quinic acid, CAS number: 77 - 95 - 2, batch number: 230822; The above drugs were all purchased from Shanghai Ronghe Medical Technology Development Co., Ltd. High-fat diet feed (60% of the energy comes from fat), normal control feed (10% of the energy comes from fat), the feed was purchased from Research Diets, Inc., USA. Drink sugar water with a concentration of 42 g / L (the sugar water ratio is 55% fructose and 45% sucrose). The triglyceride detection kit was purchased from Zhejiang Dong'ou Diagnostic Products Co., Ltd.

[0035] 2. Methods

[0036] (1)Uniform design drug screening experiment: 60 male C57BL / 6J mice were given a high-fat diet and high-sugar drinking water for 27 weeks. There were a total of 12 groups, including a normal group, a model group, and ten groups with uniform design formulas. Starting from the 19th week of modeling, the mice were randomly divided into a model group and a drug group, and intragastric administration or drinking water was continued for modeling until the 27th week. Each drug group was formulated with drugs according to the uniform design table, and drugs of each group were prepared at corresponding concentrations and intragastrically administered at a dose of 0.1 ml / 10 g body weight of mice. After 8 weeks of treatment, the mice were anesthetized by intraperitoneal injection of 3% sodium pentobarbital at a dose of 0.01 ml / g, blood was collected from the inferior vena cava, serum was separated by centrifugation, and liver tissue was taken for examination.

[0037] (2)Uniform design experimental scheme: According to the uniform design scheme, U 11 (11 6 ) table was used to calculate the drug administration. The six effective components of the six traditional Chinese medicines in the formula (chlorogenic acid, crocetin, caffeic acid, ferulic acid, gallic acid, quinic acid) were used as the investigation factors, and X1, X2, X3, X4, X5, and X6 were used to represent the six effective components of the six traditional Chinese medicines in the formula respectively. Each factor was taken at 5 levels (i.e., 5 different doses increasing in gradient), with 2 repetitions (as shown in Table 1). First, the effective doses of each drug against NAFLD reported in the literature were retrieved: the 5 levels of X1 (chlorogenic acid) were between 15 - 60 mg / kg, the 5 levels of X2 (crocetin) were between 10 - 50 mg / kg, the 5 levels of X3 (caffeic acid) were between 20 - 80 mg / kg, the 5 levels of X4 (ferulic acid) were between 20 - 80 mg / kg, the 5 levels of X5 (gallic acid) were between 25 - 100 mg / kg, and the 5 levels of X6 (quinic acid) were between 20 - 80 mg / kg. According to the uniform design table, each factor and level were arranged for the formula design, with a total of 10 groups (G1 - G10).

[0038] Table 1 is the drug administration dosage table selected by the uniform design scheme U 11 (11 6 )

[0039]

[0040] (3)Screening index: TG content in liver tissue;

[0041] (4)Statistical method: All data were statistically analyzed using the SPSS 26.0 software package. Stepwise regression analysis was used for the uniform design experiment. Measurement data in statistical description were expressed by , and count data were expressed by M(IQR). When the normality and homogeneity of variance were satisfied, comparison was made through t-test (comparison between two groups), and P < 0.05 was used as the significant level for statistical test.

[0042] 3. Results of the uniform design drug screening experiment:

[0043] The detection of TG content in liver tissues showed (as shown in Table 2) that the TG content in the model group reached 44.58 ± 4.82 mg / g, and its mean value was 3.9 times that of the normal group (11.53 ± 1.91 mg / g), with significant statistical differences between the two groups. This indicated that the model preparation was very successful, that is, the experimental model basis of the uniform design experiment was good.

[0044] The results of the triglyceride content in liver tissues and multiple statistical regression analysis are shown in Table 2 (G1 - G10, uniform design drug administration groups):

[0045] Table 2 shows the TG content in the liver tissues of mice in each group ( ).

[0046]

[0047] Note: ****P < 0.0001, vs normal group.

[0048] On the basis of the successful non - alcoholic fatty liver disease model, the TG content in liver tissues was detected. Taking the TG in liver tissues as the evaluation index and performing step - wise regression analysis, the results showed that when X1 and X4 were included in the model, the effect of reducing liver TG was the best and significant (P = 0.009) (as shown in Table 3). Therefore, the regression equation was obtained: Y = 51.176 - 0.005*X1X4. According to the coefficients, when both X1 and X4 were at the maximum doses, that is, when chlorogenic acid (X1) was 60 mg / kg and ferulic acid (X4) was 80 mg / kg, the inhibitory effect on the TG content in liver tissues was the best (Table 3 is the result of the step - wise regression of the uniform design). Combining the design of this example and the commonly used clinical doses, a compound with ferulic acid and chlorogenic acid combined in a mass ratio of 4:3 as the active ingredient was named "FC formula", which had the best effect on the non - alcoholic fatty liver disease model of mice induced by high - fat diet and high - sugar drinking water.

[0049] Table 3

[0050]

[0051] Example 2

[0052] The therapeutic effect of the screened FC formula was verified using the non - alcoholic fatty liver disease model of mice induced by high - fat diet and high - sugar drinking water

[0053] 1. Materials

[0054] (1) Animals: 64 male C57BL / 6J mice, clean - grade, weighing (21 - 25) g, purchased from Shanghai Jihui Experimental Animal Co., Ltd.

[0055] (2) Main reagents and drugs: Ferulic acid, CAS number: 537-98-4, batch number: 230812; Chlorogenic acid, CAS number: 327-97-9, batch number: 231208, purchased from Shanghai Ronghe Pharmaceutical Technology Development Co., Ltd. High-fat diet feed (60% of energy from fat), normal control feed (10% of energy from fat), the feed was purchased from Research Diets, USA. Drink sugar water with a concentration of 42 g / L (the sugar water ratio is 55% fructose and 45% sucrose). Triglyceride detection kit was purchased from Zhejiang Dong'ou Diagnostic Products Co., Ltd. ALT and AST detection kits, hematoxylin-eosin (HE) staining kit, and oil red O staining kit were purchased from Nanjing Jiancheng Bioengineering Institute.

[0056] 2. Methods

[0057] (1)Grouping and modeling: 64 male C57BL / 6J mice were given a high-fat diet and high-sugar drinking water for 18 weeks. There were 8 mice in the normal group and 56 mice in the model group. At the 12th week of modeling, they were randomly divided into the model group, the positive control drug Resmetirom group, the FC formula group (FCM group, chlorogenic acid 60 mg / kg, ferulic acid 80 mg / kg), the half-dose FC formula group (FCL group, chlorogenic acid 30 mg / kg, ferulic acid 40 mg / kg), the double-dose FC formula group (FCH group, chlorogenic acid 120 mg / kg, ferulic acid 160 mg / kg), the ferulic acid group (ferulic acid 80 mg / kg), and the chlorogenic acid group (chlorogenic acid 60 mg / kg) (8 mice in each group). They were given intragastric administration or drinking water to continue modeling until 18 weeks. The mice in the positive control drug Resmetirom group were given an intragastric dose of 5 mg / kg Resmetirom solution. The drugs in each group were prepared at corresponding concentrations and given intragastric administration at a dose of 0.1 ml / 10 g of mouse body weight. After 6 weeks of treatment, they were anesthetized by intraperitoneal injection of 3% sodium pentobarbital at a dose of 0.01 ml / g, blood was collected from the inferior vena cava, serum was separated by centrifugation, and liver tissue was taken for examination.

[0058] (2)Detection indicators: Serum ALT, AST; Liver TG; Liver tissue pathology (HE staining and oil red staining);

[0059] (3)Statistical methods: All data were statistically analyzed using the SPSS 26.0 software package. Measurement data in statistical description were expressed as , and count data were expressed as M (IQR). When the normality and homogeneity of variance were satisfied, comparison was made through t-test (comparison between two groups), and P < 0.05 was used as the significant level for statistical test.

[0060] 3. Results

[0061] Compared with the normal group, the body weight, liver tissue TG, serum ALT, and AST of the mice in the model group were significantly increased (P < 0.05). Compared with the model group, the body weight and liver tissue TG of the mice in the FCM group, FCH group, and Resmetirom group were significantly decreased (P < 0.05). At the same time, the serum ALT and AST of the mice in the FCL group, FCM group, FCH group, F group, C group, and Resmetirom group were significantly decreased (P < 0.05). Compared with the model group, FCL group, FCH group, F group, and C group, the body weight, liver tissue TG, serum ALT, and AST of the mice in the FCM group were lower than those in the model group, FCL group, FCH group, F group, and C group. At the same time, the liver tissue TG of the FCM group was significantly lower than that of the single-drug group C (P < 0.05). Compared with the positive control drug Resmetirom group, the body weight and liver tissue TG of the mice in the Resmetirom group were significantly lower than those in the FCL group, FCH group, F group, and C group (P < 0.05). However, there was no significant difference in the degree of decrease in the body weight, liver tissue TG, serum ALT, and serum AST of the mice in the FCM group and the positive control drug Resmetirom group. The results are shown in Table 4:

[0062] Table 4 Changes in serum ALT, AST activities and liver tissue TG content in mice of each group ( )

[0063]

[0064] Note: Compared with the normal group, #P < 0.0001. Compared with the model group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Compared with the FCM group, ●P < 0.05, ●●P < 0.01, ●●●P < 0.001, ●●●●P < 0.0001. Compared with the Resmetirom group, ▲P < 0.05, ▲▲P < 0.01, ▲▲▲P < 0.001, ▲▲▲▲P < 0.0001.

[0065] HE staining results showed that all model mice induced by high-fat and high-sugar diet had dense fatty degeneration of hepatocytes, aggregation and infiltration of inflammatory cells, and ballooning degeneration of some hepatocytes, indicating the success of the model. Compared with the normal group, the model group had significantly increased fatty degeneration of hepatocytes, inflammation within the lobules, ballooning degeneration of hepatocytes, and total NAS scores (P < 0.05). Compared with the model group, the FCM group and the Resmetirom group had significantly improved fatty degeneration of hepatocytes (P < 0.05). Compared with the model group, the FCL group, the FCM group, the FCH group, and the Resmetirom group had significantly reduced ballooning degeneration of hepatocytes (P < 0.05), and the FCL group, the FCM group, the FCH group, the F group, the C group, and the Resmetirom group had significantly reduced inflammation within the lobules and total NAS scores (P < 0.05). Compared with the FCM group, the FCL group, the FCH group, the F group, and the C group had significantly increased ballooning degeneration of hepatocytes (P < 0.05), and the FCL group, the FCH group, the F group, and the C group had significantly increased fatty degeneration of hepatocytes and total NAS scores (P < 0.05). Compared with the Resmetirom group, the FCL group, the FCH group, the F group, and the C group had significantly increased fatty degeneration of hepatocytes (P < 0.05), and at the same time, the FCL group, the F group, and the C group had significantly increased ballooning degeneration of hepatocytes and total NAS scores (P < 0.05), but there were no significant differences in each index between the FCM group and the positive control drug Resmetirom group. The results are shown in Table 5, Figure 1 as follows: Figure 1 It is a schematic diagram of the pathological observation results of the liver tissue of C57 mice in Example 2 (HE staining ×200), where 1, 2: normal group; 3, 4: model group; 5, 6: FCL group; 7, 8: FCM group; 9, 10: FCH group; 11, 12: F (ferulic acid) group; 13, 14: C (chlorogenic acid) group; 15, 16: Resmetirom group.

[0066] Figure 2 It is a schematic diagram of the pathological observation results of the liver tissue of C57 mice in Example 2 (oil red O staining ×200), where 1: normal group; 2: model group; 3: FCL group; 4: FCM group; 5: FCH group; 6: F (ferulic acid) group; 7: C (chlorogenic acid) group; 8: Resmetirom group. The oil red staining results also showed that the FC could significantly reduce the fatty degeneration of hepatocytes in non-alcoholic fatty liver model mice induced by high-fat diet and high-sugar drinking water.

[0067] Table 5 Comparison of NAS scores of liver tissues of mice in each group ( ).

[0068]

[0069] Note: Compared with the normal group, #P < 0.0001. Compared with the model group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Compared with the FCM group, ●P < 0.05, ●●P < 0.01, ●●●P < 0.001, ●●●●P < 0.0001. Compared with the Resmetirom group, ▲P < 0.05, ▲▲P < 0.01, ▲▲▲P < 0.001, ▲▲▲▲P < 0.0001.

[0070] The results showed that the FC formula of the present invention (a compound formula with ferulic acid and chlorogenic acid combined in a mass ratio of 4:3 as the active ingredient) has a significant therapeutic effect on non-alcoholic fatty liver disease mice induced by high-fat diet and high-sugar drinking water.

[0071] Example 3

[0072] The efficacy of the screened FC formula (a compound formula with ferulic acid and chlorogenic acid combined in a mass ratio of 4:3 as the active ingredient) was verified using a mouse non-alcoholic fatty liver model induced by methionine- and choline-deficient diet.

[0073] 1. Materials

[0074] (1) Animals: 64 male C57BL / 6J mice, clean grade, weighing (21 - 25) g, purchased from Shanghai Jihui Experimental Animal Co., Ltd.

[0075] (2) Main reagents and drugs: Ferulic acid (CAS number: 537-98-4, batch number: 230812); Chlorogenic acid (CAS number: 327-97-9, batch number: 231208); purchased from Shanghai Ronghe Medical Technology Development Co., Ltd. The methionine- and choline-deficient diet lacks choline and methionine, and the diet is purchased from Research Diets, Inc., USA. The triglyceride detection kit is purchased from Zhejiang Dong'ou Diagnostic Products Co., Ltd. The ALT, AST detection kits, hematoxylin-eosin (HE) staining kit, and oil red O staining kit are purchased from Nanjing Jiancheng Bioengineering Institute.

[0076] 2. Methods

[0077] (1)Grouping and model establishment: 64 male C57BL / 6J mice were given a diet deficient in methionine and choline for 6 weeks. There were 8 mice in the normal group and 56 mice in the model group. Starting from the second week of model establishment, the mice in the model group were randomly divided into the model group, the positive control drug Resmetirom group, the FC formula group (FCM group, chlorogenic acid 60 mg / kg, ferulic acid 80 mg / kg), the half-dose FC formula group (FCL group, chlorogenic acid 30 mg / kg, ferulic acid 40 mg / kg), the double-dose FC formula group (FCH group, chlorogenic acid 120 mg / kg, ferulic acid 160 mg / kg), the ferulic acid group (ferulic acid 80 mg / kg), and the chlorogenic acid group (chlorogenic acid 60 mg / kg) (8 mice in each group). They were given intragastric administration or drinking water to continue the model establishment until 6 weeks. The mice in the Resmetirom group were given an intragastric dose of 5 mg / kg Resmetirom solution. The drugs in each group were prepared at corresponding concentrations and given intragastric administration at a dose of 0.1 ml / 10 g of mouse body weight. After 4 weeks of treatment, they were anesthetized by intraperitoneal injection of 3% sodium pentobarbital at a dose of 0.01 ml / g. Blood was collected from the inferior vena cava, and the serum was separated by centrifugation. Liver tissues were taken for examination.

[0078] (2)Detection indexes: Serum ALT, AST; Liver TG; Liver tissue pathology (HE staining and oil red staining);

[0079] (3)Statistical method: All data were statistically analyzed using the SPSS 26.0 software package. The measurement data in statistical description were expressed as , and the count data were expressed as M (IQR). When the normality and homogeneity of variance were satisfied, comparison was made through t-test (comparison between two groups), and P < 0.05 was used as the significant level for statistical test.

[0080] 3. Results

[0081] Compared with the normal group, the body weight of mice in the model group was significantly lower than that in the normal group (P < 0.05), and the TG in the liver tissue, serum ALT and AST in the model group were significantly higher than those in the normal group (P < 0.05). Compared with the model group, in the FCL group, FCM group, FCH group, F group, C group, and Resmetirom group, the TG in the liver tissue of the FCM group decreased significantly (P < 0.05), the serum ALT in the FCM group and Resmetirom group decreased significantly (P < 0.05), and at the same time, the serum AST in the FCM group, FCH group, F group, C group, and Resmetirom group decreased significantly (P < 0.05). Compared with the FCM group, in the FCL group, FCH group, F group, and C group, the TG in the liver tissue, serum ALT, and AST in the FCM group were all lower than those in other drug-treated groups, and at the same time, the serum ALT in the FCM group was significantly lower than that in the single-drug-treated group C (P < 0.05). Compared with the Resmetirom group, in the FCL group, FCH group, and F group, the serum AST in the Resmetirom group was significantly lower than that in the FCL group, FCH group, and F group (P < 0.05). At the same time, the results showed that there was no significant difference in the degree of decrease in TG in the liver tissue, serum ALT, and serum AST between the FCM group and the positive control drug Resmetirom group. The results are shown in Table 6:

[0082] Table 6 Changes in serum ALT and AST activities and liver tissue TG content in mice of each group ( )

[0083]

[0084] Note: Compared with the normal group, #P < 0.0001. Compared with the model group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Compared with the FCM group, ●P < 0.05. Compared with the Resmetirom group, ▲P < 0.05, ▲▲P < 0.01, ▲▲▲P < 0.001, ▲▲▲▲P < 0.0001.

[0085] HE staining showed that varying degrees of hepatic steatosis, inflammatory cell infiltration, and ballooning degeneration of some hepatocytes were observed in the model mice induced by methionine- and choline-deficient diet, indicating the success of the model. Compared with the normal group, the model group showed significantly increased hepatic steatosis, intra-lobular inflammation, hepatocyte ballooning degeneration, and total NAS score (P < 0.05). Compared with the model group, the FCM group showed significantly improved hepatic steatosis (P < 0.05), and the FCL group, FCM group, C group, and Resmetirom group showed significantly reduced intra-lobular inflammation (P < 0.05). The FCL group, FCM group, FCH group, and Resmetirom group showed significantly reduced ballooning degeneration (P < 0.05), and the FCM group, FCH group, C group, and Resmetirom group showed significantly reduced total NAS score (P < 0.05). Compared with the FCM group, the FCL group, FCH group, C group, and F group showed higher indices of hepatic steatosis, intra-lobular inflammation, and hepatocyte ballooning degeneration in other drug-treated groups. The results are shown in Table 7, Figure 3 as shown Figure 3 is a schematic diagram of the pathological observation results of the liver tissue of C57 mice in Example 3 (HE staining ×200), where 1, 2: normal group; 3, 4: model group; 5, 6: FCL group; 7, 8: FCM group; 9, 10: FCH group; 11, 12: F (ferulic acid) group; 13, 14: C (chlorogenic acid) group; 15, 16: Resmetirom group.

[0086] Figure 4 is a schematic diagram of the pathological observation results of the liver tissue of C57 mice in Example 3 (Oil Red O staining ×200), where 1: normal group; 2: model group; 3: FCL group; 4: FCM group; 5: FCH group; 6: F (ferulic acid) group; 7: C (chlorogenic acid) group; 8: Resmetirom group. The Oil Red staining results also showed that the FC formula could significantly reduce hepatic steatosis in non-alcoholic fatty liver model mice induced by methionine- and choline-deficient diet.

[0087] Table 7 Comparison of NAS scores of liver tissues in each group of mice ( )

[0088]

[0089] Note: Compared with the normal group, #P < 0.0001. Compared with the model group, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. Compared with the FCM group, ●P < 0.05, ●●P < 0.01, ●●●P < 0.001, ●●●●P < 0.0001. Compared with the Resmetirom group, ▲P < 0.05, ▲▲P < 0.01, ▲▲▲P < 0.001, ▲▲▲▲P < 0.0001.

[0090] The results show that the FC formula of the present invention also has a significant therapeutic effect on mice with non-alcoholic fatty liver disease induced by methionine- and choline-deficient diet.

[0091] The above are only the preferred embodiments of the present invention, and there is no limitation to the present invention in any form. Although the present invention has been disclosed above with the preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art of this patent can make some changes or modifications to equivalent embodiments by using the technical content mentioned above within the scope of the technical solution of the present invention. However, as long as it does not depart from the content of the technical solution of the present invention, any simple modification, equivalent change and modification made to the above embodiments according to the technical essence of the present invention still fall within the scope of the present invention.

Claims

1. A Chinese medicinal composition for treating non-alcoholic fatty liver disease, comprising two Chinese medicinal ingredients, characterized in that: It is made of ferulic acid and chlorogenic acid.

2. The Chinese medicinal composition for treating non-alcoholic fatty liver disease, comprising two Chinese medicinal components, according to claim 1, characterized in that: The mass ratio of ferulic acid to chlorogenic acid is (1-10):

3.

3. The Chinese medicinal composition for treating non-alcoholic fatty liver disease composed of two Chinese medicinal ingredients according to claim 1, characterized in that: The mass ratio of ferulic acid to chlorogenic acid is 4:

3.

4. Use of the Chinese medicinal composition according to any one of claims 1 to 3 in the preparation of a medicament for treating non-alcoholic fatty liver disease.

5. Use of the Chinese medicinal composition according to any one of claims 1 to 3 in the preparation of a medicament for preventing and treating liver damage.

6. A pharmaceutical preparation, characterized in that The invention relates to a pharmaceutical preparation made from the Chinese medicinal composition according to any one of claims 1 to 3 and medically acceptable auxiliary materials.

7. The pharmaceutical preparation according to claim 6, characterized in that The dosage form of the pharmaceutical preparation is an oral preparation.

8. The pharmaceutical preparation according to claim 6, characterized in that The dosage form is selected from granules, tablets or capsules.

9. Use of the Chinese medicinal composition according to any one of claims 1 to 3 in the preparation of a medicament for treating non-alcoholic fatty liver disease.

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