CN111643533A

CN111643533A - Composition containing puerarin and application thereof

Info

Publication number: CN111643533A
Application number: CN202010737912.2A
Authority: CN
Inventors: 潘钰; 于冲; 夏海华; 张淑梅; 甄涛; 梁霆
Original assignee: Institute of Microbiology of Heilongjiang Academy of Sciences
Current assignee: Institute of Microbiology of Heilongjiang Academy of Sciences
Priority date: 2020-07-28
Filing date: 2020-07-28
Publication date: 2020-09-11

Abstract

The invention provides a composition containing puerarin and application thereof, belonging to the technical field of liver protection, wherein the composition comprises puerarin, curcumin and seabuckthorn flavone; the mass ratio of the puerarin to the curcumin to the seabuckthorn flavone is (1-2) to (1-2). The composition provided by the invention reduces the level of glutamic-oxaloacetic transaminase, the level of glutamic-pyruvic transaminase, the level of malondialdehyde and the level of triglyceride, improves the levels of superoxide dismutase and reduced glutathione, and further protects liver injury.

Description

Composition containing puerarin and application thereof

Technical Field

The invention belongs to the technical field of liver protection, and particularly relates to a puerarin-containing composition and application thereof.

Background

Alcoholic liver disease is a liver injury disease caused by excessive alcohol intake, seriously threatens the health of human bodies, and relates to a plurality of aspects such as oxidative stress, lipid peroxidation, immune inflammatory reaction and the like. Puerarin, curcumin and seabuckthorn flavone are effective components of kudzuvine root, turmeric and seabuckthorn in a list of articles which are both food and medicine and are published by the ministry of health, but the combination of the three is not reported to relieve liver injury.

Disclosure of Invention

In view of the above, the present invention aims to provide a composition containing puerarin and application thereof, and the composition provided by the present invention can protect liver injury.

In order to achieve the above purpose, the invention provides the following technical scheme:

the invention provides a composition containing puerarin, which comprises puerarin, curcumin and seabuckthorn flavone;

the mass ratio of the puerarin to the curcumin to the seabuckthorn flavone is (1-2) to (1-2).

The invention also provides application of the composition in the technical scheme in preparing a medicine for protecting liver injury.

Preferably, the liver damage is caused by carbon tetrachloride.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for reducing the content of glutamic-oxalacetic transaminase.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for reducing glutamic-pyruvic transaminase content.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for reducing the content of malondialdehyde.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for reducing the content of triglyceride.

The invention also provides application of the composition in the technical scheme in preparing a medicament for improving the content of superoxide dismutase.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for improving the content of reduced glutathione.

The invention provides a composition containing puerarin, which comprises puerarin, curcumin and seabuckthorn flavone; the mass ratio of the puerarin to the curcumin to the seabuckthorn flavone is (1-2) to (1-2). The composition provided by the invention reduces the level of glutamic-oxaloacetic transaminase, the level of glutamic-pyruvic transaminase, the level of malondialdehyde and the level of triglyceride, improves the levels of superoxide dismutase and reduced glutathione, and further protects liver injury.

Drawings

FIG. 1 shows the CCl of each pair₄Acute liver injury mice liver index, note: comparison with Control group<0.01; comparison with Model group, # p<0.05 and # # p<0.01; the comparison between the connection line groups is carried out,&p<0.05 and&&p<0.01；

FIG. 2 shows the CCl of each pair₄Effect of serum alt (a), ast (b) activity in mice with acute liver injury, injections: comparison with Control group<0.01; comparison with Model group, # p<0.05 and # # p<0.01；

FIG. 3 shows the CCl of each pair₄Effects of liver mda (a), sod (b) and gsh (c) activities in mice with acute liver injury, note: comparison with Control group<0.01; comparison with Model group, # p<0.05 and # # p<0.01; the comparison between the connection line groups is carried out,&p<0.05 and&&p<0.01；

FIG. 4 shows the CCl of each pair₄Effect of hepatic tg (a) and tcho (b) content in mice with acute liver injury, note: comparison with Control group<0.01; comparison with Model group, # p<0.05 and # # p<0.01; the comparison between the connection line groups is carried out,&p<0.05 and&&p<0.01；

FIG. 5 is the transmission electron microscope scanning image of the mouse livers of the control group, the carbon tetrachloride model group, the curcumin group and the compound group.

Detailed Description

The invention provides a composition containing puerarin, which comprises puerarin, curcumin and seabuckthorn flavone; the mass ratio of the puerarin to the curcumin to the seabuckthorn flavone is (1-2) to (1-2). The sources of the puerarin, the curcumin and the seabuckthorn flavone are not specially limited, and the puerarin, the curcumin and the seabuckthorn flavone are prepared by adopting conventional commercial products.

The invention also provides application of the composition in the technical scheme in preparing a medicine for protecting liver injury. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted. In the present invention, the liver damage is preferably caused by carbon tetrachloride.

The invention also provides application of the composition in the technical scheme in preparing a medicine for reducing the content of glutamic-oxalacetic transaminase. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted.

The invention also provides application of the composition in the technical scheme in preparation of medicines for reducing glutamic-pyruvic transaminase content. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for reducing the content of malondialdehyde. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted.

The invention also provides application of the composition in the technical scheme in preparing a medicine for reducing the content of triglyceride. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted.

The invention also provides application of the composition in the technical scheme in preparing a medicament for improving the content of superoxide dismutase. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted.

The invention also provides application of the composition in the technical scheme in preparation of a medicine for improving the content of reduced glutathione. The invention has no special limitation on the dosage form of the medicine, and the invention adopts the medically acceptable dosage forms of puerarin, curcumin and seabuckthorn flavone. The contents of puerarin, curcumin and seabuckthorn flavone in the medicine are not particularly limited, and the active substances in the conventional medicine are adopted.

The technical solutions provided by the present invention are described in detail below with reference to examples, but they should not be construed as limiting the scope of the present invention.

Example 1

1.1 test materials and apparatus

1.1.1 materials

240 Kunming mice, clean grade, 6-8 weeks old, half male and half female, 18-22g weight, provided by the safety evaluation center of Heilongjiang university of traditional Chinese medicine. Animal production license number: SCXK black 20180004.

Experimental mouse feeding conditions: mice were housed in an animal room with free access to food and water at 55% (+ -10%) relative humidity, 22 ℃ (+ -2 ℃) and a light/dark cycle of 12 hours; feeding complete nutrition pellet feed (provided by the laboratory animal center of the university of traditional Chinese medicine in Heilongjiang); the drinking water is sterilized tap water; all procedures were in accordance with the guidelines for care and use of experimental animals.

Long forceps, ophthalmic forceps, medical gloves, syringe (1mL), gauze (1.5cm × 1.5 cm).

1.1.2 Main instruments and devices

An electronic balance: BSA124S-CW model, Sidolisi scientific instruments, Beijing;

low-temperature freezing centrifuge: d-37520 type, Osterode, Germany;

an air bath oscillator: model HZQ-C, Tommin medical instruments, Harbin.

An enzyme-labeling instrument:

a micropipette:

a vortex mixer:

independent air supply system squirrel cage:

1.1.3 reagents

Puerarin, curcumin from Saian ruilin science and technology Limited; the seabuckthorn flavone is extracted and purchased from Shanxi Guanchen Biotech limited company in laboratory; ALT (Alanine Transaminase glutamic pyruvic Transaminase), AST (Aspartame Transaminase), MDA (Maleic Dialdehyde), SOD (Superoxide Dismutase), GSH (Glutathione reduced Glutathione), TG (Triglyceride) and TCSO (Total Cholesterol) determination kits are purchased from Jiangsu Nanjing Bioengineering institute; the Bradford protein concentration determination kit is purchased from Beijing Solaibao science and technology Limited; CCl₄(Carbon tetrachlororide Carbon Tetrachloride) was purchased from Tianjin Rich China chemical Co., Ltd.

1.2 animal experiments

1.2.1 model of acute carbon tetrachloride liver injury

60 mice were weighed and randomly divided into 6 groups, i.e., a blank Control group (Control), a carbon tetrachloride Model group (Model), a Puerarin group (Puerarin, abbreviated as Pue), a Curcumin group (Curcumin, abbreviated as Cur), a Seabuckthorn flavone group (Seabuckthorn flavone, abbreviated as Seaf), a complex group (Puerarin, Curcumin and Seabuckthorn flavone in a mass of 1:1:1, abbreviated as Pue + Cur + Seaf), a Puerarin group (Pue), a Curcumin group (Cur), and a Seabuckthorn flavone group (Seaf)The materials were dissolved in distilled water separately and orally administered to mice using round-head feeding needles. The mouse in the puerarin, curcumin and seabuckthorn flavone groups is administrated with corresponding materials with the dose of 300mg/kg body weight every day, the doses of the three materials in the compound group are all 100 mg/kg-bw (the total amount is 300 mg/kg-bw), and the mice in the blank control group and the carbon tetrachloride model group receive distilled water with the same volume continuously for 15 days. On day 15, acute liver injury was induced by gavage of CCl₄(0.4ml, 40% in olive oil) model was established, placebo mice were gavaged with an equal amount of olive oil, after 16h without water deprivation, weighed, blood was collected using ophthalmic forceps, mice were sacrificed using cervical dislocation, the abdominal cavity was dissected open to separate the whole liver and weighed, the liver/body weight ratio was calculated, i.e., liver index (100%): liver weight/mouse weight × 100%, after the mice blood was allowed to stand for 2 hours, serum samples were obtained by centrifugation (3000rpm, 10 minutes) and stored at-80 ℃ until assayed, liver tissue was harvested and a portion of the liver samples were fixed in 2.5% glutaraldehyde solution for histological analysis, the remaining liver samples were immediately immersed in liquid nitrogen and stored at-80 ℃ until use.

1.3 protein concentration determination

1.3.110% preparation of liver homogenate

Taking 0.1g-0.4g of mouse liver tissue block, accurately weighing, adding precooled physiological saline with the volume 9 times of the mass of the tissue block, homogenizing on ice by using a glass homogenizer, and fully grinding to homogenize the tissue. And (3) diluting the homogenate with a certain volume, measuring the protein concentration, centrifuging the rest homogenate for 10 minutes at a low-temperature centrifuge of 2000r/min, and taking the supernatant of the centrifuged homogenate for various measurements.

1.3.2Bradford kit for determination of protein concentration in homogenate

1. The protein standards were completely solubilized, 10ul was taken and diluted to 250ul with 0.9% NaCl to a final concentration of 0.2 mg/ml.

2.5 XG 250 staining solution before using 3-5 times of mixing, 1ml of 5 XG 250 staining solution, adding 4ml of double distilled water, mixing to obtain 1 XG 250 staining solution.

3. The standard is added into a 96-well plate according to 0,2,4,6,8,12,16 and 20 microliter, and PBS diluent is added to make up to 20 microliter.

4. The samples were diluted 200 fold (this was done in the previous experiments) and 20 μ l was added to the sample wells of a 96 well plate.

5. 200. mu.l of diluted 1 XG 250 staining solution was added to each well, and the mixture was allowed to stand at room temperature for 3 to 5 minutes.

6. A595 was measured with a microplate reader.

7. The protein concentration in the sample was calculated from the standard curve.

1.4 serum enzyme assay

The contents of alanine Aminotransferase (ALT) and aspartate Aminotransferase (AST) in serum are determined according to the kit instructions. Adding corresponding reagents according to an operation table and performing operation.

TABLE 1 corresponding reagents

	Assay well	Control well
			Matrix solution (. mu.l) pre-warmed at 37 ℃	20	20
Sample to be tested (ul)	5

When one sample is sucked by the measuring hole, the suction nozzle is deeply inserted into the substrate liquid at the bottom of the hole plate, and after repeated sucking, stirring and mixing, water bath or air bath at 37 ℃ is carried out for 30 minutes.

TABLE 2 addition

	Assay well	Control well
			2-4-dinitrophenylhydrazine solution (mu l)	20	20
Sample to be tested (ul)		5

When one sample is sucked in the control hole, the suction nozzle is inserted into the liquid at the bottom of the hole plate, and the sample is repeatedly sucked, beaten and mixed evenly and then is subjected to water bath or air bath at 37 ℃ for 20 minutes.

TABLE 3 addition

	Assay well	Control well
			0.4mol/L sodium hydroxide solution (. mu.l)	200	200

Gently horizontally shaking the 96-well plate, mixing, standing at room temperature for 15 minutes, measuring the wavelength of 510nm, measuring the OD value of each well by a microplate reader, and searching a standard curve to obtain the corresponding ALT/GPT activity unit (absolute OD value is the OD value of the measurement well minus the OD value of the control well).

Calculating the formula: the ALT viability (U/gprot) in the tissue is determined by the standard curve ALT viability (U/L)/protein concentration of the homogenate to be measured (gprot/L).

The AST activity (U/gprot) in the tissue is determined by the standard curve as AST activity (U/L)/protein concentration of homogenate to be measured (gprot/L).

1.5 liver index determination

1.5.1 assay of Malondialdehyde (MDA)

TABLE 4 reagents

Mixing (shaking table centrifuge tube rack)

TABLE 5 reagents

	Blank tube	Standard tube	Measuring tube	Control tube
					Reagent two (mL)	3	3	3	3
Reagent three (mL)	1	1	1
					50% glacial acetic acid (mL)				1

Covering the centrifuge tube with a cover, pricking a small hole on the cover with a needle, mixing with a vortex mixer, water-bathing at 95 deg.C for 40 min, taking out, cooling with running water, centrifuging at 3000rpm for 10 min, collecting supernatant 200 μ l, and measuring A532.

Calculating the formula: MDA activity (nmol/mgprot) ═ assay OD value-control OD value/standard OD value-blank OD value × standard concentration (10nmol/mL) ÷ test sample protein concentration (mgprot/mL) in the tissue.

1.5.2 determination of superoxide dismutase (SOD) Activity

TABLE 6 reagents

Mix well, incubate 20 minutes at 37 ℃, read at 450nm with microplate reader.

Calculating the formula: SOD inhibition (%) (a control-a control blank) - (a assay-a assay blank)/(a control-a control blank) × 100

SOD activity (U/mgprot) ═ SOD inhibition (%)/, 50% × reaction system (0.24 ml)/dilution factor (0.02 ml)/, sample protein concentration to be measured (mgprot/ml)

1.5.3 measurement of reduced Glutathione (GSH) content

TABLE 7 reagents

	Blank hole	Standard hole	Assay well
				Reagent one (ul)	100
20 μmol/L GSH Standard (μ L)		100
				Supernatant (ul)			100
Reagent two (ul)	100	100	100
				Reagent three (ul)	25	25	25

Mixing, standing for 5 min at 405nm, and measuring absorbance of each well with enzyme-labeling instrument

Calculating the formula: cell and tissue GSH content (μmol/gprot) ═ determination of OD value-blank OD value/standard OD value-blank OD value × standard tube concentration (20 μmol/L) × sample pretreatment dilution (2 times) ÷ concentration of homogenate protein to be determined (gprot/L)

1.5.4 determination of Triglyceride (TG) and Total Cholesterol (TCHO) content

Adding corresponding reagents according to an operation table

TABLE 8 reagents

Mixing, incubating at 37 deg.C for 10 min, wavelength 510nm, and measuring absorbance of each well with enzyme-labeling instrument.

Calculating the formula: triglyceride content (mmol/gprot) ═ sample OD value-blank OD value/calibration OD value-blank OD value × calibrator concentration (mmol/L) ÷ protein concentration of the sample to be tested (gprot/L).

Cholesterol content (mmol/gprot) ═ sample OD value-blank OD value/calibration OD value-blank OD value × calibrator concentration (mmol/L) ÷ protein concentration of sample to be tested (gprot/L)

1.6 histopathological Observation

Cutting mouse liver into 0.5-1mm³The sample block is immersed in 2.5% glutaraldehyde solution, and after ultrathin sections are made, each group of liver sections is observed by a projection electron microscope.

1.7 statistical treatment

The data are expressed as x ± s, and statistical analysis using graphpadprism6.0 using one-way analysis of variance or ANOVA test, with a significant statistical difference of P < 0.05.

1.8 best formula exploration test

According to the previous test results, the compound is preliminarily considered to have a certain liver protection effect and have a better effect than the materials in the formula, so that the optimal proportion is further researched to carry out an optimal formula research test.

1.8.1 CCl₄Optimal group prescription exploration test for causing acute liver injury of mice

60 mice were weighed and randomly divided into 6 groups, namely a blank Control group (Control), a carbon tetrachloride Model group (Model), and a compound group (puerarin, curcumin, and seabuckthorn flavone contents 1:1:1, abbreviated as 1Pue +1Cur +1 Seaf; contents of the three groups 2:1:1, abbreviated as 2Pue +1Cur +1 Seaf; contents of the three groups 1:2:1, abbreviated as 1Pue +2Cur +1 Seaf; contents of the three groups 1:1:2, abbreviated as 1Pue +1Cur +2 Seaf). The specific operation of the animal experiment is consistent with 1.2.1. The ALT and AST in the serum of the mouse and the MDA and SOD content in the liver are determined by the same operation method as 1.3-1.5.

2 results and analysis

2.1 Compound Pair CCl₄Effect of acute liver injury in mice

2.1.1 Compound Pair CCl₄Effect of acute liver injury mouse liver index

The liver body weight ratio, i.e., the liver index, is a general indicator of positive correlation with the degree of liver damage. Compared with a blank control group, the serum liver index of the mice in the carbon tetrachloride model group is obviously increased (p < 0.05). The success of the model building of the mouse carbon tetrachloride acute liver injury model is shown. The liver index of mice with acute liver injury caused by carbon tetrachloride (p <0.05 or p <0.01) can be obviously reduced in each group; compared with puerarin group and fructus Hippophae flavone group, the compound group can significantly reduce liver index (p <0.05) (FIG. 1).

2.1.2 Compound Pair CCl₄Influence of serum indexes AST and ALT activity of mice with acute liver injury

Compared with the blank group, the serum ALT and AST of the mouse in the carbon tetrachloride model group are obviously increased (p is less than 0.01), which indicates that the model building of the mouse carbon tetrachloride acute liver injury model is successful. The puerarin group and the curcumin group can obviously reduce serum ALT and AST (p is less than 0.05 or less than 0.01) of mice with acute liver injury caused by carbon tetrachloride. The seabuckthorn flavone group can obviously reduce the AST (p is less than 0.01) of mice with acute liver injury caused by carbon tetrachloride, and has no obvious difference (p is more than 0.05) in reducing ALT.

The compound formula is found to reduce the trend of increasing ALT and AST activities in mouse serum, and the difference is significant compared with a model group (P <0.01) (figure 2).

2.1.3 Compound Pair CCl₄Effect of MDA, SOD and GSH levels in liver tissue of mice with acute liver injury

The MDA content of the liver homogenate of the model group mouse is higher than that of the control group, and the activities of SOD and GSH are lower than those of the control group (P is less than 0.01), which indicates that the model building of the experiment is successful. The MDA content of mouse liver homogenate in the puerarin group, the curcumin group, the seabuckthorn flavone group and the compound group is lower than that in the model group (the puerarin, the curcumin and the compound group P is less than 0.01, and the seabuckthorn flavone group P is less than 0.05), and the compound group has obviously better MDA content reducing effect than other groups; SOD activity and GSH level in liver tissue homogenate of puerarin group, curcumin group and compound group are obviously higher than those of model group (curcumin, compound group P is less than 0.01, puerarin group P is less than 0.05); compared with the model group, the SOD activity of the seabuckthorn flavone group has no statistical significance (P >0.05), and the GSH level is obviously increased (P <0.05) compared with the model group; the compound composition has relatively high capacity of improving SOD activity and GSH content (figure 3).

FIG. 3. Each pair of CCl₄Acute liver injury mice liver MDA (A), SOD (B) and GSH (C) activity effect.

2.1.4 Compound Pair CCl₄TG and TCHO in liver tissue of mice with acute liver injuryInfluence of the quantity

Compared with the control group, the TG content in the liver of the mouse with the acute liver injury caused by carbon tetrachloride is obviously increased (p is less than 0.01), and the TG content in the liver of the mouse with the acute liver injury caused by carbon tetrachloride can be obviously reduced in the puerarin group, the seabuckthorn flavone group and the compound group (p is less than 0.05 or p is less than 0.01). The compound preparation has the most obvious effect. There was no significant difference in TCHO content for each group (figure 4).

2.1.5 CCl₄Acute liver injury mouse liver histopathology

The scanning image of the transmission electron microscope shows that the hepatocyte nucleus of the blank control group mouse is complete, a small amount of lipid droplets exist, and the mitochondria are normal; mouse liver cells in the carbon tetrachloride model group have the defects of nuclear shrinkage, lipid droplet increase and mitochondrial degeneration; the single material treatment group selects a curcumin group (Cur) with relatively good liver protection effect, and a transmission electron microscope image shows that mouse liver cells in the curcumin group have obvious nuclear compaction symptoms; the mice in the combination group had increased lipid droplets of hepatocytes, indicating that the hepatocytes were slightly damaged (fig. 5).

Carbon tetrachloride is an exogenous compound of a classical experimental liver injury inducing animal model, is a toxic agent of a typical chemical liver injury animal model, and has the mechanism that a large amount of free radicals CCl are generated in the metabolic process₃And CCl₃OO, which generates a highly toxic dichloromethyl radical (. CCl) upon intrahepatic activation₂) And a peroxidized methyl radical (. OOCC 1)₂) The phospholipid molecules on the liver cell membrane can be damaged to cause lipid peroxidation, the structure of the liver cell membrane is damaged, the permeability of the liver cell membrane is increased, and the purpose of liver injury is achieved.

After carbon tetrachloride enters the liver, free radicals are generated after the activation of liver microsomes to cause peroxidation, so that liver cells are damaged, a membrane structure is damaged, ALT and AST in lipid enter blood, and the activity of ALT and AST in serum is increased; the mouse serum ALT and AST content of the carbon tetrachloride model group is obviously increased compared with the control group, and the mouse serum ALT and AST of the acute liver injury caused by carbon tetrachloride can be obviously reduced by the puerarin group, the curcumin group and the compound group, so that the substances can play a role of relieving liver injury, and the liver protection effect of the seabuckthorn flavone is relatively weak. Meanwhile, the results of a scanning transmission electron microscope show that mouse liver cells in a carbon tetrachloride model group have the characteristics of nuclear compaction, mitochondrial degeneration and lipid droplet increase which are typical damaged liver cells; the result of the compound group transmission electron microscope shows that the liver cells have slight damage, which indicates that the compound has a certain liver protection function.

Oxidative stress is an unbalanced disorder caused by the production and removal of Reactive Oxygen Species (ROS). On the one hand, reactive oxygen species, as a by-product of oxidative metabolism, often damage cellular macromolecules such as DNA, lipids and proteins. The burst of ROS can cause lipid peroxidation, manifested as an increase in MDA levels. In the invention, the content of malondialdehyde in liver is obviously increased by carbon tetrachloride, the compound group obviously inhibits the production of MDA, the effect is obviously improved compared with the single action of puerarin, curcumin and seabuckthorn flavone, and the compound can alleviate the peroxidation injury of liver cells. In the present invention, carbon tetrachloride significantly reduces the amount of glutathione, the most important molecule involved in cellular antioxidant defense, and reduced GSH maintains the structural integrity of cells and organelles by scavenging some peroxides, combining with toxic foreign electrophilic species. Simultaneously, the activity of antioxidant enzyme (SOD) is reduced. The compound group shows that the compound has stronger antioxidation, increases the concentration of glutathione and increases the activity of SOD enzyme. The results of the invention show that the TG value of the liver is obviously increased compared with the control group in the carbon tetrachloride model group, which shows that carbon tetrachloride obviously affects the fat metabolism of liver cells. The TCHO content is not obviously different, and the reason for influencing the metabolism of 2 substances is different, which indicates that the liver lipid metabolism is disordered, and compared with the model group, the compound group obviously reduces the TG content and plays a role in stabilizing the lipid metabolism.

The better liver protection effect of the compound group is the result of mutual synergistic effect of a plurality of components, the action mechanism of the traditional Chinese medicine for resisting oxidative stress and hepatic fibrosis is characterized by multiple links, multiple ways and multiple targets, and relates to a plurality of cytokines and intracellular signal molecule networks. The compound can better eliminate active oxygen free radicals in vivo and prevent the organism from generating lipid peroxidation, so that the antioxidant enzyme SOD can better play the role of antioxidation. Further reducing GSH consumption, and the increase of GSH synthesis in cells can obviously inhibit the activation of HSC.

The result of the optimum proportion exploration test shows that CCl₄In a mouse acute liver injury model, the liver protection effect of the 1Pue +2Cur +1Seaf group is better than that of other proportion groups; in the ethanol induced acute liver injury model of mice, the group 2Pue +1Cur +1Seaf is numerically superior to other groups. Supposing curcumin is in CCl₄The protective effect of the curcumin on the mouse liver injury plays a more critical role, the curcumin has a better protective effect on liver fibrosis, the liver fibrosis is closely related to the generation of stellate cells (HSCs), and the research on the mechanism of the HSCs mainly focuses on the aspects of inhibiting the proliferation of the HSCs, inducing the apoptosis of the HSCs and the like at present. In normal liver tissues, HSCs are in a quiescent state, with a large number of lipid droplets within the cells; during fibrosis, HSCs are activated to produce large amounts of extracellular matrix and intracellular lipid droplets are lost.

The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.

Claims

1. A composition containing puerarin is characterized in that the composition comprises puerarin, curcumin and seabuckthorn flavone;

2. Use of the composition of claim 1 for the manufacture of a medicament for protecting against liver damage.

3. The use according to claim 2, wherein the liver damage is caused by carbon tetrachloride.

4. Use of a composition according to claim 1 for the manufacture of a medicament for reducing the level of aspartate aminotransferase.

5. Use of a composition according to claim 1 for the manufacture of a medicament for reducing glutamate pyruvate transaminase levels.

6. Use of the composition of claim 1 for the manufacture of a medicament for reducing the level of malondialdehyde.

7. Use of a composition according to claim 1 for the manufacture of a medicament for lowering triglyceride levels.

8. Use of the composition of claim 1 for the manufacture of a medicament for increasing the level of superoxide dismutase.

9. Use of the composition of claim 1 in the manufacture of a medicament for increasing reduced glutathione content.