Application of liquidambar formosana acid in preparation of anti-hepatitis B virus medicine

Abstract

The invention discloses an application of liquidambar formosana acid in preparation of anti-hepatitis B virus drugs, the liquidambar formosana acid can inhibit the replication of hepatitis B viruses, inhibit the generation and replication of hepatitis B virus antigens, and inhibit the expression of hepatitis B virus mRNA and HBV transcription factors, has a significant inhibitory effect on hepatitis B viruses, and can be used for preparing anti-hepatitis B virus drugs. A new scheme is provided for development of new anti-hepatitis B virus drugs, and the liquidambar formosana acid serving as a traditional Chinese medicinal material is rich in medicinal resource and low in cost and has important significance in research and development of the novel anti-hepatitis B virus drugs.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and in particular relates to the application of a traditional Chinese medicine compound lulutong acid in the preparation of an anti-hepatitis B virus drug. Background Art

[0002] Hepatitis B is an infectious liver disease caused by the hepatitis B virus (HBV). Its main transmission routes include blood transmission, mother-to-child transmission and sexual transmission. After the hepatitis B virus enters the liver cells, through the processes of viral replication, transcription and assembly, the virally replicated hepatitis B surface antigen, hepatitis B e antigen and hepatitis B core antigen are released on the liver cell membrane, stimulating the human immune system to identify and react. This antigen-antibody reaction on the liver cell membrane can cause damage and destruction of liver cells, resulting in a series of clinical symptoms, which can cause chronic infection and a high risk of developing cirrhosis and liver cancer.

[0003] At present, the commonly used drugs for the treatment of hepatitis B virus mainly include nucleoside analogs, interferon drugs, etc., but these drugs either have a long treatment cycle, are prone to cause drug-resistant mutations of HBV virus, or have more adverse reactions. Therefore, it is urgent to continuously develop new therapies and safer and more effective drugs on the existing basis to achieve functional cure of HBV. As an effective choice for disease treatment, Chinese herbal medicines have a significant effect on disease treatment, whether it is the use of prescriptions or the application of monomer compounds. At present, there are many schemes that have proposed anti-hepatitis B virus prescriptions or monomer compounds, which can not only improve the patient's liver function and regulate the body's immune function, but also have relatively few side effects, and patients have good compliance and tolerance.

[0004] Betulonic acid is a triterpenoid compound isolated from the Chinese herbal medicine Liquidambar formosana Hance (the dried mature infructescence of Liquidambar formosana Hance, a plant of the Hamamelidaceae family). Its molecular formula is C 30 H 46 O3, molecular weight is 454.34, structural formula is as follows:

[0005]

[0006] Studies have shown that lulutong acid has multiple biological effects such as anti-inflammatory, analgesic, antibacterial, and anti-tumor.

[0007] However, there are currently no research reports on the inhibitory effects of lumefantrine on HBV replication and transcription, as well as its anti-HBV effects. Summary of the invention

[0008] In view of this, the primary purpose of the present invention is to provide the use of passutonic acid in the preparation of anti-hepatitis B virus drugs. The present invention determines that passutonic acid has a significant inhibitory effect on hepatitis B virus (HBV), providing a new solution for the development of new drugs against hepatitis B virus. Passutonic acid, as a traditional Chinese medicinal material, has abundant medicinal resources and low cost, and is of great significance to the development of anti-hepatitis B virus drugs.

[0009] In order to achieve the above object, the present invention adopts the following technical solutions:

[0010] One aspect of the present invention provides the use of lulutonic acid in the preparation of anti-hepatitis B virus drugs.

[0011] The drug has at least one of the following effects:

[0012] a: Inhibit the replication of hepatitis B virus;

[0013] b: Inhibit the production and replication of hepatitis B virus antigen;

[0014] c: Inhibit the expression of hepatitis B virus mRNA;

[0015] d: Inhibit the expression of HBV transcription factors;

[0016] e: Treatment of hepatitis B.

[0017] Another aspect of the present invention provides an anti-HBV drug containing an effective concentration of lulutonic acid.

[0018] Beneficial effects of the present invention:

[0019] The present invention has been tested and confirmed through a large number of experiments that lulutong acid has a significant inhibitory effect on hepatitis B virus (HBV). Lulutong acid can inhibit the production and replication of hepatitis B virus antigens, inhibit the expression of hepatitis B virus mRNA and inhibit the expression of HBV transcription factors. It is also confirmed through a hepatitis B mouse model that lulutong acid has the effect of treating and controlling hepatitis B, providing a new idea for the development of hepatitis B drugs.

[0020] In addition, as a traditional Chinese medicine compound, lulutong acid has a wide source, is easy to extract, and has low cost. More importantly, it has low cytotoxicity and has a significant inhibitory effect on HBV at lower drug concentrations, which is of great significance for the development of new anti-hepatitis B virus drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 This is the result of the CCK8 cytotoxicity test in Example 1 to detect the toxicity of lulutonic acid on HepG2.215 cells.

[0022] Figure 2The results of flow cytometry determination of the drug toxicity of lulutonic acid on HepG2.215 cells in Example 2.

[0023] Figure 3 The results of the enzyme-linked immunosorbent assay in Example 3 were as follows: the inhibition of HBsAg and HBeAg production by HepG2.215 cells using lulutonic acid.

[0024] Figure 4 This is the result of real-time fluorescence quantitative determination of the inhibition of mRNA expression in HepG2.215 cells by lulutong acid in Example 4.

[0025] Figure 5 This is the result of Western blot analysis of the inhibition of HBV transcription factor expression by lulutonic acid in Example 5.

[0026] Figure 6 The results are as follows: the expression changes of HBsAg in the serum of mice after the hepatitis B mouse model was treated with lulutong acid in Example 6.

[0027] Figure 7 These are the results of immunohistochemical detection of HBcAg and HBsAg in the liver of mice after the hepatitis B mouse model was treated with lulutong acid in Example 6. DETAILED DESCRIPTION

[0028] Embodiments of the present invention are described in detail below. The embodiments described below are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0029] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art of the present invention. The terms used in the specification of the present invention herein are only for the purpose of describing specific embodiments and are not intended to limit the present invention.

[0030] The first aspect of the present invention provides the use of lulutonic acid in the preparation of anti-hepatitis B virus drugs.

[0031] Traditional Chinese medicine ingredients have natural advantages in treating hepatitis B and resisting hepatitis B virus. Traditional Chinese medicines are widely available, have low cost, and have little toxicity and side effects, and have better tolerance and compliance for patients. Therefore, screening traditional Chinese medicine ingredients with anti-hepatitis B virus effects is of great significance for the research and development of hepatitis B virus drugs. The inventors conducted a large number of experiments to screen drugs in the early stage, and finally determined that lulutong acid has a significant inhibitory effect on hepatitis B virus, which can be used to develop anti-hepatitis B virus drugs.

[0032] The second aspect of the present invention further provides an anti-hepatitis B virus drug, which contains an effective concentration of lulutonic acid.

[0033] In the present invention, the drug is an anti-HBV drug, which has at least one of the following effects a to e:

[0034] a: Inhibit the replication of hepatitis B virus;

[0035] b: Inhibit the production and replication of hepatitis B virus antigen;

[0036] c: Inhibit the expression of hepatitis B virus mRNA;

[0037] d: Inhibit the expression of HBV transcription factors;

[0038] e: Treatment of hepatitis B.

[0039] The present invention has determined through in vitro cell experiments and animal model experiments that lulutong acid can inhibit the replication of hepatitis B virus, inhibit the production and replication of hepatitis B virus antigens, inhibit the expression of hepatitis B virus mRNA and inhibit the expression of HBV transcription factors, thereby exerting a significant HBV inhibitory effect. In the present invention, the mechanism of lulutong acid against hepatitis B virus is clear, and it can treat hepatitis B and slow down the development of hepatitis B.

[0040] In the present invention, the hepatitis B virus antigen is at least one of HBsAg, HBeAg, and HBcAg.

[0041] The HBV transcription factor is at least one of PPARγ, PGC1α, HNF4α and HNF1α.

[0042] The term "effective concentration" refers to the lowest concentration at which the drug component begins to exert a therapeutic effect. In the present invention, the effective concentration is preferably the concentration at which the drug component exerts a therapeutic effect with minimal toxicity. The said exerting a therapeutic effect means that the condition of the subject is suppressed or alleviated after receiving the drug. In the present invention, it means that the replication of the hepatitis B virus is slowed down or suspended; or the hepatitis B virus antigen stops being produced, slows down or stops replicating; or the expression of the hepatitis B virus mRNA begins to decrease or is not expressed; or the expression of the HBV transcription factor begins to decrease or is not expressed; so that the subject's hepatitis B condition is controlled, alleviated or even cured.

[0043] In some specific embodiments of the present invention, the effective concentration of lulutonic acid is 5 μM to 30 μM, for example, it can be any concentration of 5 μM, 10 μM, 15 μM, 20 μM, 25 μM or 30 μM.

[0044] Preferably, the effective concentration of the lulutonic acid is 5 μM to 20 μM, for example, it can be any concentration of 5 μM, 10 μM, 15 μM, or 20 μM.

[0045] Preferably, the effective concentration of the lulutong acid is 10 μM to 20 μM. Within this concentration range, it has a significant inhibitory effect on HBV and has extremely low toxicity.

[0046] In some specific embodiments of the present invention, the effective concentration of lulutonic acid is 10 μM or 20 μM.

[0047] More preferably, the effective concentration of the lulutonic acid is 20 μM.

[0048] It is understood that the anti-HBV drug described in the present invention also includes any pharmaceutically acceptable excipients and / or carriers. These excipients and / or carriers are auxiliary components used in pharmaceutical preparations, preferably, some components that can promote the drug absorption ability of the subject, improve the biological activity of the drug, or reduce the side effects of the drug. For example, it can be a solvent (such as water, ethanol, etc.), a solubilizer or co-solvent (such as cyclodextrin, etc.), an emulsifier (such as lecithin, etc.), a colorant, a binder (such as starch, etc.), a disintegrant (such as cross-linked sodium carboxymethyl cellulose, etc.), a filler (such as lactose, etc.), a lubricant (such as magnesium stearate, etc.), a wetting agent (such as polysorbate, etc.), an osmotic pressure regulator (such as sodium chloride, etc.), a stabilizer (such as ascorbic acid, etc.), a glidant (such as colloidal silicon dioxide, etc.), an anti-caking agent (such as silicate, etc.), a flavoring agent (such as sodium saccharin, etc.), an antioxidant (such as tocopherol, etc.), a gelling agent (such as gelatin, etc.), a buffer (such as phosphate buffer, etc.), and a carrier material (such as microcrystalline cellulose, etc.), but it is not limited thereto. Any pharmaceutically acceptable excipient and / or carrier in the art can be used, and those skilled in the art can make a selection based on the pharmaceutical common sense in the art and the route of administration of the drug, etc., so there is no special limitation.

[0049] In the present invention, the administration routes of the anti-HBV drug include but are not limited to oral administration, injection, mucosal, transdermal or topical administration, nasal or inhalation administration, etc., and the corresponding dosage form is prepared according to the different administration routes. Preferably, it is an injection or oral preparation.

[0050] Furthermore, the anti-HBV drug of the present invention may also include other known active pharmaceutical ingredients with anti-HBV effects, so as to exert a better anti-HBV effect. Preferably, these active pharmaceutical ingredients are Chinese medicine ingredients, so as to be prepared into a Chinese medicine compound. It is understandable that these active pharmaceutical ingredients should have an additive or synergistic effect with lulutong acid, and will not produce toxic side effects with lulutong acid. The selection of specific drugs can be carried out by those skilled in the art by methods well known in the art. It should be noted that if it is a Chinese medicine compound, it needs to be carried out according to the Chinese medicine prescription compatibility principle of "monarch, minister, assistant and messenger".

[0051] The present invention is described below by means of specific examples. It should be noted that the following specific examples are only for illustrative purposes and do not limit the scope of the present invention in any way. In addition, unless otherwise specified, methods without specific conditions or steps are conventional methods, and the reagents and materials used can be obtained from commercial channels.

[0052] The cells, antibodies and reagents used in the examples of the present invention are specifically shown in Tables 1 to 3.

[0053] Table 1 Information on cells, viruses and experimental animals

[0054] name source HepG2.215 cell line Enzyme Research Biotechnology AAV8[HBV-D,ayw](D#2012) Paizhen Biotechnology C57BL / 6 mice Jicui Yaokang

[0055] Table 2 Antibody information

[0056] Antibody Name Part Number Manufacturer β-actin Sc-4778 Santa Cruz PGC1alpha+beta Antibody ET1702-96 Hua an PPARγ Sc-7273 Santa Cruz HNF-4α Sc-374229 Santa Cruz HNF-1α Sc-393925 Santa Cruz Ubiquitin rabbit mAb R26024 Positive Energy Goat anti-mouse IgG(H+L)Antibody 31430 Invitrogen Goat anti-rabbit IgG(H+L)Antibody 31460 Invitrogen Rabbit anti-HBsAg NB100-62652 Novus Biologicals Mouse anti-HBcAg C010103 Selvi

[0057] Table 3 Reagent information

[0058]

[0059]

[0060] It should be noted that the above information listed in the present invention is only an example made for the full disclosure of the technical solution, and does not mean that the technical solution of the present invention can only be implemented by using the above reagents, cells or antibodies. The specific scope of protection shall be subject to the claims.

[0061] Example 1 Toxicity test of lulutong acid on HepG2.215 cells

[0062] In this example, the CCK8 cytotoxicity test was used to detect the toxicity of different concentrations of passutonic acid on HepG2.215 cells to preliminarily determine the safe concentration range of passutonic acid on HepG2.215 cells. The specific steps are as follows:

[0063] HepG2.215 cells were inoculated into 96-well plates with 20,000 cells per well in DMEM growth medium containing 10% fetal bovine serum, and cultured in an incubator at 37°C and 5% CO2. When the cells grew to a confluence of about 70%, the 10mM stock solution of passepartout acid prepared by dissolving DMSO in passepartout acid dry powder was diluted with DMEM growth medium to different dosing concentrations of 5μM, 10μM, 15μM, 20μM, 25μM, 30μM, 35μM, and 40μM, respectively. The medium in the 96-well plate was replaced with 100μL of growth medium containing the drug per well. A control group (giving an equal volume of DMSO) and a blank group (without cells) were set up at the same time, with 3 replicates in each group. The cells were cultured in an incubator at 37°C and 5% CO2 for 48h. After 48 hours, discard the supernatant and add 100 μL of DMEM growth medium containing 10% CCK8 cytotoxicity detection solution. Continue to culture in an incubator at 37°C, 5% CO2 for a suitable period of time. Measure the OD value at 450nm using a microplate reader. For the results, see Figure 1 .

[0064] pass Figure 1 It can be seen that lulutong acid has low drug toxicity to HepG2.215 cells within a dosage concentration of 30 μM.

[0065] Example 2 Drug toxicity of thiamethoxam acid to HepG2.215 cells at safe concentrations

[0066] Based on the results in Example 1, flow cytometry was used in this example to further determine the drug toxicity of lulutonic acid to HepG2.215 cells at a safe dosing concentration, and the specific steps were as follows:

[0067] HepG2.215 cells were plated in a six-well plate in DMEM growth medium containing 10% fetal bovine serum, with about 500,000 cells in each well. When the HepG2.215 cells grew to a confluency of about 70% in the 6-well plate, the 10mM stock solution of 10μM, 10μM, 15μM, and 20μM of the drug were diluted with the above culture medium, and the culture medium containing 100μL of 1 ... Figure 2 .

[0068] pass Figure 2 It can be seen that compared with the control group, lulutong acid has extremely low cytotoxicity to HepG2.215 cells at a dosage concentration of less than 20 μM.

[0069] Example 3 Inhibitory effect of different concentrations of lumefantrine on the production of HBsAg and HBeAg in HepG2.215 cells

[0070] Hepatitis B surface antigen (HBsAg) is the earliest and most important marker of hepatitis B virus infection, and hepatitis B e antigen (HBeAg) is a marker of hepatitis B virus replication. In this embodiment, the inhibitory effect of lulutonic acid on the production of HBsAg and HBeAg by HepG2.215 cells at different administration concentrations was detected by enzyme-linked immunosorbent assay. The specific steps are as follows:

[0071] HepG2.215 cells were plated in 12-well plates and cultured in the same manner as in Example 1. When the HepG2.215 cells grew to a confluency of about 70% in the 12-well plate, the culture medium was discarded, and the thiabendazole was diluted to 10 μM and 20 μM with new DMEM growth medium. A control group (DMSO with the same drug volume) was set up at the same time, with 3 replicates in each group. The cells were cultured in an incubator at 37°C and 5% CO2 for 48 hours. The supernatant of the culture medium was then aspirated and tested according to the instructions of the enzyme-linked immunosorbent assay kit. The OD value was tested on an enzyme reader. The results can be found in Figure 3 .

[0072] pass Figure 3 It can be seen that with the increase of the concentration of lulutong acid, the amount of HBsAg and HBeAg produced by HepG2.215 cells decreased ( Figure 3 a and Figure 3 b), there was a significant difference compared with the control group. The results showed that lulutong acid had a significant inhibitory effect on the production of HBsAg and HBeAg in HepG2.215 cells at a concentration of 10μM and 20μM. Compared with the control group, * indicates a significant difference (P<0.05); ** indicates an extremely significant difference (P<0.01); *** indicates a more significant difference (P<0.001); **** indicates a particularly significant difference (P<0.0001).

[0073] Example 4 Inhibitory effect of different concentrations of lulutong acid on mRNA expression in HepG2.215 cells

[0074] The level of HBV mRNA reflects the activeness of HBV infection. A high level of HBV mRNA indicates active viral replication and strong infectivity. In this example, the inhibitory effect of different concentrations of lulutonic acid on mRNA expression in HepG2.215 cells was determined by real-time fluorescence quantitative PCR, and the specific steps are as follows:

[0075] Using the same cell plating and culture method as in Example 3, RNA was extracted from HepG2.215 cells 48 hours after administration, and then the reverse transcription kit ( Plus All-in-one 1st Strand cDNASynthesis SuperMix (gDNAPurge)) instructions for reverse transcription. Dilute cDNA ten-fold as a template and configure the qPCR system: 2μL diluted cDNA per well of a 96-well plate, 7μL ddH2O, 10μL SYBR, and 0.5μL each of upstream and downstream primers. The primer information involved is shown in the following table:

[0076]

[0077] According to the kit ( Real-time fluorescence quantitative PCR was performed using the qPCR program in the SYBR qPCR SuperMix Plus manual. The control group without drug administration was used as a reference to calculate the relative expression of totalRNA and pgRNA and evaluate the changes in mRNA expression. Figure 4 .

[0078] pass Figure 4 It can be seen that compared with the control group, lulutong acid at a concentration of 20 μM had a significant inhibitory effect on the mRNA expression of HepG2.215 cells ( Figure 4 a and Figure 4 b). Among them, * indicates significant difference (P<0.05); ** indicates extremely significant difference (P<0.01).

[0079] Example 5 The inhibitory effect of thiabendazole on the expression of HBV transcription factors.

[0080] HBV transcription factors play a very important role in the replication and expression of hepatitis B virus. High expression levels or active expression of HBV transcription factors will promote the replication and expression of HBV virus, thereby promoting the development of hepatitis B. In this example, Western blot was used to study the inhibitory effect of thiabendazole on the expression of HBV transcription factors (PPARγ, PGC1α, HNF4α and HNF1α), and the specific steps are as follows:

[0081] HepG2.215 cells were cultured and administered in the same manner as in Example 2. After 48 hours, the cells were collected, washed twice with PBS, and the supernatant was discarded. 200 μL of protein lysis buffer NP-40 was added to lyse the cells on ice for 40 to 60 minutes, centrifuged at 12000 rpm for 15 minutes, and the supernatant was transferred to another clean EP tube. After the concentration of the supernatant was determined by the BCA kit, protein samples of the same concentration were prepared. Then, the protein bands of different transcription factors were detected by Western blot. The test results are shown in Figure 5 .

[0082] pass Figure 5 It can be seen that lulutong acid has a significant inhibitory effect on the protein expression of HBV transcription factors at the dosage concentration of 10μM and 20μM, and has a certain dose-effect relationship.

[0083] Example 6 Construction of a hepatitis B mouse model to study the effect of lulutong acid in the hepatitis B mouse model

[0084] In this example, a hepatitis B mouse model was constructed to further study and determine the therapeutic effect of lulutonic acid on hepatitis B virus. The specific steps are as follows:

[0085] 1. Construction of hepatitis B mouse model

[0086] 6-8 weeks old CL57 mice were injected with HBV virus AAV8[HBV-D,ayw](D#2012) (PBS diluted) into the eye sockets once a week for a total of two injections. Then, the mice were bled from the eye sockets to detect HBsAg and HBeAg in the serum of the mice to determine whether the model was successfully constructed.

[0087] 2. Drug administration experiment

[0088] The successfully constructed hepatitis B mouse model was divided into an experimental group and a control group. The mice in the experimental group were intraperitoneally injected with 20μM 100μL of 10μM ...

[0089] 3. Immunohistochemical detection of HBcAg and HBsAg in mouse liver

[0090] After the mice were killed by cervical dislocation, fresh livers of mice in the control group and experimental group were fixed in 4% paraformaldehyde and embedded in paraffin. The fixed tissue samples were embedded in paraffin and 3-4 μm sections were made. The paraffin sections were dewaxed in xylene and hydrated through an alcohol gradient to remove the paraffin and restore the natural state of the tissue. The sections were treated with antigen recovery solution. The sections were treated with a 3% hydrogen peroxide aqueous solution to inactivate endogenous peroxidase, and nonspecific binding sites were blocked with a blocking solution. The sections were incubated with primary antibodies (Rabbit anti-HbsAg and Mouse anti-HbcAg) and incubated at room temperature for 60 minutes before the secondary antibodies (Goat anti-mouse IgG (H+L) Antibody and Goat anti-rabbit IgG (H+L) Antibody) labeled with horseradish peroxidase (HRP) were used to bind to the primary antibodies. DAB was used as a color developer to observe the location of the target antigen under an optical microscope.

[0091] See the results Figure 6 and Figure 7 .

[0092] in, Figure 6 The figure shows the changes in HBsAg expression in mouse serum after a period of treatment with lulutong acid. It can be seen that compared with the control group, the HBsAg expression in the serum of hepatitis B mice in the experimental group has a significant decreasing trend.

[0093] Figure 7 The figure shows the detection of HBcAg and HBsAg in the liver of mice after administration of lulutong acid. It can be seen that compared with the control group, the cell membrane and nucleus color expression in the liver cells of the experimental group of mice was weaker, which indicates that the hepatitis B condition of the mice in the experimental group treated with lulutong acid was under control, indicating that lulutong acid has obvious anti-hepatitis B virus effect.

[0094] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0095] The above-mentioned embodiments only express several implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. Application of lulutonic acid in the preparation of anti-hepatitis B virus drugs.

2. The use according to claim 1, characterized in that The drug has at least one of the following effects: a: Inhibit the replication of hepatitis B virus; b: Inhibit the production and replication of hepatitis B virus antigen; c: Inhibit the expression of hepatitis B virus mRNA; d: Inhibit the expression of HBV transcription factors; e: Treatment of hepatitis B.

3. The use according to claim 2, characterized in that The hepatitis B virus antigen is at least one of HBsAg, HBeAg, and HBcAg.

4. The use according to claim 2, characterized in that The HBV transcription factor is at least one of PPARγ, PGC1α, HNF4α and HNF1α.

5. An anti-hepatitis B virus drug, characterized in that: Contains effective concentration of lulutonic acid.

6. The anti-HBV drug according to claim 5, characterized in that: The effective concentration of the lulutonic acid is 5 μM to 30 μM; Preferably, the effective concentration of the lulutonic acid is 5 μM to 20 μM.

7. The anti-HBV drug according to claim 6, characterized in that: The effective concentration of the lulutonic acid is 10 μM to 20 μM; Preferably, the effective concentration of the lulutonic acid is 10 μM or 20 μM; Preferably, the effective concentration of lulutonic acid is 20 μM.

8. The anti-hepatitis B virus drug according to any one of claims 5 to 7, characterized in that: The drug has at least one of the following effects: a: Inhibit the replication of hepatitis B virus; b: Inhibit the production and replication of hepatitis B virus antigen; c: Inhibit the expression of hepatitis B virus mRNA; d: Inhibit the expression of HBV transcription factors; e: Treatment of hepatitis B.

9. The anti-HBV drug according to claim 8, characterized in that: The hepatitis B virus antigen is at least one of HBsAg, HBeAg, and HBcAg.

10. The anti-HBV drug according to claim 8, characterized in that: The HBV transcription factor is at least one of PPARγ, PGC1α, HNF4α and HNF1α.

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