CN108245501B

CN108245501B - Application of chained diterpene compound of lindley eupatorium in preparing anti-hepatitis B virus medicine

Info

Publication number: CN108245501B
Application number: CN201810045547.1A
Authority: CN
Inventors: 张健; 陈道峰; 任慧玲; 褚纯隽; 李显伦; 严彪; 吴双庆
Original assignee: Suzhou University
Current assignee: Suzhou University
Priority date: 2014-06-04
Filing date: 2014-06-04
Publication date: 2020-07-24
Anticipated expiration: 2034-06-04
Also published as: CN104069145A; CN104069145B; CN108113985A; CN108245501A; CN108096307B; CN108096307A; CN108159099B; CN108113985B; CN108159099A

Abstract

The invention discloses an application of a chained diterpene compound of lindley eupatorium in preparing anti-hepatitis B virus medicines. The invention prepares chain diterpenoid compounds from lindley eupatorium, and carries out anti-hepatitis B virus HBV experiment on the compounds, and the result shows that the compounds have the function of obviously inhibiting hepatitis B surface antigen HBsAg and hepatitis B core antigen HBeAg, and have better drug activity compared with the clinical drug lamivudine (3TC) for treating hepatitis B. The chained diterpene compound of the lindley eupatorium herb can be used for preparing anti-hepatitis B virus medicines.

Description

Application of chained diterpene compound of lindley eupatorium in preparing anti-hepatitis B virus medicine

Technical Field

The invention relates to an application of a chained diterpene compound of lindley eupatorium in preparing anti-hepatitis B virus medicines.

Background

The traditional Chinese medicine is mainly 1 of oral nucleoside (acid) chemical medicines such as lamivudine, adefovir, entecavir, telbivudine, tenofovir and the like which are easy to generate drug resistance, 2 of biological preparation, interferon such as interferon α and pegylated interferon α which are commonly used, but most of patients have a rebound phenomenon after drug withdrawal, the interferon is expensive (about 2-4 ten thousand yuan for each treatment course), the use is inconvenient due to gastrointestinal tract medication, side effects are easy to generate due to long-term medication, such as side effects similar to cold symptoms and thrombocytopenia, 3 of traditional Chinese medicine, kushenin, oleanolic acid and derivatives thereof, plant polysaccharides, chemical components and the like, clinical medical practices and experimental studies prove that the traditional Chinese medicine can inhibit the replication of viruses to a certain extent on hepatitis diseases, the replication of viruses of Chinese medicine is rich, the replication of kushenin Chinese medicine, the liver disease is rich in chemical components, the clinical medical practices and the research of Chinese medicine is beneficial to develop and improve the functions of hepatitis virus (11) of hepatitis B virus, especially the liver cirrhosis and liver cancer, the liver cancer is easily treated by researching the traditional Chinese medicine, the medicine has the research on the liver disease, the research on the hepatitis diseases, the research on the replication of traditional Chinese medicine, the research on the hepatitis virus replication of human body, the research on the replication of human health, the research on the research of human body, the research on the hepatitis, the research of human body, the research on the research of hepatitis virus, the research on the research of hepatitis virus replication of human body, the research on the research.

Herba Eupatorii Lindleyanum DC of Compositae is herba Eupatorii Lindleyanum, and is prepared from herba Eupatorii Lindleyanum DC. Has bitter taste and mild nature, enters liver and spleen meridians, has the functions of eliminating phlegm and stopping cough, clearing heat and releasing toxin, inducing diuresis and reducing edema and reducing blood pressure, and is used for treating cold, cough and excessive phlegm, headache, tonsillitis, bacillary dysentery and hypertension (see: the national traditional Chinese medicine administration, Chinese materia medica, Shanghai: Shanghai science and technology publisher, 1999:7,6876). The eupatorium extract has a certain effect of protecting rats with acute lung injury, the compound eupatorium capsule has a certain effect of inhibiting influenza viruses in vitro, the eupatorium syrup concentrated solution has a certain antibacterial effect on gram positive and negative bacteria and candida albicans, and the eupatorium total flavone extract has the effect of reducing blood viscosity (see: Jianzhou, Yanghui, Hoohiphyly, etc. the eupatorium has the effect of protecting rats from acute lung injury, Chinese pharmacy 2007, 18 (27): 4; Zhou Yu Dai, Wu Yan, etc. the eupatorium has the effects of resisting bacteria, relieving cough and relieving asthma, Chinese pharmacy 2001, 12 (12): 716 Bu Ru 718, Penn Ru, sinus, yellow fragrant, etc. the compound eupatorium capsule has the effect of resisting influenza viruses, Chinese patent medicine, 2008, 30 (5): 650 Bu Jing 654, Wang Jing, Qin, Cheng and Zhen Zhu Zhen, etc. in case the rat improvement of blood hyperlipidemia and the antioxidation effect, 2009, 25(2): 80-82. ) However, whether the lindley eupatorium has the effect of resisting the hepatitis B virus is not reported, and the invention finds that the lindley eupatorium has the activity of resisting the hepatitis B virus.

Disclosure of Invention

The invention aims to: provides an application of a chained diterpenoid compound of lindley eupatorium in preparing anti-hepatitis B virus medicaments.

The technical scheme of the invention is as follows: application of herba Eupatorii Lindleyani in preparing anti-HBV medicine is provided.

The lindley eupatorium herb is an ethanol extract of the lindley eupatorium herb, and the ethanol extract is obtained by adopting the following method: taking dry eupatorium lindleyanum whole grass, cutting the eupatorium lindleyanum into sections of 2-3 cm, adding 8-15 times of ethanol aqueous solution with volume fraction of 60-95%, refluxing and extracting for 1-3 hours, repeating for 1-3 times, and recovering the solvent under reduced pressure at 40-60 ℃ to obtain the eupatorium lindleyanum ethanol extract.

The lindley eupatorium herb is a lindley eupatorium herb flavone part containing jaceosidin and eupatorium xanthophyll, and the chemical structural formulas of the lindley eupatorium herb are respectively as follows:

the lindley eupatorium is a lindley eupatorium sesquiterpene part containing lindley eupatorium lactone F, lindley eupatorium lactone G, lindley eupatorium lactone H, lindley eupatorium lactone I and lindley eupatorium lactone K; the chemical structural formula is as follows:

the lindley eupatorium herb is an eluted part of lindley eupatorium herb with 70% ethanol water, and is prepared by the following method: taking dry eupatorium lindleyanum whole grass, cutting the eupatorium lindleyanum into sections of 2-3 cm, adding 8-15 times of ethanol aqueous solution with volume fraction of 60-95%, refluxing and extracting for 1-3 hours, repeating for 1-3 times, and recovering the solvent under reduced pressure at 40-60 ℃ to obtain an eupatorium lindleyanum ethanol extract; dissolving the extract in water to prepare a solution with the concentration of 0.5-1 g/ml, adsorbing the solution by weak-polarity macroporous adsorption resin, eluting by water to remove macromolecular compounds, eluting by 90-95% ethanol aqueous solution by volume fraction, and recovering the solvent at 40-60 ℃ under reduced pressure to obtain a macroporous resin part; adsorbing the obtained macroporous resin part by an ODS (oxide dispersion strengthened) reverse phase silica gel chromatographic column, eluting by 50% methanol aqueous solution by volume fraction, then eluting by 70% -75% methanol aqueous solution, recovering the solvent at 40-60 ℃ under reduced pressure, adsorbing by a 30-60 mesh polyamide chromatographic column, and eluting by 70% ethanol aqueous solution by volume fraction to obtain the lindley eupatorium 70% ethanol aqueous part.

Further, preferably, the weak polar macroporous absorption resin is AB-8, D101, HPD-100, HPD-200A, HPD-200B, HPD-300, HPD-700 or HPD-722 macroporous absorption resin;

further, preferably, the ODS reversed-phase silicA gel chromatographic column is DAISOGE L ODS reversed-phase silicA gel filler with the particle size of 50-70 um, Cosmsil ODS reversed-phase silicA gel filler with the particle size of 50-70 um or YMCODS-A reversed-phase silicA gel filler with the particle size of 50-70 um;

furthermore, preferably, the volume ratio of the water to the low-polarity macroporous adsorption resin is 8-15: 1; the volume ratio of the 90-95% ethanol aqueous solution to the macroporous adsorption resin is 8-15: 1; the mass ratio of the volume of the ODS reversed phase silica gel chromatographic column to the obtained macroporous resin part is 20-30: 1; the volume ratio of the 50% methanol aqueous solution to the 70-75% methanol aqueous solution to the ODS reversed phase silica gel chromatographic column is 5-8: 1; the mass ratio of the volume of the polyamide chromatographic column to the adsorbed sample is 20-30: 1; the volume ratio of the 70% ethanol aqueous solution to the polyamide chromatographic column is 4-10: 1.

The lindley eupatorium is a lindley eupatorium medium-chain diterpenoid compound selected from (1)3- (hydroxymethy) -1, 14, 15-trihydroxy-7, 11, 15-trimethy-2, 6, 10-hexadecainen-13-acetate and (2)3- (hydroxymethy) -1, 13, 15-trihydroxy-7, 11, 15-trimethy-2, 6, 10-hexadecainen-14-acetate, and the structural formulas of the compounds are respectively as follows:

the four extracts and two chain diterpenoid compounds are subjected to in vitro anti-hepatitis B virus HBV experiments to obtain the extract which has obvious inhibition effect on hepatitis B surface antigen HBsAg and hepatitis B core antigen HBeAg, and the extract has obvious anti-hepatitis B virus HBV effect and can be used for preparing anti-hepatitis B virus medicines.

The invention has the advantage of providing a new source for anti-hepatitis B virus medicines.

Drawings

The invention is further described with reference to the following figures and examples:

FIG. 1 is a high performance liquid chromatogram of a tremula sesquiterpene locus of example 2;

FIG. 2 is a high performance liquid chromatogram of control Eupalinolide F in example 2;

FIG. 3 is a high performance liquid chromatogram of control Eupalinolide G of example 2;

FIG. 4 is the HPLC chromatogram of control substance Eupalinolide H in example 2;

FIG. 5 is a high performance liquid chromatogram of control Eupalinolide I in example 2;

FIG. 6 is a high performance liquid chromatogram of control Eupalinolide K in example 2;

FIG. 7 is a high performance liquid chromatogram of the lindley eupatorium herb flavone site in example 2;

FIG. 8 is the HPLC chromatogram of the control Eupatorium adenophorum xanthophyll in example 2;

FIG. 9 is a high performance liquid chromatogram of a control jaceosidin from example 2;

Detailed Description

The invention is further illustrated by the following examples.

Example 1: preparation method of herba Eupatorii Lindleyani extract and herba Eupatorii Lindleyani medium chain diterpene compound

(1) Weighing 1000g of eupatorium lindleyanum dry whole herb, cutting into sections of 2-3 cm, adding 80% ethanol aqueous solution with 15 times volume, carrying out reflux extraction for 2 hours, repeating the steps for 2 times, recovering the solvent under reduced pressure at 60 ℃ to obtain 158g of eupatorium lindleyanum ethanol extract (EUP-EtOH), dissolving the eupatorium lindleyanum ethanol extract (EUP-EtOH) in about 150ml of water, adsorbing the eupatorium lindleyanum ethanol extract with 1800ml of AB-8 macroporous adsorption resin, eluting with 20000ml of water to remove macromolecular compounds, eluting with 20000ml of 95% ethanol aqueous solution, recovering the solvent under reduced pressure at 60 ℃ to obtain 61g of macroporous resin part, adsorbing the macroporous resin part with 1500ml of Cosmsil ODS reversed phase silica gel filler with 70um reversed phase silica gel chromatographic column with particle size of 70um, eluting with 50% methanol aqueous solution with 10000ml of volume, recovering the solvent under reduced pressure at 60 ℃ to obtain 21g of EUP-SQT, eluting with 10000ml of 70% methanol aqueous solution with 60 ℃ and recovering the solvent under reduced pressure at 60 ℃ to obtain 20g of extract, adsorbing the extract with 600ml of 30-60 mesh polyamide chromatographic column, eluting with 70% ethanol aqueous solution with 70 g of 60 ℃ and eluting with 357.7 g of ethanol aqueous solution to obtain 20g of eupatorium lindleyanum ethanol, and recovering the EUP ethanol aqueous solution with 4000ml of 60 ℃ to obtain.

(2) Preparation of Eupatorium lindleyanum medium-chain diterpene compound comprises drying Eupatorium lindleyanum medicinal material 2000g, reflux-extracting with 10 times of 80% ethanol-water for 2 hr each time, mixing filtrates, recovering ethanol under reduced pressure to obtain extract 250g, dissolving the extract in 100m L water, defatting with 100m L petroleum ether, extracting with chloroform twice, 100m L each time, concentrating the chloroform extraction layer under reduced pressure at 30 deg.C to obtain chloroform fraction about 11g, subjecting the chloroform fraction to 300g silicA gel column chromatography, eluting with chloroform-methanol 100:2 to obtain eluate fraction 1.08g, subjecting the chloroform fraction to 200m L Rohm resin column chromatography, eluting with 60% -80% methanol-water system, collecting 80% methanol eluate fraction 0.5g, subjecting the chloroform fraction to silicA gel column chromatography, eluting with chloroform-methanol (50:1) to obtain 2A fraction 112mg, subjecting 2A to high performance liquid chromatography, preparing easy I L C-P chromatography, subjecting to ODS-methanol chromatography (50:1) chromatography, subjecting to obtain YM-ethanol (20 mm, 20 mm/32 mm) chromatography, detecting YM C-32 mm, and separating YM/26 mm.

Example 2: detection of sesquiterpene fraction, flavone fraction and active ingredient of mid-chain diterpene compound

(1) Active ingredient detection of Eupatorium Adenophorum sesquiterpene fraction (EUP-SQT) in this example, its components were determined by HPLC (high Performance liquid chromatography), wherein mobile phase A was acetonitrile, mobile phase B was water, mobile phase conditions are shown in Table 1, column chromatography: YMC-PACK ODS-A (phi 4.6mm × 250mm,5 μm), column temperature: 28 deg.C, detection wavelength: 210nm, sample introduction amount: 20 μ l, flow rate: 1ml min^-1。

TABLE 1 HP L C chromatographic conditions

Time (min)	Mobile phase A (%)	Mobile phase B (%)
			0	30	70
5	30	70
			15	32	68
25	36	64
			40	36	64

6.02mg of reference substances, namely eupalinolide F, 14.94mg of eupalinolide G, 5.08mg of eupalinolide H, 10.04mg of eupalinolide I and 13.98mg of eupalinolide K are precisely weighed respectively, and the volume is fixed to 10ml by using a mobile phase A to serve as a stock solution. Adding mobile phase A, diluting to 1:2, 1:4, 1:8, 1:16 and 1:32, filtering with 0.22 μm filter membrane, and filtering to obtain filtrate. 5 concentrations of control were chromatographed and a standard curve was plotted, with 3 replicates per concentration point. Recording peak areas of the chromatographic peaks, and performing linear regression on the concentration (x) of the chromatographic peaks by using the peak area (y) of a reference substance to obtain a standard curve of the reference substance, wherein eupalinolide F, y is 63403 x-15.408; eupalinolide G, y 24720 x-20.065; eupalinolide h, y 79063x + 6.7811; eupalinolide I, y 239018x + 53.303; eupalinolide K, y 7913.5x + 62.857.

The EUP-SQT 6mg of eupatorium sesquiterpene prepared in example 1 was precisely weighed, the volume was determined to 10ml by mobile phase a volumetric flask, and the filtrate was filtered through a 0.22 μm membrane filter and subjected to chromatography.

FIGS. 1-6 are HPLC chromatograms of sesquiterpene positions (EUP-SQT) from Eupatorium, control products eupalinolide F, eupalinolide G, eupalinolide H, eupalinolide I and eupalinolide K.

By analyzing the high performance liquid chromatography result, eupatorium pseudochinensis (EUP-SQT) part samples containing eupatorium pseudochinensis F (eupatorium pseudochinensis F), eupatorium pseudochinensis G (eupatorium pseudochinensis G), eupatorium pseudochinensis H (eupatorium pseudochinensis H), eupatorium pseudochinensis I (eupatorium pseudochinensis I) and eupatorium pseudochinensis K (eupatorium pseudochinensis K) can be obtained, the chromatographic signal values of the samples are substituted into a standard curve, and the specific content of each component is further obtained, and the specific content is shown in Table 2.

TABLE 2 Eupatorium lappacans sesquiterpene fraction (EUP-SQT) sample containing 5 Eupatorium lappacans lactone

Sample (I)

eupalinolide F

eupalinolide G

eupalinolide H

eupalinolide I

eupalinolide K

Ratio of

Content (wt.)

4.1％

31.8％

6.8％

3.4％

47.7％

93.8％

(2) The method for detecting active ingredients of the eupatorium lindleyanum flavone part (EUP-F L A) comprises the steps of detecting the active ingredients of the eupatorium lindleyanum sesquiterpene part, wherein the conditions of the high performance liquid chromatography comprise that A mobile phase A is acetonitrile (containing 0.1% of formic acid), A mobile phase B is water (containing 0.1% of formic acid), the conditions of the mobile phase are shown in Table 3, A chromatographic column is YMC-PACK ODS-A (phi 4.6mm × 250mm,5 mu m), the column temperature is 28 ℃, the detection wavelength is 365nm, the sample injection amount is 20 mu l, and the flow rate is 1 ml/min^-1。

TABLE 3 HP L C chromatographic conditions

Time (min)	Mobile phase A (%)	Mobile phase B (%)
			0	36	64
15	36	64
			30	90	10

Precisely weighing control substances of eupatorium lucolium xanthophyll 4.67mg and jaceosidin 8.17mg respectively; and adding mobile phase A to a constant volume of 10ml to obtain stock solution, diluting with mobile phase A to obtain 1:2, 1:4, 1:8, 1:16 and 1:32, and filtering with 0.22 μm filter membrane to obtain filtrate. 5 concentrations of control were chromatographed and a standard curve was plotted, with 3 replicates per concentration point. Recording the peak area of each chromatographic peak, and performing linear regression on the concentration (x) by using the peak area (y) of a reference substance to obtain a standard curve of the reference substance, wherein eupatilin y is 52964x +19.30, and jaceosidin y is 55161x + 210.79.

6mg of the EUP-F L a sample of the EUP-F L b prepared in example 1 was weighed out precisely, and the volume was adjusted to 10ml by using mobile phase a, and the solution was filtered through a 0.22 μm filter membrane, and the filtrate was used for the measurement by chromatography.

FIGS. 7-9 are high performance liquid chromatograms of a sample of Eupatorium odoratum flavone, a control of eupatilin, and a control of jaceosidin.

Analyzing by high performance liquid chromatography to obtain eupatorium adenophorum xanthophyll and jaceosidin in the sample of eupatorium adenophorum flavone part, and substituting the chromatographic signal value of the sample into a standard curve to further obtain the specific content of each component, as shown in Table 4.

TABLE 4 content of 2 flavones in Eupatorium lindleyanum flavone part

Sample (I)	Eupatorium xanthophyll	Jaceosidin	Total ratio of
				Content (wt.)	26.0％	66.1％	92.1％

(3) Detection of active ingredients of eupatorium lindleyanum medium-chain diterpenoid compounds: the compound (a) and the compound (b) prepared in example 1 were measured for spectrum, and the spectrum data were as follows:

the compound (a) is colorless oil, and the infrared spectrum IR (KBr) shows v_max：3424，2974， 2930，1717，1653，1373，1258，1024cm^-1(ii) a HR-ESI-MS gives the molecular ion peak M/z 421.2568[ M + Na ]]⁺(calcd forC₂₂H₃₈O₆Na, 421.2561), indicating that its molecular formula is C₂₂H₃₈O₆. It is composed of¹H NMR and¹³c NMR spectra in CDCl₃The measurement in (1). The results are shown in the table1 is shown in the specification;

the compound (b) is colorless oil, and the infrared spectrum IR (KBr) shows v_max：3402，2972， 2932，1653，1576，1418，1051，1015cm^-1(ii) a HR-ESI-MS gives the molecular ion peak M/z 421.2565[ M + Na ]]⁺(calcd.forC₂₂H₃₈O₆Na, 421.2561), indicating that its molecular formula is C₂₂H₃₈O₆. It is composed of¹H NMR and¹³c NMR spectra in CDCl₃The measurement in (1). The results are shown in Table 5 below,

TABLE 5 preparation of the compounds (a), (b)¹H NMR and¹³c NMR spectrum data

Wherein the content of the first and second substances,¹h NMR and¹³c NMR was performed at 400MHz and 125MHz, respectively.

By infrared spectrum, HR-ESI-MS and,¹H NMR and¹³data analysis of C NMR spectra, in comparison with literature (see: Shuangqing Wu, Naiyu Xu, Jian Zhang, Shi Yao, Chunjun Chu. Three new acrylic reagents from Eupatorium linleyanum DC.. Journal of Asian Natural product research.2012,14(7): 652-: compound (a) is compound (1): 3- (hydroxymethy) -1, 14, 15-trihydroxy-7, 11, 15-trimethyl-2, 6, 10-hexadecacarrien-13-acetat, and the structural formula of the compound is as follows:

compound (b) is compound (2): 3- (hydroxymethy) -1, 13, 15-trihydroxy-7, 11, 15-trimethyl-2, 6, 10-hexadecacarrien-14-acetate, the structural formula of the compound is as follows:

example 3: external anti-HBV experiment of four eupatorium lindleyanum extracts and eupatorium lindleyanum medium-chain diterpene compounds

1. Experimental drugs and reagents: the chain diterpene compounds (1) and (2) were prepared in the above example 1; dimethyl sulfoxide; MTT (thiazole blue), shanghai jianlai biotechnology limited; control drug, lamivudine tablet (3 TC); kurarin Schk pharmaceuticals (Suzhou) Inc.; a cell culture medium DMEM, a growth medium and a preparation, wherein the growth medium contains 10% fetal calf serum, 380 mug/ml G418, 0.03% glutamine, penicillin, streptomycin and the like, and each 100 mug/ml;

2. the experimental equipment comprises an E L ISA kit for detecting hepatitis B surface antigen HBsAg and hepatitis B core antigen HBeAg, an incubator, a 24-hole plate and the like.

3. Cell lines: 2.2.15 cell line of Hep G2, department of education/Ministry of health, molecular virology focus laboratory, Shanghai.

4. The experimental method comprises the following steps:

2.2.15 cell lines of Hep G2 were collected at 10 × 10 per well⁵Individual cells were seeded in 24-well plates at 5% CO₂Culturing at 37 ℃ in an incubator, after 48 hours, respectively replacing the growth solution with drug culture solutions containing the compound (1) and the compound (2) which are dissolved by dimethyl sulfoxide, setting 5 concentrations of 12.5 mu g/ml to 200 mu g/ml (shown in table 2) in each drug culture solution, setting 4 parallel holes in each concentration, continuing to culture for 9 days, replacing the solution once every 3 days, wherein the drug of a positive control group is lamivudine (3TC), meanwhile, taking the culture solution without the drug as a blank control group, setting 1 hole in each of the two control groups, except the corresponding lamivudine drug of the growth solution and the culture solution without the drug, performing the same operation on the two control groups and the drug culture solution, collecting supernatant, detecting the content of HBsAg and HBeAg by using an E L ISA kit, and simultaneously determining the toxicity of the two groups of compounds on cells by using MTT, and the results are shown in table 6.

The above experimental procedures were repeated with ethanol extract of Eupatorium Adianum, Eupatorium Adianum flavone fraction, Eupatorium Adianum sesquiterpene fraction, and Eupatorium Adenanum 70% ethanol eluted fraction, respectively, to obtain the results shown in Table 6.

As can be seen from Table 6, the chain diterpenoid compounds (1) and (2) have inhibition effects on hepatitis B surface antigen HBsAg and hepatitis B core antigen HBeAg, the inhibition rates of the compounds (1) and (2) on HBsAg and HBeAg are increased along with the increase of the concentration, the inhibition rates of the compound (1) on HBsAg and HBeAg are obviously higher than 3TC at 50 mu g/m L, the inhibition rates of the compound (2) on HBsAg and HBeAg are obviously higher than 3TC at the same concentration of the compound (2) and the 3TC, and the inhibition rates of the compounds (1) and (2) on HBsAg and HBeAg are high compared with 3TC at low concentration.

The ethanol extract, flavone part, sesquiterpene part, and 70% ethanol-eluted part of Eupatorium Adianum have effects of inhibiting HBsAg and HBeAg, and increasing the inhibition rate with increasing concentration; compared with 3TC, the low-concentration Eupatorium Adenophorum hand Mazz extract, Eupatorium Adenophorum hand Mazz fraction, and Eupatorium Adenophorum hand Mazz fraction with 70% ethanol elution fraction all have high inhibitory effect on hepatitis B surface antigen HBsAg and hepatitis B core antigen HBeAg.

TABLE 6 anti-HBV test results of four groups of Eupatorium Adenophorum extracts and two groups of chain diterpene compounds

Wherein, the ratio of +: "show toxicity" means a cell viability of < 75%; /: no inhibition rate test was performed.

The above examples can obtain that the drug prepared by extracting the lindley eupatorium ethanol extract, the lindley eupatorium flavone part, the lindley eupatorium sesquiterpene part, the lindley eupatorium 70% ethanol water elution part and the two chain diterpenoid compounds from the lindley eupatorium shows obvious inhibition effect on hepatitis B surface antigen HBsAg and hepatitis B core antigen HBeAg in an in vitro anti-hepatitis B virus HBV experiment, and has better drug activity compared with the clinical drug lamivudine (3TC) for treating hepatitis B. Therefore, the ethanol extract, the flavonoid part, the sesquiterpene part, the 70% ethanol water elution part and two chain diterpene compounds extracted from the lindley eupatorium herb can be used for preparing the anti-hepatitis B virus medicines.

Claims

1. The application of the chained diterpenoid compound of the lindley eupatorium in preparing the anti-hepatitis B virus medicines is characterized in that the chained diterpenoid compound of the lindley eupatorium is selected from (1)3- (hydroxymethy) -1, 14, 15-trihydroxy-7, 11, 15-trimethyl-2, 6, 10-heptadecatrien-13-acetate and (2)3- (hydroxymethy) -1, 13, 15-trihydroxy-7, 11, 15-trimethyl-2, 6, 10-heptadecatrien-14-acetate, and the chemical structural formulas of the chained diterpenoid compound of the lindley eupatorium are respectively as follows: