CN105439878B - 1,3 indandione class compound screwtree root alkali and its medicinal usage - Google Patents
1,3 indandione class compound screwtree root alkali and its medicinal usage Download PDFInfo
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- CN105439878B CN105439878B CN201410505723.7A CN201410505723A CN105439878B CN 105439878 B CN105439878 B CN 105439878B CN 201410505723 A CN201410505723 A CN 201410505723A CN 105439878 B CN105439878 B CN 105439878B
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Abstract
The invention belongs to field of medicaments, is related to a kind of 1,3 indandione class noval chemical compounds and its purposes in anti-hepatitis virus and anti-inflammatory drug is prepared.The present invention from screwtree root platymiscium screwtree root (Helicteres.angustifolia) dry root isolated 1 by two kinds of configurations of E and Z with 1:1,3 indandione Alkaloid screwtree root alkali (helicterine) of 1 ratio composition, and confirm that it has stronger anti-hepatitis B activity using modern pharmacological research method, to HBsAg IC50It is worth for 58.3 μ g/ml, is significantly higher than positive control lamivudine, there is a certain degree of inhibitory action to HBeAg；In addition, suppressing nitric oxide generation result of the test in vitro shows its induced by endotoxin (LPS) and polyinosinic acid [poly (I:C)] the macrophage nitric oxide generation of induction has very strong inhibitory action, IC50Value is respectively 15.96 and 8.96 μM.Described screwtree root alkali can be used for preparing anti-hepatitis virus and anti-inflammatory drug.
Description
Technical field
The invention belongs to field of medicaments, is related to 1,3- indandione classes isolated in a kind of screwtree root from medicinal plant
Noval chemical compound screwtree root alkali and its purposes in anti-hepatic-B virus medicine and anti-inflammatory drug is prepared.
Background technology
Dry root or complete stool of the screwtree root for Sterculiaceae plant screwtree root (Helicteres angustifolia L.), taste
It is bitter, cold in nature, return lung, large intestine channel, can inducing diaphoresis heat-clearing, removing toxicity for detumescence, for treat cold, fever, mumps, newborn pretty young woman, measles, cough,
Dysentery, carbuncle swells.Screwtree root has and largely on screwtree root anti-hepatitis virus and controlled as the existing long history of medication among the people
The document report of inflammation is treated, and substantial amounts of clinical test proves that its is curative for effect, therefore domestic many scholars are to its anti-second
Hepatovirus and antiphlogistic effects expand numerous studies, but stay on crude extract more, up to the present, its anti-hepatitis virus and anti-
Scorching effective substance and mechanism of action are not yet clear and definite.And screwtree root is very abundant in china natural resources, present inventor intends
It is furtherd investigate, determines its anti-hepatitis virus and antiinflammatory action material, illustrates its mechanism of action, and open on this basis
Anti-hepatitis virus and anti-inflammatory drug are sent, can not only make full use of screwtree root resource, infection hepatitis B can also be reduced and suffered from
Person and the pain of inflammatory disease patients, create larger economic benefit and social benefit.
The content of the invention
It is an object of the invention to provide a kind of isolated 1,3- indandione class noval chemical compounds mountain sesame from screwtree root
Numb alkali (helicterine, 1).
The further object of the present invention, which also resides in, provides above-claimed cpd as the purposes prepared in anti-hepatic-B virus medicine.
The further object of the present invention, which also resides in, provides above-claimed cpd as the purposes prepared in anti-inflammatory drug.
Ethanol extract ethyl acetate of the present invention from screwtree root platymiscium screwtree root (H.angustifolia) dry root
Isolated 1 1,3- indandione Alkaloid in extraction position is (by the two kinds of configurations of E and Z that can not separate with 1:1 ratio group
Into), and modern pharmacological research method is applied, anti-hepatitis virus and anti-inflammatory activity evaluation have been carried out to isolated compound,
As a result confirm that it has stronger effect on hepatitics B virus in vitro activity and induced by endotoxin (LPS) and polyinosinic acid [poly (I:
C)] the macrophage system RAW264.7 generation nitric oxides (NO) of induction have stronger inhibitory action, and described compound can
For preparing anti-hepatic-B virus medicine and preparing anti-inflammatory drug.
1,3- indandiones class compound of the present invention has following chemical constitution：
Compound of the present invention is prepared by following methods：
Screwtree root dry root 25kg is taken, is crushed, 3 times (50L × 3) is extracted with 95% alcohol reflux, each 2h, merges extraction
Liquid is simultaneously concentrated to give medicinal extract 0.95kg, adds water (4L) to be suspended, respectively with isometric petroleum ether, ethyl acetate and extracting n-butyl alcohol 5 times,
Combining extraction liquid is simultaneously concentrated to dryness, and obtains acetic acid ethyl ester extract 200g.By Ethyl acetate fraction through silica gel (200-300 mesh)
Pillar layer separation, successively with methylene chloride-methanol (50:1-0:1) gradient elution, 11 fractions (Fr.1-11) are obtained, wherein flowing
Part Fr.3 (32g) again through silica gel column chromatography (methylene chloride-methanol, 30:1,20:1,10:1,5:1,3:1,1:1), MPLC (first
Alcohol-water, 20:80-80:20 gradient elutions) and purify the methods of sephadex LH-20 column chromatographys, isolated compound mountain sesame
Numb alkali (helicterine, 1, by the two kinds of configurations of E and Z that can not separate with 1:1 ratio forms).
Brief description of the drawings
Fig. 1 is the extraction separation process figure of screwtree root alkali in screwtree root root.
Embodiment
Embodiment 1 prepares screwtree root alkali
Screwtree root dry root 25kg is taken, is crushed, 3 times (50L × 3) is extracted with 95% alcohol reflux, each 2h, merges extraction
Liquid is simultaneously concentrated to give medicinal extract 0.95kg, adds water (4L) to be suspended, respectively with isometric petroleum ether, ethyl acetate and extracting n-butyl alcohol 5 times,
Combining extraction liquid is simultaneously concentrated to dryness, and obtains acetic acid ethyl ester extract 200g.By Ethyl acetate fraction through silica gel (200-300 mesh)
Pillar layer separation, successively with methylene chloride-methanol (50:1-0:1) gradient elution, 11 fractions (Fr.1-11) are obtained, wherein flowing
Part Fr.3 (32g) again through silica gel column chromatography (methylene chloride-methanol, 30:1,20:1,10:1,5:1,3:1,1:1), MPLC (first
Alcohol-water, 20:80-80:20 gradient elutions) and the purifying of the means such as sephadex LH-20 column chromatographys, it is isolated by two kinds of E and Z
Configuration is with 1:Screwtree root alkali (helicterine, the 1) composition of 1 ratio composition；
Compound 1 is yellow lump shaped crystalline (chloroform-methanol)；m.p.205-208℃；
MeOH)；Molecular formula：C14H16NO3；ESIMS：245；UV(MeOH):λmax(logε):239.5(3.41),301.4(3.27)nm；
IR(KBr)υmax(cm-1)：3318,2955,2920,2356,1694,1635,1558,1419,1308；HRESIMS
(positive)m/z246.1129[M+H]+(C14H16NO3,calcd.246.1125)；The NMR data of wherein Z configurations (1a) is
：1H-NMR(600MHz,DMSO-d6)δH:8.89 (2H, dd, J=15.7,10.6Hz, 12-NH2),7.62(1H,m,H-12),
7.43 (1H, d, J=7.9Hz, H-6), 7.32 (1H, d, J=7.9Hz, H-5), 4.27 (1H, m, H-10), 3.59 (1H, m, H-
11a),3.47(1H,overlapped,H-11b),2.56(3H,s,4-OCH3), 1.16 (3H, d, J=7.0Hz, 10-OCH3)
；13C-NMR(150MHz,DMSO-d6)δC:194.1(C-1),191.6(C-3),151.1(C-12),141.9(C-7),136.1
(C-9),136.0(C-5),135.6(C-8),133.4(C-4),132.0(C-6),104.9(C-2),66.3(C-11),34.8
(C-10),18.0(10-OCH3),17.8(4-OCH3)；The NMR data of contained E (1b) is：1H-NMR(600MHz,DMSO-
d6)δH:8.89 (2H, dd, J=15.7,10.6Hz, 12-NH2), 7.62 (1H, m, H-12), 7.43 (1H, d, J=7.9Hz, H-
6), 7.32 (1H, d, J=7.9Hz, H-5), 4.23 (1H, m, H-10), 3.59 (1H, m, H-11a), 3.47 (1H,
overlapped,H-11b),2.58(3H,s,4-OCH3), 1.17 (3H, d, J=7.0Hz, 10-OCH3)；13C-NMR
(150MHz,DMSO-d6)δC:193.9(C-3),191.4(C-1),151.0(C-12),141.6(C-7),136.5(C-8),
136.0(C-5),136.0(C-9),133.7(C-4),131.9(C-6),104.9(C-2),66.3(C-11),34.9(C-10),
18.0(10-OCH3),17.7(4-OCH3)。
The effect on hepatitics B virus in vitro of embodiment 2 (HBV) is tested
Using Hep G2 2.2.15 cell lines (Ministry of Education/medical molecular virology key lab of the Ministry of Public Health, Shanghai),
With every hole 10 × 105Individual cell is inoculated in 24 orifice plates, culture medium DMEM, and growth-promoting media contains 10% hyclone, 380 μ g/ml
G418,0.03% glutamine, each 100 μ g/ml of penicillin, streptomysin, in 5%CO237 DEG C of cultures in incubator, after 48 hours,
Changing the drug containing nutrient solution with dimethyl sulfoxide (DMSO) hydrotropy into, every kind of medicine sets 3~5 concentration, and each concentration sets 4 parallel holes, after
Continuous culture 9 days (changing liquid once in every 3 days), collect supernatant and detect HBsAg and HBeAg contents with ELISA.With not under similarity condition
The nutrient solution supernatant of drug containing is as a control group.Simultaneously use above-mentioned cell line, with mtt assay measure medicine cytotoxicity.Sun
Property control be lamivudine.As a result confirm that compound screwtree root alkali of the present invention has significant Anti-HBV effect, and it is effectively dense
Spend low, cytotoxicity is smaller, to HBsAg IC50It is worth for 58.3 μ g/ml, is significantly higher than positive control lamivudine (3TC), it is right
HBeAg has a certain degree of inhibitory action, and when concentration is 100 μ g/ml, inhibiting rate is 27.7% (table 1).
The inhibitory action that the screwtree root alkali of table 1. is secreted to HBsAg and HBeAg
/：Tested when cytotoxicity >=25% without inhibiting rate
Embodiment 3 suppresses macrophage (RAW264.7) NO generation experiments in vitro
Murine macrophage cell line (RAW264.7) cell is taken, with 1.0 × 106/ ml concentration is seeded in 96 orifice plates,
2h is cultivated at 37 DEG C.Then by test sample (1,2.5,5,10 and 20 μM) and LPS (1ng/ml) or poly (I:C) (1 μ g/ml) one
With being added in cell culture fluid, 24h is cultivated at 37 DEG C jointly, cell culture fluid is cooled down rapidly in ice.Cell is taken to train
The supernatant of nutrient solution is placed in 96 new orifice plates by every μ l of hole 50, and 50 μ l Griess reagents (25 μ l1% couple are added in every hole
The phosphoric acid solution of aminobenzene sulfonamide 2.5% and the phosphoric acid solution of 25 μ l0.1% naphthodiamides 2.5%).After placing 10min at room temperature,
Absorbance (A) value of reaction product is determined in 570nm with ELIASA.In the orifice plate of same 96, NaNO is added simultaneously2Mark
Quasi- solution is to draw calculation NO2 -The standard curve of concentration, not add LPS or poly (I:C) and sample cell culture fluid make
For blank, not to be loaded the cell culture fluids of product as negative control, using dexamethasone as positive control.Cell is determined with mtt assay
Toxic action.As a result show, compound of the present invention is to LPS and poly (I:C) the macrophage NO generations of induction have stronger
Inhibitory action, its IC50Value is respectively 15.96 and 8.96 μM, with dexamethasone (DEX) for positive control, the results showed that mountain sesame
Numb alkali has anti-inflammatory activity (as shown in table 2).
The inhibitory action that the screwtree root alkali of table 2. generates to macrophage NO
The reagent that use is tested in the present invention is techniques well known, commercially available.
Claims (2)
1. the 1,3- indandione class compound screwtree root alkali (helicterine) with following chemical constitution is preparing anti-hepatitis B
Purposes in virus drugs,
2. purposes as claimed in claim 1, it is characterised in that described 1,3- indandione class compound screwtree root alkali
(helicterine) prepared by following methods：
Screwtree root dry root is taken, is crushed, is extracted 3 times, each 2h with 95% alcohol reflux, merges extract solution and is concentrated to give medicinal extract,
Add water to be suspended, be simultaneously concentrated to dryness with isometric petroleum ether, ethyl acetate and extracting n-butyl alcohol, combining extraction liquid respectively, obtain acetic acid
Ethyl ester extract；By Ethyl acetate fraction through silica gel 200-300 mesh pillar layer separations, successively with methylene chloride-methanol 50:
1-0:1 gradient elution, 11 fraction Fr.1-11 are obtained, wherein fraction Fr.3 is again through silica gel column chromatography, using dichloromethane-first
Alcohol, 30:1,20:1,10:1,5:1,3:1,1:1 elution, MPLC, using methanol-water, 20:80-80:20 gradient elutions and
Sephadex LH-20 column chromatography methods purify, isolated compound screwtree root alkali, the screwtree root alkali by the E that can not separate and
Two kinds of configurations of Z are with 1:1 ratio forms.
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| CN201410505723.7A CN105439878B (en) | 2014-09-28 | 2014-09-28 | 1,3 indandione class compound screwtree root alkali and its medicinal usage |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0171342B1 (en) * | 1984-07-20 | 1987-10-07 | Cortial S.A. | Substituted 2-(1-amino-akylidenyl)1,3-indane diones, process for their preparation and their therapeutical use |
| CN101810653A (en) * | 2010-04-23 | 2010-08-25 | 林兴 | Application of wild sesame seed extract in preparation of anti-hepatic fibrosis medicament |
| CN102188461A (en) * | 2011-04-22 | 2011-09-21 | 林兴 | Application of screwtree root extractive in the preparation of anti-HBV medicament |
-
2014
- 2014-09-28 CN CN201410505723.7A patent/CN105439878B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0171342B1 (en) * | 1984-07-20 | 1987-10-07 | Cortial S.A. | Substituted 2-(1-amino-akylidenyl)1,3-indane diones, process for their preparation and their therapeutical use |
| CN101810653A (en) * | 2010-04-23 | 2010-08-25 | 林兴 | Application of wild sesame seed extract in preparation of anti-hepatic fibrosis medicament |
| CN102188461A (en) * | 2011-04-22 | 2011-09-21 | 林兴 | Application of screwtree root extractive in the preparation of anti-HBV medicament |
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