CN105175265A - Novel diterpenoid compound for treating liver cancer - Google Patents
Novel diterpenoid compound for treating liver cancer Download PDFInfo
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- C07C69/00—Esters of carboxylic acids; Esters of carbonic or haloformic acids
- C07C69/612—Esters of carboxylic acids having a carboxyl group bound to an acyclic carbon atom and having a six-membered aromatic ring in the acid moiety
- C07C69/618—Esters of carboxylic acids having a carboxyl group bound to an acyclic carbon atom and having a six-membered aromatic ring in the acid moiety having unsaturation outside the six-membered aromatic ring
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- C07C67/00—Preparation of carboxylic acid esters
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C67/00—Preparation of carboxylic acid esters
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Abstract
The invention discloses a novel diterpenoid compound for treating a liver cancer, belongs to the field of medicines, and particularly relates to the novel diterpenoid compound obtained by separation from a dry whole herb of plectranthus excises, and a preparation method and medical application thereof. Based on a first report, the compound can be obtained by extraction, separation and purification from the dry whole herb of the plectranthus excises, and is high in purity. An in-vitro test proves that the compound can effectively inhibit the growth and proliferation of HepG2 cells, the decrease degree and the concentration of the compound (I) present a certain dosage effect trend, and a medicine for treating the liver cancers can be further researched and developed.
Description
Technical field
The invention belongs to pharmaceutical field, be specifically related to be separated a kind of new diterpenoids compound obtained and preparation method thereof and medicinal use from the dry herb of rabdosiaexcisa.
Background technology
Rabdosiaexcisa Isodonexcisa (maxim.) hara belongs to Rabdosia plant, and widely distribute at east Asia and western part, Africa, the whole world about has 150 kinds, and about there are 90 kinds of 25 mutation in China, is distributed widely in all parts of the country.Wherein about have 30 kinds among the people can hyoscine, mainly as clearing heat and detoxicating, anticancer, anti-inflammatory, invigorating the spleen, invigorate blood circulation, antimicrobial drug uses.
Rabdosiaexcisa contains diterpene, triterpene, sterol, lipid acid and a small amount of flavones, sesquiterpene, connects the various active compositions such as hydrocarbon glucoside, wherein based on Diterpenoids from bulbus, there will be a known more than 100 and plants.In recent years, by pharmacological evaluation, people's extraction and isolation from these kind of plant, to multiple diterpene-kind compound, and proves that part diterpene compound has good antibacterial, anti-inflammatory, anti-oxidant, immunity moderation power and anti-tumor activity.
Present research shows that rabdosiaexcisa has the pharmacological actions such as antitumor, anti-oxidant, immunomodulatory, antibacterial, anti-inflammatory.The diterpene-kind compound that rabdosiaexcisa extracts can suppress the growth of human body nasopharyngeal carcinoma cell, cervical cancer cell etc. significantly.Rabdosiaexcisa extract generates the restraining effect having highly significant to the superoxide in mouse isolated viscus tissue; show that the compound wherein contained can suppress oxidizing substance in tissue juice and cell to the Oxidative demage of unsaturated fatty acids, to reach Cell protection effect.The immunological competence of body lowly easily receives the invasion of virus and bacterium, or impaired cell, downright bad cell, distortion cell can not get removing fast, just easily under attack, anti-virus ability is weak, relatively normal people easily suffers from the disease, under the same conditions or disease aggravation.Study by experiment, find that this plant can strengthen specific immunity and the non-specific immunity of mouse.The research of rabdosiaexcisa Antibacterial Constituents starts from 1954, and the ethanol extraction of this plant can suppress the growth of gram positive bacterium (Gram-positivebacteria) to have scholar's research to find.In addition, clinically worm snakebite, wound, anti-inflammatory antipyretic are had to effect for the treatment of.
Summary of the invention
The object of the present invention is to provide and a kind ofly from the dry herb of rabdosiaexcisa, be separated a kind of new diterpenoids compound obtained;
Another object of the present invention is to the preparation method that this new compound is provided;
Another object of the present invention is the medicinal use providing this compound for the preparation of Hepatoma therapy medicine.
Above-mentioned purpose of the present invention is achieved by technical scheme below:
There is the compound (I) of following structural formula,
The preparation method of described compound (I), comprise following operation steps: the dry herb of rabdosiaexcisa is pulverized by (a), extract with 70 ~ 80% alcohol heat reflux, united extraction liquid, be concentrated into without alcohol taste, use sherwood oil, ethyl acetate and water saturated n-butanol extraction successively, obtain petroleum ether extract, acetic acid ethyl ester extract and n-butyl alcohol extract respectively; Acetic acid ethyl ester extract macroporous resin removal of impurities in (b) step (a), first use 10% ethanol elution, 10 column volumes, then use 75% ethanol elution, 12 column volumes, collect 75% ethanol eluate, concentrating under reduced pressure obtains 75% ethanol elution thing medicinal extract; C in () step (b), 75% ethanol elution medicinal extract purification on normal-phase silica gel is separated, obtain 5 components successively with the methylene chloride-methanol gradient elution that volume ratio is 80:1,40:1,20:1,10:1 and 1:1; D in () step (c), component 3 is separated further by purification on normal-phase silica gel, obtain 3 components successively with the methylene chloride-methanol gradient elution that volume ratio is 25:1,20:1 and 10:1; E in () step (d), component 2 reverse phase silica gel of octadecylsilane bonding is separated, with the methanol aqueous solution isocratic elution that concentration expressed in percentage by volume is 70%, collect 9 ~ 13 column volume elutriants, elutriant concentrating under reduced pressure obtains pure compound (I).
Further, described macroporous resin is AB-8 type macroporous adsorbent resin.
Further, described alcohol heat reflux extracts the alcohol concn adopted is 75%.
A kind of pharmaceutical composition, the described compound (I) wherein containing treatment significant quantity and pharmaceutically acceptable carrier.
Described compound (I) application in the medicine preparing Hepatoma therapy.
The application of described pharmaceutical composition in the medicine preparing Hepatoma therapy.
When the compounds of this invention is used as medicine, directly can uses, or use with the form of pharmaceutical composition.
This pharmaceutical composition contains the compounds of this invention (I) for the treatment of significant quantity, and all the other are acceptable on pharmacology, nontoxic to humans and animals and pharmaceutically acceptable carrier of inertia and/or vehicle.
Described pharmaceutically acceptable carrier or vehicle are that one or more are selected from solid, semisolid and liquid diluent, filler and pharmaceutical preparation assistant agent.Pharmaceutical composition of the present invention is used with the form of per weight dose.Medicine of the present invention is applied to by form that is oral or injection the patient needing treatment.For time oral, tablet, slow releasing tablet, controlled release tablet, capsule, dripping pill, micropill, suspensoid, emulsion, powder or granule, oral liquid etc. can be made into; During for injecting, can be made into water-based or oily solution, aseptic powder injection, liposome or the emulsion etc. of sterilizing.
Accompanying drawing explanation
Fig. 1 is compound (I) structural formula;
Fig. 2 is that the theoretical ECD value of compound (I) compares with experiment ECD value;
Fig. 3 is the impact (n=3) of compound (I) on HepG2 and L02 cell proliferation.
Embodiment
Further illustrate essentiality content of the present invention below in conjunction with embodiment, but do not limit scope with this.Although be explained in detail the present invention with reference to preferred embodiment, those of ordinary skill in the art should be appreciated that and can modify to technical scheme of the present invention or equivalent replacement, and does not depart from essence and the scope of technical solution of the present invention.
Embodiment 1: compound (I) is separated preparation and structural identification
Medicinal material and reagent source: ethanol, sherwood oil, ethyl acetate, propyl carbinol, methylene dichloride are analytical pure, purchased from Shanghai Ling Feng chemical reagent company limited, methyl alcohol, analytical pure, purchased from Jiangsu Han Bang chemical reagent company limited.The dry herb of rabdosiaexcisa is purchased from Hui nationality's Chinese Medicinal Materials Markets, Guangdong, the place of production.
Preparation method: the dry herb (8kg) of rabdosiaexcisa is pulverized by (a), (25L × 3 time) are extracted with 75% alcohol heat reflux, united extraction liquid, be concentrated into without alcohol taste (6L), sherwood oil (6L × 3 time), ethyl acetate (6L × 3 time) and water saturated propyl carbinol (6L × 3 time) is used to extract successively, concentrated, obtain petroleum ether extract, acetic acid ethyl ester extract (315g) and n-butyl alcohol extract respectively; B in () step (a), acetic acid ethyl ester extract dissolved in purified water is to 2L, medical absorbent cotton filters, filtrate is separated with AB-8 type macroporous resin (1.5kg), use 10% ethanol (10L) and 75% (12L) ethanol elution successively, collect 75% ethanol eluate, concentrating under reduced pressure obtains 75% ethanol elution thing medicinal extract (155g); C in () step (b), 75% ethanol elution medicinal extract 200-300 order purification on normal-phase silica gel is separated, successively with volume ratio be 80:1 (10 column volumes), the methylene chloride-methanol gradient elution of 40:1 (8 column volumes), 20:1 (8 column volumes), 10:1 (8 column volumes) and 1:1 (5 column volumes) obtains 5 components; D component 3 (36g) 200-300 order purification on normal-phase silica gel is separated further in () step (c), successively with volume ratio be 25:1 (8 column volumes), the methylene chloride-methanol gradient elution of 20:1 (8 column volumes) and 10:1 (5 column volumes) obtains 3 components; E in () step (d), component 2 (12g) the reverse phase silica gel ODS-C18 of octadecylsilane bonding is separated, with the methanol aqueous solution isocratic elution that concentration expressed in percentage by volume is 70%, collect 9 ~ 13 column volume elutriants, elutriant concentrating under reduced pressure obtains pure compound (I) (25mg).
Structural identification: HR-ESIMS shows [M+Na]
+for m/z539.2608, can obtain molecular formula in conjunction with nuclear-magnetism feature is C
29h
40o
8, degree of unsaturation is 10.Hydrogen nuclear magnetic resonance modal data δ
h(ppm, DMSO-d
6, 500MHz): H-1 (2.52, t, J=13.5), H-1 (1.46, br, d, J=13.5), H-2 (2.51, m), H-3 (5.53, t, J=4.0), H-4 (2.08, dd, J=11.0, 4.0), H-5 (4.84, dd, J=11.0, 4.5), H-7 (1.79, t, J=15.5), H-7 (1.25, dd, J=15.5, 4.0), H-8 (1.97, m), H-8 (1.48, t, J=13.5), H-9 (3.40, dd, J=10.0, 5.5), H-11 (2.54, d, J=1.5), H-12 (4.17, d, J=1.5), H-16 (1.05, d, J=7.5), H-17 (0.71, s), H-18 (1.06, s), H-19 (0.53, s), H-20 (0.70, s), H-2 ' (7.57, br, d, J=7.0), H-3 ' (7.33, br, t, J=7.0), H-4 ' (7.35, br, t, J=7.0), H-5 ' (7.34, br, t, J=7.0), H-6 ' (7.58, br, d, J=7.0), H-7 ' (7.69, d, J=16.5), H-8 ' (6.62, d, J=16.5), 5-OH (4.04, d, J=4.5), 9-OH (3.60, d, J=5.5), 11-OH (5.37, s), 12-OH (5.37, s), 15-OH (4.55, s), carbon-13 nmr spectra data δ
c(ppm, DMSO-d
6, 125MHz): 41.7 (CH
2, 1-C), 35.8 (CH, 2-C), 78.0 (CH, 3-C), 54.1 (CH, 4-C), 63.9 (CH, 5-C), 49.5 (C, 6-C), 31.8 (CH
2, 7-C), 28.6 (CH
2, 8-C), 79.7 (CH, 9-C), 38.4 (C, 10-C), 59.0 (CH, 11-C), 56.6 (CH, 12-C), 56.6 (C, 13-C), 207.3 (C, 14-C), 85.7 (C, 15-C), 15.9 (CH
3, 16-C), 18.3 (CH
3, 17-C), 25.2 (CH
3, 18-C), 11.7 (CH
3, 19-C), 10.1 (CH
3, 20-C), 134.8 (C, 1 '-C), 128.3 (CH, 2 '-C), 127.2 (CH, 3 '-C), 130.5 (CH, 4 '-C), 129.2 (CH, 5 '-C), 128.3 (CH, 6 '-C), 145.1 (CH, 7 '-C), 118.7 (CH, 8 '-C), 167.1 (C, 9 '-C), carbon atom mark is see Fig. 1.Infrared spectra shows that this compound contains hydroxyl (3584,3500 and 3377cm
-1), carbonyl (1708 and 1689cm
-1) and aromatic ring (1604 and 1498cm
-1) group.
1this compound of HNMR spectrum display contains a trans-cinnamic acid ester group (δ H7.57 (br, d, J=7.0Hz, H-2 '), 7.33 (br, t, J=7.0Hz, H-3 '), 7.35 (br, t, J=7.0Hz, H-4 '), 7.34 (br, t, J=7.0Hz, H-5 '), 7.58 (br, d, J=7.0Hz, H-6 '), 7.69 (d, J=16.5Hz, H-7 '), 6.62 (d, J=16.5Hz, H-8 '), δ C134.8 (1 '-C), 128.3 (2 '-C), 127.2 (3 '-C), 130.5 (4 '-C), 129.2 (5 '-C), 128.3 (6 '-C), 145.1 (7 '-C), 118.7 (8 '-C), 167.1 (9 '-C)), five containing oxygen methyne (δ H5.53 (t, J=4.0Hz, H-3), 4.84 (dd, J=11.0 and 4.5Hz, H-5), 3.40 (dd, J=10.0Hz and 5.5Hz, H-9), 2.54 (d, J=1.5Hz, H-11) and 4.17 (d, J=1.5Hz, H-12)), three methylene radical, and five methyl (δ H1.05 (d, J=7.5Hz, H
3-16), 0.71 (s, H
3-17), 1.06 (s, H
3-18), 0.53 (s, H
3-19) and 0.70 (s, H
3-20)).In HMBC spectrum, the dependency of H-3 and C-9 ' confirms that this styracin ester group is positioned on C-3 position.Except trans-cinnamate acid ester moiety,
13cNMR and DEPT composes display 20 carbon signals, also comprises five quaternary carbons (carbonyl carbon δ C207.3 and containing oxygen carbon δ C85.7) except the functional unit that correspondence is above-mentioned.Above-mentioned nuclear magnetic data shows that this compound is a diterpene alcohol laurate compounds.In HMBC spectrum, H
2-1 and C-2, C-3, C-4, C-15 and C-16; H-3 and C-1, C-15, and C-9 '; H
3-16 and C-1, C-2 and C-3; The dependency of OH-15 and C-4 and C-14, and in conjunction with the chemical shift of these protons with resonance carbon, show that this Compound C-2, C-3 and C-15 position are connected with methyl, laurate fragment and hydroxyl respectively.H-4 and C-5 and C-6 in being composed by HMBC; H-5 and C-3, C-4, C-6 and C-17; H
3-17 and C-5, C-6 and C-13; H
3-20 and C-6, C-13 and C-14; OH-5 and C-4, C-5 and C-6; The dependency of OH-15 and C-14, known pimelinketone ring is connected with five-ring by C-4 with C-15, and C-5 position is connected with a hydroxyl, C-6 and C-13 position is connected with a tertiary methyl respectively.In addition, H-9 and H-11 and C-18 and C-19; H-11 and C-12 and C-13; H-12 and C-13 and C-20; H
3-17 and C-7; H
3-18 and H
3-19 and C-9, C-10 and C-11; H
3-20 and C-12; The dependency of OH-9 and C-9 and C-10 shows that this compound contains an octatomic ring, is connected with six-ring by C-6 with C-13, C-9, C-11, and C-12 position is respectively connected with a hydroxyl, and C-10 position is connected with two methyl.In NOESY spectrum, H-5 and H-12 and OH-15; H-12 and H
3the dependency of-19 shows that they are beta comfiguration.In addition, H-3 and H-2, H-4 and OH-5; H-4 and H-2, H-3 and H
3-17; H-9 and H-11 and H
3the dependency of-18 shows that these protons are α configuration.Comprehensive hydrogen spectrum, carbon spectrum, HMBC spectrum and NOESY spectrum, and document is about correlation type nuclear magnetic data, can substantially determine this compound as shown in Figure 1, steric configuration is determined further by ECD test, theoretical value and experimental value basically identical (Fig. 2).
Embodiment 2: compound (I) pharmacological action is tested
One, material and instrument
HepG2 human hepatoma cell strain is purchased from Chinese Academy of Sciences's biological chemistry and Institute of Cell Biology.L02 normal liver cell strain presented by Zhong Shan hospital of Fudan University liver cancer research.Compound (I) is made by oneself, and HPLC normalization method purity is greater than 98%.RPMI-1640 substratum, pancreatin are purchased from Gibco company.Standard calf serum is purchased from Biowest company.3-(4,5-dimethylthiazole-2)-2,5-diphenyltetrazolium bromide bromine salt (MTT), dimethyl sulfoxide (DMSO) (DMSO), trypan blue (TrypanBLue), dichlorofluorescein diacetate (DCFH-DA) is all purchased from Shanghai Chemical Reagent Co., Ltd., Sinopharm Group.Other common agents are analytical pure.
Microbalance (Switzerland plum Teller-Tuo benefit, METTLRERAE2000).Common desktop whizzer (Anting Scientific Instrument Factory, Shanghai).Digital display electric heating constant-temperature water-bath tank (Shanghai leap medical machinery factory, HH.W21.600S).Carbon dioxide cell incubator (FormaScientific company of the U.S.).High speed freezing centrifuge (Thermo company of the U.S., IECMICROMAXRF).Ultramicron ultraviolet spectrophotometer (ThermoHsher company of the U.S., NanoDrop-1000).Ultraviolet imagery system (Shanhai Furi Science and Technology Co., Ltd., FR-200A).Enzyme-linked immunosorbent assay instrument, spectrophotofluorometer (HitachiF2500).
Two, test method
1, cell cultures
Cell routine is inoculated in the RPMI-1640 substratum containing 10% (volume fraction) calf serum, is placed in 37 DEG C, 5%CO
2cultivate in the incubator of concentration.
2, compound (I) intervenes the configuration of nutrient solution
45.44mg compound (I) be dissolved in the obtained storing solution of 10mL dimethyl sulfoxide (DMSO) (DMSO) and dilute 2 times, 4 times respectively, obtaining compound (I) the application liquid of 2mmol/L, 4mmol/L and 8mmol/L respectively.Before carrying out cell intervention, 50 μ L application liquid are fully mixed with 4950 μ L cellar culture liquid (calf serum containing 10% volume fraction), obtains compound (I) and intervene nutrient solution (20 μm of ol/L, 40 μm of ol/L, 80 μm of ol/L).
3, mtt assay detects cell survival rate
The vegetative period cell of taking the logarithm is inoculated in 96 well culture plates, every hole 200 μ L (5 × 10
3cell).Cultivate 12h after cell attachment, discard original fluid, compound (I) nutrient solution adding 200 μ L different concns (mixes with cellar culture liquid with 1% volume fraction after DMSO dissolved compound (I), compound (I) final concentration is made to be 20 μm of ol/L, 40 μm of ol/L, 80 μm of ol/L), often organize 6 parallel holes.Establish 6 solvent (DMSO) control wells simultaneously.Cultivate 24h, 48h, 72h, within every 24 hours, change compound (I) and intervene nutrient solution.Intervene after terminating, suck nutrient solution, add 20 μ LMTT solution to each hole, 37 DEG C hatch 4h after suck each hole supernatant liquor, add 150 μ LDMSO, put low speed concussion 10min on shaking table, after abundant dissolving to be crystallized, with DMSO zeroing, measure each hole absorbancy with enzyme-linked immunosorbent assay instrument at 490nm place, calculate cell survival rate (survivalrate, SR).SR=experimental group A
490/ control group A
490× 100%
4, trypan blue Ran Se Measuring determines survivaling cell number
By equal amts (2 × 10
6) cell be inoculated in Tissue Culture Flask, cultivate for some time after cell attachment, discard nutrient solution, after washing twice, cell with PBS, the compound (I) adding 5mL intervenes nutrient solution (20 μm of ol/L, 40 μm of ol/L, 80 μm of ol/L).Often organize three bottles of cells, set up three bottles of solvent (DMSO) contrasts simultaneously.After cultivating 48h, use 0.25% trypsin digestion cell, collecting cell suspension, determination of trypan blue staining cell count.
5, DCFH-DA method measures ROS activity in cell
By equal amts (2 × 10
6) cell be inoculated in Tissue Culture Flask, cultivate for some time after cell attachment, suck original fluid, add 5mL compound (I) and intervene nutrient solution, after continuing to cultivate 48h, discard nutrient solution, trypsin digestion cell, collecting cell suspension is in 1.5mLEP pipe, the centrifugal 5min of 1500rpm, abandons supernatant, in precipitation, add 1mLDCFH-DA, piping and druming mixing, hatch 30min for 37 DEG C, add PBS termination reaction, centrifugal collecting precipitation is also dissolved in 1mLPBS, blow and beat uniformly cell suspension, burn light spectrophotometric determination fluorescent value with Hitachi-F2500.Excitation wavelength 488nm, emission wavelength 525nm.
Three, result and conclusion
1, Growth of Cells survival rate
HepG2 cell is after compound (I) intervention cultivates 24h, 48h, 72h, the cell survival rate of basic, normal, high dosage group is all remarkable in solvent control group (P<0.05), and the degree that declines of cell survival rate and compound (I) are intervened between concentration and be there is doses effect trend (table 1
ap<0.05VS control group;
bp<0.05VS low dose group;
cdosage group in P<0.05VS).Under same compound (I) intervenes concentration, cell survival rate presents downward trend along with the increase of intervention time.And under equal conditions, L02 Human normal hepatocyte is after compound (I) is intervened, cell survival rate is without noticeable change (table 2).Trypan Blue cell counts consistent with MTT (Fig. 3,
ap<0.05VS control group;
bp<0.05VS low dose group;
cdosage group in P<0.05VS).
2, in cell, ROS is active
HepG2 human liver cancer cell is after different concns compound (I) is intervened, and compared with control group, intracellular ROS level significantly declines (P<0.05).The results are shown in Table 3 (
ap<0.05VS control group;
bp<0.05VS low dose group).
Conclusion, the compound (I) of different concns is adopted to carry out intervention process to human liver cancer cell HepG2, show through MTT colorimetric and Trypan Blue cell counting, compound (I) effectively can suppress the growing multiplication of HepG2 cell, the survival rate of intervention group cell comparatively control group significantly reduces, and reduction degree and compound (I) concentration present certain dosage effect trend, simultaneously under same compound (I) concentration, cell survival rate presents the trend continuing to reduce along with the increase of intervention time.Meanwhile, compound (I) intervention of same dose does not produce restraining effect to L02 normal liver cell.
Table 1 compound (I) is on the impact (x ± s, n=6) of HepG2 Growth of Cells survival rate
Table 2 compound (I) is on the impact (x ± s, n=6) of L02 Growth of Cells survival rate
Table 3 compound (I) is on the impact (x ± s, n=6) of HepG2 cell ROS level
| Group | Dosage (μm ol/L) | ROS(Fluounit/mgprot) |
| Control group | 0 | 4189±204 |
| Low dosage | 20 | 3215±151 a |
| Middle dosage | 40 | 3080±196 a |
| High dosage | 80 | 2831±212 ab |
Embodiment 3
The preparation of tablet: by embodiment 1 method first obtained compound (I), and the salt utilizing organic acid to make as tartrate or citric acid or formic acid or oxalic acid etc., mineral acid example hydrochloric acid or sulfuric acid or phosphoric acid, vehicle is added, pelletizing press sheet than the ratio for 1:9 in itself and excipient weight.
Embodiment 4
The preparation of oral liquid: by embodiment 1 method first obtained compound (I), and the salt utilizing organic acid to make as tartrate or citric acid or formic acid or oxalic acid etc., mineral acid example hydrochloric acid or sulfuric acid or phosphoric acid, oral liquid method for making makes oral liquid routinely.
Embodiment 5
The preparation of capsule or granule: by embodiment 1 method first obtained compound (I), and the salt utilizing organic acid to make as tartrate or citric acid or formic acid or oxalic acid etc., mineral acid example hydrochloric acid or sulfuric acid or phosphoric acid, add vehicle in itself and excipient weight than the ratio for 1:9, make capsule or granule.
Embodiment 6
The preparation of injection liquid: by embodiment 1 method first obtained compound (I), and the salt utilizing organic acid to make as tartrate or citric acid or formic acid or oxalic acid etc., mineral acid example hydrochloric acid or sulfuric acid or phosphoric acid, inject with water routinely, essence filter, injection liquid is made in embedding sterilizing.
Embodiment 7
The preparation of aseptic powder injection: by embodiment 1 method first obtained compound (I), and the salt utilizing organic acid to make as tartrate or citric acid or formic acid or oxalic acid etc., mineral acid example hydrochloric acid or sulfuric acid or phosphoric acid, be dissolved in sterile water for injection, stirring makes molten, filter with aseptic suction funnel, aseptic essence filter again, be sub-packed in ampoule, after frozen drying, aseptic sealing by fusing obtains powder injection.
The effect of above-described embodiment is essentiality content of the present invention is described, but does not limit protection scope of the present invention with this.Those of ordinary skill in the art should be appreciated that and can modify to technical scheme of the present invention or equivalent replacement, and does not depart from essence and the protection domain of technical solution of the present invention.
Claims (7)
1. there is the compound (I) of following structural formula,
2. the preparation method of compound according to claim 1 (I), it is characterized in that comprising following operation steps: the dry herb of rabdosiaexcisa is pulverized by (a), extract with 70 ~ 80% alcohol heat reflux, united extraction liquid, be concentrated into without alcohol taste, use sherwood oil, ethyl acetate and water saturated n-butanol extraction successively, obtain petroleum ether extract, acetic acid ethyl ester extract and n-butyl alcohol extract respectively; Acetic acid ethyl ester extract macroporous resin removal of impurities in (b) step (a), first use 10% ethanol elution, 10 column volumes, then use 75% ethanol elution, 12 column volumes, collect 75% ethanol eluate, concentrating under reduced pressure obtains 75% ethanol elution thing medicinal extract; C in () step (b), 75% ethanol elution medicinal extract purification on normal-phase silica gel is separated, obtain 5 components successively with the methylene chloride-methanol gradient elution that volume ratio is 80:1,40:1,20:1,10:1 and 1:1; D in () step (c), component 3 is separated further by purification on normal-phase silica gel, obtain 3 components successively with the methylene chloride-methanol gradient elution that volume ratio is 25:1,20:1 and 10:1; E in () step (d), component 2 reverse phase silica gel of octadecylsilane bonding is separated, with the methanol aqueous solution isocratic elution that concentration expressed in percentage by volume is 70%, collect 9 ~ 13 column volume elutriants, elutriant concentrating under reduced pressure obtains pure compound (I).
3. the preparation method of compound according to claim 2 (I), is characterized in that: described macroporous resin is AB-8 type macroporous adsorbent resin.
4. the preparation method of compound according to claim 2 (I), is characterized in that: it is 75% that described alcohol heat reflux extracts the alcohol concn adopted.
5. a pharmaceutical composition, the compound according to claim 1 (I) wherein containing treatment significant quantity and pharmaceutically acceptable carrier.
6. compound according to claim 1 (I) application in the medicine preparing Hepatoma therapy.
7. the application of pharmaceutical composition according to claim 5 in the medicine preparing Hepatoma therapy.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105153084A (en) * | 2015-09-13 | 2015-12-16 | 金丽秋 | Novel diterpene compound as well as preparation method and medicinal application thereof |
| CN105254502A (en) * | 2015-11-25 | 2016-01-20 | 吕涛 | New diterpenoid compound and preparation method and medical application thereof |
| CN105481875A (en) * | 2015-12-29 | 2016-04-13 | 吴金凤 | New limonin compound as well as preparation method and medical application thereof |
| CN105534992A (en) * | 2015-12-27 | 2016-05-04 | 温州统益生物医药科技有限公司 | Medicine for protecting liver |
| CN105566251A (en) * | 2016-02-20 | 2016-05-11 | 杭州富阳伟文环保科技有限公司 | Novel leucothoe alkane type diterpene compound and preparation method and medical application thereof |
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| TIAN, YE等: "Diterpenoids with Diverse Skeletons from the Roots of Euphorbia micractina", 《JOURNAL OF NATURAL PRODUCTS》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105153084A (en) * | 2015-09-13 | 2015-12-16 | 金丽秋 | Novel diterpene compound as well as preparation method and medicinal application thereof |
| CN105254502A (en) * | 2015-11-25 | 2016-01-20 | 吕涛 | New diterpenoid compound and preparation method and medical application thereof |
| CN105534992A (en) * | 2015-12-27 | 2016-05-04 | 温州统益生物医药科技有限公司 | Medicine for protecting liver |
| CN105481875A (en) * | 2015-12-29 | 2016-04-13 | 吴金凤 | New limonin compound as well as preparation method and medical application thereof |
| CN105566251A (en) * | 2016-02-20 | 2016-05-11 | 杭州富阳伟文环保科技有限公司 | Novel leucothoe alkane type diterpene compound and preparation method and medical application thereof |
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Application publication date: 20151223 |