CN104086559A - Crystal form of diterpenoid compound - Google Patents
Crystal form of diterpenoid compound Download PDFInfo
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Abstract
The invention provides a crystal form of a compound as shown in the formula II. The crystal form belongs to an orthorhombic crystal system; the space group is P212121; the crystal cell parameters are as shown in the specification, wherein alpha is equal to 90.00 DEG, beta is equal to 99.00 DEG, and gamma is equal to 90.00 DEG; Z is equal to 4; the crystal cell volume is as shown in the specification. The invention also provides a preparation method and application of the crystal form. The compound with the crystal form in the formula II, which is separated from leonurus japonicus, has a certain inhibiting effect on platelet aggregation and has a better effect than clopidogrel bisulfate as an anti-platelet aggregation drug under the same concentration (0.1mM), so that a new choice is provided for developing a novel natural anti-platelet aggregation drug.
Description
Technical field
The present invention relates to a kind of new crystal of diterpene-kind compound.
Background technology
Motherwort Herb begins to be loaded in < < Shennong Bencaojing > >, and original name motherwort, is listed in top grade.In < < Compendium of Materia Medica > >, being referred to as " panacea of blood man ", curing mainly " threatened abortion difficult labour, retention of placenta; bruise, blood wind, blood pain; metrorrhagia and metrostaxis, hematuria ", is gynaecology's multiparity key medicine.
In Motherwort Herb, mainly contain the compositions such as alkaloid, diterpene, flavones, iridoid glycoside, benzyl carbinol glycosides, volatile oil, peptide class and fatty acid, what activity research was more at present is mainly alkaloid and diterpenes:
(1) alkaloid: mainly comprise stachydrine, syringic acid.delta.-guanidinobutyl ester, Motherwort Herb pyridine and leonurinine etc.Modern study shows that Herba Leonuri total alkaloids has a therapeutic action to cerebral ischemia re-pouring injured, and wherein, syringic acid.delta.-guanidinobutyl ester not only can be protected the myocardial cell of ischemic injuries by antioxygenation, also have neuroprotective activity.
(2) diterpene: in recent years its study of active components is increased, wherein Leoheteronin A and the obvious acetylcholine esterase inhibition activity of Leopersin G, having good prospect aspect treatment alzheimer's disease; Prehispanolone is a kind of thrombocyte antagonism factor, can anticoagulant; Leojaponin has provide protection to the pallium cell of injured with glucose deprivation; The labdane type diterpenoids that Khan obtains has anti-inflammatory action etc.
Although along with people are to the intensification of Motherwort Herb understanding and the progress of modern science and technology, the research of Motherwort Herb drug effect position and activeconstituents is carried out rapidly, diterpenes chemical constitution study more, activity research is relatively less.It is relevant that antiplatelet aggregative activity and traditional effect of Motherwort Herb " are invigorated blood circulation ", but the research of relevant anticoagulant composition extremely lacks.As can be seen here, from Motherwort Herb, obtain anticoagulant composition and there is good prospect and researching value.
Summary of the invention
The object of the present invention is to provide a kind of new crystal of furans lactonic ring analog derivative.
The invention provides the crystal formation of compound shown in formula II,
This crystal formation is rhombic system, and spacer is P2
12
12
1, unit cell parameters is
α=90.00 °,
β=90.00 °,
γ=90.00 °, Z=4, unit cell volume is
Further, the specific rotatory power of described crystal formation Chinese style I compound is [α]
20 d-58.8.
The present invention also provides the preparation method of above-mentioned crystalline form I, and it comprises following operation steps:
(1) take Motherwort Herb medicinal material, pulverize, add 95%v/v extraction using alcohol, after extracting solution decompression and solvent recovery, obtain ethanol extract;
(2) by ethanol extract water-dispersion, be extracted with ethyl acetate successively, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract;
(3) adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out and partly go up anti-phase polystyrene type resin post, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part, pass through again medium pressure liquid chromatography, octadecyl silane chromatographic column, 50-95%v/v methanol aqueous solution gradient elution, collect 70～75% methanol aqueous solution wash-out parts, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, collect elutriant, reclaim solvent, anti-phase liquid phantom preparing chromatogram on residuum, 82%v/v methanol aqueous solution wash-out, collect respectively the elutriant of different appearance times, can obtain formula II compound,
(4) modus ponens II compound, adds acetone, is placed on subsequently in the moisture eliminator that siccative is housed, and is placed to crystal and separates out, and obtains crystal formation.
Further, the mass volume ratio of formula II compound and acetone is: 10mg:0.1-1ml.
Further, the mass volume ratio of formula II compound and acetone is: 10mg:0.3-0.5ml.
Preferably, the mass volume ratio of formula II compound and acetone is: 10mg:0.4ml.
Further, described siccative is selected from discolour silica gel.The present invention also provides the purposes of above-mentioned crystalline form I in preparing platelet aggregation-against class medicine.
The present invention also provides a kind of pharmaceutical composition, and it is the preparation that contains above-mentioned crystal formation.
The present invention can adopt preparation technique means or the pharmaceutical methods of this area routine, and crystal formation of the present invention is prepared into suitable medicine type, comprises: tablet, injection, tincture, suppository, capsule, paste (ointment, ointment), ophthalmic preparation, pill, implant, syrup, mist agent (aerosol, powder inhalation, sprays), film, granule, oral solution (oral suspensions, Orally taken emulsion), powder, aural preparations, nasal formulations, lotion (irrigation, enema), liniment (paint, liniment), gelifying agent, patch etc.; Be preferably tablet, capsule, ophthalmic preparation.Wherein, described tablet is selected from lozenge, Sublingual tablet, mouth paster, chewable tablet, dispersible tablet, fuse, effervescent tablet, slow releasing tablet, controlled release tablet, enteric coated tablet etc.; Described injection is selected from injection, transfusion, freeze-dried powder, emulsion, implant, microball preparation, pellet preparations etc.; Described capsule is selected from hard capsule, soft capsule, slow releasing capsule, controlled release capsule and enteric coated capsule etc.; Described ophthalmic preparation is selected from eye drops, eye wash, intraocular injection solution, Eye ointments, ointment, gel for eye, eye mask agent, eye pill, intraocular intercalating agent etc. for eye; Described pill is selected from dripping pill, sugar-pill etc.; Described granule is selected from mix suspension grain, effervescent granule, enteric coated particles, slow-releasing granules, controlled release granule etc.; Described aural preparations is selected from ear drop, ear lotion, sprays for ear, ointment for ear, ointment for ear, gelifying agent for ear, earplug, powder for ear, pill etc. for ear; Described nasal formulations is selected from nasal drop, nasal douche, nasal spray, ointment, gel for nose, powder, powder nose inhalant, stylus etc. for nose for nose for nose.
In Motherwort Herb, the separated formula II crystal formation compound obtaining has certain restraining effect to platelet aggregation, under same concentration (0.1mM), than medicament for resisting platelet aggregation " bisulfate clopidogrel " better effects if, for the exploitation of the natural medicament for resisting platelet aggregation of novelty provides new selection.
Below by embodiment, the present invention is described in further detail, but do not limit the present invention, those skilled in the art can make according to the present invention various changes and distortion, only otherwise depart from spirit of the present invention, all should belong to the scope of claims of the present invention.
Accompanying drawing explanation
The single crystal diffraction structure iron of Fig. 1 formula II compound crystal formation
Embodiment
Embodiment 1 (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13R, 15R)-3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula II compound crystal formation)
(1) experiment material:
1. medicinal material
Motherwort Herb is picked up from Wenjiang District, Chengdu, Sichuan, is accredited as the herb of labiate Motherwort Herb Leonurus japonicus Houtt. through professor Li Min of identification and assessment of Chinese medicines teaching and research room of Chengdu University of Traditional Chinese Medicine.
2. reagent and filler
Column chromatography silica gel, 200～300 orders (SILVER REAGENT), are purchased from Qingdao Haiyang silica-gel drier factory;
Tlc silica gel G, GF
254and H (chemical pure), be purchased from Qingdao Haiyang silica-gel drier factory;
MCI gel CHP 20P, 75～150 μ m, are anti-phase polystyrene type resin, are purchased from Mitsubishi chemical company;
Sephadex LH-20 dextrane gel, is purchased from Sweden Amersham company;
GF
254silica gel Preparative TLC, is purchased from Yantai Jiang You silica gel development corporation, Ltd.;
Chromatogram methyl alcohol, 4L/ bottle, is purchased from U.S. Fisher company;
The analytical reagent such as sherwood oil, ethyl acetate, propyl carbinol, acetone, methyl alcohol, are purchased from Chengdu Ke Long chemical reagent factory.
3. laboratory apparatus
Cometro 6000 high performance liquid chromatographs (U.S. Cometro);
Waters Synapt G
2hDMS high resolution flight time mass spectrum (U.S. Waters);
Bruker-AV-400 nuclear magnetic resonance analyser (Switzerland Bruker);
Vector 22 FT-IR infrared spectrometers (Switzerland Bruker);
Shimadzu UV-260 spectrophotometry instrument (Japanese Shimadzu);
Perkin-Elmer 341 polarimetry instrument (U.S. PerkinElmer);
BP211D 100,000/electronic balance (Switzerland Sartorius);
R-210 rotatory evaporator (Switzerland BUCHI);
DZG-6050 type vacuum drying oven (the gloomy letter in Shanghai).
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 70～75% elutriants (H-3-4) that collection contains target compound, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, thin-layer chromatography is followed the trail of, collection contains target compound cut, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 82%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=48min, except desolventizing, gained solid substance is standby.
4. get solid substance 10mg, be placed in the glass sample bottle of 2ml, add 0.4ml acetone.Subsequently sample bottle is put into the moisture eliminator that discolour silica gel is housed, placed until separate out colourless needle, obtain formula II compound crystal formation.
(4) Structural Identification of composition: clear crystal; Without blackening, iodine displaing yellow, spray 10% ethanol solution of sulfuric acid is aobvious orange-yellow at 105 ℃; By HRESIMS m/z 461.2520[M+Na]
+can determine that molecular formula is C
24h
38o
7.Compound
14 unimodal signals of methyl of H-NMR demonstration (δ: 0.90,0.95,1.35,2.04), a methyl signals δ who is connected with methyne
h1.12 (d, J=6.6Hz, H
3-17), a methyl δ who is connected with methylene radical
h1.15 (t, J=7.2Hz, H
3-22), three low connects oxygen methyne signal δ
h5.10 (dd, J=6.0,3.6Hz, H-15), 4.46 (dd, J=3.0,1.8Hz, H-3), 3.78 (d, J=10.8Hz, H-7); Compound
13c-NMR and DEPT spectrum show 24 carbon signals, and DEPT spectrum shows that these carbon signals are respectively 6 methyl, 7 methylene radical (comprising 1 even oxygen carbon), 5 methynes and 6 quaternary carbon signals, wherein have 1 carbonyl (δ
c212.5) and two oxygen carbon (δ of company
c92.7,91.4).In conjunction with above information and reference (Planta Med.2008,74:1288 – 1290.), the two dimensional structure of deterministic compound is 3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.Adopt subsequently 2D NMR experiment to confirm.The relative configuration of compound is composed and is inferred by NOESY, H-5 and H-7 and H
3-18 is relevant, H-7 again with H
3-17 is relevant, shows H-5, H-7, H
3-18, H
3-17 homonymies (α direction) at ring, in like manner, H
3-20 and H-8, H
2-11, H
3-19 is relevant, shows that they are positioned at opposite side (β direction).Further adopt the experiment of X-ray single crystal diffraction to determine absolute configuration, as shown in Figure 1.Therefore, the structure of this compound is defined as (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13R, 15R)-3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone, concrete spectral data is in Table 1.
(5) proton nmr spectra (
1h-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 1.
(6) carbon-13 nmr spectra (
13c-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 2.
(7) single crystal diffraction data are in Table 3.
Table 1.
1h-NMR (600MHz) nuclear magnetic data (solvent: Me
2cO-d
6; δ: ppm; J:Hz)
Table 2.
13c-NMR (150MHz) nuclear magnetic data (is measured solvent: Me
2cO-d
6; δ: ppm)
Table 3 single crystal diffraction data
Reference examples 2 (-)-(5S, 7R, 8R, 9R, 10S, 13R, 15R)-7-hydroxyl-15-oxyethyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula III compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 70～75% elutriants (H-3-4) that collection contains target compound, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, thin-layer chromatography is followed the trail of, collection contains target compound cut, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 82%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=28min, purifying obtains 16.5mg.
(4) Structural Identification of composition: colorless oil; [α]
20 d-92.2 (c0.15, MeOH); Molecular formula: C
22h
36o
5.Its NMR data are more similar to formula II compound.The NMR data of comparative compound III and II, the ethanoyl in II and even oxygen methyne are by the MU (methylene unit) (δ in III
h1.30 and 1.13, δ
c43.2ppm) substitute.In addition, for II, the C-1 in III, C-5 (the Δ δ that deshielded respectively
c+ 6.5 ,+5.0ppm), C-2, C-4 conductively-closed (Δ δ
c-3.8 ,-3.4ppm), disclose 3 desacetoxy analogues that compound III is II, and obtained further confirmation by 2D NMR experiment.The configuration of compd B is by relatively NOESY spectrum and the optical value of B and A are determined.Infer that structure is (-)-(5S, 7R, 8R, 9R, 10S, 13R, 15R)-7-hydroxyl-15-oxyethyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.
(5) proton nmr spectra (
1h-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 1.
(6) carbon-13 nmr spectra (
13c-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 2.
Reference examples 3 (+)-(5S, 7R, 8R, 9R, 10S, 13S, 15R)-7-hydroxyl-15-oxyethyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula IV compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 70～75% elutriants (H-3-4) that collection contains target compound, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, thin-layer chromatography is followed the trail of, collection contains target compound cut, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 82%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=32min, purifying obtains 9mg.
(4) Structural Identification of composition: colorless oil, its Spectral Characteristic and compound III are closely similar, but have contrary specific optical rotation { [α]
20 d+ 114.9 (c0.20, MeOH) }. comprehensively compare HR-ESI-MS and the 2D-NMR data of III and IV, show that both have identical two dimensional structure.Further in detail relatively both are at identical deuterated solvent
1h and
13c-NMR data, find that both fine differences are: for III, H-11b in compounds Ⅳ, H-16a, H-16b, C-15 and C-16 signal move Δ δ-0.13 to High-Field respectively ,-0.08 ,-0.06 ,-0.4 and-0.5ppm.On the contrary, H-11a, H-12b, H-14a, H-14b, C-11 and C-12 signal be respectively to low mobile Δ δ+0.08 ,+0.07 ,+0.03 ,+0.06 ,+0.6 and+0.3ppm.More than show that compounds Ⅳ is the 13S-epimer of compound III, and adopt NOESY to confirm.In NOESY spectrum, H
2-14 and H
3-17 is relevant, H
2-12 and H
2-16 relevant confirmation C-13 are S configuration.Therefore, compounds Ⅳ is defined as (+)-(5S, 7R, 8R, 9R, 10S, 13S, 15R)-7-hydroxyl-15-oxyethyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.
(5) proton nmr spectra (
1h-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 1.
(6) carbon-13 nmr spectra (
13c-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 2.
Reference examples 4 (-)-(5S, 7R, 8R, 9R, 10S, 13S, 15S)-7-hydroxyl-15-methoxyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula V compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 65～70% elutriants (H-3-3) that collection contains target compound, reclaim solvent, residuum is again through Preparative TLC chromatographic separation, Shi You Mi ﹕ acetone=5 ﹕ 1 launches, scraping target product chromatographic band, with acetone 30ml wash-out, collect elutriant, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 78%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=37min, purifying obtains 1.5mg.
(4) Structural Identification of composition: colorless oil, molecular formula: C
21h
34o
5.Its chromatogram characteristic shows it and II, III, IV similar, is another pair of volution Ladanum alkane diterpene.The NMR data of comparative compound V and IV, show that oxyethyl group in IV is by the methoxy substitution (δ in V
h3.26ppm, δ
c54.7ppm).Meanwhile, H
2-14 and H-15 between 6.0 and the 4.2Hz of coupling constant in IV diminish as 5.6 in V and 1.2Hz, and in compound V, C-15 and C-16 have moved Δ δ to low field respectively
c+ 1.9 and+2.9ppm. this show that compound V is 15 'beta '-methoxy derivatives of compounds Ⅳ.Adopt subsequently 2D-NMR experiment to confirm.Adopt II, III, the definite method of IV configuration, determined the absolute configuration of compound V.Therefore, the structure of compound V is defined as (-)-(5S, 7R, 8R, 9R, 10S, 13S, 15S)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.
(5) proton nmr spectra (
1h-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 1.
(6) carbon-13 nmr spectra (
13c-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 2.
Reference examples 5 (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15S)-3-acetoxyl group-7-hydroxyl-15-methoxyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula VI compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 70～75% elutriants (H-3-4) that collection contains target compound, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, thin-layer chromatography is followed the trail of, collection contains target compound cut, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 82%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=54min, purifying obtains 1.6mg.
(4) Structural Identification of composition: colorless oil, HREI-MS m/z 447.2366[M+Na]
+prompting molecular composition is C
23h
36o
7(calcd for C
23h
36o
7na, 447.2359).The NMR data of comparative compound VI and V, show that the 3-acetoxyl group that this compound VI is V replaces analogue.In 1D-NMR spectrum, an oxygen methyne (δ of company in compound VI
h4.47ppm, δ
c78.4ppm) He one acetoxyl group (δ
h2.06ppm, δ
c170.2,21.2ppm) replaced the methylene radical in compound V.Meanwhile, in compound VI C-1 and C-5 respectively to High-Field move Δ δ-6.3 and-5.1ppm, and C-2 and C-4 are respectively to low mobile Δ δ
c+ 3.9 and+3.4ppm, prompting C-3 is that acetoxyl group replaces.The determination methods of compound structure type and II, III, IV, V are identical.Therefore, the structure of compound VI is defined as (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15S)-3-acetoxyl group-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.
(5) proton nmr spectra (
1h-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 1.
(6) carbon-13 nmr spectra (
13c-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 2.
Reference examples 6 (+)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15R)-3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula VII compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 70～75% elutriants (H-3-4) that collection contains target compound, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, thin-layer chromatography is followed the trail of, collection contains target compound cut, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 82%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=51min, purifying obtains 15mg.
(4) Structural Identification of composition: colorless oil; Molecular formula: C
24h
38o
7, M=438, HRESI MS m/z 461.2508[M+Na]
+;
1h NMR (600MHz, CD
3cOCD
3) δ: 0.89 (3H, s, H
3-20), 0.94 (3H, s, H
3-18), 1.34 (3H, s, H
3-19), 1.17 (3H, d, J=6.6Hz, H
3-17), 5.10 (1H, dd, J=5.4,3.6Hz, H-15), 4.46 (1H, br s, H-3), 3.28 (1H, s, H-5), 3.89 (1H, d, J=7.8Hz, H-16a), 3.67 (1H, d, J=7.8Hz, H-16b), 3.71,3.82 (2H, 2m, MeCH
2o), 1.15 (3H, t, J=7.2Hz, MeCH
2o);
13c NMR (150MHz, CD
3cOCD
3) δ: 26.2 (C-1), 22.9 (C-2), 78.5 (C-3), 36.6 (C-4), 52.9 (C-5), 212.7 (C-6), 78.4 (C-7), 48.2 (C-8), 92.7 (C-9), 48.5 (C-10), 30.4 (C-11), 40.6 (C-12), 91.5 (C-13), 47.8 (C-14), 104.0 (C-15), 75.6 (C-16), 20.1 (C-17), 27.1 (C-18), 22.5 (C-19), 13.6 (C-20), 170.2 (MeCO), 21.2 (MeCO), 63.4 (MeCH
2o), 15.7 (MeCH
2o).According to above data, deterministic compound VII is (+)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15R)-3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.
Reference examples 7 (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15S)-3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; Extraction, separation and purification and the Structural Identification of 15,16-bis-epoxy Ladanum alkane-6-ketone (formula VIII compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 70～75% elutriants (H-3-4) that collection contains target compound, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, thin-layer chromatography is followed the trail of, collection contains target compound cut, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 82%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=58min, purifying obtains 22mg.
(4) Structural Identification of composition: colorless oil; Molecular formula: C
24h
38o
7, M=438, HRESIMS m/z 461.2508[M+Na]
+;
1h NMR (600MHz, CD
3cOCD
3) δ: 0.90 (3H, s, H
3-17), 0.94 (3H, s, H
3-18), 1.34 (3H, s, H
3-19), 1.19 (3H, d, J=6.4Hz, H
3-20), 5.19 (1H, br d, J=5.2Hz, H-15), 4.47 (1H, br s, H-3), 3.27 (1H, s, H-5), 4.08 (1H, d, J=8.8Hz, H-16a), 3.90 (1H, d, J=8.8Hz, H-16b), 3.40,3.46 (2H, 2m, MeCH
2o), 1.13 (3H, t, J=6.8Hz, MeCH
2o);
13c NMR (CD
3cOCD
3, 100MHz) δ: 26.2 (C-1), 22.9 (C-2), 78.9 (C-3), 36.5 (C-4), 52.5 (C-5), 212.6 (C-6), 78.4 (C-7), 47.4 (C-8), 93.1 (C-9), 48.7 (C-10), 29.6 (C-11), 39.3 (C-12), 92.0 (C-13), 48.7 (C-14), 104.7 (C-15), 78.2 (C-16), 20.3 (C-17), 27.1 (C-18), 22.5 (C-19), 13.8 (C-20), 170.2 (MeCO), 63.1 (MeCH
2o) 15.7 (MeCH
2o), 21.2 (MeCO).According to above data, deterministic compound VIII is (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15S)-3-acetoxyl group-7-hydroxyl-15-oxyethyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone.
Reference examples 8 (+)-(5S, 7R, 8R, 9R, 10S, 13S, 15R)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6, extraction, separation and purification and the Structural Identification of 16-diketone (formula IX compound)
(1) experiment material: with embodiment 1
(2) extraction of medicinal material: take Motherwort Herb medicinal material 20Kg, pulverize, add 95% ethanol (3 * 160L) and extract 72 hours, extract 3 times, extracting solution after decompression and solvent recovery, obtains ethanol extract 1.2Kg after merging;
(3) separation and purification of composition:
1. take Motherwort Herb medicinal material 20kg, pulverize, add 95%v/v ethanol 160L and extract 3 times, after extracting solution decompression and solvent recovery, obtain ethanol extract 1.2kg;
2. by ethanol extract water-dispersion, use successively ethyl acetate, n-butanol extraction, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract 400g;
3. adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out part (H) 8.2g, upper anti-phase polystyrene type resin post MCI gel CHP 20P, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part (H-3) 2.1g, again by anti-phase RP-C18 medium pressure liquid chromatography, 50-95%v/v methanol aqueous solution gradient elution, thin layer is followed the trail of, 65～70% elutriants (H-3-3) that collection contains target compound, reclaim solvent, residuum is again through Preparative TLC chromatographic separation, Shi You Mi ﹕ acetone=5 ﹕ 1 launches, scraping target product chromatographic band, with acetone 30ml wash-out, collect elutriant, reclaim solvent, residuum is separated through anti-phase liquid phantom preparing chromatogram, 78%v/v methanol aqueous solution wash-out, collection retention time is t
rthe target product of=40min, purifying obtains 2.4mg.
(4) Structural Identification of composition: colorless oil, HRESIMS m/z403.2094[M+Na]
+prompting molecular formula is C
21h
32o
6.The NMR data of comparative compound IX and IV, find that their difference is mainly: in compound IX, contain interior ester carbonyl group, and the methoxyl group (δ of 15 in compound IX
h3.54ppm, δ
c57.6ppm) substituted the oxyethyl group in IV.This inference further confirms by 2D NMR.In HMBC spectrum, H
2-12 and C-9, C-11, C-14 is relevant with C-16, H
2-14 and C-12, C-13, C-15 is relevant with C-16, H-15 and C-14, C-16 is relevant with C-21, illustrates that ester carbonyl group is positioned at C-16 position.By NOESY, compose, the features such as coupling constant and specific optical rotation can be judged C-5 in compound F 17-hydroxy-corticosterone, C-7, and C-8, C-9, C-10, the absolute configuration of C-13 is identical with C.In addition J in compound IX,
14,15be 6.0 and 4.8Hz, with compound ii, III, IV, the VII close (J=6.0,4.2Hz) with 15 α-configuration, and be greater than compound V, VI, the VIII (J=5.4,1.2Hz) of 15 beta configurations.Therefore, the configuration of C-15 and compound ii in compound IX, III, IV, VII are identical.The structure of compound IX is defined as (+)-(5S, 7R, 8R, 9R, 10S, 13S, 15R)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6,16-diketone.
(5) proton nmr spectra (
1h-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 1.
(6) carbon-13 nmr spectra (
13c-NMR): Bruker-AV-600 spectrometer measures, and data are in Table 2.
By test example, illustrate beneficial effect of the present invention below.
Test example 1 has the platelet aggregation-against test of two volution labdane type diterpenoids (compound shown in formula II crystal formation and III) of 13R configuration
(1) experiment material:
1. medicine
Test-compound formula II crystal formation and formula III compound are configured to the stock solution of 10mM with DMSO ,-10 ℃ of preservations, become 0.1mM with normal saline dilution before use.Positive drug (clopidogrel bisulfate tablet), collocation method is the same.
2. animal
SD rat, clean level, male and female are regardless of, and body weight 200～240g is provided by Chengdu University of Traditional Chinese Medicine's Animal Experimental Study center.Quality of production conformity certification number: SCXK (river) 2008-11.
3. reagent
Adenosine diphosphate (ADP) disodium (adenosine-5 '-diphosphate, ADP), U.S. sigma company produces, domestic packing, 1g/ bottle.
Dimethyl sulfoxide (DMSO) (DMSO), analytical pure, 500ml/ bottle, is purchased from Chengdu Ke Long chemical reagent factory.
4. laboratory apparatus
Hyperchannel platelet aggregation instrument (APACT/2 type, German blue ripple)
Electronic balance (ESJ120-4 type, Longteng Electronic Weighing Instrument Co., Ltd., Shenyang);
Water-bath, cryogenic refrigerator etc.
(2) experimental technique:
Select adult, healthy SD rat, male and female are regardless of, after conforming, femoral artery blood sampling, with the ratio anti-freezing of 3.8% Trisodium Citrate BP and blood 1:9, then with the centrifugal 10min of 800r/min, prepare platelet rich plasma (platelet-rich plasma, PRP), remainder is prepared platelet poor plasma (platelet-poor plasma, PPP) with the centrifugal 10min of 3000r/min.With PPP zeroing, get 200 μ l PRP and add 37 ℃ of incubation 5min in opacity tube, add the tested drug solution of 20 μ l, make final concentration be 0.1mM, blank group adds 1%DMSO solution.Mix rear continuation incubation 5min, then add ADP 20 μ l (final concentration is 0.05mg/ml) induction to assemble.With platelet aggregation instrument, detect thrombocyte MA, by following formula, calculate L-Arginine (AIR):
AIR=[(blank group MA-administration group MA)/blank group MA] * 100%
3. experimental result and evaluation:
The results are shown in Table 4.
Table 4 formula II crystal formation and formula III compound on the impact of extracorporeal platelet aggregation (
)
Note: with the comparison of physiological saline group,
*p<0.05.
By research, find: when final concentration is 0.1mM, formula II crystal formation and formula III compound have obvious restraining effect (P<0.05) to the platelet aggregation of external ADP induction, inhibiting rate is respectively 19.59% and 15.27%, with positive controls inhibiting rate 16.91% quite, even higher.
Test example 2 has the platelet aggregation-against test of two volution labdane type diterpenoids (compound shown in formula V, VI, IX) of 13S configuration
(1) experiment material:
1. medicine
Test-compound (-)-(5S, 7R, 8R, 9R, 10S, 13S, 15S)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone (V), (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15S)-3-acetoxyl group-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone (VI), (+)-(5S, 7R, 8R, 9R, 10S, 13S, 15R)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6,16-diketone (IX), is configured to the stock solution of 10mM with DMSO, and-10 ℃ of preservations, become 0.1mM with normal saline dilution before use.Positive drug (clopidogrel bisulfate tablet), collocation method is the same.
2. animal
SD rat, clean level, male and female are regardless of, and body weight 200～240g is provided by Chengdu University of Traditional Chinese Medicine's Animal Experimental Study center.Quality of production conformity certification number: SCXK (river) 2008-11.
3. reagent
Adenosine diphosphate (ADP) disodium (adenosine-5 '-diphosphate, ADP), U.S. sigma company produces, domestic packing, 1g/ bottle.
Dimethyl sulfoxide (DMSO) (DMSO), analytical pure, 500ml/ bottle, is purchased from Chengdu Ke Long chemical reagent factory.
4. laboratory apparatus
Hyperchannel platelet aggregation instrument (APACT/2 type, German blue ripple)
Electronic balance (ESJ120-4 type, Longteng Electronic Weighing Instrument Co., Ltd., Shenyang);
Water-bath, cryogenic refrigerator etc.
(2) experimental technique:
Select adult, healthy SD rat, male and female are regardless of, after conforming, femoral artery blood sampling, with the ratio anti-freezing of 3.8% Trisodium Citrate BP and blood 1:9, then with the centrifugal 10min of 800r/min, prepare platelet rich plasma (platelet-rich plasma, PRP), remainder is prepared platelet poor plasma (platelet-poor plasma, PPP) with the centrifugal 10min of 3000r/min.With PPP zeroing, get 200 μ l PRP and add 37 ℃ of incubation 5min in opacity tube, add the tested drug solution of 20 μ l, make final concentration be 0.1mM, blank group adds 1%DMSO solution.Mix rear continuation incubation 5min, then add ADP 20 μ l (final concentration is 0.05mg/ml) induction to assemble.With platelet aggregation instrument, detect thrombocyte MA, by following formula, calculate L-Arginine (AIR):
AIR=[(blank group MA-administration group MA)/blank group MA] * 100%
3. experimental result and evaluation:
The results are shown in Table 5.
Table 5 compound V, VI and IX on the impact of extracorporeal platelet aggregation (
)
By research, find: when final concentration is 0.1mM, test-compound compound (-)-(5S, 7R, 8R, 9R, 10S, 13S, 15S)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone (V), (-)-(3R, 5S, 7R, 8R, 9R, 10S, 13S, 15S)-3-acetoxyl group-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6-ketone (VI), (+)-(5S, 7R, 8R, 9R, 10S, 13S, 15R)-7-hydroxyl-15-methoxyl group-9,13; 15,16-bis-epoxy Ladanum alkane-6,16-diketone (IX) has certain restraining effect to the platelet aggregation of external ADP induction, and wherein the effect trend of VI is obvious.
From above test example, in Motherwort Herb, the separated crystal formation obtaining has certain restraining effect to platelet aggregation, wherein best with formula II crystal formation compound effect, under same concentration (0.1mM), than medicament for resisting platelet aggregation " bisulfate clopidogrel " better effects if, for the exploitation of the natural medicament for resisting platelet aggregation of novelty provides new selection.
Claims (10)
1. the crystal formation of compound shown in formula II, is characterized in that:
This crystal formation is rhombic system, and spacer is P2
12
12
1, unit cell parameters is
α=90.00 °,
β=90.00 °,
γ=90.00 °, Z=4, unit cell volume is
2. crystal formation according to claim 1, is characterized in that: the specific rotatory power of described crystal formation Chinese style I compound is [α]
20 d-58.8.
3. the preparation method of crystalline form I described in claim 1 or 2, is characterized in that: it comprises following operation steps:
(1) take Motherwort Herb medicinal material, pulverize, add 95%v/v extraction using alcohol, after extracting solution decompression and solvent recovery, obtain ethanol extract;
(2) by ethanol extract water-dispersion, be extracted with ethyl acetate successively, combined ethyl acetate part, decompression and solvent recovery, obtains ethyl acetate extract;
(3) adopt silica gel column chromatography to carry out separation to ethyl acetate extract, sherwood oil: the gradient elution of acetone=(100:1)～(0:1), get sherwood oil-acetone=5:1 wash-out and partly go up anti-phase polystyrene type resin post, 50-100%v/v methanol aqueous solution gradient elution, collect 70% methanol-eluted fractions part, pass through again medium pressure liquid chromatography, octadecyl silane chromatographic column, 50-95%v/v methanol aqueous solution gradient elution, collect 70～75% methanol aqueous solution wash-out parts, reclaim solvent, residuum is separated through Sephadex LH-20 dextran gel column chromatography again, Shi You Mi ﹕ tri-Lv Jia Wan ﹕ methyl alcohol=5 ﹕ 5 ﹕ 1 wash-outs, collect elutriant, reclaim solvent, anti-phase liquid phantom preparing chromatogram on residuum, 82%v/v methanol aqueous solution wash-out, collect respectively the elutriant of different appearance times, can obtain formula II compound,
(4) modus ponens II compound, adds acetone, is placed on subsequently in the moisture eliminator that siccative is housed, and is placed to crystal and separates out, and obtains crystal formation.
4. preparation method according to claim 3, is characterized in that: the mass volume ratio of formula II compound and acetone is: 10mg:0.1-1ml.
5. preparation method according to claim 4, is characterized in that: the mass volume ratio of formula II compound and acetone is: 10mg:0.3-0.5ml.
6. preparation method according to claim 5, is characterized in that: the mass volume ratio of formula II compound and acetone is: 10mg:0.4ml.
7. preparation method according to claim 3, is characterized in that: described siccative is selected from discolour silica gel.
8. the purposes of crystalline form I in preparing platelet aggregation-against class medicine described in claim 1 or 2.
9. a pharmaceutical composition, is characterized in that: it is the preparation of crystal formation described in the claim 1 or 2 that contains effective dose.
10. pharmaceutical composition according to claim 9, is characterized in that: described preparation is selected from through gastrointestinal absorption preparation or injection formulations.
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| CN114106008A (en) * | 2021-11-26 | 2022-03-01 | 成都普思生物科技股份有限公司 | Diterpene quinone compound extracted and separated from salvia miltiorrhiza bunge and method and application thereof |
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| CN103553919A (en) * | 2013-11-05 | 2014-02-05 | 成都中医药大学 | Terpenoid and preparation method and application thereof |
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| CN104829675A (en) * | 2015-05-08 | 2015-08-12 | 成都中医药大学 | Chemical compound, crystal form, and preparation methods and applications of chemical compound and crystal form |
| CN104829675B (en) * | 2015-05-08 | 2016-09-21 | 成都中医药大学 | A kind of compound and crystal formation and preparation method thereof and purposes |
| CN114106008A (en) * | 2021-11-26 | 2022-03-01 | 成都普思生物科技股份有限公司 | Diterpene quinone compound extracted and separated from salvia miltiorrhiza bunge and method and application thereof |
| CN114106008B (en) * | 2021-11-26 | 2022-11-08 | 成都普思生物科技股份有限公司 | Diterpene quinone compound extracted and separated from salvia miltiorrhiza bunge and method and application thereof |
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Effective date of registration: 20190527 Address after: 611137 No. 1166 Liutai Avenue, Wenjiang District, Chengdu City, Sichuan Province Co-patentee after: Chengdu first pharmaceutical Co., Ltd. Patentee after: Chengdu University of Traditional Chinese Medicine Address before: 611137 No. 1166 Liutai Avenue, Wenjiang District, Chengdu City, Sichuan Province Co-patentee before: Chengdu No.1 Pharmaceutical Co., Ltd. Patentee before: Chengdu University of Traditional Chinese Medicine |
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| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20190911 Address after: No. 133, No. two, three ring road, Tianpeng Town, Pengzhou City, Pengzhou, Sichuan Patentee after: Chengdu first pharmaceutical Co., Ltd. Address before: 611137 No. 1166, Willow Road, Wenjiang District, Sichuan, Chengdu Co-patentee before: Chengdu first pharmaceutical Co., Ltd. Patentee before: Chengdu University of Traditional Chinese Medicine |