CN116063315B - Vibsane diterpenoid compound or salt thereof with eight-membered ring skeleton in coral tree, and preparation method and application thereof - Google Patents
Vibsane diterpenoid compound or salt thereof with eight-membered ring skeleton in coral tree, and preparation method and application thereof Download PDFInfo
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- -1 diterpenoid compound Chemical class 0.000 title claims abstract description 19
- 240000008135 Piscidia piscipula Species 0.000 title claims abstract description 15
- 235000016678 Erythrina glauca Nutrition 0.000 title claims abstract description 14
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 150000003839 salts Chemical class 0.000 title claims abstract description 10
- 150000001875 compounds Chemical class 0.000 claims abstract description 33
- 238000004440 column chromatography Methods 0.000 claims abstract description 15
- 239000003814 drug Substances 0.000 claims abstract description 8
- 235000019013 Viburnum opulus Nutrition 0.000 claims abstract description 7
- 101100313763 Arabidopsis thaliana TIM22-2 gene Proteins 0.000 claims abstract description 5
- 244000307537 Viburnum odoratissimum Species 0.000 claims abstract description 5
- 235000013249 Viburnum odoratissimum Nutrition 0.000 claims abstract description 5
- 238000010898 silica gel chromatography Methods 0.000 claims abstract description 4
- 238000004519 manufacturing process Methods 0.000 claims abstract description 3
- 230000003215 anti-neuroinflammatory effect Effects 0.000 claims abstract 2
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 36
- 238000004262 preparative liquid chromatography Methods 0.000 claims description 17
- 208000036110 Neuroinflammatory disease Diseases 0.000 claims description 13
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 12
- PBCJIPOGFJYBJE-UHFFFAOYSA-N acetonitrile;hydrate Chemical compound O.CC#N PBCJIPOGFJYBJE-UHFFFAOYSA-N 0.000 claims description 12
- 229940125904 compound 1 Drugs 0.000 claims description 11
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 10
- 229940125782 compound 2 Drugs 0.000 claims description 10
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 claims description 8
- 230000003959 neuroinflammation Effects 0.000 claims description 8
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- 239000000284 extract Substances 0.000 claims description 7
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- IIYFAKIEWZDVMP-UHFFFAOYSA-N tridecane Chemical compound CCCCCCCCCCCCC IIYFAKIEWZDVMP-UHFFFAOYSA-N 0.000 abstract 2
- 238000004128 high performance liquid chromatography Methods 0.000 abstract 1
- 238000000926 separation method Methods 0.000 abstract 1
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 12
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- IAZDPXIOMUYVGZ-WFGJKAKNSA-N Dimethyl sulfoxide Chemical compound [2H]C([2H])([2H])S(=O)C([2H])([2H])[2H] IAZDPXIOMUYVGZ-WFGJKAKNSA-N 0.000 description 4
- 238000004364 calculation method Methods 0.000 description 4
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- 125000004029 hydroxymethyl group Chemical group [H]OC([H])([H])* 0.000 description 3
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 3
- AICOOMRHRUFYCM-ZRRPKQBOSA-N oxazine, 1 Chemical compound C([C@@H]1[C@H](C(C[C@]2(C)[C@@H]([C@H](C)N(C)C)[C@H](O)C[C@]21C)=O)CC1=CC2)C[C@H]1[C@@]1(C)[C@H]2N=C(C(C)C)OC1 AICOOMRHRUFYCM-ZRRPKQBOSA-N 0.000 description 3
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- 229930004069 diterpene Natural products 0.000 description 2
- 150000004141 diterpene derivatives Chemical class 0.000 description 2
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- SNVLJLYUUXKWOJ-UHFFFAOYSA-N methylidenecarbene Chemical group C=[C] SNVLJLYUUXKWOJ-UHFFFAOYSA-N 0.000 description 2
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- IIYFAKIEWZDVMP-NJFSPNSNSA-N tridecane Chemical group CCCCCCCCCCCC[14CH3] IIYFAKIEWZDVMP-NJFSPNSNSA-N 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- XILIYVSXLSWUAI-UHFFFAOYSA-N 2-(diethylamino)ethyl n'-phenylcarbamimidothioate;dihydrobromide Chemical compound Br.Br.CCN(CC)CCSC(N)=NC1=CC=CC=C1 XILIYVSXLSWUAI-UHFFFAOYSA-N 0.000 description 1
- 238000005084 2D-nuclear magnetic resonance Methods 0.000 description 1
- 241000208834 Adoxaceae Species 0.000 description 1
- 235000007756 Akebia quinata Nutrition 0.000 description 1
- 240000008027 Akebia quinata Species 0.000 description 1
- 208000024827 Alzheimer disease Diseases 0.000 description 1
- 208000006820 Arthralgia Diseases 0.000 description 1
- 208000010392 Bone Fractures Diseases 0.000 description 1
- 235000014653 Carica parviflora Nutrition 0.000 description 1
- 241000243321 Cnidaria Species 0.000 description 1
- 241001450685 Corallium japonicum Species 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
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- 241000736026 Sarcandra Species 0.000 description 1
- 241000780602 Senecio Species 0.000 description 1
- 206010042674 Swelling Diseases 0.000 description 1
- SAHIZENKTPRYSN-UHFFFAOYSA-N [2-[3-(phenoxymethyl)phenoxy]-6-(trifluoromethyl)pyridin-4-yl]methanamine Chemical compound O(C1=CC=CC=C1)CC=1C=C(OC2=NC(=CC(=C2)CN)C(F)(F)F)C=CC=1 SAHIZENKTPRYSN-UHFFFAOYSA-N 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical group [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
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- UREBDLICKHMUKA-CXSFZGCWSA-N dexamethasone Chemical compound C1CC2=CC(=O)C=C[C@]2(C)[C@]2(F)[C@@H]1[C@@H]1C[C@@H](C)[C@@](C(=O)CO)(O)[C@@]1(C)C[C@@H]2O UREBDLICKHMUKA-CXSFZGCWSA-N 0.000 description 1
- 229960003957 dexamethasone Drugs 0.000 description 1
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- 125000000567 diterpene group Chemical group 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000990 heteronuclear single quantum coherence spectrum Methods 0.000 description 1
- 238000004896 high resolution mass spectrometry Methods 0.000 description 1
- 210000002865 immune cell Anatomy 0.000 description 1
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- 201000006417 multiple sclerosis Diseases 0.000 description 1
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- 231100000189 neurotoxic Toxicity 0.000 description 1
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- UMRZSTCPUPJPOJ-KNVOCYPGSA-N norbornane Chemical compound C1C[C@H]2CC[C@@H]1C2 UMRZSTCPUPJPOJ-KNVOCYPGSA-N 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 125000004430 oxygen atom Chemical group O* 0.000 description 1
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D493/00—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system
- C07D493/12—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system in which the condensed system contains three hetero rings
- C07D493/18—Bridged systems
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D301/00—Preparation of oxiranes
- C07D301/32—Separation; Purification
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D303/00—Compounds containing three-membered rings having one oxygen atom as the only ring hetero atom
- C07D303/02—Compounds containing oxirane rings
- C07D303/12—Compounds containing oxirane rings with hydrocarbon radicals, substituted by singly or doubly bound oxygen atoms
- C07D303/16—Compounds containing oxirane rings with hydrocarbon radicals, substituted by singly or doubly bound oxygen atoms by esterified hydroxyl radicals
- C07D303/17—Compounds containing oxirane rings with hydrocarbon radicals, substituted by singly or doubly bound oxygen atoms by esterified hydroxyl radicals containing oxirane rings condensed with carbocyclic ring systems having three or more relevant rings
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D493/00—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system
- C07D493/02—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system in which the condensed system contains two hetero rings
- C07D493/08—Bridged systems
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
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Abstract
Vibsane diterpenoid compounds or salts thereof with eight-membered ring skeletons in coral trees, and a preparation method and application thereof belong to the technical field of medicines, and relate to 4 vibsane diterpenoid compounds vibsanoids extracted and separated from dried leaves of a plant coral tree (Viburnum odoratissimum Ker-gawl. Var. Odoratisimum) of the genus Viburnum of the family of the five-Fucaceae, and the diterpenoid compounds vibsanoids have a mother nucleus structure of tricyclo [8.2.1.0 2,9 ] tridecane and are obtained through separation by silica gel column chromatography, HP20 macroporous resin column chromatography, ODS column chromatography and HPLC. The anti-neuroinflammatory activity of the compounds is examined by testing the inhibition effect of the compounds on NO production by LPS-induced mouse microglial cells BV-2, and the results show that the compounds A-D have different degrees of inhibition activity.
Description
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to vibsane-membered ring skeleton diterpenoid compounds vibsanoids in coral trees and a preparation method thereof, and application of the compounds in preparation of anti-neuroinflammation medicines.
Background
Coral tree (Viburnum odoratissimum Ker-gawl.var.odonatissisum): is a plant of the genus Viburnum of the family Pentaforaceae (Adoxaceae). The original products of China are mainly distributed in Fujian, hunan, guangdong, hainan and other places. The root and leaf of the Chinese medicinal herb can be used as a medicament, has the effects of clearing and activating the channels and collaterals, is mainly used for treating rheumatic arthralgia, traumatic injury, swelling and pain and fracture, and is widely used for diminishing inflammation, relieving pain and resisting rheumatism.
Neuroinflammation (Neuroinflammation): inflammation of the central nervous system, known as neuroinflammation, is a process that in a specific case is accomplished by various glia and surrounding immune cells in the central nervous system together in order to protect the brain from damage. However, if glial cells are frequently activated to overproduce immune factors or peripheral invading lymphocytes release neurotoxic factors, neuroinflammation can induce or exacerbate degenerative changes of the nervous system, such as parkinson's disease, alzheimer's disease, multiple sclerosis, and the like. The type, duration, surrounding environment, etc. of the inflammatory response seriously jeopardize the survival of neurons. Therefore, it is important to find natural medicines for treating neuroinflammation.
Disclosure of Invention
The invention aims to provide 4 vibsane diterpenoid compounds or salts thereof separated from dry leaves of a Viburnum sarcandra (Viburnum odoratissimum Ker-Gawl. Var. Odoratisimum) plant of the genus Viburnum of the family Pacifiaceae, a preparation method thereof and application of the compounds or salts thereof in preparation of anti-neuroinflammation medicines. The vibsane diterpenoid compound has a structure:
the preparation method for vibsane-membered diterpenoid compounds with eight-membered ring frameworks by using coral leaves comprises the following steps:
(1) Extracting dried leaves of coral tree with ethanol, concentrating the extractive solution to obtain extract, extracting the extract with ethyl acetate, eluting the ethyl acetate layer extract with CH 2Cl2/CH3 OH gradient (100:1-1:1) (v/v) by silica gel column chromatography to obtain Fr.1-Fr.44 fractions;
(2) Subjecting the fraction Fr.1 to HP20 column chromatography and gradient elution with ethanol water to obtain Fr.1A and Fr.1B2 fractions;
(3) The fraction Fr.1B is further eluted by ODS open column chromatography with a gradient of ethanol-water (60:40-100:0) (v/v) to obtain Fr.1B1-Fr.1B33 fractions;
(4) Fraction fr.1b3 was purified by column chromatography on silica gel with petroleum ether: ethyl acetate (50:1-1:1) (v/v) to give 7 components fr.1b31-fr.1b37;
(5) Subjecting the component Fr.1B36 to preparative liquid chromatography and eluting with methanol-water to obtain 5 subfractions Fr.1B361-Fr.1B365; purifying subfraction Fr.1B362 with acetonitrile-water by semi-preparative liquid chromatography to obtain compound 1;
(6) Fraction fr.1b2 was purified by column chromatography on silica gel with petroleum ether: ethyl acetate (50:1-1:1) (v/v) to give 6 fractions fr.1b21-fr.1b26;
(7) Subjecting the component Fr.1B24 to preparative liquid chromatography, eluting with methanol-water to obtain 4 subfractions Fr.1B241-Fr.1B244; purifying subfraction Fr.1B244 with acetonitrile-water by semi-preparative liquid chromatography to obtain compound 2; sub-fractions fr.1b241 and fr.1b242 were purified by semi-preparative liquid chromatography with acetonitrile-water to give compounds 3 and 4.
The preparation method comprises the following steps:
In the step (1), the coral tree is a coral tree (Viburnum odoratissimum Ker-Gawl. Var. Odoratisimum) of the genus Viburnum of the family Pentaforaceae. Reflux-extracting dried leaves of coral tree with 70-80% industrial ethanol.
In the step (2), the fraction Fr.1 is subjected to HP20 column chromatography and is subjected to gradient elution in a proportion of 30%,60% and 90% of ethanol water.
In the step (5), the component Fr.1B36 is eluted by methanol-water (83:17, v/v) through a preparative liquid phase chromatography to obtain 5 subfractions Fr.1B361-Fr.1B365; subfraction Fr.1B362 was purified by semi-preparative liquid chromatography using acetonitrile-water (70:30, v/v) to give compound 1.
In the step (7), the component Fr.1B24 is eluted by methanol-water (80:20, v/v) through a preparative liquid phase chromatograph to obtain 4 subfractions Fr.1B241-Fr.1B244; sub-fraction Fr.1B244 was purified by semi-preparative liquid chromatography using acetonitrile-water (65:35, v/v) to give compound 2; subfractions Fr.1B241 and Fr.1B242 were purified by semi-preparative liquid chromatography using acetonitrile-water (60:40, v/v) to give compounds 3 and 4.
The results of the system structure identification of the obtained compound are as follows:
The structure of the compounds 1-4 was identified by high resolution mass spectrometry, one-dimensional NMR, two-dimensional NMR, X-single crystal diffraction, calculated nuclear magnetism and ECD, and the related spectrograms were shown in figures 1-13.
Vibsanoid A (1) white crystals,UV (MeOH) lambda max (log epsilon): 220nm (1.32); HRESIMS gave an excimer ion peak m/z 379.2112[ M+H ] +(calcd for C21H31O6, 379.2115, which was determined to have a molecular formula of C 21H30O6 in combination with 1H,13 C-NMR data, and calculated to have an unsaturation of 7. In 1H-NMR(600MHz,DMSO-d6) spectra, δ H 5.71.71 (1 h, d, j=1.4 Hz), 2.12 (3 h, d, j=1.4 Hz) and 1.89 (3 h, d, j=1.4 Hz) are characteristic groundsel acyloxy hydrogen signals, δ H 5.64.64 (1 h, s) and 4.05 (1 h, dd, j=5.3, 3.3 Hz) are two active hydrogen signals, δ H 3.85.85 (1 h, dd, j=11.5, 5.3 Hz) and 3.17 (1 h, overlapped) are a set of oxymethylene hydrogen signals, and δ H 1.13.13 (3 h, s) and 0.89 (3 h, s) are two methyl hydrogen signals. In combination with the DEPT and HSQC spectra, 13C-NMR(150MHz,DMSO-d6) spectra showed a total of 21 carbons, including 4 methyl carbon signals, 5 methylene carbon signals, 6 methylene carbon signals and 6 quaternary carbon signals. The data indicate that the compound belongs to vibsane-type diterpenoid compounds losing 4 carbons. The structure of the compound was further confirmed by two-dimensional nuclear magnetism.
1H-1 In the H COSY spectrum, the presence of the two coupling fragments I and II is indicated by the correlation of H-6/H 2 -5 and H-8/H-9/H-10. In the HMBC spectra, H 3 -19 was observed to correlate with the presence of C-6, C-8, and H-5 with the presence of C-7, indicating that fragments I and II are linked by C-7; one significant difference compared to undecylenic ring vibsane diterpenoids is that H-10 is associated with the presence of HMBC for C-18; furthermore, in HMBC spectra, it is observed that OH-4 correlates with the presence of C-3, C-5, H-6/H 2 -18 correlates with the presence of C-4, the above data indicate the presence of the A ring structure fragment and the methyl group attached to C-7. In addition, the presence of H 3 -20 and C-10, C-11, C-1 in the HMBC spectra, the presence of H-2 and C-10, and the presence of H 2 -18 and C-2 indicate that the B ring is combined with the A ring through C-10 and C-3. Another difference compared to the undecylenic ring vibsane diterpenes reported in the literature is the correlation of H 2 -1 with C-12, C-13 and the presence of H-2/H 2-13/H2 -12 in 1H-1 H COSY observed in the HMBC spectra, indicating that the B and C rings form a bicyclo [2.2.1] heptane structural fragment. Considering the large chemical shift values of C-4 and C-8 (delta C 95.8.8 and delta C 70.6, respectively), the combination of H-8 and C-4 is related to the presence of HMBC, and therefore, it is presumed that an oxygen bridge exists between C-4 and C-8. Finally, given the chemical shift values of C-6 and C-7 (delta C 58.7.7 and delta C 54.8.8, respectively) and the remaining one unsaturation, a three-membered oxygen ring fragment is presumed to be present. H-9 is associated with the presence of HMBC for C-1', indicating that the senecio acyloxy group is attached to C-9. In summary, the planar structure of compound 1 was determined as a four-carbon vibsane type diterpene with a tricyclo [8.2.1.0 2,9 ] tridecane skeleton and named vibsanoid A.
The relative configuration of compound 1 was determined by NOESY-related and biosynthetic pathways. According to the biosynthetic pathway, the H-8 and C-11-C-12 bonds are oriented alpha, while CH 3 -7 is oriented beta. The relative configuration of the remaining chiral centers was then determined by NOESY spectroscopy. In NOESY spectra, a correlation of H-6/H 3-19,H-9/H3 -20 and H-9/H-1b was observed, indicating that H-6,H-9 and CH 3 -11 are beta oriented; the correlation of H-10/H-18a, H-18b/13a and 18-OH/H-13a indicates that these hydrogens are in the alpha orientation. 4-OH and H-8 are located on the same side of the plane, alpha oriented, due to loop tension. The relative configuration of compound 1 was further confirmed by X-single crystal diffraction experiments. And the absolute configuration of the compound was determined by comparing the measured and calculated ECD curves to be 2r,3s,4r,6r,7r,8r,9s,10r,11s.
Vibsanoid B (2) colorless oil,UV (MeOH) lambda max (log epsilon): 220nm (0.62); HRESIMS gave an excimer ion peak m/z 371.2195[ M+Na ] +(calcd for C21H32NaO4, 371.2193, which was determined to have a molecular formula of C 21H32O4 in combination with 1H,13 C-NMR data and calculated to have an unsaturation of 6. Both were found to have similar backbones by comparison with NMR data for compound 1. The difference is the position of the groundsel acyloxy group, the C-4 and C-8 oxygen bridge and the deletion of 4-OH in compound 2. 1H-1 The presence of the H-8/H 2 -9/H-10 coupling fragment was observed in the H COSY spectrum, combined with the correlation of H 3 -19 with C-6, C-8 and the correlation of H-8 with C-1' in the HMBC spectrum, indicating that the groundsel acyloxy group was attached to C-8. Whereas changes in chemical shift values in 13 C-NMR (delta C 30.3.3 and 24.3 in compound 2 and delta C 95.8.8 and 70.6 in compound 1) indicate deletions of C-4 and C-8 oxygen bridges and 4-OH.
The relative configuration of compound 2 was determined by NOESY and calculated carbon chemical shift values. In the NOESY spectrum, the correlation of H-6/H 3 -19, H-18/H-8 indicated that H-6 was in the beta orientation and that the hydroxymethyl group of C-3 was in the alpha orientation. Because of the H-10 signal overlap, its orientation cannot be determined by NOESY correlation. To determine the relative configuration of C-10, four possible configurations were calculated using quantum chemistry and the reliability of the calculation was analyzed using dp4+, the calculation showed that 2a had a higher R 2 value and dp4+ probability analysis was 100%, thus determining the relative configuration of C-10 as S. Finally, the absolute configuration of compound 2 was determined by calculation of ECD to be 2R,3R,6R,7R,8R,10S,11S.
Vibsanoid C (3) colorless oil.UV (MeOH) lambda max (log epsilon): 220nm (1.05); HRESIMS gave an excimer ion peak m/z 385.1989[ M+Na ] +(calcd for C21H30NaO5, 385.1985, which was determined to have a molecular formula of C 21H30O5 in combination with 1H,13 C-NMR data, and calculated unsaturation to be 7. Analysis of the NMR data revealed that this compound 3 also had a tricyclo [8.2.1.0 2,9 ] tridecane skeleton structure identical to that of compound 1. The difference from compound 1 is the presence of a set of linking sites for carbon-carbon double bonds (δ C 135.7.7 and 134.2) and oxygen bridges. The presence of H 3 -19 and C-6, C-7, C-8 in the HMBC spectra correlated, the presence of H-6 and C-4, and the presence of H-5 and C-7 correlated, indicated that the double bond was located between C-4 and C-7. Considering the chemical shift values of C-4 and C-7 (δ C 113.5 and 88.9) and a non-attributed oxygen atom, it is speculated that an oxygen bridge exists between C-4 and C-7. The relative configuration of compound 3 was determined by NOESY spectroscopy. In the NOESY spectra, the correlation of H-10/H 3-20,H-10/H2-18,H-10/H-1b,H-18/H-1b,H-6/H3 -19 and H-5/H-18 indicates that the hydroxymethyl groups of H-10,4-OH and C-3 are beta oriented. The smaller coupling constant (j=5.8 Hz) indicates that the carbon-carbon double bonds at the C-5 and C-6 positions are Z-type. The relative configuration of compound 3 was thus determined to be 2R,3R,4S,7R,8R,10S,11S. The absolute configuration of compound 3 was determined by calculation of ECD to be 2R,3R,4S,7R,8R,10S,11S.
Vibsanoid D (4) colorless oil.UV (MeOH) lambda max (log epsilon): 220nm (1.11); HRESIMS gave an excimer ion peak m/z 385.1996[ M+Na ] +(calcd for C21H30NaO5, 385.1985, which was determined to have a molecular formula of C 21H30O5 in combination with 1H,13 C-NMR data, and calculated unsaturation to be 7. Comparison of NMR data reveals that compound 4 is an analogue of compound 3, the main difference being that the double bond in compound 3 is replaced by a carbonyl group and the deletion of 4-OH. This is demonstrated by the correlation of H 3 -19 with C-6, the correlation of H-4 with C-6, C-7 and the correlation of H 2 -18 with C-4 and the correlation of H-5/H-6 in 1H-1 HCOSY in the HMBC spectra. The relative configuration of compound 4 was determined by NOESY. In NOESY spectra, the correlation of H-8/H-10, H-8/H-18, H-18/H-12a shows that the hydroxymethyl groups of H-8,H-10 and C-3 are oriented towards the same side and alpha is oriented towards; the correlation of H-4/H-2, H-5/H-1b indicates that H-4 is beta oriented. Thus, the relative configuration of compound 4 is defined as 2R,3S,4S,7S,8R,10R,11S. Whereas the absolute configuration is determined by comparing the measured and calculated ECDs as 2R,3S,4S,7S,8R,10R,11S.
The nuclear magnetic data assignment of compounds 1-4 is shown in Table 1
TABLE 1 1 H (600 MHz) and 13 C (150 MHz) NMR data for Compounds 1-4 (a: DMSO-d 6 and b: CDCl 3)
In addition, the anti-neuroinflammation activity of 4 vibsane diterpenoid compounds 1-4 on the mouse microglial cells BV-2 is examined, and experimental results show that the compounds 1-4 have the anti-neuroinflammation activity and can inhibit the generation of NO to different degrees at 10 mu M, so that the compounds 1-4 have the prospect of further developing medicines for treating neuroinflammation.
The invention also provides a pharmaceutical composition comprising the vibsane diterpenoid compounds 1-4 or salts thereof and a pharmaceutically acceptable carrier or excipient.
The invention has the advantages that the compounds 1-4 are all optical pure compounds with definite space configuration, and meanwhile, the anti-neuroinflammation activity is stronger, thus having further development value.
Drawings
UV spectrum of compound 1 of fig. 1;
Hresis spectrum of compound 1 of fig. 2;
UV spectrum of compound 2 of fig. 3;
Hresis spectrum of compound 2 of fig. 4;
UV spectrum of compound 3 of fig. 5;
Hresis spectrum of compound 3 of fig. 6;
UV spectrum of compound 4 of fig. 7;
hresis spectrum of compound 4 of fig. 8;
FIG. 9 critical HMBC-related and COSY-related compounds 1-4;
FIG. 10 key NOESY correlation for Compounds 1-4;
FIG. 11 single crystal plot of Compound 1;
FIG. 12 is a graph of the linear correlation of compounds 2a-2d for the measured and calculated carbon chemical shifts;
comparison of measured and calculated ECDs for compounds 1-4 of FIG. 13;
FIG. 14 effect of Compounds 1-4 on LPS-induced NO production by BV-2 cells.
Detailed Description
The examples set forth below are presented to aid one skilled in the art in a better understanding of the present invention and are not intended to limit the invention in any way.
Example 1
The preparation method of the compounds 1-4 specifically comprises the following steps:
(1) Reflux-extracting dry leaves (40 kg) of stem of Corallium japonicum Kishinouye of Viburnum of Fiveleaf of Pentaforaceae with 70% industrial ethanol for 3 times, and extracting the obtained extract with ethyl acetate. Then, the crude extract (1300 g) of the ethyl acetate layer was subjected to silica gel column chromatography with a CH 2Cl2/CH3 OH system in a volume ratio (100:1, 50:1,30:1,20:1,10:1,5:1, 1:1) gradient elution to obtain four fractions Fr.1-Fr.4.
(2) Fraction Fr.1 (80 g) was subjected to HP20 macroporous resin column chromatography, and gradient elution was carried out at a ratio of 30%,60% and 90% with ethanol water to obtain two fractions Fr.1A-Fr.1B.
(3) The fraction Fr.1B is subjected to ODS open column chromatography, and three fractions Fr.1B1-Fr.1B3 are obtained by gradient elution of ethanol-water according to the volume ratio (60:40, 70:30,80:20,90:10, 100:0).
(4) The fr.1b3 fraction obtained was purified by column chromatography on silica gel with petroleum ether: the ethyl acetate system eluted in a volume ratio (50:1, 30:1,20:1,10:1,5:1,3:1, 1:1) to yield 7 components Fr.1B31-Fr.1B37.
(5) Fr.1B36 was prepared by preparative liquid chromatography on methanol-water (83:17, v/v,7 mL/min) to give 5 subfractions Fr.1B361-Fr.1B365. Subfraction Fr.1B362 was purified by semi-preparative liquid chromatography using acetonitrile-water (70:30, v/v,2.5 mL/min) to give compound 1 (10 mg).
(6) Fr.1b2 was purified by column chromatography on silica gel with petroleum ether: ethyl acetate was eluted gradient by volume (50:1, 30:1,20:1,10:1,5:1,3:1, 1:1) to give 6 fractions fr.1b21-fr.1b26.
(7) The subfractions Fr.1B24 were purified by preparative liquid chromatography on methanol-water (80:20, v/v,7 mL/min) to give 4 subfractions Fr.1B241-Fr.1B244.Fr.1B244 was purified by semi-preparative liquid chromatography using acetonitrile-water (65:35, v/v,2.5 mL/min) to give compound 2 (7 mg); subfractions Fr.1B241 and Fr.1B242 were purified by semi-preparative liquid chromatography using acetonitrile-water (60:40, v/v,2.5 mL/min) to give compounds 3 (19.7 mg) and 4 (18.3 mg).
Example 2
Compound 1-4 screening for neuroinflammatory activity of mouse microglial cells BV-2 in vitro.
BV2 microglial cells were cultured in DMEM medium containing 10% FBS at 37℃under 5% CO 2. BV2 cells in the logarithmic growth phase were seeded at a density of 6X 10 3 cells/well in 96-well plates and cultured for 12h. Cells were treated with compounds 1-4 at a concentration of 10 μm for 1h, while dexamethasone was selected as a positive control. After that, 10. Mu.g/mL of LPS was added to the mixture to effect the reaction for 24 hours. The supernatant was taken and assayed by the addition of Griess reagent using an ELISA reader at 540 nm. The results indicate that compounds 1-4 have varying degrees of neuroinflammation inhibition on BV-2 cells (see FIG. 14).
Claims (6)
1. A vibsane-type diterpenoid compound or a salt thereof with an eight-membered ring skeleton in coral tree, which is characterized in that the vibsane-type diterpenoid compound has the structural formula:
2. A process for producing a vibsane-type diterpenoid compound having an octa-ring skeleton or a salt thereof according to claim 1, characterized in that the process for producing a vibsane-type diterpenoid compound comprises the steps of:
(1) Extracting dried leaves of coral tree with ethanol, concentrating the extractive solution to obtain extract, extracting the extract with ethyl acetate, eluting the ethyl acetate layer extract with silica gel column chromatography with CH 2Cl2/CH3 OH gradient to obtain Fr.1-Fr.4 fractions;
(2) Subjecting fraction Fr.1 to HP20 column chromatography, and gradient eluting with 30%,60% and 90% ethanol water to obtain Fr.1A and Fr.1B2 fractions;
(3) The fraction Fr.1B is further eluted by ODS open column chromatography with ethanol-water gradient of 60:40-100:0v/v to obtain Fr.1B1-Fr.1B3 fractions;
(4) Fraction fr.1b3 was purified by column chromatography on silica gel with petroleum ether: ethyl acetate 50:1-1:1v/v to give 7 fractions fr.1b31-fr.1b37;
(5) The component Fr.1B36 is eluted by methanol-water 83:17v/v through preparative liquid chromatography to obtain 5 subfractions Fr.1B361-Fr.1B365; purifying subfraction Fr.1B362 by semi-preparative liquid chromatography with acetonitrile-water 70:30v/v to obtain compound 1;
(6) Fraction fr.1b2 was purified by column chromatography on silica gel with petroleum ether: ethyl acetate 50:1-1:1v/v to give 6 fractions fr.1b21-fr.1b26;
(7) The component Fr.1B24 is eluted by a preparative liquid chromatograph with methanol-water of 80:20v/v to obtain 4 subfractions Fr.1B241-Fr.1B244; purifying subfraction Fr.1B244 by semi-preparative liquid chromatography with acetonitrile-water 65:35v/v to obtain compound 2; sub-fractions Fr.1B241 and Fr.1B242 were purified by semi-preparative liquid chromatography using acetonitrile-water 60:40v/v to give compounds 3 and 4.
3. The method for producing a vibsane-type diterpenoid compound having an octa-cyclic skeleton according to claim 2, wherein in the step (1), the coral tree is a coral tree (Viburnum odoratissimum Ker-gawl.var. Odoratisimum) belonging to the genus viburnum of the family feverfew; reflux-extracting dried leaves of coral tree with 70-80% industrial ethanol.
4. A pharmaceutical composition comprising a vibsane-type diterpenoid compound having an eight-membered ring skeleton or a salt thereof according to claim 1 and a pharmaceutically acceptable carrier or excipient.
5. Use of a vibsane-type diterpenoid compound with an octa-ring skeleton or a salt thereof according to claim 1 for the preparation of a medicament for the treatment of neuroinflammation.
6. The use of a pharmaceutical composition according to claim 4 for the preparation of an anti-neuroinflammatory agent.
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| CN109336730A (en) * | 2018-09-28 | 2019-02-15 | 南京林业大学 | Bicyclic [ 3.3.1 ] nonyl vinyl compound of one kind and preparation method thereof |
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| CN109336730A (en) * | 2018-09-28 | 2019-02-15 | 南京林业大学 | Bicyclic [ 3.3.1 ] nonyl vinyl compound of one kind and preparation method thereof |
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