CN105968150B - The preparation method of 7-O- ethyl morronisides - Google Patents
The preparation method of 7-O- ethyl morronisides Download PDFInfo
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- CN105968150B CN105968150B CN201610309791.5A CN201610309791A CN105968150B CN 105968150 B CN105968150 B CN 105968150B CN 201610309791 A CN201610309791 A CN 201610309791A CN 105968150 B CN105968150 B CN 105968150B
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- 238000002360 preparation method Methods 0.000 title claims abstract description 26
- YTZSBJLNMIQROD-SFBCHFHNSA-N Morroniside Natural products O([C@@H]1OC=C([C@H]2C[C@H](O)O[C@@H](C)[C@H]21)C(=O)OC)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O YTZSBJLNMIQROD-SFBCHFHNSA-N 0.000 claims abstract description 78
- YTZSBJLNMIQROD-UHFFFAOYSA-N (4aS)-1c-beta-D-glucopyranosyloxy-6xi-hydroxy-8t-methyl-(4ar,8ac)-5,6,8,8a-tetrahydro-1H,4aH-pyrano[3,4-c]pyran-4-carboxylic acid methyl ester Natural products C12C(C)OC(O)CC2C(C(=O)OC)=COC1OC1OC(CO)C(O)C(O)C1O YTZSBJLNMIQROD-UHFFFAOYSA-N 0.000 claims abstract description 39
- 238000010438 heat treatment Methods 0.000 claims abstract description 14
- 230000003110 anti-inflammatory effect Effects 0.000 claims abstract description 13
- 239000002260 anti-inflammatory agent Substances 0.000 claims abstract description 3
- 229940124599 anti-inflammatory drug Drugs 0.000 claims abstract description 3
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 39
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 38
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 15
- ZUKLFFYDSALIQW-MSUKCBDUSA-N Iridoid glycoside Chemical compound [H][C@]12CC[C@H](C(O)=O)[C@@]1([H])[C@H](OC1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O)OC=C2 ZUKLFFYDSALIQW-MSUKCBDUSA-N 0.000 claims description 14
- 229930182489 iridoid glycoside Natural products 0.000 claims description 14
- AMBQHHVBBHTQBF-UHFFFAOYSA-N Loganin Natural products C12C(C)C(O)CC2C(C(=O)OC)=COC1OC1OC(CO)C(O)C(O)C1O AMBQHHVBBHTQBF-UHFFFAOYSA-N 0.000 claims description 9
- 238000002425 crystallisation Methods 0.000 claims description 9
- 230000008025 crystallization Effects 0.000 claims description 9
- AMBQHHVBBHTQBF-UOUCRYGSSA-N loganin Chemical compound O([C@@H]1OC=C([C@H]2C[C@H](O)[C@H](C)[C@H]21)C(=O)OC)[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O AMBQHHVBBHTQBF-UOUCRYGSSA-N 0.000 claims description 9
- 238000010992 reflux Methods 0.000 claims description 9
- 239000012141 concentrate Substances 0.000 claims description 7
- 238000000605 extraction Methods 0.000 claims description 7
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 6
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 6
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 6
- 235000019441 ethanol Nutrition 0.000 claims description 6
- 229930182470 glycoside Natural products 0.000 claims description 6
- 150000002338 glycosides Chemical class 0.000 claims description 6
- 238000004090 dissolution Methods 0.000 claims description 5
- 239000002994 raw material Substances 0.000 claims description 5
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 3
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 claims description 2
- 229910017604 nitric acid Inorganic materials 0.000 claims description 2
- 244000107975 Strychnos nux-vomica Species 0.000 claims 1
- 239000003795 chemical substances by application Substances 0.000 claims 1
- 230000003750 conditioning effect Effects 0.000 claims 1
- 238000006243 chemical reaction Methods 0.000 abstract description 16
- 238000002474 experimental method Methods 0.000 abstract description 15
- 238000000034 method Methods 0.000 abstract description 8
- 238000000746 purification Methods 0.000 abstract description 4
- 230000001741 anti-phlogistic effect Effects 0.000 abstract description 3
- -1 ethyl morroniside Chemical compound 0.000 abstract description 3
- 239000012535 impurity Substances 0.000 abstract description 3
- 238000000338 in vitro Methods 0.000 abstract description 3
- 238000001727 in vivo Methods 0.000 abstract description 3
- 238000012216 screening Methods 0.000 abstract description 3
- 238000013461 design Methods 0.000 abstract description 2
- 238000004519 manufacturing process Methods 0.000 abstract description 2
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 abstract 3
- 108090001005 Interleukin-6 Proteins 0.000 description 9
- 239000000243 solution Substances 0.000 description 9
- 238000003756 stirring Methods 0.000 description 9
- 102100040247 Tumor necrosis factor Human genes 0.000 description 7
- IRKFOLIBBQDADK-LEYKIWRXSA-N methyl (1S,3R,4aS,8S,8aS)-3-ethoxy-1-methyl-8-[(2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy-1,3,4,4a,8,8a-hexahydropyrano[3,4-c]pyran-5-carboxylate Chemical compound CCO[C@H]1C[C@H]2[C@@H]([C@H](C)O1)[C@H](O[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O)OC=C2C(=O)OC IRKFOLIBBQDADK-LEYKIWRXSA-N 0.000 description 7
- 210000002966 serum Anatomy 0.000 description 7
- 206010061218 Inflammation Diseases 0.000 description 6
- 241000699670 Mus sp. Species 0.000 description 6
- 150000001875 compounds Chemical class 0.000 description 6
- 239000003814 drug Substances 0.000 description 6
- 230000001939 inductive effect Effects 0.000 description 6
- 230000004054 inflammatory process Effects 0.000 description 6
- 238000011160 research Methods 0.000 description 6
- 239000013078 crystal Substances 0.000 description 5
- 238000010586 diagram Methods 0.000 description 5
- 239000000706 filtrate Substances 0.000 description 5
- 238000001914 filtration Methods 0.000 description 5
- 238000001819 mass spectrum Methods 0.000 description 5
- 238000001556 precipitation Methods 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- MZOFCQQQCNRIBI-VMXHOPILSA-N (3s)-4-[[(2s)-1-[[(2s)-1-[[(1s)-1-carboxy-2-hydroxyethyl]amino]-4-methyl-1-oxopentan-2-yl]amino]-5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-3-[[2-[[(2s)-2,6-diaminohexanoyl]amino]acetyl]amino]-4-oxobutanoic acid Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC(O)=O)NC(=O)CNC(=O)[C@@H](N)CCCCN MZOFCQQQCNRIBI-VMXHOPILSA-N 0.000 description 4
- 241000209020 Cornus Species 0.000 description 4
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 4
- 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 4
- 229960003957 dexamethasone Drugs 0.000 description 4
- 241000699666 Mus <mouse, genus> Species 0.000 description 3
- 239000000470 constituent Substances 0.000 description 3
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- 235000013399 edible fruits Nutrition 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 238000002347 injection Methods 0.000 description 3
- 239000012679 serum free medium Substances 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 210000003462 vein Anatomy 0.000 description 3
- 238000002965 ELISA Methods 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 230000015572 biosynthetic process Effects 0.000 description 2
- 230000000144 pharmacologic effect Effects 0.000 description 2
- 239000000376 reactant Substances 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 230000009897 systematic effect Effects 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- NHBKXEKEPDILRR-UHFFFAOYSA-N 2,3-bis(butanoylsulfanyl)propyl butanoate Chemical compound CCCC(=O)OCC(SC(=O)CCC)CSC(=O)CCC NHBKXEKEPDILRR-UHFFFAOYSA-N 0.000 description 1
- 201000006474 Brain Ischemia Diseases 0.000 description 1
- 206010008120 Cerebral ischaemia Diseases 0.000 description 1
- 241000142975 Cornaceae Species 0.000 description 1
- 241000581650 Ivesia Species 0.000 description 1
- 206010054949 Metaplasia Diseases 0.000 description 1
- 230000021736 acetylation Effects 0.000 description 1
- 238000006640 acetylation reaction Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000006708 antioxidants Nutrition 0.000 description 1
- 210000001367 artery Anatomy 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 206010008118 cerebral infarction Diseases 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229940126678 chinese medicines Drugs 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 208000012839 conversion disease Diseases 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 239000000428 dust Substances 0.000 description 1
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 1
- 238000006266 etherification reaction Methods 0.000 description 1
- 125000005909 ethyl alcohol group Chemical group 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000007928 intraperitoneal injection Substances 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000015689 metaplastic ossification Effects 0.000 description 1
- 230000000116 mitigating effect Effects 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004112 neuroprotection Effects 0.000 description 1
- 239000003002 pH adjusting agent Substances 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000004224 protection Effects 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 238000001953 recrystallisation Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 230000005070 ripening Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 235000007586 terpenes Nutrition 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H17/00—Compounds containing heterocyclic radicals directly attached to hetero atoms of saccharide radicals
- C07H17/04—Heterocyclic radicals containing only oxygen as ring hetero atoms
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Molecular Biology (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The invention discloses the preparation methods of 7 O ethyl morronisides.7 best O ethyl morroniside preparation processes are filtered out by many experiments, including dissolving method, pH value in reaction, heating temperature, heating time etc. answers parameter, and preferably goes out best purification process, entire reaction process reasonable design, operability is strong, 7 O ethyl morronisides synthetic ratios are high, and impurity is few, and purity is high, it is at low cost, it can be achieved that industrialized production.The present invention further passes through the anti-inflammatory experiment screening in inside and outside, the results showed that 7 O ethyls morronisides are respectively provided with good antiphlogistic effects in vivo and in vitro, have the potential use for being prepared into anti-inflammatory drug.
Description
Technical field
The present invention relates to a kind of preparation methods of compound, and in particular to the preparation method of 7-O- ethyl morronisides belongs to doctor
Medicine technical field.
Background technology
Dogwood fruit is the drying and ripening pulp of Cornaceae plant Fructus Corni, be 40 kinds of extremely common rare Chinese medicines it
One.Modern research shows that iridoid glycoside constituents are the important active substances of dogwood fruit, there is anti-inflammatory, liver protection, immunosupress
Isoreactivity.Current pharmaceutical research has tentatively disclosed the work of the wherein ingredients such as new glycosides of morroniside, loganin, Fructus Corni
Property, and the pharmacological action of other iridoid glycoside constituents is still not clear at present.Literature research, which shows that morroniside has, improves sugar
The effects that sick complication of urine and mitigation cerebral ischemia, there is anti-inflammatory, anti-oxidant and neuroprotection isoreactivity.7-O- ethyls are not
Promise glycosides is a kind of iridoid glycoside constituents (content is about 0.05~0.06mg/g) in dogwood fruit, contains morroniside skeleton knot
Structure, however whether it there is the similar pharmacological activity of morroniside yet there are no systematic research.Since 7-O- ethyl morronisides are on mountain
Content is less in cornus medicine materical crude slice, is detached from medicine materical crude slice, purifies the needs that cannot preferably meet inside and outside pharmacology activity research.Cause
This, the method for the Synthesis of present invention research and development 7-O- ethyl morronisides, on this basis to the internal of 7-O- ethyl morronisides
Outer anti-inflammatory effect has carried out systematic research.
Invention content
Goal of the invention：The object of the present invention is to provide a kind of new preparation process of 7-O- ethyls morroniside, and the present invention
Investigate the inside and outside anti-inflammatory effect of 7-O- ethyl morronisides.
Technical solution：In order to achieve the goal above, the technical solution that the present invention takes is：
The preparation method of 7-O- ethyl morronisides, includes the following steps：
(1) it dissolves：Iridoid glycoside in Fructus Corni Officinalis is taken as raw material, adds the ethyl alcohol of 10~20 times of volumes of raw material weight, ultrasound
Dissolving；
(2) pH is adjusted：After ultrasonic dissolution, the pH value of solution is adjusted to 1~3 or adds in acetylation reagent；
(3) it heats：10~20h of heating and refluxing extraction filters to obtain extracting solution, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract is dissolved in ethyl acetate, is filtered, removes insoluble matter, concentration, concentrate adds in dichloromethane
Alkane places crystallization, repeats crystallization 1~3 time, is dried under reduced pressure up to 7-O- ethyl morronisides.
Preferably, the preparation method of above-described 7-O- ethyls morroniside, iridoid glycoside in Fructus Corni Officinalis are
Using Fructus Corni as raw material, iridoid glycoside in Fructus Corni Officinalis is prepared using water extracting method.Preferably, Fructus Corni ring
Contain 10~20% loganin of 20~40% morroniside of weight percent and weight percent in the total glycosides of alkene ether terpene.
As more preferred scheme, the preparation method of the 7-O- ethyl morronisides, in iridoid glycoside in Fructus Corni Officinalis
Contain 20% loganin of 40% morroniside of weight percent and weight percent.
Preferably, the preparation method of above-described 7-O- ethyls morroniside, ethyl alcohol is anhydrous in step (1)
Alcohol；Supersonic frequency is 20~60kHz, power 30kW.The present invention screens to obtain the super of best ultrasonic dissolution by many experiments
Acoustic frequency and power are greatly improved the dissolution rate of reactant, ensure reaction conversion ratio.
Preferably, the preparation method of above-described 7-O- ethyls morroniside, the pH adjusting agent in step (2)
For one kind in phosphoric acid, hydrochloric acid, sulfuric acid or nitric acid.The present invention is screened by many experiments, and it is 1~3 to adjust reactant pH value, can
Synthesis rate is improved, reaction yield is improved, the conversion ratio of 7-O- ethyl morronisides can be made more than 93%.
Preferably, the preparation method of above-described 7-O- ethyls morroniside, at 80~100 DEG C in step (3)
Heating and refluxing extraction 16h, the present invention screen different reaction temperatures and reaction time, contrast and experiment table by many experiments
It is bright, the synthetic ratio highest of 7-O- ethyl morronisides can be made in 80~100 DEG C of heating and refluxing extraction 16h, impurity is few, and yield is high.
Preferably, the preparation method of above-described 7-O- ethyls morroniside, purification step, the present invention is by big
The different solvent of experiment screening is measured, the experimental results showed that being dissolved using ethyl acetate, insoluble matter can be effectively removed, and use
Preferred dichloromethane solvent can obtain the 7-O- ethyl morronisides of more than 98% purity as recrystallization solvent.The present invention
The purification step of column chromatography complexity is needed not move through, faster, working efficiency higher is at low cost, is more advantageous to industry for purifying speed
Metaplasia is produced.
The present invention passes through the anti-inflammatory experiment in inside and outside, the results showed that 7-O- ethyls morroniside is respectively provided with well in vivo and in vitro
Antiphlogistic effects.
Advantageous effect：Compared to the prior art the preparation method of 7-O- ethyls morroniside provided by the invention has following excellent
Point：
The present invention filters out best 7-O- ethyl morroniside preparation processes by many experiments, including dissolving method, adjusts
Preferred pH value, heating temperature, heating time response parameter are saved, and preferably goes out best purification process, entire reaction process
Reasonable design, operability is strong, and 7-O- ethyl morronisides synthetic ratio is high, and impurity is few, and purity is high, at low cost, it can be achieved that industrialization
Production.
The present invention proves that 7-O- ethyls morroniside is respectively provided with very in vivo and in vitro for the first time by the anti-inflammatory experiment screening in inside and outside
Good antiphlogistic effects, can be used for being prepared into anti-inflammatory drug.
Description of the drawings
Fig. 1 is the structure diagram of 7-O- ethyls morroniside provided by the present invention.
Fig. 2 is the block diagram that 7-O- ethyls morroniside influences the Raw264.7 cell TNF secretions-α that LPS is stimulated.
Fig. 3 is that 7-O- ethyls morroniside secretes the Raw264.7 cells that LPS is stimulated the block diagram that IL-6 influences.
Fig. 4 is the block diagram that 7-O- ethyls morroniside influences LPS inducible system inflammation mice serum IL-6 levels.
Fig. 5 is the block diagram that 7-O- ethyls morroniside influences LPS inducible system inflammation mice serum TNF-α level.
Specific embodiment
According to following embodiments, the present invention may be better understood.It is however, as it will be easily appreciated by one skilled in the art that real
It applies the described specific material proportion of example, process conditions and its result and is merely to illustrate the present invention, without that should will not limit
The present invention described in detail in claims processed.
Embodiment 1
The preparation method of 7-O- ethyl morronisides, includes the following steps：
(1) it dissolves：Iridoid glycoside in Fructus Corni Officinalis (containing morroniside 40%, loganin 20%) 1kg is weighed, adds in 10L
Absolute ethyl alcohol, under normal temperature condition stirring be allowed to be uniformly dispersed；
(2) pH value is adjusted：It adds in suitable phosphoric acid,diluted and adjusts pH=3；
(3) heating reaction：Flow back 16h at 80 DEG C；Extracting solution is filtered to obtain, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract adds in ethyl acetate dissolving, filters away insoluble matter, and filtrate is concentrated into small size, to
Dichloromethane is added in concentrate to stir evenly to there is Precipitation, places crystallization, and crystal is dissolved again with ethyl acetate, adds in dichloro
Methane recrystallizes, and filtering is dried under reduced pressure to obtain compound 7-O- ethyl morroniside product 365g, conversion ratio 93%.MS prompts gainedization
Adduct molecule amount：434g/mol, high resolution mass spectrum molecular formula：C19H30O11。(1H-NMR is consistent with document report, referring to
Fortunatus SUNGHWA,H.S.,et al.Iodine-Catalyzed Etherification of
Morroniside.Chem.Pharm.Bull,2009.57(1):112-115.), structural formula such as Fig. 1.
Embodiment 2
The preparation method of 7-O- ethyl morronisides, includes the following steps：
(1) it dissolves：Weigh iridoid glycoside in Fructus Corni Officinalis (containing morroniside 40%, loganin 20%) 1kg, input extraction
Tank adds in 15L absolute ethyl alcohols, is stirred under normal temperature condition and be allowed to be uniformly dispersed；
(2) pH value is adjusted：It adds in suitable dilute sulfuric acid and adjusts PH=2；
(3) heating reaction：90 DEG C of reflux 16h；Extracting solution is filtered to obtain, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract adds in ethyl acetate dissolving, is filtered to remove insoluble matter, and filtrate is concentrated into small size, to
Dichloromethane is added in concentrate to stir evenly to there is Precipitation, places crystallization, and crystal is dissolved again with ethyl acetate, adds in dichloro
Methane recrystallizes, and filtering is dried under reduced pressure to obtain compound 7-O- ethyl morroniside product 390g, conversion ratio 98%.MS prompts gainedization
Adduct molecule amount：434, high resolution mass spectrum molecular formula：C19H30O11.Structural formula such as Fig. 1.
Embodiment 3
The preparation method of 7-O- ethyl morronisides, includes the following steps：
(1) it dissolves：Iridoid glycoside in Fructus Corni Officinalis (containing morroniside 40%, loganin 20%) 1kg is weighed, adds in 20L
Absolute ethyl alcohol, under normal temperature condition stirring be allowed to be uniformly dispersed；
(2) pH value is adjusted：It adds in suitable dilute hydrochloric acid and adjusts pH=1；
(3) heating reaction：100 DEG C of refluxing extraction 16h；Extracting solution is filtered to obtain, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract adds in ethyl acetate dissolving, filters away insoluble matter, and filtrate is concentrated into small size, to
Dichloromethane is added in concentrate to stir evenly to there is Precipitation, places crystallization, and crystal is dissolved again with ethyl acetate, adds in dichloro
Methane recrystallizes, and filtering is dried under reduced pressure to obtain compound 7-O- ethyl morroniside product 380g, conversion ratio 95%, MS prompting gainedization
Adduct molecule amount：434g/mol, high resolution mass spectrum molecular formula：C19H30O11.Structural formula such as Fig. 1.
Embodiment 4
The preparation method of 7-O- ethyl morronisides, includes the following steps：
(1) it dissolves：Iridoid glycoside in Fructus Corni Officinalis (containing morroniside 30%, loganin 15%) 1kg is weighed, adds in 10L
Absolute ethyl alcohol, under normal temperature condition stirring be allowed to be uniformly dispersed；
(2) pH value is adjusted：It adds in suitable dust technology and adjusts pH=1；
(3) heating reaction：100 DEG C of reflux 16h；Extracting solution is filtered to obtain, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract adds in ethyl acetate dissolving, filters away insoluble matter, and filtrate is concentrated into small size, to
Dichloromethane is added in concentrate to stir evenly to there is Precipitation, places crystallization, and crystal is dissolved again with ethyl acetate, adds in dichloro
Methane recrystallizes, and filtering is dried under reduced pressure to obtain 7-O- ethyl morroniside product 375g, conversion ratio 93%.MS prompting gained compounds point
Son amount：434g/mol, high resolution mass spectrum molecular formula：C19H30O11.Structural formula such as Fig. 1.
Embodiment 5
The preparation method of 7-O- ethyl morronisides, includes the following steps：
(1) it dissolves：Iridoid glycoside in Fructus Corni Officinalis (containing morroniside 30%, loganin 20%) 1kg is weighed, adds in 20L
Absolute ethyl alcohol, under normal temperature condition stirring be allowed to be uniformly dispersed；
(2) pH value is adjusted：It adds in suitable dilute hydrochloric acid and adjusts pH=1.5；
(3) heating reaction：100 DEG C of reflux 16h；Extracting solution is filtered to obtain, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract adds in ethyl acetate dissolving, filters away insoluble matter, and filtrate is concentrated into small size, to
Dichloromethane is added in concentrate to stir evenly to there is Precipitation, places crystallization, and crystal is dissolved again with ethyl acetate, adds in dichloro
Methane recrystallizes, and filtering is dried under reduced pressure to obtain 7-O- ethyl morroniside product 392g, conversion ratio 98%.MS prompting gained compounds point
Son amount：434g/mol, high resolution mass spectrum molecular formula：C19H30O11.Structural formula such as Fig. 1.
6 anti-inflammatory experiment of embodiment
1st, extracorporeal anti-inflammatory experimental method：
Experiment is divided into 5 groups, i.e. Normal group, dexamethasone positive drug group, LPS model groups, 7-O- ethyl morronisides is low
Dosage group (5 μM), 7-O- ethyl morronisides high dose group (20 μM).It takes the logarithm the RAW264.7 cells in growth period, prepared by digestion
Cell density is adjusted to 5 × 10 by cell suspension4A/ml takes 1mL to be inoculated in 24 well culture plates, and every group of 4 multiple holes are placed in
Containing 5%CO2, cultivate in 37 DEG C of incubators, until degrees of fusion is 80% or so.
Each group dose regimen is：(1) Normal group adds in complete serum free medium；(2) dexamethasone positive drug group,
It adds in containing dexamethasone (1 μ g/ml) culture medium；(3) LPS model groups add in complete serum free medium；(4) 7-O- ethyls are not
Promise glycosides is low, high dose group：It is separately added into the serum free medium containing 5,20 μM of 7-O- ethyl morronisides；Each experimental group cell is put
In containing 5%CO2, in 37 DEG C of incubators after 30min, in addition to normal group, every group of addition LPS to final concentration of 1 μ g/ml.Continue to train
It supports and draws cell supernatant afterwards for 24 hours, using ELISA method detection TNF-α, IL-6.
2nd, internal anti-inflammatory experimental method：
Experiment packet：Take BALB/C mice (male, 10-12 week old) 50, be randomly divided into 5 groups, respectively control group,
LPS model groups, dexamethasone positive controls (1.3mg/kg), 7-O- ethyl morroniside low dose groups (10mg/kg), 7-O- second
Base morroniside high dose group (30mg/kg) (tail vein injection saline+intraperitoneal injection of saline group), every group 10.
Each group dose regimen is：Normal group and LPS model group mouse tail vein injection physiological saline, remaining 3 groups of tail are quiet
Arteries and veins relative medicine.After 1h, in addition to control group, every group of mouse peritoneal injection LPS (0.8mg/kg).Blood, ice bath 10 to 15 are taken after 4h
Minute, 8000g centrifuges 10 minutes separation serum, and IL-6 and TNF-α are measured using ELISA method.
3rd, experimental result
3.1 extracorporeal anti-inflammatory experimental results：
As shown in Fig. 2, the Raw264.7 cell TNF secretions-α that 7-O- ethyls morroniside provided by the invention stimulates LPS
Influence and as shown in figure 3, the Raw264.7 cells that 7-O- ethyls morroniside provided by the invention stimulate LPS secrete IL-6's
It influences, compared with normal group, 7-O- ethyls morroniside can significantly inhibit the Raw264.7 cell TNF secretions-α stimulated due to LPS
And IL-6.
Anti-inflammatory experimental result in 3.2 bodies：
As shown in figure 4,7-O- ethyls morroniside provided by the invention is to LPS inducible system inflammation mice serum IL-6 water
Flat influence and as shown in figure 5,7-O- ethyls morroniside provided by the invention to LPS inducible system inflammation mice serums TNF-
The influence of alpha levels, during mouse vivo medicine-feeding 30mg/kg, 7-O- ethyls morroniside can inhibit LPS inducible system inflammation mouse
Blood serum IL-6 can also inhibit the release of TNF-α, and can inhibit LPS inducible system inflammation mice serums IL-6 during 10mg/kg
Release.
The technical concepts and features of embodiment of above only to illustrate the invention, its object is to allow be familiar with technique
People understands the content of present invention and is implemented, and it is not intended to limit the scope of the present invention, all real according to spirit of the invention
The equivalent change or modification that matter is done should all cover within the scope of the present invention.
Claims (6)
1. the preparation method of the 7-O- ethyl morronisides with anti-inflammatory effect, it is characterised in that include the following steps：
(1) it dissolves：Iridoid glycoside in Fructus Corni Officinalis is taken to add the ethyl alcohol of 10~20 times of volumes of raw material weight, ultrasonic dissolution for raw material；
(2) pH is adjusted：After ultrasonic dissolution, the pH value of solution is adjusted to 1~3；
(3) it heats：10~20h of heating and refluxing extraction filters to obtain extracting solution, is concentrated to give medicinal extract；
(4) it purifies, crystallize：Medicinal extract is dissolved in ethyl acetate, is filtered, removes insoluble matter, concentration, concentrate adds in dichloromethane,
Crystallization is placed, repeats crystallization 1~3 time, is dried under reduced pressure up to 7-O- ethyl morronisides；
Contain 10~20% Strychnos nux-vomica of 20~40% morroniside of weight percent and weight percent in iridoid glycoside in Fructus Corni Officinalis
Glycosides.
2. the preparation method of 7-O- ethyls morroniside according to claim 1, which is characterized in that Fructus Corni iridoid total
Contain 20% loganin of 40% morroniside of weight percent and weight percent in glycosides.
3. the preparation method of 7-O- ethyls morroniside according to claim 1, which is characterized in that ethyl alcohol is in step (1)
Absolute alcohol；Supersonic frequency is 20~60kHz, power 30kW.
4. the preparation method of 7-O- ethyls morroniside according to claim 1, which is characterized in that the pH value in step (2)
Conditioning agent is one kind in phosphoric acid, hydrochloric acid, sulfuric acid or nitric acid.
5. the preparation method of 7-O- ethyls morroniside according to claim 1, which is characterized in that in step (3) 80~
100 DEG C of heating and refluxing extraction 16h.
6. application of the 7-O- ethyl morronisides that claim 1 is prepared in anti-inflammatory drug is prepared.
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