CN110407797A - Derived from the secalonic acid class compound Secalonic acid K and preparation method of penicillium oxalicum - Google Patents
Derived from the secalonic acid class compound Secalonic acid K and preparation method of penicillium oxalicum Download PDFInfo
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- penicillium oxalicum
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- secalonic acid
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- 150000001875 compounds Chemical class 0.000 title claims abstract description 31
- 241000985513 Penicillium oxalicum Species 0.000 title claims abstract description 23
- 238000002360 preparation method Methods 0.000 title claims abstract description 9
- DRYDKQOPVBDZMQ-UHFFFAOYSA-N Secalonic acid A Natural products COC(=O)C12Oc3ccc(c(O)c3C(=O)C1=C(O)CC(C)C2O)c4ccc5OC6(C(O)C(C)CC(=C6C(=O)c5c4O)O)C(=O)OC DRYDKQOPVBDZMQ-UHFFFAOYSA-N 0.000 title abstract description 12
- NFZJAYYORNVZNI-UHFFFAOYSA-N methyl 4,8,9-trihydroxy-3-methyl-1-oxo-7-(1,5,9-trihydroxy-10a-methoxycarbonyl-6-methyl-8-oxo-6,7-dihydro-5h-xanthen-2-yl)-3,4-dihydro-2h-xanthene-4a-carboxylate Chemical class OC1C(C)CC(=O)C2=C(O)C3=C(O)C(C4=CC=C5OC6(C(=C(O)C5=C4O)C(=O)CC(C)C6O)C(=O)OC)=CC=C3OC21C(=O)OC NFZJAYYORNVZNI-UHFFFAOYSA-N 0.000 title abstract description 12
- 229930183845 Secalonic acid Natural products 0.000 title abstract description 9
- 238000000855 fermentation Methods 0.000 claims abstract description 8
- 230000004151 fermentation Effects 0.000 claims abstract description 8
- 239000003814 drug Substances 0.000 claims abstract description 7
- 229940079593 drug Drugs 0.000 claims abstract description 7
- 239000000463 material Substances 0.000 claims abstract description 6
- 208000009849 Female Genital Neoplasms Diseases 0.000 claims 1
- 206010028980 Neoplasm Diseases 0.000 abstract description 9
- 201000011510 cancer Diseases 0.000 abstract description 9
- 230000000259 anti-tumor effect Effects 0.000 abstract description 3
- 230000009702 cancer cell proliferation Effects 0.000 abstract description 3
- 239000002246 antineoplastic agent Substances 0.000 abstract description 2
- 229940041181 antineoplastic drug Drugs 0.000 abstract description 2
- 238000002474 experimental method Methods 0.000 abstract description 2
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 27
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 18
- 210000004027 cell Anatomy 0.000 description 14
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 12
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 12
- 230000000694 effects Effects 0.000 description 9
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 8
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 8
- 239000000741 silica gel Substances 0.000 description 8
- 229910002027 silica gel Inorganic materials 0.000 description 8
- 239000000287 crude extract Substances 0.000 description 7
- 239000000523 sample Substances 0.000 description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 7
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 6
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 6
- 238000000926 separation method Methods 0.000 description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 5
- 239000003795 chemical substances by application Substances 0.000 description 5
- 230000002401 inhibitory effect Effects 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 4
- -1 Secalonic acid class compound Chemical class 0.000 description 4
- 238000004587 chromatography analysis Methods 0.000 description 4
- 238000010828 elution Methods 0.000 description 4
- 239000000284 extract Substances 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 238000000746 purification Methods 0.000 description 4
- 210000004881 tumor cell Anatomy 0.000 description 4
- 238000000034 method Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 description 2
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 2
- 206010005003 Bladder cancer Diseases 0.000 description 2
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 2
- 241000233866 Fungi Species 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- 239000007836 KH2PO4 Substances 0.000 description 2
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 2
- 229930195725 Mannitol Natural products 0.000 description 2
- NFZJAYYORNVZNI-ZKHWAINJSA-N Secalonic acid A Chemical compound O[C@H]1[C@H](C)CC(=O)C2=C(O)C3=C(O)C(C4=CC=C5O[C@]6(C(=C(O)C5=C4O)C(=O)C[C@@H](C)[C@@H]6O)C(=O)OC)=CC=C3O[C@@]21C(=O)OC NFZJAYYORNVZNI-ZKHWAINJSA-N 0.000 description 2
- NFZJAYYORNVZNI-OCHURCMPSA-N Secalonic acid D Chemical compound O[C@@H]1[C@@H](C)CC(=O)C2=C(O)C3=C(O)C(C4=CC=C5O[C@@]6(C(=C(O)C5=C4O)C(=O)C[C@H](C)[C@H]6O)C(=O)OC)=CC=C3O[C@]21C(=O)OC NFZJAYYORNVZNI-OCHURCMPSA-N 0.000 description 2
- MZZSDCJQCLYLLL-UHFFFAOYSA-N Secalonsaeure A Natural products COC(=O)C12OC3C(CC1=C(O)CC(C)C2O)C(=CC=C3c4ccc(O)c5C(=O)C6=C(O)CC(C)C(O)C6(Oc45)C(=O)OC)O MZZSDCJQCLYLLL-UHFFFAOYSA-N 0.000 description 2
- MCWOXLPZYFOWRX-UHFFFAOYSA-N Stemphyperylenol Natural products OC1CC(=O)C2=C(O)C=CC3=C2C1C1=C2C3C(O)CC(=O)C2=C(O)C=C1 MCWOXLPZYFOWRX-UHFFFAOYSA-N 0.000 description 2
- 208000007097 Urinary Bladder Neoplasms Diseases 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 229940041514 candida albicans extract Drugs 0.000 description 2
- 150000001793 charged compounds Chemical class 0.000 description 2
- NEHMKBQYUWJMIP-UHFFFAOYSA-N chloromethane Chemical class ClC NEHMKBQYUWJMIP-UHFFFAOYSA-N 0.000 description 2
- 238000004440 column chromatography Methods 0.000 description 2
- 238000007796 conventional method Methods 0.000 description 2
- 238000005100 correlation spectroscopy Methods 0.000 description 2
- 239000006071 cream Substances 0.000 description 2
- 239000003480 eluent Substances 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 238000003919 heteronuclear multiple bond coherence Methods 0.000 description 2
- 239000000594 mannitol Substances 0.000 description 2
- 235000010355 mannitol Nutrition 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 2
- LPUQAYUQRXPFSQ-DFWYDOINSA-M monosodium L-glutamate Chemical compound [Na+].[O-]C(=O)[C@@H](N)CCC(O)=O LPUQAYUQRXPFSQ-DFWYDOINSA-M 0.000 description 2
- 235000013923 monosodium glutamate Nutrition 0.000 description 2
- 239000004223 monosodium glutamate Substances 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- 238000004262 preparative liquid chromatography Methods 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 230000003068 static effect Effects 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 238000002604 ultrasonography Methods 0.000 description 2
- 201000005112 urinary bladder cancer Diseases 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 230000001093 anti-cancer Effects 0.000 description 1
- 238000013096 assay test Methods 0.000 description 1
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 239000012490 blank solution Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 239000003560 cancer drug Substances 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000008025 crystallization Effects 0.000 description 1
- 239000000824 cytostatic agent Substances 0.000 description 1
- 230000001085 cytostatic effect Effects 0.000 description 1
- 238000000119 electrospray ionisation mass spectrum Methods 0.000 description 1
- 229910052564 epsomite Inorganic materials 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 150000002576 ketones Chemical class 0.000 description 1
- 239000006101 laboratory sample Substances 0.000 description 1
- 239000006193 liquid solution Substances 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- CSNNHWWHGAXBCP-UHFFFAOYSA-L magnesium sulphate Substances [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- 230000035479 physiological effects, processes and functions Effects 0.000 description 1
- 239000002504 physiological saline solution Substances 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 230000024053 secondary metabolic process Effects 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 125000003831 tetrazolyl group Chemical group 0.000 description 1
- JNELGWHKGNBSMD-UHFFFAOYSA-N xanthone Chemical group C1=CC=C2C(=O)C3=CC=CC=C3OC2=C1 JNELGWHKGNBSMD-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/78—Ring systems having three or more relevant rings
- C07D311/80—Dibenzopyrans; Hydrogenated dibenzopyrans
- C07D311/82—Xanthenes
- C07D311/84—Xanthenes with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached in position 9
- C07D311/86—Oxygen atoms, e.g. xanthones
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
- C12P17/16—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing two or more hetero rings
- C12P17/162—Heterorings having oxygen atoms as the only ring heteroatoms, e.g. Lasalocid
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Engineering & Computer Science (AREA)
- Veterinary Medicine (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Public Health (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The present invention relates to a kind of secalonic acid class compound Secalonic acid K and preparation method derived from penicillium oxalicum.The compound, which has, inhibits human cancer cell proliferation function.Its structural formula are as follows:.By fermented and cultured penicillium oxalicum (Penicillium oxalicum) IBPT-6, fermentation material is obtained, the compound is then isolated and purified out from fermentation material.It is verified by experiments, which has preferable anti-tumor activity to cancer cell.It can be used for preparing cancer cell multiplication and inhibit drug or anti-tumor drug.
Description
Technical field
The present invention relates to a kind of derived from the secalonic acid class compound Secalonic acid K of penicillium oxalicum and preparation side
Method belongs to biomedicine field.
Background technique
Secalonic acid class compound (Secalonic acids) belongs to the production of ergot pigment (Ergochrome) secondary metabolism
Object is xanthone dimer.Since Stoll etc. was in the isolated secalonic acid A from fungi in 1952
After (Secalonic acid A), the compound secalonic acid (A-I) of the series is just constantly found and studies.Rye ketone
Acid compounds have a variety of different physiological activity, with secalonic acid D(Secalonic acid D, SAD) for, 5 mg/
The SAD of ml is added in physiological saline, within the scope of 5-20 mg, it can treatment early stage bladder cancer, 50-100 mg range
It is interior, and be free from side effects generation effective in cure to more serious bladder cancer.It has been investigated that some marine fungis can be in secondary
The good secalonic acid class compound of structure novel, activity is generated in metabolic process, and there is good medicinal and industrialization prospect.
The present inventor studies and learns, penicillium oxalicum (Penicillium oxalicum) IBPT-6 has been (in 2013 12
The moon is deposited in China typical culture collection center on 25th, and address: Wuhan Wuhan University, deposit number are: CCTCC NO:M
2013714) crude extract of tunning has good cell inhibitory effect activity, studies then its active constituent.It grinds
Studying carefully secalonic acid class compound shown in discovery has anti-human cancer activity, has not yet to see the compound and presses down to the proliferation of human cancer cell
Active report is made, therefore in the market also there is not yet drug related to this.
Summary of the invention
The purpose of the present invention is to provide a kind of secalonic acid class compound Secalonic acid derived from penicillium oxalicum
K and preparation method.The compound, which has, inhibits cancer cell multiplication effect, has anticancer activity.Its structural formula are as follows:
。
The preparation method of the compound, be by fermented and cultured penicillium oxalicum (Penicillium oxalicum)
IBPT-6 obtains fermentation material, the compound is then isolated and purified out from fermentation material.Specific step is as follows:
1 fermenting and producing
By culture microorganism conventional method, take penicillium oxalicum (Penicillium oxalicum) IBPT-6 is inoculated into PDA
Cultivate 2 to 3 days, be then seeded into culture solution in 28 DEG C of incubators in solid slope culture medium, 28 DEG C static gas wave refrigerator 30 days
Afterwards, mycelium and fermentation liquid are obtained；The culture solution composition: every liter of water contains 20.0 g of mannitol, 3.0 g of yeast extract, maltose
20.0 g, 10.0 g of monosodium glutamate, glucose 10.0 g, KH2PO4 0.5 g、MgSO4·7H2O 0.3 g、NaCl 15.0 g。
The acquisition of 2 medicinal extract
With gauze by mycelium and separation of fermentative broth.Mycelium with acetone soln (contain 20% ~ 30% water) continuous ultrasound broken wall 3 times,
Filtering removal residue, obtains the mycelium crude extract containing acetone and water.Removal acetone is concentrated under reduced pressure, obtains the water-soluble of crude extract
Liquid, then ethyl acetate is added with volume ratio 1:2 and is extracted 3 times, ethyl acetate crude extract is obtained, is concentrated under reduced pressure into close dry that mycelium soaks
36.5 g of cream.
The separation and purification of 3 compounds
Mycelium medicinal extract is by 100-200 mesh silica gel mixed sample, and using petroleum ether: methylene chloride: methanol is subtracted as gradient eluent
Press silica gel chromatographic column chromatography.By simple thin-layer chromatographic analysis, merges, be separated into component A-E.Component D (5.9 g) (two
Chloromethanes: the eluate of methanol v/v=100:1) using methylene chloride: methanol carries out pressured column silica gel as gradient elution agent
Analysis merges after thin-layer chromatographic analysis and obtains five subfraction D1-D5.Component D3(1.1 g) with methanol: sour water (contains 0.1%
Trifluoroacetic acid) it is that gradient elution agent carries out reversed chromatographed on silica gel, isolated four subfraction D3-1 ~ D3-4.The Asia of D3
Component D3-2(168 mg) it is separated under the conditions of mobile phase is 50% acetonitrile (containing 0.1% trifluoroacetic acid) by semi-preparative liquid chromatography
Obtain the compound (2.3 mg).
The penicillium oxalicum (Penicillium oxalicum) IBPT-6, it has been deposited on December 25th, 2013
State's Type Tissue Collection, address: Wuhan Wuhan University, deposit number are: CCTCC NO:M 2013714.
The present invention also protects the compound to inhibit application and the chemical combination in human cancer cell hyperproliferation agent in preparation
Object is preparing the application in anti-human cancer drug.
Remarkable advantage of the invention: the secalonic acid compound shown in studying has not been reported and has significant inhibition people's cancer
Cell-proliferation activity has not yet to see the compound to the report of human cancer cell proliferation inhibition activity, therefore in the market also not yet
Seeing has drug related to this.
Detailed description of the invention
Fig. 1 Secalonic acid main COSY of K, HMBC and NOE signal.
Specific embodiment
The chemical structure of signified compound in the following example:
The fermenting and producing and separation and purification of 1 compound of embodiment
1 fermenting and producing
Produce bacterium fermented and cultured: by culture microorganism conventional method, take penicillium oxalicum (Penicillium oxalicum)
IBPT-6 (be deposited in China typical culture collection center on December 25th, 2013, address: protect by Wuhan Wuhan University
Hiding number is: CCTCC NO:M 2013714) in right amount, being inoculated into PDA solid slope culture medium and cultivates in 28 DEG C of incubators
2 to 3 days.
Take inclined-plane culture 2 to 3 days penicillium oxalicum (Penicillium oxalicum) appropriate IBPT-6, it is inoculated into dress
By 400mL culture solution, [culture solution is formed (grams per liter): mannitol 20.0, yeast extract 3.0, maltose 20.0, monosodium glutamate 10.0, Portugal
Grape sugar 10.0, KH2PO40.5, MgSO40.3, NaCl 15.0, constant volume] 1000mL conical flask in, 28 DEG C of static gas wave refrigerators
After 30 days, mycelium and fermentation liquid are obtained.
The acquisition of 2 medicinal extract
With gauze by mycelium and separation of fermentative broth.Mycelium with acetone soln (contain 20% ~ 30% water) continuous ultrasound broken wall 3 times,
Filtering removal residue, obtains the mycelium crude extract containing acetone and water.Removal acetone is concentrated under reduced pressure, obtains the water-soluble of crude extract
Liquid, then ethyl acetate is added with volume ratio 1:2 and is extracted 3 times, ethyl acetate crude extract is obtained, is concentrated under reduced pressure into close dry that mycelium soaks
36.5 g of cream.
The separation and purification of 3 compounds
Mycelium medicinal extract is by 100-200 mesh silica gel mixed sample, and using petroleum ether: methylene chloride: methanol is subtracted as gradient eluent
Press silica gel chromatographic column chromatography.By simple thin-layer chromatographic analysis, merges, be separated into component A-E.Component D (5.9 g) (two
Chloromethanes: the eluate of methanol v/v=100:1) using methylene chloride: methanol carries out pressured column silica gel as gradient elution agent
Analysis merges after thin-layer chromatographic analysis and obtains five subfraction D1-D5.Component D3(1.1 g) with methanol: sour water (contains 0.1%
Trifluoroacetic acid) it is that gradient elution agent carries out reversed chromatographed on silica gel, isolated four subfraction D3-1 ~ D3-4.The Asia of D3
Component D3-2(168 mg) it is separated under the conditions of mobile phase is 50% acetonitrile (containing 0.1% trifluoroacetic acid) by semi-preparative liquid chromatography
Obtain the compound (2.3 mg).
Compound: being yellow oil under room temperature, and high-resolution electrospray ionization mass spectrum HRESI-MS gives at m/z:639.1714
Molecular ion peak [M+H]+(calcd for C out32H31O14, 639.1714), molecular ion peak [M-H]-is provided at 637.1560
(calcd for C32H29O14, 637.1557), infer that its molecular formula is C32H30O14。1H and13C-NMR data are shown in Table 1, main
COSY, HMBC and NOE signal are shown in Fig. 1.
1 compound of table1H and13C-NMR data (500MHz1H and 125 MHz 13C, in DMSO d6)
a,b Signal can be interchanged
The test of 2 anti tumor activity in vitro of embodiment
1 laboratory sample and experimental method
The preparation test sample of sample solution is the pure compounds of separation and purification in above-described embodiment 1.Precision weighs suitable
Sample is measured, the solution of required concentration is configured to methanol, for surveying activity.
The squamous subculture of cell line and cell uses tumor cell line, and tumour cell uses the DMEM containing 10% FBS to cultivate
Base, at 37 DEG C in being passed through 5% CO2Incubator in squamous subculture.
Cell inhibitory effect activity test method
Tetrazolium (MTT) method: cell density is adjusted to every milliliter 2 × 10 by the tumour cell of logarithmic growth phase5A cell,
It is inoculated in 96 porocyte culture plates for 200 microlitres by every hole, is passed through 5% CO in 37 DEG C2Incubator in cultivate 4 hours.Every hole
2 microlitres of sample liquid or blank solution is added, after culture 24 hours, MTT liquid (every milliliter of 5 milligrams of physiology of MTT are added in every hole
Saline solution) 10 microlitres, continue culture 4 hours, 37 DEG C, 2000 revs/min are centrifuged 8 minutes, draw supernatant.DMSO is added in every hole
It each 100 microlitres, is vibrated 15 minutes on micro oscillator, until utilizing the production of MD company after crystallization is completely dissolved
SPECTRAMAX Plus type microplate reader measures extinction (OD) value of every hole at 570 nm.The sample in 96 orifice plate of same
Each concentration is respectively provided with three holes, and the another blank control that three holes are arranged and cell-free withered hole are (if drug has color to do accordingly
It is withered that drug concentration is cell-free).Each hole OD value first do it is corresponding cell-free withered, then take three hole mean OD values by IR (%)=
(ODBlank control-ODSample)/ODBlank control× 100% calculates the proliferation inhibition rate (IR%) of cell under each concentration.
2. experimental result
Cell inhibitory effect active testing result
In mtt assay test, according to the Cytostatic to tumor cell rate of the compound of various concentration, using SPSS16.0 software
Carry out data processing and calculation of half inhibitory concentration IC50Value.It the results are shown in Table 2.
Inhibitory activity (IC of 2 compound of table to cancer cell multiplication50, μM)
3. conclusion
The compound has preferable anti-tumor activity to cancer cell.It can be used as and prepare cancer cell multiplication inhibition drug or antitumor
Drug is for studying.
The foregoing is merely presently preferred embodiments of the present invention, all equivalent changes done according to scope of the present invention patent with
Modification, is all covered by the present invention.
Claims (3)
1. compound。
2. the preparation method of compound as described in claim 1, it is characterised in that: fermented and cultured penicillium oxalicum
(Penicillium oxalicum) IBPT-6, fermentation material is obtained, the compound is then isolated and purified out from fermentation material；
The penicillium oxalicum (Penicillium oxalicum) IBPT-6, Chinese Typical Representative training has been deposited on December 25th, 2013
Object collection is supported, address: Wuhan Wuhan University, deposit number are: CCTCC NO:M 2013714.
3. compound as described in claim 1 is preparing the application in gynecological tumor drug.
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| CN112107602A (en) * | 2020-09-08 | 2020-12-22 | 中国科学院烟台海岸带研究所 | A pair of twin-nitrogen containing alkaloid enantiomers, preparation and application thereof |
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| CN110407794A (en) * | 2019-04-26 | 2019-11-05 | 福州大学 | Secalonic acid K derived from penicillium oxalicum and the application on inhibition cancer cell multiplication |
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Non-Patent Citations (2)
| Title |
|---|
| LI CHEN等: "SECALONIC ACIDS H AND I, TWO NEW SECONDARY METABOLITES FROM THE MARINE-DERIVED FUNGUS PENICILLIUM OXALICUM", 《HETEROCYCLES》 * |
| LI CHEN等: "SECALONIC ACIDS J-M, FOUR NEW SECONDARY METABOLITES FROM THE MARINE-DERIVED FUNGUS PENICILLIUM OXALICUM", 《HETEROCYCLES》 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112107602A (en) * | 2020-09-08 | 2020-12-22 | 中国科学院烟台海岸带研究所 | A pair of twin-nitrogen containing alkaloid enantiomers, preparation and application thereof |
| CN112107602B (en) * | 2020-09-08 | 2021-10-15 | 中国科学院烟台海岸带研究所 | A pair of twin-nitrogen containing alkaloid enantiomers, preparation and application thereof |
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