CN116589514A - Guaiane type sesquiterpene glycoside in rhizoma atractylodis, and preparation method and application thereof - Google Patents
Guaiane type sesquiterpene glycoside in rhizoma atractylodis, and preparation method and application thereof Download PDFInfo
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- MCNAURNYDFSEML-UHFFFAOYSA-N Guaiane Natural products CC1CCC(C(C)=C)C(O)C2=C(C)C(=O)CC12 MCNAURNYDFSEML-UHFFFAOYSA-N 0.000 title claims abstract description 13
- -1 sesquiterpene glycoside Chemical class 0.000 title claims abstract description 13
- 229930182470 glycoside Natural products 0.000 title claims abstract description 12
- 238000002360 preparation method Methods 0.000 title claims description 10
- QAQCPAHQVOKALN-RMEBNNNOSA-N guaiane Chemical compound C1[C@H](C(C)C)CC[C@H](C)[C@@H]2CC[C@H](C)[C@@H]21 QAQCPAHQVOKALN-RMEBNNNOSA-N 0.000 title abstract description 9
- 229930004725 sesquiterpene Natural products 0.000 title abstract description 9
- 150000001875 compounds Chemical class 0.000 claims abstract description 32
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims abstract description 17
- 239000000741 silica gel Substances 0.000 claims abstract description 17
- 229910002027 silica gel Inorganic materials 0.000 claims abstract description 17
- 239000003814 drug Substances 0.000 claims abstract description 14
- 238000004440 column chromatography Methods 0.000 claims abstract description 12
- 229920005654 Sephadex Polymers 0.000 claims abstract description 9
- 239000012507 Sephadex™ Substances 0.000 claims abstract description 9
- 230000002441 reversible effect Effects 0.000 claims abstract description 9
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims abstract description 7
- 208000005718 Stomach Neoplasms Diseases 0.000 claims abstract description 7
- 206010017758 gastric cancer Diseases 0.000 claims abstract description 7
- 201000007270 liver cancer Diseases 0.000 claims abstract description 7
- 208000014018 liver neoplasm Diseases 0.000 claims abstract description 7
- 201000005202 lung cancer Diseases 0.000 claims abstract description 7
- 208000020816 lung neoplasm Diseases 0.000 claims abstract description 7
- 239000011347 resin Substances 0.000 claims abstract description 7
- 229920005989 resin Polymers 0.000 claims abstract description 7
- 201000011549 stomach cancer Diseases 0.000 claims abstract description 7
- 238000001953 recrystallisation Methods 0.000 claims abstract description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 78
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 44
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 39
- 239000011259 mixed solution Substances 0.000 claims description 21
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 18
- 239000002904 solvent Substances 0.000 claims description 15
- 230000002829 reductive effect Effects 0.000 claims description 9
- 241000132011 Atractylodes lancea Species 0.000 claims description 7
- 239000003480 eluent Substances 0.000 claims description 7
- 238000010828 elution Methods 0.000 claims description 7
- 239000000243 solution Substances 0.000 claims description 7
- 239000012043 crude product Substances 0.000 claims description 6
- 238000010898 silica gel chromatography Methods 0.000 claims description 6
- 206010028980 Neoplasm Diseases 0.000 claims description 5
- 238000000605 extraction Methods 0.000 claims description 5
- 239000002994 raw material Substances 0.000 claims description 5
- 238000001035 drying Methods 0.000 claims description 4
- 239000000706 filtrate Substances 0.000 claims description 3
- 238000001914 filtration Methods 0.000 claims description 3
- 238000010829 isocratic elution Methods 0.000 claims description 3
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 claims description 3
- 238000000034 method Methods 0.000 claims description 3
- 239000000047 product Substances 0.000 claims description 3
- 238000010992 reflux Methods 0.000 claims description 3
- 238000004809 thin layer chromatography Methods 0.000 claims description 3
- 241000132012 Atractylodes Species 0.000 claims description 2
- 230000009897 systematic effect Effects 0.000 claims description 2
- 230000000694 effects Effects 0.000 abstract description 9
- 229940079593 drug Drugs 0.000 abstract description 7
- 230000005764 inhibitory process Effects 0.000 abstract description 5
- 238000011160 research Methods 0.000 abstract description 5
- 150000002338 glycosides Chemical class 0.000 abstract description 4
- 230000000259 anti-tumor effect Effects 0.000 abstract description 3
- 238000011161 development Methods 0.000 abstract description 3
- 239000000126 substance Substances 0.000 abstract description 3
- 150000004354 sesquiterpene derivatives Chemical class 0.000 abstract description 2
- 239000002246 antineoplastic agent Substances 0.000 abstract 1
- 229940041181 antineoplastic drug Drugs 0.000 abstract 1
- 230000000857 drug effect Effects 0.000 abstract 1
- 238000005516 engineering process Methods 0.000 abstract 1
- 230000005918 in vitro anti-tumor Effects 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 15
- 238000001228 spectrum Methods 0.000 description 6
- 210000004881 tumor cell Anatomy 0.000 description 6
- 229910052799 carbon Inorganic materials 0.000 description 5
- TYPSVDGIQAOBAD-DZGCQCFKSA-N Atractylone Chemical compound C([C@]1(C)C2)CCC(=C)[C@@H]1CC1=C2OC=C1C TYPSVDGIQAOBAD-DZGCQCFKSA-N 0.000 description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical group [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 4
- TYPSVDGIQAOBAD-UHFFFAOYSA-N atractylone Natural products C1C2(C)CCCC(=C)C2CC2=C1OC=C2C TYPSVDGIQAOBAD-UHFFFAOYSA-N 0.000 description 4
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 3
- 238000005160 1H NMR spectroscopy Methods 0.000 description 3
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 3
- 206010061218 Inflammation Diseases 0.000 description 3
- 229960004316 cisplatin Drugs 0.000 description 3
- DQLATGHUWYMOKM-UHFFFAOYSA-L cisplatin Chemical compound N[Pt](N)(Cl)Cl DQLATGHUWYMOKM-UHFFFAOYSA-L 0.000 description 3
- 238000000990 heteronuclear single quantum coherence spectrum Methods 0.000 description 3
- 230000004054 inflammatory process Effects 0.000 description 3
- 238000005100 correlation spectroscopy Methods 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 230000029087 digestion Effects 0.000 description 2
- 238000002072 distortionless enhancement with polarization transfer spectrum Methods 0.000 description 2
- 239000012634 fragment Substances 0.000 description 2
- LHGVFZTZFXWLCP-UHFFFAOYSA-N guaiacol Chemical group COC1=CC=CC=C1O LHGVFZTZFXWLCP-UHFFFAOYSA-N 0.000 description 2
- 238000002114 high-resolution electrospray ionisation mass spectrometry Methods 0.000 description 2
- 125000001434 methanylylidene group Chemical group [H]C#[*] 0.000 description 2
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000000144 pharmacologic effect Effects 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- JUJWROOIHBZHMG-RALIUCGRSA-N pyridine-d5 Chemical compound [2H]C1=NC([2H])=C([2H])C([2H])=C1[2H] JUJWROOIHBZHMG-RALIUCGRSA-N 0.000 description 2
- 239000002689 soil Substances 0.000 description 2
- 210000000952 spleen Anatomy 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 230000004083 survival effect Effects 0.000 description 2
- 239000000341 volatile oil Substances 0.000 description 2
- KWDNQVRRYNIDTM-UHFFFAOYSA-N (1S,4S,5S,7R,10R)-10,11,14-Trihydroxyguai-3-one 11-O-D-glucopyranoside Natural products C1CC(O)(CO)C2CC(=O)C(C)C2CC1C(C)(C)OC1OC(CO)C(O)C(O)C1O KWDNQVRRYNIDTM-UHFFFAOYSA-N 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
- 239000004382 Amylase Substances 0.000 description 1
- 102000013142 Amylases Human genes 0.000 description 1
- 108010065511 Amylases Proteins 0.000 description 1
- 241000092665 Atractylodes macrocephala Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- 208000001333 Colorectal Neoplasms Diseases 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 244000166124 Eucalyptus globulus Species 0.000 description 1
- 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 1
- 241000147041 Guaiacum officinale Species 0.000 description 1
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical group CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 1
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 description 1
- 229930182816 L-glutamine Natural products 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 208000004880 Polyuria Diseases 0.000 description 1
- 244000184734 Pyrus japonica Species 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 238000005903 acid hydrolysis reaction Methods 0.000 description 1
- 235000019418 amylase Nutrition 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 230000004856 capillary permeability Effects 0.000 description 1
- 238000001460 carbon-13 nuclear magnetic resonance spectrum Methods 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 230000003833 cell viability Effects 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 206010061428 decreased appetite Diseases 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- VILAVOFMIJHSJA-UHFFFAOYSA-N dicarbon monoxide Chemical compound [C]=C=O VILAVOFMIJHSJA-UHFFFAOYSA-N 0.000 description 1
- 230000035619 diuresis Effects 0.000 description 1
- 201000006549 dyspepsia Diseases 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 230000004438 eyesight Effects 0.000 description 1
- 239000012091 fetal bovine serum Substances 0.000 description 1
- 230000002496 gastric effect Effects 0.000 description 1
- 210000001156 gastric mucosa Anatomy 0.000 description 1
- 229930182478 glucoside Natural products 0.000 description 1
- 150000008131 glucosides Chemical class 0.000 description 1
- 229940091561 guaiac Drugs 0.000 description 1
- 229960001867 guaiacol Drugs 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 238000003919 heteronuclear multiple bond coherence Methods 0.000 description 1
- 239000003864 humus Substances 0.000 description 1
- 229930195733 hydrocarbon Natural products 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 230000002147 killing effect Effects 0.000 description 1
- 238000012417 linear regression Methods 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
- 125000000325 methylidene group Chemical group [H]C([H])=* 0.000 description 1
- SNVLJLYUUXKWOJ-UHFFFAOYSA-N methylidenecarbene Chemical compound C=[C] SNVLJLYUUXKWOJ-UHFFFAOYSA-N 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 201000009240 nasopharyngitis Diseases 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000000425 proton nuclear magnetic resonance spectrum Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 201000003068 rheumatic fever Diseases 0.000 description 1
- 238000007873 sieving Methods 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- 238000002495 two-dimensional nuclear magnetic resonance spectrum Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H15/00—Compounds containing hydrocarbon or substituted hydrocarbon radicals directly attached to hetero atoms of saccharide radicals
- C07H15/18—Acyclic radicals, substituted by carbocyclic rings
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
- C07H1/06—Separation; Purification
- C07H1/08—Separation; Purification from natural products
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
- C07B2200/00—Indexing scheme relating to specific properties of organic compounds
- C07B2200/07—Optical isomers
-
- 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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- 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)
Abstract
The invention belongs to the technical field of medicines, and relates to a guaiane type sesquiterpene glycoside compound guaiane type guaiane glycoside II extracted and separated from rhizoma atractylodis rhizome by utilizing the technologies of macroporous resin, normal phase silica gel, reverse phase silica gel, sephadex column chromatography, recrystallization and the like, wherein the molecular formula of the compound is C 21 H 36 O 8 In vitro anti-tumor activity research shows that the anti-tumor compound has good anti-tumor activity, and has obvious inhibition effects on human gastric cancer cell BGC-823, human liver cancer cell HepG-2 and human lung cancer cell A549. The invention can provide a basic source of drug effect substances for the development of anti-tumor drugs and has development and application prospects.
Description
Technical Field
The invention particularly relates to a guaiane type sesquiterpene glycoside compound with a tumor cell inhibition effect.
Background
Guanzhu Zhu (rhizoma Atractylodis Guanzhu)Atractylodes japonicaKoidz.ex kitam.) is chrysanthemumThe dry root and stem of the herbaceous plant is used as a medicine, and the wild resources are rich in northeast China, and the wild woody soil with rich humus grows on the wild forest margin with the altitude of 200-800 m, under the forest, hillside fields, terraced fields and the like. The rhizoma atractylodis has the effects of strengthening spleen, eliminating dampness, drying soil, promoting diuresis, improving eyesight, dispelling wind and avoiding dirty qi, and is mainly used for treating various clinical symptoms such as inappetence, gastric dyspepsia, rheumatic arthritis, common cold and the like. Modern pharmacological research shows that the traditional Chinese medicine composition has great development potential in the aspects of resisting tumor, resisting inflammation, resisting bacteria, protecting and repairing gastric mucosa tissues and the like. In both countries of Japanese and Korean, the rhizome of Atractylodes lancea is not only used as a medicine for the Atractylodes macrocephala, but also used as a health food for eating.
The volatile oil component in rhizoma atractylodis is mainly sesquiterpene hydrocarbons, the main types of the volatile oil component are eucalyptus type, guaiane type and the like from the structural classification, some representative compounds are subjected to related pharmacological activity researches, for example, atractylone has the effects of protecting liver, resisting viruses and eliminating inflammation, apiadione can inhibit capillary permeability from being hyperfunctional and eliminate inflammation, atractylone I has the effects of improving amylase activity, enhancing intestinal digestion and absorption functions and regulating spleen and stomach, atractylone II has an inhibitory effect on human colorectal cancer cells Lovo, atractylone III also has an antiviral effect, and the number of reported compounds of the components in rhizoma atractylodis is small.
Disclosure of Invention
The invention aims to provide a novel guaiane type sesquiterpene glycoside compound and a preparation method and application thereof.
In order to achieve the above purpose, the invention is realized by adopting the following technical scheme:
the invention discloses a guaiane type sesquiterpene glycoside, which has a molecular formula as follows: c (C) 21 H 36 O 8 The common name is guaiacol II, and the structural formula is as follows:
。
the invention also provides a preparation method of the atractylis ovata guaiac glucoside II: is obtained through macroporous resin, normal phase silica gel, reverse phase silica gel and sephadex column chromatography and recrystallization treatment, and the specific preparation steps are as follows:
(1) Alcohol extraction: taking dry rhizome of rhizoma atractylodis as a raw material, properly crushing, extracting for 3 times by adopting 70% ethanol under reflux, filtering, combining the 3 filtrates, recovering the solvent under reduced pressure, and drying to obtain an extractum extract;
(2) Enriching and purifying: dispersing the extractum extract obtained in the step (1) with water to obtain a solution with the relative density of 1.25+/-0.05 g/mL, enriching and purifying by using AB-8 type macroporous resin column chromatography, eluting with water, 30% ethanol, 50% ethanol and 95% ethanol in sequence respectively, collecting 50% ethanol eluent, and recovering the solvent under reduced pressure to obtain a 50% ethanol elution part;
(3) Normal phase silica gel column chromatography: taking a 50% ethanol elution part obtained in the step (2), adopting a normal phase silica gel column, sequentially adopting a mixed solution of dichloromethane and methanol with the volume ratio of 8:1, a mixed solution of dichloromethane and methanol with the volume ratio of 5:1, and a mixed solution of dichloromethane and methanol with the volume ratio of 3:1 to perform systematic gradient elution, collecting distillate with the volume ratio of 3:1, and recovering the solvent under reduced pressure;
(4) Reversed phase silica gel column chromatography: eluting the fraction prepared in the step (3) by reverse phase silica gel ODS column chromatography sequentially with a mixed solution of methanol and water in a volume ratio of 1:2, a mixed solution of methanol and water in a volume ratio of 1:1 and a mixed solution of methanol and water in a volume ratio of 2:1, collecting eluting parts of the mixed solution of methanol and water in a volume ratio of 1:1, and merging the eluting parts after the eluting parts are detected by reverse phase silica gel thin layer chromatography, so as to sequentially obtain three parts Fr.1, F.2 and Fr.3;
(5) Sephadex column chromatography: taking the Fr.2 part obtained in the step (4), performing isocratic elution by using methanol-water with a volume ratio of 1:1.5 as a mobile phase through a sephadex column, collecting eluent, and recovering a solvent to obtain a crude product;
(6) And (3) recrystallizing: recrystallizing the crude product obtained in the step (5) twice by using a two-phase solvent system of methanol and dichloromethane=5:1 to obtain the pure product of the compound.
The invention also provides application of the atractylis ovata guaiana glycoside II in preparing medicines for preventing and treating cancers. Preferably in the preparation of medicines for preventing and treating gastric cancer, liver cancer and lung cancer.
The beneficial effects and significance of the invention are as follows: the research is to carry out intensive ingredient research and development on rhizome of rhizoma atractylodis, find out a compound with a unique chemical structure and stronger activity, and provide a novel anti-tumor medicament for clinical research.
Drawings
FIG. 1 is a chemical formula of a compound of the present invention;
FIG. 2 is a positive HR-ESI-MS spectrum of a compound of the present invention;
FIG. 3 shows a compound of the invention 1 H-NMR spectrum;
FIG. 4 shows a compound of the invention 13 C-NMR spectrum;
FIG. 5 is a DEPT spectrum of a compound of the present invention;
FIG. 6 is a HSQC spectrum of a compound of the present invention;
FIG. 7 is a HMBC pattern of a compound of the invention;
FIG. 8A compound of the invention 1 H- 1 H COSY spectrogram
FIG. 9 is a NOESY spectrum of a compound of the present invention.
Description of the embodiments
Other embodiments of the present invention will be apparent to those skilled in the art from consideration of the present disclosure, which are set forth by way of illustration only, and various modifications and improvements can be made to the present invention without departing from the spirit and scope of the invention, which is intended to be within the scope of the invention as defined by the following detailed description of the examples.
Examples
The preparation method of the compound comprises the following steps:
(1) Alcohol extraction: taking dry rhizome 10 kg of rhizoma atractylodis as a raw material, appropriately crushing the dry rhizome 10 kg of rhizoma atractylodis into coarse powder, sieving the coarse powder with a No. 1 sieve, carrying out reflux extraction for 3 times by adopting 70% ethanol for 2 h each time, wherein the weight-ethanol volume ratio of the raw materials is 1:6, filtering after extraction, merging the 3 times of filtrate, recovering the solvent under reduced pressure, and drying to obtain an extractum-like extract 2.4 kg;
(2) Enriching and purifying: dispersing the extract obtained in the step (1) with water to obtain a solution with a relative density of 1.25+/-0.05 g/mL, enriching and purifying by AB-8 type macroporous resin column chromatography (the inner diameter of the chromatographic column is 10 cm, the length is 1.80 m, the effective height of the resin is 1.35 m), eluting 3 column volumes, 2 column volumes, 3 column volumes and 2 column volumes with water, 30% ethanol and 50% ethanol respectively, collecting 50% ethanol eluent, and recovering the solvent under reduced pressure to obtain 50% ethanol eluting part 386 g;
(3) Normal phase silica gel column chromatography: taking a 50% ethanol elution part obtained in the step (2), adopting a normal phase silica gel column (the inner diameter of the chromatographic column is 6.5 and cm, the length of the chromatographic column is 1.8 and m, the effective height of silica gel is 1.2 and m), sequentially adopting a mixed solution of dichloromethane and methanol with the volume ratio of 8:1 to elute 3 column volumes, a mixed solution of dichloromethane and methanol with the volume ratio of 5:1 to elute 2.5 column volumes, a mixed solution of dichloromethane and methanol with the volume ratio of 3:1 to elute 3 column volumes, collecting distillate with the volume ratio of 3:1, and recovering solvent under reduced pressure to obtain a collected fraction 36g;
(4) Reversed phase silica gel column chromatography: taking the fraction prepared in the step (3), eluting 3 column volumes by using a methanol and water mixed solution with the volume ratio of 1:2 and eluting 2.5 column volumes by using a methanol and water mixed solution with the volume ratio of 1:1 in sequence by using reverse phase silica gel ODS column chromatography (chromatographic column with the inner diameter of 5 cm and the length of 1.5 m, wherein the effective height of reverse phase silica gel is 0.8 m), collecting eluting parts of the methanol and water mixed solution with the volume ratio of 1:1 by using reverse phase silica gel thin layer chromatography, combining the eluting parts of the methanol and water mixed solution with the volume ratio of 1:1, and sequentially obtaining three parts of Fr.1, F.2 and Fr.3;
(5) Sephadex column chromatography: taking the Fr.2 part obtained in the step (4), performing isocratic elution by using methanol-water with a volume ratio of 1:1.5 as a mobile phase through a sephadex column, eluting 4 column volumes, discarding the eluent with the first 2 column volumes, collecting the eluent with the second 2 column volumes, and recovering the solvent to obtain a crude product 42.5 mg;
(6) And (3) recrystallizing: recrystallizing the crude product obtained in the step (5) twice by using a two-phase solvent system of methanol and dichloromethane=5:1 to obtain the pure product of the compound 12.2 mg.
Examples
Identification of the Compounds: the compound of the invention is yellow flaky crystal (methanol: dichloromethane=5:1), the Molish reaction is positive, and D-glucose, [ alpha ] is detected by acid hydrolysis thin layer]25d+26 ° (c=0.15 MeOH). In the case of the HR-ESI-MS,m/zvisible [ M+NH ] at 434.27472 4 ] + The ion peak (calculated for 434.2754) indicates that the molecular weight of the compound of the invention is 416. Bonding of 1 H-NMR、 13 The molecular formula of the compound is estimated to be C by C-NMR, DEPT spectrum and the like 21 H 36 O 8 The unsaturation was calculated to be 4.
At the position of 1 H-NMR (600 MHz, Pyridine-d 5 ) In the spectrum, 4 methyl proton signals were observedδ H 1.46 (6H, s, H-12, H-13)、1.28 (3H, s, H-14)、1.11 (3H, d, J=6.6 Hz, H-15), 4 methylene proton signalsδ H 2.51 (1H, dd, J = 18.6, 9.6 Hz, H-2α)、2.98 (1H, dd, J = 18.6, 9.6 Hz, H-2β)、1.19 (1H, m, H-6α)、1.33 (1H, m, H-6β)、2.41 (1H, m, H-8α)、1.52 (1H, m, H-8β)、2.12 (1H, m, H-9α)、1.95 (1H, o, H-9β) 4 methine proton signalsδ H 1.93 (1H, t, J = 9.6 Hz, H-1)、1.87 (1H, m, H-4)、1.97 (1H, o, H-5)、2.02 (1H, mH-7) and a set of sugar proton signalsδ H 5.06 (1H, d, J = 7.8 Hz, H-1')、4.01 (1H, t, J = 8.0 Hz, H-2')、4.26 (1H, m, H-3')、4.23 (1H, m, H-4')、3.94 (1H, m, H-5')、4.35 (1H, m, H-6'α)、4.52 (1H, dd, J = 2.4, 17.1 Hz, H-6'β) Whereinδ H 5.06 (1H, d, J=8.0 Hz, H-1') is the terminal proton signal and indicates that the sugar isβConfiguration.
At the position of 13 C-NMR (150 MHz, Pyridine-d 5 ) In the spectrum, 21 carbon signals were observed, whichIn 1 carbonyl carbon signalδ C 219.2,4 methyl carbon signalsδ C 23.8, 24.5, 30.7, 12.8,5 methylene carbon signalsδ C 39.2, 36.3, 23.0, 42.5, 63.0,9 methine carbon signalsδ C 50.9, 44.0, 52.5, 50.3, 98.6, 75.4, 78.9, 71.9, 78.1,2 quaternary carbon signalsδ C 71.4, 80.4, whereinδ C 98.6, 75.4, 78.9, 71.9, 78.1, 63.0 are 6 carbon on sugar signals.
At the position of 1 H- 1 In the H COSY spectrum, H 2 -2 is related to H-1, H-4 is related to H 3 15-related, H-5 and H 2 -6 correlation, H 2 -6 is related to H-7, H-7 is related to H 2 -8 correlation, H 2 -8 and H 2 -9 correlation, deducing the presence of fragments in the parent core structure "-CH-CH 3 - "-CH2-CH-" and "-CH 2-"; h-1 'is related to H-2', H-2 'is related to H-3', H-4 'is related to H-5', H-5 'is related to H-6', and the presence of "-CH-CH-CH-" and "-CH-CH-CH" in the structure is deduced 2 - ", by 1 H-NMR、 13 The C-NMR and HSQC spectra infer that the fragment is the sugar moiety. The parent nucleus structure of the compounds of the invention and the references [ Kitajima J, kamoshit A, ishikawa T, et al Glycides ofAtractylodes Japonica.Chem Pharm Bull,2003,51(2): 152-157](1S, 4S,5S,7R, 10R) -10,11,14-trihydroxyguai-3-one 11-O-βD-glucopyranoside is highly uniform, with the main difference that there is one fewer hydroxyl group in position 14. The methyl group at position 14 was determined by DEPT and HSQC spectra. In NOESY spectra, H-1βAnd H-6%δ H 1.19 Related signals of Me-14, H-6 #δ H 2.23 Signals related to H-4, H-12/H-13, H-7 and H-12/H-13, H-5 and Me-15 indicate that the H-5, me-14, me-15, C-7 dimethyl methanol moiety isβConfiguration, H-4, H-7 and OH-10 areαConfiguration.
The structure of the compound is shown in figure 1 by combining 2D-NMR spectrum analysis, the main NMR data are shown in table 1, and the SciFinder database search determines that the compound is a novel compound, and the compound is named as Guangzhu guaiang manway glycoside II.
Examples
The MTT (tetrazolium salt) method measures the killing effect of atractylis lancea on three human tumor cells.
(1) Tumor cells: human gastric cancer cell BGC-823, human liver cancer cell HepG-2 and human lung cancer cell A549.
(2) The experimental concrete operation method comprises the following steps: human gastric cancer cell BGC-823, human liver cancer cell HepG-2 and human lung cancer cell A549 tumor cells are respectively cultured in RPMI 1640 matrix (fetal bovine serum containing 10% L-glutamine, 100 mug/mL penicillin, 100 mug/mL streptomycin). Taking the above tumor cells in logarithmic growth phase, adding 0.25% trypsin for digestion to give a concentration of 10X10 4 180 mu L of cell suspension is taken per mL and placed in a 96-well plate, each group is provided with 3 parallel compound holes, blank holes and control holes are additionally arranged, the blank holes are not inoculated with cells, the control holes are culture solutions without medicines, and cisplatin is the most positive medicine group. At 37℃with 5% CO 2 After 24-h culture in an incubator under the condition, the sample to be tested is added into the culture solution, so that the final concentration of the medicine is 5, 10, 20, 40, 80 and 160 mu mol/L. The solution continued to be 5% CO at 37 ℃C 2 Culture 48 h in incubator. After adding 20. Mu.L of 5/mg/mL MTT solution per well and continuing to culture for 4. 4 h, the culture was terminated. The supernatant was carefully aspirated, 150 μl DMSO was added per well, and the mixture was shaken on a shaker for 10min to completely dissolve the crystals. The absorbance (A) of each well was measured at 570nm using a microplate reader to calculate the cell viability inhibition: cell survival inhibition% = [1- (experimental group a-blank group a)/(control group a-blank group a)]X 100%. The data was processed using the SPSS software analysis system.
(3) Experimental results: experimental data of the survival inhibition rate of cisplatin, a positive control drug, on tumor cells at different concentrations of the novel compound are shown in table 2.
Results of the tableDisplay, linear regression calculation IC 50 The value shows that the sesquiterpene compound related by the invention, atractylis ovata guaiana glycoside II has the effect IC on human gastric cancer BGC-823 cells, human liver cancer HepG-2 cells and human lung cancer A549 cells 50 The values are 66.15 + -0.97 μmol/L and 68.06 + -1.11 μmol/L, 43.44+ -1.82 μmol/L, respectively. Cisplatin serving as positive control drug has effects IC on human gastric cancer BGC-823 cells, human liver cancer HepG-2 cells and human lung cancer A549 cells 50 The values were 5.11.+ -. 1.53. Mu. Mol/L, 4.86.+ -. 0.96. Mu. Mol/L and 10.05.+ -. 1.25. Mu. Mol/L, respectively.
In conclusion, the guaiane type sesquiterpene glycoside-rhizoma atractylodis guaiane glycoside II separated from rhizome of rhizoma atractylodis has a prospect of preparing clinical tumor treatment medicines.
Claims (3)
1. A guaiane compound isolated from rhizome of Atractylodes lancea has a molecular formula of C 21 H 36 O 8 The structural formula of the guaiane glycoside II named Guanzhong is as follows:
。
2. a process for the preparation of a compound as claimed in claim 1, wherein: the Guanzhong guaiang mandshurica II is obtained by using the rhizome of Guanzhong atractylodes rhizome as a raw material through macroporous resin, normal-phase silica gel, reverse-phase silica gel and sephadex column chromatography and recrystallization treatment, and the preparation method comprises the following steps:
(1) Alcohol extraction: taking dry rhizome of rhizoma atractylodis as a raw material, properly crushing, extracting for 3 times by adopting 70% ethanol under reflux, filtering, combining the 3 filtrates, recovering the solvent under reduced pressure, and drying to obtain an extractum extract;
(2) Enriching and purifying: dispersing the extractum extract obtained in the step (1) with water to obtain a solution with the relative density of 1.25+/-0.05 g/mL, enriching and purifying by using AB-8 type macroporous resin column chromatography, eluting with water, 30% ethanol, 50% ethanol and 95% ethanol in sequence respectively, collecting 50% ethanol eluent, and recovering the solvent under reduced pressure to obtain a 50% ethanol elution part;
(3) Normal phase silica gel column chromatography: taking a 50% ethanol elution part obtained in the step (2), adopting a normal phase silica gel column, sequentially adopting a mixed solution of dichloromethane and methanol with the volume ratio of 8:1, a mixed solution of dichloromethane and methanol with the volume ratio of 5:1, and a mixed solution of dichloromethane and methanol with the volume ratio of 3:1 to perform systematic gradient elution, collecting distillate with the volume ratio of 3:1, and recovering the solvent under reduced pressure;
(4) Reversed phase silica gel column chromatography: eluting the fraction prepared in the step (3) by reverse phase silica gel ODS column chromatography sequentially with a mixed solution of methanol and water in a volume ratio of 1:2, a mixed solution of methanol and water in a volume ratio of 1:1 and a mixed solution of methanol and water in a volume ratio of 2:1, collecting eluting parts of the mixed solution of methanol and water in a volume ratio of 1:1, and merging the eluting parts after the eluting parts are detected by reverse phase silica gel thin layer chromatography, so as to sequentially obtain three parts Fr.1, F.2 and Fr.3;
(5) Sephadex column chromatography: taking the Fr.2 part obtained in the step (4), performing isocratic elution by using methanol-water with a volume ratio of 1:1.5 as a mobile phase through a sephadex column, collecting eluent, and recovering a solvent to obtain a crude product;
(6) And (3) recrystallizing: recrystallizing the crude product obtained in the step (5) twice by using a two-phase solvent system of methanol and dichloromethane=5:1 to obtain the pure product of the compound.
3. The use of the compound of claim 1 related to atractylis ovata and guaianoside II in the preparation of medicaments for preventing and treating tumors, wherein the tumors are gastric cancer, liver cancer and lung cancer.
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