CN118206451A - Method for extracting taxane compounds from taxus cuspidata and antitumor application - Google Patents
Method for extracting taxane compounds from taxus cuspidata and antitumor application Download PDFInfo
- Publication number
- CN118206451A CN118206451A CN202410394630.5A CN202410394630A CN118206451A CN 118206451 A CN118206451 A CN 118206451A CN 202410394630 A CN202410394630 A CN 202410394630A CN 118206451 A CN118206451 A CN 118206451A
- Authority
- CN
- China
- Prior art keywords
- silica gel
- thin layer
- ethyl acetate
- layer chromatography
- petroleum ether
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 244000162450 Taxus cuspidata Species 0.000 title claims abstract description 18
- 235000009065 Taxus cuspidata Nutrition 0.000 title claims abstract description 18
- DKPFODGZWDEEBT-QFIAKTPHSA-N taxane Chemical class C([C@]1(C)CCC[C@@H](C)[C@H]1C1)C[C@H]2[C@H](C)CC[C@@H]1C2(C)C DKPFODGZWDEEBT-QFIAKTPHSA-N 0.000 title claims abstract description 17
- 238000000034 method Methods 0.000 title claims abstract description 9
- 230000000259 anti-tumor effect Effects 0.000 title abstract description 8
- 150000001875 compounds Chemical class 0.000 claims abstract description 33
- 239000000126 substance Substances 0.000 claims abstract description 6
- SRCZQMGIVIYBBJ-UHFFFAOYSA-N ethoxyethane;ethyl acetate Chemical compound CCOCC.CCOC(C)=O SRCZQMGIVIYBBJ-UHFFFAOYSA-N 0.000 claims description 28
- 239000003208 petroleum Substances 0.000 claims description 28
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 17
- 239000000741 silica gel Substances 0.000 claims description 17
- 229910002027 silica gel Inorganic materials 0.000 claims description 17
- 238000010898 silica gel chromatography Methods 0.000 claims description 16
- 238000004809 thin layer chromatography Methods 0.000 claims description 16
- 238000001514 detection method Methods 0.000 claims description 15
- 238000010828 elution Methods 0.000 claims description 14
- 238000012746 preparative thin layer chromatography Methods 0.000 claims description 13
- 229940123237 Taxane Drugs 0.000 claims description 12
- -1 cinnamoyl Chemical group 0.000 claims description 11
- 125000003668 acetyloxy group Chemical group [H]C([H])([H])C(=O)O[*] 0.000 claims description 10
- 125000004423 acyloxy group Chemical group 0.000 claims description 10
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 9
- 238000002156 mixing Methods 0.000 claims description 7
- 239000000243 solution Substances 0.000 claims description 7
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 6
- DQXKOHDUMJLXKH-PHEQNACWSA-N (e)-n-[2-[2-[[(e)-oct-2-enoyl]amino]ethyldisulfanyl]ethyl]oct-2-enamide Chemical compound CCCCC\C=C\C(=O)NCCSSCCNC(=O)\C=C\CCCCC DQXKOHDUMJLXKH-PHEQNACWSA-N 0.000 claims description 4
- HTSGKJQDMSTCGS-UHFFFAOYSA-N 1,4-bis(4-chlorophenyl)-2-(4-methylphenyl)sulfonylbutane-1,4-dione Chemical compound C1=CC(C)=CC=C1S(=O)(=O)C(C(=O)C=1C=CC(Cl)=CC=1)CC(=O)C1=CC=C(Cl)C=C1 HTSGKJQDMSTCGS-UHFFFAOYSA-N 0.000 claims description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 4
- NLFBCYMMUAKCPC-KQQUZDAGSA-N ethyl (e)-3-[3-amino-2-cyano-1-[(e)-3-ethoxy-3-oxoprop-1-enyl]sulfanyl-3-oxoprop-1-enyl]sulfanylprop-2-enoate Chemical compound CCOC(=O)\C=C\SC(=C(C#N)C(N)=O)S\C=C\C(=O)OCC NLFBCYMMUAKCPC-KQQUZDAGSA-N 0.000 claims description 4
- 238000000605 extraction Methods 0.000 claims description 4
- 241001116500 Taxus Species 0.000 claims description 3
- WGLUMOCWFMKWIL-UHFFFAOYSA-N dichloromethane;methanol Chemical compound OC.ClCCl WGLUMOCWFMKWIL-UHFFFAOYSA-N 0.000 claims description 3
- 238000010829 isocratic elution Methods 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- 238000002953 preparative HPLC Methods 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- 239000002246 antineoplastic agent Substances 0.000 claims description 2
- 229940041181 antineoplastic drug Drugs 0.000 claims description 2
- 239000007864 aqueous solution Substances 0.000 claims description 2
- 230000006837 decompression Effects 0.000 claims description 2
- 238000001914 filtration Methods 0.000 claims description 2
- 238000004128 high performance liquid chromatography Methods 0.000 claims description 2
- 238000002386 leaching Methods 0.000 claims description 2
- 238000000926 separation method Methods 0.000 claims description 2
- 238000000746 purification Methods 0.000 claims 1
- 238000011160 research Methods 0.000 abstract description 9
- 230000005764 inhibitory process Effects 0.000 abstract description 7
- 210000004881 tumor cell Anatomy 0.000 abstract description 4
- 238000011161 development Methods 0.000 abstract description 2
- 230000000694 effects Effects 0.000 abstract description 2
- 239000002547 new drug Substances 0.000 abstract description 2
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 16
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 15
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 9
- 238000005160 1H NMR spectroscopy Methods 0.000 description 8
- OKKJLVBELUTLKV-MZCSYVLQSA-N Deuterated methanol Chemical compound [2H]OC([2H])([2H])[2H] OKKJLVBELUTLKV-MZCSYVLQSA-N 0.000 description 8
- 239000007787 solid Substances 0.000 description 8
- 239000003814 drug Substances 0.000 description 7
- 210000004027 cell Anatomy 0.000 description 6
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 4
- 229940079593 drug Drugs 0.000 description 4
- 230000002401 inhibitory effect Effects 0.000 description 4
- 239000002398 materia medica Substances 0.000 description 4
- 239000002609 medium Substances 0.000 description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 3
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 3
- 229910052739 hydrogen Inorganic materials 0.000 description 3
- 239000001257 hydrogen Substances 0.000 description 3
- 238000001228 spectrum Methods 0.000 description 3
- 206010006187 Breast cancer Diseases 0.000 description 2
- 208000026310 Breast neoplasm Diseases 0.000 description 2
- 108010087230 Sincalide Proteins 0.000 description 2
- 241001149649 Taxus wallichiana var. chinensis Species 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 238000010609 cell counting kit-8 assay Methods 0.000 description 2
- 229960001592 paclitaxel Drugs 0.000 description 2
- IZTQOLKUZKXIRV-YRVFCXMDSA-N sincalide Chemical compound C([C@@H](C(=O)N[C@@H](CCSC)C(=O)NCC(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(N)=O)NC(=O)[C@@H](N)CC(O)=O)C1=CC=C(OS(O)(=O)=O)C=C1 IZTQOLKUZKXIRV-YRVFCXMDSA-N 0.000 description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 208000032612 Glial tumor Diseases 0.000 description 1
- 206010018338 Glioma Diseases 0.000 description 1
- ZDZOTLJHXYCWBA-VCVYQWHSSA-N N-debenzoyl-N-(tert-butoxycarbonyl)-10-deacetyltaxol Chemical compound O([C@H]1[C@H]2[C@@](C([C@H](O)C3=C(C)[C@@H](OC(=O)[C@H](O)[C@@H](NC(=O)OC(C)(C)C)C=4C=CC=CC=4)C[C@]1(O)C3(C)C)=O)(C)[C@@H](O)C[C@H]1OC[C@]12OC(=O)C)C(=O)C1=CC=CC=C1 ZDZOTLJHXYCWBA-VCVYQWHSSA-N 0.000 description 1
- 229930012538 Paclitaxel Natural products 0.000 description 1
- 108010019160 Pancreatin Proteins 0.000 description 1
- 208000004880 Polyuria Diseases 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 201000009361 ascariasis Diseases 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229940125904 compound 1 Drugs 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 206010012601 diabetes mellitus Diseases 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 230000001079 digestive effect Effects 0.000 description 1
- 230000035619 diuresis Effects 0.000 description 1
- 229960003668 docetaxel Drugs 0.000 description 1
- 230000000857 drug effect Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000012091 fetal bovine serum Substances 0.000 description 1
- 229930003935 flavonoid Natural products 0.000 description 1
- 235000017173 flavonoids Nutrition 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000011389 fruit/vegetable juice Nutrition 0.000 description 1
- 229930182470 glycoside Natural products 0.000 description 1
- 230000005918 in vitro anti-tumor Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 208000017169 kidney disease Diseases 0.000 description 1
- 229930013686 lignan Natural products 0.000 description 1
- 235000009408 lignans Nutrition 0.000 description 1
- 230000027939 micturition Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000010413 mother solution Substances 0.000 description 1
- 229940055695 pancreatin Drugs 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000001953 recrystallisation Methods 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- RCINICONZNJXQF-MZXODVADSA-N taxol Chemical compound O([C@@H]1[C@@]2(C[C@@H](C(C)=C(C2(C)C)[C@H](C([C@]2(C)[C@@H](O)C[C@H]3OC[C@]3([C@H]21)OC(C)=O)=O)OC(=O)C)OC(=O)[C@H](O)[C@@H](NC(=O)C=1C=CC=CC=1)C=1C=CC=CC=1)O)C(=O)C1=CC=CC=C1 RCINICONZNJXQF-MZXODVADSA-N 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention relates to a method for extracting taxane compounds from taxus cuspidata and an anti-tumor application, wherein the method researches the chemical components of the regenerable branches of the taxus cuspidata, separates and identifies a series of taxane compounds, the structure of which is shown as a formula I, a formula II or a formula III, researches the anti-tumor activity of the separated taxane compounds, and the compounds of the invention have an inhibition effect on tumor cells, thus laying a foundation for the deep research of the active components of the taxus cuspidata and the development of new drugs.
Description
Technical Field
The invention belongs to the technical field of natural medicines, and particularly relates to an extraction method and anti-tumor application of a taxane compound in taxus cuspidata.
Background
The taxus cuspidata (Taxus cuspidata Siebold & Zucc) is rich in natural resources in China, and is mainly distributed in Jilin vintage, zhang Ancai and Changbai mountain areas, and also distributed in Japan, korean and Russian. In traditional Chinese medicine, taxus chinensis is recorded in famous medical miscellaneous records, and in famous medical miscellaneous records such as new revised materia medica, materia medica schema, materia medica drawing channel, materia medica Puchen, and Chinese medicine big dictionary. The seeds, bark, stems, leaves and branches can be used as medicines, wherein the medicines used for the branches and leaves are beneficial to promoting urination and dredging channels, inducing diuresis and detumescence, and inhibiting diabetes and treating kidney diseases. In the dictionary of Chinese medicine, taxus chinensis seeds are used for treating food stagnation and expelling ascariasis.
In modern pharmacological research, more than 200 compounds, including taxane compounds, lignan compounds, flavonoid compounds, steroid compounds, glycoside compounds and the like, are separated from different parts of taxus northeast, wherein the taxane compounds are mainly used. The existing mature taxane medicines comprise taxol and docetaxel, and have good anti-tumor activity.
In view of the research on the type and the drug effect of the taxane compounds in the taxus cuspidata, the invention develops related research.
Disclosure of Invention
The invention aims to overcome the defects in the prior art and provide a novel extraction method and anti-tumor activity of the taxane compounds in the taxus cuspidata.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
The first aspect of the invention provides application of a taxane compound in taxus cuspidata in preparation of antitumor drugs, wherein the structure of the taxane compound is shown as a formula I, a formula II or a formula III:
Wherein,
R 1 is selected from H, OH;
r 2 is selected from H, OH, C1-C5 acyloxy, phenoxyacyl;
r 3 is selected from OH, C1-C5 acyloxy, cinnamoyl;
R 4 is selected from OH, C1-C5 acyloxy;
r 5 is selected from OH, C1-C5 acyloxy, phenoxyacyl;
the connecting bond of R6 can be a double bond or a single bond, and when the connecting bond is a double bond, O is formed; when the linkage is a single bond, it is selected from OH, C1-C5 straight or branched acyloxy.
As a further improvement of the present invention, said R 2 is selected from H, OH or acetoxy; the R 3 is selected from OH, acetoxy or cinnamoyl; the R 4 is selected from OH or acetoxy; the R 5 is selected from OH, acetoxy or phenoxyacyl; the bonding bond of R 6 can be a double bond or a single bond, and when the bonding bond is a double bond, O is the bonding bond; when the bond is a single bond, it is selected from OH, acetoxy.
As a further improvement of the present invention, a compound selected from the group consisting of:
the second aspect of the present invention provides a method for extracting the above compound, comprising the steps of:
s1, alcohol extraction and enrichment, namely crushing the reproducible branches of the taxus cuspidata, leaching with an alcohol aqueous solution, filtering the extracting solution, and concentrating under reduced pressure to obtain an enriched substance after concentrating under reduced pressure.
S2, separating and purifying, namely separating the concentrated substance after decompression concentration by a medium-pressure silica gel column, and separating and purifying by using a preparation thin layer chromatography and/or HPLC.
The step of separating and purifying S2 is further improved as follows:
Step one:
Separating by a medium-pressure silica gel column: chromatographic conditions: petroleum ether-ethyl acetate 15% -70% gradient elution is carried out for 600min, and the flow rate is: 50mL/min, collecting each effluent, and combining the effluent according to thin layer chromatography detection to obtain 13 components, which are recorded as components 1-13;
Subjecting the obtained component 4 to preparative thin layer chromatography, eluting with petroleum ether-ethyl acetate (3:1) to obtain four bands, and obtaining compound III from the second band;
step two:
Mixing the component 8 obtained in the first step with 100-200 mesh silica gel, separating by using a medium pressure 300-400 mesh silica gel column chromatography, performing gradient elution for 230min by using 9% -33% of petroleum ether-ethyl acetate, and performing isocratic elution for 15min by using 50% of petroleum ether-ethyl acetate at the flow rate: 35mL/min, collecting each effluent, and combining the effluent according to thin layer chromatography detection to obtain 9 components, namely 8-1 to 8-9;
The obtained component 8-5 is subjected to thin layer chromatography dichloromethane-methanol (9:1) to obtain five bands, and the third band is used for obtaining a compound I-4;
Step three:
the component 8-8 obtained in the second step is separated out, and is recrystallized by acetone after repeated washing to obtain a compound I-1;
And step four:
Mixing the component 11 obtained in the first step with 100-200 mesh silica gel, separating by using a medium pressure 300-400 mesh silica gel column chromatography, and performing gradient elution with 30% -75% of petroleum ether-ethyl acetate for 200min at a flow rate: collecting each effluent at 35mL/min, and combining the effluent according to thin layer chromatography detection to obtain 8 components which are designated as components 11-1 to 11-8;
Subjecting the obtained component 11-3 to preparative thin layer chromatography to obtain six bands of petroleum ether-ethyl acetate (2:3), and obtaining compound I-3 from the fourth band;
step five:
Repeating silica gel column chromatography for the component 11-5 obtained in the fourth step, and performing preparative thin layer chromatography to obtain three bands of petroleum ether-ethyl acetate (2:3), wherein the third band is used for obtaining a compound I-6;
step six:
Selecting the component 11-6 obtained in the step four, and repeating silica gel column chromatography to obtain a compound I-5;
step seven:
Selecting the component 12 obtained in the first step, mixing 100-200 silica gel, separating by using medium pressure silica gel column chromatography, and performing gradient elution in 300-400 meshes of silica gel by using 18% -55% of petroleum ether-ethyl acetate for 150min at the flow rate: 35mL/min, collecting each effluent, and combining the effluent according to thin layer chromatography detection to obtain 13 components, namely 12-1 to 12-13;
Subjecting the obtained component 12-3 to preparative thin layer chromatography to obtain six bands of petroleum ether-ethyl acetate (2:3), and obtaining compound I-2 from the second band;
step eight:
And (3) performing preparative HPLC on the component 12-8 obtained in the step seven, wherein the chromatographic condition is that acetonitrile is 45% -70%, gradient elution is performed for 30min, the flow rate is 4mL/min, the detection wavelength is 254nm, and the compound II is obtained, and t R =1.5 min.
The beneficial effects of adopting above-mentioned technical scheme to produce lie in:
The invention researches the chemical components of the reproducible branches of the taxus northeast, separates and identifies a series of taxane compounds.
The invention carries out anti-tumor activity research on the separated taxane compounds, and the proved compounds have the inhibition effect on tumor cells, thus laying a foundation for deep research on the active ingredients of taxus cuspidata and development of new drugs.
Drawings
FIG. 1 inhibition of proliferation of MCF-7 tumor cell lines by example 8 of the present invention;
FIG. 2 inhibition of MDA-MB-453 cell lines by example 8 of the present invention.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention will be more fully described below with reference to the specific embodiments.
Unless otherwise indicated, the equipment used in this embodiment is conventional, the reagents used are conventional, all commercially available, and the methods of operation used are described in textbooks in the field.
Example 1
95G of the taxus cuspidata renewable small branch extract is separated by medium-pressure silica gel column chromatography, petroleum ether-ethyl acetate 15% -70% gradient elution is carried out for 600min, and the flow rate is that: 50mL/min, collecting the effluent, and combining the effluent according to thin layer chromatography detection to obtain 13 components Fr.1-Fr.13.
Fr.4 (14.2 mg) was subjected to preparative thin layer chromatography, petroleum ether-ethyl acetate (3:1) to give four bands, and compound III (5 mg) was obtained from the second band.
The mother solution of Fr.8 is about 8.4g, 4g of sample-mixing silica gel (100-200 meshes), 160g of separating silica gel (300-400 meshes) by using medium-pressure silica gel column chromatography, gradient elution of 9% -33% of petroleum ether-ethyl acetate for 230min, isocratic elution of 50% of petroleum ether-ethyl acetate for 15min and flow rate: and (3) collecting each effluent at 35mL/min, combining the effluent according to thin layer chromatography detection to obtain 9 components, and recording the components as Fr.8-1-Fr.8-9.
Fr.8-5 by preparative thin layer chromatography dichloromethane-methanol (9:1) gave five bands, compound 6 (17 mg) from the third band; fr.8- (8) had crystals precipitated, and compound I-1 (4 mg) was obtained by repeated washing and acetone recrystallization.
Fr.11 is stirred on 100-200 meshes of silica gel, is separated by medium-pressure 300-400 meshes of silica gel column chromatography, and is eluted by 30% -75% of petroleum ether-ethyl acetate gradient for 200min with the flow rate: collecting each effluent at 35mL/min, and combining the effluent according to thin layer chromatography detection to obtain 8 components (Fr.11 (1-8);
Fr.11-3 by preparative thin layer chromatography, petroleum ether-ethyl acetate (2:3) gave six bands, from the fourth band compound I-3 (8.6 mg). Fr.11-5 after repeated silica gel column chromatography, three bands were obtained by preparative thin layer chromatography, petroleum ether-ethyl acetate (2:3), and Compound I-6 (9.2 mg) was obtained from the third band. Fr.11-6 was subjected to silica gel column chromatography twice, and eluted with petroleum ether-ethyl acetate (1:3) to give Compound I-5 (8.8 mg).
Fr.12 about 6.3g, 6g of sample-mixed silica gel (100-200 meshes), separation by medium pressure silica gel column chromatography, 150g of separated silica gel (300-400 meshes), petroleum ether-ethyl acetate 18% -55% gradient elution for 150min, flow rate: and (3) collecting each effluent at 35mL/min, combining the effluent according to thin layer chromatography detection to obtain 13 components, and recording the components as Fr.12-1 to Fr.12-13.Fr.12-3 by preparative thin layer chromatography, petroleum ether-ethyl acetate (2:3) gave six bands, from the second band compound I-2 (9.7 mg). Fr.12-8 is subjected to preparative HPLC, gradient elution is carried out for 30min under the chromatographic condition of 45% -70% acetonitrile, the flow rate is 4mL/min, the detection wavelength is 254nm, the compound II (1.9 mg, tR=1.5 min) is obtained,
The morphology and structure of the obtained novel taxane compounds are characterized as follows:
compound I-1 9-Deacetyltaxol A
White powdery solid (methanol) with molecular formulas C24H34O7.1H-NMR(600MHz,CD3OD)δ：1.89(1H,d,J＝7.8Hz,H-1),5.58(1H,dd,J＝6.0,2.4Hz,H-2),3.63(1H,d,J＝5.6Hz,H-3),4.12(1H,s,H-5),1.87(1H,m,H-6α),1.76(1H,m,H-6β),1.63(2H,m,H-7),6.06(1H,d,J＝10.4Hz,H-9),5.89(1H,d,J＝10.4Hz,H-10),2.79(1H,dd,J＝19.0,6.2Hz,H-14α),2.44(1H,d,J＝19.8Hz,H-14β),1.11(3H,s,H-16),1.73(3H,s,H-17),2.20(3H,s,H-18),0.89(3H,s,H-19),4.78(1H,s,H-20a),5.18(3H,s,H-20b),2.06,2.09( 3H,s,OCOCH3×2).13C-NMR(150MHz,CD3OD)δ：48.4(C-1),69.9(C-2),40.9(C-3),147.2(C-4),75.4(C-5),30.3(C-6),24.4(C-7),44.5(C-8),73.2(C-9),76.0(C-10),149.7(C-11),138.1(C-12),200.8(C-13),35.7(C-14),37.6(C-15),36.1(C-16),26.4(C-17),13.0(C-18),16.6(C-19),113.1(C-20),20.0,19.5,170.2,170.9(OCOCH3×2).
Compound I-2 1-hydroxy-taxol A
White powdery solid (dichloromethane) with a molecular formula of C26H36O9.1H-NMR(600MHz,CDCl3)δ：5.61(1H,d,J＝7.5Hz,H-2),3.74(1H,d,J＝6.4Hz,H-3),4.18(1H,s,H-5),1.75(1H,m,H-6α),1.63(1H,m,H-6β),1.84(1H,td,J＝13.6,4.7Hz,H-7α),1.59(1H,m,H-7β),5.91(1H,d,J＝10.3Hz,H-9),6.16(1H,d,J＝10.4Hz,H-10),2.83(1H,d,J＝19.5Hz,H-14α),2.60(1H,d,J＝18.5Hz,H-14β),1.69(3H,s,H-16),1.21(3H,s,H-17),2.23(3H,s,H-18),0.91(3H,s,H-19),4.64(1H,s,H-20a),5.16(3H,s,H-20b),2.07,2.09,2.13( each 3H, s, OCOCH 3 X3. Compounds are known, only hydrogen spectra and literature controls are made to determine structure.
Compound I-3 5 alpha, 9 alpha-dihydroxy-2 alpha, 10 beta, 13 alpha-triacetoxy-4 (20), 11-diene taxane
White powdery solid (methanol) with a molecular formula of C26H38O8. 1H-NMR(600MHz,CDCl3)δ：1.75(1H,m,H-1),5.39(1H,dd,J＝5.8,2.0Hz,H-2),3.51(1H,d,J＝5.7Hz,H-3),4.24(1H,s,H-5),1.76(1H,m,H-6α),1.61(1H,m,H-6β),1.51(1H,m,H-7),4.21(1H,d,J＝9.8Hz,H-9),5.88(1H,d,J＝9.9Hz,H-10),5.75(1H,dd,J＝10.5,5.1Hz,H-13),2.64(1H,ddd,J＝15.6,10.5,8.8Hz,H-14α),1.53(1H,dd,J＝15.8,5.3Hz,H-14β),1.02(3H,s,H-16),1.59(3H,s,H-17),2.14(3H,s,H-18),1.05(3H,s,H-19),5.22(1H,br.s,H-20a),4.89(1H,s,H-20b),2.10,2.09,2.05( each 3H, s, OCOCH 3 X3.
Compound I-4 9-Deacetyltaxol E
White powdery solid (methanol), the known compounds of the formula C35H44O9. 1H-NMR(600MHz,CDCl3)δ：1.73(1H,m,H-1),5.42(1H,dd,J＝6.1,2.2Hz,H-2),3.34(1H,d,J＝6.1Hz,H-3),5.47(1H,br.s,H-5),1.88(1H,ddd,J＝14.4,4.5,2.4Hz,H-6α),1.82(dd,J＝8.6,2.1Hz,1H,H-6β),1.95(1H,ddd,J＝13.4,4.5,2.4Hz,H-7α),1.66(1H,m,H-7β),4.34(d,J＝10.0Hz,1H,H-9),5.86(1H,d,J＝10.1Hz,H-10),5.78(1H,t,J＝6.0Hz,H-13),2.65(1H,dt,J＝15.1,9.4Hz,H-14α),1.47(1H,dd,J＝15.2,6.9Hz,H-14β),1.10(3H,s,H-16),1.63(3H,s,CH3-17),2.31(s,3H,CH3-18),1.09(3H,s,H-19),5.38(1H,s,H-20a),5.01(1H,s,H-20b),2.10,2.03,1.78( each 3H,s,OCOCH3×3),6.68(1H,d,J＝16.1Hz,H-22),7.77(1H,d,J＝16.0Hz,H-23),7.51(2H,m,H-25),7.40(3H,m,H-26,27). were identified, and only the hydrogen spectrum and literature controls were used to identify the structure.
Compound I-5 5 alpha, 13 alpha diacetoxy-9 alpha, 10 beta dihydroxy-4 (20), 11-diene taxane
White powdery solid (dichloromethane) with molecular formula C24H36O6.1H-NMR(600MHz,CDCl3)δ：1.82(1H,m,H-1),1.70(1H,m,H-2),2.94(1H,d,J＝8.0Hz,H-3),5.35(1H,s,H-5),1.78-1.80(2H,m,H-6),1.75(1H,m,H-7),1.55(1H,m,H-7),4.14(1H,d,J＝9.6Hz,H-9),4.80(1H,d,J＝9.6Hz,H-10),5.89(1H,d,J＝8.0Hz,H-13),1.07(1H,dd,J＝15.2,8.3Hz,H-14α),2.66(1H,dt,J＝14.4,9.8Hz,H-14β),1.52(3H,s,H-16),1.17(3H,s,H-17),1.95(3H,d,J＝1.5Hz,H-18),0.91(3H,s,H-19),5.19(1H,s,H-20a),4.82(3H,s,H-20b),2.15,2.07( 3H,s,5,13-COCH3).13C-NMR(150MHz,CDCl3)δ：40.5(C-1),28.2(C-2),38.0(C-3),149.4(C-4),76.7(C-5),27.6(C-6),26.1(C-7),42.8(C-8),78.7(C-9),72.0(C-10),138.8(C-11),133.8(C-12),71.0(C-13),31.9(C-14),39.4(C-15),27.3(C-16),31.5(C-17),14.9(C-18),18.1(C-19),113.5(C-20),21.8,21.4,170.0,170.6(OCOCH3×2).
Compound I-6 2 alpha, 9 alpha, 10 beta-triacetoxy-5 alpha, 13 alpha-dihydroxy-4 (20), 11-diene taxane
White powdery solid (methanol) with a molecular formula of C26H38O8.1H-NMR(600MHz,CDCl3)δ：1.75(1H,m,H-1),5.42(1H,dd,J＝5.9,1.9Hz,H-2),3.46(1H,d,J＝5.7Hz,H-3),4.24(1H,s,H-5),1.68(1H,m,H-6α),1.93(1H,m,H-7),5.76(1H,d,J＝10.2Hz,H-9),6.05(1H,d,J＝10.3Hz,H-10),4.35(1H,dd,J＝8.9,4.4Hz,H-13),2.70(1H,m,H-14α),1.58(1H,dd,J＝16.1,4.5Hz,H-14β),1.66(3H,s,H-16),0.93(3H,s,H-17),2.26(3H,s,H-18),0.85(3H,s,H-19),4.84(1H,br.s,H-20a),5.21(1H,br.s,H-20b),2.01,2.04,2.05( each 3H, s, OCOCH 3 X3. Compounds are known, only hydrogen spectra and literature controls are made to determine structure.
Compound II 2 alpha, 5 alpha, 13 alpha-triacetoxy-7 beta-hydroxy-9, 10-dione-2 (3.fwdarw.20) rearranged taxane
White powdery solid (dichloromethane) with molecular formula C26H34O9.1H-NMR(600MHz,CD3OD)δ：1.47(1H,dd,J＝12.1,2.5Hz,H-1),5.62(1H,d,J＝7.0Hz,H-2),5.88(1H,d,J＝9.1Hz,H-3),5.63(1H,s,H-5),1.98(1H,dd,J＝14.1,1.4Hz,H-6α),2.18(1H,ddd,J＝14.1,5.7,3.4Hz,H-6β),4.21(2H,dd,J＝12.5,5.5Hz,H-7),5.37(1H,d,J＝9.3Hz,H-13),2.70(1H,m,H-14α),2.43(1H,dd,J＝15.6,1,8Hz,H-14β),1.18(3H,s,H-16),1.17(3H,s,H-17),1.70(3H,s,H-18),1.94(1H,m,H-19a),2.74(1H,d,J＝8.2Hz,H-19b),1.51(3H,s,H-20),2.11,2.01,2.12( 3H,s,2,5,14-OCOCH3).13C-NMR(150MHz,CD3OD)δ：48.1(C-1),72.5(C-2),122.1(C-3),140.9(C-4),71.3(C-5),38.0(C-6),66.8(C-7),54.2(C-8),201.3(C-9),211.0(C-10),137.5(C-11),148.4(C-12),68.6(C-13),27.9(C-14),37.5(C-15),33.0(C-16),24.5(C-17),18.8(C-18),32.2(C-19),26.0(C-20),21.2,20.8,20.8,172.1,171.7,171.7(OCOCH3×3).
Compound III 10 beta-benzoyloxy-1 beta, 5 alpha-dihydroxy-7 beta, 9 alpha, 13 alpha-triacetoxy-4 (20), 11-diene-11 (15- > 1) rearranged taxane
Colorless amorphous solid (methanol) of molecular formula C33H42O10.1H-NMR(600MHz,CDCl3)δ：1.48(1H,d,J＝14.2Hz,H-2),2.42(1H,dd,J＝14.1,9.4Hz,H-2),2.91(1H,d,J＝9.4Hz,H-3),4.37(1H,dd,J＝3.8,1.6Hz,H-5),1.98(1H,ddd,J＝14.2,5.3,1.6Hz,H-6α),1.84(1H,ddd,J＝14.2,11.3,3.8Hz,H-6β),5.67(1H,dd,J＝11.3,5.3Hz,H-7),6.06(1H,s,H-9),6.64(1H,d,J＝10.4Hz,H-10),5.46(1H,t,J＝7.3Hz,H-13),1.31(1H,dd,J＝14.1,5.0Hz,H-14α),2.52(1H,dd,J＝14.1,7.4Hz,H-14β),1.07(3H,s,H-16),1.35(3H,s,H-17),2.03(3H,s,H-18),0.88(1H,m,H-19),4.80(1H,s,H-20a),5.15(1H,s,H-20b),2.06,1.76,2.05( 3H,s,7,9,13-OCOCH3).13C-NMR(150MHz,CDCl3)δ：63.3(C-1),29.2(C-2),37.6(C-3),150.3(C-4),72.5(C-5),36.0(C-6),69.7(C-7),45.1(C-8),77.2(C-9),70.1(C-10),136.4(C-11),147.5(C-12),79.7(C-13),44.1(C-14),75.7(C-15),26.9(C-16),24.8(C-17),12.1(C-18),12.8(C-19),111.5(C-20),21.4,20.7,21.1,169.9,170.0,171.0(7,9,13-OCOCH3×3),164.1,129.2,129.5,128.3,133.3(10-OCOPh).
Example 1 of biological Activity
The CCK-8 method is adopted to detect the in vitro anti-tumor activity of the compound. Two breast cancer tumor cell lines MCF-7 and MDA-MB-453 are inoculated into RPMI 1640 culture solution containing 10% fetal bovine serum, cultured at 37 ℃ under the condition of 5% CO 2 saturated humidity, and subcultured after digestion of pancreatin digestive juice, and cells in logarithmic growth phase are adopted for experiments. All compounds to be tested were prepared with DMSO at a concentration of 10M, the cells were resuspended individually, and then seeded in 96-well plates at 5X 10 3 cells per well, 10. Mu.L of CCK-8 solution per well was added and incubation was continued at 37℃for 48h, and absorbance at 450nm was measured using a microplate reader. Inhibition (%) = (OD control-OD experimental)/(OD control-OD blank) ×100%.
The test results show that the compound has inhibitory activity on two human breast cancer cells (MCF-7, MDA-MB-453) and human glioma cells (U251) at the concentration of 10M. Compound 1 had moderate inhibitory activity against MCF-7 (see fig. 1) with an inhibition rate of 38.03%; compound 8 has moderate inhibitory activity against MDA-MB-453 (see FIG. 2), with an inhibition rate of 25.78%.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and are not limiting; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit and scope of the technical solutions of the embodiments of the present invention.
Claims (5)
1. The application of taxane compounds in taxus cuspidata in preparing antitumor drugs is characterized in that the taxane compounds have the structure shown in formula I, formula II or formula III:
Wherein,
R 1 is selected from H, OH;
r 2 is selected from H, OH, C1-C5 acyloxy, phenoxyacyl;
r 3 is selected from OH, C1-C5 acyloxy, cinnamoyl;
R 4 is selected from OH, C1-C5 acyloxy;
r 5 is selected from OH, C1-C5 acyloxy, phenoxyacyl;
the connecting bond of R6 can be a double bond or a single bond, and when the connecting bond is a double bond, O is formed; when the linkage is a single bond, it is selected from OH, C1-C5 straight or branched acyloxy.
2. The use of taxanes in taxus cuspidata according to claim 1, wherein R 2 is selected from H, OH or acetoxy; the R 3 is selected from OH, acetoxy or cinnamoyl; the R 4 is selected from OH or acetoxy; the R 5 is selected from OH, acetoxy or phenoxyacyl; the bonding bond of R 6 can be a double bond or a single bond, and when the bonding bond is a double bond, O is the bonding bond; when the bond is a single bond, it is selected from OH, acetoxy.
3. The use of taxanes in taxus cuspidata according to claim 1, wherein formula I is selected from the following compounds:
4. A method for extracting taxanes from taxus cuspidate for use according to claim 3, comprising the steps of:
s1, alcohol extraction and enrichment, namely crushing the reproducible branches of the taxus cuspidata, leaching with an alcohol aqueous solution, filtering the extracting solution, and concentrating under reduced pressure to obtain an enriched substance after concentrating under reduced pressure.
S2, separating and purifying, namely separating the concentrated substance after decompression concentration by a medium-pressure silica gel column, and separating and purifying by using a preparation thin layer chromatography and/or HPLC.
5. The method according to claim 4, wherein the step of S2 separation and purification is selected from any one or more of the following steps:
Step one:
Separating by a medium-pressure silica gel column: chromatographic conditions: petroleum ether-ethyl acetate 15% -70% gradient elution is carried out for 600min, and the flow rate is: 50mL/min, collecting each effluent, and combining the effluent according to thin layer chromatography detection to obtain 13 components, which are recorded as components 1-13;
Subjecting the obtained component 4 to preparative thin layer chromatography, eluting with petroleum ether-ethyl acetate (3:1) to obtain four bands, and obtaining compound III from the second band;
step two:
Mixing the component 8 obtained in the first step with 100-200 mesh silica gel, separating by using a medium pressure 300-400 mesh silica gel column chromatography, performing gradient elution for 230min by using 9% -33% of petroleum ether-ethyl acetate, and performing isocratic elution for 15min by using 50% of petroleum ether-ethyl acetate at the flow rate: 35mL/min, collecting each effluent, and combining the effluent according to thin layer chromatography detection to obtain 9 components, namely 8-1 to 8-9;
The obtained component 8-5 is subjected to thin layer chromatography dichloromethane-methanol (9:1) to obtain five bands, and the third band is used for obtaining a compound I-4;
Step three:
the component 8-8 obtained in the second step is separated out, and is recrystallized by acetone after repeated washing to obtain a compound I-1;
And step four:
Mixing the component 11 obtained in the first step with 100-200 mesh silica gel, separating by using a medium pressure 300-400 mesh silica gel column chromatography, and performing gradient elution with 30% -75% of petroleum ether-ethyl acetate for 200min at a flow rate: collecting each effluent at 35mL/min, and combining the effluent according to thin layer chromatography detection to obtain 8 components which are designated as components 11-1 to 11-8;
Subjecting the obtained component 11-3 to preparative thin layer chromatography to obtain six bands of petroleum ether-ethyl acetate (2:3), and obtaining compound I-3 from the fourth band;
step five:
Repeating silica gel column chromatography for the component 11-5 obtained in the fourth step, and performing preparative thin layer chromatography to obtain three bands of petroleum ether-ethyl acetate (2:3), wherein the third band is used for obtaining a compound I-6;
step six:
Selecting the component 11-6 obtained in the step four, and repeating silica gel column chromatography to obtain a compound I-5;
step seven:
Selecting the component 12 obtained in the first step, mixing 100-200 silica gel, separating by using medium pressure silica gel column chromatography, and performing gradient elution in 300-400 meshes of silica gel by using 18% -55% of petroleum ether-ethyl acetate for 150min at the flow rate: 35mL/min, collecting each effluent, and combining the effluent according to thin layer chromatography detection to obtain 13 components, namely 12-1 to 12-13;
Subjecting the obtained component 12-3 to preparative thin layer chromatography to obtain six bands of petroleum ether-ethyl acetate (2:3), and obtaining compound I-2 from the second band;
step eight:
And (3) performing preparative HPLC on the component 12-8 obtained in the step seven, wherein the chromatographic condition is that acetonitrile is 45% -70%, gradient elution is performed for 30min, the flow rate is 4mL/min, the detection wavelength is 254nm, and the compound II is obtained, and t R =1.5 min.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410394630.5A CN118206451A (en) | 2024-04-02 | 2024-04-02 | Method for extracting taxane compounds from taxus cuspidata and antitumor application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410394630.5A CN118206451A (en) | 2024-04-02 | 2024-04-02 | Method for extracting taxane compounds from taxus cuspidata and antitumor application |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN118206451A true CN118206451A (en) | 2024-06-18 |
Family
ID=91447369
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410394630.5A Pending CN118206451A (en) | 2024-04-02 | 2024-04-02 | Method for extracting taxane compounds from taxus cuspidata and antitumor application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN118206451A (en) |
-
2024
- 2024-04-02 CN CN202410394630.5A patent/CN118206451A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN114031579B (en) | Preparation and application of daphnane diterpenoid compounds in lilac daphne flower buds | |
| CN112479887B (en) | Ester compound in purslane and extraction and separation method and application thereof | |
| CN103088083A (en) | Method for preparing anti-inflammation and anti-tumor activity sesquiterpene lactone compounds | |
| CN107698510A (en) | The alkaloid compound and extracting method extracted from blue or green bamboo mark | |
| CN118206451A (en) | Method for extracting taxane compounds from taxus cuspidata and antitumor application | |
| CN113214214B (en) | Preparation method and application of terpenoid in Atractylodes lancea | |
| CN113278026B (en) | Lignin compound with anti-tumor activity and preparation method and application thereof | |
| CN107474027A (en) | The propionic acid of straw mushroom fructification active component 2 (5H) furanone 4 and application | |
| CN109180632B (en) | A method for preparing compound separated from radix Tripterygii Wilfordii | |
| CN112724113A (en) | Flavonoid compound with anti-tumor activity and preparation method and application thereof | |
| CN114685420B (en) | Compound with anti-tumor activity and preparation method and application thereof | |
| CN118126050B (en) | 6-Methoxy carbazole alkaloid compound and preparation method and application thereof | |
| CN113248384B (en) | Euphorbia lathyris diterpene alkane type compound and preparation method and application thereof | |
| CN113666816B (en) | Peking euphorbia diterpenoid compound with anti-tumor activity and preparation method and application thereof | |
| CN110903269A (en) | Cadinane sesquiterpene compound and preparation method thereof | |
| CN112125804B (en) | Diterpenoid of euphorbia lathyris, preparation method and anti-leukemia application thereof | |
| CN113735698B (en) | Tricyclic diterpenoid compound with antitumor activity, and preparation method and application thereof | |
| CN116903578B (en) | Phenolic acid compound in Glechoma hederacea as well as extraction and separation method and application thereof | |
| CN116217649B (en) | Steroid compound in sophora japonica linn and preparation method and application thereof | |
| CN114573656B (en) | Preparation method and application of polyoxy oleanane type triterpene compound sub-merino triterpene C | |
| CN113234084B (en) | Compound with anti-rotavirus activity in Meadowrue of Crane celebration and preparation method thereof | |
| CN117771237B (en) | Application and preparation method of abietane diterpenoid compound | |
| CN115536532B (en) | Crotonane diterpenoid compound, preparation method, pharmaceutical composition and application thereof | |
| CN117304032B (en) | Steroid sapogenin compound in radix clinopodii, and preparation method and application thereof | |
| CN110305081B (en) | Dihydro agarofuran type sesquiterpene and preparation and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination |