CN109232710A - The preparation method of a kind of special different steroid alkaloid and its derivative - Google Patents
The preparation method of a kind of special different steroid alkaloid and its derivative Download PDFInfo
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- CN109232710A CN109232710A CN201811348938.7A CN201811348938A CN109232710A CN 109232710 A CN109232710 A CN 109232710A CN 201811348938 A CN201811348938 A CN 201811348938A CN 109232710 A CN109232710 A CN 109232710A
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- 229930003352 steroid alkaloid Natural products 0.000 title claims abstract description 13
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- NEMWYOKGHGSVSC-MSSYMPDSSA-N hupehenine Chemical compound C([C@@H]1[C@H](O)C[C@@H]23)[C@@H](O)CC[C@]1(C)[C@H]3C[C@@H]1[C@H]2CC[C@H]2[C@@H](C)[C@@H]3CC[C@H](C)CN3C[C@H]21 NEMWYOKGHGSVSC-MSSYMPDSSA-N 0.000 claims abstract description 9
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- 125000001972 isopentyl group Chemical group [H]C([H])([H])C([H])(C([H])([H])[H])C([H])([H])C([H])([H])* 0.000 description 1
- 125000001449 isopropyl group Chemical group [H]C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 125000005956 isoquinolyl group Chemical group 0.000 description 1
- 125000001786 isothiazolyl group Chemical group 0.000 description 1
- CTAPFRYPJLPFDF-UHFFFAOYSA-N isoxazole Chemical compound C=1C=NOC=1 CTAPFRYPJLPFDF-UHFFFAOYSA-N 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 201000005202 lung cancer Diseases 0.000 description 1
- 208000020816 lung neoplasm Diseases 0.000 description 1
- FBBDOOHMGLLEGJ-UHFFFAOYSA-N methane;hydrochloride Chemical compound C.Cl FBBDOOHMGLLEGJ-UHFFFAOYSA-N 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 230000036457 multidrug resistance Effects 0.000 description 1
- 125000001971 neopentyl group Chemical group [H]C([*])([H])C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- 239000012044 organic layer Substances 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 125000002971 oxazolyl group Chemical group 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 125000004430 oxygen atom Chemical group O* 0.000 description 1
- 230000020477 pH reduction Effects 0.000 description 1
- 125000003538 pentan-3-yl group Chemical group [H]C([H])([H])C([H])([H])C([H])(*)C([H])([H])C([H])([H])[H] 0.000 description 1
- IUKLSMSEHKDIIP-UHFFFAOYSA-N petine Natural products CC1(O)C2CCC3C4CC(O)C5CC(O)CCC5(C)C4CC3C2CN2C1CCC(C)C2 IUKLSMSEHKDIIP-UHFFFAOYSA-N 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 125000001436 propyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 125000003373 pyrazinyl group Chemical group 0.000 description 1
- 125000003226 pyrazolyl group Chemical group 0.000 description 1
- 125000004076 pyridyl group Chemical group 0.000 description 1
- 125000005493 quinolyl group Chemical group 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 125000002914 sec-butyl group Chemical group [H]C([H])([H])C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 125000003548 sec-pentyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- CHQMHPLRPQMAMX-UHFFFAOYSA-L sodium persulfate Substances [Na+].[Na+].[O-]S(=O)(=O)OOS([O-])(=O)=O CHQMHPLRPQMAMX-UHFFFAOYSA-L 0.000 description 1
- 150000003431 steroids Chemical class 0.000 description 1
- 201000011549 stomach cancer Diseases 0.000 description 1
- 238000013456 study Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 125000004434 sulfur atom Chemical group 0.000 description 1
- 229960001603 tamoxifen Drugs 0.000 description 1
- 125000000999 tert-butyl group Chemical group [H]C([H])([H])C(*)(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- 125000001973 tert-pentyl group Chemical group [H]C([H])([H])C([H])([H])C(*)(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- 125000000335 thiazolyl group Chemical group 0.000 description 1
- 125000001544 thienyl group Chemical group 0.000 description 1
- QERYCTSHXKAMIS-UHFFFAOYSA-N thiophene-2-carboxylic acid Chemical compound OC(=O)C1=CC=CS1 QERYCTSHXKAMIS-UHFFFAOYSA-N 0.000 description 1
- 125000001425 triazolyl group Chemical group 0.000 description 1
- 239000000052 vinegar Substances 0.000 description 1
- 235000021419 vinegar Nutrition 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07J—STEROIDS
- C07J71/00—Steroids in which the cyclopenta(a)hydrophenanthrene skeleton is condensed with a heterocyclic ring
- C07J71/0036—Nitrogen-containing hetero ring
- C07J71/0042—Nitrogen only
- C07J71/0052—Nitrogen only at position 16(17)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- 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
- Y02P—CLIMATE CHANGE MITIGATION TECHNOLOGIES IN THE PRODUCTION OR PROCESSING OF GOODS
- Y02P20/00—Technologies relating to chemical industry
- Y02P20/50—Improvements relating to the production of bulk chemicals
- Y02P20/55—Design of synthesis routes, e.g. reducing the use of auxiliary or protecting groups
Landscapes
- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses the preparation methods of a kind of special different steroid alkaloid and its derivative, including then further prepare corresponding derivative with directly extracting separation from Hupeh Fritillary Bulb coarse powder or synthesizing different steroid alkaloid by hupehenine first.Different steroid alkaloid of the invention and its derivative have anti-tumor activity, have certain application prospect.
Description
Technical field
Design medicine chemical field of the present invention, and in particular to the preparation of a kind of special different steroid alkaloid and its derivative
Method.
Background technique
Due to aging of population, environmental pollution, social spirit pressure is big, and the factors such as unsound life style cause me
The Cancer Mortality of state is high.According to the status and trend report of Chinese tumour in 2017 of National Cancer Center publication
Accuse display: China's tumor incidence ranks forefront five successively at present are as follows: lung cancer, gastric cancer, liver cancer, cancer of the esophagus and colorectal cancer.Lung
It is the first that cancer, breast cancer occupy Chinese male respectively, women falls ill.Therefore cancer is seriously threatening the people of the world and Chinese people
Health.Dominant reactive part is alkaloid in Hupeh Fritillary Bulb, and in recent years for its inverse cancer cell multidrug resistance, cholinolytic is living
Property, the correlative studys such as cholinesterase inhibition are more, but anti-tumor aspect research is less.Previous research finds the different steroid in part
Body alkaloid have antitumor activity, as peimine can inhibit MCF-7/TAM (human breast cancer cell of resistance to tamoxifen) and
The proliferation of AML-KG-1a (acute myeloid leukemia cell), and can induce its apoptosis.But its activity is lower, it is right at us
Have found the cis- different steroid alkaloid ingredient of cevine type of a kind of special D/E ring in the research process of Hupeh Fritillary Bulb, it is this kind of at
Divide C-5, C-6 double bond structure with feature, they have apparent anti-tumor activity, we expand research to this, with it
The active material for obtaining more more effectively treatment tumours, provides more effective antitumour drugs for clinical application.
Summary of the invention
The object of the invention is that providing one kind has anti-tumor activity C-5, C-6 double bond structure containing feature, spy simultaneously
The preparation method of the cis- different steroid alkaloid of cevine type of different D/E ring and its derivative.
Technical scheme is as follows:
The preparation method of different steroid alkaloid shown in formula I, includes the following steps:
Hupeh Fritillary Bulb coarse powder solvent extraction is concentrated under reduced pressure to give total medicinal extract, and total medicinal extract is stood after being dispersed with sour water, supernatant
Liquid adjusts pH to more than 9, extract is obtained by extraction with organic solvent, extract is by the isolated Formula Iization
Close object.
The present invention also provides another the method for preparing compound of formula I, include the following steps:
1) 3 hydroxyls of hupehenine (hupehenine) are protected to obtain compound 2 with silicon substrate；
2) dehydration of compound 2 obtains compound 3；
3) compound 3 is deprotected to obtain compound of formula I
Preferably, step 1) includes: hupehenine in the presence of organic base and 4-dimethylaminopyridine
(hupehenine) it is reacted with silated reagent, obtains compound 2, the silated reagent is selected from: tert-butyldimethylsilyl chloride silicon
Alkane, tri isopropyl chlorosilane, chlorotriethyl silane or tert-butyl diphenyl chlorosilane.
Preferably, step 3) is included in hydrochloric acid, hydrofluoric acid, pyridine hydrofluoride, tetrabutyl ammonium fluoride or cesium fluoride
In the presence of be deprotected to obtain compound of formula I.
Preferably, step 2) includes being dissolved in compound 2 and 4-dimethylaminopyridine in dry pyridine and phosphorus oxychloride
Reaction obtains compound 3.
The present invention also provides the preparation methods of Formula II compound, include the following steps:
Compound of formula I and carboxylic acid, acid anhydrides or acyl chloride reaction production II compound,
Wherein, R1It is selected from :-C (O) R2Or-S (O)2R3；
R2Selected from C1-C6Alkyl or optionally by one or more R4Replace following group: C5-C10Heteroaryl or C6-C10Virtue
Base；
R3Selected from C1-C6Alkyl or optionally by one or more R4Substituted C6-C10Aryl；
R4Selected from C1-C6Alkyl or halogen.
Preferably, R2Selected from C1-C6Alkyl, C5-C10Heteroaryl or optionally by one or more R4Substituted C6-C10Virtue
Base.
Preferably, R3Selected from optionally by one or more R4Substituted C6-C10Aryl；It more selects excellent by C1-C6Alkyl or halogen
The phenyl that element replaces.
The present invention also provides the preparation methods of formula III compound, include the following steps:
Compound of formula I and thionyl chloride occur substitution reaction and obtain Formula II compound；
The present invention also provides the preparation methods of Formula V compound, include the following steps:
Compound of formula I reacts production IV compound with paratoluensulfonyl chloride, and formula IV compound reacts life with potassium tert-butoxide
At Formula V compound；
Term definition and explanation:
Unless otherwise indicated, group and the term definition recorded in present specification and claims, including its work
For recorded in the definition of example, illustrative definition, preferred definition, table definition, particular compound determines in embodiment
Justice etc., can any combination and combination each other.Group definition and compound structure after such combination and combination, should
Belong in the range of present specification record.
Term " C1-C6 " is interpreted as the preferred direct-connected or branch saturation monovalent hydrocarbon for indicating to have 1-6 carbon atom, example
Such as methyl, ethyl, propyl, butyl, amyl, hexyl, isopropyl, isobutyl group, sec-butyl, tert-butyl, isopentyl, 2- methyl fourth
Base, 1- methyl butyl, 1- ethyl propyl, 1,2- dimethyl propyl, neopentyl, 1,1- dimethyl propyl, 4- methyl amyl, 3- first
Base amyl, 2- methyl amyl, 1- methyl amyl, 2- ethyl-butyl, 1- ethyl-butyl, 3,3- dimethylbutyl, 2,2- dimethyl
Butyl, 1,1- dimethylbutyl, 2,3- dimethylbutyl, 1,3- dimethylbutyl or 1,2- dimethylbutyl etc. or theirs is different
Structure body.
Term " C6-C10Aryl " refers to the group of the derivative containing 6 to 10 carbon atoms to aromatic hydrocarbons.The example of such group
Including but not limited to phenyl, benzyl, naphthalene.
Term " C5-C10Heteroaryl ", which refers to, to be contained 5 to 10 carbon atoms in its ring and contains 1 to 4 each independently
Heteroatomic aromatic heterocyclic group selected from O, S and N.Condition is on the ring of the group without two adjacent O atoms or two
A adjacent S atom.The heterocyclic group includes fused benzo ring system.C5-C10The example of heteroaryl include but is not limited to pyridyl group,
Imidazole radicals, furyl, pyrimidine radicals, pyrazolyl, triazolyl, pyrazinyl, tetrazole radical, furyl, thienyl, oxazolyl, isoxazole
Base, thiazolyl, isothiazolyl, pyrrole radicals, quinolyl, isoquinolyl, indyl, benzimidazolyl, benzofuranyl, diaza
Naphthalene, isoindolyl, purine radicals, benzothienyl, benzothiazolyl.
Specific embodiment
The preparation of embodiment 1:CW-1
1) after dry Hupeh Fritillary Bulb (Fritillariahupehensis Hsiao et K.C.) being broken into coarse powder,
80% ethyl alcohol heating and refluxing extraction 2 times, each 3h.It is concentrated under reduced pressure after extracting solution twice is merged, obtains total medicinal extract；
2) 3% hydrochloric acid solution is first added into total medicinal extract makes its acidification, stands after being sufficiently stirred, takes supernatant, ammonium hydroxide
Alkalization to pH value is greater than 9.Petroleum ether (10L × 3) after being saturated with water, petroleum ether/ethyl acetate (1:1,10L × 3) and vinegar
Acetoacetic ester (10L × 3) is to lye successively stage extraction；
3) alkaloid of water saturated petroleum ether portion passes through purification on normal-phase silica gel with cyclohexane/ethyl acetate/diethylamine
Chromatographic column isolates and purifies to obtain CW-1 (lake shellfish C prime) repeatedly.
Lake shellfish C prime hydrogen carbon modal data:
The preparation of embodiment 2:CW-2
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, CH is added thereto2Cl2(5ml) makees solvent, then plus
Enter triethylamine (140 μ L, 1mmol), aceticanhydride (66 μ L, 0.7mmol), 4-dimethylaminopyridine (DMAP, 12mg, 0.1mmol), it should
5h is stirred at room temperature in reaction.After TCL monitors fully reacting, water quenching reaction is added into reaction solution.Reaction solution adds H2O2With
CH2Cl2(1:1) is extracted (2 times), and CH is merged2Cl2Layer, with the NaHCO of 1:13It washes twice, is washed with water twice, finally uses nothing
Aqueous sodium persulfate dries, filters, and vacuum distillation obtains crude product.Silica gel column chromatography separation, washing and dehydrating integrated machine ratio are petroleum ether: acetone=
6:1, at the same in eluant, eluent be added 1 ‰ triethylamine, finally obtain product 200.7mg, yield 91%.ESI-MS,m/z:
440.20[M+H]+
The preparation of embodiment 3:CW-3
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, anhydrous CH is added thereto2Cl2(5ml) makees solvent,
It adds n-butyric acie (91 μ L, 1mmol), 1- ethyl-(3- dimethylaminopropyl) phosphinylidyne diimmonium salt hydrochlorate is added at 0 DEG C
(EDCI, 575mg, 3mmol) is eventually adding 4-dimethylaminopyridine (DMAP, 12mg, 0.1mmol).Reaction solution is in nitrogen
Under protection, 2h is reacted at room temperature.After TCL monitors fully reacting, add 10% NaHCO3Solution stirs quenching reaction.Reaction solution adds
H2O2And CH2Cl2(1:1) is extracted (2 times), and CH is merged2Cl2Layer, with the saturation NaCl of 1:12It washes twice, then uses H2O2Wash two
Time, it is finally dried, filtered with anhydrous sodium sulfate, vacuum distillation obtains crude product.Silica gel column chromatography separation, washing and dehydrating integrated machine ratio are stone
Oily ether: acetone=6:1, while in eluant, eluent be added 1 ‰ triethylamine, finally obtain product 210.5mg, yield 90.1%.
ESI-MS,m/z:468.20[M+H]+
The preparation of embodiment 4:CW-4
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, anhydrous CH is added thereto2Cl2(5ml) makees molten
Agent adds triethylamine (280 μ L, 2mmol), trimethyl-aceyl chloride (123 μ L, 1mmol), 4-dimethylaminopyridine (DMAP,
12mg, 0.1mmol), reaction solution reacts 5h under the protection of nitrogen at room temperature.After TCL monitors fully reacting, to reaction solution
In plus water quenching reaction.Reaction solution adds H2O2And CH2Cl2(1:1) is extracted (2 times), and CH is merged2Cl2Layer, with the saturation of 1:1
NaHCO3It washes twice, is washed with water twice, is finally dried, filtered with anhydrous sodium sulfate, vacuum distillation obtains crude product.Silicagel column
Chromatographic isolation, washing and dehydrating integrated machine ratio are petroleum ether: acetone=4:1, while 1 ‰ triethylamine being added in eluant, eluent, are finally obtained
Product 211.2mg, yield 87.7%.ESI-MS,m/z:482.30[M+H]+
The preparation of embodiment 5:CW-5
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, anhydrous CH is added thereto2Cl2(5ml) makees solvent,
It adds n-caproic acid (179 μ L, 1mmol), 1- ethyl-(3- dimethylaminopropyl) phosphinylidyne diimmonium salt hydrochlorate is added at 0 DEG C
(EDCI, 575mg, 3mmol) is eventually adding 4-dimethylaminopyridine (DMAP, 12mg, 0.1mmol).Reaction solution is in nitrogen
Under protection, 2h is reacted at room temperature.After TCL monitors fully reacting, add 10% NaHCO3Solution stirs quenching reaction.Reaction solution adds
H2O2And CH2Cl2(1:1) is extracted (2 times), and CH is merged2Cl2Layer, with the saturation NaCl of 1:12It washes twice, then uses H2O2Wash two
Time, it is finally dried, filtered with anhydrous sodium sulfate, vacuum distillation obtains crude product.Silica gel column chromatography separation, washing and dehydrating integrated machine ratio are stone
Oily ether: acetone=6:1, while in eluant, eluent be added 1 ‰ triethylamine, finally obtain product 221.7mg, yield 89.5%.
ESI-MS,m/z:496.30[M+H]+。
The preparation of embodiment 6:CW-6
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, anhydrous CH is added thereto2Cl2(5ml) makees solvent,
It adds 2- thiophenic acid (128mg, 1mmol), 1- ethyl-(3- dimethylaminopropyl) phosphinylidyne diimmonium salt is added at 0 DEG C
Hydrochlorate (EDCI, 575mg, 3mmol) is eventually adding 4-dimethylaminopyridine (DMAP, 12mg, 0.1mmol).Reaction solution is in nitrogen
Under the protection of gas, 3.5h is reacted at room temperature.After TCL monitors fully reacting, add 10% NaHCO3Solution stirs quenching reaction.Instead
Liquid is answered to add H2O2And CH2Cl2(1:1) is extracted (2 times), and CH is merged2Cl2Layer, with the saturation NaCl of 1:12It washes twice, then uses
H2O2It washes twice, is finally dried, filtered with anhydrous sodium sulfate, vacuum distillation obtains crude product.Silica gel column chromatography separation, washing and dehydrating integrated machine
Ratio is petroleum ether: acetone=6:1, while 1 ‰ triethylamine being added in eluant, eluent, finally obtains product 231.4mg, yield
91.2%.ESI-MS,m/z:508.20[M+H]+
The preparation of embodiment 7:CW-7
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, be added thereto anhydrous CH2Cl2 (5ml) make it is molten
Agent adds pyromucic acid (112mg, 1mmol), and 1- ethyl-(3- dimethylaminopropyl) phosphinylidyne diimine is added at 0 DEG C
Hydrochloride (EDCI, 575mg, 3mmol) is eventually adding 4-dimethylaminopyridine (DMAP, 12mg, 0.1mmol).Reaction solution exists
Under the protection of nitrogen, 6h is reacted at room temperature.After TCL monitors fully reacting, 10% NaHCO3 solution is added to stir quenching reaction.Instead
It answers liquid that H2O2 and CH2Cl2 (1:1) is added to be extracted (2 times), merges CH2Cl2 layers, washed twice with the saturation NaCl2 of 1:1, then use
H2O2 is washed twice, is finally dried, filtered with anhydrous sodium sulfate, and vacuum distillation obtains crude product.Silica gel column chromatography separation, washing and dehydrating integrated machine
Ratio is petroleum ether: acetone=6:1, while 1 ‰ triethylamine being added in eluant, eluent, finally obtains product 223.5mg, yield
91%.ESI-MS,m/z:492.20[M+H]+。
The preparation of embodiment 8:CW-8
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, anhydrous CH is added thereto2Cl2(5ml) makees molten
Agent adds triethylamine (280 μ L, 2mmol), chlorobenzoyl chloride (116 μ L, 1mmol), 4-dimethylaminopyridine (DMAP, 12mg,
0.1mmol), reaction solution is under the protection of nitrogen, and reaction is stayed overnight at room temperature.After TCL monitors fully reacting, add into reaction solution
Water quenching reaction.Reaction solution adds H2O2And CH2Cl2(1:1) is extracted (2 times), and CH is merged2Cl2Layer, with the saturation of 1:1
NaHCO3It washes twice, is washed with water twice, is finally dried, filtered with anhydrous sodium sulfate, vacuum distillation obtains crude product.Silicagel column
Chromatographic isolation, washing and dehydrating integrated machine ratio are petroleum ether: acetone=5:1, while 1 ‰ triethylamine being added in eluant, eluent, are finally obtained
Product 238.2mg, yield 95%.ESI-MS,m/z:502.20[M+H]
The preparation of embodiment 9:CW-9
CW-1 (200mg, 0.5mmol) is weighed in 10ml round-bottomed flask, anhydrous CH is added thereto2Cl2(5ml) makees molten
Agent adds triethylamine (280 μ L, 2mmol), p-methyl benzene sulfonic chloride (286mg, 1.5mmol), 4-dimethylaminopyridine
(DMAP, 60mg, 0.5mmol), reaction solution is under the protection of nitrogen, and reaction is stayed overnight at room temperature.After TCL monitors fully reacting,
Add 10%NaHCO into reaction solution3Quenching reaction.Reaction solution adds H2O2And CH2Cl2(1:1) is extracted (2 times), is merged
CH2Cl2Layer, is washed twice with the saturation NaCl of 1:1, is washed with water twice, is finally dried, filtered with anhydrous sodium sulfate, is evaporated under reduced pressure
Obtain crude product.Silica gel column chromatography separation, eluant, eluent ratio is petroleum ether: acetone=8:1, while 1 ‰ being added in eluant, eluent
Triethylamine, finally obtain product 238.2mg, yield 95%.ESI-MS,m/z:552.20[M+H]+。
The preparation of embodiment 10:CW-10
Compound CW-9 (300mg, 0.53mmol) 10ml round-bottomed flask is taken, dry benzene (1.59ml) is added, adds
Potassium tert-butoxide (500mg, 5.3mmol) is added under nitrogen protection in DMSO (2.65ml), and 1h, TLC detection reaction are reacted at 60 DEG C
Completely, raw material disappears.With saturation NaHCO3It is quenched, EA is extracted three times, and EA layers respectively through saturated salt solution and water washing, anhydrous sulphur
Sour sodium is dry, and solvent is recovered under reduced pressure.Crude product is purified by silica gel column chromatography, and eluant, eluent ratio is PE:EA=7:1, while being washed
1 ‰ triethylamine is added in de- agent, finally obtains product Compound 176mg, yield 85%.ESI-MS,m/z:380.20[M+H
]+。
The preparation of embodiment 11:CW-11
It takes compound CW-1 (200mg, 0.5mmol) to be dissolved in dry CH2Cl2 (5ml), is slowly added dropwise two at 0 DEG C
Chlorine sulfoxide (SOCl2,0.36ml, 5mmol), after adding, reaction solution moves to room temperature, and 3h, TLC detection are reacted under the protection of nitrogen
Fully reacting, raw material disappear.Saturation NaHCO3 is added dropwise into reaction solution to generate to bubble-free, layering extracts, water layer CH2Cl2
It is extracted twice, merges organic phase, for organic layer respectively through saturated salt solution and water washing, anhydrous sodium sulfate is dry, is recovered under reduced pressure molten
Agent.Crude product is purified by silica gel column chromatography, and eluant, eluent ratio is PE:EA=10:1, while 1 ‰ three second being added in eluant, eluent
Amine finally obtains product 200mg, yield 96%.ESI-MS,m/z:416.10[M+H]+
Biological activity test
1, anti-human non-small cell lung cancer cell (A549) proliferation activity test
1) test material: common different steroid alkaloid in the compound of the embodiment of the present invention, fritillaria, 5 FU 5 fluorouracil and
Non-small cell lung carcinoma cell (A549)
2) cancer cell line culture
By A549 cell be placed in 10% FBS, 1% penicillin, 1% streptomysin DMEM culture medium in, 37 DEG C,
It is cultivated in 5%CO2 saturated humidity incubator.It after cell covers with bottom of bottle, removes with regard to culture medium, PBS is added to clean, then with 2.5%
Trypsin digestion 3min, culture medium containing serum is added and terminates digestion, is blown and beaten cell by culture bottle is low with pipette tips,
It is centrifuged 3min under 1200r/min and removes supernatant, replaces culture solution, sub-bottle culture, it is outstanding that cell is made in logarithmic growth phase cell
Liquid, for testing.
3) the anti tumor activity in vitro experiment of A549 cell
A experimental group and drug-treated
The each test compound of this experiment is all divided into 8 groups, including 6 administration groups, 1 control group and 1 blank
Group.
B is inoculated with cancer cell in 96 orifice plates
The cancer cell for selecting logarithmic growth phase, through A549 cell is blown and beaten from bottom of bottle after the completion of trypsin digestion,
Be transferred in centrifuge tube, 3min be centrifuged under 1200r/min and removes culture medium, the strain of every hole be added corresponding culture medium be configured to 5 ×
104The cell suspension of/ml is seeded in 96 well culture plates, and every hole is inoculated with 100ul, and the hole of 96 well culture plate surroundings adds PBS, no
Connect cell, be placed in 37 DEG C, 5%CO2, saturated humidity incubator culture for 24 hours.
C delivers medicine to 96 orifice plates
Plating cells can carry out cell administration after for 24 hours, control group and zeroing group are separately added into not drug containing, contain correspondence
Dissolving the 100 μ L of culture medium of the solvent of drug, experimental group is added the culture solution 100 μ L of different pharmaceutical concentration gradient, and each dose
Amount sets 6 parallel holes, cultivates 48h in 5%CO2, the sterile constant temperature cabinet of 37 DEG C of saturated humidity after giving medicine.
D colorometric assay
48h is cultivated, hole is added the MTT sterile solution of the 5mg/ml of 20ul Fresh, is placed in 37 DEG C, 5%CO2, saturation
4h is cultivated in the incubator of humidity.Liquid carefully is discarded supernatant, and 150ulDMSO dissolution purple crystal precipitating is added, by 96 orifice plates
Be placed on shaking table be protected from light at a slow speed, horizontal concussion 10min, the light absorption value in each hole is then detected under 490nm wavelength with microplate reader.
Calculate the growth inhibition ratio under each concentration, GI (growth inhibition ratio)=1- drug control group OD value/control group OD
Value × 100%, wherein items OD value has deducted blank group experiment value.This experiment is positive right with 5 FU 5 fluorouracil (5-Fu)
According to each compound is averaged in triplicate, and experimental result is indicated with average value ± standard deviation
The administration concentration and IC50 of 1 compound of table
As known from Table 1, the anti-human non-small cell lung cancer cell of the compound of the present invention (A549) proliferation activity is apparently higher than it
His fritillaria derivative and positive control 5 FU 5 fluorouracil.
2, resisting leukemia K 562 cell strain proliferation activity is tested
1) test material: common different steroid alkaloid in the compound of the embodiment of the present invention, fritillaria, harringtonine and
People's chronic myelogenous leukemia cell strain (K562)
2) cancer cell line culture
By K562 cell be placed in 10% FBS, 1% penicillin, 1% streptomysin RPMI-1640 culture medium in,
37 DEG C, cultivate in 5%CO2 saturated humidity incubator.After cell covers with bottom of bottle, cell pipette tips are blown and beaten, 1200r/
It is centrifuged 3min under min and removes supernatant, replaces culture solution, sub-bottle culture, logarithmic growth phase cell is made cell suspension, is used for
Experiment.
3) the anti tumor activity in vitro experiment of K562 cell
A experimental group and drug-treated
The each test compound of this experiment is all divided into 8 groups, including 6 administration groups, 1 control group and 1 blank
Group.
B is inoculated with cancer cell in 96 orifice plates
The cancer cell for selecting logarithmic growth phase is transferred in centrifuge tube after blowing and beating K562 cell from bottom of bottle,
Be centrifuged 3min under 1200r/min and remove culture medium, the strain of every hole be added corresponding culture medium be configured to 5 × 104/ml cell it is outstanding
Liquid is seeded in 96 well culture plates, and every hole is inoculated with 100ul, and the hole of 96 well culture plate surroundings adds PBS, do not connect cell, be placed in 37
DEG C, the incubator culture of 5%CO2, saturated humidity for 24 hours.
C delivers medicine to 96 orifice plates
Plating cells can carry out cell administration after for 24 hours, control group and zeroing group are separately added into not drug containing, contain correspondence
Dissolving the 100 μ L of culture medium of the solvent of drug, experimental group is added the culture solution 100 μ L of different pharmaceutical concentration gradient, and each dose
Amount sets 6 parallel holes, cultivates 48h in 5%CO2, the sterile constant temperature cabinet of 37 DEG C of saturated humidity after giving medicine.
D colorometric assay
48h is cultivated, the sterile solution containing 5mg/mlMTT of 20ul Fresh is added in hole, is placed in 37 DEG C, 5%CO2, satisfies
4h is cultivated in the incubator of humidity.Liquid carefully is discarded supernatant, and 150ulDMSO dissolution purple crystal precipitating is added, by 96 holes
Plate be placed on shaking table be protected from light at a slow speed, horizontal concussion 10min, the light absorption value in each hole is then detected under 490nm wavelength with microplate reader.
Calculate the growth inhibition ratio under each concentration, GI (growth inhibition ratio)=1- drug control group OD value/control group OD
Value × 100%, wherein items OD value has deducted blank group experiment value.This experiment is using harringtonine as positive control, each
Compound is averaged in triplicate, and experimental result indicates (x ± s) with average value ± standard deviation.
The administration concentration and IC50 of 2 compound of table
As known from Table 2, the proliferation activity of the compound of the present invention resisting leukemia K 562 cell is apparently higher than as control
Other fritillaria derivatives.
3, anti-human marrow shape thyroid gland duct tumor cell (TT) increment activity test
1) test material: common different steroid alkaloid in the compound of the embodiment of the present invention, fritillaria, 5-fluor-uracil and
People's marrow shape thyroid gland duct tumor cell (TT)
2) cancer cell line culture
By TT cell be placed in 10% FBS, 1% penicillin, 1% streptomysin DMEM culture medium in, 37 DEG C,
It is cultivated in 5%CO2 saturated humidity incubator.It after cell covers with bottom of bottle, removes with regard to culture medium, PBS is added to clean, then with 2.5%
Trypsin digestion 3min, culture medium containing serum is added and terminates digestion, is blown and beaten cell by culture bottle is low with pipette tips,
It is centrifuged 3min under 1200r/min and removes supernatant, replaces culture solution, sub-bottle culture, it is outstanding that cell is made in logarithmic growth phase cell
Liquid, for testing.
3) the anti tumor activity in vitro experiment of TT cell
A experimental group and drug-treated
The each test compound of this experiment is all divided into 9 groups, including 7 administration groups, 1 control group and 1 blank
Group.
B is inoculated with cancer cell in 96 orifice plates
The cancer cell for selecting logarithmic growth phase turns through blowing and beating TT cell from bottom of bottle after the completion of trypsin digestion
Move on in centrifuge tube, 3min be centrifuged under 1200r/min and removes culture medium, the strain of every hole be added corresponding culture medium be configured to 2 ×
The cell suspension of 104/ml is seeded in 96 well culture plates, and every hole is inoculated with 100ul, and the hole of 96 well culture plate surroundings adds PBS, no
Connect cell, be placed in 37 DEG C, 5%CO2, saturated humidity incubator culture for 24 hours.
C delivers medicine to 96 orifice plates
Plating cells can carry out cell administration after for 24 hours, control group and zeroing group are separately added into not drug containing, contain correspondence
Dissolving the 100 μ L of culture medium of the solvent of drug, experimental group is added the culture solution 100 μ L of different pharmaceutical concentration gradient, and each dose
Amount sets 6 parallel holes, cultivates 48h in 5%CO2, the sterile constant temperature cabinet of 37 DEG C of saturated humidity after giving medicine.
D colorometric assay
48h is cultivated, hole is added the sterile solution of the 5mg/mlMTT of 20ul Fresh, is placed in 37 DEG C, 5%CO2, saturation
4h is cultivated in the incubator of humidity.Liquid carefully is discarded supernatant, and 150ulDMSO dissolution purple crystal precipitating is added, by 96 orifice plates
Be placed on shaking table be protected from light at a slow speed, horizontal concussion 10min, the light absorption value in each hole is then detected under 490nm wavelength with microplate reader.
Calculate the growth inhibition ratio under each concentration, GI (growth inhibition ratio)=1- drug control group OD value/control group OD
Value × 100%, wherein items OD value has deducted blank group experiment value.This experiment is using 5 FU 5 fluorouracil as positive control, each
Compound is averaged in triplicate, and experimental result is indicated with average value ± standard deviation
The administration concentration and IC50 of 3 compound of table
As known from Table 3, the compound of the present invention people marrow shape thyroid gland duct tumor cell (TT) proliferation activity is apparently higher than
Other fritillaria derivatives and positive control 5 FU 5 fluorouracil.
Finally, it is stated that the above examples are only used to illustrate the technical scheme of the present invention and are not limiting, although referring to compared with
Good embodiment describes the invention in detail, those skilled in the art should understand that, it can be to skill of the invention
Art scheme is modified or replaced equivalently, and without departing from the objective and range of technical solution of the present invention, should all be covered at this
In the scope of the claims of invention.
Claims (10)
1. the preparation method of different steroid alkaloid shown in formula I, which comprises the steps of:
Hupeh Fritillary Bulb coarse powder solvent extraction is concentrated under reduced pressure to give total medicinal extract, and total medicinal extract is stood after being dispersed with sour water, supernatant tune
PH to more than 9 is saved, extract is obtained by extraction with organic solvent, extract is by the isolated Formula Compound I.
2. a kind of the method for preparing compound of formula I, which comprises the steps of:
1) 3 hydroxyls of hupehenine (hupehenine) are protected to obtain compound 2 with silicon substrate；
2) dehydration of compound 2 obtains compound 3；
3) compound 3 is deprotected to obtain compound of formula I
3. method as claimed in claim 2, which is characterized in that step 1) includes: to exist in organic base and 4-dimethylaminopyridine
Under, hupehenine (hupehenine) is reacted with silated reagent, obtains compound 2, and the silated reagent is selected from: tert-butyl
Dimethylchlorosilane, tri isopropyl chlorosilane, chlorotriethyl silane or tert-butyl diphenyl chlorosilane.
4. method as claimed in claim 3, which is characterized in that step 3) is included in hydrochloric acid, hydrofluoric acid, pyridine hydrofluoride, four
Compound of formula I is deprotected to obtain in the presence of butyl ammonium fluoride or cesium fluoride.
5. such as Claims 2 or 3 the method, which is characterized in that step 2) includes that compound 2 and 4-dimethylaminopyridine is molten
Compound 3 is obtained with phosphorus oxychloride reaction in dry pyridine.
6. the preparation method of Formula II compound, which comprises the steps of:
Compound of formula I and carboxylic acid, acid anhydrides or acyl chloride reaction production II compound,
Wherein, R1It is selected from :-C (O) R2Or-S (O)2R3；
R2Selected from C1-C6Alkyl or optionally by one or more R4Replace following group: C5-C10Heteroaryl or C6-C10Aryl；
R3Selected from C1-C6Alkyl or optionally by one or more R4Substituted C6-C10Aryl；
R4Selected from C1-C6Alkyl or halogen.
7. preparation method as claimed in claim 6, which is characterized in that R2Selected from C1-C6Alkyl, C5-C10Heteroaryl or optionally by one
A or multiple R4Substituted C6-C10Aryl.
8. preparation method as claimed in claims 6 or 7, which is characterized in that R3Selected from optionally by one or more R4Substituted C6-
C10Aryl.
9. the preparation method of formula III compound, which comprises the steps of:
Compound of formula I and thionyl chloride occur substitution reaction and obtain Formula II compound；
10. the preparation method of Formula V compound, which comprises the steps of:
Compound of formula I reacts production IV compound with paratoluensulfonyl chloride, and formula IV compound reacts production V with potassium tert-butoxide
Compound；
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| CN112386600A (en) * | 2020-12-03 | 2021-02-23 | 天津贝猫科技有限公司 | Application of Hupeimine in preparation of medicine for preventing acute kidney injury |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1740187A (en) * | 2004-08-23 | 2006-03-01 | 华中科技大学同济医学院 | Application of peininine and its derivative in preparing antitumor medicine |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1740187A (en) * | 2004-08-23 | 2006-03-01 | 华中科技大学同济医学院 | Application of peininine and its derivative in preparing antitumor medicine |
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| Title |
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| KADOTA, S: "《A STEROIDAL ALKALOID FROM VERATRUM OBLONGUM》", 《PHYTOCHEMISTRY》 * |
| 张鹏: "《湖北贝母茎叶中的生物碱类成分研究》", 《中草药》 * |
| 肖崇厚: "《中药化学》", 30 June 1997, 上海科学技术出版社 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112386600A (en) * | 2020-12-03 | 2021-02-23 | 天津贝猫科技有限公司 | Application of Hupeimine in preparation of medicine for preventing acute kidney injury |
| CN112386600B (en) * | 2020-12-03 | 2023-02-28 | 天津贝猫科技有限公司 | Application of Hupeimine in preparation of medicine for preventing acute kidney injury |
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