CN116987031A - Single benzyl isoquinoline alkaloid, preparation method and application thereof, and pharmaceutical composition - Google Patents
Single benzyl isoquinoline alkaloid, preparation method and application thereof, and pharmaceutical composition Download PDFInfo
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- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 239000008194 pharmaceutical composition Substances 0.000 title claims abstract description 7
- 229930015408 benzyl-isoquinoline alkaloid Natural products 0.000 title description 2
- 150000005516 benzylisoquinolines Chemical class 0.000 title description 2
- 150000001875 compounds Chemical class 0.000 claims abstract description 56
- 208000002193 Pain Diseases 0.000 claims abstract description 9
- IZTUINVRJSCOIR-UHFFFAOYSA-N benzylisoquinoline Chemical class N=1C=CC2=CC=CC=C2C=1CC1=CC=CC=C1 IZTUINVRJSCOIR-UHFFFAOYSA-N 0.000 claims abstract description 6
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 6
- 201000010099 disease Diseases 0.000 claims abstract description 5
- 239000003814 drug Substances 0.000 claims abstract description 5
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 36
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical group OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 30
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 claims description 30
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 27
- 239000003513 alkali Substances 0.000 claims description 19
- 239000002253 acid Substances 0.000 claims description 18
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 13
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 12
- 238000010828 elution Methods 0.000 claims description 12
- 239000000463 material Substances 0.000 claims description 12
- 230000005526 G1 to G0 transition Effects 0.000 claims description 10
- 239000000284 extract Substances 0.000 claims description 10
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 9
- ZMANZCXQSJIPKH-UHFFFAOYSA-N Triethylamine Chemical compound CCN(CC)CC ZMANZCXQSJIPKH-UHFFFAOYSA-N 0.000 claims description 9
- 235000011114 ammonium hydroxide Nutrition 0.000 claims description 9
- 238000000605 extraction Methods 0.000 claims description 9
- 238000001914 filtration Methods 0.000 claims description 8
- 229910052739 hydrogen Inorganic materials 0.000 claims description 8
- 239000001257 hydrogen Substances 0.000 claims description 8
- 238000002953 preparative HPLC Methods 0.000 claims description 7
- 208000019901 Anxiety disease Diseases 0.000 claims description 6
- 230000036506 anxiety Effects 0.000 claims description 6
- 239000002245 particle Substances 0.000 claims description 6
- 241000245050 Menispermum Species 0.000 claims description 5
- 125000004435 hydrogen atom Chemical group [H]* 0.000 claims description 5
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 5
- HQVFCQRVQFYGRJ-UHFFFAOYSA-N formic acid;hydrate Chemical compound O.OC=O HQVFCQRVQFYGRJ-UHFFFAOYSA-N 0.000 claims description 4
- SVWLIIFHXFGESG-UHFFFAOYSA-N formic acid;methanol Chemical compound OC.OC=O SVWLIIFHXFGESG-UHFFFAOYSA-N 0.000 claims description 4
- 238000000034 method Methods 0.000 claims description 4
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 claims description 4
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 4
- 238000010992 reflux Methods 0.000 claims description 4
- AVBGNFCMKJOFIN-UHFFFAOYSA-N triethylammonium acetate Chemical compound CC(O)=O.CCN(CC)CC AVBGNFCMKJOFIN-UHFFFAOYSA-N 0.000 claims description 4
- 125000003342 alkenyl group Chemical group 0.000 claims description 3
- 125000003147 glycosyl group Chemical group 0.000 claims description 3
- 238000010438 heat treatment Methods 0.000 claims description 3
- 239000012535 impurity Substances 0.000 claims description 3
- 238000005342 ion exchange Methods 0.000 claims description 3
- 238000002156 mixing Methods 0.000 claims description 3
- 238000002791 soaking Methods 0.000 claims description 3
- GHQPBDDZGPAVJP-UHFFFAOYSA-N azanium;methanol;hydroxide Chemical compound N.O.OC GHQPBDDZGPAVJP-UHFFFAOYSA-N 0.000 claims description 2
- 229910052736 halogen Inorganic materials 0.000 claims description 2
- 150000002367 halogens Chemical class 0.000 claims description 2
- 150000002431 hydrogen Chemical class 0.000 claims 3
- 239000002585 base Substances 0.000 claims 2
- 238000003810 ethyl acetate extraction Methods 0.000 claims 2
- ZCYVEMRRCGMTRW-UHFFFAOYSA-N 7553-56-2 Chemical compound [I] ZCYVEMRRCGMTRW-UHFFFAOYSA-N 0.000 claims 1
- WKBOTKDWSSQWDR-UHFFFAOYSA-N Bromine atom Chemical compound [Br] WKBOTKDWSSQWDR-UHFFFAOYSA-N 0.000 claims 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 claims 1
- PXGOKWXKJXAPGV-UHFFFAOYSA-N Fluorine Chemical compound FF PXGOKWXKJXAPGV-UHFFFAOYSA-N 0.000 claims 1
- 239000002671 adjuvant Substances 0.000 claims 1
- 125000003545 alkoxy group Chemical group 0.000 claims 1
- GDTBXPJZTBHREO-UHFFFAOYSA-N bromine Substances BrBr GDTBXPJZTBHREO-UHFFFAOYSA-N 0.000 claims 1
- 229910052794 bromium Inorganic materials 0.000 claims 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims 1
- 229910052801 chlorine Inorganic materials 0.000 claims 1
- 239000000460 chlorine Substances 0.000 claims 1
- 239000003937 drug carrier Substances 0.000 claims 1
- 229910052731 fluorine Inorganic materials 0.000 claims 1
- 239000011737 fluorine Substances 0.000 claims 1
- 229910052740 iodine Inorganic materials 0.000 claims 1
- 239000011630 iodine Substances 0.000 claims 1
- 238000004519 manufacturing process Methods 0.000 claims 1
- 238000011321 prophylaxis Methods 0.000 claims 1
- 238000011282 treatment Methods 0.000 claims 1
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- 108700023159 delta Opioid Receptors Proteins 0.000 abstract description 6
- 102000048124 delta Opioid Receptors Human genes 0.000 abstract description 6
- 230000000144 pharmacologic effect Effects 0.000 abstract description 3
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- 238000012360 testing method Methods 0.000 abstract description 2
- 239000010410 layer Substances 0.000 description 22
- 239000000243 solution Substances 0.000 description 12
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 6
- 238000005516 engineering process Methods 0.000 description 6
- 229930013397 isoquinoline alkaloid Natural products 0.000 description 5
- JFJZZMVDLULRGK-URLMMPGGSA-O tubocurarine Chemical compound C([C@H]1[N+](C)(C)CCC=2C=C(C(=C(OC3=CC=C(C=C3)C[C@H]3C=4C=C(C(=CC=4CCN3C)OC)O3)C=21)O)OC)C1=CC=C(O)C3=C1 JFJZZMVDLULRGK-URLMMPGGSA-O 0.000 description 5
- 239000000841 delta opiate receptor agonist Substances 0.000 description 4
- 231100000673 dose–response relationship Toxicity 0.000 description 4
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- WGIDFDFAOQVAHY-UFPGJGBJSA-N 4-[(S)-[(2S,5R)-2,5-dimethyl-4-prop-2-enyl-1-piperazinyl]-phenylmethyl]-N,N-diethylbenzamide Chemical compound C1=CC(C(=O)N(CC)CC)=CC=C1[C@H](C=1C=CC=CC=1)N1[C@@H](C)CN(CC=C)[C@H](C)C1 WGIDFDFAOQVAHY-UFPGJGBJSA-N 0.000 description 3
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- BJWWOUUGCAPHOV-UHFFFAOYSA-N 1,3-dibenzylisoquinoline Chemical compound C=1C2=CC=CC=C2C(CC=2C=CC=CC=2)=NC=1CC1=CC=CC=C1 BJWWOUUGCAPHOV-UHFFFAOYSA-N 0.000 description 2
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 2
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 2
- 206010058019 Cancer Pain Diseases 0.000 description 2
- AQASRZOCERRGBL-UHFFFAOYSA-N Dauricine Natural products CN1CCC2=CC(OC)=C(OC)C=C2C1CC1=CC=C(O)C(OC2=CC=C(C=C2)CC2N(C)CCC=3C=C(C(=CC=32)OC)OC)=C1 AQASRZOCERRGBL-UHFFFAOYSA-N 0.000 description 2
- 239000007995 HEPES buffer Substances 0.000 description 2
- 239000012981 Hank's balanced salt solution Substances 0.000 description 2
- 238000005481 NMR spectroscopy Methods 0.000 description 2
- 239000004480 active ingredient Substances 0.000 description 2
- 239000000556 agonist Substances 0.000 description 2
- 230000008485 antagonism Effects 0.000 description 2
- 239000005557 antagonist Substances 0.000 description 2
- BZKUYNBAFQJRDM-UHFFFAOYSA-N aporphine Chemical compound C12=CC=CC=C2CC2N(C)CCC3=CC=CC1=C32 BZKUYNBAFQJRDM-UHFFFAOYSA-N 0.000 description 2
- 125000004432 carbon atom Chemical group C* 0.000 description 2
- 238000001460 carbon-13 nuclear magnetic resonance spectrum Methods 0.000 description 2
- 238000005100 correlation spectroscopy Methods 0.000 description 2
- AQASRZOCERRGBL-ROJLCIKYSA-N dauricine Chemical compound CN1CCC2=CC(OC)=C(OC)C=C2[C@H]1CC1=CC=C(O)C(OC2=CC=C(C=C2)C[C@H]2N(C)CCC=3C=C(C(=CC=32)OC)OC)=C1 AQASRZOCERRGBL-ROJLCIKYSA-N 0.000 description 2
- 238000000586 desensitisation Methods 0.000 description 2
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- 239000002024 ethyl acetate extract Substances 0.000 description 2
- 235000019253 formic acid Nutrition 0.000 description 2
- 238000003919 heteronuclear multiple bond coherence Methods 0.000 description 2
- -1 hydroxy, carboxy Chemical group 0.000 description 2
- 238000002372 labelling Methods 0.000 description 2
- 238000001819 mass spectrum Methods 0.000 description 2
- 229930014626 natural product Natural products 0.000 description 2
- 108020003175 receptors Proteins 0.000 description 2
- 102000005962 receptors Human genes 0.000 description 2
- 238000002836 resonant waveguide grating Methods 0.000 description 2
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- 125000006727 (C1-C6) alkenyl group Chemical group 0.000 description 1
- 125000004191 (C1-C6) alkoxy group Chemical group 0.000 description 1
- UICBHOXXGLYZJH-UHFFFAOYSA-N 5,6-dihydroisoquinolino[2,1-b]isoquinolin-7-ium Chemical compound C1=CC=C2CC[N+]3=CC4=CC=CC=C4C=C3C2=C1 UICBHOXXGLYZJH-UHFFFAOYSA-N 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 208000024172 Cardiovascular disease Diseases 0.000 description 1
- WQZGKKKJIJFFOK-IVMDWMLBSA-N D-allopyranose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@H](O)[C@@H]1O WQZGKKKJIJFFOK-IVMDWMLBSA-N 0.000 description 1
- 125000000824 D-ribofuranosyl group Chemical group [H]OC([H])([H])[C@@]1([H])OC([H])(*)[C@]([H])(O[H])[C@]1([H])O[H] 0.000 description 1
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- 241000218164 Menispermaceae Species 0.000 description 1
- 241000244939 Menispermum dauricum Species 0.000 description 1
- 208000027520 Somatoform disease Diseases 0.000 description 1
- QCMQEZNBBPGFKQ-UHFFFAOYSA-N Thalisopynine Natural products CN1CCC2=C(OC)C(OC)=C(OC)C3=C2C1CC1=C3C=C(OC)C(O)=C1 QCMQEZNBBPGFKQ-UHFFFAOYSA-N 0.000 description 1
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- 239000013543 active substance Substances 0.000 description 1
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- 230000008484 agonism Effects 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- 125000000217 alkyl group Chemical group 0.000 description 1
- 230000003288 anthiarrhythmic effect Effects 0.000 description 1
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- 125000002519 galactosyl group Chemical group C1([C@H](O)[C@@H](O)[C@@H](O)[C@H](O1)CO)* 0.000 description 1
- 125000002791 glucosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 description 1
- 150000004676 glycans Polymers 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
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- FVRABHGHBLRNNR-UHFFFAOYSA-N liriodenine Natural products O=C1C=CC=c2c1cc3nccc4cc5OCOc5c2c34 FVRABHGHBLRNNR-UHFFFAOYSA-N 0.000 description 1
- 125000003796 lyxosyl group Chemical group C1([C@@H](O)[C@@H](O)[C@H](O)CO1)* 0.000 description 1
- 125000000311 mannosyl group Chemical group C1([C@@H](O)[C@@H](O)[C@H](O)[C@H](O1)CO)* 0.000 description 1
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- 238000005259 measurement Methods 0.000 description 1
- 230000003340 mental effect Effects 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 125000000250 methylamino group Chemical group [H]N(*)C([H])([H])[H] 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- INAXVFBXDYWQFN-XHSDSOJGSA-N morphinan Chemical compound C1C2=CC=CC=C2[C@]23CCCC[C@H]3[C@@H]1NCC2 INAXVFBXDYWQFN-XHSDSOJGSA-N 0.000 description 1
- 235000013557 nattō Nutrition 0.000 description 1
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- 229940002612 prodrug Drugs 0.000 description 1
- 239000000651 prodrug Substances 0.000 description 1
- 238000000425 proton nuclear magnetic resonance spectrum Methods 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
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- 125000000969 xylosyl group Chemical group C1([C@H](O)[C@@H](O)[C@H](O)CO1)* 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D217/00—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems
- C07D217/12—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems with radicals, substituted by hetero atoms, attached to carbon atoms of the nitrogen-containing ring
- C07D217/18—Aralkyl radicals
- C07D217/20—Aralkyl radicals with oxygen atoms directly attached to the aromatic ring of said aralkyl radical, e.g. papaverine
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/22—Anxiolytics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/24—Antidepressants
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
-
- 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
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Veterinary Medicine (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Pain & Pain Management (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Neurology (AREA)
- Neurosurgery (AREA)
- Rheumatology (AREA)
- Psychiatry (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention relates to a monobenzyl isoquinoline alkaloid from radix puerariae of bats, a preparation method and application thereof, and a pharmaceutical composition, wherein the test of an opioid Delta receptor target point shows that the compound has certain pharmacological activity, and can be used for preparing medicaments for preventing and/or treating diseases such as pain. Attachment (New compound structure)
Description
Technical Field
The invention belongs to the field of natural pharmaceutical chemistry, and relates to a preparation method of a monobenzyl isoquinoline alkaloid and application thereof in preventing or treating diseases such as pain, anxiety, depression and the like.
Background
The root and stem of Menispermaceae plant (Menispermum dauricum DC) is a traditional Chinese medicine, and is also called rhizoma Menispermi. Is used for treating tonsillitis, rheumatic arthritis, diarrhea, dysentery, gastroenteritis, cardiovascular diseases, thrombosis diseases, etc. The main component of the radix puerariae of bats is isoquinoline alkaloid and shows wide biological activity such as antiarrhythmic, antibacterial, anti-inflammatory, anti-tumor and the like. The isoquinoline alkaloid in radix Puerariae comprises multiple subclasses such as dibenzyl isoquinoline, benzyl isoquinoline, aporphine, protoberberine and morphinane. Wherein the content of dibenzyl isoquinoline alkaloid is high, and is about more than 85% of liposoluble alkaloid, represented by dauricine and dauricine Ge Sulin.
The technology of label-free cell integrated pharmacology (CLIP) based on an optical sensor is a novel phenotypic screening technology, and the principle of the technology is that when cells are subjected to external stimulus (such as drug small molecules are combined with receptors), many physiological actions generated by the cells bring about redistribution of cell internal mass, the change of the mass distribution is changed through a resonance waveguide grating (Resonant Waveguide Grating, RWG) biosensor, and an output signal is a change of reflection wavelength, so that a visualized dynamic mass reset (Dynamic mass redistribution, DMR) signal is formed, and the experimental operation process can be carried out on an Epic system. The CLIP technology has the characteristics of no labeling (no labeling of receptor and compound), no bias (phenotypic screening), high throughput, no damage, high spatial-temporal resolution (change of reflection wavelength in the whole measurement time range), and the like, and thus, the CLIP technology is widely applied in various fields including screening and discovery of natural product active molecules.
The radix puerariae hepiali is rich in isoquinoline alkaloid, and in view of the important pharmacological activity of isoquinoline alkaloid widely reported at present, the novel active isoquinoline alkaloid is found to be very significant from the radix puerariae hepiali by combining with the CLIP technology.
Disclosure of Invention
The invention provides a preparation method of a monobenzyl isoquinoline alkaloid and application thereof in preventing or treating diseases such as pain, anxiety, depression and the like, wherein the structural general formula of the compound is shown as follows:
wherein:
r1 to R6 are each independently selected from hydrogen, halogen, hydroxy, carboxy, C1-C6 alkoxy, glycosyl, C1-C6 alkenyl, and the like.
Further, R1 is selected from hydrogen or methyl, and R2-R6 are selected from hydrogen, hydroxyl or methoxy.
Further, R1 is selected from methyl, R2 and R3 are selected from methoxy, R4 is selected from hydrogen, and the compound of formula (A) is shown as (B).
Further, R5 and R6 are selected from hydroxyl groups, and the compound of formula (B) is shown as formula (C).
Further, the steric configuration of the compound of formula (C) is preferably 1R.
The invention also provides a process for the preparation of a compound of formula (I), characterized in that it comprises the steps of:
(1) Extracting medicinal materials: 1 kg-100 kg of asiatic moonseed rhizome dry medicinal material, adding 6-10L of ethanol with the volume fraction of 50% -90% into each kg of medicinal material for soaking for 1-24 h, and heating to 50%Filtering for 1-3 h to obtain an extracting solution; reflux-extracting for 1-5 times, mixing the extracting solutions, concentrating the obtained extracting solution to 0.3-0.6L per kilogram of medicinal material to obtain rhizoma Menispermi extracting solution;
(2) Preparing total alkali: adding sulfuric acid with the volume concentration of 0.1% -3% into the rhizoma Menispermi extract to adjust the pH to 1-4, adding ethyl acetate with the volume of 1-3 times of the volume of the acid-adjusted extract to extract, layering to obtain an ethyl acetate extract layer and an acid water layer, extracting for 1-5 times, adding ammonia water with the mass concentration of 25% -28% into the acid water layer to adjust the pH to 8-10, adding n-butanol with the volume of 1-3 times of the volume of the alkali-adjusted sample to extract, layering to obtain n-butanol and an alkali water layer, extracting for 1-5 times, merging the organic layers, concentrating to obtain rhizoma Menispermi crude alkali, and decolorizing and removing impurities through an ion exchange chromatographic column to obtain refined total alkali;
(3) Separating and purifying the total alkali obtained in the step (2) by adopting a reversed phase column filled with C18HCE (particle size 5-60 um), wherein the volume ratio is 0: 100-100: 0 (volume concentration 0.01-5%) formic acid-methanol/(volume concentration 0.01-5%) formic acid-water solution, the elution gradient is 0-80 minutes, 0-95% A. 8 fractions were collected in time, F1 (RT: 0-17 min), F2 (RT: 17-24 min), F3 (RT: 24-28 min), F4 (RT: 28-32.5 min), F5 (RT: 32.5-38.5 min), F6 (RT: 38.5-52 min), F7 (RT: 52-62 min), F8 (RT: 62-74 min), respectively;
(4) Preparing the subfraction F6 obtained in the step (3) by reversed phase preparative HPLC, wherein a chromatographic column is a C8CE stationary phase (5-60 mu m, 20X 250-100X 250 mm), a mobile phase A is (volume fraction 0.01-10%) ammonia water-methanol, B is (volume fraction 0.01-10%) (ammonia water with mass concentration of 25-28%) water, elution gradient is 0-80 minutes, 0% A-95% A is total to obtain 8 subfractions, 8 subfractions are obtained by collecting the total according to time, and F6-1-F6-8 are respectively F6-1 (RT: 2-7 min), F6-2 (RT: 7-11 min), F6-3 (RT: 11-13 min), F6-4 (RT: 13-17 min), F6-5 (RT: 17-22 min), F6-6 (RT: 22-27 min), F6-7 (RT: 27-31 min), F6-8 (RT: 31-35 min);
(5) Preparing the subfraction F6-6 obtained in the step (4) by preparative HPLC, wherein a chromatographic column adopts a C18CE stationary phase (5-60 mu m, 4.6X1250-20X 250 mm), a mobile phase is methanol (A) and water (B) (each containing triethylamine acetate with the volume fraction of 0.01-10% (the volume ratio of the triethylamine to the acetic acid is 1:1-1:5), the elution gradient is 0-60 min,0% A-90% A, the obtained fraction F6-6-4 (peak corresponding to RT:22.5 min) passes through a C18HCE stationary phase (5-60 mu m, 4.6X1250-20X 250 mm), the mobile phase A is formic acid-methanol with the volume fraction of 0.01-1%, and the mobile phase B is formic acid-water with the volume fraction of 0.01-1%, and the elution gradient is 0-40 min,0% A-90% A, so as to obtain the peak corresponding to the compound of the formula (I) (RT: 15 min);
in the invention, the following components are added: the glycosyl group includes, but is not limited to, glucosyl, glucuronyl, mannosyl, galactosyl, allose, fructosyl, sorbosyl, fuosyl, rhamnosyl, chicken natto, arabinosyl, lyxosyl, xylosyl, ribosyl, and various disaccharide and polysaccharide groups formed from the monosaccharides; the C is 1 ～C 6 Alkyl of (C) means C 1 、C 2 、C 3 、C 4 、C 5 、C 6 I.e. straight-chain or branched alkyl having 1 to 6 carbon atoms; c (C) 1 ～C 6 Alkenyl of (C) 1 、C 2 、C 3 、C 4 、C 5 、C 6 I.e. a straight-chain or branched alkenyl group having 1 to 6 carbon atoms, a straight-chain or branched alkenyl group having 1 to 6 double bonds.
It is another object of the present invention to provide a novel monobenzyl isoquinoline alkaloid, or a crystal form thereof, or an isomer thereof, or a pharmaceutically acceptable salt thereof, or a solvate thereof, or a prodrug thereof, or a metabolite thereof as an active ingredient, or an application of any one or more of the above as a Delta receptor agonist in the preparation of medicaments for preventing and/or treating pain, anxiety, depression and the like.
The pain disorders include, but are not limited to, traumatic pain, neuropathic pain, cancer pain, mental (psychological) pain, preferably inflammatory pain and cancer pain.
By pharmaceutical composition is meant that one or more compounds of the invention may be used in combination with each other, or alternatively, the compounds of the invention may be used in combination with any other active agent. If a group of compounds is used, the compounds may be administered to a subject simultaneously, separately or sequentially. The amount of the active ingredient (i.e., the compound of the present invention) in the pharmaceutical composition of the present invention may be specifically applied according to the condition of the patient and the condition diagnosed by the doctor, the dosage or concentration of the active compound is adjusted within a wide range, and the content of the active compound is 1% to 90% of the pharmaceutical composition.
Drawings
FIG. 1 preparation scheme for Compound I
FIG. 2 ESI of Compound I + First-order mass spectrogram
FIG. 3 Compound I 1 H NMR spectrum
FIG. 4 Compound I 13 C NMR spectrum
FIG. 5 Compound I 1 H, 1 H-COSY and HMBC correlation
FIG. 6 dose response curves for agonism, desensitization and antagonism of Compound I on HEK-293-Delta cells, +.representing concentration-response curves for Compound I, +.
Detailed Description
The following examples are intended to illustrate the invention and not to limit the invention further, which may be practiced in any of the ways described in this summary.
Preparation examples of the compounds of formula (I) according to the invention:
preparation of Compounds
100kg of asiatic moonseed rhizome medicinal material,
(1) Extracting medicinal materials: weighing 100kg rhizoma Menispermi, adding 1000L 70% ethanol, soaking for 24h,60Extracting under reflux for 2 hr, filtering to obtain extractive solution 1, adding 1000L of concentrated solutionEthanol with a degree of 70%, 60->Extracting under reflux for 2 hr, filtering to obtain extractive solution 2, adding 1000L 70% ethanol, and 60 ∈>Extracting for 2h by thermal reflux, filtering to obtain an extracting solution 3, combining the extracting solutions 1-3, and concentrating to 50L to obtain the rhizoma Menispermi extracting solution.
(2) Preparing total alkali: adding dilute sulfuric acid with the volume concentration of 1% into the rhizoma Menispermi extract to adjust the pH to 2-3, adding ethyl acetate with the volume of the extract after the acid adjustment to extract for the first time, standing and layering to obtain an ethyl acetate extract layer 1 and an acid water layer 1, continuously adding dilute sulfuric acid with the volume concentration of 0.2% into the acid water layer 1 to adjust the pH to 2-3, adding ethyl acetate with the volume of the extract after the acid adjustment to extract for the second time to obtain an ethyl acetate layer 2 and an acid water layer 2, adding dilute sulfuric acid with the volume concentration of 1% into the acid water layer 2 to adjust the pH to 2-3, and then adding ethyl acetate with the volume of the same as the extract after the acid adjustment to extract for the third time to obtain an ethyl acetate layer 3 and an acid water layer 3. Adding ammonia water with the mass concentration of 25-28% into the acid water layer 3 to adjust the pH value to 9-10, adding n-butanol with the same volume as that of the alkali-adjusted sample for extraction, standing and layering to obtain an n-butanol layer 1 and an alkali water layer 1, adding ammonia water with the mass concentration of 25-28% into the alkali water layer 1 to adjust the pH value to 9-10, and then adding n-butanol with the same volume as that of the alkali-adjusted extract for extraction for the second time to obtain an n-butanol layer 2 and an alkali water layer 2. Then ammonia water with the mass concentration of 25% -28% is added into the alkaline water layer 2 to adjust the pH value to 9-10, then n-butanol with the same volume as the extraction liquid after the alkaline adjustment is added for extraction for the third time, thus obtaining n-butanol layer 3 and alkaline water layer 3, the n-butanol layer 1-3 is combined and concentrated into extract, methanol is adopted to be redissolved to 12L to obtain a crude alkali sample, 10ml of concentrated solution is taken to measure the solid content to be about 500g/L, and the crude asiatic moonseed is calculated to obtain about 6.0kg of crude asiatic moonseed, which accounts for 6.0% of the mass of the medicinal material. In this calculation, about 1L of crude alkali sample re-dissolved in methanol is taken, 1L of pure water is added for dilution and dissolution, the supernatant is obtained by centrifugal filtration, then the supernatant is subjected to ion exchange Q column of agarose gel matrix, and the refined total alkali is obtained by decolorization and impurity removal, wherein the recovery rate is about 97%.
(3) Preparing the total alkali obtained in the step (2) by adopting reversed phase preparative HPLC, separating and purifying a chromatographic column stationary phase which is C18HCE (particle diameter 10um, diameter and height 100X 250 mm) by adopting a reversed phase column, wherein the flow rate is 300mL/min, the mobile phase is methanol (A) and water (B) (each of which contains formic acid with volume concentration of 0.1 percent), and the elution gradient is 0-10min and 0 percent B (volume ratio); 10-20min,10% B;20-35min,20% B;35-45min,25% B;45-60min,30% B; eluting with 60-75min and 90% B to obtain 8 fractions of F1 (RT: 0-17 min), F2 (RT: 17-24 min), F3 (RT: 24-28 min), F4 (RT: 28-32.5 min), F5 (RT: 32.5-38.5 min), F6 (RT: 38.5-52 min), F7 (RT: 52-62 min) and F8 (RT: 62-74 min), respectively;
(4) Preparing the subfraction F6 obtained in the step (3) by reversed phase preparative HPLC, wherein a chromatographic column is a C8CE stationary phase (particle size of 10 mu m, diameter and height of 100X 250 mm), the flow rate is 300mL/min, a mobile phase is methanol (A) and water (B, ammonia water with the mass concentration of 25% -28% with the volume fraction of 0.03%), the elution gradient is 0-30 min,10% -95% A (linear gradient), 30-40 min and 95% A (volume ratio), 8 subfractions are obtained, namely F6-1 (RT: 2-7 min), F6-2 (RT: 7-11 min), F6-3 (RT: 11-13 min), F6-4 (RT: 13-17 min), F6-5 (RT: 17-22 min), F6-6 (RT: 22-27 min), F6-7 (RT: 27-31 min) and F6-8 (RT: 31-35 min) respectively;
(5) Preparing the sub fraction F6-6 obtained in the step (4) by preparative HPLC, wherein a chromatographic column adopts a C18CE stationary phase (particle size 7 μm, diameter and height of 10×250 mm), flow rate is 3mL/min, mobile phase is methanol (A) and water (B) (each containing triethylamine acetate with volume fraction of 0.05% (volume ratio of acetic acid to triethylamine is 1:3)), elution gradient is 0-10min,20% -60% A (linear gradient), 10-25min,60% A,25min-40min,90% A (volume ratio), fraction F6-6-1 (peak corresponding to RT:18 min), F6-6-2 (peak corresponding to RT:19.5 min), F6-6-3 (peak corresponding to RT:21 min), and F6-6-4 (peak corresponding to RT:22.5 min) are obtained. The obtained fractions F6-6-4 respectively pass through a C18HCE stationary phase (particle size 7 μm, diameter and height 10X 250 mm), mobile phases A and B are methanol and water (each containing 0.2% formic acid by volume fraction), elution gradient is 0-40 min,0% A-90% A (linear gradient), and compound I (peak corresponding to RT:15 min) is obtained by peak collection;
the preparation scheme of the compound I is shown in figure 1.
(5) The plane structure of the compound I is determined by ultraviolet, mass spectrum and nuclear magnetism characterization, the three-dimensional configuration of the compound I is determined by electronic round dichroism (ECD), the structural formula of the compound I is shown in the formula (I), and the identification information is as follows:
compound I:4mg, C 26 H 29 NO 5 MW:435.2046, white powder, was dissolved in methanol.
1 H NMR(CD 3 OD,600MHz):δ4.38(1H,dd,J＝10.0,4.5Hz,H-1),3.31(1H,m,H-3α),3.68(1H,m,H-3β),3.13(1H,ddd,J＝17.6,10.8,7.0Hz H-4α),3.01(1H,m,H-4β),6.79(1H,s,H-5),5.92(1H,s,H-8),3.37(1H,dd,J＝13.2,4.5Hz,H-9α),2.99(1H,dd,J＝13.2,10.0Hz,H-9β),7.07(1H,br d,J＝8.5Hz,H-10),6.88(1H,br d,J＝8.5Hz,H-11),6.88(1H,br d,J＝8.5Hz,H-13),7.07(1H,br d,J＝8.5Hz,H-14),6.93(1H,d,J＝8.3Hz,H-3′),6.95(1H,d,J＝2.0Hz,H-4′),4.48(2H,s,H-7′),2.89(3H,s,N-Me),3.80(3H,s,CH 3 O-6),3.49(3H,s,CH 3 O-7)。
ECD(MeOH)λ max (△ε)211(-33.62),233(-16.96),287(-4.52)nm；
ESI of Compound I + A primary mass spectrum, 1 H NMR 13 C NMR spectrum 1 H, 1 The correlation of H-COSY and HMBC is shown in FIGS. 2-5.
Activity test examples:
the sample is the prepared new compound (compound I); HEK293-delta cells were derived from HEK293, a cell overexpressing delta receptor used by the institute of chemical and physical, dalian, china academy of sciences, and from laboratory transfection, and reference was constructed (Xu, F.F.; zhou, H.; liu, X.M.; zhang, X.L.; A.C.;wang, z.w.; hou, t.; wang, j.x.; qu, l.l.; zhang, p.y.; piao, h.l.; liang, X.M., label-free cell phenotypic study of FFA and FFA1 and discovery of novel agonists of FFA4 from natural products.Rsc Advances 2019,9 (26), 15073-15083); SNC162 (cat# 1529) and SDM25N (cat# 1410) were purchased from Tocres corporation; DMEM high-sugar broth (cat# C11995500 BT) was purchased from ThermoFisher company, and fetal bovine serum (cat# 04000101A) was purchased from Shenyang Hui Bai Biotech Co., ltd; balanced salt solutions HBSS (cat# 14065-056) and HEPES (cat# 15630-080) were purchased from Gibco corporation. The detection platform is Kang Ningdi third generationThe signal detected by the imager is the wavelength shift caused by the Dynamic Mass Reset (DMR) of the cell.
Inoculating HEK293-delta cells in logarithmic growth phaseIn a well (total number of wells 384) biosensor microplate, the volume of the seeded cell suspension per well was 40. Mu.L, and the number of cells seeded per well was 2.5X10 4 Next, 384-well plates were placed in a cell incubator (containing CO at a volume concentration of 5%) 2 Culturing at 37deg.C for 22-24 hr, sucking off culture solution from 384-well microplate inoculated with HEK293-delta cells when cell fusion degree reaches 95%, adding 30 μl HBSS buffer (containing 20mM HEPES) into each well, and placing in->The high throughput high content activity screening system was equilibrated and then tested.
The experimental steps are carried out in the following three steps:
(1) Excitation experiments: a baseline of 2min was established and then compound I (10. Mu.L) was added to different wells at different final concentrations (100. Mu.M, 50. Mu.M, 25. Mu.M, 12.5. Mu.M, 6.25. Mu.M, 3.125. Mu.M, 1.563. Mu.M, 0.781. Mu.M, 0.391. Mu.M, 0.195. Mu.M, 0.098. Mu.M, 0.049. Mu.M, 0.024. Mu.M) and continued monitoring for 1h as shown in FIG. 6The dose response curve of Compound I (Compound I) is of the monophasic "S" type and reaches a saturated response, corresponding to EC 50 The value was 2.5.+ -. 0.3. Mu.M.
(2) Desensitization experiment: after pretreatment of HEK293-Delta cells with 10. Mu.L of Compound I (10. Mu.L) at various final concentrations (100. Mu.M, 50. Mu.M, 25. Mu.M, 12.5. Mu.M, 6.25. Mu.M, 3.125. Mu.M, 1.563. Mu.M, 0.781. Mu.M, 0.391. Mu.M, 0.195. Mu.M, 0.098. Mu.M, 0.049. Mu.M, 0.024. Mu.M) for 1h, a 2min baseline was established, and then 10. Mu.L (400 nM) of Delta receptor-specific agonist SNC162 was added to each well, and the results after continued monitoring for 1h are shown in FIG. 6, compound I was able to completely desensitize the DMR response signal of Delta receptor agonist SNC162 (Compound I-SNC 162), which was dose-dependent, IC was shown 50 The value was 45.3.+ -. 5.5. Mu.M.
(3) Antagonism experiment: the results of continuing monitoring for 1 hour after 10. Mu.L of different final concentrations (32. Mu.M, 16. Mu.M, 8. Mu.M, 4. Mu.M, 2. Mu.M, 1. Mu.M, 0.5. Mu.M, 0.25. Mu.M, 0.125. Mu.M, 0.063. Mu.M, 0.031. Mu.M, 0.016. Mu.M, 0.008. Mu.M) of the specific antagonist SDM25N of Delta receptor were added to the different wells, HEK293-Delta cells were pretreated for 1 hour, and a 2min baseline was established, followed by 10. Mu.L of Compound I (fixed final concentration of 50. Mu.M) was added to each well as shown in FIG. 6. Delta receptor selective antagonist SDM25N is capable of completely inhibiting the DMR response signal of Compound I on HEK293-Delta cells (SDM25N_CompoundI) and exhibits dose dependency, IC thereof 50 The value was 0.31.+ -. 0.09. Mu.M, which means that the compound was a Delta agonist. Current studies indicate that opioid Delta receptors are associated with pain, anxiety, depression, and other conditions. The compound of the invention has different parent nucleus structure from the existing Delta agonist, and is important for pharmacological research and clinical application of diseases such as pain, anxiety, depression and the like.
Claims (9)
1. A monobenzyl isoquinoline alkaloid has a structure shown in a formula (A):
wherein:
r1 to R6 are each independently selected from one or more of hydrogen, halogen (one or more of fluorine, chlorine, bromine and iodine), hydroxyl, carboxyl, C1 to C6 alkoxy, glycosyl and C1 to C6 alkenyl.
2. A compound according to claim 1, characterized in that: r1 is selected from hydrogen or methyl, and R2-R6 are respectively and independently selected from hydrogen, hydroxyl or methoxy.
3. A compound according to claim 2, characterized in that: r1 is selected from methyl, R2 and R3 are selected from methoxy, R4 is selected from hydrogen, and the compound shown in the formula (A) is shown in the formula (B);
4. a compound according to claim 1 or 3, characterized in that: r5 and R6 are selected from hydroxyl, and the compound shown in the formula (B) is shown in the formula (C):
5. a compound according to claim 4, characterized in that: the compound of formula (C) preferably has a steric configuration selected from 1R.
6. A process for preparing a compound according to claim 5, comprising the steps of:
(1) Extracting medicinal materials: 1 kg-100 kg of asiatic moonseed rhizome, adding 6-10L of ethanol with the volume fraction of 50% -90% into each kg of medicinal materials, soaking for 1-24 h, heating to 50-90 ℃ for reflux extraction for 1-3 h, and filtering to obtain an extracting solution; reflux-extracting for 1-5 times, mixing the extracting solutions, concentrating the obtained extracting solution to 0.3-0.6L per kilogram of medicinal material to obtain rhizoma Menispermi extracting solution;
(2) Preparing total alkali: adding sulfuric acid with the volume concentration of 0.1% -3% into the rhizoma Menispermi extract in the step (1) to adjust the pH to 1-4, adding ethyl acetate with the volume of 1-3 times of the volume of the acid-adjusted extract for extraction, layering to obtain an ethyl acetate extraction layer and an acid water layer, extracting for 1-5 times (namely, re-extracting the acid water layer for 0-4 times), adding ammonia water with the mass concentration of 25% -28% into the acid water layer to adjust the pH to 8-10, adding n-butanol with the volume of 1-3 times of the volume of the alkali-adjusted sample for extraction, layering to obtain an n-butanol layer and an alkali water layer, extracting for 1-5 times (namely, re-extracting the alkali water layer for 0-4 times), merging the n-butanol layer, concentrating to obtain rhizoma Menispermi crude alkali, and decolorizing and removing impurities through an ion exchange chromatographic column to obtain refined total alkali;
(3) Separating and purifying the total alkali obtained in the step (2) by adopting a reversed phase column filled with C18HCE (particle size 5-60 um), wherein the volume ratio is 0: 100-100: 0 (volume concentration 0.01-5%) formic acid-methanol/(volume concentration 0.01-5%) formic acid-water solution, the elution gradient is 0-80 minutes, 0-95% A. 8 fractions were collected in time, F1 (RT: 0-17 min), F2 (RT: 17-24 min), F3 (RT: 24-28 min), F4 (RT: 28-32.5 min), F5 (RT: 32.5-38.5 min), F6 (RT: 38.5-52 min), F7 (RT: 52-62 min), F8 (RT: 62-74 min), respectively;
(4) Preparing the subfraction F6 obtained in the step (3) by reversed phase preparative HPLC, wherein a chromatographic column is a C8CE stationary phase (5-60 mu m, 20X 250-100X 250 mm), a mobile phase A is (volume fraction 0.01-10%) ammonia water-methanol, B is (volume fraction 0.01-10%) (ammonia water with mass concentration of 25-28%) water, elution gradient is 0-80 minutes, 0-95% A is collected by time to obtain 8 subfractions, F6-1-F6-8 are respectively F6-1 (RT: 2-7 min), F6-2 (RT: 7-11 min), F6-3 (RT: 11-13 min), F6-4 (RT: 13-17 min), F6-5 (RT: 17-22 min), F6-6 (RT: 22-27 min), F6-7 (RT: 27-31 min), F6-8 (RT: 31-35 min);
(5) Preparing the subfraction F6-6 obtained in the step (4) by preparative HPLC, wherein a chromatographic column adopts a C18CE stationary phase (5-60 mu m, 4.6X1250-20X 250 mm), a mobile phase is methanol (A) and water (B) (each containing 0.01-10% of triethylamine acetate (the volume ratio of the triethylamine acetate to the triethylamine is 1:1-1:5), the elution gradient is 0-60 min,0% of A-90% of A, the obtained fraction F6-6-4 (peak corresponding to RT:22.5 min) passes through a C18HCE stationary phase (5-60 mu m, 4.6X1250-20X 250 mm), the mobile phase A is formic acid-methanol (0.01-1%) and the mobile phase B is formic acid-water (0.01-1%), and the elution gradient is 0-40 min,0% of A-90% of A, so as to obtain the compound of the formula (I) (peak corresponding to RT:15 min).
7. A process for producing a compound according to claim 6, wherein,
when reflux-extracting for 2-5 times in the step (1), filtering the materials in the previous reflux-extracting process, adding 6-10L of 50-90% ethanol in volume fraction per kilogram of medicinal materials into the filtered filter residue, heating to 50-90 ℃ for reflux-extracting for 1-3 h, and filtering to obtain an extracting solution; mixing the extractive solutions;
when ethyl acetate is extracted for 2-5 times in the step (2), firstly, sulfuric acid with the volume concentration of 0.1% -3% is adopted to adjust the pH value of the acid water layer layered last time to 1-4, ethyl acetate with the volume of 1-3 times of the volume of the acid water layer is added into the acid water layer for extraction, and the ethyl acetate extraction layer and the acid water layer are obtained through layering; the weak base is ammonia water with the mass concentration of 10-25%;
when n-butanol is extracted for 2-5 times, firstly, weak base is adopted to adjust the pH value of the alkaline water layer which is layered last time to 8-10, and n-butanol with the volume of 1-3 times of the volume of the alkaline water layer is added into the alkaline water layer for extraction and layering, so that n-butanol and an alkaline water layer are obtained.
8. Use of a compound according to claim 5 for the preparation of a medicament for the prophylaxis and/or treatment of one or more diseases selected from pain, anxiety and depression.
9. A pharmaceutical composition characterized by: the compound of claim 5 is formulated (e.g., mixed) with a pharmaceutically acceptable carrier or adjuvant.
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