CN106420902B - 胀果甘草提取物及甘草查耳酮a的保肝活性及药物新用途 - Google Patents
胀果甘草提取物及甘草查耳酮a的保肝活性及药物新用途 Download PDFInfo
- Publication number
- CN106420902B CN106420902B CN201610894553.5A CN201610894553A CN106420902B CN 106420902 B CN106420902 B CN 106420902B CN 201610894553 A CN201610894553 A CN 201610894553A CN 106420902 B CN106420902 B CN 106420902B
- Authority
- CN
- China
- Prior art keywords
- licochalcone
- glycyrrhiza
- extract
- glycyrrhiza inflata
- liver
- 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.)
- Active
Links
- 241001278898 Glycyrrhiza inflata Species 0.000 title abstract description 49
- KAZSKMJFUPEHHW-UHFFFAOYSA-N (2E)-3-[5-(1,1-dimethyl-2-propenyl)-4-hydroxy-2-methoxyphenyl]-1-(4-hdyroxyphenyl)-2-propen-1-one Natural products COC1=CC(O)=C(C(C)(C)C=C)C=C1C=CC(=O)C1=CC=C(O)C=C1 KAZSKMJFUPEHHW-UHFFFAOYSA-N 0.000 title abstract description 43
- IUCVKTHEUWACFB-UHFFFAOYSA-N Licochalcone A Natural products COC1=CC=C(C(C)(C)C=C)C=C1C=CC(=O)C1=CC=C(O)C=C1 IUCVKTHEUWACFB-UHFFFAOYSA-N 0.000 title abstract description 43
- KAZSKMJFUPEHHW-DHZHZOJOSA-N Licochalcone A Chemical compound COC1=CC(O)=C(C(C)(C)C=C)C=C1\C=C\C(=O)C1=CC=C(O)C=C1 KAZSKMJFUPEHHW-DHZHZOJOSA-N 0.000 title abstract description 43
- 239000000284 extract Substances 0.000 title abstract description 43
- 230000000694 effects Effects 0.000 title abstract description 32
- 210000004185 liver Anatomy 0.000 title description 18
- 206010067125 Liver injury Diseases 0.000 claims abstract description 24
- 239000003814 drug Substances 0.000 claims abstract description 22
- 229940079593 drug Drugs 0.000 claims abstract description 15
- 239000000126 substance Substances 0.000 claims abstract description 14
- 150000001875 compounds Chemical class 0.000 claims abstract description 11
- 231100000753 hepatic injury Toxicity 0.000 claims abstract description 11
- 239000000203 mixture Substances 0.000 claims abstract description 6
- 150000003839 salts Chemical class 0.000 claims abstract description 5
- VZGDMQKNWNREIO-UHFFFAOYSA-N tetrachloromethane Chemical group ClC(Cl)(Cl)Cl VZGDMQKNWNREIO-UHFFFAOYSA-N 0.000 claims description 44
- 238000002360 preparation method Methods 0.000 claims description 4
- 239000003937 drug carrier Substances 0.000 claims description 2
- 230000002265 prevention Effects 0.000 claims description 2
- 240000008917 Glycyrrhiza uralensis Species 0.000 abstract description 14
- 235000000554 Glycyrrhiza uralensis Nutrition 0.000 abstract description 14
- 239000000469 ethanolic extract Substances 0.000 abstract description 12
- 238000011160 research Methods 0.000 abstract description 6
- 150000002148 esters Chemical class 0.000 abstract description 4
- 239000000651 prodrug Substances 0.000 abstract description 4
- 229940002612 prodrug Drugs 0.000 abstract description 4
- 239000012453 solvate Substances 0.000 abstract description 4
- 230000001154 acute effect Effects 0.000 abstract description 3
- 231100000234 hepatic damage Toxicity 0.000 abstract description 3
- 230000008818 liver damage Effects 0.000 abstract description 3
- 230000004792 oxidative damage Effects 0.000 abstract description 3
- 230000005779 cell damage Effects 0.000 abstract description 2
- 208000037887 cell injury Diseases 0.000 abstract description 2
- 230000001590 oxidative effect Effects 0.000 abstract description 2
- 208000037816 tissue injury Diseases 0.000 abstract description 2
- 230000036541 health Effects 0.000 abstract 1
- 230000009897 systematic effect Effects 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 25
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 22
- 235000006200 Glycyrrhiza glabra Nutrition 0.000 description 22
- 244000303040 Glycyrrhiza glabra Species 0.000 description 19
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 15
- 239000002609 medium Substances 0.000 description 15
- 101000588302 Homo sapiens Nuclear factor erythroid 2-related factor 2 Proteins 0.000 description 13
- 102100031701 Nuclear factor erythroid 2-related factor 2 Human genes 0.000 description 13
- 238000000034 method Methods 0.000 description 12
- 239000000243 solution Substances 0.000 description 12
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 11
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 9
- 239000000463 material Substances 0.000 description 9
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 8
- 241000202807 Glycyrrhiza Species 0.000 description 8
- 108700008625 Reporter Genes Proteins 0.000 description 8
- 230000004913 activation Effects 0.000 description 8
- 231100000439 acute liver injury Toxicity 0.000 description 8
- LPLVUJXQOOQHMX-QWBHMCJMSA-N glycyrrhizinic acid Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@H](O[C@@H]1O[C@@H]1C([C@H]2[C@]([C@@H]3[C@@]([C@@]4(CC[C@@]5(C)CC[C@@](C)(C[C@H]5C4=CC3=O)C(O)=O)C)(C)CC2)(C)CC1)(C)C)C(O)=O)[C@@H]1O[C@H](C(O)=O)[C@@H](O)[C@H](O)[C@H]1O LPLVUJXQOOQHMX-QWBHMCJMSA-N 0.000 description 8
- 235000011477 liquorice Nutrition 0.000 description 8
- 230000001681 protective effect Effects 0.000 description 8
- 230000006378 damage Effects 0.000 description 7
- 241001465754 Metazoa Species 0.000 description 6
- 241000699666 Mus <mouse, genus> Species 0.000 description 5
- 210000004369 blood Anatomy 0.000 description 5
- 239000008280 blood Substances 0.000 description 5
- 238000001514 detection method Methods 0.000 description 5
- 210000003494 hepatocyte Anatomy 0.000 description 5
- 230000005764 inhibitory process Effects 0.000 description 5
- 230000000144 pharmacologic effect Effects 0.000 description 5
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 4
- 108020004414 DNA Proteins 0.000 description 4
- 235000001453 Glycyrrhiza echinata Nutrition 0.000 description 4
- 235000017382 Glycyrrhiza lepidota Nutrition 0.000 description 4
- 108060001084 Luciferase Proteins 0.000 description 4
- 239000005089 Luciferase Substances 0.000 description 4
- 238000002835 absorbance Methods 0.000 description 4
- -1 chalcone compound Chemical class 0.000 description 4
- 230000003013 cytotoxicity Effects 0.000 description 4
- 231100000135 cytotoxicity Toxicity 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 4
- 208000006454 hepatitis Diseases 0.000 description 4
- LTINPJMVDKPJJI-UHFFFAOYSA-N iodinated glycerol Chemical compound CC(I)C1OCC(CO)O1 LTINPJMVDKPJJI-UHFFFAOYSA-N 0.000 description 4
- 229940010454 licorice Drugs 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 238000001228 spectrum Methods 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- 102100036475 Alanine aminotransferase 1 Human genes 0.000 description 3
- 108010082126 Alanine transaminase Proteins 0.000 description 3
- 108010003415 Aspartate Aminotransferases Proteins 0.000 description 3
- 102000004625 Aspartate Aminotransferases Human genes 0.000 description 3
- DQFBYFPFKXHELB-UHFFFAOYSA-N Chalcone Natural products C=1C=CC=CC=1C(=O)C=CC1=CC=CC=C1 DQFBYFPFKXHELB-UHFFFAOYSA-N 0.000 description 3
- 102000003855 L-lactate dehydrogenase Human genes 0.000 description 3
- 108700023483 L-lactate dehydrogenases Proteins 0.000 description 3
- 231100000354 acute hepatitis Toxicity 0.000 description 3
- 230000003078 antioxidant effect Effects 0.000 description 3
- 235000005513 chalcones Nutrition 0.000 description 3
- 208000019425 cirrhosis of liver Diseases 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 230000007850 degeneration Effects 0.000 description 3
- 239000002552 dosage form Substances 0.000 description 3
- 238000000605 extraction Methods 0.000 description 3
- 239000012091 fetal bovine serum Substances 0.000 description 3
- 229940069445 licorice extract Drugs 0.000 description 3
- 210000005228 liver tissue Anatomy 0.000 description 3
- 239000006166 lysate Substances 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 235000018102 proteins Nutrition 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 210000002966 serum Anatomy 0.000 description 3
- 239000011550 stock solution Substances 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 230000004083 survival effect Effects 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- 238000005303 weighing Methods 0.000 description 3
- 108700032225 Antioxidant Response Elements Proteins 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 241000196324 Embryophyta Species 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- DPXJVFZANSGRMM-UHFFFAOYSA-N acetic acid;2,3,4,5,6-pentahydroxyhexanal;sodium Chemical compound [Na].CC(O)=O.OCC(O)C(O)C(O)C(O)C=O DPXJVFZANSGRMM-UHFFFAOYSA-N 0.000 description 2
- 230000003213 activating effect Effects 0.000 description 2
- 239000004480 active ingredient Substances 0.000 description 2
- 230000009692 acute damage Effects 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000003963 antioxidant agent Substances 0.000 description 2
- 235000006708 antioxidants Nutrition 0.000 description 2
- 239000002775 capsule Substances 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 239000001768 carboxy methyl cellulose Substances 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 231100000012 chronic liver injury Toxicity 0.000 description 2
- 239000002285 corn oil Substances 0.000 description 2
- 235000005687 corn oil Nutrition 0.000 description 2
- 239000008367 deionised water Substances 0.000 description 2
- 229910021641 deionized water Inorganic materials 0.000 description 2
- 238000010828 elution Methods 0.000 description 2
- SRCZQMGIVIYBBJ-UHFFFAOYSA-N ethoxyethane;ethyl acetate Chemical compound CCOCC.CCOC(C)=O SRCZQMGIVIYBBJ-UHFFFAOYSA-N 0.000 description 2
- 239000002024 ethyl acetate extract Substances 0.000 description 2
- 230000003203 everyday effect Effects 0.000 description 2
- 239000008187 granular material Substances 0.000 description 2
- 238000010438 heat treatment Methods 0.000 description 2
- 206010073071 hepatocellular carcinoma Diseases 0.000 description 2
- 230000002443 hepatoprotective effect Effects 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 229910052739 hydrogen Inorganic materials 0.000 description 2
- 239000001257 hydrogen Substances 0.000 description 2
- 230000008595 infiltration Effects 0.000 description 2
- 238000001764 infiltration Methods 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 238000002372 labelling Methods 0.000 description 2
- 201000007270 liver cancer Diseases 0.000 description 2
- 208000014018 liver neoplasm Diseases 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- 230000003647 oxidation Effects 0.000 description 2
- 238000007254 oxidation reaction Methods 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- 239000006187 pill Substances 0.000 description 2
- 238000010992 reflux Methods 0.000 description 2
- 230000001105 regulatory effect Effects 0.000 description 2
- 239000000741 silica gel Substances 0.000 description 2
- 229910002027 silica gel Inorganic materials 0.000 description 2
- 235000019812 sodium carboxymethyl cellulose Nutrition 0.000 description 2
- 229920001027 sodium carboxymethylcellulose Polymers 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 238000000967 suction filtration Methods 0.000 description 2
- 239000003826 tablet Substances 0.000 description 2
- DQFBYFPFKXHELB-VAWYXSNFSA-N trans-chalcone Chemical compound C=1C=CC=CC=1C(=O)\C=C\C1=CC=CC=C1 DQFBYFPFKXHELB-VAWYXSNFSA-N 0.000 description 2
- 229910021642 ultra pure water Inorganic materials 0.000 description 2
- 239000012498 ultrapure water Substances 0.000 description 2
- 210000003462 vein Anatomy 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 1
- 229930195730 Aflatoxin Natural products 0.000 description 1
- XWIYFDMXXLINPU-UHFFFAOYSA-N Aflatoxin G Chemical compound O=C1OCCC2=C1C(=O)OC1=C2C(OC)=CC2=C1C1C=COC1O2 XWIYFDMXXLINPU-UHFFFAOYSA-N 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 102000002004 Cytochrome P-450 Enzyme System Human genes 0.000 description 1
- 108010015742 Cytochrome P-450 Enzyme System Proteins 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 206010016654 Fibrosis Diseases 0.000 description 1
- 229920002527 Glycogen Polymers 0.000 description 1
- 206010019668 Hepatic fibrosis Diseases 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 239000012097 Lipofectamine 2000 Substances 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- SEBFKMXJBCUCAI-UHFFFAOYSA-N NSC 227190 Natural products C1=C(O)C(OC)=CC(C2C(OC3=CC=C(C=C3O2)C2C(C(=O)C3=C(O)C=C(O)C=C3O2)O)CO)=C1 SEBFKMXJBCUCAI-UHFFFAOYSA-N 0.000 description 1
- 229930040373 Paraformaldehyde Natural products 0.000 description 1
- 239000004952 Polyamide Substances 0.000 description 1
- GAMYVSCDDLXAQW-AOIWZFSPSA-N Thermopsosid Natural products O(C)c1c(O)ccc(C=2Oc3c(c(O)cc(O[C@H]4[C@H](O)[C@@H](O)[C@H](O)[C@H](CO)O4)c3)C(=O)C=2)c1 GAMYVSCDDLXAQW-AOIWZFSPSA-N 0.000 description 1
- 108090000340 Transaminases Proteins 0.000 description 1
- 102000003929 Transaminases Human genes 0.000 description 1
- 108091023040 Transcription factor Proteins 0.000 description 1
- 102000040945 Transcription factor Human genes 0.000 description 1
- 208000025865 Ulcer Diseases 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000002671 adjuvant Substances 0.000 description 1
- 239000005409 aflatoxin Substances 0.000 description 1
- 150000001413 amino acids Chemical group 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 230000000843 anti-fungal effect Effects 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 229940121375 antifungal agent Drugs 0.000 description 1
- 239000011230 binding agent Substances 0.000 description 1
- 238000012742 biochemical analysis Methods 0.000 description 1
- 238000010876 biochemical test Methods 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 229940098773 bovine serum albumin Drugs 0.000 description 1
- 210000005252 bulbus oculi Anatomy 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 230000003833 cell viability Effects 0.000 description 1
- 231100000481 chemical toxicant Toxicity 0.000 description 1
- 230000009693 chronic damage Effects 0.000 description 1
- 230000007882 cirrhosis Effects 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000005336 cracking Methods 0.000 description 1
- 230000000120 cytopathologic effect Effects 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 238000001784 detoxification Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- WGLUMOCWFMKWIL-UHFFFAOYSA-N dichloromethane;methanol Chemical compound OC.ClCCl WGLUMOCWFMKWIL-UHFFFAOYSA-N 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000007884 disintegrant Substances 0.000 description 1
- 235000013399 edible fruits Nutrition 0.000 description 1
- 239000003623 enhancer Substances 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 239000010408 film Substances 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 229930003944 flavone Natural products 0.000 description 1
- 150000002212 flavone derivatives Chemical class 0.000 description 1
- 235000011949 flavones Nutrition 0.000 description 1
- 229930182486 flavonoid glycoside Natural products 0.000 description 1
- 150000007955 flavonoid glycosides Chemical class 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 238000003304 gavage Methods 0.000 description 1
- 229940096919 glycogen Drugs 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 210000003128 head Anatomy 0.000 description 1
- 235000013402 health food Nutrition 0.000 description 1
- 238000007490 hematoxylin and eosin (H&E) staining Methods 0.000 description 1
- 230000002440 hepatic effect Effects 0.000 description 1
- 206010019692 hepatic necrosis Diseases 0.000 description 1
- 230000007866 hepatic necrosis Effects 0.000 description 1
- 231100000283 hepatitis Toxicity 0.000 description 1
- 231100000334 hepatotoxic Toxicity 0.000 description 1
- 230000003082 hepatotoxic effect Effects 0.000 description 1
- 239000003906 humectant Substances 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 210000004969 inflammatory cell Anatomy 0.000 description 1
- 230000002757 inflammatory effect Effects 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 230000037356 lipid metabolism Effects 0.000 description 1
- 230000003859 lipid peroxidation Effects 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 210000005229 liver cell Anatomy 0.000 description 1
- 231100000832 liver cell necrosis Toxicity 0.000 description 1
- 208000019423 liver disease Diseases 0.000 description 1
- 210000001853 liver microsome Anatomy 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 239000000314 lubricant Substances 0.000 description 1
- 201000004792 malaria Diseases 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 239000012982 microporous membrane Substances 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 230000017074 necrotic cell death Effects 0.000 description 1
- 231100000065 noncytotoxic Toxicity 0.000 description 1
- 230000002020 noncytotoxic effect Effects 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 238000001543 one-way ANOVA Methods 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 239000012188 paraffin wax Substances 0.000 description 1
- 229920002866 paraformaldehyde Polymers 0.000 description 1
- 210000004738 parenchymal cell Anatomy 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 230000010412 perfusion Effects 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N phenol group Chemical group C1(=CC=CC=C1)O ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 1
- 150000002989 phenols Chemical class 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 229920002647 polyamide Polymers 0.000 description 1
- 235000013824 polyphenols Nutrition 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000010298 pulverizing process Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000007790 scraping Methods 0.000 description 1
- 238000011894 semi-preparative HPLC Methods 0.000 description 1
- 230000009528 severe injury Effects 0.000 description 1
- SEBFKMXJBCUCAI-HKTJVKLFSA-N silibinin Chemical compound C1=C(O)C(OC)=CC([C@@H]2[C@H](OC3=CC=C(C=C3O2)[C@@H]2[C@H](C(=O)C3=C(O)C=C(O)C=C3O2)O)CO)=C1 SEBFKMXJBCUCAI-HKTJVKLFSA-N 0.000 description 1
- 238000010898 silica gel chromatography Methods 0.000 description 1
- 229960004245 silymarin Drugs 0.000 description 1
- 235000017700 silymarin Nutrition 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 125000006850 spacer group Chemical group 0.000 description 1
- 239000012086 standard solution Substances 0.000 description 1
- 230000007863 steatosis Effects 0.000 description 1
- 231100000240 steatosis hepatitis Toxicity 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 239000000829 suppository Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000003440 toxic substance Substances 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- ZBZJXHCVGLJWFG-UHFFFAOYSA-N trichloromethyl(.) Chemical compound Cl[C](Cl)Cl ZBZJXHCVGLJWFG-UHFFFAOYSA-N 0.000 description 1
- 229930182493 triterpene saponin Natural products 0.000 description 1
- 150000008130 triterpenoid saponins Chemical class 0.000 description 1
- 231100000397 ulcer Toxicity 0.000 description 1
- 238000004704 ultra performance liquid chromatography Methods 0.000 description 1
- 230000035899 viability Effects 0.000 description 1
- 230000009385 viral infection Effects 0.000 description 1
- 210000001835 viscera Anatomy 0.000 description 1
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 description 1
- 239000003643 water by type Substances 0.000 description 1
Images
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/48—Fabaceae or Leguminosae (Pea or Legume family); Caesalpiniaceae; Mimosaceae; Papilionaceae
- A61K36/484—Glycyrrhiza (licorice)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/12—Ketones
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying
Landscapes
- Health & Medical Sciences (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Life Sciences & Earth Sciences (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Alternative & Traditional Medicine (AREA)
- Biotechnology (AREA)
- Botany (AREA)
- Medical Informatics (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Medicines Containing Plant Substances (AREA)
- Acyclic And Carbocyclic Compounds In Medicinal Compositions (AREA)
Abstract
本发明公开了胀果甘草乙醇提取物及甘草查耳酮A的医药新用途。发明人发现胀果甘草与其他药用甘草存在明显的化学成分差异，继而对胀果甘草乙醇提取物及其中重要成分甘草查耳酮A的系统研究，发现其具有保护氧化型细胞损伤、组织损伤及急性化学性肝损伤的显著活性。基于以上发现，这一提取物及该活性化合物或其药学上可接受的盐、酯、溶剂化物、立体异构体、互变异构体、前药以及其混合物可用于制备治疗/预防氧化性损伤和/或急性化学性肝损伤药物及保健品。
Description
技术领域
本发明涉及胀果甘草乙醇提取物及甘草查耳酮A的提取方法，以及胀果甘草提取物与甘草查耳酮A的保肝药物新用途，具体涉及它们在保护氧化性肝损伤、急性肝炎导致的急性肝损伤中的应用。
背景技术
肝脏是人体内脏中最大的实质性器官，担负解毒、储存肝糖、代谢脂质等重要生理功能，为维持机体生命活动发挥至关重要的作用。诸多因素都会对肝脏造成损伤，常见的有药物损伤、肝炎等病毒感染、酒精摄取过量、黄曲霉毒素等有毒化学物质摄入，等等。长期的肝损伤往往会导致不同程度的肝细胞坏死、脂肪变性、肝硬化甚至肝癌。因此，预防和治疗肝损伤是临床上肝病治疗的重要环节之一，是减少肝纤维化、肝坏死、肝硬化以及肝癌等疾病发生和发展的基础。临床上有许多保肝药物及方法，但由于目前的治疗手段常常存在毒副作用，各类保肝药的使用受到限制。因此，从传统天然药物中发现有效的新型保肝药物具有重要的现实意义和良好的应用前景。
四氯化碳(CCl4)是一种常用的肝毒性试剂，研究表明，四氯化碳进入体内经过肝微粒体细胞色素P450酶代谢，形成三氯甲基自由基，造成脂质过氧化、氨基酸功能基团受损、核酸转化和突变，致使细胞的正常生命活动受到破坏。四氯化碳造成的急性损伤严重时可能导致动物死亡，长期的慢性损伤会造成明显的肝纤维化症状。四氯化碳造成的急性和慢性肝损伤模型广泛用于保肝药物研究。
甘草(Glycyrrhizae Radix et Rhizoma)是常用中药，具有保肝、抗溃疡、抗病毒、抗氧化、抗炎、抗菌、抗疟等药理活性，含有黄酮、黄酮苷、三萜皂苷等化学成分。中国药典(2015年版)规定甘草来源于豆科植物乌拉尔甘草Glycyrrhiza uralensis Fisch.、胀果甘草G.inflata Bat.和光果甘草G.glabra L.的干燥根和根茎。我国甘草的主流品种为乌拉尔甘草，研究较多。胀果甘草的化学成分与药理活性研究相对较少。我们的研究表明，胀果甘草与光果甘草、乌拉尔甘草提取物的低极性部位存在明显的化学成分差异，且对于一些药理模型，其活性明显优于光果甘草和乌拉尔甘草。
甘草及其制剂在临床广泛地应用于保肝。然而，大多数文献并未明确地标明甘草的原植物品种(即三种甘草属植物不予区分)；少量研究虽然明确了甘草的种属，但大多来自乌拉尔甘草和光果甘草(Jung等，BMC Complem Altern M,2016,16,DOI:10.1186/s12906-016-0997-0；Yin G等，Fish Physiol Biochem,2011,37,209-216)。胀果甘草提取物的保肝活性少有报道，仅有6个三萜皂苷类成分被报道在细胞模型上有较好的保肝活性(Zheng YF等，Molecules,2015,20,6273-6283)。对于胀果甘草的低极性提取物及单体成分，主要是游离酚类化合物，其保肝活性未见报道。
甘草查耳酮A是胀果甘草中含量最高的游离酚类成分之一，是一种连有异戊烯基的查耳酮类化合物，被报道具有抗肿瘤、抗炎、抗氧化、抗真菌、调节免疫等多种药理活性，并经小鼠和细胞实验证实有调节肝脂质代谢的活性(Quan HY等,Fitoterapia 2013,86,208-216)。甘草查耳酮A保护氧化性损伤和化学性肝损伤的活性未见报道。
本发明公开胀果甘草提取物及甘草查耳酮A的多种保肝药理活性。
发明内容
本发明的目的在于克服现有的技术及资源局限，提供了胀果甘草乙醇提取物及甘草查耳酮A保肝新用途。
本发明之胀果甘草提取物，其特征在于，提取对象为胀果甘草的根和根茎。
本发明之胀果甘草提取物，其特征在于，提取过程为：称取胀果甘草，粉碎。加入5～10倍体积的95％乙醇，加热回流3～4h，抽滤。滤渣按上法分别再用95％和75％乙醇提取1次。合并提取液，减压浓缩至无醇味，得总浸膏，即为胀果甘草乙醇提取物。浸膏用适量的去离子水混悬，用2～3倍量乙酸乙酯萃取4遍。回收乙酸乙酯，得到乙酸乙酯部位浸膏，即为胀果甘草提取物乙酸乙酯相浸膏。
本发明人在对胀果甘草进行提取后，研究了其对Nrf2的激活作用，以及其在动物体内保护CCl4造成的急性肝损伤活性，结果显示胀果甘草提取物具有激活Nrf2的抗氧化作用，以及降低CCl4造成的急性肝炎肝损伤作用，并且降低肝损伤导致的转氨酶升高的保肝活性。
本发明之胀果甘草提取物可用于制备具有保肝作用的药物和保健食品，在剂型方面可采用各种剂型，如片剂、颗粒剂、散剂、胶囊或丸剂等剂型。
本发明之胀果甘草提取物可用于制备具有保肝作用的药物和保健食品，在形状方面可采用各种形状。
本发明提供了甘草中化合物甘草查耳酮A的保肝新用途。此化合物的结构式如下：
本发明所提供的化合物的药物新用途指化合物本身或其药学上可接受的盐、酯、溶剂化物、立体异构体、互变异构体、前药中的至少一种，以及其混合物在制备具有如下(1)-(2)中至少一种功能的药物中的应用：(1)用于预防和治疗氧化造成的细胞损伤、组织损伤；(2)预防和治疗化学性肝损伤。
以上述活性化合物或其药学上可接受的盐、酯、溶剂化物、立体异构体、互变异构体、前药以及它的混合物为有效成分制备的药物也属于本发明的保护范围。
需要的时候，在上述药物中还可以加入一种或多种药学上可接受的载体或辅料。所述载体或辅料包括药学领域常规的稀释剂、赋形剂、填充剂、粘合剂、湿润剂、崩解剂、吸收促进剂、表面活性剂、吸附载体、润滑剂等。
利用上述活性化合物或其药学上可接受的盐、酯、溶剂化物、立体异构体、互变异构体以及前药作为活性成分，单独或组合使用，或与其它药物、辅料等配合制备成各种剂型，包括但不限于片剂、散剂、丸剂、注射剂、胶囊剂、膜剂、栓剂、膏剂、冲剂等多种形式。上述各种剂型的药物均可以按照药学领域的常规方法制备。
附图说明
图1为胀果甘草药材的DNA品种鉴定结果。
图2为胀果甘草、光果甘草、乌拉尔甘草乙醇提取物乙酸乙酯相浸膏的指纹图谱。
图3为甘草查耳酮A的氢谱。
图4为甘草查耳酮A的碳谱。
图5为胀果甘草提取物和甘草查耳酮A的Nrf2激活活性。
图6为胀果甘草、光果甘草、乌拉尔甘草乙醇提取物的Nrf2激活活性，
*p<0.05，**p<0.01，***p<0.001与第一批胀果甘草组比较。
图7为甘草查耳酮A对HepG2、MCF7、SW480、A549四种细胞的细胞毒性。
图8为胀果甘草提取物和甘草查耳酮A对四氯化碳造成的HepG2细胞急性损伤的保护活性，*p<0.05，**p<0.01，***p<0.001与模型组比较。
图9为胀果甘草提取物和甘草查耳酮A对四氯化碳造成的小鼠急性肝损伤的保护活性，*p<0.05，**p<0.01，***p<0.001与模型组比较。
图10为胀果甘草提取物和甘草查耳酮A对四氯化碳造成的小鼠急性肝损伤的保护活性中各组动物肝脏HE染色病理切片，其中1)对照组，2)模型组，3)阳性药组，4)甘草查耳酮A高剂量保护组，5)甘草查耳酮A低剂量保护组，6)甘草提取物高剂量保护组，7)甘草提取物低剂量保护组。
具体实施方式
下面结合实施例进一步阐述本发明，应理解，以下实施例仅用于说明本发明而不用于限制本发明的保护范围。
下述实施例中所使用的实验方法如无特殊说明，均为常规方法。
下述实施例中所用的材料、试剂等，如无特殊说明，均可从商业途径得到。
实施例1、胀果甘草的DNA品种鉴定。
一、实验材料和方法。
用植物基因组DNA提取试剂盒(天根生化科技有限公司，北京)提取样品的总DNA，PCR扩增其ITS序列及trnH-psbA转录间隔区序列并进行测序。引物序列为：ITS-F,GAAGGATCATTGTCGATGCC；ITS-R,GCGTTCAAAGACGCCTATTGG；trnH-forward，ACG GGA ATT GAACCC GCG CA；Gly-trnHR1，CAT ATG ACT TCA CAA TGT AAA ATC。
二、实验结果。
胀果甘草的DNA品种鉴定结果如图1所示。根据文献总结的甘草品种与基因型之间的对应关系(Kondo K,Biol Pharm Bull 2007,30,1497-1502)，该样品ITS基因型为I-2型，trnH-psbA为T-3型，故鉴定为胀果甘草。
实施例2、胀果甘草成分分析及胀果甘草浸膏、LCA的制备。
一、实验材料和方法。
本方法中提及的所有化学试剂均购自北京化工厂。
提取物制备：称取胀果甘草干燥根20kg，粉碎。加入5～10倍量95％乙醇，加热回流3～4h，抽滤。滤渣按上法分别再用95％和75％乙醇提取1次。合并提取液，减压浓缩至无醇味，得总浸膏，即为胀果甘草乙醇提取物。浸膏用适量的去离子水混悬，用2～3倍量乙酸乙酯萃取4遍。回收乙酸乙酯，得到乙酸乙酯部位浸膏。
甘草查耳酮A分离：取乙酸乙酯部位浸膏溶解，加入900g硅胶(100-200目)拌样，分两次上样于6kg硅胶柱(100-200目)，采用石油醚-乙酸乙酯(100:0,50:1,20:1,10:1,8:1,6:1,4:1,2:1,1:1,0:1,v/v)梯度洗脱，根据TLC和HPLC检测结果合并成7个流份(A-G)。流份E主要含查耳酮A，经硅胶柱色谱分离，以石油醚-乙酸乙酯(100:0,50:1,10:1,6:1,4:1,2:1,1:1v/v)作为流动相梯度洗脱，经TLC和HPLC检测结果合并得到3个流份(E1-E3)。流份E3先过聚酰胺柱，以二氯甲烷-甲醇(100:0,50:1,10:1,6:1,4:1,2:1,1:1v/v)作为流动相梯度洗脱；再用甲醇溶解得到主含LCA的流份，过LH-20柱纯化，甲醇作洗脱剂，最后经半制备HPLC纯化得到甘草查耳酮A。
提取物检测：称取甘草提取物粉末2mg，溶于1mL甲醇中，用0.22μm微孔滤膜过滤，进样。
色谱条件：
仪器：Agilent 1290高效液相色谱仪；
色谱柱：Acquity UPLC HSS T3柱(1.8μm,2.1×150mm)，VanGuard预柱(1.8μm,2.1×5mm)(Waters,MA,USA)；
流动相：乙腈(A)-0.1％甲酸(B)；
洗脱程序：0-7.5min,10-25％A；7.5-38min,25-38％A；38-53min,38-47％A；53-64min,47-57％A；64-72min,57-75％A；72-75min,75-80％A,85min停止。
检测波长：254nm。
二、实验结果。
同法提取的胀果甘草、光果甘草、乌拉尔甘草三种甘草乙醇提取物乙酸乙酯相浸膏成分指纹图谱见图2。
甘草查耳酮A的氢谱和碳谱见图3-4。
实施例3、胀果甘草提取物及甘草查耳酮A对Nrf2的激活作用。
一、实验材料和方法。
HepG2细胞采用Lipofectamine 2000(Life Technologies公司)稳定转染Nrf2萤光素酶报告基因，得到的细胞称之为HepG2C8细胞。HepG2C8细胞用10％胎牛血清的MEM培养基培养，维持温度37℃，5％CO2饱和湿度，使用0.25％胰蛋白酶-EDTA消化传代，按2×105/孔接种于24孔板12小时后，吸去培养基，给药处理。
给药处理：每孔分别加入含2.5μM、5μM、10μM、20μM、40μM甘草查耳酮A或50μg/mL甘草提取物的MEM培养基500μL，平行2个副孔。空白组加入含等量DMSO的MEM培养基500μL。培养6小时。
报告基因检测：吸去孔中的培养基，每孔加入200μL的PBS清洗，再分别加入100μL报告基因裂解液，4℃裂解10min，用枪头将孔底细胞全部刮下，与裂解液一起转移入1.5mLEP管中，4℃10000g离心10min，上清分别进行以下两部分操作：
1.蛋白浓度的测定：
标准曲线的制备：96孔板中分别加入0,2,4,8,10μL 1mg/mL牛血清白蛋白标准液，用超纯水补足至20μL。
样品蛋白浓度测试：96孔板中分别加入5μL上清，加15μL超纯水补足20μL。
A液和B液按50：1配制BCA溶液，每孔中加200μL，37℃烘箱中孵育30min，酶标仪测定570nm下各孔吸光度。
2.萤光素酶检测：
分别吸取50μL样品上清于96孔白板中，空白对照加入50μL平行空白报告基因裂解液，用酶标仪检测。
萤光素酶活性强度计算：激活强度＝(样品每孔萤光强度值-空白对照孔的平均萤光强度值)/(对应的蛋白浓度×DMSO孔平均萤光强度值)
分别计算同一化合物两平行样品结果的平均值及其SD值，即得该化合物对报告基因的激活强度及其SD值。
另取不同批次的胀果甘草、光果甘草、乌拉尔甘草的乙醇提取物进行实验，测定其Nrf2激活活性。
二、实验结果
Nrf2是调节体内氧化还原平衡的重要转录因子，其通过抗氧化响应元件(ARE)控制细胞II相药物代谢酶和抗氧化基因的表达，激活Nrf2信号通路可以减轻各种因素引起的细胞氧化损伤。本实施例采用Nrf2萤光素酶报告基因的方法，测定甘草提取物及甘草查耳酮A的Nrf2激活作用。如图5所示，50μg/mL甘草提取物对Nrf2报告基因有激活作用，达到3.8倍。甘草查耳酮A也可以剂量依赖地激活Nrf2报告基因活性，在10μM时的活性最强，达到4.07倍。
对不同批次胀果甘草、光果甘草、乌拉尔甘草的乙醇提取物Nrf2激活活性比较，如图6所示，两批胀果甘草提取物的活性均达到2.8倍以上，而光果甘草和乌拉尔甘草提取物活性最大值只有1.7倍。胀果甘草对Nrf2的激活活性明显高于光果甘草和乌拉尔甘草。
实施例4、胀果甘草浸膏及甘草查耳酮A对HepG2细胞活力的影响。
一、实验材料和方法。
HepG2人肝癌细胞购自美国菌种保存中心(ATCC)。HepG2细胞用10％胎牛血清的DMEM培养基培养，维持温度37℃，5％CO2饱和湿度，使用0.25％胰蛋白酶-EDTA消化传代，按1×104/孔接种于96孔板12小时后，吸去培养基。分别加入含10μM甘草查耳酮A的DMEM培养基100μL，继续培养24小时，向每孔加入10μL MTS溶液(美国Promega公司)，继续培养2～4小时，酶标仪测定490nm下各孔吸光度。
以空白组为0％，对照组为100％，分别计算各组的细胞活力抑制率及SD值。
同样方法对甘草查耳酮A的MCF7、SW480、A549的细胞毒性进行测试。
二、实验结果
10μM甘草查耳酮A对HepG2、MCF7、SW480、A549的细胞毒性均较弱，如图7所示。10μM甘草查耳酮A对HepG2细胞活力抑制率为13.5％，对MCF7和SW480的细胞活性抑制率分别为9.8％，7.0％，细胞毒性较低；而对A549的抑制率甚至为0，即几乎无细胞毒性。
实施例5、甘草查耳酮A对四氯化碳导致肝细胞HepG2损伤的保护作用。
一、实验材料和方法。
四氯化碳储液配置：DMSO与四氯化碳按1：1(v/v)混合，涡旋得到50％CCl4储备液。向DMEM培养基中加入储备液得到含四氯化碳浓度0.35％的培养基。
HepG2人肝癌细胞购自美国菌种保存中心(ATCC)。HepG2培养于含10％胎牛血清的DMEM培养基中，按1×104/孔种于96孔板中，培养12h后将培养基吸出，分组加入以下液体(100μL/孔，3复孔/组)：
空白组：不含四氯化碳的空白DMEM培养基(加入不含细胞空白孔)；
空白对照组：含与加药组同等浓度的DMSO的空白DMEM培养基；
模型组：含与加药组同等浓度的DMSO的含0.35％四氯化碳的DMEM培养基；
给药组：加入终浓度为10μM甘草查耳酮A的含0.35％四氯化碳的DMEM培养基；
培养6h后，每孔加入10μL MTS溶液，避光孵育2～4h，490nm下测各孔吸光度。
以空白组为0％，对照组为100％，分别计算各组的细胞存活率及SD值，对化合物的保护作用进行评价，由吸光度值计算得到各组孔中细胞存活率。
二、实验结果。
如图8所示，10μM甘草查耳酮A对0.35％CCl4损伤的HepG2起到显著保作用，可以将四氯化碳损伤后的细胞存活率从53％升高至75％，且保护作用呈现明显的剂量依赖性。说明甘草查耳酮A具有显著保护急性肝损伤活性，可用于制备保护急性肝炎肝损伤药物。
实施例6、胀果甘草提取物及甘草查耳酮A对四氯化碳造成的急性肝损伤的保护作用。
一、实验材料和方法。
4-6周雄性ICR鼠购自北京大学医学部实验动物部，给予12h光照12h黑暗，恒温，正常饲料及水饲养，于无菌环境中适应4天后进行实验。
动物随机分为7组(n＝6)，1)对照组及2)模型组动物每天按10ml/kg灌胃0.3％羧甲基纤维素钠溶液，3)阳性药组每天按10ml/kg灌胃10mg/ml的水飞蓟素混悬液(混于0.3％羧甲基纤维素钠溶液)，4)甘草查耳酮A低剂量保护组及5)甘草查耳酮A高剂量保护组每天按10ml/kg分别灌胃1mg/ml、5mg/ml的甘草查耳酮A混悬液(给药剂量10mg/kg、50mg/kg)，6)胀果甘草提取物低剂量保护组、7)胀果甘草提取物高剂量保护组每天按10ml/kg分别灌胃5mg/ml、20mg/ml的胀果甘草提取物混悬液(给药剂量50mg/kg、200mg/kg)。连续给药7天，第7天灌胃6h后：1)对照组腹腔注射玉米油(7ml/kg)，2)～9)组腹腔注射CCl4造模(0.2％v/v,7ml/kg,溶于玉米油)，24h后摘眼球取血，心脏灌流后取肝组织。血液样品室温静置2h后4℃8000g离心15min取血清。
血清样品进行血生化分析，检测ALT、AST、LDH指标。肝脏组织经多聚甲醛固定后进行石蜡包埋、切片、HE染色、拍照。
二、实验结果。
实验数据计量资料均以均数±标准差表示，两均数的比较使用SPSS16.0统计软件中的one-way ANOVA分析，*p<0.05，**p<0.01，***p<0.001与模型组比较。
肝细胞受损后会释放谷丙转氨酶(ALT)，谷草转氨酶(AST)和乳酸脱氢酶(LDH)入血，血清中的这些酶浓度反应肝损伤程度。如图9所示，血生化结果显示胀果甘草提取物及甘草查耳酮A均有明显的保护效果以及剂量依赖，显著保护了四氯化碳造成的急性肝损伤。
在光学显微镜下，无损伤的小鼠肝组织切片中，可见肝细胞大小、形态均正常，无肝实质性细胞病变，中央静脉及汇管区正常，如图10所示。四氯化碳损伤后的模型组，小鼠肝组织中中央静脉周围肝细胞无正常细胞形态，肝细胞肿大，部分细胞呈气球样变性，有炎症细胞浸润现象。经给药胀果甘草提取物或甘草查耳酮A后，肝细胞坏死程度显著减轻，炎症浸润程度降低，低剂量给药组部分细胞仍呈现气球样变性，高剂量给药组几乎无气球样变性。与血生化结果吻合，说明胀果甘草提取物及甘草查耳酮A均有明显保护肝损伤的效果。
Claims (3)
1.下式所示的化合物或其药学上可接受的盐中的至少一种或混合物在制备治疗/预防化学性肝损伤药物中的应用
2.如权利要求1中所述的应用，其特征在于化学性肝损伤类型为四氯化碳诱发的肝损伤。
3.如权利要求1中所述的应用，其特征在于治疗/预防化学性肝损伤的化合物，其进一步包含医药上可接受的载体。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610894553.5A CN106420902B (zh) | 2016-10-13 | 2016-10-13 | 胀果甘草提取物及甘草查耳酮a的保肝活性及药物新用途 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610894553.5A CN106420902B (zh) | 2016-10-13 | 2016-10-13 | 胀果甘草提取物及甘草查耳酮a的保肝活性及药物新用途 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106420902A CN106420902A (zh) | 2017-02-22 |
| CN106420902B true CN106420902B (zh) | 2020-01-03 |
Family
ID=58175141
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610894553.5A Active CN106420902B (zh) | 2016-10-13 | 2016-10-13 | 胀果甘草提取物及甘草查耳酮a的保肝活性及药物新用途 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106420902B (zh) |
Families Citing this family (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109674848A (zh) * | 2019-02-22 | 2019-04-26 | 毕节市第一人民医院 | 一种甘草提取物的制备方法与用途 |
| CN112999260A (zh) * | 2021-03-18 | 2021-06-22 | 华南农业大学 | 一种降低动物氧化应激损伤的中药提取物及其制备方法和应用 |
| CN117100739B (zh) * | 2022-05-17 | 2024-04-12 | 北京慧宝源生物技术股份有限公司 | 负调节Nrf2信号通路的小分子组合物及其应用 |
| CN114887069B (zh) * | 2022-06-07 | 2024-06-07 | 广州白云山敬修堂药业股份有限公司 | 一种降低药物肝毒性的甘草炭包衣材料及其制备方法和用途 |
| CN115317386A (zh) * | 2022-09-21 | 2022-11-11 | 江西沁华蔓医药科技有限公司 | 提拉紧致抗衰老精华液及其制备方法 |
| CN115531276A (zh) * | 2022-11-09 | 2022-12-30 | 江西沁华蔓医药科技有限公司 | 透皮保湿精华液及其制备方法 |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2009269889A (ja) * | 2008-05-09 | 2009-11-19 | Maruzen Pharmaceut Co Ltd | グルタチオン産生促進剤、グルタチオンの欠乏に起因する疾患の予防・治療剤及び飲食品 |
| JP2013103925A (ja) * | 2011-11-16 | 2013-05-30 | Minofuaagen Seiyaku:Kk | 肝臓線維化抑制剤 |
| CN103547262A (zh) * | 2011-04-21 | 2014-01-29 | 庆熙大学校产学协力团 | 甘草查耳酮a的新用途 |
-
2016
- 2016-10-13 CN CN201610894553.5A patent/CN106420902B/zh active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2009269889A (ja) * | 2008-05-09 | 2009-11-19 | Maruzen Pharmaceut Co Ltd | グルタチオン産生促進剤、グルタチオンの欠乏に起因する疾患の予防・治療剤及び飲食品 |
| CN103547262A (zh) * | 2011-04-21 | 2014-01-29 | 庆熙大学校产学协力团 | 甘草查耳酮a的新用途 |
| JP2013103925A (ja) * | 2011-11-16 | 2013-05-30 | Minofuaagen Seiyaku:Kk | 肝臓線維化抑制剤 |
Non-Patent Citations (4)
| Title |
|---|
| Lico A enhances Nrf2-mediated defense mechanisms against t-BHP-induced oxidative stress and cell death via Akt and ERK activation in Raw 264.7 cells;Hongming Lv等;《Oxidative medicine and cellular longevity》;20151231;第2015卷;第1页摘要 * |
| Licochalcone A regulates hepatic lipid metabolism through activation of AMP-activated protein kinase;Hai Yan Quan等;《Fitoterapia》;20130314;第86卷;第208页摘要 * |
| 甘草黄酮的分离鉴定、药效及其指纹图谱研究;崔永明;《中国博士学位论文全文数据库 医药卫生科技辑》;20091215(第12期);第E057-13页 正文第5页左栏第3个结构式、第8页 第1.4.4节第1段 * |
| 胀果甘草化学成分的分离与鉴定;孙良明;《安徽医药》;20131231;第17卷(第7期);第1121页摘要，右栏第2-3段、图1 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106420902A (zh) | 2017-02-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106420902B (zh) | 胀果甘草提取物及甘草查耳酮a的保肝活性及药物新用途 | |
| Zhang et al. | Extraction of bio-active components from Rhodiola sachalinensis under ultrahigh hydrostatic pressure | |
| Shaker et al. | Silymarin, the antioxidant component and Silybum marianum extracts prevent liver damage | |
| Bhardwaj et al. | Rhodiola: An overview of phytochemistry and pharmacological applications | |
| CN107854522B (zh) | 一种组合物及其制备方法和用途 | |
| CN110772547A (zh) | 文王一支笔提取物在制备治疗肝炎药物中的应用 | |
| CN107137434B (zh) | 一种药物组合物及其制备方法和在制备抗柯萨奇病毒药物中的应用 | |
| CN108096307B (zh) | 野马追乙醇提取物在制备抗乙肝病毒药物中的应用 | |
| CN101028322B (zh) | 毛裂蜂斗菜提取物用于制备防治肿瘤疾病药物的用途 | |
| CN1803179B (zh) | 一种中药组合物及其制备方法和有效成分的检测方法 | |
| US20160213730A1 (en) | Identification of natural plant extracts harboring anti-hepatitis c virus ns5b polymerase activity | |
| CN107129478B (zh) | 一种半萜内酯类化合物及其制备方法和应用 | |
| CN102626465B (zh) | 一种龙柴方微丸及其制备方法 | |
| Gamde et al. | Histopathological and biochemical effects of aqueous leaf extract of Cadaba farinosa on the liver of adult wistar rats | |
| Wang et al. | Chemical constituents and hepatoprotective properties of Rhododendron simsii Planch extract in Con A-induced autoimmune hepatitis | |
| CN101500587A (zh) | 抗病毒产品 | |
| CN112618584A (zh) | 四块瓦提取物在制备抗乙型肝炎病毒和/或防治乙型肝炎的药物中的应用 | |
| CN107434795B (zh) | 一种预防脂肪肝的化合物及其制备方法和应用 | |
| Alavian et al. | Influence of Nutrients, Bioactive Compounds, and Plant Extracts in Liver Diseases | |
| CN108379495B (zh) | 高良姜提取物在制备预防和/或治疗非酒精性脂肪肝制剂中的用途 | |
| CN1969937B (zh) | 一种治疗肝炎的药物组合物 | |
| Jian-bo et al. | Anti-Hepatitis B virus activity of flavonoids from Marchantia convoluta | |
| CN100574764C (zh) | 一种抗肝炎药物组合物及其制备方法 | |
| Sodani | Histopathological changes of mice liver induced by an Aloe vera whole leaf extract | |
| CN110885385B (zh) | 翼首草毒素a及其应用和低肝损伤毒性翼首草提取物的制备方法 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |