CN112876522A - Method for extracting saussurea involucrate flavonoid composition - Google Patents
Method for extracting saussurea involucrate flavonoid composition Download PDFInfo
- Publication number
- CN112876522A CN112876522A CN201911205352.XA CN201911205352A CN112876522A CN 112876522 A CN112876522 A CN 112876522A CN 201911205352 A CN201911205352 A CN 201911205352A CN 112876522 A CN112876522 A CN 112876522A
- Authority
- CN
- China
- Prior art keywords
- column
- flavonoid composition
- eluent
- extraction method
- components
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 229930003935 flavonoid Natural products 0.000 title claims abstract description 48
- 150000002215 flavonoids Chemical class 0.000 title claims abstract description 48
- 235000017173 flavonoids Nutrition 0.000 title claims abstract description 48
- 239000000203 mixture Substances 0.000 title claims abstract description 44
- 241000133134 Saussurea Species 0.000 title claims abstract description 20
- 238000000034 method Methods 0.000 title claims description 19
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 53
- 238000000605 extraction Methods 0.000 claims abstract description 22
- BNWJOHGLIBDBOB-UHFFFAOYSA-N myristicin Chemical compound COC1=CC(CC=C)=CC2=C1OCO2 BNWJOHGLIBDBOB-UHFFFAOYSA-N 0.000 claims abstract description 20
- IVCZEZUJCMWBBR-UHFFFAOYSA-N 7-O-beta-D-glucopyranosyl-7,3',4'-trihydroxyflavone Natural products OC1C(O)C(O)C(CO)OC1OC1=CC=C2C(=O)C=C(C=3C=C(O)C(O)=CC=3)OC2=C1 IVCZEZUJCMWBBR-UHFFFAOYSA-N 0.000 claims abstract description 10
- JMGZEFIQIZZSBH-UHFFFAOYSA-N Bioquercetin Natural products CC1OC(OCC(O)C2OC(OC3=C(Oc4cc(O)cc(O)c4C3=O)c5ccc(O)c(O)c5)C(O)C2O)C(O)C(O)C1O JMGZEFIQIZZSBH-UHFFFAOYSA-N 0.000 claims abstract description 10
- OVSQVDMCBVZWGM-IDRAQACASA-N Hirsutrin Natural products O([C@H]1[C@H](O)[C@H](O)[C@H](O)[C@@H](CO)O1)C1=C(c2cc(O)c(O)cc2)Oc2c(c(O)cc(O)c2)C1=O OVSQVDMCBVZWGM-IDRAQACASA-N 0.000 claims abstract description 10
- FVQOMEDMFUMIMO-UHFFFAOYSA-N Hyperosid Natural products OC1C(O)C(O)C(CO)OC1OC1C(=O)C2=C(O)C=C(O)C=C2OC1C1=CC=C(O)C(O)=C1 FVQOMEDMFUMIMO-UHFFFAOYSA-N 0.000 claims abstract description 10
- IVTMALDHFAHOGL-UHFFFAOYSA-N eriodictyol 7-O-rutinoside Natural products OC1C(O)C(O)C(C)OC1OCC1C(O)C(O)C(O)C(OC=2C=C3C(C(C(O)=C(O3)C=3C=C(O)C(O)=CC=3)=O)=C(O)C=2)O1 IVTMALDHFAHOGL-UHFFFAOYSA-N 0.000 claims abstract description 10
- GXMWXESSGGEWEM-UHFFFAOYSA-N isoquercitrin Natural products OCC(O)C1OC(OC2C(Oc3cc(O)cc(O)c3C2=O)c4ccc(O)c(O)c4)C(O)C1O GXMWXESSGGEWEM-UHFFFAOYSA-N 0.000 claims abstract description 10
- PEFNSGRTCBGNAN-QNDFHXLGSA-N luteolin 7-O-beta-D-glucoside Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=CC(O)=C2C(=O)C=C(C=3C=C(O)C(O)=CC=3)OC2=C1 PEFNSGRTCBGNAN-QNDFHXLGSA-N 0.000 claims abstract description 10
- QZOVLVSTWSTHQN-UHFFFAOYSA-N luteolin 7-O-glucoside Natural products OCC1OC(Oc2cc(O)c3C(=O)C=C(C(=O)c3c2)c4ccc(O)c(O)c4)C(O)C(O)C1O QZOVLVSTWSTHQN-UHFFFAOYSA-N 0.000 claims abstract description 10
- KBGKQZVCLWKUDQ-UHFFFAOYSA-N luteolin-glucoside Natural products OC1C(O)C(O)C(CO)OC1OC1=CC(O)=CC2=C1C(=O)C=C(C=1C=C(O)C(O)=CC=1)O2 KBGKQZVCLWKUDQ-UHFFFAOYSA-N 0.000 claims abstract description 10
- OVSQVDMCBVZWGM-QSOFNFLRSA-N quercetin 3-O-beta-D-glucopyranoside Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C(C=2C=C(O)C(O)=CC=2)OC2=CC(O)=CC(O)=C2C1=O OVSQVDMCBVZWGM-QSOFNFLRSA-N 0.000 claims abstract description 10
- FDRQPMVGJOQVTL-UHFFFAOYSA-N quercetin rutinoside Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC=2C(C3=C(O)C=C(O)C=C3OC=2C=2C=C(O)C(O)=CC=2)=O)O1 FDRQPMVGJOQVTL-UHFFFAOYSA-N 0.000 claims abstract description 10
- IKGXIBQEEMLURG-BKUODXTLSA-N rutin Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](C)O[C@@H]1OC[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](OC=2C(C3=C(O)C=C(O)C=C3OC=2C=2C=C(O)C(O)=CC=2)=O)O1 IKGXIBQEEMLURG-BKUODXTLSA-N 0.000 claims abstract description 10
- ALABRVAAKCSLSC-UHFFFAOYSA-N rutin Natural products CC1OC(OCC2OC(O)C(O)C(O)C2O)C(O)C(O)C1OC3=C(Oc4cc(O)cc(O)c4C3=O)c5ccc(O)c(O)c5 ALABRVAAKCSLSC-UHFFFAOYSA-N 0.000 claims abstract description 10
- 235000005493 rutin Nutrition 0.000 claims abstract description 10
- 229960004555 rutoside Drugs 0.000 claims abstract description 10
- 238000004108 freeze drying Methods 0.000 claims abstract description 4
- 235000019441 ethanol Nutrition 0.000 claims description 20
- 239000003480 eluent Substances 0.000 claims description 16
- 239000011347 resin Substances 0.000 claims description 14
- 229920005989 resin Polymers 0.000 claims description 14
- 241001145025 Saussurea involucrata Species 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 239000002904 solvent Substances 0.000 claims description 8
- 238000010828 elution Methods 0.000 claims description 7
- 239000012535 impurity Substances 0.000 claims description 6
- 238000002156 mixing Methods 0.000 claims description 6
- 239000000843 powder Substances 0.000 claims description 5
- 239000012488 sample solution Substances 0.000 claims description 5
- 238000005406 washing Methods 0.000 claims description 5
- 238000011049 filling Methods 0.000 claims description 4
- 239000007789 gas Substances 0.000 claims description 4
- 239000008367 deionised water Substances 0.000 claims description 3
- 229910021641 deionized water Inorganic materials 0.000 claims description 3
- 238000001914 filtration Methods 0.000 claims description 3
- 238000011068 loading method Methods 0.000 claims description 3
- 238000002791 soaking Methods 0.000 claims description 3
- 241000196324 Embryophyta Species 0.000 claims description 2
- 238000010438 heat treatment Methods 0.000 claims description 2
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen group Chemical group [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 claims description 2
- 238000010298 pulverizing process Methods 0.000 claims description 2
- 238000005070 sampling Methods 0.000 claims description 2
- 239000003814 drug Substances 0.000 abstract description 39
- 229940079593 drug Drugs 0.000 abstract description 31
- 230000000694 effects Effects 0.000 abstract description 16
- 240000002853 Nelumbo nucifera Species 0.000 abstract description 14
- 235000006508 Nelumbo nucifera Nutrition 0.000 abstract description 14
- 235000006510 Nelumbo pentapetala Nutrition 0.000 abstract description 14
- 230000003285 pharmacodynamic effect Effects 0.000 abstract description 11
- 206010003246 arthritis Diseases 0.000 abstract description 6
- 238000002360 preparation method Methods 0.000 abstract description 6
- 238000011160 research Methods 0.000 abstract description 5
- 230000004044 response Effects 0.000 abstract description 2
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 50
- 241000699670 Mus sp. Species 0.000 description 33
- 238000012360 testing method Methods 0.000 description 20
- 229960000583 acetic acid Drugs 0.000 description 17
- 238000002474 experimental method Methods 0.000 description 16
- 241000699666 Mus <mouse, genus> Species 0.000 description 15
- 230000008961 swelling Effects 0.000 description 12
- 150000002500 ions Chemical class 0.000 description 11
- 241001465754 Metazoa Species 0.000 description 10
- 239000007924 injection Substances 0.000 description 10
- 238000002347 injection Methods 0.000 description 10
- 210000003371 toe Anatomy 0.000 description 10
- 238000010172 mouse model Methods 0.000 description 9
- 208000009386 Experimental Arthritis Diseases 0.000 description 8
- 239000002504 physiological saline solution Substances 0.000 description 8
- 239000000126 substance Substances 0.000 description 8
- 230000008569 process Effects 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- 210000003462 vein Anatomy 0.000 description 7
- 239000007788 liquid Substances 0.000 description 6
- 239000000463 material Substances 0.000 description 6
- 239000003643 water by type Substances 0.000 description 6
- 238000004458 analytical method Methods 0.000 description 5
- 150000001875 compounds Chemical class 0.000 description 5
- 238000003860 storage Methods 0.000 description 5
- 238000010586 diagram Methods 0.000 description 4
- 230000000857 drug effect Effects 0.000 description 4
- 210000002683 foot Anatomy 0.000 description 4
- 239000012362 glacial acetic acid Substances 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 238000003305 oral gavage Methods 0.000 description 4
- 239000000047 product Substances 0.000 description 4
- 239000000523 sample Substances 0.000 description 4
- 238000012353 t test Methods 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- 239000002671 adjuvant Substances 0.000 description 3
- 230000000202 analgesic effect Effects 0.000 description 3
- 238000010171 animal model Methods 0.000 description 3
- 230000037396 body weight Effects 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000013329 compounding Methods 0.000 description 3
- 239000000284 extract Substances 0.000 description 3
- 238000003304 gavage Methods 0.000 description 3
- 230000000762 glandular Effects 0.000 description 3
- 238000000465 moulding Methods 0.000 description 3
- 239000003826 tablet Substances 0.000 description 3
- 230000001225 therapeutic effect Effects 0.000 description 3
- 238000004704 ultra performance liquid chromatography Methods 0.000 description 3
- 238000011725 BALB/c mouse Methods 0.000 description 2
- 208000032843 Hemorrhage Diseases 0.000 description 2
- 206010023232 Joint swelling Diseases 0.000 description 2
- 102000011842 Serrate-Jagged Proteins Human genes 0.000 description 2
- 108010036039 Serrate-Jagged Proteins Proteins 0.000 description 2
- 238000000692 Student's t-test Methods 0.000 description 2
- XBJFCYDKBDVADW-UHFFFAOYSA-N acetonitrile;formic acid Chemical compound CC#N.OC=O XBJFCYDKBDVADW-UHFFFAOYSA-N 0.000 description 2
- 229930013930 alkaloid Natural products 0.000 description 2
- 230000000740 bleeding effect Effects 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 229960001193 diclofenac sodium Drugs 0.000 description 2
- 239000002552 dosage form Substances 0.000 description 2
- 235000013399 edible fruits Nutrition 0.000 description 2
- 239000007941 film coated tablet Substances 0.000 description 2
- HQVFCQRVQFYGRJ-UHFFFAOYSA-N formic acid;hydrate Chemical compound O.OC=O HQVFCQRVQFYGRJ-UHFFFAOYSA-N 0.000 description 2
- 238000003384 imaging method Methods 0.000 description 2
- 239000007928 intraperitoneal injection Substances 0.000 description 2
- 238000005259 measurement Methods 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
- 238000004806 packaging method and process Methods 0.000 description 2
- 239000012264 purified product Substances 0.000 description 2
- 230000000384 rearing effect Effects 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 238000007789 sealing Methods 0.000 description 2
- 239000008354 sodium chloride injection Substances 0.000 description 2
- JGMJQSFLQWGYMQ-UHFFFAOYSA-M sodium;2,6-dichloro-n-phenylaniline;acetate Chemical compound [Na+].CC([O-])=O.ClC1=CC=CC(Cl)=C1NC1=CC=CC=C1 JGMJQSFLQWGYMQ-UHFFFAOYSA-M 0.000 description 2
- 238000007619 statistical method Methods 0.000 description 2
- 239000007939 sustained release tablet Substances 0.000 description 2
- 239000012085 test solution Substances 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 238000005303 weighing Methods 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
- 241000208838 Asteraceae Species 0.000 description 1
- 241000131317 Capitulum Species 0.000 description 1
- 229920000742 Cotton Polymers 0.000 description 1
- 108010022337 Leucine Enkephalin Proteins 0.000 description 1
- 208000037093 Menstruation Disturbances Diseases 0.000 description 1
- 206010027339 Menstruation irregular Diseases 0.000 description 1
- 206010027514 Metrorrhagia Diseases 0.000 description 1
- 108010084238 NAD+ peroxidase Proteins 0.000 description 1
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical compound [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 description 1
- 206010053476 Traumatic haemorrhage Diseases 0.000 description 1
- 210000001015 abdomen Anatomy 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 235000013361 beverage Nutrition 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 235000019658 bitter taste Nutrition 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 239000002775 capsule Substances 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000012937 correction Methods 0.000 description 1
- 238000013480 data collection Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 229930182478 glucoside Natural products 0.000 description 1
- 150000008131 glucosides Chemical class 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 201000001881 impotence Diseases 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 238000007912 intraperitoneal administration Methods 0.000 description 1
- MWDZOUNAPSSOEL-UHFFFAOYSA-N kaempferol Natural products OC1=C(C(=O)c2cc(O)cc(O)c2O1)c3ccc(O)cc3 MWDZOUNAPSSOEL-UHFFFAOYSA-N 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 210000003127 knee Anatomy 0.000 description 1
- URLZCHNOLZSCCA-UHFFFAOYSA-N leu-enkephalin Chemical compound C=1C=C(O)C=CC=1CC(N)C(=O)NCC(=O)NCC(=O)NC(C(=O)NC(CC(C)C)C(O)=O)CC1=CC=CC=C1 URLZCHNOLZSCCA-UHFFFAOYSA-N 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 210000003141 lower extremity Anatomy 0.000 description 1
- LRDGATPGVJTWLJ-UHFFFAOYSA-N luteolin Natural products OC1=CC(O)=CC(C=2OC3=CC(O)=CC(O)=C3C(=O)C=2)=C1 LRDGATPGVJTWLJ-UHFFFAOYSA-N 0.000 description 1
- IQPNAANSBPBGFQ-UHFFFAOYSA-N luteolin Chemical compound C=1C(O)=CC(O)=C(C(C=2)=O)C=1OC=2C1=CC=C(O)C(O)=C1 IQPNAANSBPBGFQ-UHFFFAOYSA-N 0.000 description 1
- 235000009498 luteolin Nutrition 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 230000005906 menstruation Effects 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 125000000963 oxybis(methylene) group Chemical group [H]C([H])(*)OC([H])([H])* 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 230000005855 radiation Effects 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 201000003068 rheumatic fever Diseases 0.000 description 1
- 239000011435 rock Substances 0.000 description 1
- 229930004725 sesquiterpene Natural products 0.000 description 1
- 150000004354 sesquiterpene derivatives Chemical class 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 239000004575 stone Substances 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 235000019605 sweet taste sensations Nutrition 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 229910021642 ultra pure water Inorganic materials 0.000 description 1
- 239000012498 ultrapure water Substances 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
- C07H1/06—Separation; Purification
- C07H1/08—Separation; Purification from natural products
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P19/00—Drugs for skeletal disorders
- A61P19/02—Drugs for skeletal disorders for joint disorders, e.g. arthritis, arthrosis
-
- 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
- A61P25/04—Centrally acting analgesics, e.g. opioids
-
- 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
- C07D—HETEROCYCLIC COMPOUNDS
- C07D317/00—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms
- C07D317/08—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3
- C07D317/44—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D317/46—Heterocyclic compounds containing five-membered rings having two oxygen atoms as the only ring hetero atoms having the hetero atoms in positions 1 and 3 ortho- or peri-condensed with carbocyclic rings or ring systems condensed with one six-membered ring
- C07D317/48—Methylenedioxybenzenes or hydrogenated methylenedioxybenzenes, unsubstituted on the hetero ring
- C07D317/62—Methylenedioxybenzenes or hydrogenated methylenedioxybenzenes, unsubstituted on the hetero ring with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to atoms of the carbocyclic ring
- C07D317/64—Oxygen atoms
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H17/00—Compounds containing heterocyclic radicals directly attached to hetero atoms of saccharide radicals
- C07H17/04—Heterocyclic radicals containing only oxygen as ring hetero atoms
- C07H17/06—Benzopyran radicals
- C07H17/065—Benzo[b]pyrans
- C07H17/07—Benzo[b]pyran-4-ones
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicinal Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Rheumatology (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Pain & Pain Management (AREA)
- Molecular Biology (AREA)
- Neurosurgery (AREA)
- Biomedical Technology (AREA)
- Immunology (AREA)
- Orthopedic Medicine & Surgery (AREA)
- Neurology (AREA)
- Physical Education & Sports Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The invention relates to an extraction method of a snow lotus flavonoid composition, which comprises the following steps: (1) alcohol extraction; (2) fine extraction; (3) and (5) freeze-drying. The extraction method can obtain the saussurea involucrate flavonoid composition containing four components, wherein the four components are respectively as follows: rutin, myristicin, isoquercitrin, and luteolin-7-o-glucoside. Researches show that the flavonoid composition has pharmacodynamic activity, can be used for preparing various pain relieving and pain easing medicines and arthritis treatment medicines for clinical application, and has the effects of high pharmacodynamic activity, small clinical dosage, quick response, low side effect and the like compared with a Chinese patent medicine preparation prepared by directly applying saussurea involucrate.
Description
Technical Field
The invention relates to the technical field of compositions with medical application, in particular to an extraction method and application of a snow lotus flavonoid composition.
Background
Snow lotus (the name of Saussurea involucrata (kar. et Kir.) Sch. -Bip.) has the shape similar to lotus flower, so the name of snow lotus, which is called snow lotus for short, is obtained. Is perennial herb of saussurea of Compositae, and has a height of 15-35 cm. The rhizome is thick and the neck is left with most of the brown leaves. Thick and strong stem without hair. Dense leaves, basal leaves and cauline leaves without stems, oval leaves or oval ovate leaves; the most upper part is in the shape of leaf bud, membranous and light yellow, and surrounds the general inflorescence, and sharp teeth are arranged on the edge. 10-20 head inflorescences, and spherical total inflorescences are densely arranged at the top of the stem. The involucre is hemispherical and has the diameter of 1 cm; 3-4 layers of bract, purple brown at edge or all. The floret is purple. The slim fruit is oblong. The crown hair is white. The flower and fruit period is 7-9 months. It is distributed in the alpine region of many places in China, and Russia and Kazakhstan are also distributed. Grow in rock cracks, stone walls and gravel beaches near high mountain snow lines, and have an altitude of 2400 + 4000 meters. Tianshan produced the most and the best quality. Can be used as a medicine and also has certain ornamental value.
The snow lotus herb has warm nature, sweet and bitter taste, enters liver, spleen and kidney meridians, has the functions of dispelling cold, strengthening yang, regulating menstruation and stopping bleeding, and is used for treating impotence, weak waist and knees, irregular menstruation, metrorrhagia and metrostaxis, leukorrhagia, rheumatic arthritis, traumatic hemorrhage and other diseases. Has effects in dredging channels, promoting blood circulation, expelling cold, removing dampness, stopping bleeding, relieving swelling, and removing toxic materials. Snow lotus herb contains protein, amino acid, flavonoid, alkaloid, etc. However, how to generate and take effect of the unique drug effect is always a difficult point and a blind point in the research field.
Disclosure of Invention
The snow lotus herb flavonoid composition takes snow lotus herb as a research object, deeply researches main pharmacodynamic components of snow lotus herb, successfully discovers the most important pharmacodynamic components and compatibility of the snow lotus herb, and provides the snow lotus herb flavonoid composition and the application thereof on the basis.
The property of the saussurea involucrate adopted by the invention is observed by naked eyes as follows: the stem is upright and hairless. Basal leaves are not seen; the leaf is oblong or oval, 7-20 cm long, 3-6 cm wide, and has serrate edge and glandular hair on both sides; the cauline leaves have no stems; the uppermost part is in the shape of bract, membranous, yellow, oblong or oval long round, 16 cm long, 7 cm wide, blunt top, with serrate edge, and the two sides are surrounded by short mollissima and glandular hair, which surround the total inflorescence. 13 capitulums, spherical total inflorescences with no small pedicel or short small pedicel at the stem tip. The involucre is hemispherical, and the diameter is 1-1.5 cm; involucre 4 layers, oval outer layer, elliptical middle layer, and linear inner layer. The top of all bracts is sharp, the edge is dark purple, and the outside is short, soft and glandular hair. The small flower is blue-purple, the length of the small flower is 1.8 cm, the length of the tube part is 8 mm, and the length of the eave part is 10 mm. 2 layers of crown hair, light brown and short outer layer; coarse, 5 mm long, inner layer long, feather-like, 1.2 cm long.
Saussurea involucrate has strong medicinal action, but the chemical components of saussurea involucrate are complex, and the most main medicinal components are the difficulty of research. The known chemical components of the saussurea involucrate comprise flavonoids, alkaloids, sesquiterpenes, volatile oil and the like, wherein the flavonoids are the main chemical components of the saussurea involucrate, have obvious biological activities of eliminating superoxide anion free radicals, hydroxyl free radicals and resisting oxidation of NADH peroxidase, and have stronger effects of resisting oxidation, radiation and aging and the like. Therefore, the invention firstly extracts and analyzes the flavonoid substances in the saussurea involucrate. After long-time complex extraction, analysis and pharmacodynamic tests, four flavonoid components extracted from saussurea involucrate are innovatively found to have strong pharmacodynamic activity, and are respectively: rutin, myristicin, isoquercitrin, and luteolin-7-o-glucoside.
Meanwhile, when the extracted components of the saussurea involucrate flavonoid are analyzed, the weight compatibility of the four components is rutin: 28-32; myristicin: 0.8-1.2; isoquercitrin: 1.8-2.2; luteolin-7-o-glucoside: 6.8-7.2, the combination has the strongest pharmacodynamic activity, and more preferably, the compatibility is as follows: rutin: 30, of a nitrogen-containing gas; myristicin: 1; isoquercitrin: 2; luteolin-7-o-glucoside: 7.
after the types and the compatibility of the flavonoid components of the saussurea involucrate are obtained, the extraction method of the components can be carried out by adopting known extraction methods of various traditional Chinese medicine extracts, wherein one preferable extraction method is as follows:
the method comprises the following steps:
(1) alcohol extraction: pulverizing whole plant of saussurea involucrata, adding 10 times of 70% ethanol, heating and reflux-extracting for 3 times, each time for 1h, filtering, mixing extractive solutions, recovering solvent under reduced pressure, and concentrating to 1/50 of original solution volume as sample solution;
(2) fine extraction: sampling the sample solution, loading the pretreated resin column, and eluting, wherein the elution step comprises: washing the resin column with water for 3BV to remove impurities, and then eluting with 15% ethanol for 8BV to remove impurities, wherein the elution flow rate is 3 BV/h; finally eluting with 40% ethanol for 6BV at a flow rate of 3BV/h, and respectively collecting 40% of eluent of 1-2BV, 40% of eluent of 3-4BV and 40% of eluent of 5-6 BV;
(3) freeze-drying: mixing the collected eluates, recovering solvent under reduced pressure, and lyophilizing to obtain powder.
The resin column in the step (2) adopts AB-8 macroporous resin, and the pretreatment step comprises the following steps: soaking the column in absolute ethyl alcohol for 24h, filling the column in a wet method, eluting the column with the ethanol at the flow rate of about 2.5BV/h until the eluent is not turbid after being added with water with the volume of 1:5, and finally washing the column with deionized water at the same flow rate of about 5BV until the eluent has no alcohol smell for later use. The BV refers to the volume of resin loaded in the resin column, called bed volume or bed volume (bed volume), abbreviated BV.
The extraction method can obtain the saussurea involucrate flavonoid composition containing four components, wherein the four components are respectively as follows: rutin, myristicin, isoquercitrin, and luteolin-7-o-glucoside.
The invention proves the pharmacodynamic activity of the flavonoid composition through a large number of animal experiments, can be used for preparing various pain-relieving and pain-relieving medicines for clinical application and arthritis treatment medicines, and has the effects of high pharmacodynamic activity, small clinical dosage, quick response, low side effect and the like compared with a Chinese patent medicine preparation prepared by directly applying saussurea involucrate. The medicament dosage form can adopt conventional tablets or capsules, and can also be prepared into novel medicament dosage forms with higher absorption rate or better storage property according to the characteristics of the medicament components.
Drawings
FIG. 1 is a BPI diagram (good repeatability of two times of sample injection) of saussurea involucrata purified substance in positive ion mode;
FIG. 2 is a BPI diagram (good repeatability of two times of sample injection) of saussurea involucrata purified substance in negative ion mode;
FIG. 3 is a BPI plot in positive ion mode;
FIG. 4 shows the mice administered by oral gavage;
FIG. 5 is the tail vein administration of mice;
FIG. 6 is intraperitoneal administration of mice;
FIG. 7 is a mouse writhing;
FIG. 8 is a photograph of toe swelling;
fig. 9 is a photograph of toe swelling measurements.
Detailed Description
Embodiments of the present invention are described below with reference to the drawings. Elements and features depicted in one drawing or one embodiment of the invention may be combined with elements and features shown in one or more other drawings or embodiments. It should be noted that the figures and description omit representation and description of components or processes that are not relevant to the present invention and that are known to those of ordinary skill in the art for the sake of clarity.
Example 1 extraction of flavonoids from saussurea involucrata
1.1 alcohol extraction of saussurea involucrata medicinal material
Taking 6 saussurea involucrata medicinal materials, weighing the following materials before and after powdering: no. 1: 28.5g, 28.3 g; no. 2: 16.3g, 16.5 g; no. 3: 13.3g, 13.5 g; no. 4: 15.6g, 15.7 g; no. 5: 23.1g, 23.2 g; no. 6: 14.6g and 14.7 g. And (3) the total powder is as follows: extracting with 10 times of 70% ethanol under reflux for 3 times (1 hr each time), filtering with 8 layers of gauze, and mixing extractive solutions.
1.2 preparing the sample loading solution
Recovering solvent from the extractive solution under reduced pressure to obtain about 70mL of sample solution.
2 fine lifting
2.1 AB-8 macroporous resin (95% ethanol soaked for 24h) is selected, the inner diameter of the column is about 2.5cm, and the ratio of the diameter to the height is 1: 10. Soaking the macroporous resin in absolute ethyl alcohol for 24h, and then filling the column with a wet method, wherein a layer of cotton is required to be plugged at the bottom of the resin column before filling the column, and the column is soaked in absolute ethyl alcohol. Eluting with ethanol at about 2.5BV/h until the eluate plus five times water (1:5) is clear of turbidity. The eluent is washed by deionized water with the same flow rate of about 5BV until the eluent has no alcohol smell for standby.
2.2, the water-washed liquid level is leveled, and then the sample is loaded.
2.3 washing the resin column with water for 3BV to remove impurities, and then eluting with 15% ethanol for 8BV to remove impurities, wherein the elution flow rate is 3 BV/h. Finally eluting with 40% ethanol for 6BV at a flow rate of 3BV/h, and collecting 40% of eluent of 1-2BV, 40% of eluent of 3-4BV and 40% of eluent of 5-6 BV.
And 2.4, decompressing the eluent, recovering the solvent, and freeze-drying by using a freeze dryer to obtain a powdery product.
Example 2 analytical characterization of the powdered product obtained in example 1
1. Instruments and reagents
Waters acquisition UPLC Class H ultra high performance liquid chromatograph (Waters corporation, USA), Waters G2Q-TOF/MS high resolution mass spectrometer (Waters corporation, USA), Waters acquisition UPLC BEH C18 chromatographic column (2.1mm × 100mm, 1.7 μm) (Waters corporation, USA); data processing software 4.1 MassLynx; acetonitrile (mass spectrum grade, Fisher Scientific); drochen ultrapure water (guangzhou drochen food & beverage limited)); formic acid (quality spectrum grade, Fisher Scientific Co., Ltd.)
2 method
2.1 preparation of the solution
Preparation of a test solution: precisely weighing 0.02054g of the total extract, placing in a 10mL volumetric flask, adding 40% ethanol to a constant volume, mixing uniformly, centrifuging at a high speed of 12000r/min for 15min, and taking supernatant to obtain a test solution.
2.2 conditions of analysis
A chromatographic column: WatersAcquity UPLC BEH C18(2.1 mm. times.100 mm, 1.7 μm); the detection wavelength is 360 nm; the injection volume was 2. mu.L. Mobile phase: the A phase is 0.1% acetonitrile formate, the B phase is 0.1% formic acid water, gradient elution is adopted, and the specific elution conditions are as shown in the following table.
Chromatographic conditions are as follows: flow rate: 0.3 mL/min; column temperature: 30 deg.C
| Time (min) | A (0.1% formic acid acetonitrile)% | B (0.1% formic acid water)% |
| 0 | 5 | 95 |
| 4 | 9 | 91 |
| 10 | 13 | 87 |
| 20 | 16 | 84 |
| 30 | 17 | 83 |
| 35 | 28 | 72 |
| 40 | 95 | 5 |
| 43 | 5 | 95 |
Mass spectrometry conditions, using ESI ion source MSE positive ion mode: the ion source temperature is 120 ℃; the temperature of desolventizing gas is 400 ℃; the desolventizing air flow rate is 600L/h; the flow rate of the gas in the taper hole is 50L/h; the taper hole voltage is 40V; a capillary tube with a voltage of 3.0kV high-energy channel; the collision voltage is 20-75 eV. MSE scan mode: the mass scanning range m/z is 50-1200; the scan time was 0.2 s. To ensure accurate mass determination, mass correction during data collection using leucine-enkephalin as an external standard can produce fragment ions M/z 556.2771[ M + H ] +. Data acquisition and analysis were performed using MassLynx V4.1 software.
3 results and analysis
The BPI diagram of the saussurea involucrata purified product in positive ion mode is shown in figure 1, and the BPI diagram of the saussurea involucrata purified product in negative ion mode is shown in figure 2.
The chromatogram in positive ion mode was resolved, see FIG. 3.
The positive ion mode analysis results are shown in table 1:
example 3 Components and compatibility of saussurea involucrate flavonoid composition
The purified saussurea involucrate obtained by the extraction method mainly contains 4 flavonoid substances which are respectively shown in the following steps: rutin: myristicin: isoquercitrin: luteolin-7-o-glucoside, wherein the weight ratio of luteolin to glucoside is 30:1: 2: 7.
example 4 efficacy test (analgesic effect) of flavonoid composition of example 3
The pharmacodynamics of the flavonoid composition of example 3 was evaluated based on the therapeutic effect of the flavonoid composition of example 3 on an acetic acid-induced mouse pain model.
The method comprises the step of randomly dividing mice into 5 groups, namely a model control group (7 mice), a positive drug control group, a flavonoid composition high-dose group in example 3, a flavonoid composition medium-dose group in example 3 and a flavonoid composition low-dose group in example 3, wherein 9 mice are respectively divided into 9 mice, and a flavonoid composition tail vein administration group (7 mice) in example 3 is respectively adopted. Divided into groups and administered on the same day, 1 time per day for 3 days, and the control group is administered with physiological saline for intragastric administration under the same conditions. After 1 hour of the last administration, each mouse was intraperitoneally injected with 0.2ml of 1% acetic acid (glacial acetic acid), and the number of times of typical writhing of each mouse was observed within 20 minutes, and the results were counted by using a comparative t-test between groups.
The result shows that the flavonoid composition in the example 3 can obviously reduce the writhing frequency of mice when being applied to the model experiment of the mice pain caused by acetic acid, and has good dose-effect relationship dependence.
The conclusion is that the flavonoid composition in the example 3 has obvious analgesic effect.
1 test Material
1.1 test animals
BALB/c mice (SPF grade), Experimental animals technology, Inc. of Wei Tony Hua, Beijing. Production license of experimental animal: SCXK (Jing) 2016-: days 42-62, males 62, animal certification number: no. 1100111911022372.
1.2 drugs and reagents
1.2.1 flavonoid composition of example 3, light earthy yellow powder, dry stored away from the sun, dosed orally by gavage in the test at 60mg/kg, 30mg/kg, 15mg/kg, dissolved in physiological saline, dosed by volume: 0.1ml/10 g.
1.2.2 positive drugs, diclofenac sodium sustained-release tablets (trade name: hibilin), produced by Beijing Nouhua pharmaceutical industry Co., Ltd., national drug standard H10980297, dry storage in dark, light pink triangular film coated tablets, 75 mg/tablet, the dosage in the experiment is: 5 mg/kg. Dissolving with normal saline, and administering via oral gavage.
1.2.3 solvent: sodium chloride injection, specification: 250ml 2.25g, property: liquid, storage conditions: and (4) sealing and preserving and packaging: soft-packed bag, approved document No.: h13023201, production unit: shijiazhuang four drugs Co Ltd
1.2.4 modeling medicine: acetic acid (glacial acetic acid) (analytical purity content 99.5%) with the English name of acetic acid, molecular formula of C2H4O 2; CH3COOH, molecular weight 60.05 (international atomic weight in 1985), density 1.0492 boiling point 117.9 ℃, CAS number 64-19-7 Tianjin Cheng Yuan Chemicals, Inc
1.3 test apparatus
Electronic balance Sartorius, model: BSA 3202S-CW;
electronic balance, METTLER-TOLEDO, model: LE 204;
small animal body weight balance, shanghai yuping scientific instruments ltd, model: YP 1002; a liquid transfer device: eppendorf (eduncut);
IVC mouse rearing cage, Suzhou teaching cage products
High definition imaging System, Canon EOS 5D
2. Experimental methods
2.1 preparation of acetic acid-induced pain model in mice
12 Balb/c mice are purchased, acetic acid (glacial acetic acid) solution is prepared by using physiological saline, the concentration is respectively set to be 0.6%, 0.7%, 0.8% and 1%, 3 mice are respectively injected into each concentration, each mouse is subjected to intraperitoneal injection administration by 0.2ml, a pre-experiment is constructed as a model, the pre-experiment is immediately observed after administration, the frequency of typical body twisting of each mouse within 20 minutes after administration is recorded by using a video camera, and the administration dosage with proper body twisting frequency is found out. The results of preliminary experiments show that 0.7% of the administered dose is the most appropriate dose.
2.2 Experimental method for model drug effect of acetic acid induced pain in mice
50 Balb/c mice were selected, and randomly divided into 7 model negative control groups (given physiological saline) and 7 positive drug control groups (5mg/kg) according to body weight, 9 mice each group, three lavage dose groups (60mg/kg dose group, 30mg/kg dose group, 15mg/kg dose group) of the flavonoid composition of test drug example 3, 9 mice each group, and 7 groups of the flavonoid composition of test drug example 3 by tail vein injection at 30mg/kg dose. The administration frequency of each group is 1 time per day for 3 days, and the control group is administered with physiological saline under the same condition. After 1 hour from the last administration, 0.7% acetic acid (glacial acetic acid) was intraperitoneally injected into each mouse at a dose of 0.2 ml/mouse, and the number of times of the occurrence of typical writhing in each mouse was observed within 20 minutes, and the writhing inhibition rate was calculated.
2.3 photograph of Experimental Process
FIG. 4 shows the administration of the drug by oral gavage, FIG. 5 shows the administration of the drug by tail vein injection, FIG. 6 shows the administration of the drug by intraperitoneal injection, and FIG. 7 shows the writhing of the mouse (manifestations: reaction of belly indent, trunk hind limb stretch, hip lift, etc.).
3. Results of the experiment
3.1 statistical methods
The data is processed by adopting an SPSS13.0 statistical software package, and the data of each group are subjected to addition and subtraction of standard deviation by mean
Represents; the results were compared using an interclass comparison t test, P<0.05 is statistically significant, P<0.01 is that the difference has significant statistical significance
3.2 statistical results
TABLE 2 Effect on acetic acid-induced pain model in mice
Comparison with model control group: p < 0.05; p <0.01
Table 2 the results show: after acetic acid injection, the number of writhing of mice in each administration group is obviously less than that of a model control group, and the writhing number of the mice in each administration group is obviously different from that of the model control group (P <0.05 and P < 0.01).
4 conclusion
In the acetic acid induced mouse pain model drug effect experiment, the flavonoid composition high, medium and low dose groups and the tail vein middle dose group in the example 3 have obvious pain formation effects, and each administration group can greatly reduce the body writhing times of mice and show good dose-effect correlation.
Example 5 efficacy test (arthritis therapeutic effect) of flavonoid composition of example 3
The efficacy of the flavonoid composition of example 3 was evaluated on the basis of the therapeutic effect of the flavonoid composition of example 3 on a freund's complete adjuvant (CFA) arthritis-causing mouse model.
Method, mice are injected with 30 mu L of Freund's complete adjuvant into the skin of the left hind toe at one time to establish an adjuvant arthritis mouse model. The success of the model building is judged by the obvious swelling of the model building side, the contralateral side and the foot sole of the front paw, the affected joint deformity, the inconvenient movement and the obvious reduction of the physical quality of the mouse. After 24h of molding, mice which failed in molding were removed, and all mice which succeeded in molding were randomly divided into 5 groups, which were respectively a model control group (7 mice), a positive drug control group, a flavonoid composition high dose group of example 3, a flavonoid composition medium dose group of example 3, and a flavonoid composition low dose group of example 3, 9 mice per group, and a flavonoid composition tail vein administration group of example 3 (7 mice). Divided into groups and administered on the same day, 1 time per day for 10 days, and the control group is administered with physiological saline for intragastric administration under the same conditions. The animal status was recorded, toe swelling thickness was measured with a vernier caliper, data was recorded, and results were counted using a comparative t-test between groups.
The result shows that the flavonoid composition in the example 3 can obviously reduce the joint swelling degree of mice when being applied to adjuvant arthritis mouse model experiments, and has good dose-effect relationship dependence.
The conclusion is that the flavonoid composition in the example 3 has obvious effect of reducing the joint swelling.
1 test Material
1.1 test animals
BALB/c mice (SPF grade), Schbefu (Beijing) Biotechnology, Inc. Production license of experimental animal: SCXK (Jing) 2016-: days 42-56, 50 males, animal certification number: no. 1103241911021019.
1.2 drugs and reagents
1.2.1 flavonoid composition of example 3, yellow-white powder, dry stored in the dark, dosed orally by gavage at 60mg/kg, 30mg/kg, 15mg/kg in the test, dissolved in physiological saline, dosed by the volume: 0.1ml/10 g.
1.2.2 positive drugs, diclofenac sodium sustained-release tablets (trade name: hibilin), produced by Beijing Nouhua pharmaceutical industry Co., Ltd., national drug standard H10980297, dry storage in dark, light pink triangular film coated tablets, 75 mg/tablet, the dosage in the experiment is: 5 mg/kg. Dissolving with normal saline, and administering via oral gavage.
1.2.3 solvent: sodium chloride injection, specification: 250ml 2.25g, property: liquid, storage conditions: and (4) sealing and preserving and packaging: soft-packed bag, approved document No.: h13023201, production unit: shijiazhuang four drugs Co Ltd
1.3 test apparatus
Electronic balance Sartorius, model: BSA 3202S-CW;
electronic balance, METTLER-TOLEDO, model: LE 204;
small animal body weight balance, shanghai yuping scientific instruments ltd, model: YP 1002; a liquid transfer device: eppendorf (eduncut);
IVC mouse rearing cage, Suzhou teaching cage products
High definition imaging System, Canon EOS 5D
2. Experimental methods
2.1 preparation of adjuvant arthritis mouse model
Mice were injected subcutaneously with 30 μ L of freund's complete adjuvant in one injection in the left hind toe to establish an adjuvant arthritis mouse model. The success of the model making is judged by the obvious swelling of the side, the contralateral side and the foot sole of the front paw of the model making of the mouse, the affected joint deformity, the inconvenient movement and the obvious reduction of the physical quality
2.2 adjuvant arthritis mouse model drug effect experimental method
After random grouping, the test drug was divided into 7 (saline administration), a positive drug control group (5mg/kg), 9 per group, three gavage dose groups (60mg/kg dose group, 30mg/kg dose group, 15mg/kg dose group) of test drug, 9 per group, and 7 tail vein injection groups of test drug at 30mg/kg dose. The administration frequency of each group is 1 time per day for 10 days, and the control group is administered with physiological saline under the same condition. The state of the animals was observed periodically, and the swelling thickness of the toes of the mice was measured with a vernier caliper. The photograph of toe swelling is shown in fig. 8, and the photograph of toe swelling measurement is shown in fig. 9.
3. Results of the experiment
3.1 statistical methods
The data is processed by adopting an SPSS13.0 statistical software package, and the data of each group are subjected to addition and subtraction of standard deviation by mean
Represents; the results were compared using an interclass comparison t test, P<A difference of 0.05 is statistically significant.
3.2 statistical results
TABLE 3 Effect on adjuvant arthritis mouse model
Comparison with model control group: p <0.05
Table 3 the results show: the dose groups all inhibit the toe swelling of arthritis mice to different degrees, wherein the dose group of drug X and the high dose group of drug X have significant difference compared with the model control group (P < 0.05).
4 conclusion
In an adjuvant arthritis mouse model efficacy experiment, the flavonoid composition high, medium and low dose groups and the medium dose injection group in the embodiment 3 have the effect of inhibiting toe swelling of mice and show good dose-effect correlation, wherein the compound high dose group and the compound medium dose group have significance.
Example 6
To further prove the effect of the snow lotus flavonoid composition, in example 6, based on the compatibility of the components obtained in example 3, four commercially available compounds are adopted to compound the composition in the application, and then a pharmacodynamic test is performed, wherein the test process is consistent with the test processes of examples 4 and 5, and is not repeated here, and the compounding ratio and the test results are shown in the following table:
TABLE 4 composition Components and compounding ratios (by weight)
| Components | Rutin | Myristicin | Isoquercitrin | Luteolin-7-o-glucoside |
| Compounding ratio | 30 | 1 | 2 | 7 |
TABLE 5 Effect on acetic acid-induced pain model in mice
Comparison with model control group: p < 0.05; p <0.01
TABLE 6 Effect on adjuvant arthritis mouse model
| Group of | Dosage (mg/kg) | Animal number (only) | Swelling and proliferation rate% |
| Model control group | - | 7 | 14.45±11.80 |
| Drug X high dose | 60 | 9 | -1.40±11.37* |
| Drug X Medium dose | 30 | 9 | -2.45±8.99** |
| Drug X Medium dose injection | 30 | 7 | -2.58±10.96* |
| Drug X Low dose | 15 | 9 | -1.67±11.83* |
| Positive drug | 5 | 9 | -0.05±7.99* |
Comparison with model control group: p <0.05
As can be seen from tables 5 and 6, the flavonoid composition directly compounded by the four compounds still has good analgesic effect and arthritis treatment effect.
Although the present invention and its advantages have been described in detail, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims. Moreover, the scope of the present application is not intended to be limited to the particular embodiments of the process, machine, means, methods and steps described in the specification. As one of ordinary skill in the art will readily appreciate from the disclosure of the present invention, processes, machines, means, methods, or steps, presently existing or later to be developed that perform substantially the same function or achieve substantially the same result as the corresponding embodiments described herein may be utilized according to the present invention. Accordingly, the appended claims are intended to include within their scope such processes, devices, means, methods, or steps.
Claims (5)
1. The extraction method of the saussurea involucrate flavonoid composition is characterized by comprising the following steps:
(1) alcohol extraction: pulverizing whole plant of saussurea involucrata, adding 10 times of 70% ethanol, heating and reflux-extracting for 3 times, each time for 1h, filtering, mixing extractive solutions, recovering solvent under reduced pressure, and concentrating to 1/50 of original solution volume as sample solution;
(2) fine extraction: sampling the sample solution, loading the pretreated resin column, and eluting, wherein the elution step comprises: washing the resin column with water for 3BV to remove impurities, and then eluting with 15% ethanol for 8BV to remove impurities, wherein the elution flow rate is 3 BV/h; finally eluting with 40% ethanol for 6BV at a flow rate of 3BV/h, and respectively collecting 40% of eluent of 1-2BV, 40% of eluent of 3-4BV and 40% of eluent of 5-6 BV;
(3) freeze-drying: mixing the collected eluates, recovering solvent under reduced pressure, and lyophilizing to obtain powder.
2. The extraction method according to claim 1, wherein the resin column in the step (2) is AB-8 macroporous resin, and the pretreatment step comprises: soaking the column in absolute ethyl alcohol for 24h, filling the column in a wet method, eluting the column with the ethanol at the flow rate of about 2.5BV/h until the eluent is not turbid after being added with water with the volume of 1:5, and finally washing the column with deionized water at the same flow rate of about 5BV until the eluent has no alcohol smell for later use.
3. The extraction method as claimed in claim 1 or 2, wherein the saussurea involucrate flavonoid composition comprises four components, which are respectively: rutin, myristicin, isoquercitrin, and luteolin-7-o-glucoside.
4. The extraction method according to claim 3, wherein the four components are present in the composition in the following weight ratios:
rutin: 28-32; myristicin: 0.8-1.2; isoquercitrin: 1.8-2.2; luteolin-7-o-glucoside: 6.8-7.2.
5. The extraction method according to claim 4, wherein the four components are present in the composition in the following weight ratios:
rutin: 30, of a nitrogen-containing gas; myristicin: 1; isoquercitrin: 2; luteolin-7-o-glucoside: 7.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201911205352.XA CN112876522A (en) | 2019-11-29 | 2019-11-29 | Method for extracting saussurea involucrate flavonoid composition |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201911205352.XA CN112876522A (en) | 2019-11-29 | 2019-11-29 | Method for extracting saussurea involucrate flavonoid composition |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN112876522A true CN112876522A (en) | 2021-06-01 |
Family
ID=76039070
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201911205352.XA Pending CN112876522A (en) | 2019-11-29 | 2019-11-29 | Method for extracting saussurea involucrate flavonoid composition |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN112876522A (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101209274A (en) * | 2007-12-25 | 2008-07-02 | 新疆维吾尔自治区药物研究所 | Saussurea involucrata extract and saussurea involucrata catablasm and producing method thereof |
| CN102106889A (en) * | 2011-02-21 | 2011-06-29 | 贾正平 | Application of snow saussurea and snow saussurea extract in preparation of anti-hypoxia medicaments and method for preparing snow saussurea extract |
| CN103330728A (en) * | 2013-07-17 | 2013-10-02 | 贾正平 | Preparation method and application of saussurea involucrate flavonoid capsule |
-
2019
- 2019-11-29 CN CN201911205352.XA patent/CN112876522A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101209274A (en) * | 2007-12-25 | 2008-07-02 | 新疆维吾尔自治区药物研究所 | Saussurea involucrata extract and saussurea involucrata catablasm and producing method thereof |
| CN102106889A (en) * | 2011-02-21 | 2011-06-29 | 贾正平 | Application of snow saussurea and snow saussurea extract in preparation of anti-hypoxia medicaments and method for preparing snow saussurea extract |
| CN103330728A (en) * | 2013-07-17 | 2013-10-02 | 贾正平 | Preparation method and application of saussurea involucrate flavonoid capsule |
Non-Patent Citations (1)
| Title |
|---|
| 景临林 等: "天山雪莲抗缺氧活性成分研究", 中药材, vol. 38, no. 1, pages 199 - 201 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| EP1463412B1 (en) | Composition comprising wenguanguo extracts, methods for preparing same and uses therof | |
| CN101002841B (en) | Effective components of rose, its preparing method and use | |
| CN106963853B (en) | Total flavonoid aglycone extract of Sichuan blackberry lily and preparation method and application thereof | |
| CN102228506A (en) | Composition of malaytea scurfpea extract as well as preparation method and use thereof | |
| CN100374120C (en) | Powder of flenabane and its preparation method as well as application in making drugs | |
| JP2009514960A (en) | Herbal extract powder and its preparation and use | |
| CN101057925B (en) | Preparation technology for 'jieguqili' capsule | |
| KR20160117425A (en) | Solid dispersion containing desmodium styracifolium (osb.) merr. flavonoids, method of preparing same, and use thereof | |
| WO2008145064A1 (en) | The method for a sequoyitol-containing extract obtaining from the genus of trifolium, sobyean and ginkgo biloba and use thereof | |
| CN105079308A (en) | Common rockvine herb extract as well as preparation method and pharmaceutical application of extract | |
| CN104815025A (en) | Preparation process and quality control method of cassia twig poria cocos preparation | |
| CN113663029A (en) | Compound wild buckwheat rhizome composition | |
| CN106074588A (en) | The combination of Rhizoma Paridis forrestii monomer saponin and pharmaceutical composition and its application in pharmacy | |
| CN1775254A (en) | Medicinal preparation for treating deaf and preparing method | |
| CN112870209B (en) | Saussurea involucrata flavonoid composition | |
| CN100444849C (en) | New use of tribulus terrestris extraction | |
| KR20130074121A (en) | Ginseng prosapogenin high concentration containing sanchi ginseng preparation using sonication and process for thereof | |
| CN112876522A (en) | Method for extracting saussurea involucrate flavonoid composition | |
| CN112870243A (en) | Application of snow lotus flavonoid composition in preparation of pain relieving and pain easing medicines | |
| CN112870242A (en) | Application of saussurea involucrate flavonoid composition in preparation of medicine for treating arthritis diseases | |
| CN101904894B (en) | Application of lamiophlomis rotate total glycosides in preparing medicines | |
| CN102641342B (en) | A kind of Chinese medicine extract and preparation method for the treatment of nephropathy | |
| CN110448651B (en) | Preparation method of Tibetan traditional Chinese medicine composition for treating liver diseases, composition and granules containing composition | |
| CN102861278B (en) | Lipid-lowering extract from effective parts of sharpleaf galangal fruit and preparation and application thereof | |
| CN101974012B (en) | Novel compound ethyl brevicate with pharmaceutical activity |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20210601 |
|
| WD01 | Invention patent application deemed withdrawn after publication |