CN111533680A - Folium Mori extract and folium Mori multicomponent mixture with blood sugar lowering effect prepared from the same - Google Patents
Folium Mori extract and folium Mori multicomponent mixture with blood sugar lowering effect prepared from the same Download PDFInfo
- Publication number
- CN111533680A CN111533680A CN202010470759.1A CN202010470759A CN111533680A CN 111533680 A CN111533680 A CN 111533680A CN 202010470759 A CN202010470759 A CN 202010470759A CN 111533680 A CN111533680 A CN 111533680A
- Authority
- CN
- China
- Prior art keywords
- mulberry leaf
- extract
- ethanol solution
- extraction
- mulberry
- 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
- 239000000284 extract Substances 0.000 title claims abstract description 105
- 239000000203 mixture Substances 0.000 title claims abstract description 34
- 210000004369 blood Anatomy 0.000 title abstract description 16
- 239000008280 blood Substances 0.000 title abstract description 16
- 230000000694 effects Effects 0.000 title description 12
- 240000000249 Morus alba Species 0.000 claims abstract description 182
- 235000008708 Morus alba Nutrition 0.000 claims abstract description 182
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 170
- 238000000605 extraction Methods 0.000 claims abstract description 99
- 238000000034 method Methods 0.000 claims abstract description 64
- 150000004676 glycans Chemical class 0.000 claims abstract description 51
- 229920001282 polysaccharide Polymers 0.000 claims abstract description 51
- 239000005017 polysaccharide Substances 0.000 claims abstract description 51
- 229930003944 flavone Natural products 0.000 claims abstract description 46
- 235000011949 flavones Nutrition 0.000 claims abstract description 46
- 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 claims abstract description 45
- 150000002212 flavone derivatives Chemical class 0.000 claims abstract description 45
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 claims abstract description 45
- 230000002378 acidificating effect Effects 0.000 claims abstract description 29
- 238000002360 preparation method Methods 0.000 claims abstract description 15
- 239000002994 raw material Substances 0.000 claims abstract description 12
- 238000000746 purification Methods 0.000 claims description 29
- 239000003480 eluent Substances 0.000 claims description 23
- 238000002481 ethanol extraction Methods 0.000 claims description 22
- 239000011347 resin Substances 0.000 claims description 22
- 229920005989 resin Polymers 0.000 claims description 22
- 239000000843 powder Substances 0.000 claims description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 17
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 claims description 12
- 238000011068 loading method Methods 0.000 claims description 12
- 239000012153 distilled water Substances 0.000 claims description 10
- 239000000047 product Substances 0.000 claims description 8
- 239000003729 cation exchange resin Substances 0.000 claims description 6
- 239000003456 ion exchange resin Substances 0.000 claims description 6
- 229920003303 ion-exchange polymer Polymers 0.000 claims description 6
- 238000010828 elution Methods 0.000 claims description 5
- 238000002156 mixing Methods 0.000 claims description 4
- 238000001556 precipitation Methods 0.000 claims description 4
- VHUUQVKOLVNVRT-UHFFFAOYSA-N Ammonium hydroxide Chemical compound [NH4+].[OH-] VHUUQVKOLVNVRT-UHFFFAOYSA-N 0.000 claims description 3
- 235000011114 ammonium hydroxide Nutrition 0.000 claims description 3
- 230000001376 precipitating effect Effects 0.000 claims description 3
- 239000012264 purified product Substances 0.000 claims description 2
- 239000003814 drug Substances 0.000 abstract description 7
- 230000002218 hypoglycaemic effect Effects 0.000 abstract description 6
- 230000001603 reducing effect Effects 0.000 abstract description 6
- 229940079593 drug Drugs 0.000 abstract description 5
- LXBIFEVIBLOUGU-JGWLITMVSA-N duvoglustat Chemical compound OC[C@H]1NC[C@H](O)[C@@H](O)[C@@H]1O LXBIFEVIBLOUGU-JGWLITMVSA-N 0.000 description 187
- LXBIFEVIBLOUGU-UHFFFAOYSA-N Deoxymannojirimycin Natural products OCC1NCC(O)C(O)C1O LXBIFEVIBLOUGU-UHFFFAOYSA-N 0.000 description 93
- 239000000243 solution Substances 0.000 description 53
- 241000700159 Rattus Species 0.000 description 28
- 239000000523 sample Substances 0.000 description 25
- 206010012601 diabetes mellitus Diseases 0.000 description 14
- 238000002474 experimental method Methods 0.000 description 13
- 230000008569 process Effects 0.000 description 13
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 10
- 239000002253 acid Substances 0.000 description 10
- 238000012360 testing method Methods 0.000 description 10
- 238000005303 weighing Methods 0.000 description 9
- 210000002966 serum Anatomy 0.000 description 8
- 239000003153 chemical reaction reagent Substances 0.000 description 7
- 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 description 6
- 206010022489 Insulin Resistance Diseases 0.000 description 6
- 238000002835 absorbance Methods 0.000 description 6
- 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 description 6
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 6
- 239000011344 liquid material Substances 0.000 description 6
- 239000000463 material Substances 0.000 description 6
- 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 description 6
- 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 description 6
- 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 description 6
- 235000005493 rutin Nutrition 0.000 description 6
- 229960004555 rutoside Drugs 0.000 description 6
- 238000012216 screening Methods 0.000 description 6
- 239000012086 standard solution Substances 0.000 description 6
- 102000004190 Enzymes Human genes 0.000 description 5
- 108090000790 Enzymes Proteins 0.000 description 5
- 238000010521 absorption reaction Methods 0.000 description 5
- 239000003963 antioxidant agent Substances 0.000 description 5
- 230000003078 antioxidant effect Effects 0.000 description 5
- 238000010790 dilution Methods 0.000 description 5
- 239000012895 dilution Substances 0.000 description 5
- 229940088598 enzyme Drugs 0.000 description 5
- 229930003935 flavonoid Natural products 0.000 description 5
- 235000017173 flavonoids Nutrition 0.000 description 5
- 239000012488 sample solution Substances 0.000 description 5
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 4
- DDRJAANPRJIHGJ-UHFFFAOYSA-N creatinine Chemical compound CN1CC(=O)NC1=N DDRJAANPRJIHGJ-UHFFFAOYSA-N 0.000 description 4
- 239000012043 crude product Substances 0.000 description 4
- 230000003544 deproteinization Effects 0.000 description 4
- 150000002215 flavonoids Chemical class 0.000 description 4
- 210000004153 islets of langerhan Anatomy 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- 210000004185 liver Anatomy 0.000 description 4
- 230000014759 maintenance of location Effects 0.000 description 4
- 239000013641 positive control Substances 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 108010074051 C-Reactive Protein Proteins 0.000 description 3
- 102100032752 C-reactive protein Human genes 0.000 description 3
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 3
- 102000004877 Insulin Human genes 0.000 description 3
- 108090001061 Insulin Proteins 0.000 description 3
- 208000017170 Lipid metabolism disease Diseases 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 230000006378 damage Effects 0.000 description 3
- 238000004042 decolorization Methods 0.000 description 3
- 239000008103 glucose Substances 0.000 description 3
- 229940125396 insulin Drugs 0.000 description 3
- 238000002386 leaching Methods 0.000 description 3
- 230000003859 lipid peroxidation Effects 0.000 description 3
- XZWYZXLIPXDOLR-UHFFFAOYSA-N metformin Chemical compound CN(C)C(=N)NC(N)=N XZWYZXLIPXDOLR-UHFFFAOYSA-N 0.000 description 3
- 230000036542 oxidative stress Effects 0.000 description 3
- 230000009467 reduction Effects 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 208000001072 type 2 diabetes mellitus Diseases 0.000 description 3
- 102000011690 Adiponectin Human genes 0.000 description 2
- 108010076365 Adiponectin Proteins 0.000 description 2
- 102000016938 Catalase Human genes 0.000 description 2
- 108010053835 Catalase Proteins 0.000 description 2
- 102000006587 Glutathione peroxidase Human genes 0.000 description 2
- 108700016172 Glutathione peroxidases Proteins 0.000 description 2
- 108010023302 HDL Cholesterol Proteins 0.000 description 2
- DKNPRRRKHAEUMW-UHFFFAOYSA-N Iodine aqueous Chemical compound [K+].I[I-]I DKNPRRRKHAEUMW-UHFFFAOYSA-N 0.000 description 2
- 108010028554 LDL Cholesterol Proteins 0.000 description 2
- 102000016267 Leptin Human genes 0.000 description 2
- 108010092277 Leptin Proteins 0.000 description 2
- WSMYVTOQOOLQHP-UHFFFAOYSA-N Malondialdehyde Chemical compound O=CCC=O WSMYVTOQOOLQHP-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- REFJWTPEDVJJIY-UHFFFAOYSA-N Quercetin Chemical compound C=1C(O)=CC(O)=C(C(C=2O)=O)C=1OC=2C1=CC=C(O)C(O)=C1 REFJWTPEDVJJIY-UHFFFAOYSA-N 0.000 description 2
- ZSJLQEPLLKMAKR-UHFFFAOYSA-N Streptozotocin Natural products O=NN(C)C(=O)NC1C(O)OC(CO)C(O)C1O ZSJLQEPLLKMAKR-UHFFFAOYSA-N 0.000 description 2
- 102000019197 Superoxide Dismutase Human genes 0.000 description 2
- 108010012715 Superoxide dismutase Proteins 0.000 description 2
- YVNQAIFQFWTPLQ-UHFFFAOYSA-O [4-[[4-(4-ethoxyanilino)phenyl]-[4-[ethyl-[(3-sulfophenyl)methyl]amino]-2-methylphenyl]methylidene]-3-methylcyclohexa-2,5-dien-1-ylidene]-ethyl-[(3-sulfophenyl)methyl]azanium Chemical compound C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C(=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S(O)(=O)=O)C)C=2C(=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S(O)(=O)=O)C)C=C1 YVNQAIFQFWTPLQ-UHFFFAOYSA-O 0.000 description 2
- PNNCWTXUWKENPE-UHFFFAOYSA-N [N].NC(N)=O Chemical compound [N].NC(N)=O PNNCWTXUWKENPE-UHFFFAOYSA-N 0.000 description 2
- 239000004480 active ingredient Substances 0.000 description 2
- 210000001789 adipocyte Anatomy 0.000 description 2
- 229930013930 alkaloid Natural products 0.000 description 2
- 239000003472 antidiabetic agent Substances 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 210000004027 cell Anatomy 0.000 description 2
- 235000012000 cholesterol Nutrition 0.000 description 2
- 230000000295 complement effect Effects 0.000 description 2
- NKLPQNGYXWVELD-UHFFFAOYSA-M coomassie brilliant blue Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=C1 NKLPQNGYXWVELD-UHFFFAOYSA-M 0.000 description 2
- 229940109239 creatinine Drugs 0.000 description 2
- 238000007865 diluting Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000012869 ethanol precipitation Methods 0.000 description 2
- -1 flavonoid compound Chemical class 0.000 description 2
- 235000021588 free fatty acids Nutrition 0.000 description 2
- 238000004108 freeze drying Methods 0.000 description 2
- 230000002757 inflammatory effect Effects 0.000 description 2
- 239000007928 intraperitoneal injection Substances 0.000 description 2
- 150000002500 ions Chemical class 0.000 description 2
- 230000003907 kidney function Effects 0.000 description 2
- NRYBAZVQPHGZNS-ZSOCWYAHSA-N leptin Chemical compound O=C([C@H](CO)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CO)NC(=O)CNC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](N)CC(C)C)CCSC)N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](CS)C(O)=O NRYBAZVQPHGZNS-ZSOCWYAHSA-N 0.000 description 2
- 229940039781 leptin Drugs 0.000 description 2
- 229940118019 malondialdehyde Drugs 0.000 description 2
- 229960003105 metformin Drugs 0.000 description 2
- 238000005457 optimization Methods 0.000 description 2
- 210000000496 pancreas Anatomy 0.000 description 2
- 210000005084 renal tissue Anatomy 0.000 description 2
- LPXPTNMVRIOKMN-UHFFFAOYSA-M sodium nitrite Chemical compound [Na+].[O-]N=O LPXPTNMVRIOKMN-UHFFFAOYSA-M 0.000 description 2
- ZSJLQEPLLKMAKR-GKHCUFPYSA-N streptozocin Chemical compound O=NN(C)C(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O ZSJLQEPLLKMAKR-GKHCUFPYSA-N 0.000 description 2
- 229960001052 streptozocin Drugs 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 230000002195 synergetic effect Effects 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 210000005239 tubule Anatomy 0.000 description 2
- 238000002137 ultrasound extraction Methods 0.000 description 2
- BNGXYYYYKUGPPF-UHFFFAOYSA-M (3-methylphenyl)methyl-triphenylphosphanium;chloride Chemical compound [Cl-].CC1=CC=CC(C[P+](C=2C=CC=CC=2)(C=2C=CC=CC=2)C=2C=CC=CC=2)=C1 BNGXYYYYKUGPPF-UHFFFAOYSA-M 0.000 description 1
- MZOFCQQQCNRIBI-VMXHOPILSA-N (3s)-4-[[(2s)-1-[[(2s)-1-[[(1s)-1-carboxy-2-hydroxyethyl]amino]-4-methyl-1-oxopentan-2-yl]amino]-5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-3-[[2-[[(2s)-2,6-diaminohexanoyl]amino]acetyl]amino]-4-oxobutanoic acid Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CC(O)=O)NC(=O)CNC(=O)[C@@H](N)CCCCN MZOFCQQQCNRIBI-VMXHOPILSA-N 0.000 description 1
- HIXDQWDOVZUNNA-UHFFFAOYSA-N 2-(3,4-dimethoxyphenyl)-5-hydroxy-7-methoxychromen-4-one Chemical compound C=1C(OC)=CC(O)=C(C(C=2)=O)C=1OC=2C1=CC=C(OC)C(OC)=C1 HIXDQWDOVZUNNA-UHFFFAOYSA-N 0.000 description 1
- CWVRJTMFETXNAD-FWCWNIRPSA-N 3-O-Caffeoylquinic acid Natural products O[C@H]1[C@@H](O)C[C@@](O)(C(O)=O)C[C@H]1OC(=O)\C=C\C1=CC=C(O)C(O)=C1 CWVRJTMFETXNAD-FWCWNIRPSA-N 0.000 description 1
- GOZMBJCYMQQACI-UHFFFAOYSA-N 6,7-dimethyl-3-[[methyl-[2-[methyl-[[1-[3-(trifluoromethyl)phenyl]indol-3-yl]methyl]amino]ethyl]amino]methyl]chromen-4-one;dihydrochloride Chemical compound Cl.Cl.C=1OC2=CC(C)=C(C)C=C2C(=O)C=1CN(C)CCN(C)CC(C1=CC=CC=C11)=CN1C1=CC=CC(C(F)(F)F)=C1 GOZMBJCYMQQACI-UHFFFAOYSA-N 0.000 description 1
- 102000014777 Adipokines Human genes 0.000 description 1
- 108010078606 Adipokines Proteins 0.000 description 1
- MQVRGDZCYDEQML-UHFFFAOYSA-N Astragalin Natural products C1=CC(OC)=CC=C1C1=C(OC2C(C(O)C(O)C(CO)O2)O)C(=O)C2=C(O)C=C(O)C=C2O1 MQVRGDZCYDEQML-UHFFFAOYSA-N 0.000 description 1
- 102100026189 Beta-galactosidase Human genes 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- PZIRUHCJZBGLDY-UHFFFAOYSA-N Caffeoylquinic acid Natural products CC(CCC(=O)C(C)C1C(=O)CC2C3CC(O)C4CC(O)CCC4(C)C3CCC12C)C(=O)O PZIRUHCJZBGLDY-UHFFFAOYSA-N 0.000 description 1
- 206010019708 Hepatic steatosis Diseases 0.000 description 1
- 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 description 1
- 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 description 1
- 108010059881 Lactase Proteins 0.000 description 1
- 241000218231 Moraceae Species 0.000 description 1
- 241000218213 Morus <angiosperm> Species 0.000 description 1
- HDPISCIDKUMRDI-UHFFFAOYSA-M N(=O)[O-].[Na+].[N+](=O)([O-])[O-].[Al+3] Chemical compound N(=O)[O-].[Na+].[N+](=O)([O-])[O-].[Al+3] HDPISCIDKUMRDI-UHFFFAOYSA-M 0.000 description 1
- CWVRJTMFETXNAD-KLZCAUPSSA-N Neochlorogenin-saeure Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O CWVRJTMFETXNAD-KLZCAUPSSA-N 0.000 description 1
- 101710184309 Probable sucrose-6-phosphate hydrolase Proteins 0.000 description 1
- ZVOLCUVKHLEPEV-UHFFFAOYSA-N Quercetagetin Natural products C1=C(O)C(O)=CC=C1C1=C(O)C(=O)C2=C(O)C(O)=C(O)C=C2O1 ZVOLCUVKHLEPEV-UHFFFAOYSA-N 0.000 description 1
- HWTZYBCRDDUBJY-UHFFFAOYSA-N Rhynchosin Natural products C1=C(O)C(O)=CC=C1C1=C(O)C(=O)C2=CC(O)=C(O)C=C2O1 HWTZYBCRDDUBJY-UHFFFAOYSA-N 0.000 description 1
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 1
- 102400000472 Sucrase Human genes 0.000 description 1
- 101710112652 Sucrose-6-phosphate hydrolase Proteins 0.000 description 1
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 1
- 102000000852 Tumor Necrosis Factor-alpha Human genes 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 239000000478 adipokine Substances 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- 102000016679 alpha-Glucosidases Human genes 0.000 description 1
- 108010028144 alpha-Glucosidases Proteins 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 210000002469 basement membrane Anatomy 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 108010005774 beta-Galactosidase Proteins 0.000 description 1
- 230000000975 bioactive effect Effects 0.000 description 1
- 229940098773 bovine serum albumin Drugs 0.000 description 1
- 238000012512 characterization method Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 229940074393 chlorogenic acid Drugs 0.000 description 1
- CWVRJTMFETXNAD-JUHZACGLSA-N chlorogenic acid Chemical compound O[C@@H]1[C@H](O)C[C@@](O)(C(O)=O)C[C@H]1OC(=O)\C=C\C1=CC=C(O)C(O)=C1 CWVRJTMFETXNAD-JUHZACGLSA-N 0.000 description 1
- FFQSDFBBSXGVKF-KHSQJDLVSA-N chlorogenic acid Natural products O[C@@H]1C[C@](O)(C[C@@H](CC(=O)C=Cc2ccc(O)c(O)c2)[C@@H]1O)C(=O)O FFQSDFBBSXGVKF-KHSQJDLVSA-N 0.000 description 1
- 235000001368 chlorogenic acid Nutrition 0.000 description 1
- BMRSEYFENKXDIS-KLZCAUPSSA-N cis-3-O-p-coumaroylquinic acid Natural products O[C@H]1C[C@@](O)(C[C@@H](OC(=O)C=Cc2ccc(O)cc2)[C@@H]1O)C(=O)O BMRSEYFENKXDIS-KLZCAUPSSA-N 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 150000002016 disaccharides Chemical class 0.000 description 1
- 229940000406 drug candidate Drugs 0.000 description 1
- 239000012636 effector Substances 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 239000003797 essential amino acid Substances 0.000 description 1
- 235000020776 essential amino acid Nutrition 0.000 description 1
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 1
- 239000003777 experimental drug Substances 0.000 description 1
- 150000002213 flavones Chemical class 0.000 description 1
- 238000003304 gavage Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 235000009200 high fat diet Nutrition 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 201000001421 hyperglycemia Diseases 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 235000011073 invertase Nutrition 0.000 description 1
- 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 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
- JPUKWEQWGBDDQB-QSOFNFLRSA-N kaempferol 3-O-beta-D-glucoside Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C(C=2C=CC(O)=CC=2)OC2=CC(O)=CC(O)=C2C1=O JPUKWEQWGBDDQB-QSOFNFLRSA-N 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 229940116108 lactase Drugs 0.000 description 1
- 238000011031 large-scale manufacturing process Methods 0.000 description 1
- 230000031700 light absorption Effects 0.000 description 1
- 210000005228 liver tissue Anatomy 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 235000010755 mineral Nutrition 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 150000002772 monosaccharides Chemical class 0.000 description 1
- 230000004792 oxidative damage Effects 0.000 description 1
- 210000004923 pancreatic tissue Anatomy 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
- OQUKIQWCVTZJAF-UHFFFAOYSA-N phenol;sulfuric acid Chemical compound OS(O)(=O)=O.OC1=CC=CC=C1 OQUKIQWCVTZJAF-UHFFFAOYSA-N 0.000 description 1
- 229930000732 piperidine alkaloid Natural products 0.000 description 1
- 150000003053 piperidines Chemical class 0.000 description 1
- 150000008442 polyphenolic compounds Chemical class 0.000 description 1
- 235000013824 polyphenols Nutrition 0.000 description 1
- 230000000291 postprandial effect Effects 0.000 description 1
- NLKNQRATVPKPDG-UHFFFAOYSA-M potassium iodide Substances [K+].[I-] NLKNQRATVPKPDG-UHFFFAOYSA-M 0.000 description 1
- 229960001285 quercetin Drugs 0.000 description 1
- 235000005875 quercetin Nutrition 0.000 description 1
- 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 description 1
- 238000011552 rat model Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000007873 sieving Methods 0.000 description 1
- 210000000813 small intestine Anatomy 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
- 230000008719 thickening Effects 0.000 description 1
- 150000003626 triacylglycerols Chemical class 0.000 description 1
- UFTFJSFQGQCHQW-UHFFFAOYSA-N triformin Chemical compound O=COCC(OC=O)COC=O UFTFJSFQGQCHQW-UHFFFAOYSA-N 0.000 description 1
- 210000003934 vacuole Anatomy 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 238000003809 water extraction Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D211/00—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings
- C07D211/04—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
- C07D211/06—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
- C07D211/36—Heterocyclic compounds containing hydrogenated pyridine rings, not condensed with other rings with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members 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 ring carbon atoms
- C07D211/40—Oxygen atoms
- C07D211/44—Oxygen atoms attached in position 4
- C07D211/46—Oxygen atoms attached in position 4 having a hydrogen atom as the second substituent in position 4
-
- 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
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
- A61K31/445—Non condensed piperidines, e.g. piperocaine
-
- 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
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
-
- 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)
- A61K36/60—Moraceae (Mulberry family), e.g. breadfruit or fig
- A61K36/605—Morus (mulberry)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
- A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
-
- 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
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/55—Liquid-liquid separation; Phase separation
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Pharmacology & Pharmacy (AREA)
- Medicinal Chemistry (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Epidemiology (AREA)
- Diabetes (AREA)
- Engineering & Computer Science (AREA)
- General Chemical & Material Sciences (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Obesity (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Endocrinology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Emergency Medicine (AREA)
- Alternative & Traditional Medicine (AREA)
- Biotechnology (AREA)
- Botany (AREA)
- Medical Informatics (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The invention relates to a mulberry leaf extract and a mulberry leaf multi-component mixture with the hypoglycemic effect prepared from the mulberry leaf extract. Particularly provides a mulberry leaf DNJ extract which is prepared by the following method: extracting folium Mori with acidic ethanol solution; the concentration of the ethanol solution is 65-75%, and the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 15-30 mL/g. The extraction method provided by the invention is simple to operate and mild in condition, and under the specific extraction condition of the invention, the yield and purity of DNJ in the obtained mulberry leaf DNJ extract are remarkably improved. The mulberry leaf DNJ extract is further used as a raw material, and is mixed with a mulberry leaf flavone extract and a mulberry leaf polysaccharide extract in a specific ratio to prepare a mulberry leaf multi-component mixture, so that the mulberry leaf multi-component mixture has an excellent blood sugar reducing effect and has a good application prospect in preparation of blood sugar reducing medicines.
Description
Technical Field
The invention belongs to the field of traditional Chinese medicine extraction, and particularly relates to a mulberry leaf extract and a mulberry leaf multi-component mixture with a hypoglycemic effect prepared from the mulberry leaf extract.
Background
Folium Mori is the leaf of Morus alba L of Morus of Moraceae, and contains essential amino acids, vitamins, minerals, flavonoids, alkaloids, polysaccharides, polyphenols, and other bioactive components. The composition comprises a polyhydroxy alkaloid, namely 1-Deoxynojirimycin (DNJ), which can be combined with diglycosidase such as maltase, sucrase, lactase and the like in the small intestine, so that disaccharide can not be further decomposed into monosaccharide to be absorbed by an organism, and the phenomenon of rapid rise of postprandial blood sugar is obviously inhibited, and the occurrence and development of diabetes are delayed. In addition, according to the records of the literature 'multi-component synergistic hypoglycemic effect of mulberry leaves, Wangdeli and Jiangjie, etc.', the mulberry leaf polysaccharide, flavone and DNJ mixture can achieve the synergistic hypoglycemic effect by alleviating oxidative damage in a diabetic mouse, repairing damaged islet cells and improving the resistance of insulin effector cells of the body. Therefore, DNJ is effectively extracted from the mulberry leaves, and the DNJ has very important value for preparing the medicine for treating diabetes.
At present, the research reports on the DNJ extraction process are less, and the existing extraction process mainly comprises a microwave-assisted method, a dilute acid extraction method and an ultrasonic-assisted method. The literature "orthogonal optimization process for extracting 1-deoxynojirimycin acid water in mulberry leaves" discloses a method for extracting DNJ in mulberry leaves by using acidic aqueous solution, and the optimal process is as follows: the granularity of mulberry leaf powder is 60 meshes, the extraction temperature is 55 ℃, the pH value is 3, the material-liquid ratio of hydrochloric acid solution is 1:20(g/mL), the extraction time is 1.5h, the DNJ yield is 0.062%, and the purity is 19.7%. The document "response surface method optimization of a process for extracting 1-deoxynojirimycin in mulberry leaves by ultrasonic assistance" discloses an ultrasonic-assisted extraction method using water as a solvent, wherein under the optimal extraction process conditions: the extraction time is 60min, the extraction temperature is 75 ℃, the ultrasonic power is 350W, the liquid-material ratio is 16mL/g, and the maximum extraction amount of DNJ is 1.873 mg/g.
The extraction methods all adopt water as an extraction solvent, so that the difficulty of removing the solvent in post-treatment is increased, and the extraction methods are not suitable for large-scale production; in addition, the yield of DNJ in the mulberry leaf extract obtained by the method is low, and the production cost is increased. Therefore, there is a need for an extraction process that can improve the extraction yield and purity of DNJ from mulberry leaf extract.
Disclosure of Invention
The invention aims to provide a mulberry leaf DNJ extract and a mulberry leaf multi-component mixture with the hypoglycemic effect prepared from the same.
The invention provides a mulberry leaf DNJ extract which is prepared by the following method: extracting folium Mori with acidic ethanol solution; the concentration of the ethanol solution is 65-75%, and the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 15-30 mL/g.
Further, the pH value of the acidic ethanol solution is 1-3, the concentration of the ethanol solution is 70-71%, the volume-mass ratio of the ethanol solution to the mulberry leaves is 20-26 mL/g, and the mulberry leaves are mulberry leaf powder which is ground and sieved by a 60-mesh sieve;
and/or the extraction temperature is 45-65 ℃ during extraction, and preferably 50-61 ℃.
Further, the extraction method is an extraction method, and comprises the following steps: extracting mulberry leaves serving as a raw material by using an ethanol solution with the pH value of 1-3; the concentration of the ethanol solution is 70% -71%, the extraction temperature is 60-61 ℃, the volume-mass ratio of the ethanol solution to the mulberry leaves is 20-21 mL/g, the extraction times are 1-2 times, and the extraction time is 148-150 min each time.
Further, the extraction method is an ultrasonic-assisted acidic ethanol extraction method, and comprises the following steps: taking mulberry leaves as a raw material, and extracting the mulberry leaves by using an ethanol solution with the pH of 2-3 under the ultrasonic wave with the power of 320-322W; the concentration of the ethanol solution is 70%, the extraction temperature is 50 ℃, the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 20mL/g, the extraction times are 1-2 times, and the extraction time is 30-31 min each time.
Further, the extraction method is a microwave-assisted acidic ethanol extraction method, and comprises the following steps: taking mulberry leaves as a raw material, and extracting the mulberry leaves by using an ethanol solution with the pH of 2-3 under the microwave with the power of 300-306W; the concentration of the ethanol solution is 70%, the volume-mass ratio of the ethanol solution to the mulberry leaves is 25-26 mL/g, the extraction times are 1-2 times, and the extraction time is 70s each time.
Further, the method also comprises a purification step, wherein the purification method comprises the following steps: purifying the extracted product by macroporous resin and/or ion exchange resin; preferably, the macroporous resin is NKA-9 macroporous resin, and the ion exchange resin is 001 × 7 cation exchange resin.
Further, when the macroporous resin is used for purification, the loading conditions are as follows: the sample concentration is 1.5mg/mL, the pH value is 8, and the sample flow rate is 1.5 BV/h; the elution conditions were as follows: the eluent is 75% ethanol, and the flow rate of the eluent is 2 BV/h;
and/or, when the purified product is subjected to ion exchange resin, the loading conditions are as follows: the sample concentration is 0.8mg/mL, the pH value is 2, and the sample flow rate is 1.5 BV/h; the elution conditions were as follows: the eluent is 1mol/L ammonia water, and the flow rate of the eluent is 1 BV/h.
The invention also provides a mulberry leaf multi-component mixture, which is obtained by mixing the mulberry leaf DNJ extract, the mulberry leaf flavone extract and the mulberry leaf polysaccharide extract; preferably, in the mulberry leaf multi-component mixture, the weight ratio of DNJ, flavone and polysaccharide is 1:6: 8.
Further, the mulberry leaf flavone extract is prepared by the following method: extracting folium Mori with ethanol solution; the concentration of the ethanol solution is 70%, the extraction temperature is 88 ℃, the volume-mass ratio of the ethanol solution to the mulberry leaves is 30mL/g, and the extraction time is 2.5 h;
or, the mulberry leaf flavone extract is prepared by the following method: extracting folium Mori as raw material with ethanol solution under ultrasonic wave of power 126W; the concentration of the ethanol solution is 51%, the extraction temperature is 70 ℃, the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 25mL/g, and the extraction time is 36 min;
and/or the mulberry leaf polysaccharide extract is prepared by the following method: extracting folium Mori with water; the extraction temperature is 82 ℃, the volume-to-mass ratio of water to mulberry leaves is 17mL/g, and the extraction time is 74 min.
Further, the preparation method of the mulberry leaf flavone extract also comprises the following purification steps: purifying the extracted product by AB-8 macroporous resin; preferably, the sample loading concentration during purification is 3mg/mL, and the sample loading flow rate is 2 BV/h; the eluent is 70% ethanol, and the flow rate of the eluent is 2 BV/h;
and/or the preparation method of the mulberry leaf polysaccharide extract further comprises the following purification steps: precipitating the product obtained after extraction with ethanol solution; preferably, the concentration of the ethanol solution is 95%, the volume of the ethanol solution is 3 times of that of the polysaccharide extract, the precipitation temperature is 2-4 ℃, and the precipitation time is 12 hours;
preferably, the preparation method of the mulberry leaf polysaccharide extract further comprises the following purification steps: purifying the system precipitated by the ethanol solution by AB-8 macroporous resin; preferably, the sample concentration during purification is 0.7mg/mL, the sample flow rate is 3BV/h, the eluent is distilled water, and the eluent flow rate is 2 BV/h.
Unless otherwise specified, the concentrations of the mixed solution of the present invention are volume concentrations, for example, 70% ethanol is a 70% ethanol solution.
The invention provides 3 methods for preparing a mulberry leaf DNJ extract, and experiments prove that: (1) when acidic ethanol is used for leaching, under the following specific extraction conditions: the ethanol concentration is 70-71%, the extraction time is 148-150 min, the liquid-material ratio is 20-21 mL/g, the extraction temperature is 60-61 ℃, and the DNJ yield in the obtained extract is up to 2.939 mg/g; (2) when the ultrasonic-assisted acidic ethanol extraction method is adopted for extraction, the extraction is carried out under the following specific extraction conditions: the ultrasonic power is 320-322W, the extraction time is 30-31 min, the liquid-material ratio is 2mL/g, the extraction temperature is 50 ℃, and the yield of DNJ in the obtained extract is up to 3.203 mg/g; (3) when the microwave-assisted acidic ethanol extraction method is adopted for extraction, the extraction is carried out under the following specific extraction conditions: the microwave power is 300-306W, the extraction time is 70s, the liquid-material ratio is 25-26 mL/g, and the yield of DNJ in the obtained extract is up to 3.097 mg/g.
The invention also utilizes NKA-9 macroporous resin and 001 multiplied by 7 cation exchange resin to purify the obtained mulberry leaf DNJ extract, and obviously improves the purity of DNJ in the extract under specific purification conditions.
In addition, the mulberry leaf multi-component mixture obtained by mixing the mulberry leaf DNJ extract prepared by the method as a raw material with a mulberry leaf flavone extract and a mulberry leaf polysaccharide extract in a specific ratio (the weight ratio of DNJ to flavone to polysaccharide is controlled to be 1:6:8) has an excellent blood sugar reducing effect, and has a good application prospect in preparation of blood sugar reducing medicines.
The extraction method provided by the invention is simple to operate and mild in condition, and under the specific extraction condition of the invention, the yield and purity of DNJ in the obtained mulberry leaf DNJ extract are obviously improved. The mulberry leaf DNJ extract has good prospect in preparing hypoglycemic drugs with the active ingredients only containing mulberry leaf DNJ or the mixture of mulberry leaf polysaccharide, flavone and DNJ.
Obviously, many modifications, substitutions, and variations are possible in light of the above teachings of the invention, without departing from the basic technical spirit of the invention, as defined by the following claims.
The present invention will be described in further detail with reference to the following examples. This should not be understood as limiting the scope of the above-described subject matter of the present invention to the following examples. All the technologies realized based on the above contents of the present invention belong to the scope of the present invention.
Detailed Description
The raw materials and equipment used in the invention are known products and are obtained by purchasing commercial products.
NKA-9 macroporous resin is purchased from the adsorption material science and technology company of Baowen, Hebei, with the specific surface area of 450-.
White mulberry (Morus alba L.) leaves are cleaned and dried in the shade, crushed by a universal crusher, sieved by a 60-mesh sieve to obtain mulberry leaf powder, and stored away from light for later use.
Example 1 extraction of folium Mori DNJ by acidic ethanol extraction
Weighing 2g of mulberry leaf powder, placing the mulberry leaf powder in a flat-bottomed flask, adding 42mL of ethanol with the concentration of 71% and the pH value adjusted by hydrochloric acid, extracting for 148min at the extraction temperature of 61 ℃, and extracting for 2 times to obtain the mulberry leaf DNJ extract 1. The DNJ yield was 2.939 (mg/g).
Example 2 extraction of folium Mori DNJ by ultrasonic-assisted acidic ethanol extraction
Weighing 2g of mulberry leaf powder, placing the mulberry leaf powder in a flat-bottomed flask, adding 40mL of ethanol with the concentration of 70% and the pH value adjusted by hydrochloric acid, extracting for 31min under the conditions that the extraction temperature is 50 ℃ and the ultrasonic power is 322W, and extracting for 2 times to obtain the mulberry leaf DNJ extract 2. The DNJ yield was 3.203 (mg/g).
Example 3 extraction of folium Mori DNJ by microwave-assisted acidic ethanol extraction
Weighing 2g of mulberry leaf powder, placing the mulberry leaf powder in a flat-bottomed flask, respectively adding 52mL of ethanol with the concentration of 70% and the pH value of 2 adjusted by hydrochloric acid, extracting for 70s under the microwave power of 306W, and extracting for 2 times to obtain the mulberry leaf DNJ extract 3. The DNJ yield was 3.097 (mg/g).
Example 4 purification of a folium Mori DNJ extract
The mulberry leaf DNJ extract 1(DNJ purity 8.2%) prepared in example 1 was used as a crude product, and was purified by NKA-9 macroporous resin. The sample loading conditions were as follows: the sample concentration is 1.5mg/mL, the pH value is 8, and the flow rate is 1.5 BV/h; the eluent is 75% ethanol, and the flow rate of the eluent is 2 BV/h.
After the purification process, the DNJ purity of the obtained mulberry leaf DNJ extract is increased from 8.2% to 28.4%, and the purity is improved by 3.5 times.
Example 5 further purification of folium Mori DNJ extract
The purified mulberry leaf DNJ extract (DNJ purity 28.4%) obtained in example 4 was used as a crude product and further purified using 001X 7 cation exchange resin. The sample loading conditions were as follows: the pH value of the sample is 2, the concentration is 0.8mg/mL, and the flow rate is 1.5 BV/h; the eluent is 1mol/L ammonia water, and the elution flow rate is 1 BV/h.
After the further purification process, the DNJ purity of the obtained mulberry leaf DNJ extract is increased from 28.4% to 70.4%, and the purity is further improved by 2.5 times.
Example 6 extraction of flavones from mulberry leaves by ethanol extraction
Weighing 2g of mulberry leaf powder, placing the mulberry leaf powder in a flat-bottomed flask, adding 60mL of 70% ethanol, and extracting for 2.5h at the extraction temperature of 88 ℃ to obtain the mulberry leaf flavone extract 1. The flavone yield was 4.16%.
The yield of flavone is the mass of flavone in the mulberry leaf flavone extract/the mass of mulberry leaf powder.
Example 7 ultrasonic-assisted extraction of flavonoids from mulberry leaves by ethanol extraction
Weighing 2g of mulberry leaf powder, placing the mulberry leaf powder in a flat-bottomed flask, adding 50mL of 51% ethanol, and extracting for 36min under the conditions that the ultrasonic temperature is 70 ℃ and the ultrasonic power is 126W to obtain the mulberry leaf flavone extract 2. The flavone yield was 3.79%.
Example 8 purification of folium Mori flavone extract
The mulberry leaf flavone extract 1 (the flavone purity is 8.49%) prepared in example 6 is used as a crude product, and AB-8 macroporous resin is adopted for purification. The sample loading conditions were as follows: the sample concentration is 3mg/mL, and the flow rate is 2 BV/h; the eluent is 70% ethanol, and the flow rate of the eluent is 2 BV/h.
After the purification process, the purity of the flavone in the mulberry leaf flavone extract is increased from 8.49% to 52.34%, and the purity is improved by 6.16 times.
The extract purified by the AB-8 macroporous resin is subjected to component analysis by an HPLC method, and is found to mainly contain 5 flavonoids, and the content of each flavonoid is as follows: 86.50mg/g chlorogenic acid, 60.94mg/g isoquercitrin, 35.12mg/g astragalin, 23.75mg/g rutin, 14.87mg/g quercetin.
Example 9 extraction of polysaccharides from Mulberry leaves Using Water extraction ethanol precipitation
(1) Preparation of mulberry leaf polysaccharide extract
Sieving folium Mori powder with 40 mesh sieve, and defatting with petroleum ether. Weighing 2g of mulberry leaf powder, placing the mulberry leaf powder into a round-bottom flask, adding distilled water, and extracting for 2 times under the conditions that the liquid-material ratio is 17:1mL/g, the extraction temperature is 82 ℃, and the extraction time is 74 min. Centrifuging the obtained extractive solution, collecting supernatant, and vacuum concentrating for 3 times to obtain folium Mori polysaccharide concentrated solution.
(2) Ethanol precipitation
And (2) adding the mulberry leaf polysaccharide concentrated solution obtained in the step (1) into 95% ethanol solution with the volume being 3 times that of the concentrated solution, standing and precipitating for 12 hours at low temperature (2-4 ℃), and performing high-speed centrifugal separation (3000r/min, 15min) to obtain a mulberry leaf polysaccharide extract, wherein the polysaccharide yield is 14.24%.
The yield of the polysaccharide is the mass of the polysaccharide in the mulberry leaf polysaccharide extract/the mass of the mulberry leaf powder.
Example 10 purification of polysaccharides from Mulberry leaves
The ethanol-precipitated polysaccharide extract of mulberry leaves obtained in example 9 was used as a crude product, and was purified using AB-8 macroporous resin. The sample loading conditions were as follows: the sample concentration is 0.7mg/mL, and the flow rate is 3 BV/h; the eluent is distilled water, and the flow rate of the eluent is 2 BV/h.
After the purification process, the purity of the polysaccharide of the mulberry leaf polysaccharide extract is 32.6%, the yield of the polysaccharide is 19.3%, the retention rate of the polysaccharide is 56.5%, the deproteinization rate is 68.4%, and the decoloration rate is 50.8%.
EXAMPLE 11 preparation of a Multi-component mixture of Mulberry leaves
Mixing the DNJ extract of mulberry leaves obtained in any one of examples 1 to 5, the flavone extract of mulberry leaves obtained in any one of examples 6 to 8 and the polysaccharide extract of mulberry leaves obtained in any one of examples 9 to 10, and controlling DNJ: and (3) flavone: the weight ratio of the polysaccharides is 1:6:8, and the mulberry leaf multi-component mixture is obtained.
The beneficial effects of the present invention are demonstrated by the following experimental examples.
Experimental example 1 test of DNJ content and purity in Mulberry leaf DNJ extract
1.1 preparation of reagents
(1) DNJ standard solution preparation: accurately weighing 5mg of DNJ standard substance into a beaker, adding a small amount of 0.05mol/L hydrochloric acid for dissolving, adding a certain amount of distilled water, transferring into a 50mL volumetric flask, washing the beaker for three times, transferring the beaker and a glass rod into the volumetric flask together, and metering to 50mL to obtain a DNJ standard solution of 0.1 mg/mL.
(2) Preparing a wagner reagent: weighing l g of I2And 10g KI, dissolving in a small amount of distilled water in a beaker, adding a small amount of acetic acid, heating, diluting to 100mL with distilled water, and storing in a brown jar for later use.
1.2 determination of the specific absorption wavelength
DNJ belongs to a piperidine alkaloid, forms a complex with wagner reagent under an acidic condition, and can be detected under an ultraviolet condition. And (3) taking 5mL of 0.l mg/mL DNJ standard solution into a 25mL volumetric flask, adding a small amount of wagner reagent, and scanning in a wavelength range of 240nm-420nm of an ultraviolet spectrophotometer to determine the specific absorption wavelength of DNJ.
1.3 drawing a standard curve
Respectively sucking 10 muL, 20 muL, 30 muL, 40 muL and 50 muL of DNJ standard sample solution, adding equal amount of iodine-potassium iodide reagent as a color developing agent into 5 tubes with plugs of different numbers, adding distilled water to make the total volume of the sample solution be 5mL, adding equal amount of iodine-potassium iodide reagent into another tube with plugs, gradually adding distilled water to adjust the volume of the solution to be 5mL, and using the solution as a blank control. The absorbance of each test tube solution was measured at the maximum absorption wavelength, and a standard curve was drawn with DNJ concentration as the abscissa and absorbance as the ordinate.
1.4 determination of the DNJ content
(1) Adding a certain amount of sample solution into a 25mL volumetric flask, adding a wagner reagent, fixing the volume, oscillating, measuring the absorbance at the maximum absorption wavelength, calculating the corresponding sample concentration according to a standard curve, and calculating the DNJ yield (namely the DNJ content) according to the following formula:
c: standard Curve DNJ concentration (μ g/mL)
n: dilution factor
V: sample volume (mL)
W: mulberry leaf mass (g)
1.5 determination of the DNJ purity
Taking a mulberry leaf DNJ extract to be detected, and calculating according to the following formula:
c: standard Curve DNJ concentration (mg/mL)
n: dilution factor
V: sample volume (mL)
M: oven dried weight (g) of sample to be tested
1.6 results of the experiment
The standard curve of DNJ was obtained using the method described above: 0.1641x-0.0013, R20.9991. The DNJ yields in the examples were calculated separately and are shown in table 1. The yield of DNJ extracted by adopting the ultrasonic-assisted acid ethanol method is 3.203mg/g, which is improved by 9.0% compared with the yield of DNJ extracted by adopting the acid ethanol method; the yield of DNJ extracted by adopting the microwave-assisted acid ethanol method is 3.097mg/g, which is improved by 5.4 percent compared with the acid ethanol method.
As shown in Table 2, the DNJ purity of the mulberry leaf DNJ extract obtained by purifying the NKA-9 macroporous resin in example 4 is increased from 8.2% to 28.4%, and the purity is improved by 3.5 times. Example 5 the DNJ purity of the mulberry leaf DNJ extract further purified by 001 × 7 cation exchange resin was increased from 28.4% to 70.4%, which was further increased 2.5 times.
TABLE 1 DNJ yield calculation results for each example
Sample (I) | Extraction method | DNJ yield (mg/g) |
Example 1 | Acid ethanol extraction method | 2.939 |
Example 2 | Ultrasonic-assisted acidic ethanol leaching method | 3.203 |
Example 3 | Microwave-assisted acidic ethanol extraction method | 3.097 |
TABLE 2 calculation of DNJ purity for each example
Sample (I) | Extraction method | DNJ purity |
Example 1 | Acid ethanol extraction method | 8.2% |
Example 4 | NKA-9 macroporous resin purification | 28.4% |
Example 5 | 001 × 7 cation exchange resin purification | 70.4% |
Experimental example 2 testing of flavone content and purity in flavone extract from Mori folium
2.1 principle of aluminum nitrate-sodium nitrite method
After the flavonoid compound reacts with the aluminum nitrate, the formed flavone aluminum salt complex ion is yellow, and the color depth of the complex ion is in a certain proportion relation with the flavone content, so that the quantitative determination can be realized.
2.2 drawing Standard Curve
Weighing 10.00mg of rutin standard substance, dissolving in 65% ethanol solution, and diluting to 50mL to obtain 0.2mg/mL rutin standard solution. Respectively sucking 0, 0.5mL, 1.0mL, 2.0mL, 3.0mL and 4.0mL rutin standard solution into a 10mL volumetric flask, and adding 0.5mL NaNO2(1:20), shaking and standing for 6min, adding 0.5mL Al (NO)3)3(1:10), shaking uniformly and standing for 6min, adding 4mL of 1mol/L NaOH solution and 65% ethanol solution to constant volume, shaking uniformly and standing for 10-20 min, taking the first tube solution as a blank control, and measuring absorbance at the wavelength of 510 nm. And (4) drawing a standard curve by taking the concentration of the rutin standard solution as a horizontal coordinate and the absorbance as a vertical coordinate.
2.3 measurement of flavone content in Mulberry leaves
Taking 0.5mL of sample solution to be detected, putting the sample solution into a volumetric flask, operating according to a method for drawing a standard curve, and calculating the flavone content (namely the flavone yield) according to a formula. The formula is as follows:
in the formula: c-standard curve concentration of flavone (mg/mL)
n-dilution multiple
V-volume of sample liquid (mL)
W-Mulberry leaf quality (g)
2.4 purity determination of folium Mori flavone
Taking the mulberry leaf flavone extract to be detected, and calculating the purity of the flavone according to the following formula.
In the formula: c-standard curve concentration of flavone (mg/mL)
n-dilution multiple
V-volume of sample liquid (mL)
M-weight of extract after lyophilization (g)
2.5 results of the experiment
Obtaining standard rutin yeast by the methodLine: a (y) 10.949(x) +0.0138, R20.9991. The flavone yield of the extract obtained in example 6 was calculated to be 4.16%, and the flavone yield of the extract obtained in example 7 was calculated to be 3.79%.
In example 8, the purity of the flavone extract purified by AB-8 macroporous resin is increased from 8.49% to 52.34%, and the purity is improved by 6.16 times.
Experimental example 3 testing of polysaccharide content and purity in polysaccharide extract of Mori folium
3.1 polysaccharide Retention assay
The polysaccharide content was determined by phenol-sulfuric acid method and the polysaccharide retention was calculated as follows.
3.2 determination of the deproteinization Rate
Protein content was determined by Coomassie Brilliant blue method. Preparation of Coomassie brilliant blue G-250: 100mg of Coomassie brilliant blue G-250 was weighed, dissolved in 50mL of 90% ethanol, 100mL of 85% (W/V) phosphoric acid was added, and the volume was adjusted to 1000mL with distilled water. Preparing 100 mu g/mL bovine serum albumin, respectively taking 0mL, 0.1mL, 0.2mL, 0.3mL, 0.4mL, 0.5mL and 0.6mL in a test tube, supplementing 1mL with 0.15M NaCl solution, adding 5mL of Coomassie brilliant blue solution prepared in advance, standing for 10min, measuring the light absorption value at 595nm, and drawing a standard curve. The deproteinization rate was calculated by the following formula.
3.3 determination of decolorization ratio
The preparation method comprises the steps of preparing a mulberry leaf crude polysaccharide solution with the concentration of 1%, and carrying out visible-ultraviolet full-wavelength scanning on the solution, wherein the result shows that the solution has no maximum absorption wavelength. According to the complementary color principle, the solution presents a color which is the complementary color of the light absorbed by the solution, and the solution mainly absorbs the visible light in the blue waveband because the polysaccharide solution is orange yellow after being diluted before and after decolorization. Therefore, the wavelength of 450nm at the center of this wavelength band was selected as the detection wavelength, and the absorbance of the solution was measured. And the decolorization ratio was calculated according to the following formula.
3.4 determination of purity of purified polysaccharides from Mulberry leaves
Taking a mulberry leaf polysaccharide extract to be detected, and calculating the purity of polysaccharide according to the following formula:
in the formula: C-Standard Curve polysaccharide concentration (mg/mL)
n-dilution multiple
V-volume of sample liquid (mL)
M-weight of extract after lyophilization (g)
3.5 results of the experiment
After the AB-8 macroporous resin purification process, the purity of the polysaccharide of the mulberry leaf extract obtained in example 10 is 32.6%, the yield of the polysaccharide is 19.3%, the polysaccharide retention rate is 56.5%, the deproteinization rate is 68.4%, and the decoloration rate is 50.8%.
Experimental example 4 screening of folium Mori DNJ extraction Using acidic ethanol
1. Experimental methods
Referring to the process of example 1, the ethanol concentration, extraction time, liquid-to-material ratio, and extraction temperature were changed according to table 3 to extract DNJ extracts from mulberry leaves, respectively, and the DNJ yield in each process was calculated according to the method of experimental example 1. Repeat 5 times under each process condition and take the average.
2. Results of the experiment
As shown in Table 3, it can be seen that the highest DNJ yield of the obtained extract was obtained when the mulberry leaves were extracted with acidic ethanol, wherein the ethanol concentration was 70 to 71%, the extraction time was 148 to 150min, the liquid-to-material ratio was 20 to 21mL/g, and the extraction temperature was 60 to 61 ℃.
TABLE 3 screening experiment results of the process for extracting DNJ from mulberry leaves by acidic ethanol method
Experimental example 5 Process screening for extracting folium Mori DNJ by ultrasonic-assisted acidic ethanol extraction method
1. Experimental methods
Referring to the process of example 2, the ultrasonic power, extraction time, liquid-to-material ratio, and extraction temperature were changed according to table 4 to extract respective mulberry DNJ extracts, and the DNJ yield in each process was calculated according to the method of experimental example 1.
2. Results of the experiment
As shown in Table 4, it can be seen that, when mulberry leaves are extracted by the ultrasonic-assisted acidic ethanol extraction method, the highest DNJ yield is obtained in the obtained extract when the ultrasonic power is 320-322W, the extraction time is 30-31 min, the liquid-material ratio is 20mL/g, and the extraction temperature is 50 ℃.
TABLE 4 technological screening test results of DNJ extraction from mulberry leaves by ultrasonic-assisted acidic ethanol extraction method
Experimental example 6 Process screening for extracting folium Mori DNJ by microwave-assisted acidic ethanol extraction method
1. Experimental methods
Referring to the process of example 3, the microwave power, extraction time, and liquid-to-material ratio were changed according to table 5 to extract respective mulberry leaf DNJ extracts, and the DNJ yield in each process was calculated according to the method of experimental example 1.
2. Results of the experiment
As shown in Table 5, it can be seen that when the mulberry leaves are extracted by the microwave-assisted acidic ethanol extraction method, the highest DNJ yield is obtained in the obtained extract when the microwave power is 300-306W, the extraction time is 70s, and the liquid-to-material ratio is 25-26 mL/g.
TABLE 5 technological screening test results of extracting DNJ from mulberry leaves by microwave-assisted acidic ethanol extraction method
Experimental example 7 characterization of hypoglycemic Effect of Multi-component mixture of Mulberry leaves
1. Experimental methods
Experimental drugs: the multi-component mixture of mulberry leaves obtained in example 11; the positive drug is metformin.
Establishing a diabetic rat model: healthy male SD rats are taken and fed with high-fat feed for 6 weeks, and are subjected to intraperitoneal injection once with streptozotocin solution (30 mg/kg. BW), fasting blood glucose is measured after 72 hours, and the success of the model building of the diabetic rats is determined according to the index that the fasting blood glucose is more than or equal to 11.1 mmol/L.
Mulberry leaf multicomponent mixture (100 mg/kg. d. BW) or positive drug (185 mg/kg. d. BW) was fed (gavage) to modeled SD male rats for a total of 6 weeks of treatment. The modeled SD male rats fed the same volume of saline served as the model control group, and the normal SD rats fed the same volume of saline served as the blank control group.
After 6 weeks of treatment, rats were tested for blood glucose levels, insulin levels; testing rat serum for total cholesterol, triglycerides, LDL-C and HDL-C; antioxidant enzymes in rat serum were tested: activities of superoxide dismutase SOD, glutathione peroxidase GSH-Px, catalase CAT, lipid peroxidation malondialdehyde MDA level; testing of adipocytokines in rat serum: free fatty acids, TNF- α, leptin, and inflammatory factor C-reactive protein levels; testing the kidney function indexes of urea nitrogen and creatinine level in the serum of the rat; rat liver tissue, pancreas tissue, kidney tissue sections were observed.
2. Results of the experiment
After 6 weeks, the blood sugar level and the insulin level of the rats in the mulberry leaf multi-component mixture group are only 26.06 percent and 53.30 percent (P is less than 0.05) of those in the model control group, are equivalent to those in the positive drug metformin control group (P is more than 0.05), and have better blood sugar reduction effect.
After 6 weeks, the total cholesterol, triglyceride, LDL-C and HDL-C in the serum of rats in the mulberry leaf multi-component mixture group are 79.10%, 49.50%, 74.24% and 1.33 times (P <0.05) of those in the model control group respectively, and have no significant difference (P >0.05) with the blank control group and the positive control group, which indicates that the mulberry leaf multi-component mixture can effectively improve the lipid metabolism disorder of the diabetic rats.
After 6 weeks, antioxidant enzymes in serum of rats of the mulberry leaf multicomponent mixture group: the activities of superoxide dismutase SOD, glutathione peroxidase GSH-Px and catalase CAT are respectively 1.26 times, 1.34 times and 1.36 times of the model group (P <0.05), the level of lipid peroxidation malondialdehyde MDA is 84.06 percent of the model group (P <0.05), and the activity is not significantly different from that of a positive control group (P > 0.05). The mulberry leaf multi-component mixture improves the oxidative stress state in the diabetic rat body by improving the antioxidant enzyme activity in the diabetic rat body and inhibiting lipid peroxidation.
After 6 weeks, the ratio of adipocyte factor in serum of rats in the mulberry leaf multicomponent mixture group: free fatty acid, TNF-alpha, leptin and inflammatory factor C-reactive protein levels were reduced to 68.63%, 88.25%, 84.15% and 41.03% (P <0.05) of the model control group, while adipokine adiponectin levels increased 1.16-fold (P <0.05) of the model group, all without significant difference from the positive control metformin group (P > 0.05). The mulberry leaf extract can effectively reduce the abnormal high expression of fat cell factors and C-reactive protein in the body of a diabetic rat and improve the insulin resistance state; and activates adiponectin expression, enhancing insulin sensitivity.
After 6 weeks, the kidney function indexes of urea nitrogen and creatinine levels in the serum of rats in the mulberry leaf multi-component mixture group are 71.31% and 62.23% of the model group respectively (P <0.05), and are close to the positive control group and have no statistical difference (P > 0.05).
After 6 weeks, the liver fat vacuoles of rats in the mulberry leaf multi-component mixture group are obviously reduced, the cells are orderly arranged, the shapes are relatively complete, and the liver lobule structures are complete. The islet cells are clear in boundary and regular in arrangement, and the number of islet cells is increased. The glomerulus volume is slightly increased, renal tubules are not transparent, the thickening of the glomerulus and renal tubule basement membrane is obviously reduced, and the pathological state of renal tissues is improved. The mulberry leaf extract is prompted to have the effects of improving the hepatic steatosis of diabetic rats caused by high-fat diet combined with intraperitoneal injection of streptozotocin, repairing damaged islet cells, relieving the damage of continuous hyperglycemia to the kidney and the like.
Therefore, the mulberry leaf multi-component mixture prepared by the invention can relieve oxidative stress injury of tissues such as liver, pancreas and the like, improve insulin resistance state and enhance insulin sensitivity by improving the activity of antioxidant enzyme in a diabetic rat, thereby effectively reducing the blood sugar level of the diabetic rat, improving lipid metabolism disorder of the rat and having excellent blood sugar reduction effect.
In conclusion, the invention provides a mulberry leaf DNJ extract which is prepared by extracting mulberry leaves serving as raw materials by using an acidic ethanol solution; the concentration of the ethanol solution is 65-75%, and the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 15-30 mL/g. In the method, the extraction mode can be leaching, ultrasonic-assisted acid ethanol extraction or microwave-assisted acid ethanol extraction. Under the specific extraction conditions of the invention, the yield and purity of DNJ in the obtained mulberry leaf DNJ extract are obviously improved. The mulberry leaf DNJ extract has good prospect in preparing hypoglycemic drugs with the active ingredients only containing mulberry leaf DNJ or the mixture of mulberry leaf polysaccharide, flavone and DNJ.
Particularly, the mulberry leaf multi-component mixture consisting of the mulberry leaf DNJ extract, the mulberry leaf flavone extract and the mulberry leaf polysaccharide extract in a ratio of 1:6:8 can improve the activity of antioxidant enzymes in a diabetic rat body, relieve oxidative stress damage of tissues such as liver, pancreas and the like, improve the insulin resistance state and enhance insulin sensitivity, thereby effectively reducing the blood sugar level of the diabetic rat, improving lipid metabolism disorder of the rat, having excellent blood sugar reduction effect and wide application prospect.
Claims (10)
1. A mulberry leaf DNJ extract is characterized in that: the preparation method comprises the following steps: extracting folium Mori with acidic ethanol solution; the concentration of the ethanol solution is 65-75%, and the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 15-30 mL/g.
2. The extract as claimed in claim 1, wherein: the pH value of the acidic ethanol solution is 1-3, the concentration of the ethanol solution is 70-71%, the volume-mass ratio of the ethanol solution to the mulberry leaves is 20-26 mL/g, and the mulberry leaves are mulberry leaf powder which is crushed and then sieved by a 60-mesh sieve;
and/or the extraction temperature is 45-65 ℃ during extraction, and preferably 50-61 ℃.
3. Extract according to claim 1 or 2, characterized in that: the extraction method is an extraction method and comprises the following steps: extracting mulberry leaves serving as a raw material by using an ethanol solution with the pH value of 1-3; the concentration of the ethanol solution is 70% -71%, the extraction temperature is 60-61 ℃, the volume-mass ratio of the ethanol solution to the mulberry leaves is 20-21 mL/g, the extraction times are 1-2 times, and the extraction time is 148-150 min each time.
4. Extract according to claim 1 or 2, characterized in that: the extraction method is an ultrasonic-assisted acidic ethanol extraction method, and comprises the following steps: taking mulberry leaves as a raw material, and extracting the mulberry leaves by using an ethanol solution with the pH of 2-3 under the ultrasonic wave with the power of 320-322W; the concentration of the ethanol solution is 70%, the extraction temperature is 50 ℃, the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 20mL/g, the extraction times are 1-2 times, and the extraction time is 30-31 min each time.
5. Extract according to claim 1 or 2, characterized in that: the extraction method is a microwave-assisted acidic ethanol extraction method and comprises the following steps: taking mulberry leaves as a raw material, and extracting the mulberry leaves by using an ethanol solution with the pH of 2-3 under the microwave with the power of 300-306W; the concentration of the ethanol solution is 70%, the volume-mass ratio of the ethanol solution to the mulberry leaves is 25-26 mL/g, the extraction times are 1-2 times, and the extraction time is 70s each time.
6. The extract according to any one of claims 1 to 5, wherein: the method also comprises a purification step, wherein the purification method comprises the following steps: purifying the extracted product by macroporous resin and/or ion exchange resin; preferably, the macroporous resin is NKA-9 macroporous resin, and the ion exchange resin is 001 × 7 cation exchange resin.
7. The extract as claimed in claim 6, wherein: when the macroporous resin is used for purification, the loading conditions are as follows: the sample concentration is 1.5mg/mL, the pH value is 8, and the sample flow rate is 1.5 BV/h; the elution conditions were as follows: the eluent is 75% ethanol, and the flow rate of the eluent is 2 BV/h;
and/or, when the purified product is subjected to ion exchange resin, the loading conditions are as follows: the sample concentration is 0.8mg/mL, the pH value is 2, and the sample flow rate is 1.5 BV/h; the elution conditions were as follows: the eluent is 1mol/L ammonia water, and the flow rate of the eluent is 1 BV/h.
8. A mulberry leaf multi-component mixture is characterized in that: the mulberry leaf multicomponent mixture is obtained by mixing the mulberry leaf DNJ extract as claimed in any one of claims 1 to 7 with a mulberry leaf flavone extract and a mulberry leaf polysaccharide extract; preferably, in the mulberry leaf multi-component mixture, the weight ratio of DNJ, flavone and polysaccharide is 1:6: 8.
9. The multi-component mulberry leaf mixture of claim 8, wherein: the mulberry leaf flavone extract is prepared by the following method: extracting folium Mori with ethanol solution; the concentration of the ethanol solution is 70%, the extraction temperature is 88 ℃, the volume-mass ratio of the ethanol solution to the mulberry leaves is 30mL/g, and the extraction time is 2.5 h;
or, the mulberry leaf flavone extract is prepared by the following method: extracting folium Mori as raw material with ethanol solution under ultrasonic wave of power 126W; the concentration of the ethanol solution is 51%, the extraction temperature is 70 ℃, the volume-to-mass ratio of the ethanol solution to the mulberry leaves is 25mL/g, and the extraction time is 36 min;
and/or the mulberry leaf polysaccharide extract is prepared by the following method: extracting folium Mori with water; the extraction temperature is 82 ℃, the volume-to-mass ratio of water to mulberry leaves is 17mL/g, and the extraction time is 74 min.
10. The multi-component mulberry leaf mixture of claim 9, wherein: the preparation method of the mulberry leaf flavone extract also comprises the following purification steps: purifying the extracted product by AB-8 macroporous resin; preferably, the sample loading concentration during purification is 3mg/mL, and the sample loading flow rate is 2 BV/h; the eluent is 70% ethanol, and the flow rate of the eluent is 2 BV/h;
and/or the preparation method of the mulberry leaf polysaccharide extract further comprises the following purification steps: precipitating the product obtained after extraction with ethanol solution; preferably, the concentration of the ethanol solution is 95%, the volume of the ethanol solution is 3 times of that of the polysaccharide extract, the precipitation temperature is 2-4 ℃, and the precipitation time is 12 hours;
preferably, the preparation method of the mulberry leaf polysaccharide extract further comprises the following purification steps: purifying the system precipitated by the ethanol solution by AB-8 macroporous resin; preferably, the sample concentration during purification is 0.7mg/mL, the sample flow rate is 3BV/h, the eluent is distilled water, and the eluent flow rate is 2 BV/h.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202010470759.1A CN111533680A (en) | 2020-05-28 | 2020-05-28 | Folium Mori extract and folium Mori multicomponent mixture with blood sugar lowering effect prepared from the same |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202010470759.1A CN111533680A (en) | 2020-05-28 | 2020-05-28 | Folium Mori extract and folium Mori multicomponent mixture with blood sugar lowering effect prepared from the same |
Publications (1)
Publication Number | Publication Date |
---|---|
CN111533680A true CN111533680A (en) | 2020-08-14 |
Family
ID=71974485
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202010470759.1A Pending CN111533680A (en) | 2020-05-28 | 2020-05-28 | Folium Mori extract and folium Mori multicomponent mixture with blood sugar lowering effect prepared from the same |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN111533680A (en) |
Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN112546091A (en) * | 2020-12-16 | 2021-03-26 | 承德医学院 | Preparation process and detection method of mulberry leaf extract dropping pill |
CN112603941A (en) * | 2021-01-13 | 2021-04-06 | 湘潭市中心医院 | Preparation method of traditional Chinese medicine extract for regulating glycolipid metabolism |
CN113712206A (en) * | 2021-09-16 | 2021-11-30 | 湖南希尔天然药业有限公司 | A composition containing folium Mori DNJ and folium Mori peptide and its preparation method |
CN114539132A (en) * | 2022-04-13 | 2022-05-27 | 重庆工商大学 | Method for performing DNJ (deoxyribose nucleic acid) extraction on mulberry leaves by hydrothermal acid control |
CN115316497A (en) * | 2022-08-15 | 2022-11-11 | 广东省农业科学院蚕业与农产品加工研究所 | Mulberry leaf extract, mandarin fish feed and application thereof |
Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101209284A (en) * | 2007-12-24 | 2008-07-02 | 天津大学 | Method for continuously extracting alkaloids, flavone and polysaccharide active components from mulberry leaves |
CN101412703A (en) * | 2008-11-17 | 2009-04-22 | 江苏科技大学 | Composite extracting technique for coproduction of mulberry tea flavone, polysaccharide and alkaloid |
CN102101840A (en) * | 2011-02-15 | 2011-06-22 | 杭州惠远实业有限公司 | Method for extracting and separating high-purity 1-Deoxynojirimycin from folium mori |
CN103755623A (en) * | 2014-01-03 | 2014-04-30 | ***广州总医院 | Response surface method optimized method for extracting 1-deoxynojirimycin in mulberry leaves with acidic ethanol solution |
CN107536871A (en) * | 2017-09-13 | 2018-01-05 | 中国科学院动物研究所 | A kind of preparation method for Traditional Chinese medicine for decreasing blood sugar extract |
CN114539132A (en) * | 2022-04-13 | 2022-05-27 | 重庆工商大学 | Method for performing DNJ (deoxyribose nucleic acid) extraction on mulberry leaves by hydrothermal acid control |
-
2020
- 2020-05-28 CN CN202010470759.1A patent/CN111533680A/en active Pending
Patent Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN101209284A (en) * | 2007-12-24 | 2008-07-02 | 天津大学 | Method for continuously extracting alkaloids, flavone and polysaccharide active components from mulberry leaves |
CN101412703A (en) * | 2008-11-17 | 2009-04-22 | 江苏科技大学 | Composite extracting technique for coproduction of mulberry tea flavone, polysaccharide and alkaloid |
CN102101840A (en) * | 2011-02-15 | 2011-06-22 | 杭州惠远实业有限公司 | Method for extracting and separating high-purity 1-Deoxynojirimycin from folium mori |
CN103755623A (en) * | 2014-01-03 | 2014-04-30 | ***广州总医院 | Response surface method optimized method for extracting 1-deoxynojirimycin in mulberry leaves with acidic ethanol solution |
CN107536871A (en) * | 2017-09-13 | 2018-01-05 | 中国科学院动物研究所 | A kind of preparation method for Traditional Chinese medicine for decreasing blood sugar extract |
CN114539132A (en) * | 2022-04-13 | 2022-05-27 | 重庆工商大学 | Method for performing DNJ (deoxyribose nucleic acid) extraction on mulberry leaves by hydrothermal acid control |
Non-Patent Citations (9)
Title |
---|
季涛,等: "基于α-葡萄糖苷酶抑制活性评价桑叶多组分药效相互作用研究", 《中国中药杂志》 * |
张月,等: "桑叶黄酮提取物制备及其体外抗氧化作用研究", 《中国食品添加剂》 * |
杨小蓉,等: "桑叶DNJ的分离纯化及其生物活性研究进展", 《食品工业》 * |
玄光善,等: "桑叶有效成分降糖作用研究", 《食品科学》 * |
王兴婷,等: "桑叶DNJ提取物体外降血糖及抗氧化作用研究", 《食品科技》 * |
王德萍,等: "桑叶多组分协同降血糖作用", 《现代预防医学》 * |
苏楠,等: "响应面分析法优化桑叶中1-脱氧野尻霉素的稀酸乙醇浸提工艺", 《中国实验方剂学杂志》 * |
贺伟强,等: "桑叶活性成分药理作用研究进展", 《北方园艺》 * |
鲁慧,等: "新疆桑叶生物活性成分种属与季节差异性研究", 《食品科技》 * |
Cited By (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN112546091A (en) * | 2020-12-16 | 2021-03-26 | 承德医学院 | Preparation process and detection method of mulberry leaf extract dropping pill |
CN112603941A (en) * | 2021-01-13 | 2021-04-06 | 湘潭市中心医院 | Preparation method of traditional Chinese medicine extract for regulating glycolipid metabolism |
CN113712206A (en) * | 2021-09-16 | 2021-11-30 | 湖南希尔天然药业有限公司 | A composition containing folium Mori DNJ and folium Mori peptide and its preparation method |
CN114539132A (en) * | 2022-04-13 | 2022-05-27 | 重庆工商大学 | Method for performing DNJ (deoxyribose nucleic acid) extraction on mulberry leaves by hydrothermal acid control |
CN114539132B (en) * | 2022-04-13 | 2023-09-12 | 重庆工商大学 | DNJ method for hydrothermally acid-controlled alcohol extraction of mulberry leaves |
CN115316497A (en) * | 2022-08-15 | 2022-11-11 | 广东省农业科学院蚕业与农产品加工研究所 | Mulberry leaf extract, mandarin fish feed and application thereof |
CN115316497B (en) * | 2022-08-15 | 2023-05-09 | 广东省农业科学院蚕业与农产品加工研究所 | Mulberry leaf extract, mandarin fish feed and application thereof |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN111533680A (en) | Folium Mori extract and folium Mori multicomponent mixture with blood sugar lowering effect prepared from the same | |
CN109078042B (en) | Method for extracting active ingredients from radix puerariae through compound biological enzymolysis | |
CN107029120B (en) | Preparation method of dendrobium and polygonatum sibiricum soft extract | |
CN108047343B (en) | Preparation method and application of fritillaria pallidiflora total polysaccharide | |
CN110101728B (en) | Combined extraction method of purslane polysaccharide and total flavonoids based on micelle medium treatment | |
CN115869345B (en) | Shennong chrysanthemum flavonoid compound and preparation method and application thereof | |
CN113278088A (en) | Sargassum fusiforme polysaccharide with obvious intestinal mucosa repair activity and preparation method and application thereof | |
CN113024685A (en) | Low-molecular-weight Dictyophora indusiata (Vent. Ex pers) Fisch trum-Dictyophora (Vent. Ex pers) Fisch trum et Schott polysaccharide, and preparation method and application thereof | |
CN110974870B (en) | Preparation method of crude extract of mulberry polyphenol | |
CN114848701B (en) | Preparation method of phyllanthus emblica extract | |
CN110066349B (en) | Low-molecular-weight blackberry polysaccharide and preparation method thereof | |
CN114177218B (en) | Mulberry leaf extract rich in 1-deoxynojirimycin and preparation method thereof | |
CN114699468A (en) | Preparation method of vine tea extract | |
CN108358822B (en) | Method for continuously extracting multiple active ingredients from defective and defective wolfberry fruits | |
CN114712416B (en) | Method for efficiently and synchronously extracting flavone, alkaloid and polyphenol in lotus leaves by using water-borne method | |
CN116925163A (en) | Method for obtaining rose glycoside by low-frequency ultrasound | |
CN113694152B (en) | Method for obtaining coix seed extracting solution by high-stability enzymolysis method | |
CN112442136A (en) | Method for extracting functional components from tremella | |
CN114904294A (en) | Preparation method of high-yield tea flavone | |
CN114632105A (en) | Comprehensive utilization method and application of peony seed meal | |
CN109464500B (en) | Method for extracting flavonoids from mulberry leaves by using complex enzyme | |
CN111035622B (en) | Angelica keiskei chalcone microcapsule and preparation method thereof | |
CN112220776A (en) | Chickpea saponin microcapsule and preparation method and application thereof | |
NL2032187B1 (en) | Novel anti-hypoxia polysaccharide from dendrobium officinale, as well as preparation method by steam explosion and application thereof | |
CN115191489B (en) | Preparation method of gastrodia elata protein emulsion |
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 | ||
RJ01 | Rejection of invention patent application after publication | ||
RJ01 | Rejection of invention patent application after publication |
Application publication date: 20200814 |