CN110964659A - 降血糖功效的植物乳杆菌及其筛选方法和应用药剂及食品 - Google Patents
降血糖功效的植物乳杆菌及其筛选方法和应用药剂及食品 Download PDFInfo
- Publication number
- CN110964659A CN110964659A CN201910709658.2A CN201910709658A CN110964659A CN 110964659 A CN110964659 A CN 110964659A CN 201910709658 A CN201910709658 A CN 201910709658A CN 110964659 A CN110964659 A CN 110964659A
- Authority
- CN
- China
- Prior art keywords
- cells
- lactic acid
- lactobacillus plantarum
- acid bacteria
- solution
- 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
- 238000000034 method Methods 0.000 title claims abstract description 42
- 240000006024 Lactobacillus plantarum Species 0.000 title claims abstract description 36
- 235000013965 Lactobacillus plantarum Nutrition 0.000 title claims abstract description 36
- 229940072205 lactobacillus plantarum Drugs 0.000 title claims abstract description 36
- 238000012216 screening Methods 0.000 title claims abstract description 33
- 239000008280 blood Substances 0.000 title claims abstract description 27
- 210000004369 blood Anatomy 0.000 title claims abstract description 27
- 230000001603 reducing effect Effects 0.000 title claims abstract description 24
- 235000000346 sugar Nutrition 0.000 title claims abstract description 23
- 239000003814 drug Substances 0.000 title claims abstract description 10
- 235000013305 food Nutrition 0.000 title claims description 3
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 claims abstract description 114
- 241000894006 Bacteria Species 0.000 claims abstract description 70
- 239000004310 lactic acid Substances 0.000 claims abstract description 57
- 235000014655 lactic acid Nutrition 0.000 claims abstract description 57
- 230000001580 bacterial effect Effects 0.000 claims abstract description 21
- 230000000694 effects Effects 0.000 claims abstract description 15
- 238000002474 experimental method Methods 0.000 claims abstract description 12
- 239000006041 probiotic Substances 0.000 claims abstract description 10
- 235000018291 probiotics Nutrition 0.000 claims abstract description 10
- 238000001727 in vivo Methods 0.000 claims abstract description 9
- 238000004321 preservation Methods 0.000 claims abstract description 9
- 230000000529 probiotic effect Effects 0.000 claims abstract description 8
- 230000003178 anti-diabetic effect Effects 0.000 claims abstract description 7
- 238000012795 verification Methods 0.000 claims abstract description 6
- 230000003647 oxidation Effects 0.000 claims abstract description 3
- 238000007254 oxidation reaction Methods 0.000 claims abstract description 3
- 239000000243 solution Substances 0.000 claims description 32
- 239000000523 sample Substances 0.000 claims description 28
- 241000700159 Rattus Species 0.000 claims description 27
- 239000001963 growth medium Substances 0.000 claims description 21
- 241000186660 Lactobacillus Species 0.000 claims description 20
- 229940039696 lactobacillus Drugs 0.000 claims description 20
- 238000006243 chemical reaction Methods 0.000 claims description 17
- 239000006228 supernatant Substances 0.000 claims description 17
- 239000003833 bile salt Substances 0.000 claims description 16
- 239000012530 fluid Substances 0.000 claims description 16
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 16
- 238000012258 culturing Methods 0.000 claims description 15
- 230000002496 gastric effect Effects 0.000 claims description 14
- 239000000725 suspension Substances 0.000 claims description 13
- 238000000855 fermentation Methods 0.000 claims description 12
- 230000004151 fermentation Effects 0.000 claims description 12
- 208000001072 type 2 diabetes mellitus Diseases 0.000 claims description 12
- 238000002835 absorbance Methods 0.000 claims description 11
- 239000000284 extract Substances 0.000 claims description 11
- 230000031700 light absorption Effects 0.000 claims description 11
- 206010012601 diabetes mellitus Diseases 0.000 claims description 10
- 239000008103 glucose Substances 0.000 claims description 10
- 238000002156 mixing Methods 0.000 claims description 10
- 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 claims description 9
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 claims description 9
- 230000005764 inhibitory process Effects 0.000 claims description 9
- 230000004083 survival effect Effects 0.000 claims description 9
- 102100023374 Forkhead box protein M1 Human genes 0.000 claims description 8
- 101000907578 Homo sapiens Forkhead box protein M1 Proteins 0.000 claims description 8
- 102100024295 Maltase-glucoamylase Human genes 0.000 claims description 8
- 108010028144 alpha-Glucosidases Proteins 0.000 claims description 8
- 230000003859 lipid peroxidation Effects 0.000 claims description 8
- 239000007788 liquid Substances 0.000 claims description 8
- 235000004213 low-fat Nutrition 0.000 claims description 8
- 238000005259 measurement Methods 0.000 claims description 8
- 239000000203 mixture Substances 0.000 claims description 8
- 108090000790 Enzymes Proteins 0.000 claims description 7
- 102000004190 Enzymes Human genes 0.000 claims description 7
- 229940088598 enzyme Drugs 0.000 claims description 7
- 239000002609 medium Substances 0.000 claims description 7
- 239000007787 solid Substances 0.000 claims description 7
- 238000005406 washing Methods 0.000 claims description 7
- TUJKJAMUKRIRHC-UHFFFAOYSA-N hydroxyl Chemical compound [OH] TUJKJAMUKRIRHC-UHFFFAOYSA-N 0.000 claims description 6
- 230000003914 insulin secretion Effects 0.000 claims description 6
- 239000002504 physiological saline solution Substances 0.000 claims description 6
- 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 claims description 6
- 238000002965 ELISA Methods 0.000 claims description 5
- 239000003472 antidiabetic agent Substances 0.000 claims description 5
- 239000003963 antioxidant agent Substances 0.000 claims description 5
- 230000003078 antioxidant effect Effects 0.000 claims description 5
- 235000006708 antioxidants Nutrition 0.000 claims description 5
- 238000001514 detection method Methods 0.000 claims description 5
- 210000001035 gastrointestinal tract Anatomy 0.000 claims description 5
- 230000000968 intestinal effect Effects 0.000 claims description 5
- 210000004153 islets of langerhan Anatomy 0.000 claims description 5
- 238000009630 liquid culture Methods 0.000 claims description 5
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims description 4
- -1 DPPH free radical Chemical class 0.000 claims description 4
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 claims description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 4
- 241001465754 Metazoa Species 0.000 claims description 4
- 230000003115 biocidal effect Effects 0.000 claims description 4
- 239000007853 buffer solution Substances 0.000 claims description 4
- 238000003304 gavage Methods 0.000 claims description 4
- 230000015784 hyperosmotic salinity response Effects 0.000 claims description 4
- 239000000843 powder Substances 0.000 claims description 4
- 238000012360 testing method Methods 0.000 claims description 4
- YNJBWRMUSHSURL-UHFFFAOYSA-N trichloroacetic acid Chemical compound OC(=O)C(Cl)(Cl)Cl YNJBWRMUSHSURL-UHFFFAOYSA-N 0.000 claims description 4
- 108010019160 Pancreatin Proteins 0.000 claims description 3
- 238000004113 cell culture Methods 0.000 claims description 3
- 238000010276 construction Methods 0.000 claims description 3
- 235000009200 high fat diet Nutrition 0.000 claims description 3
- 230000002401 inhibitory effect Effects 0.000 claims description 3
- 210000000936 intestine Anatomy 0.000 claims description 3
- 229940055695 pancreatin Drugs 0.000 claims description 3
- 238000005502 peroxidation Methods 0.000 claims description 3
- RVBUGGBMJDPOST-UHFFFAOYSA-N 2-thiobarbituric acid Chemical compound O=C1CC(=O)NC(=S)N1 RVBUGGBMJDPOST-UHFFFAOYSA-N 0.000 claims description 2
- BTJIUGUIPKRLHP-UHFFFAOYSA-N 4-nitrophenol Chemical compound OC1=CC=C([N+]([O-])=O)C=C1 BTJIUGUIPKRLHP-UHFFFAOYSA-N 0.000 claims description 2
- IFBHRQDFSNCLOZ-IIRVCBMXSA-N 4-nitrophenyl-α-d-galactoside Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1OC1=CC=C([N+]([O-])=O)C=C1 IFBHRQDFSNCLOZ-IIRVCBMXSA-N 0.000 claims description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 2
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 claims description 2
- 229930182816 L-glutamine Natural products 0.000 claims description 2
- OYHQOLUKZRVURQ-HZJYTTRNSA-N Linoleic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O OYHQOLUKZRVURQ-HZJYTTRNSA-N 0.000 claims description 2
- 241000700157 Rattus norvegicus Species 0.000 claims 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 claims description 2
- 230000009471 action Effects 0.000 claims description 2
- 230000003698 anagen phase Effects 0.000 claims description 2
- 235000010323 ascorbic acid Nutrition 0.000 claims description 2
- 229960005070 ascorbic acid Drugs 0.000 claims description 2
- 239000011668 ascorbic acid Substances 0.000 claims description 2
- 238000004364 calculation method Methods 0.000 claims description 2
- 239000013553 cell monolayer Substances 0.000 claims description 2
- 230000004663 cell proliferation Effects 0.000 claims description 2
- 238000005119 centrifugation Methods 0.000 claims description 2
- 229960001701 chloroform Drugs 0.000 claims description 2
- 238000001816 cooling Methods 0.000 claims description 2
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 claims description 2
- 235000018417 cysteine Nutrition 0.000 claims description 2
- 239000012153 distilled water Substances 0.000 claims description 2
- 239000000839 emulsion Substances 0.000 claims description 2
- 230000003203 everyday effect Effects 0.000 claims description 2
- 238000000605 extraction Methods 0.000 claims description 2
- 239000011790 ferrous sulphate Substances 0.000 claims description 2
- 235000003891 ferrous sulphate Nutrition 0.000 claims description 2
- 239000012091 fetal bovine serum Substances 0.000 claims description 2
- 230000006870 function Effects 0.000 claims description 2
- 230000012010 growth Effects 0.000 claims description 2
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 claims description 2
- 229910000359 iron(II) sulfate Inorganic materials 0.000 claims description 2
- OYHQOLUKZRVURQ-IXWMQOLASA-N linoleic acid Natural products CCCCC\C=C/C\C=C\CCCCCCCC(O)=O OYHQOLUKZRVURQ-IXWMQOLASA-N 0.000 claims description 2
- 235000020778 linoleic acid Nutrition 0.000 claims description 2
- 238000000465 moulding Methods 0.000 claims description 2
- 239000002244 precipitate Substances 0.000 claims description 2
- 239000012488 sample solution Substances 0.000 claims description 2
- CDBYLPFSWZWCQE-UHFFFAOYSA-L sodium carbonate Substances [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 claims description 2
- 229910000029 sodium carbonate Inorganic materials 0.000 claims description 2
- GNBVPFITFYNRCN-UHFFFAOYSA-M sodium thioglycolate Chemical compound [Na+].[O-]C(=O)CS GNBVPFITFYNRCN-UHFFFAOYSA-M 0.000 claims description 2
- 229940046307 sodium thioglycolate Drugs 0.000 claims description 2
- 239000008223 sterile water Substances 0.000 claims description 2
- 230000004936 stimulating effect Effects 0.000 claims description 2
- 229960001052 streptozocin Drugs 0.000 claims description 2
- 239000000126 substance Substances 0.000 claims description 2
- 230000035899 viability Effects 0.000 claims description 2
- 230000000638 stimulation Effects 0.000 claims 2
- XPFJYKARVSSRHE-UHFFFAOYSA-K trisodium;2-hydroxypropane-1,2,3-tricarboxylate;2-hydroxypropane-1,2,3-tricarboxylic acid Chemical compound [Na+].[Na+].[Na+].OC(=O)CC(O)(C(O)=O)CC(O)=O.[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O XPFJYKARVSSRHE-UHFFFAOYSA-K 0.000 claims 2
- 101100191768 Caenorhabditis elegans pbs-4 gene Proteins 0.000 claims 1
- 241001052560 Thallis Species 0.000 claims 1
- 238000003287 bathing Methods 0.000 claims 1
- 230000007910 cell fusion Effects 0.000 claims 1
- 239000012295 chemical reaction liquid Substances 0.000 claims 1
- 238000007865 diluting Methods 0.000 claims 1
- 235000020776 essential amino acid Nutrition 0.000 claims 1
- 239000003797 essential amino acid Substances 0.000 claims 1
- 230000002779 inactivation Effects 0.000 claims 1
- 238000002372 labelling Methods 0.000 claims 1
- 239000011259 mixed solution Substances 0.000 claims 1
- 239000008055 phosphate buffer solution Substances 0.000 claims 1
- 238000000338 in vitro Methods 0.000 abstract description 7
- 239000002953 phosphate buffered saline Substances 0.000 description 11
- 230000002218 hypoglycaemic effect Effects 0.000 description 9
- 238000011160 research Methods 0.000 description 7
- 206010022489 Insulin Resistance Diseases 0.000 description 6
- 238000000513 principal component analysis Methods 0.000 description 5
- ULGZDMOVFRHVEP-RWJQBGPGSA-N Erythromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=O)[C@H](C)C[C@@](C)(O)[C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 ULGZDMOVFRHVEP-RWJQBGPGSA-N 0.000 description 4
- 239000003223 protective agent Substances 0.000 description 4
- 230000002000 scavenging effect Effects 0.000 description 4
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 230000002292 Radical scavenging effect Effects 0.000 description 3
- 239000003242 anti bacterial agent Substances 0.000 description 3
- 229940088710 antibiotic agent Drugs 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 description 2
- WQZGKKKJIJFFOK-SVZMEOIVSA-N (+)-Galactose Chemical compound OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@H]1O WQZGKKKJIJFFOK-SVZMEOIVSA-N 0.000 description 2
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 2
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 2
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 2
- 229930195725 Mannitol Natural products 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- DRTQHJPVMGBUCF-XVFCMESISA-N Uridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-XVFCMESISA-N 0.000 description 2
- 108010059993 Vancomycin Proteins 0.000 description 2
- PYMYPHUHKUWMLA-LMVFSUKVSA-N aldehydo-D-ribose Chemical compound OC[C@@H](O)[C@@H](O)[C@@H](O)C=O PYMYPHUHKUWMLA-LMVFSUKVSA-N 0.000 description 2
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 description 2
- 238000010171 animal model Methods 0.000 description 2
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 229960005091 chloramphenicol Drugs 0.000 description 2
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 2
- 229940099352 cholate Drugs 0.000 description 2
- BHQCQFFYRZLCQQ-OELDTZBJSA-N cholic acid Chemical compound C([C@H]1C[C@H]2O)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC(O)=O)C)[C@@]2(C)[C@@H](O)C1 BHQCQFFYRZLCQQ-OELDTZBJSA-N 0.000 description 2
- 229960002227 clindamycin Drugs 0.000 description 2
- KDLRVYVGXIQJDK-AWPVFWJPSA-N clindamycin Chemical compound CN1C[C@H](CCC)C[C@H]1C(=O)N[C@H]([C@H](C)Cl)[C@@H]1[C@H](O)[C@H](O)[C@@H](O)[C@@H](SC)O1 KDLRVYVGXIQJDK-AWPVFWJPSA-N 0.000 description 2
- 229960003276 erythromycin Drugs 0.000 description 2
- 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 2
- 239000000594 mannitol Substances 0.000 description 2
- 235000010355 mannitol Nutrition 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 230000008569 process Effects 0.000 description 2
- 150000003254 radicals Chemical class 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- JQXXHWHPUNPDRT-WLSIYKJHSA-N rifampicin Chemical compound O([C@](C1=O)(C)O/C=C/[C@@H]([C@H]([C@@H](OC(C)=O)[C@H](C)[C@H](O)[C@H](C)[C@@H](O)[C@@H](C)\C=C\C=C(C)/C(=O)NC=2C(O)=C3C([O-])=C4C)C)OC)C4=C1C3=C(O)C=2\C=N\N1CC[NH+](C)CC1 JQXXHWHPUNPDRT-WLSIYKJHSA-N 0.000 description 2
- 229960001225 rifampicin Drugs 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- OESFSXYRSCBAQJ-UHFFFAOYSA-M sodium;3-carboxy-3,5-dihydroxy-5-oxopentanoate;2-hydroxypropane-1,2,3-tricarboxylic acid Chemical compound [Na+].OC(=O)CC(O)(C(O)=O)CC(O)=O.OC(=O)CC(O)(C(O)=O)CC([O-])=O OESFSXYRSCBAQJ-UHFFFAOYSA-M 0.000 description 2
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Chemical compound CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 2
- 229960003165 vancomycin Drugs 0.000 description 2
- MYPYJXKWCTUITO-LYRMYLQWSA-N vancomycin Chemical compound O([C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=C2C=C3C=C1OC1=CC=C(C=C1Cl)[C@@H](O)[C@H](C(N[C@@H](CC(N)=O)C(=O)N[C@H]3C(=O)N[C@H]1C(=O)N[C@H](C(N[C@@H](C3=CC(O)=CC(O)=C3C=3C(O)=CC=C1C=3)C(O)=O)=O)[C@H](O)C1=CC=C(C(=C1)Cl)O2)=O)NC(=O)[C@@H](CC(C)C)NC)[C@H]1C[C@](C)(N)[C@H](O)[C@H](C)O1 MYPYJXKWCTUITO-LYRMYLQWSA-N 0.000 description 2
- MYPYJXKWCTUITO-UHFFFAOYSA-N vancomycin Natural products O1C(C(=C2)Cl)=CC=C2C(O)C(C(NC(C2=CC(O)=CC(O)=C2C=2C(O)=CC=C3C=2)C(O)=O)=O)NC(=O)C3NC(=O)C2NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(CC(C)C)NC)C(O)C(C=C3Cl)=CC=C3OC3=CC2=CC1=C3OC1OC(CO)C(O)C(O)C1OC1CC(C)(N)C(O)C(C)O1 MYPYJXKWCTUITO-UHFFFAOYSA-N 0.000 description 2
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 description 1
- 206010067484 Adverse reaction Diseases 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- GUBGYTABKSRVRQ-UHFFFAOYSA-N D-Cellobiose Natural products OCC1OC(OC2C(O)C(O)C(O)OC2CO)C(O)C(O)C1O GUBGYTABKSRVRQ-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-CUHNMECISA-N D-Cellobiose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-CUHNMECISA-N 0.000 description 1
- RFSUNEUAIZKAJO-VRPWFDPXSA-N D-Fructose Natural products OC[C@H]1OC(O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-VRPWFDPXSA-N 0.000 description 1
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 1
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 1
- WQZGKKKJIJFFOK-QTVWNMPRSA-N D-mannopyranose Chemical compound OC[C@H]1OC(O)[C@@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-QTVWNMPRSA-N 0.000 description 1
- SRBFZHDQGSBBOR-IOVATXLUSA-N D-xylopyranose Chemical compound O[C@@H]1COC(O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-IOVATXLUSA-N 0.000 description 1
- 206010012735 Diarrhoea Diseases 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 241000192125 Firmicutes Species 0.000 description 1
- RFSUNEUAIZKAJO-ARQDHWQXSA-N Fructose Chemical compound OC[C@H]1O[C@](O)(CO)[C@@H](O)[C@@H]1O RFSUNEUAIZKAJO-ARQDHWQXSA-N 0.000 description 1
- 229930182566 Gentamicin Natural products 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 1
- 102000006587 Glutathione peroxidase Human genes 0.000 description 1
- 108700016172 Glutathione peroxidases Proteins 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 description 1
- 229920000057 Mannan Polymers 0.000 description 1
- OVRNDRQMDRJTHS-CBQIKETKSA-N N-Acetyl-D-Galactosamine Chemical compound CC(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@H](O)[C@@H]1O OVRNDRQMDRJTHS-CBQIKETKSA-N 0.000 description 1
- MBLBDJOUHNCFQT-UHFFFAOYSA-N N-acetyl-D-galactosamine Natural products CC(=O)NC(C=O)C(O)C(O)C(O)CO MBLBDJOUHNCFQT-UHFFFAOYSA-N 0.000 description 1
- AYRXSINWFIIFAE-UHFFFAOYSA-N O6-alpha-D-Galactopyranosyl-D-galactose Natural products OCC1OC(OCC(O)C(O)C(O)C(O)C=O)C(O)C(O)C1O AYRXSINWFIIFAE-UHFFFAOYSA-N 0.000 description 1
- 230000010718 Oxidation Activity Effects 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- NGFMICBWJRZIBI-JZRPKSSGSA-N Salicin Natural products O([C@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@H](CO)O1)c1c(CO)cccc1 NGFMICBWJRZIBI-JZRPKSSGSA-N 0.000 description 1
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 102000019197 Superoxide Dismutase Human genes 0.000 description 1
- 108010012715 Superoxide dismutase Proteins 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000003044 adaptive effect Effects 0.000 description 1
- 230000006838 adverse reaction Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- WQZGKKKJIJFFOK-DVKNGEFBSA-N alpha-D-glucose Chemical compound OC[C@H]1O[C@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-DVKNGEFBSA-N 0.000 description 1
- NGFMICBWJRZIBI-UHFFFAOYSA-N alpha-salicin Natural products OC1C(O)C(O)C(CO)OC1OC1=CC=CC=C1CO NGFMICBWJRZIBI-UHFFFAOYSA-N 0.000 description 1
- 229940024606 amino acid Drugs 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000003064 anti-oxidating effect Effects 0.000 description 1
- 238000009635 antibiotic susceptibility testing Methods 0.000 description 1
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 235000015278 beef Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- DRTQHJPVMGBUCF-PSQAKQOGSA-N beta-L-uridine Natural products O[C@H]1[C@@H](O)[C@H](CO)O[C@@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-PSQAKQOGSA-N 0.000 description 1
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 description 1
- 229940093761 bile salts Drugs 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 238000002815 broth microdilution Methods 0.000 description 1
- 229910000019 calcium carbonate Inorganic materials 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 239000006143 cell culture medium Substances 0.000 description 1
- 229960002626 clarithromycin Drugs 0.000 description 1
- AGOYDEPGAOXOCK-KCBOHYOISA-N clarithromycin Chemical compound O([C@@H]1[C@@H](C)C(=O)O[C@@H]([C@@]([C@H](O)[C@@H](C)C(=O)[C@H](C)C[C@](C)([C@H](O[C@H]2[C@@H]([C@H](C[C@@H](C)O2)N(C)C)O)[C@H]1C)OC)(C)O)CC)[C@H]1C[C@@](C)(OC)[C@@H](O)[C@H](C)O1 AGOYDEPGAOXOCK-KCBOHYOISA-N 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- YXVFQADLFFNVDS-UHFFFAOYSA-N diammonium citrate Chemical compound [NH4+].[NH4+].[O-]C(=O)CC(O)(C(=O)O)CC([O-])=O YXVFQADLFFNVDS-UHFFFAOYSA-N 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 238000003028 enzyme activity measurement method Methods 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 235000021107 fermented food Nutrition 0.000 description 1
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 210000004051 gastric juice Anatomy 0.000 description 1
- 229960002518 gentamicin Drugs 0.000 description 1
- DLRVVLDZNNYCBX-CQUJWQHSSA-N gentiobiose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC[C@@H]1[C@@H](O)[C@H](O)[C@@H](O)C(O)O1 DLRVVLDZNNYCBX-CQUJWQHSSA-N 0.000 description 1
- 229960002989 glutamic acid Drugs 0.000 description 1
- 230000002641 glycemic effect Effects 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 229940126904 hypoglycaemic agent Drugs 0.000 description 1
- 230000000415 inactivating effect Effects 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 description 1
- 229940061634 magnesium sulfate heptahydrate Drugs 0.000 description 1
- SCVOEYLBXCPATR-UHFFFAOYSA-L manganese(II) sulfate pentahydrate Chemical compound O.O.O.O.O.[Mn+2].[O-]S([O-])(=O)=O SCVOEYLBXCPATR-UHFFFAOYSA-L 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- CWKLZLBVOJRSOM-UHFFFAOYSA-N methyl pyruvate Chemical compound COC(=O)C(C)=O CWKLZLBVOJRSOM-UHFFFAOYSA-N 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 229940049954 penicillin Drugs 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 235000021110 pickles Nutrition 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 230000029537 positive regulation of insulin secretion Effects 0.000 description 1
- GNSKLFRGEWLPPA-UHFFFAOYSA-M potassium dihydrogen phosphate Chemical compound [K+].OP(O)([O-])=O GNSKLFRGEWLPPA-UHFFFAOYSA-M 0.000 description 1
- LWIHDJKSTIGBAC-UHFFFAOYSA-K potassium phosphate Substances [K+].[K+].[K+].[O-]P([O-])([O-])=O LWIHDJKSTIGBAC-UHFFFAOYSA-K 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 238000012847 principal component analysis method Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- NGFMICBWJRZIBI-UJPOAAIJSA-N salicin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@H]1OC1=CC=CC=C1CO NGFMICBWJRZIBI-UJPOAAIJSA-N 0.000 description 1
- 229940120668 salicin Drugs 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 239000001632 sodium acetate Substances 0.000 description 1
- 235000017281 sodium acetate Nutrition 0.000 description 1
- 239000000600 sorbitol Substances 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 230000003068 static effect Effects 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 150000008163 sugars Chemical class 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- 229960002180 tetracycline Drugs 0.000 description 1
- 229930101283 tetracycline Natural products 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- DRTQHJPVMGBUCF-UHFFFAOYSA-N uracil arabinoside Natural products OC1C(O)C(CO)OC1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-UHFFFAOYSA-N 0.000 description 1
- 229940045145 uridine Drugs 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 239000012138 yeast extract Substances 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
- A61K35/741—Probiotics
- A61K35/744—Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
- A61K35/747—Lactobacilli, e.g. L. acidophilus or L. brevis
-
- 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
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P39/00—General protective or antinoxious agents
- A61P39/06—Free radical scavengers or antioxidants
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/11—Lactobacillus
- A23V2400/169—Plantarum
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/225—Lactobacillus
- C12R2001/25—Lactobacillus plantarum
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Microbiology (AREA)
- Wood Science & Technology (AREA)
- Mycology (AREA)
- Pharmacology & Pharmacy (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Biochemistry (AREA)
- Veterinary Medicine (AREA)
- Diabetes (AREA)
- Virology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Tropical Medicine & Parasitology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Biomedical Technology (AREA)
- General Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Epidemiology (AREA)
- Emergency Medicine (AREA)
- Endocrinology (AREA)
- Hematology (AREA)
- Obesity (AREA)
- Molecular Biology (AREA)
- Polymers & Plastics (AREA)
- Nutrition Science (AREA)
- Toxicology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
本发明提供一种植物乳杆菌,该植物乳杆菌的保藏编号为CCTCC M 2019591,拉丁文名称为Lactobacillus plantarum CT152;提供一种具有降血糖功能的药剂,该药剂采用上述的植物乳杆菌。并提供一种植物乳杆菌的筛选方法,包括如下步骤:S1:进行益生指标测定;S2:进行抗氧化能力测定;S3:进行乳酸菌的抗糖尿病特性的检测;S4:进行体内实验的验证。本发明***地建立起降糖乳酸菌的体外高效筛选方法,通过PCA和AHP结合的方式筛选得到一株有较好降糖效果的发明菌株CCTCC M 2019591,并得到了体内实验的效果验证。
Description
技术领域
本发明具体涉及一种降血糖功效的植物乳杆菌及其筛选方法和应用药剂。
背景技术
国际糖尿病联盟公布了第八版的全球糖尿病地图。结果显示,全球糖尿病成人患者(20-79岁)从2000年的1.51亿,到2017年已达到4.25亿,增加近2 倍。预计到2045年,糖尿病患者可能达到6.29亿。而中国的糖尿病患者位居世界第一,糖尿病被公认为世界第三大致死性疾病,其发病率呈逐年增加趋势,且Ⅱ型糖尿病患者的比例高达90%以上。目前主要通过药物降低血糖来达到治疗Ⅱ型糖尿病的目的。针对糖尿病的降糖药物通常会引起一系列的不良反应,如腹泻、耐药性和继发性失效等问题。近年来由于乳酸菌的天然性和安全性,逐渐成为糖尿病防治的研究热点。
乳酸菌(Lactic acid bacteria,LAB)指发酵糖类,主要产物为乳酸的一类无芽孢、革兰氏染色阳性细菌的总称。近几年研究发现,乳酸菌作为定居肠道中的有益菌群具有多种生理功能;维持肠道微生态平衡,影响血糖代谢等等。目前针对降糖乳酸菌的全面***的体外筛选方法仍未建立,体内的动物实验筛选受时间、价格方面的因素受到了一定的限制,因此亟待对于目前的体外降糖乳酸菌的筛选方法进行评估和整合,建立一套全面地、***地降糖乳酸菌的筛选方法,为乳酸菌降糖的机制研究提供一定的技术和方法支撑。之前陈佩等研究者运用主成分分析法从体外筛选得到2株降糖乳酸菌,并且在体内进行了验证其降糖效果,但是其筛选指标不够全面,如并未对乳酸菌的安全性进行评估。本发明根据前期研究者的体外功能菌的筛选指标进行整合和补充,运用主成分分析法(PCA)和层次分析法(AHP)相结合,建立高效降糖乳酸菌的筛选方法,并且在体内得到了证实,这为乳酸菌降糖的机制研究提供一定的技术和方法支撑。
发明内容
本发明解决的技术问题之一是体外降糖乳酸菌的筛选方法的建立;本发明要解决的技术问题之二是提供应用上述筛选方法得到的一株经过体内验证得降糖菌株Lactobacillus plantarum CT152,它在体外模拟的胃肠液和胆盐环境下具有较高的存活率;且具有一定的抗氧化和抗糖尿病的特性;本发明要解决的技术问题之二是提供一种应用到该降糖菌株Lactobacillus plantarum CT152的降血糖药剂。
为达到上述要求,本发明采取的技术方案是:提供一种植物乳杆菌,该植物乳杆菌的保藏编号为CCTCC NO: M 2019591,拉丁文名称为Lactobacillus plantarum CT152,保藏地为中国典型培养物保藏中心,保藏地地址为湖北省武汉市武昌区八一路229号武汉大学校内,保藏日期为2019年7月30日;提供一种具有降血糖功能的药剂,采用上述的植物乳杆菌;提供一种植物乳杆菌的筛选方法,包括如下步骤:
S1:进行益生指标测定,包括模拟的胃肠液中的存活率测定、胆盐耐受力测定、对Caco-2细胞的粘附能力测定及抗生素耐药性检测;
S2:进行抗氧化能力测定,包括乳酸菌清除DPPH能力的测定、乳酸菌清除羟自由基能力测定、乳酸菌抗脂质过氧化能力测定、乳酸菌的还原能力测定及乳酸菌的GSH-Px和T-SOD的酶活测定;
S3:进行乳酸菌的抗糖尿病特性的检测,包括大鼠肠道α-glucosidase活力抑制率的测定和乳酸菌干预高糖刺激胰岛细胞分泌胰岛素实验;以及
S4:进行体内实验的验证,包括2型糖尿病模型大鼠的构建及进行灌胃治疗。
本发明提供的体外降糖乳酸菌的高效筛选方法,可以在一定程度上为后续的功能性乳酸菌的筛选提供技术和方法支撑,通过此方法,筛选得到一株潜在的降糖菌株CCTCC M2019591。在益生方面,发明菌株的胃肠液耐受能力很好,存活率达到了97.62%,且胆盐的耐受和对于Caco-2细胞的粘附能力也较阳性对照菌株强,粘附率达到18.67%;菌株的抗氧化能力也很高,无细胞提取物和完整细胞在DPPH的清除能力上达到了15%-16%左右。清除羟自由基能力方面,发明菌株与商业对照菌株也能力相当,对于抗脂质过氧化能力,发明菌株发酵上清和完整细胞都更强,分别达到8.48%和15.64%,还有GSH-Px和T-SOD酶活力方面,菌株CCTCC M 2019591也较强。此外,关于乳酸菌对于大鼠肠道提取的α-glucosidase活力的抑制率方面,发明菌株达到15%左右的抑制率,且在用该菌株干预条件下,可以刺激其胰岛细胞INS-1在高糖条件下提高产胰岛素的能力,达到空白的1.41倍,从而发挥其降血糖功能。并且本方法在筛选指标中,对菌株的抗生素耐药性进行了检测,保证菌株没有可转移的耐药基因,是一株潜在的安全性功能性菌株。在通过PCA和AHP相结合的方面来综合评价其降糖能力的时候,发明菌株得分较商业对照菌株LGG的得分更高,说明发明菌株的综合降糖能力明显高于商业菌株,并且在动物实验中也得到了相应的效果验证。
附图说明
此处所说明的附图用来提供对本申请的进一步理解,构成本申请的一部分,在这些附图中使用相同的参考标号来表示相同或相似的部分,本申请的示意性实施例及其说明用于解释本申请,并不构成对本申请的不当限定。在附图中:
图1示意性地示出了根据本申请一个实施例的植物乳杆菌对INS-1细胞胰岛素分泌量的影响的示意图。
图2示意性地示出了根据本申请一个实施例的植物乳杆菌对Caco-2细胞的粘附率示意图。
图3示意性地示出了根据本申请一个实施例灌胃四周大鼠随机血糖变化值的示意图。
图4示意性地示出了根据本申请一个实施例中灌胃六周大鼠随机血糖变化值。
具体实施方式
为使本申请的目的、技术方案和优点更加清楚,以下结合附图及具体实施例,对本申请作进一步地详细说明。
在以下描述中,对“一个实施例”、“实施例”、“一个示例”、“示例”等等的引用表明如此描述的实施例或示例可以包括特定特征、结构、特性、性质、元素或限度,但并非每个实施例或示例都必然包括特定特征、结构、特性、性质、元素或限度。另外,重复使用短语“根据本申请的一个实施例”虽然有可能是指代相同实施例,但并非必然指代相同的实施例。
为简单起见,以下描述中省略了本领域技术人员公知的某些技术特征。
根据本申请的一个实施例,提供一种植物乳杆菌,该植物乳杆菌的保藏编号为CCTCC M 2019591,拉丁文名称为Lactobacillus plantarum CT152;并提供一种一种具有降血糖功能的药剂,该药剂采用权利要求1所述的植物乳杆菌。
根据本申请的一个实施例,提供一种植物乳杆菌的筛选方法,该植物乳杆菌的筛选方法具体包括如下步骤:
(1)益生指标测定:
1)模拟的胃肠液中的存活率:将乳酸菌用0.85%生理盐水重悬后,在模拟胃液(pH3.0)中将其菌液密度调节为1×109CFU/mL。混匀后将其放置于37℃培养1,2和3h后分别计活菌数。在模拟胃液中培养3h后,取1mL培养液加入至9mL模拟肠液(pH 8.0)中,混匀后于37℃培养。分别于2、4和 8h检测活菌数。用MRS固体培养基37℃培养48h,计算其存活率。
存活率(%)=[log CFU N1/log CFU N]×100%;
N1=经过模拟胃肠液处理之后的乳酸菌活菌数,N0=未处理前乳酸菌的活菌数。
2)胆盐耐受力:将培养18h的乳酸菌按2%-5%分别接种于含有或不含有 0.3%胆盐的MRS-THIO(MRS含有0.2%的巯基乙酸钠)培养基中。混合物于 37℃培养16-24h,每隔一段时间在600nm;处测吸光值,记录不加胆盐和加了胆盐的培养基吸光值达到0.3个单位时各自所需的时间,这两个时间差即为乳酸菌在胆盐中生长的延迟时间(LT),LT越短说明乳酸菌,耐受胆盐的能力越强。
3)对Caco-2细胞的粘附能力:Caco-2细胞生长在含有20%胎牛血清,1%L- 谷氨酰胺,1%非必需氨基酸和1%双抗的高糖DMEM培养液中,37℃,5%CO2 的培养箱中进行培养,隔天更换一次培养液,等细胞增殖融合率达80%左右时,以含0.02%EDTA的0.25%胰酶消化细胞,按1:2-1:3传代。取对数生长期细胞进行试验。
将新鲜培养的乳酸菌用PBS洗两次,调节细菌浓度为1×109CFU/mL,悬浮于100mg/mL异硫氰酸荧光素(FIFC)中,37℃暗处作用0.5-1h,用PBS洗涤 4次去除未结合的FIFC。将Caco-2细胞以4×105-5×105cell/孔接种于6孔板, 37℃培养16-24h。将FIFC标记的乳酸菌重悬于DMEM培养基中,调整其密度为1×109CFU/mL。将此菌悬液加入到Caco-2细胞单层中,37℃作用1-2h,用无菌PBS洗涤3次,将未粘附的菌洗脱掉,之后每个培养的孔中加入胰酶作用,待细胞完全脱落,加入0.4mL完全培养基终止反应。收集液体,用多功能酶标仪测定其荧光强度来计算其细菌的黏附率。A和A0分别表示乳酸菌粘附细胞前后的荧光强度。
粘附率(%)=Log A/Log A0×100%。
4)抗生素耐药性检测:
依据欧洲微生物药物敏感委员会(http://www.eucast.org)关于微生物对抗生素的敏感阈值X(http://www.eucast.org/mic_distributions/),分析菌株的抗生素耐药性。当分离菌株的最小抑菌质量浓度(Minimal Inhibitory Concentration,MIC)≤ Xμg/mL时,为敏感菌株;反之,则为耐药菌株。
MIC测定采用微量肉汤稀释法。分别将常用的抗生素配制成相应的贮存液, MRS液体培养基2倍梯度稀释成使用液。向无菌的96孔板中加入198μL含不同浓度抗生素的MRS液体培养基后,接种2μL菌株的培养液(约1.0× 107CFU/mL),37℃静止培养24h后,统计不同菌株的MIC。每组重复3次,并设置对照组。
(2)抗氧化能力测定:
1)乳酸菌清除DPPH能力的测定:反应体系中加入1mL乳酸菌的菌悬液,无细胞提取物或者发酵上清液,再加入1mL 0.2mmol/L DPPH的无水乙醇,震荡充分混匀,在室温下避光反应30min,再于6000rpm离心10min,取其上清,517nm波长处测定样品吸光度(OD值)。用等体积的生理盐水代替样品溶液作为对照组,并以等体积的生理盐水和无水乙醇的混合液作为空白调零。按照以下公式计算:
DPPH自由基的清除率=[1-A517(样品)/A517(对照)]×100%
2)乳酸菌清除羟自由基能力测定:根据南京建成生物公司的羟自由基试剂盒测定,不做按照其说明书严格执行。
3)乳酸菌抗脂质过氧化能力测定:0.5mL pH 7.4的PBS中加入1mL亚油酸乳化液,加入0.2mL 0.01%的硫酸亚铁和0.01%的抗坏血酸,再加入 0.2m-0.5mL发酵上清,菌悬液或者细胞提取物,混匀后放置于37℃水浴中反应12h,取2mL反应液加入0.2mL 0.4%的三氯乙酸,2mL 0.8%的硫代巴比妥酸和0.2mL 0.4%BHT,震荡充分混匀后,在100℃下反应30min,直至样品冷却后再加入2mL三氯甲烷进行抽提,离心10min后,收集上清。532nm 波长下测定样品OD值,以PBS作为空白对照。按照下式计算乳酸菌抗脂质过氧化的能力:抗脂质过氧化率=[1-A532(样品)/A532(空白)]×100%。
4)乳酸菌的还原能力测定:0.5mL菌悬液或细胞提取物中加入1%铁***0.5mL,0.5mL PBS(pH 6.6),震荡充分混匀,于50℃中水浴20min,样品替换为蒸馏水后可作为对照组。在冰浴中急速冷却后,加入0.5mL 10%的三氯乙酸,3000rpm离心10min,取其1mL上清后加入1mL 0.1%的Fe CL3,反应10min后在700nm波长下测定样品的OD值。结果采用半胱氨酸作为标准表征还原力,计算公式如下:还原能力=[A700(样品)-A700(对照)/A700(对照)]× 100%。
5)乳酸菌的GSH-Px和T-SOD的酶活测定:
用南京建成的试剂盒检测乳酸菌发酵上清和细胞提取物中GSH-Px和 T-SOD的酶活,具体操作严格按照试剂盒说明书。
(3)乳酸菌的抗糖尿病特性
1)大鼠肠道α-glucosidase活力抑制率的测定
在150μL PBS(0.1M pH 6.8)中加入75μL 20mM的PNPG溶液及25μL,待测乳酸菌的发酵上清,无细胞提取物和完整细胞,将混合物于37℃水浴10 min。加入50μL大鼠肠道提取的α-glucosidase溶液(0.17U/mL)继续反应10 min。加入1mL 0.1M Na2CO3作为反应终止液。并将反应液于405nm处测其吸光值,吸光值与对硝基酚(PNP
)的游离量成正比。反应体系中采用pH 6.8的0.1M PBS作为α-glucosidase溶液及待测样品的空白对照,采用以下公式计算样品的抑制活性。
酶抑制率(%)=[1-(C-D)/(A-B)]×100%
其中:A为含有α-glucosidase溶液但不含样品的测定吸光
B为不含α-glucosidase溶液及待测样品的测定吸光值
C为含有α-glucosidase溶液及待测样品的测定吸光值
D为不含α-glucosidase溶液但含待测样品的测定吸光值
2)乳酸菌干预高糖刺激胰岛细胞分泌胰岛素实验
乳酸菌在MRS液体培养基中厌氧传代2次,每次37℃下孵育18h,使菌株活力达到最高值。对菌悬液以灭菌的PBS缓冲液进行梯度稀释,采用平板倾注法,在MRS固体培养基上37℃培养24-48h,测定活菌数(保证每个平板 30-300的活菌菌落)。根据菌粉活菌数,配制1×109CFU/mL的菌悬液,经过 120kw超声破碎,然后65℃、30-60min灭活处理,8000×g离心获得菌体混合物沉淀。用不含双抗的INS-1细胞培养液进行重悬,作为GSIS实验刺激细胞的培养液,储存于4℃待用.
INS-1细胞以4×105-5×105cells/mL的密度接种于6孔板,孵育过夜待细胞贴壁,然后换用含有灭活菌液的细胞培养液孵育16-24h,对细胞进行不同益生菌株的干预作用。各组细胞处理结束后,细胞用磷酸盐缓冲液(PBS)洗2 次,加入葡萄糖浓度16.7mmol/L的高糖INS-1完全培养液,37℃下孵育0.5-1 h。然后收集细胞上清液,用ELISA法检测各组在高葡萄糖刺激下的胰岛素的分泌量。ELISA检测步骤按照试剂盒说明书进行,吸光度采用酶标仪(450nm 处)进行检测。以OD值为纵坐标(Y),标准物浓度为横坐标(X),绘制标准曲线,并根据此计算其他样品的浓度。
(4)体内实验的验证
1)2型糖尿病模型大鼠的构建
所有实验动物的喂养程序都经过四川大学华西动物房的许可。实验动物房常年保持适宜的温度和湿度,并严格遵循12h白天和12h黑夜的循环标准。70 只雄性6-8周龄的Wistar大鼠,用基础饲料适应性喂养5天后随机分为低脂组和高脂组。前四周,除低脂组喂食基础饲料外,其它各组均喂食高脂饲料,并于每周固定时间进行大鼠的体重测量记录。
第五周,所有大鼠禁食不禁水12-18h,除低脂组外,其余各组大鼠均按照 30-40mg/kg体重注射新鲜配制的链脲霉素STZ。将一定量STZ溶于柠檬酸- 柠檬酸钠缓冲液中(pH 4.5),现配现用,冰浴保存,低脂组注射柠檬酸-柠檬酸钠缓冲液中(pH 4.5)。造模后72h期间保证充足的无菌水及高脂饮食,并每日更换垫料,72h后随机血糖高于16.7mmol/L或者空腹血糖高于11.7mmol/L 的为造模成功的糖尿病大鼠。
2)灌胃治疗
将成模大鼠随机分为模型组、LGG组、CT152组,模型组用0.85%的生理盐水作为对照,LGG和CCTCC M 2019591以1×109CFU/day进行为期6-8周的灌胃治疗。
体内实验的验证中灌胃6周大鼠随机血糖变化值如图4所示。
乳酸菌菌剂的制备:1)将CCTCC M 2019591菌株接种于固体斜面培养基上,在37℃培养16-24h进行活化,然后取培养好的固体斜面培养基,在无菌条件下接种于种子液体培养基中,在37℃培养16-24h,制得一级种子液;按照2-5%的接种量,将一级种子液接种于种子液体培养基中,在37℃条件下,静置培养16-24h,然后收集发酵液。2)发酵结束后,立即将发酵液离心并用清水清洗,如此反复2-3遍后,按照菌:保护剂为1:1-1:2(保护剂可为常用的海藻糖、脱脂乳粉、山梨醇、甘露醇等或其为原料的复配保护剂)比例混合后进行冷冻干燥,粉碎,即成为乳酸菌菌剂。
本发明提供一种广泛***地降糖乳酸菌的筛选方法,且用此方法筛选得到一株具有益生、高氧化活性和潜在降糖功效的植物乳杆菌,该菌株分离筛选自传统发酵食品泡菜中,命名为Lactobacillus plantarum CT152,保藏编号为 CCTCC M 2019591,保藏日期为2019年7月30日,保藏于中国武汉市中国典型培养物保藏中心。
该菌株在MRS培养基上,呈圆形,表面光滑,细密,色白,偶尔呈浅黄或深黄色,菌落大小均匀。生理特性:能利用N-乙酰基-D半乳糖胺,龙胆二糖,α-D-葡萄糖,D-纤维二糖,D-果糖,甘露糖,甘露醇,麦芽糖,β-甲基-D-葡萄糖,D-洛酮糖,D-核糖,水杨苷,D-海藻糖,D-木糖,丙酮酸甲酯,丙三醇,尿苷。不能利用淀粉,甘露聚糖,D-半乳糖,L-海藻糖,L-谷氨酸等等。
本发明从泡菜和发酵酸奶中分离纯化得到120株乳酸菌,通过耐酸和耐胆盐初筛得到18株候选的实验菌株,通过检测益生指标(模拟胃肠液的耐受能力、胆盐耐受能力、对Caco-2细胞的粘附能力);抗氧化性指标(对DPPH自由基的清除能力、羟自由基的清除能力、抗脂质过氧化能力、还原能力,以及对谷胱甘肽过氧化物酶GSH-Px和总超氧化物歧化酶T-SOD活力的测定);抗糖尿病指标(α-葡萄糖苷酶抑制率的测定和在乳酸菌干预下大鼠胰岛瘤细胞INS-1产胰岛素能力测定)来综合评价候选20株乳酸菌的降血糖能力,通过主成分分析法 (PCA)分析得到益生、抗氧化、抗糖尿病的指数得分情况,再运用层次分析法(AHP)得到降糖综合指标得分,以商业菌株LGG(Lactobacillus.rhamnosus GG ATCC 53103)为对照,得到较LGG综合得分较高者为筛选出来的降糖菌株。
本发明所采用的培养基配方(1L):蛋白胨10g,葡萄糖20g,酵母提取物 5g,牛肉膏10g,磷酸二氢钾2g,柠檬酸氢铵2g,乙酸钠5g,七水硫酸镁0.5g,五水硫酸锰0.25g,吐温801mL,琼脂20g(固体培养),碳酸钙7.5g(固体分离培养)。
本实施例的各项测试记录如下表1-表8所示,其中:
表1为乳酸菌对肠胃液的耐受能力测定;
表2为乳酸菌对胆盐耐受能力测定;
表3乳酸菌DPPH清除和羟自由基清除能力;
表4乳酸菌抗脂质过氧化和还原能力测定;
表5乳酸菌发酵上清和细胞内GSH-Px和T-SOD的活力;
表6乳酸菌对大鼠肠道提取的α-glucosidase活力的抑制率;
表7乳酸菌抗生素敏感性测试结果;
表8通过PCA和AHP方法综合降糖指数得分情况;
表1:
表2:
表3:
表4:
表5:
表6:
表7:
GEN,庆大霉素;AMP,青霉素;KM,卡拉霉素;STR,链霉素;TET,四环素;
ERY,红霉素;CLI,克林霉素;VAN,万古霉素;RIF,利福平;CM,氯霉素.
表8:
以上所述实施例仅表示本发明的若干实施方式,其描述较为具体和详细,但并不能理解为对本发明范围的限制。应当指出的是,对于本领域的普通技术人员来说,在不脱离本发明构思的前提下,还可以做出若干变形和改进,这些都属于本发明保护范围。因此本发明的保护范围应该以所述权利要求为准。
Claims (12)
1.一种植物乳杆菌,其特征在于:该植物乳杆菌的保藏编号为CCTCC M 2019591,拉丁文名称为Lactobacillus plantarum CT152。
2.一种具有降血糖功能的药品及食品,其特征在于:采用权利要求1所述的植物乳杆菌。
3.一种植物乳杆菌的筛选方法,其特征在于,包括如下步骤:
S1:进行益生指标测定,包括模拟的胃肠液中的存活率测定、胆盐耐受力测定、对Caco-2细胞的粘附能力测定及抗生素耐药性检测;
S2:进行抗氧化能力测定;
S3:进行乳酸菌的抗糖尿病特性的检测,包括大鼠肠道α-glucosidase活力抑制率的测定和乳酸菌干预高糖刺激胰岛细胞分泌胰岛素实验。
4.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述进行抗氧化能力测定的步骤,具体包括:乳酸菌清除DPPH能力的测定、乳酸菌清除羟自由基能力测定、乳酸菌抗脂质过氧化能力测定、乳酸菌的还原能力测定及乳酸菌的GSH-Px和T-SOD的酶活测定。
5.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:还包括进行体内实验验证的步骤,具体包括2型糖尿病模型大鼠的构建及进行灌胃治疗。
6.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述模拟的胃肠液中的存活率测定的具体步骤包括:将乳酸菌用0.85%生理盐水重悬后,在pH为3的模拟胃液中将其菌液密度调节为1×109CFU/mL,混匀后将其放置于37℃培养1,2和3h后分别计活菌数,在模拟胃液中培养3h后,取1mL培养液加入至9mL pH为8的模拟肠液中,混匀后于37℃培养,分别于2、4和8h检测活菌数,用MRS固体培养基37℃培养24-48h,计算其存活率,
存活率(%)=[log CFU N1/log CFU N]×100%
N1=经过模拟胃肠液处理之后的乳酸菌活菌数,N0=未处理前乳酸菌的活菌数。
7.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述胆盐耐受力测定的步骤包括:培养18h-24h的乳酸菌按2-5%分别接种于含有或不含有0.3%胆盐的MRS-THIO培养基中,混合物于37℃培养16-24h,每隔一段时间在600nm;处测吸光值,记录不加胆盐和加了胆盐的培养基吸光值达到0.3个单位时各自所需的时间,这两个时间差即为乳酸菌在胆盐中生长的延迟时间,延迟时间设为LT,LT越短说明乳酸菌,耐受胆盐的能力越强;所述MRS-THIO培养基中中的MRS含有0.2%的巯基乙酸钠。
8.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述对Caco-2细胞的粘附能力测定的步骤包括:Caco-2细胞生长在含有20%胎牛血清,1%L-谷氨酰胺,1%非必需氨基酸和1%双抗的高糖DMEM培养液中,37℃,5%CO2的培养箱中进行培养,隔天更换一次培养液,等细胞增殖融合率达80%左右时,以含0.02%EDTA的0.25%胰酶消化细胞,按1:2-1:3传代,取对数生长期细胞进行试验;
将新鲜培养的乳酸菌用PBS洗两次,调节细菌浓度为1×109CFU/mL,悬浮于100mg/mL异硫氰酸荧光素中,37℃暗处作用0.5-1h,用PBS洗涤4次去除未结合的FIFC,将Caco-2细胞以4×105-5×105cell/孔接种于6孔板,37℃培养16-24h,将FIFC标记的乳酸菌重悬于DMEM培养基中,调整其密度为1×109CFU/mL,将此菌悬液加入到Caco-2细胞单层中,37℃作用1-2h,用无菌PBS洗涤3次,将未粘附的菌洗脱掉,之后每个培养的孔中加入胰酶作用,待细胞完全脱落,加入0.4mL完全培养基终止反应,收集液体,用多功能酶标仪测定其荧光强度来计算其细菌的黏附率;
粘附率(%)=Log A/Log A0×100%;
其中,A和A0分别表示乳酸菌粘附细胞前后的荧光强度。
9.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述乳酸菌清除DPPH能力测定的步骤包括:反应体系中加入1mL乳酸菌的菌悬液,无细胞提取物或者发酵上清液,再加入1mL0.2mmol/L DPPH的无水乙醇,震荡充分混匀,在室温下避光反应30min,再于6000rpm离心10min,取其上清,517nm波长处测定样品吸光度既OD值,用等体积的生理盐水代替样品溶液作为对照组,并以等体积的生理盐水和无水乙醇的混合液作为空白调零,按照以下公式计算:
DPPH自由基的清除率=[1-A517(样品)/A517(对照)]×100%;
所述乳酸菌抗脂质过氧化能力测定的步骤包括:0.5mL pH 7.4的PBS中加入1mL亚油酸乳化液,加入0.2mL0.01%的硫酸亚铁和0.01%的抗坏血酸,再加入0.5mL发酵上清,菌悬液或者细胞提取物,混匀后放置于37℃水浴中反应12h,取2mL反应液加入0.2mL0.4%的三氯乙酸,2mL0.8%的硫代巴比妥酸和0.2mL0.4%BHT,震荡充分混匀后,在100℃下反应30min,直至样品冷却后再加入2mL三氯甲烷进行抽提,离心10min后,收集上清,532nm波长下测定样品OD值,以PBS作为空白对照,按照下式计算乳酸菌抗脂质过氧化的能力:
抗脂质过氧化率=[1-A532(样品)/A532(空白)]×100%;
所述乳酸菌的还原能力测定的步骤包括:0.5mL菌悬液或细胞提取物中加入1%铁***0.5mL,0.5mLph为6.6的PBS,震荡充分混匀,于50℃中水浴20min,样品替换为蒸馏水后可作为对照组,在冰浴中急速冷却后,加入0.5mL10%的三氯乙酸,3000rpm离心10min,取其1mL上清后加入1mL0.1%的FeCL3,反应10min后在700nm波长下测定样品的OD值,结果采用半胱氨酸作为标准表征还原力,计算公式如下:
还原能力=[A700(样品)-A700(对照)/A700(对照)]×100%。
10.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述大鼠肠道α-glucosidase活力抑制率测定的步骤包括:在150μL PBS(0.1M pH6.8)中加入75μL20mM的PNPG溶液及25μL,PBS为0.1M ph值为6.8。待测乳酸菌的发酵上清,无细胞提取物和完整细胞,将混合物于37℃水浴10min,加入50μL大鼠肠道提取的α-glucosidase溶液,α-glucosidase溶液为0.17U/mL,继续反应10min,加入1mL0.1M Na2CO3作为反应终止液,并将反应液于405nm处测其吸光值,吸光值与对硝基酚的游离量成正比,反应体系中采用pH6.8的0.1M PBS作为α-glucosidase溶液及待测样品的空白对照,采用以下公式计算样品的抑制活性,
酶抑制率(%)=[1-(C-D)/(A-B)]×100%;
其中:A为含有α-glucosidase溶液但不含样品的测定吸光;
B为不含α-glucosidase溶液及待测样品的测定吸光值;
C为含有α-glucosidase溶液及待测样品的测定吸光值;
D为不含α-glucosidase溶液但含待测样品的测定吸光值。
11.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述乳酸菌干预高糖刺激胰岛细胞分泌胰岛素实验的步骤包括:乳酸菌在MRS液体培养基中厌氧传代2次,每次37℃下孵育18h,使菌株活力达到最高值,对菌悬液以灭菌的PBS缓冲液进行梯度稀释,采用平板倾注法,在MRS固体培养基上37℃培养24-48h,测定活菌数,并保证每个平板30-300的活菌菌落,根据菌粉活菌数,配制1×109CFU/mL的菌悬液,经过120kw超声破碎,然后65℃、30-60min灭活处理,8000×g离心获得菌体混合物沉淀,用不含双抗的INS-1细胞培养液进行重悬,作为GSIS实验刺激细胞的培养液,储存于4℃待用,
INS-1细胞以4×105-5×105cells/mL的密度接种于6孔板,孵育过夜待细胞贴壁,然后换用含有灭活菌液的细胞培养液孵育16-24h,对细胞进行不同益生菌株的干预作用,各组细胞处理结束后,细胞用磷酸盐缓冲液洗2次,加入葡萄糖浓度16.7mmol/L的高糖INS-1完全培养液,37℃下孵育0.5-1h,然后收集细胞上清液,用ELISA法检测各组在高葡萄糖刺激下的胰岛素的分泌量,ELISA检测步骤按照试剂盒说明书进行,吸光度采用酶标仪在450nm处进行检测,以OD值为纵坐标Y,标准物浓度为横坐标X,绘制标准曲线,并根据此计算其他样品的浓度。
12.根据权利要求3所述的植物乳杆菌的筛选方法,其特征在于:所述2型糖尿病模型大鼠的构建的步骤包括:将70只雄性6-8周龄的Wistar大鼠,用基础饲料适应性喂养5天后随机分为低脂组和高脂组,前四周,除低脂组喂食基础饲料外,其它各组均喂食高脂饲料,并于每周固定时间进行大鼠的体重测量记录;
第五周,所有大鼠禁食不禁水12-18h,除低脂组外,其余各组大鼠均按照30-40mg/kg体重注射新鲜配制的链脲霉素STZ,将一定量STZ溶于pH为4.5的柠檬酸-柠檬酸钠缓冲液中,现配现用,冰浴保存,低脂组注射ph为4.5的柠檬酸-柠檬酸钠缓冲液中,造模后72h期间保证充足的无菌水及高脂饮食,并每日更换垫料,72h后随机血糖高于16.7mmol/L或者空腹血糖高于11.7mmol/L的为造模成功的糖尿病大鼠;
将成模大鼠随机分为模型组、LGG组、CT152组,模型组用0.85%的生理盐水作为对照,LGG和CCTCCM2019591以1×109CFU/day进行为期6-8周的灌胃治疗。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201910709658.2A CN110964659A (zh) | 2019-08-02 | 2019-08-02 | 降血糖功效的植物乳杆菌及其筛选方法和应用药剂及食品 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201910709658.2A CN110964659A (zh) | 2019-08-02 | 2019-08-02 | 降血糖功效的植物乳杆菌及其筛选方法和应用药剂及食品 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN110964659A true CN110964659A (zh) | 2020-04-07 |
Family
ID=70028382
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN201910709658.2A Pending CN110964659A (zh) | 2019-08-02 | 2019-08-02 | 降血糖功效的植物乳杆菌及其筛选方法和应用药剂及食品 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN110964659A (zh) |
Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109022313A (zh) * | 2018-08-02 | 2018-12-18 | 中国农业科学院兰州兽医研究所 | 一株植物乳杆菌 |
CN112402465A (zh) * | 2020-12-18 | 2021-02-26 | 四川大学华西医院 | 一种植物乳杆菌n-1在制备预防和/或改善良性***增生的产品及用途 |
KR20220006864A (ko) * | 2020-07-09 | 2022-01-18 | 한국식품연구원 | 락토바실러스 플란타룸 k97 및 이의 용도 |
CN114437997A (zh) * | 2022-04-07 | 2022-05-06 | 山东向日葵生物工程有限公司 | 一种植物乳杆菌sf-l38及其在制备血糖控制产品中的应用 |
CN114752529A (zh) * | 2022-04-29 | 2022-07-15 | 科郦有限公司 | 植物乳杆菌hom3201菌株及其活菌制剂、制备方法和用途 |
Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN105567586A (zh) * | 2015-12-21 | 2016-05-11 | 南昌大学 | 一株具有抗糖尿病功能的植物乳杆菌及其应用 |
CN106834196A (zh) * | 2017-04-10 | 2017-06-13 | 吉林省农业科学院 | 植物乳杆菌ch126及其在制备降糖功能性食品中的应用 |
CN107502575A (zh) * | 2017-09-20 | 2017-12-22 | 中国农业科学院农产品加工研究所 | 一株具有α‑葡萄糖苷酶高抑制活性的植物乳杆菌 |
CN108018236A (zh) * | 2017-12-18 | 2018-05-11 | 富乐顿生物工程科技(北京)有限公司 | 一种降低α-葡萄糖苷酶活性的空间植物乳杆菌SS18-37及其应用 |
-
2019
- 2019-08-02 CN CN201910709658.2A patent/CN110964659A/zh active Pending
Patent Citations (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN105567586A (zh) * | 2015-12-21 | 2016-05-11 | 南昌大学 | 一株具有抗糖尿病功能的植物乳杆菌及其应用 |
CN106834196A (zh) * | 2017-04-10 | 2017-06-13 | 吉林省农业科学院 | 植物乳杆菌ch126及其在制备降糖功能性食品中的应用 |
CN107502575A (zh) * | 2017-09-20 | 2017-12-22 | 中国农业科学院农产品加工研究所 | 一株具有α‑葡萄糖苷酶高抑制活性的植物乳杆菌 |
CN108018236A (zh) * | 2017-12-18 | 2018-05-11 | 富乐顿生物工程科技(北京)有限公司 | 一种降低α-葡萄糖苷酶活性的空间植物乳杆菌SS18-37及其应用 |
Non-Patent Citations (1)
Title |
---|
TING CAI ET AL.: "In vitro evaluation by PCA and AHP of potential antidiabetic properties of lactic acid bacteria isolated from traditional fermented food", 《LWT - FOOD SCIENCE AND TECHNOLOGY》 * |
Cited By (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109022313A (zh) * | 2018-08-02 | 2018-12-18 | 中国农业科学院兰州兽医研究所 | 一株植物乳杆菌 |
CN109022313B (zh) * | 2018-08-02 | 2021-08-24 | 中国农业科学院兰州兽医研究所 | 一株植物乳杆菌 |
KR20220006864A (ko) * | 2020-07-09 | 2022-01-18 | 한국식품연구원 | 락토바실러스 플란타룸 k97 및 이의 용도 |
KR102570432B1 (ko) | 2020-07-09 | 2023-08-25 | 한국식품연구원 | 락토바실러스 플란타룸 k97 및 이의 용도 |
CN112402465A (zh) * | 2020-12-18 | 2021-02-26 | 四川大学华西医院 | 一种植物乳杆菌n-1在制备预防和/或改善良性***增生的产品及用途 |
CN112402465B (zh) * | 2020-12-18 | 2022-12-30 | 四川大学华西医院 | 一种植物乳杆菌n-1在制备预防和/或改善良性***增生的产品及用途 |
CN114437997A (zh) * | 2022-04-07 | 2022-05-06 | 山东向日葵生物工程有限公司 | 一种植物乳杆菌sf-l38及其在制备血糖控制产品中的应用 |
CN114437997B (zh) * | 2022-04-07 | 2022-06-10 | 山东向日葵生物工程有限公司 | 一种植物乳杆菌sf-l38及其在制备血糖控制产品中的应用 |
CN114752529A (zh) * | 2022-04-29 | 2022-07-15 | 科郦有限公司 | 植物乳杆菌hom3201菌株及其活菌制剂、制备方法和用途 |
CN114752529B (zh) * | 2022-04-29 | 2023-12-19 | 科郦有限公司 | 植物乳杆菌hom3201菌株及其活菌制剂、制备方法和用途 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN110964659A (zh) | 降血糖功效的植物乳杆菌及其筛选方法和应用药剂及食品 | |
CN102597214B (zh) | 新型植物乳杆菌及含该植物乳杆菌的组合物 | |
CN110835616B (zh) | 副干酪乳杆菌gks6的活性物质、含其的组合物及其促进长寿的用途 | |
Kojima et al. | Combining prebiotics and probiotics to develop novel synbiotics that suppress oral pathogens | |
CN110157647B (zh) | 一种能够缓解焦虑、改善睡眠的短乳杆菌及其用途 | |
EP3715448A2 (en) | Novel lactic acid bacteria and use thereof | |
CN109182184B (zh) | 一株犬源乳酸片球菌及其应用 | |
CN109182162B (zh) | 一株具有抗氧化能力的植物乳杆菌及应用 | |
CN101744841A (zh) | 用于治疗炎性疾病的双歧杆菌 | |
CA2609617A1 (en) | Feline probiotic lactobacilli | |
CN109234189A (zh) | 一株具有抗氧化能力的植物乳杆菌菌株bx62及其应用 | |
CN110577912B (zh) | 一种格氏乳杆菌及其在制备发酵乳中的应用 | |
CN113308421B (zh) | 一种植物乳杆菌bufx及其在代谢综合征中的应用 | |
CN114574406B (zh) | 鼠李糖乳杆菌菌株wka55及其在制备防治酒精性肝损伤制品方面的用途与产品 | |
CN110833565B (zh) | 植物乳杆菌gkm3的活性物质、含其的组合物及其促进长寿的用途 | |
Garcia-Mazcorro et al. | Are there indigenous Saccharomyces in the digestive tract of livestock animal species? Implications for health, nutrition and productivity traits | |
CN115322932B (zh) | 一株具有解酒醒酒能力的植物乳植杆菌和应用 | |
CN112154202A (zh) | 模拟肠道环境 | |
CN113322216A (zh) | 一种副干酪乳杆菌b111h及其在代谢综合征中的应用 | |
CN113444656B (zh) | 一种具有抗衰老功能的菌株及其应用 | |
Aklilu et al. | Identification and biotyping of Escherichia coli from diarrheic lambs in and around Debre Birhan town, Ethiopia | |
CN112708574B (zh) | 保加利亚乳杆菌tci904、其组合物及其用于减少体重的用途 | |
TWI669124B (zh) | 芽孢乳酸菌用於製備代謝重金屬及保護肝臟醫藥組成物之用途 | |
Bobga et al. | Evaluation of the anti-diabetic potential of probiotic Lactobacillus fermentum (PRI 29) isolated from cameroonian fermented cow milk in alloxan induced diabetes type-1 mice model | |
TWI788633B (zh) | 新穎乳酸菌用於製備嘌呤化合物降解劑之用途 |
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: 20200407 |