KR20200130889A - Novel Lactobacillus plantarum KCC-35 - Google Patents
Novel Lactobacillus plantarum KCC-35 Download PDFInfo
- Publication number
- KR20200130889A KR20200130889A KR1020190055227A KR20190055227A KR20200130889A KR 20200130889 A KR20200130889 A KR 20200130889A KR 1020190055227 A KR1020190055227 A KR 1020190055227A KR 20190055227 A KR20190055227 A KR 20190055227A KR 20200130889 A KR20200130889 A KR 20200130889A
- Authority
- KR
- South Korea
- Prior art keywords
- strain
- kcc
- kacc
- present
- lactobacillus plantarum
- Prior art date
Links
- 240000006024 Lactobacillus plantarum Species 0.000 title claims abstract description 17
- 235000013965 Lactobacillus plantarum Nutrition 0.000 title claims abstract description 12
- 229940072205 lactobacillus plantarum Drugs 0.000 title claims abstract description 12
- 239000006041 probiotic Substances 0.000 claims abstract description 36
- 235000018291 probiotics Nutrition 0.000 claims abstract description 36
- 230000000529 probiotic effect Effects 0.000 claims abstract description 23
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 claims abstract description 18
- 235000013305 food Nutrition 0.000 claims abstract description 16
- 239000004310 lactic acid Substances 0.000 claims abstract description 9
- 235000014655 lactic acid Nutrition 0.000 claims abstract description 9
- 235000021107 fermented food Nutrition 0.000 claims abstract description 8
- 238000000034 method Methods 0.000 claims description 16
- 238000002360 preparation method Methods 0.000 claims description 7
- 230000000813 microbial effect Effects 0.000 claims description 5
- 244000144972 livestock Species 0.000 claims description 3
- 230000000843 anti-fungal effect Effects 0.000 abstract description 15
- 238000000855 fermentation Methods 0.000 abstract description 11
- 230000004151 fermentation Effects 0.000 abstract description 11
- 230000000844 anti-bacterial effect Effects 0.000 abstract description 10
- 239000003242 anti bacterial agent Substances 0.000 abstract description 8
- 238000004519 manufacturing process Methods 0.000 abstract description 8
- 241000894006 Bacteria Species 0.000 abstract description 7
- 108090000790 Enzymes Proteins 0.000 abstract description 7
- 102000004190 Enzymes Human genes 0.000 abstract description 7
- 235000017587 Medicago sativa ssp. sativa Nutrition 0.000 abstract description 5
- 230000003078 antioxidant effect Effects 0.000 abstract description 5
- 150000001720 carbohydrates Chemical class 0.000 abstract description 5
- 235000014633 carbohydrates Nutrition 0.000 abstract description 5
- 241000219823 Medicago Species 0.000 abstract description 4
- 239000002253 acid Substances 0.000 abstract description 4
- 210000000941 bile Anatomy 0.000 abstract description 3
- 230000033228 biological regulation Effects 0.000 abstract description 3
- 239000003795 chemical substances by application Substances 0.000 abstract 1
- 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 18
- 238000004458 analytical method Methods 0.000 description 14
- 239000000243 solution Substances 0.000 description 13
- 238000002835 absorbance Methods 0.000 description 12
- 238000002474 experimental method Methods 0.000 description 12
- 210000004027 cell Anatomy 0.000 description 10
- 238000012360 testing method Methods 0.000 description 10
- 235000012000 cholesterol Nutrition 0.000 description 9
- 230000000052 comparative effect Effects 0.000 description 9
- 244000005700 microbiome Species 0.000 description 9
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 8
- 235000013406 prebiotics Nutrition 0.000 description 8
- 239000003833 bile salt Substances 0.000 description 7
- 230000000694 effects Effects 0.000 description 7
- 239000002609 medium Substances 0.000 description 7
- 239000000843 powder Substances 0.000 description 7
- 230000002292 Radical scavenging effect Effects 0.000 description 6
- 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 description 6
- 229940088598 enzyme Drugs 0.000 description 6
- 239000007788 liquid Substances 0.000 description 6
- 238000012216 screening Methods 0.000 description 6
- 108020004465 16S ribosomal RNA Proteins 0.000 description 5
- 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 5
- 229920001202 Inulin Polymers 0.000 description 5
- MUPFEKGTMRGPLJ-RMMQSMQOSA-N Raffinose Natural products O(C[C@H]1[C@@H](O)[C@H](O)[C@@H](O)[C@@H](O[C@@]2(CO)[C@H](O)[C@@H](O)[C@@H](CO)O2)O1)[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 MUPFEKGTMRGPLJ-RMMQSMQOSA-N 0.000 description 5
- MUPFEKGTMRGPLJ-UHFFFAOYSA-N UNPD196149 Natural products OC1C(O)C(CO)OC1(CO)OC1C(O)C(O)C(O)C(COC2C(C(O)C(O)C(CO)O2)O)O1 MUPFEKGTMRGPLJ-UHFFFAOYSA-N 0.000 description 5
- 229940088710 antibiotic agent Drugs 0.000 description 5
- 239000008103 glucose Substances 0.000 description 5
- JYJIGFIDKWBXDU-MNNPPOADSA-N inulin Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)OC[C@]1(OC[C@]2(OC[C@]3(OC[C@]4(OC[C@]5(OC[C@]6(OC[C@]7(OC[C@]8(OC[C@]9(OC[C@]%10(OC[C@]%11(OC[C@]%12(OC[C@]%13(OC[C@]%14(OC[C@]%15(OC[C@]%16(OC[C@]%17(OC[C@]%18(OC[C@]%19(OC[C@]%20(OC[C@]%21(OC[C@]%22(OC[C@]%23(OC[C@]%24(OC[C@]%25(OC[C@]%26(OC[C@]%27(OC[C@]%28(OC[C@]%29(OC[C@]%30(OC[C@]%31(OC[C@]%32(OC[C@]%33(OC[C@]%34(OC[C@]%35(OC[C@]%36(O[C@@H]%37[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O%37)O)[C@H]([C@H](O)[C@@H](CO)O%36)O)[C@H]([C@H](O)[C@@H](CO)O%35)O)[C@H]([C@H](O)[C@@H](CO)O%34)O)[C@H]([C@H](O)[C@@H](CO)O%33)O)[C@H]([C@H](O)[C@@H](CO)O%32)O)[C@H]([C@H](O)[C@@H](CO)O%31)O)[C@H]([C@H](O)[C@@H](CO)O%30)O)[C@H]([C@H](O)[C@@H](CO)O%29)O)[C@H]([C@H](O)[C@@H](CO)O%28)O)[C@H]([C@H](O)[C@@H](CO)O%27)O)[C@H]([C@H](O)[C@@H](CO)O%26)O)[C@H]([C@H](O)[C@@H](CO)O%25)O)[C@H]([C@H](O)[C@@H](CO)O%24)O)[C@H]([C@H](O)[C@@H](CO)O%23)O)[C@H]([C@H](O)[C@@H](CO)O%22)O)[C@H]([C@H](O)[C@@H](CO)O%21)O)[C@H]([C@H](O)[C@@H](CO)O%20)O)[C@H]([C@H](O)[C@@H](CO)O%19)O)[C@H]([C@H](O)[C@@H](CO)O%18)O)[C@H]([C@H](O)[C@@H](CO)O%17)O)[C@H]([C@H](O)[C@@H](CO)O%16)O)[C@H]([C@H](O)[C@@H](CO)O%15)O)[C@H]([C@H](O)[C@@H](CO)O%14)O)[C@H]([C@H](O)[C@@H](CO)O%13)O)[C@H]([C@H](O)[C@@H](CO)O%12)O)[C@H]([C@H](O)[C@@H](CO)O%11)O)[C@H]([C@H](O)[C@@H](CO)O%10)O)[C@H]([C@H](O)[C@@H](CO)O9)O)[C@H]([C@H](O)[C@@H](CO)O8)O)[C@H]([C@H](O)[C@@H](CO)O7)O)[C@H]([C@H](O)[C@@H](CO)O6)O)[C@H]([C@H](O)[C@@H](CO)O5)O)[C@H]([C@H](O)[C@@H](CO)O4)O)[C@H]([C@H](O)[C@@H](CO)O3)O)[C@H]([C@H](O)[C@@H](CO)O2)O)[C@@H](O)[C@H](O)[C@@H](CO)O1 JYJIGFIDKWBXDU-MNNPPOADSA-N 0.000 description 5
- 229940029339 inulin Drugs 0.000 description 5
- 108090000623 proteins and genes Proteins 0.000 description 5
- MUPFEKGTMRGPLJ-ZQSKZDJDSA-N raffinose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO[C@@H]2[C@@H]([C@@H](O)[C@@H](O)[C@@H](CO)O2)O)O1 MUPFEKGTMRGPLJ-ZQSKZDJDSA-N 0.000 description 5
- 229920001817 Agar Polymers 0.000 description 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- SRBFZHDQGSBBOR-IOVATXLUSA-N D-xylopyranose Chemical compound O[C@@H]1COC(O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-IOVATXLUSA-N 0.000 description 4
- 240000004296 Lolium perenne Species 0.000 description 4
- 239000008272 agar Substances 0.000 description 4
- 230000002776 aggregation Effects 0.000 description 4
- 238000004220 aggregation Methods 0.000 description 4
- 210000004051 gastric juice Anatomy 0.000 description 4
- 239000008188 pellet Substances 0.000 description 4
- 239000004460 silage Substances 0.000 description 4
- 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 4
- 241000228193 Aspergillus clavatus Species 0.000 description 3
- 241000186660 Lactobacillus Species 0.000 description 3
- 240000004658 Medicago sativa Species 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 241000228150 Penicillium chrysogenum Species 0.000 description 3
- 241000825258 Scopulariopsis brevicaulis Species 0.000 description 3
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 3
- 240000006394 Sorghum bicolor Species 0.000 description 3
- 235000011684 Sorghum saccharatum Nutrition 0.000 description 3
- 230000028654 Type IV pili-dependent aggregation Effects 0.000 description 3
- 240000008042 Zea mays Species 0.000 description 3
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 3
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 3
- 230000002378 acidificating effect Effects 0.000 description 3
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 3
- 239000006285 cell suspension Substances 0.000 description 3
- 238000012512 characterization method Methods 0.000 description 3
- 235000005822 corn Nutrition 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- MGJZITXUQXWAKY-UHFFFAOYSA-N diphenyl-(2,4,6-trinitrophenyl)iminoazanium Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1N=[N+](C=1C=CC=CC=1)C1=CC=CC=C1 MGJZITXUQXWAKY-UHFFFAOYSA-N 0.000 description 3
- 230000007760 free radical scavenging Effects 0.000 description 3
- 230000002496 gastric effect Effects 0.000 description 3
- 210000001035 gastrointestinal tract Anatomy 0.000 description 3
- 230000036541 health Effects 0.000 description 3
- BVIDQAVCCRUFGU-UHFFFAOYSA-M methyl sulfate;trimethyl(1-phenothiazin-10-ylpropan-2-yl)azanium Chemical compound COS([O-])(=O)=O.C1=CC=C2N(CC(C)[N+](C)(C)C)C3=CC=CC=C3SC2=C1 BVIDQAVCCRUFGU-UHFFFAOYSA-M 0.000 description 3
- 239000006872 mrs medium Substances 0.000 description 3
- UHPMCKVQTMMPCG-UHFFFAOYSA-N 5,8-dihydroxy-2-methoxy-6-methyl-7-(2-oxopropyl)naphthalene-1,4-dione Chemical compound CC1=C(CC(C)=O)C(O)=C2C(=O)C(OC)=CC(=O)C2=C1O UHPMCKVQTMMPCG-UHFFFAOYSA-N 0.000 description 2
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 241001225321 Aspergillus fumigatus Species 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 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 2
- 108020004414 DNA Proteins 0.000 description 2
- 241000223218 Fusarium Species 0.000 description 2
- 235000010624 Medicago sativa Nutrition 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- CTQNGGLPUBDAKN-UHFFFAOYSA-N O-Xylene Chemical compound CC1=CC=CC=C1C CTQNGGLPUBDAKN-UHFFFAOYSA-N 0.000 description 2
- 239000004098 Tetracycline Substances 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 230000000996 additive effect Effects 0.000 description 2
- 102000005840 alpha-Galactosidase Human genes 0.000 description 2
- 108010030291 alpha-Galactosidase Proteins 0.000 description 2
- 229940121375 antifungal agent Drugs 0.000 description 2
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 2
- 229940091771 aspergillus fumigatus Drugs 0.000 description 2
- 230000008901 benefit Effects 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
- 102000006995 beta-Glucosidase Human genes 0.000 description 2
- 108010047754 beta-Glucosidase Proteins 0.000 description 2
- 229940093761 bile salts Drugs 0.000 description 2
- 230000003115 biocidal effect Effects 0.000 description 2
- 238000004364 calculation method Methods 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 230000008859 change Effects 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
- 230000005757 colony formation Effects 0.000 description 2
- 230000003247 decreasing effect Effects 0.000 description 2
- 229960003964 deoxycholic acid Drugs 0.000 description 2
- 229930195733 hydrocarbon Natural products 0.000 description 2
- 150000002430 hydrocarbons Chemical class 0.000 description 2
- 210000000936 intestine Anatomy 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 229930027917 kanamycin Natural products 0.000 description 2
- 229960000318 kanamycin Drugs 0.000 description 2
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 2
- 229930182823 kanamycin A Natural products 0.000 description 2
- 229940039696 lactobacillus Drugs 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 229960000564 nitrofurantoin Drugs 0.000 description 2
- NXFQHRVNIOXGAQ-YCRREMRBSA-N nitrofurantoin Chemical compound O1C([N+](=O)[O-])=CC=C1\C=N\N1C(=O)NC(=O)C1 NXFQHRVNIOXGAQ-YCRREMRBSA-N 0.000 description 2
- 244000052769 pathogen Species 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- FHHPUSMSKHSNKW-SMOYURAASA-M sodium deoxycholate Chemical compound [Na+].C([C@H]1CC2)[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 FHHPUSMSKHSNKW-SMOYURAASA-M 0.000 description 2
- 229960005322 streptomycin Drugs 0.000 description 2
- 229960002180 tetracycline Drugs 0.000 description 2
- 229930101283 tetracycline Natural products 0.000 description 2
- 235000019364 tetracycline Nutrition 0.000 description 2
- 150000003522 tetracyclines Chemical class 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 239000008096 xylene Substances 0.000 description 2
- 241000228197 Aspergillus flavus Species 0.000 description 1
- 239000002028 Biomass Substances 0.000 description 1
- 206010006187 Breast cancer Diseases 0.000 description 1
- 208000026310 Breast neoplasm Diseases 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- 206010010774 Constipation Diseases 0.000 description 1
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 1
- 241000194032 Enterococcus faecalis Species 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- VGGSQFUCUMXWEO-UHFFFAOYSA-N Ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 description 1
- 239000005977 Ethylene Substances 0.000 description 1
- 206010016654 Fibrosis Diseases 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241000223221 Fusarium oxysporum Species 0.000 description 1
- 208000018522 Gastrointestinal disease Diseases 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 206010020751 Hypersensitivity Diseases 0.000 description 1
- 206010020772 Hypertension Diseases 0.000 description 1
- 208000022559 Inflammatory bowel disease Diseases 0.000 description 1
- 201000010538 Lactose Intolerance Diseases 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 102000057297 Pepsin A Human genes 0.000 description 1
- 108090000284 Pepsin A Proteins 0.000 description 1
- 241000589517 Pseudomonas aeruginosa Species 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- 241000191967 Staphylococcus aureus Species 0.000 description 1
- 229930003268 Vitamin C Natural products 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000008186 active pharmaceutical agent Substances 0.000 description 1
- 230000032683 aging Effects 0.000 description 1
- 208000026935 allergic disease Diseases 0.000 description 1
- 230000007815 allergy Effects 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 230000001093 anti-cancer Effects 0.000 description 1
- 230000002790 anti-mutagenic effect Effects 0.000 description 1
- 230000003262 anti-osteoporosis Effects 0.000 description 1
- 238000011482 antibacterial activity assay Methods 0.000 description 1
- 238000011483 antifungal activity assay Methods 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000003149 assay kit Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 230000007882 cirrhosis Effects 0.000 description 1
- 208000019425 cirrhosis of liver Diseases 0.000 description 1
- 210000001072 colon Anatomy 0.000 description 1
- 208000029742 colonic neoplasm Diseases 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- -1 controllability Substances 0.000 description 1
- 239000012228 culture supernatant Substances 0.000 description 1
- 235000015140 cultured milk Nutrition 0.000 description 1
- 235000014048 cultured milk product Nutrition 0.000 description 1
- 235000013365 dairy product Nutrition 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 210000002249 digestive system Anatomy 0.000 description 1
- 229940032049 enterococcus faecalis Drugs 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 244000078673 foodborn pathogen Species 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 235000013376 functional food Nutrition 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 239000007789 gas Substances 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 230000007366 host health Effects 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 238000011835 investigation Methods 0.000 description 1
- 208000002551 irritable bowel syndrome Diseases 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 230000037353 metabolic pathway Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 235000005900 nutrient bioavailability Nutrition 0.000 description 1
- 230000001717 pathogenic effect Effects 0.000 description 1
- 229940111202 pepsin Drugs 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 230000008092 positive effect Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 238000011002 quantification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000002000 scavenging effect Effects 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 210000000813 small intestine Anatomy 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 239000012137 tryptone Substances 0.000 description 1
- 230000035899 viability Effects 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C 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
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
-
- 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
- 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
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/32—Foods, ingredients or supplements having a functional effect on health having an effect on the health of the digestive tract
- A23V2200/3204—Probiotics, living bacteria to be ingested for action in the digestive tract
-
- 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
-
- A23Y2220/67—
-
- C12R1/25—
-
- 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
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Polymers & Plastics (AREA)
- Microbiology (AREA)
- Zoology (AREA)
- Organic Chemistry (AREA)
- Food Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Genetics & Genomics (AREA)
- Wood Science & Technology (AREA)
- Biochemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- General Health & Medical Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Mycology (AREA)
- Nutrition Science (AREA)
- Virology (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Physiology (AREA)
- Animal Husbandry (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Description
본 발명은 신규 유산균 락토바실러스 플란타럼 KCC-35 균주에 관한 것으로, 구체적으로 알팔파 사료에서 분리된 신규 유산균 락토바실러스 플란타럼(Lactobacillus plantarum) KCC-35 균주(미생물 수탁번호 KACC 92222P)에 관한 것이다.The present invention relates to a novel lactic acid bacteria Lactobacillus plantarum KCC-35 strain, specifically, to a novel lactic acid bacteria Lactobacillus plantarum ( Lactobacillus plantarum ) KCC-35 strain (microbial accession number KACC 92222P) isolated from alfalfa feed. .
사람의 위장관은 복잡하고 강력한 미생물 환경을 갖고 있다. 이러한 미생물 환경은 나이, 건강 및 생활양식에 따라 변화하며, 병원체와 유해음식물질의 작용을 저해하고 숙주에게 유익한 효과를 촉진하는 중요한 역할을 한다. 이러한 효과는 위장관 내 미생물의 경향과 관련이 있으며, 위장관의 이러한 미생물의 균형은 위장병의 위험을 낮출 수 있다. 프로바이오틱스의 이점으로는 락토오스 불내성, 영양분의 생체이용률 및 알레르기 예방 개선이 포함되며, 항-돌연변이, 항암, 항-고콜레스테롤, 항-고혈압, 항-골다공증 및 면역기능조절 효과가 보고된 바 있다. 또한 프로바이오틱스는 염증성 장 질환, 과민성 장증후군, 간경변, 변비를 없애고 결장암, 간암 및 유방암 위험을 줄일 수 있는 것으로도 보고된 바 있다.The human gastrointestinal tract has a complex and powerful microbial environment. These microbial environments change with age, health, and lifestyle, and play an important role in inhibiting the action of pathogens and harmful food substances and promoting beneficial effects to the host. These effects are related to the tendency of microbes in the gastrointestinal tract, and the balance of these microbes in the gastrointestinal tract can lower the risk of gastrointestinal diseases. Advantages of probiotics include improved lactose intolerance, nutrient bioavailability and allergy prevention, and anti-mutagenic, anti-cancer, anti-high cholesterol, anti-hypertension, anti-osteoporosis, and immune function modulating effects have been reported. It has also been reported that probiotics can eliminate inflammatory bowel disease, irritable bowel syndrome, cirrhosis, and constipation and reduce the risk of colon, liver and breast cancer.
현재, 건강 증진 프로바이오틱 식품의 장점은 전세계적으로 널리 받아들여지고 있다. FAO/WHO에 따르면, 건강 증진용 프로바이오틱스는 충분한 양(적어도 106 ~ 107 cfu/g)으로 섭취하면 숙주의 건강에 긍정적인 영향을 미치는 생존 가능한 활성 미생물이다. 최근 연구에서 죽은 미생물의 면역학적 특성이 보고된 바 있다. 대부분의 프로바이오틱 미생물은 유산균을 생산하는 박테리아이며 그중 Lactobacillus는 인간의 건강을 향상시키는 가장 기본적인 속균 중 하나이다. 따라서 여러 락토바실러스 종들의 게놈 서열이 규명되었으며 광범위한 발효식품에 사용되어 왔다. 또한 이러한 락토바실러스와 관련된 대사경로, 유전자의 조절 및 기능이 보고되었다. 대부분의 연구자들은 프로바이오틱스를 전달하기 위한 가장 좋은 방법이 특히 발효유와 요구르트를 통한 발효유제품이라고 생각해왔다.Currently, the benefits of health-promoting probiotic foods are widely accepted worldwide. According to FAO/WHO, health-promoting probiotics are viable, active microorganisms that, when consumed in sufficient amounts (at least 10 6 to 10 7 cfu/g), have a positive effect on the health of the host. In recent studies, the immunological properties of dead microorganisms have been reported. Most probiotic microorganisms are lactic acid-producing bacteria, of which Lactobacillus is one of the most basic genus bacteria that improve human health. Therefore, the genome sequence of several Lactobacillus species has been identified and has been used in a wide range of fermented foods. In addition, the metabolic pathways related to Lactobacillus, regulation and function of genes have been reported. Most researchers have thought that the best way to deliver probiotics is fermented milk products, especially fermented milk and yogurt.
오늘날 연구자들은 새로운 바이오틱스를 생산하는 것에 더 많은 관심을 가지고 있으며, 비유제품 프로바이오틱스는 사람을 위한 프로바이오틱 기능성 식품의 개발을 위한 좋은 기반이 될 수 있을 것으로 생각한다. 잘 특성화된 프로바이오틱 미생물 중 몇몇은 전세계에서 상업적으로 이용가능하다. 그러나 신규한 미생물의 스크리닝은 여전히 발효식품산업으로부터 큰 관심을 받고 있다.Today, researchers are more interested in producing new biotics, and they believe that non-dairy probiotics could be a good basis for the development of probiotic functional foods for humans. Several of the well characterized probiotic microorganisms are commercially available worldwide. However, screening of novel microorganisms is still receiving great attention from the fermented food industry.
이에 본 발명자는 알팔파 사일리지로부터 프로바이오틱스의 우수한 특성을 갖는 신규 미생물을 스크리닝하였으며, 이의 결과 신규 유산균 락토바실러스 플란타럼 KCC-35 균주가 항박테리아 및 항진균 활성, 다양한 탄수화물의 발효능, 다양한 효소 생산능, 항균제를 이용한 조절성, 내산성, 내담즙성, 응집성, 소수성, 항산화 활성 등 매우 우수한 프로바이오틱스의 특성을 나타냄을 확인하고 본 발명을 완성하게 되었다.Accordingly, the present inventors screened new microorganisms having excellent properties of probiotics from alfalfa silage, and as a result, the new lactic acid bacteria Lactobacillus plantarum KCC-35 strain has antibacterial and antifungal activity, fermentation capacity of various carbohydrates, various enzyme production capacity, The present invention was completed by confirming that it exhibits very excellent properties of probiotics such as controllability, acid resistance, bile resistance, cohesiveness, hydrophobicity, and antioxidant activity using an antibacterial agent.
따라서 본 발명의 주된 목적은 프로바이오틱스로서의 우수한 특성을 갖는 새로운 균주를 제공하는데 있다.Therefore, the main object of the present invention is to provide a new strain having excellent properties as probiotics.
본 발명의 다른 목적은 상기 균주를 이용한 프로바이오틱스 생균제제, 식품 및 사료를 제공하는데 있다.Another object of the present invention is to provide a probiotic probiotic, food and feed using the strain.
본 발명의 한 양태에 따르면, 본 발명은 신규 유산균 락토바실러스 플란타럼(Lactobacillus plantarum) KCC-35 균주(미생물 수탁번호 KACC 92222P)를 제공한다.According to one aspect of the present invention, the present invention provides a novel lactic acid bacteria Lactobacillus plantarum KCC-35 strain (microbial accession number KACC 92222P).
본 발명의 다른 양태에 따르면, 본 발명은 상기 균주를 포함하는 프로바이오틱스 생균제제를 제공한다.According to another aspect of the present invention, the present invention provides a probiotic probiotic preparation comprising the strain.
본 발명의 생균제제에 있어서, 사람 또는 가축용 프로바이오틱스 생균제제인 것이 바람직하다.In the probiotic preparation of the present invention, it is preferably a probiotic probiotic preparation for human or livestock.
본 발명의 또 다른 양태에 따르면, 본 발명은 상기 균주 또는 상기 균주의 배양액을 포함하는 식품을 제공한다.According to another aspect of the present invention, the present invention provides a food comprising the strain or a culture solution of the strain.
본 발명의 또 다른 양태에 따르면, 본 발명은 상기 균주로 발효하여 제조된 발효식품을 제공한다.According to another aspect of the present invention, the present invention provides a fermented food manufactured by fermenting with the strain.
본 발명의 또 다른 양태에 따르면, 본 발명은 상기 균주 또는 상기 균주의 배양액을 포함하는 사료를 제공한다.According to another aspect of the present invention, the present invention provides a feed comprising the strain or a culture solution of the strain.
본 발명의 또 다른 양태에 따르면, 본 발명은 상기 균주로 발효하여 제조된 발효사료를 제공한다.According to another aspect of the present invention, the present invention provides a fermented feed prepared by fermenting with the strain.
본 발명의 균주는 항박테리아 및 항진균 활성, 다양한 탄수화물의 발효능, 다양한 효소 생산능, 항균제를 이용한 조절성, 내산성, 내담즙성, 응집성, 소수성, 항산화 활성 등 매우 우수한 프로바이오틱스의 특성을 갖는 신규 유산균이므로, 프로바이오틱스 생균제제, 또는 식품이나 사료에 포함시키거나 발효식품의 발효균으로 매우 유용하다.The strain of the present invention is a novel lactic acid bacteria having very excellent properties of probiotics such as antibacterial and antifungal activity, fermentation ability of various carbohydrates, various enzyme production ability, regulation using antibacterial agents, acid resistance, bile resistance, cohesiveness, hydrophobicity, antioxidant activity, etc. Therefore, it is very useful as a probiotic probiotic, or as a fermentation bacteria in foods or feeds or fermented foods.
도 1은 본 발명 균주(KCC-35)의 항박테리아 활성을 실험한 결과이다. KACC, KACC 91016 균주.
도 2는 본 발명 균주(KCC-35)의 항진균 활성을 실험한 결과이다. KACC, KACC 91016 균주; S.B, Scopulariopsis brevicaulis; A.C, Aspergillus clavatus; P.C, Penicillium chrysogenum; A.F, Aspergillus fumigatus; F.O, Fusarium oxisporum.
도 3은 본 발명 균주(KCC-35)의 다양한 pH에 대한 저항성을 실험한 결과이다. KACC, KACC 91016 균주.
도 4는 본 발명 균주(KCC-35)의 위액 환경에 대한 저항성을 실험한 결과이다. KACC, KACC 91016 균주.
도 5는 본 발명 균주(KCC-35)의 담즙염 내성을 실험한 결과이다. KACC, KACC 91016 균주.
도 6은 본 발명 균주(KCC-35)의 소수성을 실험한 결과이다. KACC, KACC 91016 균주.
도 7은 본 발명 균주(KCC-35)의 자동응집 활성을 실험한 결과이다. KACC, KACC 91016 균주.
도 8은 본 발명 균주(KCC-35)의 프리바이오틱스 사용성을 실험한 결과이다. KACC, KACC 91016 균주.
도 9는 본 발명 균주(KCC-35)의 콜레스테롤 동화성을 실험한 결과이다. 파란색 그래프, KCC-35; 붉은색 그래프, KACC 91016.
도 10은 본 발명 균주(KCC-35)의 과산화수소 라디칼 소거활성을 실험한 결과이다. KACC, KACC 91016 균주.
도 11은 본 발명 균주(KCC-35)의 DPPH 라디칼 소거활성을 실험한 결과이다. vit-c, 비타민C; KACC, KACC 91016 균주.
도 12는 본 발명 균주(KCC-35)의 저수분 IRG 사료의 발효능을 실험한 결과이다. KACC, KACC 91016 균주.
도 13은 본 발명 균주(KCC-35)의 저수분 옥수수 사료의 발효능을 실험한 결과이다. KACC, KACC 91016 균주.
도 14는 본 발명 균주(KCC-35)의 저수분 수수 사료의 발효능을 실험한 결과이다. KACC, KACC 91016 균주.1 is a result of testing the antibacterial activity of the strain of the present invention (KCC-35). KACC, KACC 91016 strain.
Figure 2 is a result of testing the antifungal activity of the strain of the present invention (KCC-35). KACC, KACC 91016 strain; SB, Scopulariopsis brevicaulis ; AC, Aspergillus clavatus ; PC, Penicillium chrysogenum ; AF, Aspergillus fumigatus ; FO, Fusarium oxisporum .
3 is a result of testing the resistance of the strain of the present invention (KCC-35) to various pH. KACC, KACC 91016 strain.
Figure 4 is a result of testing the resistance of the present strain (KCC-35) to the gastric juice environment. KACC, KACC 91016 strain.
5 is a result of testing the bile salt resistance of the strain of the present invention (KCC-35). KACC, KACC 91016 strain.
6 is a result of testing the hydrophobicity of the strain of the present invention (KCC-35). KACC, KACC 91016 strain.
7 is a result of testing the auto-aggregation activity of the strain of the present invention (KCC-35). KACC, KACC 91016 strain.
8 is a result of testing the usability of prebiotics of the present strain (KCC-35). KACC, KACC 91016 strain.
9 is a result of an experiment on cholesterol assimilation of the strain of the present invention (KCC-35). Blue graph, KCC-35; Red graph, KACC 91016.
10 is a result of an experiment of the hydrogen peroxide radical scavenging activity of the strain of the present invention (KCC-35). KACC, KACC 91016 strain.
11 is a result of an experiment of the DPPH radical scavenging activity of the strain of the present invention (KCC-35). vit-c, vitamin C; KACC, KACC 91016 strain.
12 is a result of the experiment of the fermentation ability of the low moisture IRG feed of the present invention strain (KCC-35). KACC, KACC 91016 strain.
13 is a result of experimentation of the fermentation ability of the low moisture corn feed of the present strain (KCC-35). KACC, KACC 91016 strain.
14 is a result of an experiment of the fermentation ability of the low moisture sorghum feed of the present invention strain (KCC-35). KACC, KACC 91016 strain.
본 발명의 신규 유산균인 락토바실러스 플란타럼(Lactobacillus plantarum) KCC-35 균주는 알팔파(Alfalfa, Medicago sativa) 사일리지에서 분리되었으며, 농촌진흥청 국립농업과학원 미생물은행(KACC: Korean Agricultural Culture Collection)에 미생물 수탁번호 KACC 92222P로 기탁되어 있다.The novel lactobacillus plantarum of the present invention, Lactobacillus plantarum , KCC-35 strain was isolated from alfalfa ( Medicago sativa ) silage, and a microorganism was entrusted to the Korean Agricultural Culture Collection (KACC) of the Rural Development Administration. It is deposited under the number KACC 92222P.
본 발명에서 확인된 본 발명 균주의 16S rRNA 유전자의 일부 염기서열은 서열번호 1과 같으며, NCBI Genbank에 KP091751.1(accession number)로 기탁되어 있다. 이 염기서열을 통한 계통 분류결과를 바탕으로 본 발명의 균주는 락토바실러스 플란타럼에 속하는 새로운 균주로 분류될 수 있다.Some nucleotide sequences of the 16S rRNA gene of the strain of the present invention identified in the present invention are the same as SEQ ID NO: 1, and are deposited as KP091751.1 (accession number) in NCBI Genbank. The strain of the present invention can be classified as a new strain belonging to Lactobacillus plantarum based on the result of phylogenetic classification through this nucleotide sequence.
본 발명의 균주는 Escherichia coli, Enterococcus faecalis, Staphylococcus aureus 및 Pseudomonas aeruginosa에 대해 항박테리아 활성을 나타내며, Scopulariopsis brevicaulis, Aspergillus clavatus, Penicillium chrysogenum, Aspergillus fumigatus, Fusarium oxisporum에 대해 항진균 활성을 나타낼 수 있다.The strain of the present invention exhibits antibacterial activity against Escherichia coli , Enterococcus faecalis , Staphylococcus aureus, and Pseudomonas aeruginosa , and can exhibit antibacterial activity against Scopulariopsis brevicaulis , Aspergillus clavatus , Penicillium chrysogenum , Aspergillus fumigatus , and Fusarium oxisporum .
본 발명의 균주는 자일로스(xylose), 글루코스, 만노스(mannose) 등의 탄수화물(표 1 참조)을 발효할 수 있다. 따라서 이들 탄소원이 함유된 재료를 발효하는데 이용할 수 있다.The strain of the present invention can ferment carbohydrates (see Table 1) such as xylose, glucose, and mannose. Therefore, it can be used to ferment the material containing these carbon sources.
본 발명의 균주는 α-갈락토시다제(α-galactosidase), β-글루코시다제(β-glucosidase) 등의 효소를 생산할 수 있다. 따라서 이들 효소의 활성이 필요한 분야에 사용될 수 있다.The strain of the present invention can produce enzymes such as α-galactosidase and β-glucosidase. Therefore, it can be used in fields requiring the activity of these enzymes.
본 발명의 균주는 클로람페니콜(chloramphenicol), 니트로푸란토인(nitrofurantoin), 스트렙토마이신(streptomycin) 등의 항생제에 감수성을 나타내므로 이들 항생제를 이용하여 생장을 조절할 수 있으며, 카나마이신(kanamycin), 테트라사이클린(tetracycline) 등의 항생제에 저항성을 나타낼 수 있다.Since the strain of the present invention is sensitive to antibiotics such as chloramphenicol, nitrofurantoin, and streptomycin, growth can be controlled using these antibiotics, kanamycin, tetracycline, etc. ), and other antibiotics.
본 발명의 균주는 pH 2 ~ 3의 위액 환경에서도 생존할 수 있고 담즙염이 높은 농도로 존재하는 조건에서도 생존할 수 있어, 동물의 위나 소장 등의 소화기관을 지나는 동안 생존하여 장까지 살아있는 채로 도달할 가능성이 높다.The strain of the present invention can survive in a gastric environment of
본 발명의 균주는 우수한 응집력 및 소수성을 나타내므로 동물의 장내에 정착하는데 유리하다.Since the strain of the present invention exhibits excellent cohesiveness and hydrophobicity, it is advantageous to settle in the intestine of animals.
본 발명의 균주는 라피노스, 이눌린과 같은 프리바이오틱스를 이용할 수 있다.The strain of the present invention may use prebiotics such as raffinose and inulin.
본 발명의 균주는 콜레스테롤 동화성 및 항산화 활성이 우수하여 프로바이오틱스로서 유용하게 활용할 수 있다.The strain of the present invention is excellent in cholesterol assimilation and antioxidant activity and can be usefully utilized as probiotics.
본 발명의 균주는 수분이 낮은 조건에서도 여러 가지 사료작물을 발효할 수 있어 발효사료의 제조를 위한 유산균으로 유용하게 활용할 수 있다.The strain of the present invention can ferment various feed crops even under low moisture conditions, and thus can be effectively utilized as lactic acid bacteria for the production of fermented feed.
상기와 같이 본 발명의 균주는 프로바이오틱스로서의 매우 우수한 특성을 가지므로, 프로바이오틱스 생균제제로 이용할 수 있다.As described above, the strain of the present invention has very excellent properties as probiotics, and thus can be used as probiotic probiotics.
이때 생균제제는 본 발명의 균주 자체 또는 균주의 배양물로 이루어질 수 있으며, 제제화의 용이성을 위한 부형제 등의 첨가제가 포함된 형태일 수도 있다.At this time, the probiotic preparation may be made of the strain itself or a culture of the strain of the present invention, and may be in a form containing an additive such as an excipient for ease of formulation.
또한 본 발명의 균주 또는 이의 배양물은 식품이나 사료의 재료 또는 첨가제로 이용될 수 있다. 이때 배양물은 균체가 포함되지 않은 형태로도 이용할 수 있으나, 프로바이오틱스로서의 효과를 기대하기 위해서는 균체가 포함된 형태로 이용하는 것이 바람직하다.In addition, the strain of the present invention or a culture thereof may be used as a material or additive for food or feed. At this time, the culture may be used in a form that does not contain the cells, but in order to expect the effect as a probiotic, it is preferable to use the culture in a form containing the cells.
또한 본 발명의 균주는 발효식품 또는 발효사료의 발효균주로도 이용할 수 있다. 이때 발효식품 또는 발효사료에는 표 1에서와 같이 본 발명의 균주가 발효할 수 있는 성분이 함유된 원료를 사용하는 것이 바람직하다.In addition, the strain of the present invention can also be used as a fermented strain of fermented food or fermented feed. At this time, it is preferable to use a raw material containing a fermentable ingredient by the strain of the present invention as shown in Table 1 for fermented food or fermented feed.
이하, 실시예를 통하여 본 발명을 더욱 상세히 설명하기로 한다. 이들 실시예는 단지 본 발명을 예시하기 위한 것이므로, 본 발명의 범위가 이들 실시예에 의해 제한되는 것으로 해석되지는 않는다.Hereinafter, the present invention will be described in more detail through examples. Since these examples are for illustrative purposes only, the scope of the present invention is not to be construed as being limited by these examples.
[실시예][Example]
1. 방법1. Method
1-1. 균주의 분리 및 예비 스크리닝1-1. Isolation and preliminary screening of strains
대한민국 천안에 위치한 국립축산과학원으로부터 발효된 알팔파(Alfalfa, Medicago sativa) 사일리지(silage) 샘플을 수집하였다. 1g의 사일리지 샘플을 멸균수로 연속희석(107)하고, Ilavenil 등(J Sci Food Agric. 96(2):593-601)과 같은 방법으로 균주 분리를 시도하였다.Alfalfa ( Medicago sativa ) fermented from the National Institute of Livestock Science in Cheonan, Korea, silage samples were collected. 1 g of the silage sample was serially diluted (10 7 ) with sterile water, and strain isolation was attempted in the same manner as Ilavenil et al. ( J Sci Food Agric. 96(2):593-601).
성장 프로파일 분석 및 항진균 실험결과를 바탕으로, 효과적인 균주를 추가 특성분석을 위해 선별하고 이를 KCC-35로 명명하였다. 균주를 하룻밤 배양하여 생화학 및 생리학적 특성 분석을 위해 사용하였다.Based on the growth profile analysis and antifungal test results, an effective strain was selected for further characterization and named KCC-35. The strain was cultured overnight and used for biochemical and physiological characterization.
Megazyme 어세이 키트(Bray, Co. WIcklow, Ireland)를 발효산의 정량에 사용하였고, API 50 CH 및 API-ZYM 키트(Marcy-I' Etoile, France)를 탄수화물 발효 및 효소 생산 분석에 사용하였다. 항생제 민감성 스크리닝은 Valan Arasu 등(J Appl Microbiol. 115(5):1172-1185)의 디스크 확산법을 적용하였다.Megazyme assay kit (Bray, Co. WIcklow, Ireland) was used for quantification of fermented acid, and
본 실험 및 이하의 실험에서 비교균주로는 대한민국 등록특허 제10-0496022호의 KACC 91016(락토바실러스 플랜타럼 NLRI 201)을 사용하였다.In this experiment and the following experiments, KACC 91016 (Lactobacillus plantarum NLRI 201) of Korean Patent Registration No. 10-0496022 was used as a comparative strain.
1-2. 분자적 특성 분석 및 기탁1-2. Molecular characterization and deposit
16s rRNA 유전자 서열분석을 Solgent 사(서울, 한국)를 통해 Sanger의 방법에 따라 수행하였다. KCC-35 균주의 게놈 DNA를 추출하고, QIAquick® 키트(Qiagen Ltd., Crawley, UK)로 정제하였으며, 유니버셜 프라이머 27F(5'-AGA GTT TGA TCG TGG CTC AG-3') 및 1492R(3'-GCT TAC CTT GTT ACG ACT T-5')을 사용하여 증폭산물의 서열을 분석하였다. NCBI gene bank의 데이터베이스에서 BLAST를 사용하여 KCC-35의 16s rRNA 서열을 얼라인 분석하였으며, 이 서열을 NCBI Genebank에 기탁하였다.16s rRNA gene sequencing was performed by Solgent (Seoul, Korea) according to Sanger's method. Genomic DNA of the KCC-35 strain was extracted and purified with QIAquick® kit (Qiagen Ltd., Crawley, UK), and universal primers 27F (5'-AGA GTT TGA TCG TGG CTC AG-3') and 1492R (3' -GCT TAC CTT GTT ACG ACT T-5') was used to analyze the sequence of the amplification product. The 16s rRNA sequence of KCC-35 was aligned and analyzed using BLAST in the database of the NCBI gene bank, and this sequence was deposited in the NCBI Genebank.
1-3. 항진균 활성 분석1-3. Antifungal activity assay
Fusarium Oxysporum, Aspergillus clavatus, Penicillium chrysogenum, Scopulariopsis brevicaulis 및 Aspergillus Flavus을 KACC(전주, 한국)로부터 입수하고, 약간의 변형이 수반된 Ilavenil 등(J Sci Food Agric. 96(2):593-601)의 pour-plate 방법으로 항진균 활성을 스크리닝하였다. KCC-35 균주에 의한 진균의 성장 저해 및 진균 바이오매스는 Valan Arasu 등(J Appl Microbiol. 115(5):1172-1185)의 방법으로 분석하였다. Fusarium Oxysporum, Aspergillus clavatus, Penicillium chrysogenum, Scopulariopsis brevicaulis and Aspergillus Flavus were obtained from KACC (Jeonju, Korea), and poured by Ilavenil et al. ( J Sci Food Agric. 96(2):593-601) with slight modifications . Antifungal activity was screened by the -plate method. Growth inhibition and fungal biomass by the KCC-35 strain were analyzed by the method of Valan Arasu et al. ( J Appl Microbiol. 115(5):1172-1185).
1-4. 항박테리아 활성 분석1-4. Antibacterial activity assay
Ilavenil 등(J Sci Food Agric. 96(2):593-601) 및 Gaudana 등(Br J Nutr. 103:1620-1628)의 아가 스팟 테스트(agar spot test)를 사용하여 KCC-35 균주의 항박테리아 활성을 측정하였다.Antibacterial of KCC-35 strain using the agar spot test of Ilavenil et al. ( J Sci Food Agric. 96(2):593-601) and Gaudana et al. ( Br J Nutr. 103:1620-1628) The activity was measured.
KCC-35 균주의 24시간 배양액(5㎕)을 MRS 배지 상에 접종하고, 37℃에서 24시간 배양하였다. 이후 병원균의 하룻밤 배양액을 각 소프트 아가(tryptone soy 및 brain heart infusion)와 혼합하고, KCC-35의 24시간 배양된 콜로니가 함유된 MRS 한천배지에 부은 다음 37℃에서 24시간 배양하여 저해구역의 반지름을 측정하였다.A 24-hour culture solution (5 µl) of the KCC-35 strain was inoculated onto the MRS medium and cultured at 37°C for 24 hours. After that, the overnight culture of the pathogen was mixed with each soft agar (tryptone soy and brain heart infusion), poured on MRS agar medium containing colonies cultured for 24 hours of KCC-35, and incubated at 37°C for 24 hours to determine the radius of the inhibition zone. Was measured.
1-5. 항진균 활성의 생리학적 요인 분석1-5. Analysis of physiological factors of antifungal activity
KCC-35 균주의 항진균 활성 상에서 온도 및 pH에 따른 효과를 조사하였다. 균주를 MRS 액체배지에서 서로 다른 온도 및 pH 상에서 배양하고, 배양 상층액의 항진균 활성을 Valan Arasu 등(J Appl Microbiol. 115(5):1172-1185)의 배양액 마이크로희석법으로 조사하였다.The effect of temperature and pH on the antifungal activity of the KCC-35 strain was investigated. The strains were cultured in MRS liquid medium at different temperatures and pH, and the antifungal activity of the culture supernatant was investigated by the microdilution method of the culture solution of Valan Arasu et al. ( J Appl Microbiol. 115(5):1172-1185).
1-6. pH 환경에 대한 저항성 분석1-6. Analysis of resistance to pH environments
Ilavenil 등(J Sci Food Agric. 96:593-601)의 방법에 따라 KCC-35 균주의 낮은 pH에 대한 내성을 조사하였다. 배양된 균주를 다양한 pH의 MRS 배양액에 접종하고, 37℃에서 48시간 배양하였다. 배양 후 각 배양액 100㎕를 pour plate 방법으로 MRS 배지에 접종하고, 37℃에서 48시간 배양하여 콜로니를 계수하였다.The resistance of the KCC-35 strain to low pH was investigated according to the method of Ilavenil et al. ( J Sci Food Agric. 96:593-601). The cultured strains were inoculated into MRS culture solutions of various pH and cultured at 37°C for 48 hours. After cultivation, 100 µl of each culture solution was inoculated into the MRS medium by a pour plate method, and cultured at 37° C. for 48 hours to count colonies.
1-7. 위의 pH에 대한 내성 분석1-7. Analysis of tolerance to gastric pH
Charteris 등(J Appl Microbiol. 84:759-768)의 방법에 따라 KCC-35 균주의 위액에 대한 저항성을 조사하였다. 펩신(pepsin, 3㎎/㎖), 0.5%(w/v) NaCl 및 물을 사용하여 위액을 준비하고, 1mol/L HCl을 사용하여 pH를 약 2 및 3으로 맞추었다. 30㎖의 균주 배양액을 4℃에서 6000xg로 20분간 원심분리하고, 펠렛을 수득하여 K2HPO4(50mmol/L, 6.5pH)로 3회 세척하였다. 펠렛을 재현탁한 다음 1㎖의 현탁액을 9㎖의 위액과 혼합하고 37℃에서 3시간 배양하였다. 배양전 및 배양후의 총 생균을 계수하였다.The resistance of the KCC-35 strain to gastric juice was investigated according to the method of Charteris et al. ( J Appl Microbiol. 84:759-768). Gastric juice was prepared using pepsin (3 mg/ml), 0.5% (w/v) NaCl and water, and the pH was adjusted to about 2 and 3 using 1 mol/L HCl. 30 ml of the strain culture solution was centrifuged at 6000xg at 4° C. for 20 minutes, and a pellet was obtained and washed three times with K 2 HPO 4 (50 mmol/L, 6.5 pH). After resuspending the pellet, 1 ml of the suspension was mixed with 9 ml of gastric juice and incubated at 37°C for 3 hours. Total viable cells before and after culture were counted.
1-8. 농축된 담즙염 환경에 대한 내성 분석1-8. Analysis of tolerance to a concentrated bile salt environment
KCC-35 균주의 배양액을 0.5% 소듐 디옥시콜레이트 및 0.3% 옥스갈(DCA Sigma, St Louis, MO, USA)과 같은 담즙염이 함유된 MRS 액체배지에 접종하고 37℃에서 배양하였다. 일정 시간 간격(24시간 및 48시간 후)으로 배양액을 분취하고 600nm에서 흡광도를 측정하였다. 담즙염을 첨가하지 않은 MRS 배지를 대조군으로 사용하였다.The culture medium of the KCC-35 strain was inoculated into MRS liquid medium containing bile salts such as 0.5% sodium deoxycholate and 0.3% oxgal (DCA Sigma, St Louis, MO, USA) and cultured at 37°C. The culture solution was aliquoted at regular time intervals (after 24 hours and 48 hours) and absorbance was measured at 600 nm. MRS medium to which bile salt was not added was used as a control.
1-9. 콜로니 형성능(자동응집) 스크리닝1-9. Colony formation ability (automatic aggregation) screening
Del Re 등(Letters in Applied Microbiology, 31:438-442)의 방법으로 표면 상호작용을 통한 세포막의 응집을 조사하였다. 균주 배양액을 8000xg로 10분간 원심분리하고, 펠렛을 108 CFU/㎖로 PBS(pH 7)에 재현탁하였다. 세포 현탁액 4㎖을 10초간 볼텍싱하고 상온에서 3시간 배양하여 자동응집을 조사하였다. 각 시간에 상층의 세포현탁액 100㎕을 PBS 3.9㎖과 혼합하고 600nm에서 흡광도를 측정하였다. 자동응집은 다음의 계산식으로 계산하였다.The aggregation of cell membranes through surface interaction was investigated by the method of Del Re et al ( Letters in Applied Microbiology , 31:438-442). The strain culture was centrifuged at 8000xg for 10 minutes, and the pellet was resuspended in PBS (pH 7) at 10 8 CFU/ml. 4 ml of the cell suspension was vortexed for 10 seconds and incubated for 3 hours at room temperature to investigate auto-aggregation. At each time, 100 µl of the cell suspension in the upper layer was mixed with 3.9 ml of PBS, and absorbance was measured at 600 nm. Auto-aggregation was calculated by the following calculation formula.
[계산식][formula]
1-(At/A0) x 1001-(A t /A 0 ) x 100
At : 각 배양시간(1, 2, 3시간)에 따른 흡광도A t : absorbance according to each culture time (1, 2, 3 hours)
A0 : 0시간의 흡광도A 0 : absorbance at 0 hours
1-10. 접착력 스크리닝1-10. Adhesion screening
Rosenberg 등(FEMS Microbiology Letters, 9:29-33)의 방법에 따라 KCC-35 균주의 소수성을 조사하였다. 세포표면 소수성은 인산완충액으로부터 탄화수소에 결합하는 미생물의 능력을 부여한다. 1일간 배양한 균주배양액을 5000xg로 15분간 원심분리하고, 세포를 수집하여 PBS(pH 7)로 세척하였다. 흡광도는 540nm에서 측정하였다. 세포현탁액 1㎖를 동부피의 탄화수소(자일렌 및 클로로포름)와 혼합하고, 540nm에서 흡광도를 측정하였다. 30초 후 수성용액의 흡광도(ODfinal)를 측정하고 초기값(ODinitial)과 비교하였다. 소수성은 다음 계산식으로 계산하였다.The hydrophobicity of the KCC-35 strain was investigated according to the method of Rosenberg et al. ( FEMS Microbiology Letters , 9:29-33). Cell surface hydrophobicity imparts the ability of microorganisms to bind to hydrocarbons from the phosphate buffer. The strain culture solution cultured for 1 day was centrifuged at 5000xg for 15 minutes, and the cells were collected and washed with PBS (pH 7). The absorbance was measured at 540 nm. 1 ml of the cell suspension was mixed with an eastern coat of hydrocarbons (xylene and chloroform), and absorbance was measured at 540 nm. After 30 seconds, the absorbance (OD final ) of the aqueous solution was measured and compared with the initial value (OD initial ). Hydrophobicity was calculated by the following formula.
소수성% = (ODinitial - ODfinal) / ODinitial x 100Hydrophobicity% = (OD initial -OD final ) / OD initial x 100
1-11. 콜레스테롤 동화성 분석1-11. Cholesterol assimilation analysis
Ilavenil 등(J Sci Food Agric, 96:593-601)의 방법에 따라 KCC-35 균주의 콜레스테롤 동화성을 조사하였다. 0.3% 옥스갈 및 100㎍/㎖의 수용성 콜레스테롤을 함유하는 MRS 액체배지를 준비하고, 3개의 50㎖ 튜브에 나누어 담은 다음 각각 KCC-35 균주를 접종하였다. 접종한 배양액을 37℃에서 6, 12, 24시간 배양한 다음, 튜브를 10000xg로 15분간 원심분리하고, 상등액에 콜레스테롤의 존재여부를 조사하였다.Cholesterol assimilation of the KCC-35 strain was investigated according to the method of Ilavenil et al. ( J Sci Food Agric , 96:593-601). An MRS liquid medium containing 0.3% oxgal and 100 μg/ml of water-soluble cholesterol was prepared, divided into three 50 ml tubes, and inoculated with KCC-35 strains, respectively. The inoculated culture was incubated at 37°C for 6, 12, and 24 hours, then the tube was centrifuged at 10000xg for 15 minutes, and the presence of cholesterol in the supernatant was examined.
1-12. 프리바이오틱스 사용성 분석1-12. Prebiotics usability analysis
본 실험은 Ilavenil 등(J Sci Food Agric, 96:593-601)의 방법을 사용하여 수행하였다. 본 실험에서 MRS 액체배지에는 2% 라피노스(raffinose), 글루코스(glucose) 및 이눌린(inulin)이 첨가되었다. 새롭게 배양된 KCC-35 균주 배양액을 프리바이오틱스가 첨가된 액체배지에 첨가하고 37℃에서 24시간 배양하였다. 글루코스를 첨가한 MRS 액체배지를 대조군으로 사용하였다. 최종 흡광도는 600nm에서 측정하였다. 균주에 의한 프리바이오틱스의 사용성은 계산식 [((MRSp - MRSb) x 100) / MRSg - MRSb)] 로 계산하였다. 이때 MRSp는 라피노스 및 이눌린이 처리된 실험군의 흡광도이고, MRSb는 탄소원이 없는 실험군의 흡광도(음성대조군)이며, MRSg는 글루코스를 처리한 실험군의 흡광도(양성대조군)이다.This experiment was performed using the method of Ilavenil et al. ( J Sci Food Agric , 96:593-601). In this experiment, 2% raffinose, glucose, and inulin were added to the MRS liquid medium. The newly cultured KCC-35 strain culture was added to the liquid medium to which prebiotics were added, and cultured at 37°C for 24 hours. MRS liquid medium to which glucose was added was used as a control. The final absorbance was measured at 600 nm. The usability of prebiotics by strain was calculated by the calculation formula [((MRSp-MRSb) x 100) / MRSg-MRSb)]. At this time, MRSp is the absorbance of the experimental group treated with raffinose and inulin, MRSb is the absorbance of the experimental group without a carbon source (negative control), and MRSg is the absorbance of the experimental group treated with glucose (positive control).
1-13. DPPH법에 의한 자유라디칼 소거력 분석1-13. Analysis of free radical scavenging force by DPPH method
Li 등(Food Chemistry, 135:1914-1919)의 방법에 약간의 변경을 적용하여 KCC-35 균주의 DPPH(2,2-Diphenyl-1-picrylhydrazyl) 라디칼 소거활성을 측정하였다. The DPPH (2,2-Diphenyl-1-picrylhydrazyl) radical scavenging activity of the KCC-35 strain was measured by applying a slight change to the method of Li et al. ( Food Chemistry , 135:1914-1919).
다양한 농도(10, 20, 30, 40 및 50㎍)의 균주 배양액(109CFU/㎖)을 1㎖의 DPPH 용액(0.05mM)과 혼합하고, 상온에서 30분간 암실에서 배양하였다. DPPH 용액을 대조군(control)으로 사용하였다. 메탄올과 균주세포의 혼합액은 Blank로 사용하였다. 흡광도는 517nm에서 측정하였다. DPPH 라디칼 소거활성은 다음 계산식으로 계산하였다.Various concentrations (10, 20, 30, 40 and 50 ㎍) of the strain culture solution (10 9 CFU/ml) was mixed with 1 ml of DPPH solution (0.05 mM), and incubated in the dark for 30 minutes at room temperature. DPPH solution was used as a control. A mixture of methanol and strain cells was used as a blank. The absorbance was measured at 517 nm. DPPH radical scavenging activity was calculated by the following formula.
DPPH 소거활성(%) = [(Asample - Ablank) / Acontrol] x 100DPPH scavenging activity (%) = [(A sample -A blank ) / A control ] x 100
1-14. 과산화수소 라디칼 소거력 분석1-14. Hydrogen peroxide radical scavenging ability analysis
Li 등(Food Chemistry, 135:1914-1919)의 방법에 따라 과산화수소 저항성을 조사하였다. 균주 배양액 10㎖을 원심분리하고 펠렛을 수집한 다음 각기 다른 농도(0.5, 1.0 및 1.5mM)의 H2O2가 함유된 10㎖의 0.9% NaCl과 혼합하였다. 각 샘플을 희석하고 BCP 한천플레이트를 사용하여 초기 및 37℃ 배양 30분 후의 균체의 생존력을 측정하였다. 실험은 3반복 수행하였다.Hydrogen peroxide resistance was investigated according to the method of Li et al. ( Food Chemistry , 135:1914-1919). 10 ml of the strain culture solution was centrifuged and the pellet was collected, and then mixed with 10 ml of 0.9% NaCl containing H 2 O 2 at different concentrations (0.5, 1.0 and 1.5 mM). Each sample was diluted and the viability of the cells was measured initially and after 30 minutes of incubation at 37° C. using a BCP agar plate. The experiment was performed in triplicate.
1-15. 저수분 사료분말 상의 적용1-15. Application on low moisture feed powder
이탈리안라이그라스(IRG), 옥수수 및 수수 분말을 NIAS로부터 입수하여 에틸렌 가스로 멸균하였다. 멸균조건에서 상온 건조하여 상기 분말의 수분함량을 30%로 고정시켰다. 각 분말을 비접종 및 KCC-35 접종그룹으로 나누었다. 비접종 그룹은 상기 분말만을 발효분석에 사용하였고, 접종그룹은 1x10-4/250g의 KCC-35를 나누어 첨가하였다. 각 그룹당 3개의 샘플을 준비하고 공기흐름이 차단되도록 단단히 밀봉하였다. 상기 샘플들을 혐기조건에서 45일간 보관하고, 발효된 사료분말의 미생물수 및 pH를 조사하였다.Italian ryegrass (IRG), corn and sorghum powder were obtained from NIAS and sterilized with ethylene gas. It was dried at room temperature under sterilization conditions to fix the moisture content of the powder at 30%. Each powder was divided into non-vaccinated and KCC-35 inoculated groups. In the non-inoculated group, only the powder was used for fermentation analysis, and in the inoculated group, 1×10 -4 /250 g of KCC-35 was divided and added. Three samples were prepared for each group and tightly sealed to block airflow. The samples were stored for 45 days under anaerobic conditions, and the number of microorganisms and pH of the fermented feed powder were examined.
1-16. 통계 분석1-16. Statistical analysis
모든 수치데이터는 3회의 독립적인 실험으로부터 얻었고, 이들 데이터의 분석은 SPSS 소프트웨어(SPSS-16 Inc., Chicago, IL, USA)를 사용하여 수행하였다. 결과는 평균±표준편차로 나타내었다. 평균값들 사이의 유의적인 차이는 P < 0.05를 유의성 레벨로 하는 다중분석을 사용하여 분석하였다.All numerical data were obtained from three independent experiments, and analysis of these data was performed using SPSS software (SPSS-16 Inc., Chicago, IL, USA). Results are expressed as mean±standard deviation. Significant differences between the mean values were analyzed using a multiplex analysis with P <0.05 as the significance level.
2. 결과2. Results
2-1. 분리 및 스크리닝2-1. Separation and screening
뚜렷한 형태학적 특성을 바탕으로 KCC-35 균주를 분리하였다. 기본 확인 실험을 통해 분리된 균주가 그람양성, 비운동성, 원형, 카탈라아제 음성 및 중온성임을 확인하였다. 이를 바탕으로 분자적 특성, 항진균 및 프로바이오틱스 잠재력에 대해 추가로 실험하였다. 16S rRNA 서열 분석을 통해 KCC-35의 분자적 특성을 분석한 결과, Lactobacillus plantarum에 속하는 것으로 나타났다. 이 16S rRNA 서열은 NCBI 유전자 은행 데이터베이스에 KP091751.1로 기탁되었다.KCC-35 strain was isolated based on distinct morphological characteristics. Through the basic confirmation experiment, it was confirmed that the isolated strains were Gram-positive, non-motile, circular, catalase-negative and mesophilic. Based on this, the molecular properties, antifungal and probiotic potential were further tested. As a result of analyzing the molecular characteristics of KCC-35 through 16S rRNA sequence analysis, it was found to belong to Lactobacillus plantarum . This 16S rRNA sequence was deposited as KP091751.1 in the NCBI Gene Bank database.
2-2. 생화학적 특성 및 항생제 감수성2-2. Biochemical properties and antibiotic sensitivity
KCC-35 균주는 자일로스(xylose), 글루코스, 만노스(mannose) 등을 발효하며(표 1 참조), α-갈락토시다제(α-galactosidase), β-글루코시다제(β-glucosidase)와 같은 효소를 생산하는 것으로 나타났다(표 2 참조). 또한 KCC-35 균주는 클로람페니콜(chloramphenicol), 니트로푸란토인(nitrofurantoin), 스트렙토마이신(streptomycin) 등의 항생제에 감수성을 나타내고, 카나마이신(kanamycin), 테트라사이클린(tetracycline) 등의 항생제에 저항성을 나타내었다(표 3 참조).The KCC-35 strain fermented xylose, glucose, mannose, etc. (see Table 1), α-galactosidase, β-glucosidase, and It was found to produce the same enzyme (see Table 2). In addition, the KCC-35 strain showed sensitivity to antibiotics such as chloramphenicol, nitrofurantoin, and streptomycin, and resistance to antibiotics such as kanamycin and tetracycline ( See Table 3).
2-3. 항박테리아 활성2-3. Antibacterial activity
식품매개 병원균에 대한 항박테리아 활성을 조사한 결과, KCC-35는 비교균주인 KACC 91016 균주에 비해 우수한 항박테리아 활성이 있는 것으로 나타났다(도 1 참조).As a result of examining the antibacterial activity against food-borne pathogens, KCC-35 was found to have superior antibacterial activity compared to the comparative strain, KACC 91016 (see FIG. 1).
2-4. 항진균 활성2-4. Antifungal activity
진균에 대한 항진균 활성을 조사한 결과, KCC-35는 비교균주인 KACC 91016 균주에 비해 우수한 항진균 활성이 있는 것으로 나타났다(도 2 참조). 또한, pH 및 온도에 따른 KCC-35의 항진균 활성을 조사한 결과에서도 우수한 항진균 활성이 있는 것으로 나타났다(표 4 및 5 참조).As a result of examining the antifungal activity against fungi, it was found that KCC-35 has superior antifungal activity compared to the comparative strain, KACC 91016 (see FIG. 2). In addition, the results of investigation of the antifungal activity of KCC-35 according to pH and temperature showed excellent antifungal activity (see Tables 4 and 5).
2-5. 산성조건 및 담즙염 내성2-5. Acidic conditions and bile salt resistance
KCC-35의 다양한 pH 환경에 대한 내성을 조사한 결과, 산성 환경이 될수록 콜로니의 성장이 감소하기는 하지만 우수한 산성조건 내성을 나타내었다. 특히, 비교균주인 KACC 91016 균주에 비해 산성조건에 대한 내성이 우수한 것으로 나타났다(도 3 참조).As a result of investigating the resistance of KCC-35 to various pH environments, it showed excellent resistance to acidic conditions, although the growth of colonies decreased with increasing acidity. In particular, it was found to have excellent resistance to acidic conditions compared to the comparative strain, KACC 91016 (see FIG. 3).
또한, pH2 및 pH3의 위액 환경에서도 우수한 내성을 나타내었으며(도 4 참조), 0.3% oxgall 및 0.5% sodium deoxycholate의 높은 농도의 담즙염 환경에서도 우수한 내성을 나타내었다(도 5 참조).In addition, it exhibited excellent tolerance in gastric fluid environments of pH2 and pH3 (see FIG. 4), and exhibited excellent tolerance in bile salt environments of high concentrations of 0.3% oxgall and 0.5% sodium deoxycholate (see FIG. 5).
2-6. 세포응집 및 소수성2-6. Cell aggregation and hydrophobicity
KCC-35의 세포응집은 시간에 따라 증가하며, 비교균주인 KACC 91016 균주에 비해 응집력이 더 우수한 것으로 나타났다(도 7 참조).Cell aggregation of KCC-35 increased with time, and it was found that the cohesive power was better than that of the comparative strain, KACC 91016 (see FIG. 7).
소수성 또한 비교균주인 KACC 91016 균주에 비해 높고, 특히 자일렌에 비해 클로로포름에 대해 높은 소수성을 나타냈다(도 6 참조).Hydrophobicity was also higher than that of the comparative strain, KACC 91016, and particularly, showed higher hydrophobicity to chloroform than xylene (see FIG. 6).
2-7. 프리바이오틱 이용성 및 콜레스테롤 동화성2-7. Prebiotic availability and cholesterol assimilation
라피노스 및 이눌린을 사용하여 프리바이오틱 이용성을 조사한 결과, KCC-35는 비교균주인 KACC 91016 균주에 비해 프리바이오틱 이용성이 우수하고, 이눌린에 비해 라피노스를 더 잘 이용하는 것으로 나타났다(도 8 참조).As a result of examining the prebiotic availability using raffinose and inulin, it was found that KCC-35 has superior prebiotic availability compared to the comparative strain, KACC 91016, and uses raffinose better than inulin (see FIG. 8).
또한, 비교균주인 KACC 91016 균주에 비해 콜레스테롤 동화성도 우수한 것으로 나타났다(도 9 참조).In addition, it was found that cholesterol assimilation was excellent compared to the comparative strain, KACC 91016 (see FIG. 9).
2-8. 항산화 효과2-8. Antioxidative effect
과산화수소 라디칼 및 DPPH 라디칼 소거활성 분석 방법을 이용하여 자유라디칼 소거활성을 조사한 결과, KCC-35는 비교균주인 KACC 91016 균주에 비해 자유라디칼 소거활성이 우수한 것으로 나타났다(도 10 및 11 참조).As a result of investigating the free radical scavenging activity using hydrogen peroxide radical and DPPH radical scavenging activity analysis method, KCC-35 was found to have superior free radical scavenging activity compared to the comparative strain KACC 91016 (see FIGS. 10 and 11).
2-9. 저수분 사료의 발효성2-9. Fermentability of low moisture feed
IRG, 옥수수 및 수수 분말을 사용하여 혐기조건에서 저수분 사료의 발효성을 조사한 결과, 저수분 사료에 KCC-35를 접종하였을 때 콜로니 생성이 증가하고, 발효로 인해 pH가 낮아진 것으로 나타났다(도 12 내지 14 참조). 이는 KCC-35가 저수분 사료를 효율적으로 발효할 수 있음을 의미한다.As a result of investigating the fermentability of low moisture feed in anaerobic conditions using IRG, corn and sorghum powder, colony formation increased when KCC-35 was inoculated in low moisture feed, and pH decreased due to fermentation (FIG. 12) To 14). This means that KCC-35 can efficiently ferment low moisture feed.
+ : 양성반응, - : 음성반응, KACC : KACC 91016 균주+: Positive reaction,-: negative reaction, KACC: KACC 91016 strain
+ : 약한 생산, ++ : 보통의 생산, +++ : 강한 생산, KACC : KACC 91016 균주+: weak production, ++: moderate production, +++: strong production, KACC: KACC 91016 strain
S : 감수성(>10mm), R : 저항성, KACC : KACC 91016 균주S: susceptibility (>10mm), R: resistance, KACC: KACC 91016 strain
KACC : KACC 91016 균주KACC: KACC 91016 strain
KACC : KACC 91016 균주KACC: KACC 91016 strain
<110> REPUBLIC OF KOREA(MANAGEMENT : RURAL DEVELOPMENT ADMINISTRATION) <120> Novel Lactobacillus plantarum KCC-35 <130> PA-D18243 <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1215 <212> DNA <213> Unknown <220> <223> Lactobacillus plantarum KCC-35 <400> 1 ggggtgtggc ggccgccaat agatgcagtc gaacgaactc tggtattgat tggtgcttgc 60 atcatgattt acatttgagt gagtggcgaa ctggtgagta acacgtggga aacctgccca 120 gaagcggggg ataacacctg gaaacagatg ctaataccgc ataacaactt ggaccgcatg 180 gtccgagctt gaaagatggc ttcggctatc acttttggat ggtcccgcgg cgtattagct 240 agatggtggg gtaaccggtc tcacccattg gcaatgatat cgtagccgac ctgagagggt 300 aatcggccac attgggactg agacacggcc caaactccta cgggaggcag cagtagggaa 360 tcttccacaa tggacgaaag tctgatggag caacgcccgc gtgagtgaag aagggtttcg 420 gctcgtaaaa ctctgttgtt aaagaagaac atatctgaga gtaactgttc aggtattgac 480 ggtatttaac ccagaaagcc acggctaact acgtgccagc agccgcggta atacgtaggt 540 ggcaagcgtt gtccggattt attgggcgta aagcgagcgc aggcggtttt ttaagtctga 600 tgtgaaagcc ttcggctcaa ccgaagaagt gcatcggaaa ctgggaaact tgagtgcaga 660 agaggacagt ggaactccat gtgtagcggt gaaatgcgta gatatatgga agaacaccag 720 tggcgaaggc ggctgtctgg tctgtaactg acgctgaggc tcgaaagtat gggtagcaaa 780 caggattaga taccctggta gtccataccg taaacgatga atgctaagtg ttggagggtt 840 tccgcccttc agtgctgcag ctaacgcatt aagcattccg cctggggagt acggccgcaa 900 ggctgaaact caaaggaatt gacgggggcc cgcacaagcg gtggagcatg tggtttaatt 960 cgaagctacg cgaagaaccc ttaccaggtc ttgacatact atgcaaatct aagagattag 1020 acgttccctt cggggacatg gatacaggtg gtgcatggct gtcgtcagct cgtgtcgtga 1080 gatgttgggt taagtcccgc aacgaggcgc aacccttatt atcagttgcc cagcattaag 1140 ttgggcactc tggtgagact gccggtgaca aaccggagga aggtggggat gacgtcaaat 1200 catcatgccc cttat 1215 <110> REPUBLIC OF KOREA(MANAGEMENT: RURAL DEVELOPMENT ADMINISTRATION) <120> Novel Lactobacillus plantarum KCC-35 <130> PA-D18243 <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1215 <212> DNA <213> Unknown <220> <223> Lactobacillus plantarum KCC-35 <400> 1 ggggtgtggc ggccgccaat agatgcagtc gaacgaactc tggtattgat tggtgcttgc 60 atcatgattt acatttgagt gagtggcgaa ctggtgagta acacgtggga aacctgccca 120 gaagcggggg ataacacctg gaaacagatg ctaataccgc ataacaactt ggaccgcatg 180 gtccgagctt gaaagatggc ttcggctatc acttttggat ggtcccgcgg cgtattagct 240 agatggtggg gtaaccggtc tcacccattg gcaatgatat cgtagccgac ctgagagggt 300 aatcggccac attgggactg agacacggcc caaactccta cgggaggcag cagtagggaa 360 tcttccacaa tggacgaaag tctgatggag caacgcccgc gtgagtgaag aagggtttcg 420 gctcgtaaaa ctctgttgtt aaagaagaac atatctgaga gtaactgttc aggtattgac 480 ggtatttaac ccagaaagcc acggctaact acgtgccagc agccgcggta atacgtaggt 540 ggcaagcgtt gtccggattt attgggcgta aagcgagcgc aggcggtttt ttaagtctga 600 tgtgaaagcc ttcggctcaa ccgaagaagt gcatcggaaa ctgggaaact tgagtgcaga 660 agaggacagt ggaactccat gtgtagcggt gaaatgcgta gatatatgga agaacaccag 720 tggcgaaggc ggctgtctgg tctgtaactg acgctgaggc tcgaaagtat gggtagcaaa 780 caggattaga taccctggta gtccataccg taaacgatga atgctaagtg ttggagggtt 840 tccgcccttc agtgctgcag ctaacgcatt aagcattccg cctggggagt acggccgcaa 900 ggctgaaact caaaggaatt gacgggggcc cgcacaagcg gtggagcatg tggtttaatt 960 cgaagctacg cgaagaaccc ttaccaggtc ttgacatact atgcaaatct aagagattag 1020 acgttccctt cggggacatg gatacaggtg gtgcatggct gtcgtcagct cgtgtcgtga 1080 gatgttgggt taagtcccgc aacgaggcgc aacccttatt atcagttgcc cagcattaag 1140 ttgggcactc tggtgagact gccggtgaca aaccggagga aggtggggat gacgtcaaat 1200 catcatgccc cttat 1215
Claims (7)
상기 생균제제는 사람 또는 가축용 프로바이오틱스 생균제제인 것을 특징으로 하는 생균제제.The method of claim 2,
The probiotic preparation is a probiotic probiotic preparation for human or livestock.
Fermented feed produced by fermenting with the strain of claim 1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020190055227A KR102234499B1 (en) | 2019-05-10 | 2019-05-10 | Novel Lactobacillus plantarum KCC-35 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR1020190055227A KR102234499B1 (en) | 2019-05-10 | 2019-05-10 | Novel Lactobacillus plantarum KCC-35 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR20200130889A true KR20200130889A (en) | 2020-11-23 |
| KR102234499B1 KR102234499B1 (en) | 2021-03-31 |
Family
ID=73680405
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR1020190055227A KR102234499B1 (en) | 2019-05-10 | 2019-05-10 | Novel Lactobacillus plantarum KCC-35 |
Country Status (1)
| Country | Link |
|---|---|
| KR (1) | KR102234499B1 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20220030842A (en) * | 2020-09-03 | 2022-03-11 | 대한민국(농촌진흥청장) | Lactobacillus plantarum kcc-47 strain and composition comprising the same |
| ES2916398A1 (en) * | 2020-12-30 | 2022-06-30 | Univ Valencia | Lactobacillus plantarum strain, use as probiotic and bioactive product derived from it (Machine-translation by Google Translate, not legally binding) |
| CN116731939A (en) * | 2023-08-15 | 2023-09-12 | 山东利邦牧业股份有限公司 | Lactobacillus plantarum and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20160026948A (en) * | 2016-02-19 | 2016-03-09 | 대한민국(농촌진흥청장) | Lactobacillus plantarum KCC-19 and composition comprising the same |
| KR20170049292A (en) * | 2015-10-28 | 2017-05-10 | 대한민국(농촌진흥청장) | Lactobacillus plantarum KCC-26 and composition comprising the same |
| KR20180020742A (en) * | 2016-08-19 | 2018-02-28 | 대한민국(농촌진흥청장) | Lactobacillus plantarum KCC-32 and composition comprising the same |
-
2019
- 2019-05-10 KR KR1020190055227A patent/KR102234499B1/en active IP Right Grant
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20170049292A (en) * | 2015-10-28 | 2017-05-10 | 대한민국(농촌진흥청장) | Lactobacillus plantarum KCC-26 and composition comprising the same |
| KR20160026948A (en) * | 2016-02-19 | 2016-03-09 | 대한민국(농촌진흥청장) | Lactobacillus plantarum KCC-19 and composition comprising the same |
| KR20180020742A (en) * | 2016-08-19 | 2018-02-28 | 대한민국(농촌진흥청장) | Lactobacillus plantarum KCC-32 and composition comprising the same |
Non-Patent Citations (5)
| Title |
|---|
| Age. Res. Rev. 9, 107-116. |
| Best Pract. Res. Clin. Gastroenterol. 18, 299-313. |
| Cell 124, 837-848. |
| Genbank Accession No.KP091751.1(2015.03.23.)* * |
| J. Nutr. Health Aging 5, 80-91. |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20220030842A (en) * | 2020-09-03 | 2022-03-11 | 대한민국(농촌진흥청장) | Lactobacillus plantarum kcc-47 strain and composition comprising the same |
| ES2916398A1 (en) * | 2020-12-30 | 2022-06-30 | Univ Valencia | Lactobacillus plantarum strain, use as probiotic and bioactive product derived from it (Machine-translation by Google Translate, not legally binding) |
| WO2022144483A1 (en) * | 2020-12-30 | 2022-07-07 | Universitat De València (60%) | Lactobacillus plantarum strain, use as a probiotic and bioactive product derived therefrom |
| CN116731939A (en) * | 2023-08-15 | 2023-09-12 | 山东利邦牧业股份有限公司 | Lactobacillus plantarum and application thereof |
| CN116731939B (en) * | 2023-08-15 | 2023-11-07 | 山东利邦牧业股份有限公司 | Lactobacillus plantarum and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| KR102234499B1 (en) | 2021-03-31 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Nami et al. | Hypocholesterolaemic activity of a novel autochthonous potential probiotic Lactobacillus plantarum YS5 isolated from yogurt | |
| KR100911115B1 (en) | Novel lactic acid bacteria having acid resistance, bile-salt resistance and anti-bacterial effect and composition containing the same | |
| KR101399712B1 (en) | Novel fermented milk product and use thereof | |
| KR102234499B1 (en) | Novel Lactobacillus plantarum KCC-35 | |
| Kalui et al. | Functional characteristics of Lactobacillus plantarum and Lactobacillus rhamnosus from ikii, a Kenyan traditional fermented maize porridge | |
| EP1859022A1 (en) | Novel acid tolerant lactobacillus sakei probio- 65 with the ability of growth suppression of pathogenic microorganisms and the anti-allergic effect | |
| Le et al. | Identification of a novel potential probiotic Lactobacillus plantarum FB003 isolated from salted-fermented shrimp and its effect on cholesterol absorption by regulation of NPC1L1 and PPARα | |
| CN110564638A (en) | Lactobacillus reuteri with probiotic characteristics and application thereof | |
| JP4456160B2 (en) | Novel lactic acid strains and their use | |
| KR20080109585A (en) | Novel plant origin lactic acid bacteria having acid resistance, bile salt-resistance, salt-resistance and anti-bacterial effect and composition containing the same | |
| CN103060222A (en) | Bacillus subtilis B27 with probiotic effect and application thereof | |
| Rezaei et al. | Isolation of lactic acid probiotic strains from Iranian camel milk: technological and antioxidant properties | |
| CN114752529B (en) | Lactobacillus plantarum HOM3201 strain and viable bacteria preparation, preparation method and application thereof | |
| CN112195122A (en) | Selenium-enriched animal bifidobacterium for inhibiting tumor growth and application thereof | |
| CN106635916B (en) | Acetobacter orientalis YZD-09 and application thereof | |
| KR101451810B1 (en) | Novel Strains of Lactobacillus plantarum K255 with High Productivity of Gamma-Aminobutyric Acid | |
| Yetiman et al. | Assessment of Genomic and Metabolic Characteristics of Cholesterol-Reducing and GABA Producer Limosilactobacillus fermentum AGA52 Isolated from Lactic Acid Fermented Shalgam Based on “In Silico” and “In Vitro” Approaches | |
| KR101345248B1 (en) | Lactic acid bacteria for inhibiting the pathogenic bacteria and composition containing the same | |
| Xu et al. | Probiotic activity of ropy Lactiplantibacillus plantarum NA isolated from Chinese northeast sauerkraut and comparative evaluation of its live and heat-killed cells on antioxidant activity and RAW 264.7 macrophage stimulation | |
| KR19990079232A (en) | Lactobacillus plantarum pmeo 08 (KF-11028) microbe having cholesterol lowering ability | |
| KR102207949B1 (en) | Novel Lactobacillus plantarum KCC-36 | |
| KR20190047756A (en) | A composition comprising lactobacillus probiotics and a functional feed additive comprising the same | |
| KR102141569B1 (en) | Novel Lactobacillus plantarum KCC-34 | |
| KR101201338B1 (en) | A feed additive containing novel Lactobacillus reuteri | |
| KR20150101789A (en) | Bacillus subtilis BK418 strain having complex enzyme productivity and antifungal activity and uses thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| AMND | Amendment | ||
| E601 | Decision to refuse application | ||
| X091 | Application refused [patent] | ||
| AMND | Amendment | ||
| X701 | Decision to grant (after re-examination) | ||
| GRNT | Written decision to grant |