CN102212495A - Lactobacillus acidophilus and application thereof - Google Patents
Lactobacillus acidophilus and application thereof Download PDFInfo
- Publication number
- CN102212495A CN102212495A CN2011100951179A CN201110095117A CN102212495A CN 102212495 A CN102212495 A CN 102212495A CN 2011100951179 A CN2011100951179 A CN 2011100951179A CN 201110095117 A CN201110095117 A CN 201110095117A CN 102212495 A CN102212495 A CN 102212495A
- Authority
- CN
- China
- Prior art keywords
- lactobacillus acidophilus
- lactobacterium acidophilum
- preparation
- bionate
- pro
- 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.)
- Granted
Links
- 240000001046 Lactobacillus acidophilus Species 0.000 title claims abstract description 94
- 235000013956 Lactobacillus acidophilus Nutrition 0.000 title claims abstract description 94
- 229940039695 lactobacillus acidophilus Drugs 0.000 title claims abstract description 94
- 238000002360 preparation method Methods 0.000 claims abstract description 50
- 241001465754 Metazoa Species 0.000 claims abstract description 44
- 244000005700 microbiome Species 0.000 claims abstract description 18
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims abstract description 12
- 210000000936 intestine Anatomy 0.000 claims abstract description 11
- 230000036039 immunity Effects 0.000 claims abstract description 6
- 230000029087 digestion Effects 0.000 claims abstract description 5
- 238000010521 absorption reaction Methods 0.000 claims abstract description 4
- 241000894006 Bacteria Species 0.000 claims description 66
- 230000000968 intestinal effect Effects 0.000 claims description 46
- 241000209094 Oryza Species 0.000 claims description 40
- 235000007164 Oryza sativa Nutrition 0.000 claims description 40
- 235000009566 rice Nutrition 0.000 claims description 40
- 235000020183 skimmed milk Nutrition 0.000 claims description 34
- 206010012735 Diarrhoea Diseases 0.000 claims description 33
- 235000015099 wheat brans Nutrition 0.000 claims description 18
- 229930006000 Sucrose Natural products 0.000 claims description 15
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose 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)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 15
- 239000005720 sucrose Substances 0.000 claims description 15
- 238000009395 breeding Methods 0.000 claims description 13
- 230000001488 breeding effect Effects 0.000 claims description 13
- 239000000126 substance Substances 0.000 claims description 12
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 claims description 10
- 229920002774 Maltodextrin Polymers 0.000 claims description 10
- 239000005913 Maltodextrin Substances 0.000 claims description 10
- 229940035034 maltodextrin Drugs 0.000 claims description 10
- 239000001506 calcium phosphate Substances 0.000 claims description 9
- 229910000389 calcium phosphate Inorganic materials 0.000 claims description 9
- 235000011010 calcium phosphates Nutrition 0.000 claims description 9
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 claims description 9
- 230000000050 nutritive effect Effects 0.000 claims description 7
- 241000588724 Escherichia coli Species 0.000 claims description 5
- 229920002472 Starch Polymers 0.000 claims description 5
- 235000011187 glycerol Nutrition 0.000 claims description 5
- 239000008107 starch Substances 0.000 claims description 5
- 235000019698 starch Nutrition 0.000 claims description 5
- 239000003112 inhibitor Substances 0.000 claims description 4
- 230000009466 transformation Effects 0.000 claims description 3
- 235000015097 nutrients Nutrition 0.000 abstract description 31
- 230000012010 growth Effects 0.000 abstract description 30
- 239000000654 additive Substances 0.000 abstract description 8
- 239000000203 mixture Substances 0.000 abstract description 8
- 201000010099 disease Diseases 0.000 abstract description 4
- 238000012216 screening Methods 0.000 abstract description 4
- 238000006243 chemical reaction Methods 0.000 abstract description 2
- 230000002708 enhancing effect Effects 0.000 abstract description 2
- 238000009472 formulation Methods 0.000 abstract description 2
- 230000001228 trophic effect Effects 0.000 abstract description 2
- 241000186660 Lactobacillus Species 0.000 description 103
- 229940039696 lactobacillus Drugs 0.000 description 93
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 40
- 238000012360 testing method Methods 0.000 description 33
- 238000010790 dilution Methods 0.000 description 27
- 239000012895 dilution Substances 0.000 description 27
- 230000004083 survival effect Effects 0.000 description 27
- 230000001580 bacterial effect Effects 0.000 description 25
- 235000013372 meat Nutrition 0.000 description 25
- 238000000034 method Methods 0.000 description 25
- 235000014347 soups Nutrition 0.000 description 25
- 239000007788 liquid Substances 0.000 description 24
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 23
- 230000002354 daily effect Effects 0.000 description 22
- 230000000694 effects Effects 0.000 description 22
- 229920001817 Agar Polymers 0.000 description 20
- 239000008272 agar Substances 0.000 description 20
- 241000282898 Sus scrofa Species 0.000 description 19
- 238000000855 fermentation Methods 0.000 description 18
- 230000004151 fermentation Effects 0.000 description 18
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 18
- 239000000843 powder Substances 0.000 description 18
- 239000000243 solution Substances 0.000 description 18
- 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 17
- 239000008103 glucose Substances 0.000 description 17
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 15
- 239000012153 distilled water Substances 0.000 description 15
- 210000004913 chyme Anatomy 0.000 description 13
- 235000019621 digestibility Nutrition 0.000 description 13
- 239000001963 growth medium Substances 0.000 description 13
- 239000000047 product Substances 0.000 description 13
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 12
- 229910052802 copper Inorganic materials 0.000 description 12
- 239000010949 copper Substances 0.000 description 12
- 230000008569 process Effects 0.000 description 12
- 235000014590 basal diet Nutrition 0.000 description 11
- 239000003085 diluting agent Substances 0.000 description 11
- 210000001630 jejunum Anatomy 0.000 description 11
- 229910019142 PO4 Inorganic materials 0.000 description 10
- 239000001888 Peptone Substances 0.000 description 10
- 108010080698 Peptones Proteins 0.000 description 10
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 10
- 239000002253 acid Substances 0.000 description 10
- 238000004500 asepsis Methods 0.000 description 10
- 235000010633 broth Nutrition 0.000 description 10
- 210000001035 gastrointestinal tract Anatomy 0.000 description 10
- 235000019319 peptone Nutrition 0.000 description 10
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 10
- 239000010452 phosphate Substances 0.000 description 10
- 238000004321 preservation Methods 0.000 description 10
- 239000001632 sodium acetate Substances 0.000 description 10
- 229960004249 sodium acetate Drugs 0.000 description 10
- 235000017281 sodium acetate Nutrition 0.000 description 10
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 9
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 9
- SRBFZHDQGSBBOR-QMKXCQHVSA-N alpha-L-arabinopyranose Chemical compound O[C@H]1CO[C@@H](O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-QMKXCQHVSA-N 0.000 description 9
- 150000001413 amino acids Chemical class 0.000 description 9
- 229910002091 carbon monoxide Inorganic materials 0.000 description 9
- 229940099352 cholate Drugs 0.000 description 9
- 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 9
- 230000001079 digestive effect Effects 0.000 description 9
- 239000011790 ferrous sulphate Substances 0.000 description 9
- 235000003891 ferrous sulphate Nutrition 0.000 description 9
- 210000003405 ileum Anatomy 0.000 description 9
- 238000011081 inoculation Methods 0.000 description 9
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 9
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 9
- 235000014655 lactic acid Nutrition 0.000 description 9
- 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 9
- 229940061634 magnesium sulfate heptahydrate Drugs 0.000 description 9
- 239000011591 potassium Substances 0.000 description 9
- 229910052700 potassium Inorganic materials 0.000 description 9
- 210000002784 stomach Anatomy 0.000 description 9
- 108010046845 tryptones Proteins 0.000 description 9
- 239000011701 zinc Substances 0.000 description 9
- 229910052725 zinc Inorganic materials 0.000 description 9
- 241000252983 Caecum Species 0.000 description 8
- 229960000583 acetic acid Drugs 0.000 description 8
- 210000004534 cecum Anatomy 0.000 description 8
- 210000001072 colon Anatomy 0.000 description 8
- 239000000306 component Substances 0.000 description 8
- 230000036541 health Effects 0.000 description 8
- 239000002054 inoculum Substances 0.000 description 8
- 239000004310 lactic acid Substances 0.000 description 8
- SQQMAOCOWKFBNP-UHFFFAOYSA-L manganese(II) sulfate Chemical compound [Mn+2].[O-]S([O-])(=O)=O SQQMAOCOWKFBNP-UHFFFAOYSA-L 0.000 description 8
- 210000004400 mucous membrane Anatomy 0.000 description 8
- 239000006041 probiotic Substances 0.000 description 8
- 235000018291 probiotics Nutrition 0.000 description 8
- 241000193830 Bacillus <bacterium> Species 0.000 description 7
- 241000305071 Enterobacterales Species 0.000 description 7
- 210000000941 bile Anatomy 0.000 description 7
- 238000001035 drying Methods 0.000 description 7
- 210000001198 duodenum Anatomy 0.000 description 7
- 208000015181 infectious disease Diseases 0.000 description 7
- 238000004519 manufacturing process Methods 0.000 description 7
- 239000002574 poison Substances 0.000 description 7
- 231100000614 poison Toxicity 0.000 description 7
- 235000010482 polyoxyethylene sorbitan monooleate Nutrition 0.000 description 7
- 229920000053 polysorbate 80 Polymers 0.000 description 7
- 238000000926 separation method Methods 0.000 description 7
- 239000001509 sodium citrate Substances 0.000 description 7
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 7
- FRXSZNDVFUDTIR-UHFFFAOYSA-N 6-methoxy-1,2,3,4-tetrahydroquinoline Chemical compound N1CCCC2=CC(OC)=CC=C21 FRXSZNDVFUDTIR-UHFFFAOYSA-N 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- XLOMVQKBTHCTTD-UHFFFAOYSA-N Zinc monoxide Chemical compound [Zn]=O XLOMVQKBTHCTTD-UHFFFAOYSA-N 0.000 description 6
- 238000002474 experimental method Methods 0.000 description 6
- 239000000463 material Substances 0.000 description 6
- 239000002207 metabolite Substances 0.000 description 6
- 230000000529 probiotic effect Effects 0.000 description 6
- 238000012545 processing Methods 0.000 description 6
- 238000011160 research Methods 0.000 description 6
- 238000005507 spraying Methods 0.000 description 6
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 5
- 241000282887 Suidae Species 0.000 description 5
- 241001148470 aerobic bacillus Species 0.000 description 5
- 239000011248 coating agent Substances 0.000 description 5
- 238000000576 coating method Methods 0.000 description 5
- 235000013305 food Nutrition 0.000 description 5
- 239000008101 lactose Substances 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- 238000002156 mixing Methods 0.000 description 5
- 239000002245 particle Substances 0.000 description 5
- 230000000452 restraining effect Effects 0.000 description 5
- 238000005070 sampling Methods 0.000 description 5
- 239000002689 soil Substances 0.000 description 5
- 238000003860 storage Methods 0.000 description 5
- 238000009631 Broth culture Methods 0.000 description 4
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 4
- 102000016938 Catalase Human genes 0.000 description 4
- 108010053835 Catalase Proteins 0.000 description 4
- 235000019750 Crude protein Nutrition 0.000 description 4
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 4
- 238000010171 animal model Methods 0.000 description 4
- 230000000845 anti-microbial effect Effects 0.000 description 4
- 230000003115 biocidal effect Effects 0.000 description 4
- 230000037396 body weight Effects 0.000 description 4
- 238000004364 calculation method Methods 0.000 description 4
- 229910052799 carbon Inorganic materials 0.000 description 4
- 238000011161 development Methods 0.000 description 4
- 239000003814 drug Substances 0.000 description 4
- 230000003203 everyday effect Effects 0.000 description 4
- 239000012530 fluid Substances 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 230000014759 maintenance of location Effects 0.000 description 4
- BPMVRAQIQQEBLN-OBPBNMOMSA-N methyl n-[(e)-(1-hydroxy-4-oxidoquinoxalin-4-ium-2-ylidene)methyl]iminocarbamate Chemical compound C1=CC=C2N(O)C(=C/N=NC(=O)OC)/C=[N+]([O-])C2=C1 BPMVRAQIQQEBLN-OBPBNMOMSA-N 0.000 description 4
- 238000011169 microbiological contamination Methods 0.000 description 4
- 238000000386 microscopy Methods 0.000 description 4
- 235000013379 molasses Nutrition 0.000 description 4
- 239000006916 nutrient agar Substances 0.000 description 4
- 239000011574 phosphorus Substances 0.000 description 4
- 229910052698 phosphorus Inorganic materials 0.000 description 4
- 239000006152 selective media Substances 0.000 description 4
- 238000005406 washing Methods 0.000 description 4
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 3
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 3
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 3
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 239000007995 HEPES buffer Substances 0.000 description 3
- 241000282414 Homo sapiens Species 0.000 description 3
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 3
- 241000186604 Lactobacillus reuteri Species 0.000 description 3
- PLXBWHJQWKZRKG-UHFFFAOYSA-N Resazurin Chemical compound C1=CC(=O)C=C2OC3=CC(O)=CC=C3[N+]([O-])=C21 PLXBWHJQWKZRKG-UHFFFAOYSA-N 0.000 description 3
- DPDMMXDBJGCCQC-UHFFFAOYSA-N [Na].[Cl] Chemical compound [Na].[Cl] DPDMMXDBJGCCQC-UHFFFAOYSA-N 0.000 description 3
- 239000003674 animal food additive Substances 0.000 description 3
- 239000011575 calcium Substances 0.000 description 3
- 229910052791 calcium Inorganic materials 0.000 description 3
- 229960000427 carbadox Drugs 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 230000006378 damage Effects 0.000 description 3
- 238000013016 damping Methods 0.000 description 3
- 230000007423 decrease Effects 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 210000004051 gastric juice Anatomy 0.000 description 3
- 244000144972 livestock Species 0.000 description 3
- 239000012533 medium component Substances 0.000 description 3
- 229930182817 methionine Natural products 0.000 description 3
- 244000005706 microflora Species 0.000 description 3
- 239000011259 mixed solution Substances 0.000 description 3
- 235000021238 nutrient digestion Nutrition 0.000 description 3
- 239000002953 phosphate buffered saline Substances 0.000 description 3
- 239000002504 physiological saline solution Substances 0.000 description 3
- 230000002265 prevention Effects 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 150000003839 salts Chemical class 0.000 description 3
- 239000011787 zinc oxide Substances 0.000 description 3
- GHOKWGTUZJEAQD-ZETCQYMHSA-N (D)-(+)-Pantothenic acid Chemical compound OCC(C)(C)[C@@H](O)C(=O)NCCC(O)=O GHOKWGTUZJEAQD-ZETCQYMHSA-N 0.000 description 2
- GVJHHUAWPYXKBD-UHFFFAOYSA-N (±)-α-Tocopherol Chemical compound OC1=C(C)C(C)=C2OC(CCCC(C)CCCC(C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-UHFFFAOYSA-N 0.000 description 2
- 241000304886 Bacilli Species 0.000 description 2
- 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 2
- 230000005526 G1 to G0 transition Effects 0.000 description 2
- 241000287828 Gallus gallus Species 0.000 description 2
- 238000003794 Gram staining Methods 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- XUJNEKJLAYXESH-REOHCLBHSA-N L-Cysteine Chemical compound SC[C@H](N)C(O)=O XUJNEKJLAYXESH-REOHCLBHSA-N 0.000 description 2
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 2
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 2
- 241000186840 Lactobacillus fermentum Species 0.000 description 2
- PVNIIMVLHYAWGP-UHFFFAOYSA-N Niacin Chemical compound OC(=O)C1=CC=CN=C1 PVNIIMVLHYAWGP-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- AUNGANRZJHBGPY-SCRDCRAPSA-N Riboflavin Chemical compound OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-SCRDCRAPSA-N 0.000 description 2
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 2
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 2
- 239000006035 Tryptophane Substances 0.000 description 2
- 238000005903 acid hydrolysis reaction Methods 0.000 description 2
- 230000000996 additive effect Effects 0.000 description 2
- 239000004411 aluminium Substances 0.000 description 2
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 2
- 229910052782 aluminium Inorganic materials 0.000 description 2
- 230000000844 anti-bacterial effect Effects 0.000 description 2
- 230000010165 autogamy Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 230000004071 biological effect Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 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 2
- 238000012258 culturing Methods 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- AIUDWMLXCFRVDR-UHFFFAOYSA-N dimethyl 2-(3-ethyl-3-methylpentyl)propanedioate Chemical class CCC(C)(CC)CCC(C(=O)OC)C(=O)OC AIUDWMLXCFRVDR-UHFFFAOYSA-N 0.000 description 2
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 2
- 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 2
- 230000035622 drinking Effects 0.000 description 2
- 210000000981 epithelium Anatomy 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 230000037406 food intake Effects 0.000 description 2
- 235000012631 food intake Nutrition 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 238000003306 harvesting Methods 0.000 description 2
- 229940001882 lactobacillus reuteri Drugs 0.000 description 2
- 230000007774 longterm Effects 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 2
- CXKWCBBOMKCUKX-UHFFFAOYSA-M methylene blue Chemical compound [Cl-].C1=CC(N(C)C)=CC2=[S+]C3=CC(N(C)C)=CC=C3N=C21 CXKWCBBOMKCUKX-UHFFFAOYSA-M 0.000 description 2
- 229960000907 methylthioninium chloride Drugs 0.000 description 2
- 230000000813 microbial effect Effects 0.000 description 2
- 238000010899 nucleation Methods 0.000 description 2
- 235000021049 nutrient content Nutrition 0.000 description 2
- 229940066779 peptones Drugs 0.000 description 2
- 235000013406 prebiotics Nutrition 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000000241 respiratory effect Effects 0.000 description 2
- 238000005096 rolling process Methods 0.000 description 2
- 210000003296 saliva Anatomy 0.000 description 2
- 238000010186 staining Methods 0.000 description 2
- 239000007858 starting material Substances 0.000 description 2
- 229910052717 sulfur Inorganic materials 0.000 description 2
- 239000011593 sulfur Substances 0.000 description 2
- 150000003467 sulfuric acid derivatives Chemical class 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 239000013589 supplement Substances 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 238000009271 trench method Methods 0.000 description 2
- 229960004799 tryptophan Drugs 0.000 description 2
- 241001148471 unidentified anaerobic bacterium Species 0.000 description 2
- 229930003231 vitamin Natural products 0.000 description 2
- 235000013343 vitamin Nutrition 0.000 description 2
- 239000011782 vitamin Substances 0.000 description 2
- 229940088594 vitamin Drugs 0.000 description 2
- 230000003442 weekly effect Effects 0.000 description 2
- 235000019786 weight gain Nutrition 0.000 description 2
- 230000004584 weight gain Effects 0.000 description 2
- FPIPGXGPPPQFEQ-UHFFFAOYSA-N 13-cis retinol Natural products OCC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-UHFFFAOYSA-N 0.000 description 1
- -1 13g Substances 0.000 description 1
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 description 1
- LRDIEHDJWYRVPT-UHFFFAOYSA-N 4-amino-5-hydroxynaphthalene-1-sulfonic acid Chemical compound C1=CC(O)=C2C(N)=CC=C(S(O)(=O)=O)C2=C1 LRDIEHDJWYRVPT-UHFFFAOYSA-N 0.000 description 1
- ZCYVEMRRCGMTRW-UHFFFAOYSA-N 7553-56-2 Chemical compound [I] ZCYVEMRRCGMTRW-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose 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)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 241000186000 Bifidobacterium Species 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- GHOKWGTUZJEAQD-UHFFFAOYSA-N Chick antidermatitis factor Natural products OCC(C)(C)C(O)C(=O)NCCC(O)=O GHOKWGTUZJEAQD-UHFFFAOYSA-N 0.000 description 1
- WTEVQBCEXWBHNA-UHFFFAOYSA-N Citral Natural products CC(C)=CCCC(C)=CC=O WTEVQBCEXWBHNA-UHFFFAOYSA-N 0.000 description 1
- AUNGANRZJHBGPY-UHFFFAOYSA-N D-Lyxoflavin Natural products OCC(O)C(O)C(O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-UHFFFAOYSA-N 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- IMQLKJBTEOYOSI-GPIVLXJGSA-N Inositol-hexakisphosphate Chemical compound OP(O)(=O)O[C@H]1[C@H](OP(O)(O)=O)[C@@H](OP(O)(O)=O)[C@H](OP(O)(O)=O)[C@H](OP(O)(O)=O)[C@@H]1OP(O)(O)=O IMQLKJBTEOYOSI-GPIVLXJGSA-N 0.000 description 1
- LEVWYRKDKASIDU-IMJSIDKUSA-N L-cystine Chemical compound [O-]C(=O)[C@@H]([NH3+])CSSC[C@H]([NH3+])C([O-])=O LEVWYRKDKASIDU-IMJSIDKUSA-N 0.000 description 1
- AYFVYJQAPQTCCC-GBXIJSLDSA-N L-threonine Chemical compound C[C@@H](O)[C@H](N)C(O)=O AYFVYJQAPQTCCC-GBXIJSLDSA-N 0.000 description 1
- 240000001929 Lactobacillus brevis Species 0.000 description 1
- 241001468157 Lactobacillus johnsonii Species 0.000 description 1
- 240000006024 Lactobacillus plantarum Species 0.000 description 1
- 235000013965 Lactobacillus plantarum Nutrition 0.000 description 1
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 1
- XUYPXLNMDZIRQH-LURJTMIESA-N N-acetyl-L-methionine Chemical compound CSCC[C@@H](C(O)=O)NC(C)=O XUYPXLNMDZIRQH-LURJTMIESA-N 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- SCKXCAADGDQQCS-UHFFFAOYSA-N Performic acid Chemical compound OOC=O SCKXCAADGDQQCS-UHFFFAOYSA-N 0.000 description 1
- IMQLKJBTEOYOSI-UHFFFAOYSA-N Phytic acid Natural products OP(O)(=O)OC1C(OP(O)(O)=O)C(OP(O)(O)=O)C(OP(O)(O)=O)C(OP(O)(O)=O)C1OP(O)(O)=O IMQLKJBTEOYOSI-UHFFFAOYSA-N 0.000 description 1
- 241000589516 Pseudomonas Species 0.000 description 1
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 206010042566 Superinfection Diseases 0.000 description 1
- 108010008038 Synthetic Vaccines Proteins 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 1
- 239000004473 Threonine Substances 0.000 description 1
- FPIPGXGPPPQFEQ-BOOMUCAASA-N Vitamin A Natural products OC/C=C(/C)\C=C\C=C(\C)/C=C/C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-BOOMUCAASA-N 0.000 description 1
- QYSXJUFSXHHAJI-XFEUOLMDSA-N Vitamin D3 Natural products C1(/[C@@H]2CC[C@@H]([C@]2(CCC1)C)[C@H](C)CCCC(C)C)=C/C=C1\C[C@@H](O)CCC1=C QYSXJUFSXHHAJI-XFEUOLMDSA-N 0.000 description 1
- 229930003427 Vitamin E Natural products 0.000 description 1
- 229930003448 Vitamin K Natural products 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 230000003044 adaptive effect Effects 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- FPIPGXGPPPQFEQ-OVSJKPMPSA-N all-trans-retinol Chemical compound OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-OVSJKPMPSA-N 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- QGZKDVFQNNGYKY-UHFFFAOYSA-O ammonium group Chemical group [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 210000000436 anus Anatomy 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 238000009360 aquaculture Methods 0.000 description 1
- 244000144974 aquaculture Species 0.000 description 1
- 238000004380 ashing Methods 0.000 description 1
- BWKOZPVPARTQIV-UHFFFAOYSA-N azanium;hydron;2-hydroxypropane-1,2,3-tricarboxylate Chemical compound [NH4+].OC(=O)CC(O)(C(O)=O)CC([O-])=O BWKOZPVPARTQIV-UHFFFAOYSA-N 0.000 description 1
- 235000015278 beef Nutrition 0.000 description 1
- 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 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000036760 body temperature Effects 0.000 description 1
- 238000009835 boiling Methods 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 235000014633 carbohydrates Nutrition 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 1
- 229910000366 copper(II) sulfate Inorganic materials 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000007850 degeneration Effects 0.000 description 1
- 239000002274 desiccant Substances 0.000 description 1
- 238000001514 detection method Methods 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
- 208000010643 digestive system disease Diseases 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 238000004043 dyeing Methods 0.000 description 1
- 230000000688 enterotoxigenic effect Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- XTLNYNMNUCLWEZ-UHFFFAOYSA-N ethanol;propan-2-one Chemical compound CCO.CC(C)=O XTLNYNMNUCLWEZ-UHFFFAOYSA-N 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 210000003608 fece Anatomy 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 238000011010 flushing procedure Methods 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 230000001408 fungistatic effect Effects 0.000 description 1
- WIGCFUFOHFEKBI-UHFFFAOYSA-N gamma-tocopherol Natural products CC(C)CCCC(C)CCCC(C)CCCC1CCC2C(C)C(O)C(C)C(C)C2O1 WIGCFUFOHFEKBI-UHFFFAOYSA-N 0.000 description 1
- 230000002496 gastric effect Effects 0.000 description 1
- 239000012362 glacial acetic acid Substances 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 230000007412 host metabolism Effects 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 238000007654 immersion Methods 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- PNDPGZBMCMUPRI-UHFFFAOYSA-N iodine Chemical compound II PNDPGZBMCMUPRI-UHFFFAOYSA-N 0.000 description 1
- 239000011630 iodine Substances 0.000 description 1
- 229910052740 iodine Inorganic materials 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 230000002147 killing effect Effects 0.000 description 1
- 230000006651 lactation Effects 0.000 description 1
- 229940072205 lactobacillus plantarum Drugs 0.000 description 1
- 235000012204 lemonade/lime carbonate Nutrition 0.000 description 1
- WPBNNNQJVZRUHP-UHFFFAOYSA-L manganese(2+);methyl n-[[2-(methoxycarbonylcarbamothioylamino)phenyl]carbamothioyl]carbamate;n-[2-(sulfidocarbothioylamino)ethyl]carbamodithioate Chemical compound [Mn+2].[S-]C(=S)NCCNC([S-])=S.COC(=O)NC(=S)NC1=CC=CC=C1NC(=S)NC(=O)OC WPBNNNQJVZRUHP-UHFFFAOYSA-L 0.000 description 1
- 239000012092 media component Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 238000012543 microbiological analysis Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 210000004877 mucosa Anatomy 0.000 description 1
- WTEVQBCEXWBHNA-YFHOEESVSA-N neral Chemical compound CC(C)=CCC\C(C)=C/C=O WTEVQBCEXWBHNA-YFHOEESVSA-N 0.000 description 1
- 229960003512 nicotinic acid Drugs 0.000 description 1
- 235000001968 nicotinic acid Nutrition 0.000 description 1
- 239000011664 nicotinic acid Substances 0.000 description 1
- 239000002417 nutraceutical Substances 0.000 description 1
- 235000021436 nutraceutical agent Nutrition 0.000 description 1
- 235000003170 nutritional factors Nutrition 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 229940055726 pantothenic acid Drugs 0.000 description 1
- 235000019161 pantothenic acid Nutrition 0.000 description 1
- 239000011713 pantothenic acid Substances 0.000 description 1
- 230000007918 pathogenicity Effects 0.000 description 1
- SHUZOJHMOBOZST-UHFFFAOYSA-N phylloquinone Natural products CC(C)CCCCC(C)CCC(C)CCCC(=CCC1=C(C)C(=O)c2ccccc2C1=O)C SHUZOJHMOBOZST-UHFFFAOYSA-N 0.000 description 1
- 239000000467 phytic acid Substances 0.000 description 1
- 229940068041 phytic acid Drugs 0.000 description 1
- 235000002949 phytic acid Nutrition 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920003023 plastic Polymers 0.000 description 1
- 230000000069 prophylactic effect Effects 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 229940124551 recombinant vaccine Drugs 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 230000001373 regressive effect Effects 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 235000019192 riboflavin Nutrition 0.000 description 1
- 239000002151 riboflavin Substances 0.000 description 1
- 229960002477 riboflavin Drugs 0.000 description 1
- 230000000630 rising effect Effects 0.000 description 1
- 229910052711 selenium Inorganic materials 0.000 description 1
- 239000011669 selenium Substances 0.000 description 1
- 230000001568 sexual effect Effects 0.000 description 1
- GNBVPFITFYNRCN-UHFFFAOYSA-M sodium thioglycolate Chemical compound [Na+].[O-]C(=O)CS GNBVPFITFYNRCN-UHFFFAOYSA-M 0.000 description 1
- GOLXNESZZPUPJE-UHFFFAOYSA-N spiromesifen Chemical compound CC1=CC(C)=CC(C)=C1C(C(O1)=O)=C(OC(=O)CC(C)(C)C)C11CCCC1 GOLXNESZZPUPJE-UHFFFAOYSA-N 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 239000008399 tap water Substances 0.000 description 1
- 235000020679 tap water Nutrition 0.000 description 1
- 235000015193 tomato juice Nutrition 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 108010050327 trypticase-soy broth Proteins 0.000 description 1
- 239000000273 veterinary drug Substances 0.000 description 1
- 235000019155 vitamin A Nutrition 0.000 description 1
- 239000011719 vitamin A Substances 0.000 description 1
- QYSXJUFSXHHAJI-YRZJJWOYSA-N vitamin D3 Chemical compound C1(/[C@@H]2CC[C@@H]([C@]2(CCC1)C)[C@H](C)CCCC(C)C)=C\C=C1\C[C@@H](O)CCC1=C QYSXJUFSXHHAJI-YRZJJWOYSA-N 0.000 description 1
- 235000005282 vitamin D3 Nutrition 0.000 description 1
- 239000011647 vitamin D3 Substances 0.000 description 1
- 235000019165 vitamin E Nutrition 0.000 description 1
- 229940046009 vitamin E Drugs 0.000 description 1
- 239000011709 vitamin E Substances 0.000 description 1
- 235000019168 vitamin K Nutrition 0.000 description 1
- 239000011712 vitamin K Substances 0.000 description 1
- 150000003721 vitamin K derivatives Chemical class 0.000 description 1
- 229940045997 vitamin a Drugs 0.000 description 1
- 229940021056 vitamin d3 Drugs 0.000 description 1
- 150000003722 vitamin derivatives Chemical class 0.000 description 1
- 229940046010 vitamin k Drugs 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
Images
Abstract
The invention discloses a lactobacillus acidophilus and application thereof. The lactobacillus acidophilus is lactobacillus acidophilus and is collected in the China General Microbiology Center of Committee for Culture Collection of Microorganisms on 11th March, 2011; and the registered number of the lactobacillus acidophilus in the collection center is CGMCC No.4649. The lactobacillus acidophilus and a lactobacillus acidophilus preparation disclosed by the invention are mainly used as additives of animal fodder and can also be used for adjusting the microecological balance in animal intestines so that the product has a function of enhancing the specific immunity function to prevent and treat diseases and is also capable of providing trophic factors, accelerating digestion and absorption of nutrients, accelerating the animal growth and increasing the fodder conversion rate. Meanwhile, by screening the formulation, the cost of the preparation disclosed by the invention is low and the quality of the product is stable.
Description
Technical field
The present invention relates to a strain Lactobacterium acidophilum and an application thereof, particularly contain Pro-Bionate, feed, the Its Preparation Method And Use of this Lactobacterium acidophilum, belong to biological technical field.
Background technology
Add microbiotic in the feed and promote the history in existing more than 50 year of growth of animal, the intensification Developing of Animal Industry has been made major contribution.Constantly strengthen along with the raising of people's living standard with to the attention degree of health, people begin to find that antibiotic feed additive is bringing the negative effect that becomes increasingly conspicuous that the behind hid of great economic benefit, the autogenous infection and the superinfection that cause as microbiotic, chemical sproof generation, problem such as residual in the destruction of normal intestinal flora and livestock product and the environment, these all will bring serious threat to aquaculture, fodder industry and animal, and then jeopardize human beings'health by food chain.Therefore, forbid that the cry that Antibiotic Additive uses is more and more higher in livestock industry, the work of researching and developing new microbiotic substitute is extremely urgent.
In recent years, the research and development of antibiotics substitute probiotic bacterium, zymin, acid supplement etc. become the focus of green feed additive in recent years, and wherein probiotic bacterium receives much concern.What bright clear grade significantly descends to lactobacillus (Lactobacillus) quantity in the diarrhoea intestine of young pigs of discovering of sick, strong intestine of young pigs flora, and coliform count significantly rises.Use the lactobacillus probiotics can adjust the micro ecology of gastrointestinal tract balance, produce bacterial classification so lactobacillus is a kind of up-and-coming probiotics.But lactobacillus is the general name that a class can produce the bacterium of a large amount of lactic acid from fermentable carbohydrates, wherein lactobacillus genus and genus bifidobacterium are the dominant microfloras in the animal gastrointestinal tract, the probiotic formulations applicating history of these Pseudomonas the earliest, kind is a lot, comprising lactobacillus-fermented feed, lactobacillus powder and milk-acid bacteria extract.Lactobacillus acidophilus, lactobacillus johnsonii, fermentation lactobacillus, saliva lactobacillus, short and small lactobacillus, Lai Shi lactobacillus, lactobacillus plantarum and cellobiose lactobacillus can be arranged as the lactobacillus of fodder additives.Modern development in science and technology makes people more deep to the understanding of lactobacillus excellent characteristic, and is without stop to the upsurge of its development and use.In livestock industry, the development and use of lactobacillus also deepen constantly, and in conjunction with the feature of lactobacillus, hew out various application approaches.Lactobacillus distributes wide, contains a large amount of lactobacilluss in animal digestive tract.Studies show that some lactobacillus can be survived, and animal is had prebiotic effect in the digestive tube of animal.Totally 43 kinds of the safe microorganism kinds that U.S. FDA and AAFCO announced in 1989, wherein lactobacillus accounts for 12 kinds, and The Ministry of Agriculture of the People's Republic of China, MOA announced in 12 kinds of microorganism fodders that allow to use in 1999 has 3 kinds to be lactobacillus.In the U.S., the product of the industrial application of probiotics 57% contains lactobacillus genus.The lactasinum that the Kang Bai of Dalian Medical College etc. develop is a kind of drying agent of lactobacillus, good stability between preservation period, and every gram product contains lactobacillus alive about 10
7CFU, piglet oral 3-4 gram every day can effectively be treated diarrhoea.At present the bacterial classification of the normal microflora in chitling road has: Lactobacillus acidophilus (Lactobacillus acidophilus), lactobacillus reuteri (L.reuteri), short lactobacillus (L.brevis), cellobiose lactobacillus (L.cellobiosus), fermentation lactobacillus (L.fermentum), saliva lactobacillus (L.salivirius), wherein most importantly Lactobacillus acidophilus and lactobacillus reuteri.The quantity of lactobacillus, kind and position have important effect for prevention and control disease.Lactobacillus energy synthesise vitamins, supplement food digestion promotes host's metabolism, overcomes decay process, reduces host's blood cholesterol levels, strengthens the physiological significances such as tolerance of host to lactose, and does not have pathogenicity bo.Lactobacillus can suppress the activity of enteron aisle spoilage organism, and lactobacillus produces lactic acid can reduce the pH value, suppresses other microorganism growth.
The utilization of lactic acid bacteria class Products Development has become one of fodder industry and has enlivened the field, but also have some problem demanding prompt solutions such as unstable product quality, number of viable is not enough, digestibility is low and cost is high.Therefore, cultivation reproductivity is strong, tolerance strong, the lactobacillus series products of bacterial classification to produce output height, steady quality, have greater activity in body temperature of the higher digestive ferment of generation, to improve its utilization ratio, improve technology simultaneously, reduce production costs, improve the competitiveness of product in market and all very meaningful for Developing of Animal Industry and human beings'health.
Summary of the invention
The purpose of this invention is to provide a strain Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and application thereof.
Lactobacterium acidophilum provided by the present invention (Lactobacillus acidophilus) AI2021 is numbered CGMCC No.4649 registering on the books of China Committee for Culture Collection of Microorganisms common micro-organisms center.This Lactobacterium acidophilum strong stress resistance, anti-assorted bacterium ability be strong, have probiotic properties.
Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 has following characteristic:
(1), the survival rate after pH2.0 handles 6 hours is 69.3%;
(2), in 75 ℃ of water-baths among the 15min survival rate be 62.7%;
(3), place at room temperature that survival rate is respectively 60% after 1 month
(4), inoculate broth culture at 37 ℃, 5%CO
2Cultivate in the incubator that to reach the highest growth concentration in back 24 hours be 10
11Cfu/ml (Fig. 1).
Another object of the present invention provides a kind of Pro-Bionate.Said preparation can substitute existing feeding antibiotic, and has disease-resistant, somatotrophic effect.
Pro-Bionate provided by the present invention, its activeconstituents is described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021.
Described preparation has following at least a purposes:
1) product of microecological balance in the animal intestine is reconciled in preparation;
2) preparation strengthens the product of non-specific immunity;
3) improve the animal-feed transformation efficiency;
4) promote digestion and the absorption of animal to nutritive substance;
5) preparation reduces the product of animal diarrhea disease percentage;
6) preparation improves the product of breeding performonce fo animals.
Above-mentioned Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 or preparation can be used for preparing the intestinal bacteria inhibitor.
Intestinal bacteria in the described intestinal bacteria inhibitor specifically can be intestinal bacteria K88, e. coli k99 or intestinal bacteria 987P.
Described raising breeding performonce fo animals can be the production performance that improves weanling pig, is embodied as to improve piglet average daily gain and/or the average daily ingestion amount of piglet.
Described reduction animal diarrhea disease percentage specifically can be presented as the coli-infection grice diarrhoea rate that reduces; Described reduction diarrhoea index specifically can be presented as and reduce coli-infection grice diarrhoea index.
Described preparation specifically can be made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and carrier; Described carrier specifically can be at least a in calcium phosphate, powdered rice hulls, wheat bran, rice bran, skim-milk, maltodextrin, sucrose, glycerine and the starch.
In the described preparation, the proportioning of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and described carrier can be: (10
7-10
10) cfu: (0.3-5) gram carrier.
Described carrier can pass through micronization processes.Through after the micronization processes, 90% powder granularity is less than 100 μ m, and median size is less than 50 μ m, and described median size is 50% diameter of particle.
Described preparation specifically can be following any:
A) the Pro-Bionate A that is made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and calcium phosphate, among the described Pro-Bionate A, the proportioning of the two is 5 gram calcium phosphate: (10
7-10
10) the described Lactobacterium acidophilum of cfu (Lactobacillus acidophilus) AI2021;
B) the Pro-Bionate B that is made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and powdered rice hulls, among the described Pro-Bionate B, the proportioning of the two is 0.5 gram powdered rice hulls: (10
7-10
10) the described Lactobacterium acidophilum of cfu (Lactobacillus acidophilus) AI2021;
C) the Pro-Bionate C that is made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate C, the proportioning of the two is the described carrier of 0.3 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls and skim-milk, and the mass ratio of described powdered rice hulls and skim-milk is 2: 1;
D) the Pro-Bionate D that is made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate D, the proportioning of the two is the described carrier of 0.3 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls, skim-milk and maltodextrin, and the mass ratio of described powdered rice hulls, skim-milk and maltodextrin is 1: 1: 1;
E) the Pro-Bionate E that is made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate E, the proportioning of the two is the described carrier of 0.3 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu (Lactobacillus acidophilus) AI2021; Described carrier is made up of skim-milk and wheat bran, and the mass ratio of described skim-milk and wheat bran is 1: 1;
F) the Pro-Bionate F that is made up of described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate F, the proportioning of the two is the described carrier of 0.4 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls, skim-milk and wheat bran, and the mass ratio of described powdered rice hulls, skim-milk and wheat bran is 2: 1: 1.
The animal-feed of Lactobacterium acidophilum of the present invention (Lactobacillus acidophilus) AI2021 or described preparation can be used as additive and is used to prepare animal-feed, and animal wherein includes but not limited to various animals such as pig, ox, sheep, chicken.This feed has and the similar function of antibiotic feed, but the side effect of antibiotic-free feed.
In the described animal-feed, specifically can contain 10 in the described animal-feed of every gram
7Described Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 that CFU is above.
Lactobacterium acidophilum of the present invention and Pro-Bionate thereof are mainly as the fodder additives of animal, it can substitute the microbiotic in the existing animal daily ration, regulate microecological balance in the animal intestine, thereby have the prophylactic effect of the non-specific immune function of enhancing, the while can also provide nutritional factor, promotion is nutraceutical digests and assimilates, reduces diarrhoea, promotes growth of animal and improve food conversion ratio, improve the production performance of weanling pig.
Lactobacterium acidophilum of the present invention and Pro-Bionate thereof play a role in health care to control animal digestive system disease, can stimulate its gi tract to grow to growing animal simultaneously, so it is applied in the effect that can play disease-resistant growth-promoting in the feed as fodder additives.Simultaneously, thereby Lactobacterium acidophilum of the present invention and Pro-Bionate thereof have no drug resistance and medicine is residual in animal product human beings'health being produced the possibility of potential harm, is a kind of promising green feed additive.
Describe the present invention in detail below in conjunction with specific embodiment, described enforcement is used in to be understood rather than restriction the present invention.
The preservation explanation
Strain name: Lactobacterium acidophilum
Latin name: Lactobacillus acidophilus
Strain number: AI2021
Preservation mechanism: China Committee for Culture Collection of Microorganisms common micro-organisms center
Preservation mechanism is called for short: CGMCC
Address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City
Preservation date: on March 11st, 2011
The preservation center numbering of registering on the books: CGMCC No.4649
Description of drawings
Fig. 1 is the fermentation growth curve of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021;
Fig. 2 is the dull and stereotyped synoptic diagram that Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 suppresses the intestinal bacteria experiment
Embodiment
Lactobacterium acidophilum of the present invention (Lactobacillus acidophilus) AI2021, the Hungates that can be inoculated in Rogosa SL agar earlier rolls in the pipe substratum and cultivates, and the inoculum that obtains is inoculated in the Rogosa SL meat soup anaerobic culture medium and cultivates.
Described nutrient agar wherein is for being fit to any substratum that Lactobacterium acidophilum is cultivated, and which kind of substratum those skilled in the art know is fit to the Lactobacterium acidophilum cultivation.Specifically can select for use the Hungates of Rogosa SL agar to roll the pipe substratum, the Hungates of Rogosa SL agar described here rolls the pipe substratum and is interpreted as on the basis of the complete selective medium of Rogosa SL, the substratum that the suitable Lactobacterium acidophilum of suitably revising is cultivated.The substratum that described suitable Lactobacterium acidophilum is cultivated can be the substratum of any suitable Lactobacterium acidophilum cultivation of prior art, this dawn known to those skilled in the art, specifically can be: Tryptones 1-20g, extractum carnis 1-20g, yeast soaks powder 1-15g, glucose 1-20g, pectinose 1-15g, sucrose 1-15g, sodium-acetate 5-25g, Sodium Citrate 1-10g, potassium primary phosphate 1-15g, magnesium sulfate heptahydrate 0.1-1.5g, four water manganous sulfate 0.1-1.5g, ferrous sulfate 0.01-0.10g, tween-80 0.1-2.0mL, agar 5-20g, distilled water 100-2500mL; Be preferably Tryptones 5-15g, extractum carnis 5-15g, yeast soak powder 3-10g, glucose 5-15g, pectinose 3-10g, sucrose 3-10g, sodium-acetate 10-20g, Sodium Citrate 1-5g, potassium primary phosphate 1-10g, magnesium sulfate heptahydrate 0.1-1g, four water manganous sulfate 0.1-1g, ferrous sulfate 0.01-0.05g, tween-80 0.5-1.5mL, agar 10-15g, distilled water 500-2000mL; Tryptones 10g more preferably, extractum carnis 10g, yeast soak powder 5g, glucose 10g, pectinose 5g, sucrose 5g, sodium-acetate 15g, Sodium Citrate 2g, potassium primary phosphate 6g, magnesium sulfate heptahydrate 0.58g, four water manganous sulfate 0.25g, ferrous sulfate 0.03g, tween-80 1mL, agar 13g, distilled water 1000mL.
Liquid nutrient medium wherein is for being fit to any substratum that Lactobacterium acidophilum is cultivated, those skilled in the art also know this liquid nutrient medium, preferred meat soup anaerobic culture medium wherein, meat soup anaerobic culture medium described here is interpreted as on the basis of ordinary broth anaerobic culture medium, the suitable substratum of revising, the substratum of described suitable modification can be the substratum of any suitable Lactobacterium acidophilum cultivation of prior art, this dawn known to those skilled in the art, preferred Rogosa SL meat soup anaerobic culture medium, wherein the prescription of substratum is: Tryptones 5-15g, yeast soak powder 3-10g, glucose 5-15g, pectinose 3-10g, sucrose 3-10g, sodium-acetate 10-20g, Sodium Citrate 1-5g, potassium primary phosphate 1-10g, magnesium sulfate heptahydrate 0.1-1g, four water manganous sulfate 0.1-1g, ferrous sulfate 0.01-0.05g, tween-80 0.5-1.5mL, distilled water 500-2000mL; Be preferably Tryptones 10g, yeast soaks powder 5g, glucose 10g, pectinose 5g, sucrose 5g, sodium-acetate 15g, Sodium Citrate 2g, potassium primary phosphate 6g, magnesium sulfate heptahydrate 0.58g, four water manganous sulfate 0.25g, ferrous sulfate 0.03g, tween-80 1mL, distilled water 1000mL.
Use above-mentioned substratum, those skilled in the art generally know concrete culturing process, and the Hungates that is preferably above-mentioned Rogosa SL nutrient agar here rolls in the pipe and cultivates, and preferably in incubator, under 37 ℃, cultivate 15-72 hour; Lactobacterium acidophilum enlarged culturing wherein preferably in Rogosa SL meat soup anaerobic culture medium, was cultivated 15-80 hour down for 37 ℃.The pH value of above-mentioned substratum can be 6-7.
Described Lactobacterium acidophilum CGMCC No.4649 strain liquid is the strain liquid through one time fermentation, Secondary Fermentation or three fermentation gained.The carbon N/P ratio is 100: 5～10: 0.5～5 in the substratum of wherein said one time fermentation, Secondary Fermentation or three fermentations; Preferred 100: 10: 2.8.
Described Pro-Bionate's substratum is any suitable making Pro-Bionate's substratum, and its medium component is preferably: molasses, sucrose, glucose, three's weight ratio are 20-50 part: 10-70 part: 10-30 part.
Described Pro-Bionate's substratum is any suitable making Pro-Bionate's substratum, and its medium component also comprises peptone on the basis of above-mentioned substratum, and the weight ratio of itself and molasses is 10-25 part: 15-55 part.
Described Pro-Bionate's substratum is any suitable making Pro-Bionate's substratum, and its medium component also comprises phosphoric acid and/or soluble phosphate on the basis of above-mentioned substratum, and the weight ratio of itself and molasses is 10-25: 15-55.
Described aftertreatment be with Lactobacterium acidophilum bacterium liquid through once, secondary, three fermentations expand numerously, and zymocyte liquid is added in the phosphate buffered saline buffer, spraying drying gets final product, the pH of described phosphate buffered saline buffer is 6.5～6.7.
Animal-feed of the present invention comprises: conventional animal feed and Pro-Bionate's preparation of the present invention.Wherein Pro-Bionate's content is at least 0.001%, is preferably 0.001-3%, more preferably 0.01%-2%, most preferably 0.05%-0.2%.Pro-Bionate of the present invention can cooperate by same any on the market conventional animal feed at present, and animal-feed protection domain promptly of the present invention is not limited by the conventional animal feed kind that cooperates.
Yet conventional animal feed wherein can be preferably daily ration, and more preferably pig is used daily ration with daily ration, ruminating animal with daily ration and aquatic animal with daily ration, chicken.For different animals, the prescription of its daily ration has nothing in common with each other, and still, disclosed any daily ration can be as animal-feed of the present invention in the prior art, and its prescription all is the common practise in this field.
Employed experimental technique is ordinary method if no special instructions among the following embodiment.
Used material, reagent etc. if no special instructions, all can obtain from commercial channels among the following embodiment.
Separation and the evaluation of embodiment 1, Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021
Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 extraction from pig is gastrointestinal tract mucous, separation, screening, purifying obtain.
1.1 test materials
Get the gastrointestinal tract mucous 5cm of healthy piglet
2(stomach, duodenum, jejunum, ileum, caecum and colon), in Sterilization cup, the chyme with on the stroke-physiological saline solution flushing mucous membrane is equipped with the mucous membrane immersion in the beaker of 15mL HEPES damping fluid, behind tweezers waggle mucous membrane 5min, obtains supernatant liquor.
1.2 substratum
1.2.1Rogosa the substratum of SL agar
Tryptones 10g, extractum carnis 10g, yeast soak powder 5g, glucose 10g, pectinose 5g, sucrose 5g, sodium-acetate 15g, Sodium Citrate 2g, potassium primary phosphate 6g, magnesium sulfate heptahydrate 0.58g, four water manganous sulfate 0.25g, ferrous sulfate 0.03g, tween-80 1mL, agar 13g is settled to 1000mL with distilled water.
1.2.2Rogosa SL meat soup anaerobic culture medium
Tryptones 10g, yeast soak powder 5g, extractum carnis 10g, glucose 10g, pectinose 5g, sucrose 5g, sodium-acetate 15g, Sodium Citrate 2g, potassium primary phosphate 6g, magnesium sulfate heptahydrate 0.58g, four water manganous sulfate 0.25g, ferrous sulfate 0.03g, tween-80 1mL uses distilled water 1000mL.
1.3 gramstaining
Draw a small amount of Rogosa SL broth culture with asepsis injector, drop on the slide glass, oven dry is fixing gently on spirit lamp flame.Drip violet staining liquid, dye 1min, washing; Drip the Gram's iodine solution mordant dyeing, effect 1min, washing; Drip acetone ethanol mixed solution (acetone: decolouring 30s 95% ethanol=3: 7), washing; Drip husky yellow staining fluid and redye 1min, washing is waited to do, and observes on ordinary optical microscope, and it is red negative that thalline becomes, purple positive.The bacillus that is Gram-positive form unanimity further carries out catalase test.
1.4 catalase test
Make Rogosa SL slant medium, get the about 0.2m L injection of culture Rogosa SL medium slant, 5%CO are housed
2Incubator is cultivated 24h for 37 ℃, grow bacterium colony after, 3% superoxol is added drop-wise on the bacterium colony, be negative if there is bubble to produce explanation, be positive if there is bubble to produce explanation.Can tentatively think lactobacillus genus by the culture that anaerobism on the Rogosa SL is cultivated through Gram-positive and catalase test feminine gender.
1.5 acid resistance seed selection
Preparation contains the Rogosa SL broth culture of pH2.0 and pH2.5, pH3.0, pH3.5, and the bacterial classification of 24h has been cultivated in inoculation respectively, and inoculum size 1% is observed after cultivating 24h in 37 ℃.If the muddiness that substratum becomes illustrates that this bacterial strain is acidproof.Carry out gradient dilution to 10 from inoculation beginning and 8h with asepsis injector taking-up 1mL
-5After, to get the 0.3mL diluent and go up evenly coating at MRS agar (pH5.2), plate is placed on 37 ℃, 5%CO
2In the incubator, cultivate 24h, calculate survival rate.Microscopy is seen whether microbiological contamination then.The initial separation sample retention of the Lactobacterium acidophilum that can grow is standby at-80 ℃ of refrigerators.
1.6 bile tolerance performance seed selection
Preparation contains the malt extract medium of pig cholate 0.3%, and the bacterial classification of 24h has been cultivated in inoculation, and inoculum size 1% is observed behind 28 ℃ of cultivation 24h.If the muddiness that substratum becomes illustrates this bacterial strain bile tolerance.Microscopy is seen whether microbiological contamination then.The initial separation sample retention of the bacterial strain that can grow is standby at-80 ℃ of refrigerators.
1.7 anti-high-copper performance seed selection
Preparation contains the broth culture of 0.1% cupric sulfate pentahydrate, and the strain culture of 24h has been cultivated in inoculation, and inoculum size 1% is observed behind 28 ℃ of cultivation 12-72h.If the muddiness that substratum becomes illustrates the anti-high-copper of this bacterial strain.Microscopy is seen whether microbiological contamination then.The initial separation sample retention of the Lactobacterium acidophilum that can grow is standby at-80 ℃ of refrigerators.
1.8 anti-high zinc performance seed selection
Preparation contains the meat soup anaerobic culture medium of 1% zinc oxide, and the lactobacillus culture of 24h has been cultivated in inoculation, and inoculum size 1% is observed behind 28 ℃ of cultivation 12-72h.If the muddiness that substratum becomes illustrates the anti-high zinc of this bacterial strain.Microscopy is seen whether microbiological contamination then.The initial separation sample retention of the Lactobacterium acidophilum that can grow is standby at-80 ℃ of refrigerators.
2. result and discussion
2.1 separation of Lactobacterium acidophilum and evaluation
This test utilizes from healthy piglet is gastrointestinal tract mucous that the complete selective medium of Rogosa SL filters out 7009 strain lactobacillus strains after persalt is proofreaied and correct.In the HEPES damping fluid, shake 5min with piglet is gastrointestinal tract mucous, draw the 0.2mL supernatant liquor with asepsis injector and pack into and contain rolling of Rogosa SL agar and cultivate 24h in the pipe, choose white colony again and in the aseptic Rogosa SL of anaerobism meat soup, cultivate.Through gramstaining and catalase test, prove that lactobacillus is the GI dominant microflora of healthy piglet, illustrate through the gauged Rogosa SL of hydrochloric acid substratum to be fit to separate gastrointestinal tract mucous lactobacillus.
The screening of bacterial classification is and loaded down with trivial details process.The seed selection process of most lactic acid bacteria is to separate probiotic bacterium, the bacterial classification that so just can obtain to have good field planting effect from the digestive tube epithelium.
2.2 acid resistance seed selection
By the acid resistance seed selection, have 573 strain lactobacilluss can be in the Rogosa of pH2.0 SL substratum growth and breeding, mortality is 30%.This illustrates that GI most of Lactobacterium acidophilum acid resistance is better, has 70% can breed in the gi tract of piglet.Lactobacillus generally by field planting at digestive tube epithelium generation probiotic properties.Therefore, if bacterial classification does not have tolerance to be difficult to the production performance of animal is produced good effect to hydrochloric acid in gastric juice.
2.3 bile tolerance performance seed selection
To carry out the seed selection of bile tolerance performance by 573 strain lactobacilluss of acid resistance seed selection, have 95 strain lactobacilluss can be in containing the Rogosa SL substratum of cholate 0.3% growth and breeding, mortality is 83%.Studies show that cholate is a more adverse factors of the ratio hydrochloric acid in gastric juice that runs into of intestine lactobacillus in animal gastrointestinal tract.Chou and Weimer discover that be effective with cholate selectivity domestication lactobacillus to the tolerance of cholate, the energy force rate parental generation lactobacillus of the lactobacillus bile tolerance that domestication obtains afterwards through the several generations cholate is strong, lactobacillus produces tolerance to the cholate performance easily, and have certain heredity, the bacterium that selects bile tolerance has great importance aborning.
2.4 anti-high-copper performance seed selection
To carry out the seed selection of anti-high-copper performance by 95 strain lactobacilluss of bile tolerance performance seed selection, have 79 strain lactobacilluss can be in the Rogosa of 250mg/kg copper SL substratum growth and breeding, mortality is 17%.Prove that this lactobacillus has stronger anti-high copper feature.Because the high-copper daily ration has somatotrophic effect, be the daily ration of the commonplace use of present China, the Lactobacterium acidophilum of therefore selecting to have anti-high-copper is extremely necessary.
2.5 anti-high zinc performance seed selection
To carry out the seed selection of anti-high zinc performance by 79 strain lactobacilluss of anti-high-copper performance seed selection, have 71 strain lactobacilluss can be in containing the meat soup anaerobic culture medium of 1% zinc oxide growth and breeding, mortality is 10%.High zinc has and prevents to suffer from diarrhoea, promotes the effect of growing, and therefore utilizes zinc oxide to prepare pig starter feed in China, and especially the producer of weanling pig feed is a lot.Therefore the lactic bacilli strains that selects anti-high zinc also is necessary.
3. brief summary
Obtaining 71 strains by this test, to derive from piglet gastrointestinal tract mucous, and environment in the digestive tube is had the lactobacillus acidophilus strains of certain resistivity, and this lays the foundation for later seed selection work.
The biological Characteristics Study of embodiment 2, fermentation lactobacillus
1.1 growth curve
MRS meat soup: peptone, 10g; Beef extract, 10g; Yeast soaks powder, 5g; Dipotassium hydrogen phosphate, 2g; Citric acid two ammoniums, 2g; Glucose, 20g; Magnesium sulfate heptahydrate, 0.58g; Four water manganous sulfates, 0.25g; Sodium-acetate, 5g; Distilled water, 1000mL, dress 300mL MRS meat soup in the 500mL Erlenmeyer flask.By 1% inoculum size inoculating lactobacillus culture, at preceding 24h every 1h, after the 24h the 28th, 32,36,40,44, the 48h sampling, after getting the 1mL nutrient solution and carrying out gradient dilution, 10
-2～10
-6Extent of dilution is got the 0.3mL diluent and is gone up evenly coating at MRS agar (pH5.2), and each gradient is made 3 parallel samples, and plate is placed on 37 ℃, 5%CO
2Incubator in, cultivate 24h, the extent of dilution of the colony number 50～150 in the ware of making even is as calculating usefulness, each gradient is made 3 parallel samples, with the mean value ecbatic.Every milliliter of bacterial concentration is represented with logarithmic value.
1.2 acidproof survival rate
MRS meat soup prescription is regulated pH to 5.4 with among the embodiment 2 1.1 with Glacial acetic acid, again with concentrated hydrochloric acid with pH regulator to 2.0.Place Hungates to roll pipe, each pipe dress 20mL meat soup is made the aseptic meat soup of anaerobism.The back that cools down adds the culture of 1mL lactobacillus 16h in every pipe, when beginning, 2h, 4h, 6h and 8h take a sample respectively, measures survival rate.After taking out 1mL and carry out gradient dilution with asepsis injector, 10
-4～10
-7Extent of dilution is got the 0.3mL diluent and is gone up evenly coating at MRS agar (pH5.2), and each gradient is made 3 parallel samples, and plate is placed on 37 ℃, 5%CO
2Incubator in, cultivate 24h, the plate count of the colony number 50～150 in the ware of making even is with the mean value ecbatic.
The calculation formula of survival rate is:
S
Acid=n
x/ n
0
S
AcidFor handle the lactobacillus survival rate of back different time through pH2.0; n
0For pH2.0 handles preceding every milliliter of viable count; n
xBe every milliliter of viable count behind pH2.0 processing 2,4,6, the 8h.
1.3 heat-resisting survival rate
MRS meat soup prescription is with step 1.1, and regulating pH is 6.7, places Hungates to roll pipe, and each pipe dress 20mL meat soup is made the aseptic meat soup of anaerobism.Add the culture after the 1mL lactobacillus is cultivated 16h in every pipe, be placed in 37 ℃ of incubators behind the 16h, sampling is carried out live bacterial count and is cultivated, and the Hungates pipe is placed on rapidly in 75 ℃ of water-baths heats 15min simultaneously, and live bacterial count is carried out in sampling.After taking out 1mL and carry out gradient dilution with asepsis injector, 10
-4～10
-7Extent of dilution is got the 0.3mL diluent and is gone up evenly coating at MRS agar (pH5.2), and each gradient is made 3 parallel samples, and plate is placed on 37 ℃, 5%CO
2In the incubator, cultivate 24h, the colony number in the ware of making even is 50～150 plate count, with the mean value ecbatic.
The calculation formula of survival rate is:
S
Heat=n
1/ n
0
S
HeatFor through the lactobacillus survival rate after 75 ℃ of processing; n
0Be every milliliter of viable count before the heat treated; n
1Be every milliliter of viable count behind the heat treated 15min.
1.4 storage survival rate
MRS meat soup prescription is with step 1.1, and regulating pH is 6.7, places Hungates to roll pipe, each pipe is placed the MRS meat soup of 20mL, make the aseptic meat soup of anaerobism after, add the culture that the 1mL lactobacillus is cultivated 16h in every pipe, be placed in 37 ℃ of incubators behind the 16h, sampling 1mL carries out live bacterial count.Place under the room temperature after 1 month, sampling 1mL carries out live bacterial count and cultivates.After method of counting carries out gradient dilution with asepsis injector taking-up 1mL, 10
-4～10
-7Extent of dilution is got the 0.3mL diluent and is gone up evenly coating at MRS agar (pH5.2), and each gradient is made 3 parallel samples, and plate is placed on 37 ℃, 5%CO
2Incubator in, cultivate 24h, get colony number and be 50～150 plate count, with the mean value ecbatic.
The calculation formula of survival rate is:
S
Storage=n
1/ n
0
S
StorageBe the lactobacillus survival rate after preserving through 1 month; n
0Be every milliliter of viable count before preserving; n
1For preserving every milliliter of viable count after 1 month.
2 statistical methods
Adopt the common homogeneous design software of developing of Chinese homogeneous design association and Hong Kong Baptist University's statistical research and referral centre 4.0 editions, separating of each factor carried out optimization with quadratic regression and progressively regressive method.
3 results and discussion
3.1 Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 growth curve
Growth curve mainly reflects a kind of microbial growth characteristic, and microbial growth generally experiences lag period, logarithmic phase, stationary phase and decline phase four-stage, and this is a kind of typical growth curves model.Be the adaptive process of microorganism to new growing environment lag period, in this course, it is constant or descend that microorganism shows as quantity, and himself macromole and micromolecular composition are adjusted, and also can produce specific material such as enzyme simultaneously and wait and adapt to new environment.Logarithmic phase be microorganism to after the new environmental adaptation, growth and breeding speed is the stage of geometricprogression, is a fastest stage of quantity growth, shows as the increase of thalline quantity and weight.But arrived the latter stage of logarithmic phase, because the thalli growth metabolism is to the consumption of nutritive substance and the accumulation of toxic products, the growth and breeding speed of bacterium descends.Be the stage that rate of bacterial growth and rate of death tend to balance stationary phase.The decline phase bacterial number obviously descends.Measure growth curve and have vital role for definite suitable fermentation time
The growth curve of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 as shown in Figure 1, from Fig. 1 as seen, the concentration of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 the time slowly rises in beginning, through after the 3h from 10
5The individual order of magnitude rises rapidly, surpasses 10 to 18h
11The individual order of magnitude, to 32h, the number of bacteria level begins slowly to drop to 10
11Below the order of magnitude, major cause is that substratum nutritive substance density loss and harmful meta-bolites increase cause the dead increase of thalline, and the thalline quantity of growth and breeding is more more than dead thalline, causes total number of viable to reduce.From growth curve as can be seen, be 18～23h after cultivation the best harvesting time of Lactobacterium acidophilum, and as can be seen from the figure viable count maintains 10
11The order of magnitude reaches about 15h, and maintains 10
10The order of magnitude reaches about 26h.
3.2 the acidproof survival rate of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021
Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 sees Table 1 through the cell concentration that pH2.0 handles different time.
As seen from Table 1, at the preceding 6h that pH2.0 handles, the quantity of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 descends slowly, and survival rate drops to 69.3% from 87.9.Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 is that a strain separates the bacterial classification from the piglet jejunal mucous membrane, this survival rate should be an ideal comparatively for its anti-restraining effect of crossing hydrochloric acid in gastric juice or killing action, shows that also this bacterial strain can have the viable count of sufficient amount to arrive jejunum.
Table 1 pH2.0 handles the influence (unit: CFU/ml) of different time to live lactobacillus acidophilus concentration
3.3 the heat-resisting survival rate of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021
Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 handles 15min survival rate afterwards through 75 ℃ and reaches 62.7%.The pelleting temperature of general pig starter feed is between 70 ℃～85 ℃, after high temperature resistant survival rate low also be lactobacillus as one of major limitation sexual factor of fodder additives, according to the research of Fuller (1989), lactobacillus does not have viable bacteria after handling 10min through 75 ℃ basically.From heat-resisting survival rate, the fermentation lactobacillus of this test seed selection can tolerate the high temperature when granulating, and will have future preferably as fodder additives.
3.4 Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 storage tolerance survival rate
After one month storage, the survival rate of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 is 60%.
4 brief summaries
Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 18 to 23h is suitable thalline harvesting time after fermentation.Heat-resisting survival rate is 62.7%, and the storage tolerance survival rate is 60%, acidproof survival rate 69.3%, comprehensive degeneration-resistant survival rate 25.7%.
1.1 the lactobacillus meta-bolites suppresses the seed selection of intestinal bacteria ability
Intestinal bacteria (K88, K99 and 987P) are available from China Veterinary Drugs Supervisory Inst..K88 at the preserving number of Chinese medicine bacterium preservation administrative center is: CMCC44742, K99 at the preserving number of Chinese medicine bacterium preservation administrative center is: CMCC44820,987P at the preserving number of Chinese medicine bacterium preservation administrative center is: CMCC44317.
Lactobacillus: the Lactobacterium acidophilum of acidproof, the anti-high zinc of 71 strains, anti-high-copper and cholate.
Substratum: Mai Kangkai (MacConkay) substratum.
Add the about 15mL of MRS meat soup in the Hungates pipe, carbonating is made aseptic anaerobism MRS meat soup.Inoculate the 71 strains anti-high-copper of acidproof, anti-high zinc that is separated in the healthy piglet stomach and the Lactobacterium acidophilum of cholate in each pipe respectively, lactobacillus is cultivated the back through 18h and is inoculated in the meat soup by 1%, and after cultivating 24h in 37 ℃ of incubators, it is standby to put 4 ℃ of refrigerators preservations.
Suppress coli test: make the Mai Kangkai substratum, behind 121 ℃ of autoclaving 15min, asepticly topple over plate after the heating for dissolving.In the line of plate bottom plate is divided into three parallel zones with marking pen, each zone loop-carrier streak inoculation intestinal bacteria nutrient solution (4.0 * 10
8CFU/mL) in media surface, dig one then in the place of the about 3em of anomaly ware center line and widely be the ditch of 0.5em, ditch is vertical with the inoculation line, with the hot loop-carrier benefit end, as shown in Figure 2.Ditch is filled up (can not overflow) with different lactobacillus cultures respectively, after putting 37 ℃ of cultivations of common incubator 18h, the position of intestinal bacteria bacterium colony appears in observation, and colibacillary bacterium colony is red, measures from the nearest intestinal bacteria bacterium colony of ditch and the distance of ditch.Each lactobacillus is made two parallel samples, with the mean value ecbatic.
1.2 lactobacillus and the seed selection of intestinal bacteria mixed culture restraining effect
Intestinal bacteria (K88, K99 and 987P) source is saved together with 1.1.
Lactobacillus: 71 strain intestines meta-bolitess have stronger inhibiting lactobacillus to intestinal bacteria.
Colibacillary detection substratum: Mai Kangkai (MacConkay) substratum and 1.1 together.
Lactobacillus is inoculated in the aseptic MRS meat soup of anaerobism, behind 37 ℃ of cultivation 24h, puts 4 ℃ of refrigerators and preserves standby.
1.3 the different concns lactobacillus is to colibacillary inhibition
Select a strain lactobacillus at random, get 5mL culture (8.2 * 10
9CFU/mL) respectively with 5,10, the 15mL nutrient broth mixes, and makes 3 nutrient broths that contain different lactobacillus culture concentration, the inoculation intestinal bacteria, inoculum size is 5% of a nutrient solution, puts 37 ℃, 5%CO
2 Behind incubator cultivation 4,8 and the 18h, take out nutrient solution with asepsis injector respectively, nutrient solution becomes 10 through gradient dilution
-2～10
-5After, each extent of dilution is made 4 parallel samples, get the 0.3mL diluent, evenly coat on the maconkey agar flat board with " L " rod, place 37 ℃, common incubator is cultivated 18h, gets colony number 50～150 plate count, with the mean value ecbatic, calculate colibacillary quantity in every milliliter of nutrient solution.
2.3.2 contain lactobacillus and colibacillary quantitative relation in the nutrient broth of 1/4 lactobacillus
Get 5mL lactobacillus culture (10
8CFU/mL) mix with the 15mL nutrient broth respectively, inoculation 24h culture of Escherichia coli, inoculum size is respectively 10% (10 of nutrient solution
7CFU/mL), the quantity of lactobacillus is colibacillary 10 times in the initial mixed solution, and the pH that regulates nutrient solution with 0.1N NaOH solution is 7.0, after putting 37 ℃ of incubators and cultivating 24h, takes out nutrient solution with asepsis injector respectively, and nutrient solution is through gradient dilution to 10
-5, 10
-6, 10
-7After, to get the 0.3mL diluent and evenly coat on the MRS nutrient agar with " L " rod, the nutrient solution of each gradient is made 3 parallel samples, and MRS nutrient agar (pH5.4) is put 37 ℃, 5%CO
2Incubator is cultivated 18h, gets colony number 50～150 plate count, represents the quantity of lactobacillus with mean value, calculates the quantity of lactobacillus in every milliliter of nutrient solution.Directly get the 1mL nutrient solution through gradient dilution to 10 after cultivating 24h with asepsis injector
-2After, get the 0.3mL diluent and place on the maconkey agar, evenly be coated with " L " rod, each pipe is made 3 parallel samples, and flat board is placed 37 ℃, and common incubator is cultivated 18h, calculates each dull and stereotyped intestinal bacteria colony number, with the mean value ecbatic.
2 results
2.1 the lactobacillus meta-bolites suppresses colibacillary seed selection
The lactobacillus meta-bolites suppresses colibacillary effect to be index from the nearest intestinal bacteria bacterium colony of ditch and the distance of ditch, and distance is big more, illustrates that fungistatic effect is good more.The plate trench method that this test is adopted is the improvement to the plate borehole method of classics, ultimate principle is to be placed on antimicrobial substance porous in the ditch to cross agar and suppress intestinal bacteria, along with the increase from the ditch distance, the concentration of the antimicrobial substance in porous past is just more little.The distance of intestinal bacteria bacterium colony and ditch is big more, illustrates that the bacteriostasis of the contained antimicrobial substance of lactobacillus meta-bolites is strong more.Can measure the lactobacillus meta-bolites simultaneously to the colibacillary restraining effect of 3 strains with the plate trench method at same plate, and can only measure the colibacillary restraining effect of a strain at same plate with plate borehole method.63 strain lactobacilluss to the antibacterial distance of intestinal bacteria K88 and K99 between 1.0～2.6cm.The further quantitative relation of they and intestinal bacteria mixed culture of research.Wherein, Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 is respectively 2.0cm, 2.3cm, 1.8cm to the antibacterial distance of intestinal bacteria K88,987P and K99.
2.2 the quantitative relation of lactobacillus and intestinal bacteria vitro culture
2.2.1 different ratios Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 culture and the quantitative relation of intestinal bacteria under the mixed culture condition
The results are shown in Table 2.
Contain different ratios Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 nutrient solution in table 2 nutrient broth to colibacillary restraining effect (unit: CFU/mL)
2.2.2 lactobacillus and the intestinal bacteria quantitative relation under the mixed culture condition
Quantitative relation (the unit: CFU/mL) of table 3 lactobacillus and intestinal bacteria mixed culture
As can be seen from Table 3, the lactobacillus of different concns is similar to colibacillary restraining effect during to the intestinal bacteria mixed culture, illustrates that the nutrient broth that contains 25% Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 is enough to be used for doing to suppress colibacillary test.Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and intestinal bacteria see Table 8 with the quantitative relation of intestinal bacteria mixed culture in the nutrient broth that contains 25% Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 culture.As seen from the table, AI2021 has stronger bacteriostasis relatively to three kinds of serotype intestinal bacteria.Quantitative relation when external test tube is cultivated between different microorganisms can be used as the interactional in vivo a kind of index of reflection microorganism.
The bacterial strain that is numbered AI2021 that the present invention screens has following biological property:
Table 4 Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 biological characteristics
According to above biological property, AI2021 is accredited as Lactobacterium acidophilum (Lactobacillus acidophilus).With this bacterial strain on March 11st, 2011 in China Committee for Culture Collection of Microorganisms common micro-organisms center (CGMCC) preservation, the preservation center numbering of registering on the books: CGMCC No.4649.
1, Lactobacterium acidophilum CGMCC No.4649 is inoculated in the glucose nutrient broth, its component is: peptone 10g, extractum carnis 5g, sodium-chlor 5g, glucose 2.5g, distilled water 1000ml, and regulate pH to 6.7 with acetic acid, put common incubator 37 degree and cultivate after 18-24 hour, use the sterilized water gradient dilution, obtaining concentration is (10
7-10
10) strain liquid of cfu/g.
By weight, will contain 5 parts of mixing of 1 part of strain liquid, calcium phosphate of Lactobacillus acidophilus CGMCC No.4649, drying is pulverized, and promptly obtains the Pro-Bionate A that is made up of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and calcium phosphate.Among this Pro-Bionate A, the proportioning of the two is 5 gram calcium phosphate: 10
10Cfu Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021.
2, Lactobacterium acidophilum CGMCC No.4649 is inoculated in the Rogosa SL meat soup cultivates, its component comprises: 10g Tryptones, 5g yeast soak powder, 10g glucose, 5g pectinose, 5g sucrose, 15g sodium acetate, 2g Diammonium citrate, 6g potassium primary phosphate, 0.30g anhydrous magnesium sulfate, 0.12g manganous sulfate, 0.03g ferrous sulfate, 1mL tween-80,1000ml distilled water, and regulate pH to 6.7 with acetic acid; Put common incubator and cultivate after 18-24 hour for 37 ℃, use the sterilized water gradient dilution, obtaining concentration is (10
7-10
10) strain liquid of cfu/g.
By weight, to contain 5 parts of mixing of 10 parts of strain liquids, powdered rice hulls of Lactobacterium acidophilum CGMCC No.4649, spraying drying, powdered rice hulls carried out micronization processes in advance, promptly obtained the Pro-Bionate B that is made up of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and powdered rice hulls.Among this Pro-Bionate B, the proportioning of the two is 0.5 gram powdered rice hulls: 10
7The described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021.
3, place fermentor tank to carry out one grade fermemtation with cultivating the Lactobacterium acidophilum CGMCC No.4649 that obtains in the step 1, cultivate after 18-24 hour for 37 ℃, use the sterilized water gradient dilution, obtaining concentration is (10
7-10
10) strain liquid of cfu/g.The fermentation tank culture medium component is skim-milk, peptone, sucrose, lactose, glycerine, starch, and six weight ratio is 10 parts: 15 parts: 1 part: 3 parts: 1 part: 2 parts.Other adds suitable quantity of water, and regulates pH to 6.7 with acetic acid.The carbon N/P ratio is 100: 5: 0.5 in the substratum.
By weight, 10 parts of strain liquids, 2 parts of the powdered rice hulls that will contain Lactobacterium acidophilum CGMCC No.4649,1 part of skim-milk, spraying drying, powdered rice hulls passes through micronization processes (90% powder granularity is less than 100 μ m, and median size is less than 50 μ m, and described median size is 50% diameter of particle) in advance, promptly obtain the Pro-Bionate C that is made up of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and carrier, the proportioning of the two is the described carrier of 0.3 gram: 10
8Cfu Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls and skim-milk, and the mass ratio of described powdered rice hulls and skim-milk is 2: 1.
4, place seeding tank to carry out one grade fermemtation with cultivating the Lactobacterium acidophilum CGMCC No.4649 that obtains in the step 1, place the production jar to carry out second order fermentation then, use the sterilized water gradient dilution, obtaining concentration is (10
7-10
10) strain liquid of cfu/g.Two-stage fermentation jar nutrient media components is 12 parts of powdered rice hulls, 10 parts of peptones, and 10 parts of skim-milks, 1 part of sucrose, 2 parts of lactose, 1 part of glycerine, 3 parts of starch mix, and other adds suitable quantity of water.And regulate pH to 6.7 with acetic acid.The carbon N/P ratio is 100: 10: 5 in the substratum.
By weight, to contain 1 part of mixing of 1 part of 1 part of 10 parts of strain liquids, powdered rice hulls, maltodextrin, skim-milk of Lactobacterium acidophilum CGMCC No.4649, spraying drying, powdered rice hulls passes through micronization processes in advance, and (90% powder granularity is less than 100 μ m, median size is less than 50 μ m, described median size is 50% diameter of particle), promptly obtain the Pro-Bionate D that is made up of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and carrier, the proportioning of the two is the described carrier of 0.3 gram: 10
9Cfu Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls, skim-milk and maltodextrin, and the mass ratio of described powdered rice hulls, skim-milk and maltodextrin is 1: 1: 1.
5, place seeding tank to carry out fermenting twice continuously with cultivating the Lactobacterium acidophilum CGMCC No.4649 that obtains in the step 2, place the production jar to carry out three grade fermemtation then, use the sterilized water gradient dilution, obtaining concentration is (10
7-10
10) strain liquid of cfu/g.The fermentation tank culture medium component is 12 parts of powdered rice hulls, 6 parts of peptones, 10 parts of skim-milks, 1 part of sucrose, 1 part in molasses, and 2 parts of lactose, 1 part of glycerine, 3 parts of starch, the carbon N/P ratio is 100: 7: 2.Add entry, and regulate pH to 6.7 with acetic acid.
By weight, to contain 1 part of mixing of 1 part in 6 parts of strain liquids, wheat bran, skim-milk of Lactobacterium acidophilum CGMCC No.4649, spraying drying, powdered rice hulls passes through micronization processes in advance, and (90% powder granularity is less than 100 μ m, median size is less than 50 μ m, described median size is 50% diameter of particle), promptly obtain the Pro-Bionate E that forms by yogurt acidfast bacilli (Lactobacillus acidophilus) AI2021 and carrier, the proportioning of the two is the described carrier of 0.3 gram: 10
7Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021; Described carrier is made up of skim-milk and wheat bran, and the mass ratio of described skim-milk and wheat bran is 1: 1.
6, contain Lactobacterium acidophilum CGMCC No.4649 strain liquid preparation method with step 4.
By weight, to contain 1 part of 1 part in 2 parts of 10 parts of strain liquids, powdered rice hulls, wheat bran, skim-milk, the mixing of Lactobacterium acidophilum CGMCC No.4649, spraying drying, powdered rice hulls and wheat bran pass through micronization processes in advance, and (90% powder granularity is less than 100 μ m, median size is less than 50 μ m, described median size is 50% diameter of particle), promptly obtain the Pro-Bionate F that is made up of Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 and carrier, the proportioning of the two is the described carrier of 0.4 gram: 10
10Cfu Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls, skim-milk and wheat bran, and the mass ratio of described powdered rice hulls, skim-milk and wheat bran is 2: 1: 1.
Experimental example 5, Pro-Bionate are to the influence of piglet production performance.
1.1 materials and methods:
Experimental animal
Choose 36 of the DLY ternary hybrid piglets of 28 ± 2d wean, mean body weight 7.56 ± 0.52kg (creep feed is raised lactation drylot feeding nutriment 7 days in the wean back), distinguishing component at random by body weight is 2 treatment group, every group of 6 repetitions, each repeats 3 piglets.
1.2 Pro-Bionate
Adopt the Pro-Bionate A-F of embodiment 4.
1.3 test grouping
It is two processing that test is divided into, and handling 1 is test group, any among the Pro-Bionate A-F of basal diet interpolation weight ratio 0.1% embodiment 4; Handling 2 is control group, and each arrangement of handling of basal diet sees Table 5.The composition and the nutritive ingredient of basal diet see Table 5.
The basal diet of table 5 growth test is formed and trophic level
Annotate:
1. crude protein, Methionin, methionine(Met), Gelucystine, Threonine, calcium and phosphorus are measured value.
2. per kilogram Preblend provides: vitamin A, 11,000IU; Vitamin D3 500,000 I.U/GM, 1503IU; Vitamin-E, 44.1IU; Vitamin K, 4.0mg; Riboflavin, 5.22mg; Pantothenic acid, 20.0mg; Nicotinic acid, 26.0mg; Vitamin B12,0.01mg; Manganese, 35.0mg; Iron, 100.0mg; Zinc, 90.0mg; Copper, 16.5mg; Iodine, 0.30mg; Selenium, 0.30mg.
1.4 feeding and management
Test in national feed Engineering Technical Research Centre respiratory chamber and carry out.21 days trial periods.The duration of test feeding piglet is in fully closed child care piglet house, and the temperature in the house remains on 24～27 ℃.Free choice feeding, each hurdle circle are equipped with the duck-beak type water fountain and freely drink water for piglet.1% Preblend autogamy of basal diet does not contain any microbiotic.The immunity of piglet is undertaken by the immune programme for children of pig routine veterinary transmissible disease, the strict health and epidemic prevention system of carrying out of feeding piglet control measures.
1.5 sample collection and processing
Test claims the piglet individual weight when beginning, 7d, 14d and 21d, record day weight gain and feed food consumption weekly.
1.6 data statistics
(SPSS Inc., USA) statistics is handled in the check of the independent sample t of statistical software to test all The data SPSS12.0.
2. result and discussion
2.1 the Pro-Bionate to weaned piglet after the influence of growth performance
Table 6 Pro-Bionate E is to the influence of weanling pig growth performance
Annotate: the table intermediate value is a mean+SD.
As seen from Table 6, wean back 1wk, 2wks and 3wks, piglet average daily gain (ADG) comparison of test group be you can well imagine high by 3.27%, 3.17% and 3.21% significant difference (P＜0.05) according to component, average daily ingestion amount (ADFI) comparison according to component you can well imagine high by 3.17%, 3.29% and 3.16%, significant difference (P＜0.05)).
Carry out same experiment with Pro-Bionate A-D and F and also obtained identical conclusion.
Experimental example 6, Pro-Bionate are to the influence of coli-infection grice diarrhoea.
1. materials and methods
The Pro-Bionate E of embodiment 4.
1.1 experimental animal and feeding and management
Select the health Du * length * big commodity hybridization piglet of 12 28 ± 2d wean, body weight is 7.65 ± 1.10kg.Be divided into 2 groups at random, single cage is raised in metabolic cage, and the room temperature of respiratory chamber is controlled at 25 ℃～27 ℃, 6 piglets of treatment group oral liquid acidophilic's Lemonal of drinking water every day, and drinking every day into, the sum of Lactobacterium acidophilum CGMCC No.4649 is about 10
7CFU, 6 piglet drinking public water supplies of control group in contrast.(total count is 10 with the mixed-culture medium of 20mL e. coli k99, K88 and 987P (source with embodiment 3) to test 8d
8CFU) oral challenge, the colibacillary of three kinds of serotypes is in equal proportions in this mixed solution, a twice-daily, 1d attacks poison back pig and only still freely drinks tap water and search for food altogether, measures and attacks poison back diarrhoea disease time and diarrhoea index, trial period 14d.
1.2 test daily ration
The test piglet basal diet of searching for food does not contain any microbiotic, prepares with reference to the requirement that NRC (1998) recommends.Embodiment 5 tables 5 are seen in the basal diet configuration.
1.3 diarrhoea index
The diarrhoea index is that the diarrhoea head multiply by corresponding diarrhoea score, and the diarrhoea score is calculated according to the diarrhoea standards of grading of table 7.Diarrhea rate is that the inferior piglet number divided by every day of the total head of the interior piglet of suffering from diarrhoea of 7d multiply by 7 after attacking poison.
Table 7 diarrhoea standards of grading
1.4 statistical study
SPSS12.0 is adopted in the statistical study of data, and (SPSSInc., USA) statistical study is carried out in the independent sample t-check of software.
2 results and discussion
Test-results sees Table 8, and the test group diarrhea disease percentage descends 69.6% than control group, the diarrhoea index decreased 67.Control group 2d after attacking poison promptly has first piglet to suffer from diarrhoea, and experimental group is just found first grice diarrhoea until attack poison back 5d.In China, many experimental studies have proved the diarrhoea that probiotics can effectively be prevented and treated sucking piglets and weanling pig, and some effect is better than microbiotic or recombinant vaccine.The Lactobacterium acidophilum lactobacillus of the lactic acid producing rhzomorph that contains in the Pro-Bionate has a good Chinese People's Anti-Japanese Military and Political College enterobacteria characteristic external, according to have in the Hoefling report newborn piglet diarrhoea 26% since enterotoxigenic Escherichia coli cause.The test group piglet is 7d before attacking poison, the oral always water that contains Lactobacterium acidophilum, and Lactobacterium acidophilum exists in a large number at gi tract chyme and mucous membrane, and at specific gi tract microhabitat performance prebiotic effect.Piglet is subjected to intestinal bacteria when attacking poison; because Lactobacterium acidophilum can suppress colibacillary growth and breeding; thereby protected body to avoid the grievous injury that a large amount of breedings of intestinal bacteria cause effectively, reduced the severity of diarrhea disease percentage and diarrhoea, postponed the diarrhoea disease time simultaneously.
Table 8 Pro-Bionate E is to the influence of coli-infection diarrhea of weaned piglets
Carry out same experiment with other five kinds of Pro-Bionates of embodiment 4 and also obtained identical conclusion.
Experimental example 7, Pro-Bionate are to the influence of early-weaned piglets gi tract microorganism species, nutrient digestibility.
Materials and methods
1.1 experimental animal and daily ration are handled
Select 36 28 ± 2d weanling pigs, body weight 7.65 ± 1.09kg, by completely random block design animal experiment, test divides 2 processing, and each handles 6 repetitions, each repeats 3 piglets, high-rise online flat supporting, the hurdle circle is of a size of 2.0 * 2.0m, and every circle is a repetition, each handles 3 circle boars, 3 circle sows.Handling 1 is test group, and basal diet adds the Pro-Bionate F of 0.1% (mass content) embodiment 4; Handling 2 is control group, and basal diet adds the 50mg/kg carbadox, and each arrangement of handling sees Table 5.The duration of test feeding piglet is in fully closed child care piglet house, and the temperature in the house remains on 24～27 ℃.Free choice feeding, each hurdle circle are equipped with the duck-beak type water fountain and freely drink water for piglet.1% Preblend autogamy of basal diet does not contain any microbiotic.The immunity of piglet is undertaken by the immune programme for children of pig routine veterinary transmissible disease, the strict health and epidemic prevention system of carrying out of feeding piglet control measures.Trial period 21d claims the piglet individual weight when on-test, 7d, 14d and 21d, record day weight gain and feed food consumption weekly.In the end the diatomite of interpolation 0.5% is as the digestibility of indicator mensuration nutritive substance in 1 all daily rations, and the basal diet composition sees Table 5.
The arrangement of table 9 experimental animal
1.2 sample collection and processing
Each repeats to get a piglet to test 21d, fetches the intestines chyme after butchering and measures amino acid digestibility.Asepsis is got stomach, each 5cm of duodenum, jejunum, ileum, caecum and colon is long, after the ligation of two ends, be placed in-80 ℃ of refrigerators, be used to measure the microorganism species of content and mucous membrane, the content of getting stomach, duodenum, jejunum epimere, jejunum stage casing, jejunum hypomere, ileum, caecum, the colon section of falling and colon liter section is loaded in the plastics tubing, is placed in-80 ℃ of refrigerators.Testing last three days is that the fresh excreta of just having discharged from anus is collected by unit with the circle, is placed in-20 ℃ of refrigerators, measures the nutrient digestion rate.
1.3 microbiological analysis
The aseptic gi tract chyme 1g that takes by weighing different sites is dissolved in the stroke-physiological saline solution of 99mL, places 5～8 little granulated glass spherees in the bottle of stroke-physiological saline solution.And then carry out 10 times of dilutions step by step, until 10-7,10
-5～10
-7Each extent of dilution is got the 0.3mL diluent and carry out lactobacillus, bifidus bacillus and anaerobic bacteria culture in being rolled the pipe substratum accordingly.10
-2～10
-5Each extent of dilution is got the 0.3mL diluent carries out intestinal bacteria, enterobacteria and aerobic bacteria in corresponding plate culture medium counting.Mucous membrane microbioassay method is with reference to the method for Rojas and Conway (1996), the aseptic 1cm2 gastrointestinal mucosa of getting, wash mucous membrane to there not being the visible chyme with aseptic PBS solution, blot lip-deep water with aseptic filter paper, with aseptic nipper tissue block is immersed the aseptic HEPES damping fluid of 10mL then, and constantly 5min is washed in vibration, and then carries out 10 times of dilutions step by step, up to 10-3.At each extent of dilution, get the 0.3mL diluent in corresponding flat board or roll the pipe substratum in.Microorganism is cultivated with selective medium, and the making of substratum is with reference to the method for Chen Tianshou (1995), and intestinal bacteria and enterobacteria substratum are with Yihong methylene blue (EMB) substratum (peptone, 10g; Lactose, 10g; Dipotassium hydrogen phosphate, 2g; Agar, 13g; Yihong-Y, 0.4g; Methylene blue, 0.065g; Distilled water, 1000mL; PH, 7.1), the lactobacillus complete selective medium of Rogosa SL (Tryptones, 10g; Yeast soaks powder, 5g; Glucose, 10g; Pectinose, 5g; Sucrose, 5g; Sodium-acetate, 15g; The citric acid ammonium, 2g; Potassium primary phosphate, 6g; Magnesium sulfate heptahydrate, 0.58g; Four water manganous sulfates, 0.25g; Ferrous sulfate, 0.03g; Tween-80,1g; Agar, 13g; 0.1% resazurin, 1mL; Distilled water, 1000mL; Regulate pH to 5.2 with acetic acid).Bifidus bacillus is used bifidus bacillus substratum (BM): glucose, 20g; Trypticase, 20g; Yeast soaks powder, 10g; Peptone, 10g; Tomato juice, 333mL; Tween-80,2g; Distilled water, 1000mL; PH 6.6, lime carbonate, 10g; Halfcystine, 0.5g; 0.1% resazurin, 1mL.Aerobic bacteria and anerobe are used aerobic bacteria and anaerobic bacteria culture base (junket peptone, 15g; Glucose, 5g; The L-Gelucystine, 0.5g; Sodium thioglycollate, 0.5g; Yeast soaks powder, 5g; Sodium-chlor, 2.5g; 0.1% resazurin, 1mL; Agar, 13g, distilled water, 1000mL; PH7.1), anerobe, lactobacillus and bifidus bacillus substratum are made into Hungates and roll pipe, and aerobic bacteria, intestinal bacteria and enterobacteria are cultivated with dull and stereotyped.Hungates pipe and substratum plate are placed on 37 ℃ of incubators, count behind the cultivation 48h.Each extent of dilution is made three flat boards or is rolled pipe, with the flat board of 50～150 bacterium colonies or the extent of dilution work counting usefulness of rolling pipe.
1.4 chemical analysis takes out ight soil from refrigerator, after to be unit with the circle with ight soil mix, get and be positioned over about 300g in the aluminium box, the aluminium box is placed on forced air drying 96h in 65 ℃ the baking oven.After daily ration and faecal samples are pulverized, cross 40 mesh sieves.The chyme sample carries out lyophilize, is used to measure amino acid whose daily ration sample and chyme and pulverizes 60 mesh sieves.Thick moisture, crude protein, coarse ash, calcium and the total phosphorus of daily ration and excrement measured with reference to State Standard of the People's Republic of China GB/T6435-1986 (2001), GB/T 6432-1994 (2001), GB/T 6438-1992 (2001), GB/T 6436-1992 (2001) and GB/T 6437-1992 (2001) respectively.Wherein, crude protein adopts the KJELTEC1035 fully-automatic analyzer to measure.
Energy adopts the full-automatic energometry instrument of PARR1281 (PARR Instrument Corp., the U.S.) to measure.Amino acid (removing sulfur-containing amino acid and tryptophane) is measured with automatic analyzer for amino acids (L-8800 of Hitachi, Japan) behind 110 ℃ of following 6mol/L hydrochloric acid hydrolysis 24h by standard GB/T18246-2000 (2001); Sulfur-containing amino acid (comprising methionine(Met) and halfcystine) after using peroxyformic acid (1mL hydrogen peroxide+9mL formic acid) oxidation 16h under 110 ℃, uses the hydrochloric acid hydrolysis method to measure according to standard GB/T15399-1994 (1996); Tryptophane uses high pressure liquid chromatography (Tianjin, island LC-10A, Japan) to measure behind 110 ℃ of following 4mol/L sodium hydroxide hydrolysis 24h according to standard GB/T18246-2000 (2001).The method of (1974) such as employing McCarthy is measured the salt acid insoluble ash.Promptly get about 10g sample, boil 30min with 100mL 4N HCl after, with no ash content filter paper filtering, wash until anacidity, then more than 650 ℃ of following ashing 6h with boiling water.
1.5 digestibility calculation formula
Calculate the all-digestive tract digestibility of amino acid ileal digestibility or nutrient as follows:
D＝100-(C1×P2)÷(C2×P1)×100
Wherein, D=amino acid ileal digestibility or nutrient all-digestive tract digestibility (%),
C1=cattle salt acid insoluble ash to be measured content (%),
Nutrient content in P2=chyme or the ight soil (%),
Salt acid insoluble ash content in C2=chyme or the ight soil,
P1=feed nutrient content to be measured (%).
1.6 data statistics
(SPSS Inc., USA) statistics is handled in the check of the independent sample t of statistical software to test all The data SPSS9.0for Windows.
2 results and discussion
2.1 Pro-Bionate F is to the influence of weanling pig digestive tube chyme microorganism species
As can be seen from Table 10, compare with carbadox, in piglet digestive tube chyme, experimental group makes the intestinal bacteria quantity at positions such as piglet stomach, caecum and colon significantly raise (P＜0.01), but reduce (P＜0.01) at ileum, and do not have significant difference between the two at jejunum.Enterobacteria quantity raises (P＜0.01) significantly at jejunum, caecum and the colon utmost point, and significantly raise at ileum (P＜0.05), and significantly reduce (P＜0.01) at duodenum does not have significant difference between two groups in stomach.Lactobacillus quantity all raises (P＜0.01) than the carbadox group utmost point significantly at stomach, duodenum, jejunum and ileum, and (P＜0.05) significantly raises at caecum.Bifidus bacillus quantity extremely significantly raises (P＜0.01) at stomach, jejunum, ileum and caecum, and (P＜0.05) significantly raises at duodenum and colon.The anerobe sum raises (P＜0.01) significantly at stomach, duodenum, jejunum and the ileum utmost point, in caecum and then significantly rising (P＜0.05) of colon.Muralidhara etc. (1977) find the to feed lactobacillus quantity of intestine of young pigs tissue of lactic acid lactobacillus rises the decline of intestinal bacteria quantity.Report lactobacilluss such as Silva can produce unknown antimicrobial substance and suppress intestinal bacteria.Some research reports show that lactobacillus can reduce enterobacteria and the intestinal bacteria quantity in weanling pig enteron aisle and the ight soil, and aspect the inhibition growth in piglets, these two kinds of bacterium play an important role.This shows that lactobacillus preparation can improve the quantity of lactobacillus, bifidus bacillus, anerobe, intestinal bacteria, enterobacteria and aerobic bacteria in the chyme of the most positions of digestive tube.
Table 10 Pro-Bionate F is to the influence (unit of weanling pig digestive tube chyme microorganism species; 1gCFU/g, weight in wet base)
Annotate: the table intermediate value is a mean+SD.
2.2 Pro-Bionate F is to the influence of nutrient digestion rate
As can be seen from Table 11, compare with carbadox, Pro-Bionate F has significantly improved the all-digestive tract apparent digestibility (P＜0.05) of crude protein and total phosphorus in the daily ration, but all-digestive tract apparent digestibilities such as energy, dry-matter, organism and calcium and amino acid terminal ileum apparent digestibility all had no significant effect (P＞0.05).Phytic acid in the feedstuff raw material is to the inhibition that is absorbed with of phosphorus, and lactic acid can be alleviated this inhibition (Fuller, 1989).Lactobacterium acidophilum has the ability of higher generation lactic acid in this preparation when in-vitro screening.The result shows that the Pro-Bionate is also little to the influence of the nutrient digestion rate of weanling pig.
The compound lactobacillus preparation of table 11 is to the influence (unit: %) of weanling pig daily ration nutrient apparent digestibility
Annotate: the table intermediate value is a mean+SD.
Experimental example 8, different strain liquid carrier adsorb prepared preparation long term storage stability at room temperature
2, experimental technique
Lactobacterium acidophilum CGMCC No.4649 is inoculated in the glucose nutrient broth, its component is: peptone 10g, extractum carnis 5g, sodium-chlor 5g, glucose 2.5g, distilled water 1000ml, and regulate pH to 6.7 with acetic acid, put common incubator 37 degree and cultivate after 18-24 hour, use the sterilized water gradient dilution, obtaining concentration is 10
10The strain liquid of cfu/g.
The strain liquid carrier is respectively powdered rice hulls, wheat bran, skim-milk, maltodextrin.The proportioning of carrier and Lactobacterium acidophilum CGMCC No.4649 is 0.5 gram carrier in the preparation that is made into carrier: 10
9Cfu Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021.
Be made into and measure spawn activity (concentration by bacterial classification in the preparation embodies) behind the preparation at once, placed for 1,6,12,18 week respectively under the normal temperature then after, measure spawn activity more respectively; Calculate and place the per-cent that the back spawn activity accounts for the preceding spawn activity of placement.
3, experimental result
As shown in table 12
Table 12 different strain liquid carrier adsorbs the Journal of Sex Research steady in a long-term of prepared preparation
As seen preparation of the present invention is more stable in carrier.
Claims (10)
1. Lactobacterium acidophilum (Lactobacillus acidophilus) AI2021 is numbered CGMCC No.4649 registering on the books of China Committee for Culture Collection of Microorganisms common micro-organisms center.
2. Pro-Bionate, it is characterized in that: the activeconstituents of described preparation is the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021.
3. preparation according to claim 2 is characterized in that: described preparation has following at least a purposes:
1) product of microecological balance in the animal intestine is reconciled in preparation;
2) preparation strengthens the product of non-specific immunity;
3) improve the animal-feed transformation efficiency;
4) promote digestion and the absorption of animal to nutritive substance;
5) preparation reduces the product of animal diarrhea disease percentage;
6) preparation improves the product of breeding performonce fo animals.
4. according to claim 2 or 3 described preparations, it is characterized in that: described preparation is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and carrier; Described carrier is at least a in calcium phosphate, powdered rice hulls, wheat bran, rice bran, skim-milk, maltodextrin, sucrose, glycerine and the starch.
5. preparation according to claim 4 is characterized in that: in the described preparation, the proportioning of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and described carrier is: (10
7-10
10) cfu: (0.3-5) gram carrier;
Described preparation specifically can be A)-in F) any:
A) the Pro-Bionate A that is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and calcium phosphate, among the described Pro-Bionate A, the proportioning of the two is 5 gram calcium phosphate: (10
7-10
10) the described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021;
B) the Pro-Bionate B that is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and powdered rice hulls, among the described Pro-Bionate B, the proportioning of the two is 0.5 gram powdered rice hulls: (10
7-10
10) the described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021;
C) the Pro-Bionate C that is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate C, the proportioning of the two is the described carrier of 0.3 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls and skim-milk, and the mass ratio of described powdered rice hulls and skim-milk is 2: 1;
D) the Pro-Bionate D that is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate D, the proportioning of the two is the described carrier of 0.3 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls, skim-milk and maltodextrin, and the mass ratio of described powdered rice hulls, skim-milk and maltodextrin is 1: 1: 1;
E) the Pro-Bionate E that is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate E, the proportioning of the two is the described carrier of 0.3 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021; Described carrier is made up of skim-milk and wheat bran, and the mass ratio of described skim-milk and wheat bran is 1: 1;
F) the Pro-Bionate F that is made up of the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 and described carrier, among the described Pro-Bionate F, the proportioning of the two is the described carrier of 0.4 gram: (10
7-10
10) the described Lactobacterium acidophilum of cfu claim 1 (Lactobacillus acidophilus) AI2021; Described carrier is made up of powdered rice hulls, skim-milk and wheat bran, and the mass ratio of described powdered rice hulls, skim-milk and wheat bran is 2: 1: 1.
6. the animal-feed that contains arbitrary described preparation among the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 or the claim 1-5.
7. animal-feed according to claim 6 is characterized in that: in the described animal-feed, contain 10 in the described animal-feed of every gram
7The described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 that CFU is above.
8. the following at least a purposes of arbitrary described preparation among the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 or the claim 1-5:
1) product of microecological balance in the animal intestine is reconciled in preparation;
2) preparation strengthens the product of non-specific immunity;
3) improve the animal-feed transformation efficiency;
4) promote digestion and the absorption of animal to nutritive substance;
5) preparation reduces the product of animal diarrhea disease percentage;
6) preparation improves the product of breeding performonce fo animals.
9. the application of arbitrary described preparation in preparation intestinal bacteria inhibitor among the described Lactobacterium acidophilum of claim 1 (Lactobacillus acidophilus) AI2021 or the claim 1-5.
10. application according to claim 9, the intestinal bacteria in the described intestinal bacteria inhibitor are intestinal bacteria K88, e. coli k99 or intestinal bacteria 987P.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011100951179A CN102212495B (en) | 2011-04-15 | 2011-04-15 | Lactobacillus acidophilus and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011100951179A CN102212495B (en) | 2011-04-15 | 2011-04-15 | Lactobacillus acidophilus and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102212495A true CN102212495A (en) | 2011-10-12 |
| CN102212495B CN102212495B (en) | 2013-04-10 |
Family
ID=44744036
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011100951179A Expired - Fee Related CN102212495B (en) | 2011-04-15 | 2011-04-15 | Lactobacillus acidophilus and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102212495B (en) |
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102640863A (en) * | 2012-02-09 | 2012-08-22 | 西华大学 | Micro-ecological preparation for ornamental fish feed and preparation method of micro-ecological preparation |
| CN103555611A (en) * | 2013-09-27 | 2014-02-05 | 上海源耀生物股份有限公司 | Lactobacillus acidophilus culture feed additive and preparation method thereof |
| CN103555634A (en) * | 2013-11-11 | 2014-02-05 | 山东康地恩生物科技有限公司 | Lactobacillus acidophilus and applications thereof |
| CN103602614A (en) * | 2013-11-11 | 2014-02-26 | 南京曜动节能环保科技有限公司 | Preparation and application of composite microecological feed additive |
| CN103783281A (en) * | 2012-10-30 | 2014-05-14 | 上海市农业科学院 | Health growth-promotion feed additive replacing antibiotic and preparation method thereof |
| CN106011037A (en) * | 2016-08-05 | 2016-10-12 | 福州大学 | Lactobacillus acidophilus and application thereof |
| CN106191044A (en) * | 2016-08-05 | 2016-12-07 | 福建农林大学 | A kind of gene engineering method improving lactein plnK yield |
| CN106350468A (en) * | 2016-09-08 | 2017-01-25 | 济南康多宝生物技术有限公司 | Novel lactobacillus acidophilus |
| CN109717245A (en) * | 2017-10-31 | 2019-05-07 | 内蒙古伊利实业集团股份有限公司 | Only make the probiotics fermention cream and preparation method thereof of leavening with lactobacillus acidophilus |
| CN110229770A (en) * | 2019-07-19 | 2019-09-13 | 福建省农业科学院农业工程技术研究所 | A kind of bacillus subtilis formulation and preparation method |
| CN111575223A (en) * | 2020-05-20 | 2020-08-25 | 江南大学 | Method for reducing secretion of surface substances of lactobacillus rhamnosus |
| CN112266880A (en) * | 2020-10-20 | 2021-01-26 | 青岛普罗百世生物科技有限公司 | Lactobacillus acidophilus and application thereof in weaned pig feed |
| CN115806895A (en) * | 2021-09-16 | 2023-03-17 | 我的轻食有限公司 | Lactobacillus acidophilus TW01 strain, and probiotic composition and application thereof |
| CN116083300A (en) * | 2022-12-08 | 2023-05-09 | 玫斯江苏宠物食品科技有限公司 | Probiotic composite preparation for preventing and treating canine diarrhea and application thereof |
| CN116640705A (en) * | 2023-07-21 | 2023-08-25 | 山东威曼宠物食品有限公司 | Lactobacillus acidophilus King45 for improving pet hair, and microbial inoculum and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101463329A (en) * | 2009-01-07 | 2009-06-24 | 北京龙科方舟生物工程技术中心 | Saccharomyces cerevisiae, yeast preparation including the same, feed, and preparation and use thereof |
| CN101554204A (en) * | 2008-04-11 | 2009-10-14 | 北京龙科方舟生物工程技术中心 | Fermentation lactobacillus preparation |
-
2011
- 2011-04-15 CN CN2011100951179A patent/CN102212495B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101554204A (en) * | 2008-04-11 | 2009-10-14 | 北京龙科方舟生物工程技术中心 | Fermentation lactobacillus preparation |
| CN101463329A (en) * | 2009-01-07 | 2009-06-24 | 北京龙科方舟生物工程技术中心 | Saccharomyces cerevisiae, yeast preparation including the same, feed, and preparation and use thereof |
Non-Patent Citations (2)
| Title |
|---|
| 《中国畜牧杂志》 20030630 黄沧海 益生乳酸杆菌抑制大肠杆菌的研究 p27-28 1-3 第39卷, 第6期 * |
| 《黑龙江畜牧兽医》 20050228 黄沧海 饲用乳酸杆菌菌种益生特性的选育 p21-23 1-3 , 第2期 * |
Cited By (22)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102640863B (en) * | 2012-02-09 | 2013-04-17 | 西华大学 | Preparation method of micro-ecological preparation for ornamental fish feed |
| CN102640863A (en) * | 2012-02-09 | 2012-08-22 | 西华大学 | Micro-ecological preparation for ornamental fish feed and preparation method of micro-ecological preparation |
| CN103783281A (en) * | 2012-10-30 | 2014-05-14 | 上海市农业科学院 | Health growth-promotion feed additive replacing antibiotic and preparation method thereof |
| CN103555611A (en) * | 2013-09-27 | 2014-02-05 | 上海源耀生物股份有限公司 | Lactobacillus acidophilus culture feed additive and preparation method thereof |
| CN103555634A (en) * | 2013-11-11 | 2014-02-05 | 山东康地恩生物科技有限公司 | Lactobacillus acidophilus and applications thereof |
| CN103602614A (en) * | 2013-11-11 | 2014-02-26 | 南京曜动节能环保科技有限公司 | Preparation and application of composite microecological feed additive |
| CN103555634B (en) * | 2013-11-11 | 2015-02-18 | 山东康地恩生物科技有限公司 | Lactobacillus acidophilus and applications thereof |
| CN103602614B (en) * | 2013-11-11 | 2015-09-23 | 南京曜动节能环保科技有限公司 | A kind of preparations and applicatio of composite microorganism type forage additives |
| CN106011037B (en) * | 2016-08-05 | 2019-09-13 | 福州大学 | A kind of lactobacillus acidophilus and its application |
| CN106011037A (en) * | 2016-08-05 | 2016-10-12 | 福州大学 | Lactobacillus acidophilus and application thereof |
| CN106191044A (en) * | 2016-08-05 | 2016-12-07 | 福建农林大学 | A kind of gene engineering method improving lactein plnK yield |
| CN106350468A (en) * | 2016-09-08 | 2017-01-25 | 济南康多宝生物技术有限公司 | Novel lactobacillus acidophilus |
| CN106350468B (en) * | 2016-09-08 | 2019-05-10 | 济南康多宝生物技术有限公司 | A kind of acidophilic lactobacillus |
| CN109717245A (en) * | 2017-10-31 | 2019-05-07 | 内蒙古伊利实业集团股份有限公司 | Only make the probiotics fermention cream and preparation method thereof of leavening with lactobacillus acidophilus |
| CN110229770A (en) * | 2019-07-19 | 2019-09-13 | 福建省农业科学院农业工程技术研究所 | A kind of bacillus subtilis formulation and preparation method |
| CN111575223A (en) * | 2020-05-20 | 2020-08-25 | 江南大学 | Method for reducing secretion of surface substances of lactobacillus rhamnosus |
| CN112266880A (en) * | 2020-10-20 | 2021-01-26 | 青岛普罗百世生物科技有限公司 | Lactobacillus acidophilus and application thereof in weaned pig feed |
| CN115806895A (en) * | 2021-09-16 | 2023-03-17 | 我的轻食有限公司 | Lactobacillus acidophilus TW01 strain, and probiotic composition and application thereof |
| CN116083300A (en) * | 2022-12-08 | 2023-05-09 | 玫斯江苏宠物食品科技有限公司 | Probiotic composite preparation for preventing and treating canine diarrhea and application thereof |
| CN116083300B (en) * | 2022-12-08 | 2023-10-17 | 玫斯江苏宠物食品科技有限公司 | Probiotic composite preparation for preventing and treating canine diarrhea and application thereof |
| CN116640705A (en) * | 2023-07-21 | 2023-08-25 | 山东威曼宠物食品有限公司 | Lactobacillus acidophilus King45 for improving pet hair, and microbial inoculum and application thereof |
| CN116640705B (en) * | 2023-07-21 | 2023-09-19 | 山东威曼宠物食品有限公司 | Lactobacillus acidophilus King45 for improving pet hair, and microbial inoculum and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102212495B (en) | 2013-04-10 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102212495B (en) | Lactobacillus acidophilus and application thereof | |
| CN102304489B (en) | Lactobacillus reuteri strain and application thereof | |
| CN101463329B (en) | Saccharomyces cerevisiae, yeast preparation including the same, feed, and preparation and use thereof | |
| CN102373172B (en) | Enterococcus faecium and application thereof | |
| CN101638627B (en) | Bacillus subtilis and application thereof in biological feed additives | |
| CN102399733B (en) | Lactobacillus johnsonii, microbial inoculum, application and premix thereof | |
| CN102876614B (en) | Bacillus licheniformis and application of bacillus licheniformis | |
| CN102747003B (en) | Screening and application of probiotic Enterococcus faecium | |
| CN102409015B (en) | Composite micro-ecological preparation as well as premixed material and application of preparation in feed additive | |
| CN103392911B (en) | Feed-use high-activity lactobacillus solid preparation and preparation method thereof | |
| CN103981118B (en) | A kind of bacillus subtilis feed addictive and its preparation method and application | |
| CN104293696B (en) | One strain enterococcus faecalis HEW-A131 and application thereof | |
| CN103070285B (en) | Microbe feed additive and preparation method thereof | |
| CN103005159A (en) | Preparation method of ginkgo leaf biological feed additive | |
| CN102226157A (en) | Lactobacillus fermentum and application thereof | |
| CN101974463B (en) | Lactobacillus reuteri and composite viable bacteria preparation thereof | |
| CN105685970A (en) | Compound nutritious food capable of improving whole digestive tract | |
| WO2008023580A1 (en) | Animal feed additive | |
| CN102391962B (en) | Enterococcus faecium preparation agent and manufacturing technology thereof | |
| CN104862254A (en) | Enterococcus faecium HEW-A588 and application thereof | |
| CN105994941A (en) | Non-antibiotic feed prepared through microbial fermentation | |
| CN104673726A (en) | Porcine lactobacillus acidophilus freeze-drying preparation and application thereof | |
| CN109554356A (en) | A kind of mycotoxin biodegradation agent and its application containing glucose oxidase and saccharomyces cerevisiae | |
| CN114806978B (en) | Lactobacillus johnsonii SXDT-23 and application thereof | |
| CN102907585B (en) | A kind of middle pig probiotic vitamin premix and batch |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C53 | Correction of patent of invention or patent application | ||
| CB02 | Change of applicant information |
Address after: 100081 Haidian District, Zhongguancun, South Street, No. 1, No. 12, building, room feed, room 205 Applicant after: BEIJING LONGKEFANGZHOU BIO-ENGINEERING TECHNOLOGY CO.,LTD. Address before: 100193 Beijing Old Summer Palace West Road, Haidian District, No. 2 Applicant before: Peking Sinagri Yin Thai Biotechnology Co.,Ltd. |
|
| COR | Change of bibliographic data |
Free format text: CORRECT: APPLICANT; FROM: BEIJING LONGKE FANGZHOU BIOENGINEERING TECHNOLOGY CENTER TO: BEIJING LONGKE FANGZHOU BIOENGINEERING TECHNOLOGY CO., LTD. |
|
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| EE01 | Entry into force of recordation of patent licensing contract |
Application publication date: 20111012 Assignee: NANCHANG AONONG BIOLOGICAL SCIENCE & TECHNOLOGY Co.,Ltd. Assignor: BEIJING LONGKEFANGZHOU BIO-ENGINEERING TECHNOLOGY CO.,LTD. Contract record no.: 2013990000763 Denomination of invention: Lactobacillus acidophilus and application thereof Granted publication date: 20130410 License type: Exclusive License Record date: 20131118 |
|
| LICC | Enforcement, change and cancellation of record of contracts on the licence for exploitation of a patent or utility model | ||
| EC01 | Cancellation of recordation of patent licensing contract |
Assignee: NANCHANG AONONG BIOLOGICAL SCIENCE & TECHNOLOGY Co.,Ltd. Assignor: BEIJING LONGKEFANGZHOU BIO-ENGINEERING TECHNOLOGY CO.,LTD. Contract record no.: 2013990000763 Date of cancellation: 20180626 |
|
| EC01 | Cancellation of recordation of patent licensing contract | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20130410 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |