CN106432419B - 一种乳酸片球菌来源的抗菌六肽及其制备方法 - Google Patents
一种乳酸片球菌来源的抗菌六肽及其制备方法 Download PDFInfo
- Publication number
- CN106432419B CN106432419B CN201610728511.4A CN201610728511A CN106432419B CN 106432419 B CN106432419 B CN 106432419B CN 201610728511 A CN201610728511 A CN 201610728511A CN 106432419 B CN106432419 B CN 106432419B
- Authority
- CN
- China
- Prior art keywords
- lactein
- antibacterial peptide
- antibacterial
- escherichia coli
- feed
- 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.)
- Active
Links
- 230000000844 anti-bacterial effect Effects 0.000 title abstract description 12
- 238000002360 preparation method Methods 0.000 title abstract description 10
- 241000191998 Pediococcus acidilactici Species 0.000 title abstract description 6
- 239000003910 polypeptide antibiotic agent Substances 0.000 claims abstract description 36
- 241000588724 Escherichia coli Species 0.000 claims abstract description 28
- 241000186779 Listeria monocytogenes Species 0.000 claims abstract description 8
- 241000191967 Staphylococcus aureus Species 0.000 claims abstract description 8
- 241000894006 Bacteria Species 0.000 claims description 24
- 238000000034 method Methods 0.000 claims description 10
- 239000000203 mixture Substances 0.000 claims description 6
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 5
- 239000013598 vector Substances 0.000 claims description 5
- 238000010353 genetic engineering Methods 0.000 claims description 4
- 230000005764 inhibitory process Effects 0.000 claims description 4
- 239000003755 preservative agent Substances 0.000 claims description 4
- 230000002335 preservative effect Effects 0.000 claims description 4
- 201000010099 disease Diseases 0.000 claims description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 3
- 102000040430 polynucleotide Human genes 0.000 claims 2
- 108091033319 polynucleotide Proteins 0.000 claims 2
- 239000002157 polynucleotide Substances 0.000 claims 2
- 238000003745 diagnosis Methods 0.000 claims 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 claims 1
- 239000010931 gold Substances 0.000 claims 1
- 229910052737 gold Inorganic materials 0.000 claims 1
- 230000002401 inhibitory effect Effects 0.000 abstract description 15
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 11
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 abstract description 10
- 244000046052 Phaseolus vulgaris Species 0.000 abstract description 9
- 235000010627 Phaseolus vulgaris Nutrition 0.000 abstract description 9
- -1 DPPH free radical Chemical class 0.000 abstract description 6
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 abstract description 6
- 240000004808 Saccharomyces cerevisiae Species 0.000 abstract description 5
- 239000003242 anti bacterial agent Substances 0.000 abstract description 5
- 229940088710 antibiotic agent Drugs 0.000 abstract description 5
- 230000002000 scavenging effect Effects 0.000 abstract description 5
- OUUQCZGPVNCOIJ-UHFFFAOYSA-M Superoxide Chemical compound [O-][O] OUUQCZGPVNCOIJ-UHFFFAOYSA-M 0.000 abstract description 4
- 230000035755 proliferation Effects 0.000 abstract description 4
- 230000002829 reductive effect Effects 0.000 abstract description 4
- 240000006024 Lactobacillus plantarum Species 0.000 abstract description 3
- 235000013965 Lactobacillus plantarum Nutrition 0.000 abstract description 3
- 229940072205 lactobacillus plantarum Drugs 0.000 abstract description 3
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 22
- 241001465754 Metazoa Species 0.000 description 14
- 230000000694 effects Effects 0.000 description 14
- 230000003385 bacteriostatic effect Effects 0.000 description 10
- 230000003115 biocidal effect Effects 0.000 description 10
- 108090000765 processed proteins & peptides Proteins 0.000 description 9
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 8
- 238000000926 separation method Methods 0.000 description 8
- 239000000126 substance Substances 0.000 description 8
- 238000000855 fermentation Methods 0.000 description 7
- 230000004151 fermentation Effects 0.000 description 7
- 239000007788 liquid Substances 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 238000000108 ultra-filtration Methods 0.000 description 7
- 239000006041 probiotic Substances 0.000 description 6
- 235000018291 probiotics Nutrition 0.000 description 6
- 239000006071 cream Substances 0.000 description 5
- 239000000287 crude extract Substances 0.000 description 5
- 238000002474 experimental method Methods 0.000 description 5
- 238000004108 freeze drying Methods 0.000 description 5
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 5
- 239000011149 active material Substances 0.000 description 4
- 238000005571 anion exchange chromatography Methods 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 239000002054 inoculum Substances 0.000 description 4
- 239000004310 lactic acid Substances 0.000 description 4
- 235000014655 lactic acid Nutrition 0.000 description 4
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 4
- 239000000243 solution Substances 0.000 description 4
- 244000063299 Bacillus subtilis Species 0.000 description 3
- 235000014469 Bacillus subtilis Nutrition 0.000 description 3
- 241000194031 Enterococcus faecium Species 0.000 description 3
- 241000192125 Firmicutes Species 0.000 description 3
- 241000191996 Pediococcus pentosaceus Species 0.000 description 3
- 238000000502 dialysis Methods 0.000 description 3
- 238000001641 gel filtration chromatography Methods 0.000 description 3
- 239000002609 medium Substances 0.000 description 3
- 238000012545 processing Methods 0.000 description 3
- 230000001954 sterilising effect Effects 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 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 2
- 206010059866 Drug resistance Diseases 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 102000004190 Enzymes Human genes 0.000 description 2
- 230000002411 adverse Effects 0.000 description 2
- 230000000845 anti-microbial effect Effects 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 238000004587 chromatography analysis Methods 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 235000014113 dietary fatty acids Nutrition 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 229930195729 fatty acid Natural products 0.000 description 2
- 239000000194 fatty acid Substances 0.000 description 2
- 150000004665 fatty acids Chemical class 0.000 description 2
- 235000013305 food Nutrition 0.000 description 2
- 235000019253 formic acid Nutrition 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 239000012634 fragment Substances 0.000 description 2
- 238000005227 gel permeation chromatography Methods 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 150000002500 ions Chemical class 0.000 description 2
- 244000144972 livestock Species 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 244000000010 microbial pathogen Species 0.000 description 2
- 239000002773 nucleotide Substances 0.000 description 2
- 125000003729 nucleotide group Chemical group 0.000 description 2
- 230000003647 oxidation Effects 0.000 description 2
- 238000007254 oxidation reaction Methods 0.000 description 2
- 239000001301 oxygen Substances 0.000 description 2
- 229910052760 oxygen Inorganic materials 0.000 description 2
- 235000019629 palatability Nutrition 0.000 description 2
- 244000144977 poultry Species 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 102000004196 processed proteins & peptides Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 230000002441 reversible effect Effects 0.000 description 2
- 238000002390 rotary evaporation Methods 0.000 description 2
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Inorganic materials [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 2
- 238000001228 spectrum Methods 0.000 description 2
- 238000003860 storage Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- FFRBMBIXVSCUFS-UHFFFAOYSA-N 2,4-dinitro-1-naphthol Chemical compound C1=CC=C2C(O)=C([N+]([O-])=O)C=C([N+]([O-])=O)C2=C1 FFRBMBIXVSCUFS-UHFFFAOYSA-N 0.000 description 1
- 102000044503 Antimicrobial Peptides Human genes 0.000 description 1
- 108700042778 Antimicrobial Peptides Proteins 0.000 description 1
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 108010062877 Bacteriocins Proteins 0.000 description 1
- 241001478240 Coccus Species 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 102000009123 Fibrin Human genes 0.000 description 1
- 108010073385 Fibrin Proteins 0.000 description 1
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 1
- 241001506991 Komagataella phaffii GS115 Species 0.000 description 1
- 241000186660 Lactobacillus Species 0.000 description 1
- 241000186781 Listeria Species 0.000 description 1
- 241000896292 Odontothrips loti Species 0.000 description 1
- 241000235342 Saccharomycetes Species 0.000 description 1
- 235000009754 Vitis X bourquina Nutrition 0.000 description 1
- 235000012333 Vitis X labruscana Nutrition 0.000 description 1
- 240000006365 Vitis vinifera Species 0.000 description 1
- 235000014787 Vitis vinifera Nutrition 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 239000003674 animal food additive Substances 0.000 description 1
- 238000005349 anion exchange Methods 0.000 description 1
- 230000002421 anti-septic effect Effects 0.000 description 1
- 239000006030 antibiotic growth promoter Substances 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 244000052616 bacterial pathogen Species 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000023555 blood coagulation Effects 0.000 description 1
- 235000021466 carotenoid Nutrition 0.000 description 1
- 150000001747 carotenoids Chemical class 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 239000002131 composite material Substances 0.000 description 1
- 230000009514 concussion Effects 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 238000009313 farming Methods 0.000 description 1
- 235000019197 fats Nutrition 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 229950003499 fibrin Drugs 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 125000002485 formyl group Chemical class [H]C(*)=O 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 238000012826 global research Methods 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 241000411851 herbal medicine Species 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000004957 immunoregulator effect Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- JEIPFZHSYJVQDO-UHFFFAOYSA-N iron(III) oxide Inorganic materials O=[Fe]O[Fe]=O JEIPFZHSYJVQDO-UHFFFAOYSA-N 0.000 description 1
- 229940039696 lactobacillus Drugs 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 238000005453 pelletization Methods 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 235000013406 prebiotics Nutrition 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 150000003254 radicals Chemical class 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 108091008146 restriction endonucleases Proteins 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
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 239000013589 supplement Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 235000021081 unsaturated fats Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 150000003722 vitamin derivatives Chemical class 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N37/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids
- A01N37/18—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing the group —CO—N<, e.g. carboxylic acid amides or imides; Thio analogues thereof
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N37/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids
- A01N37/44—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing at least one carboxylic group or a thio analogue, or a derivative thereof, and a nitrogen atom attached to the same carbon skeleton by a single or double bond, this nitrogen atom not being a member of a derivative or of a thio analogue of a carboxylic group, e.g. amino-carboxylic acids
- A01N37/46—N-acyl derivatives
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/50—Isolated enzymes; Isolated proteins
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
- A23B4/00—General methods for preserving meat, sausages, fish or fish products
- A23B4/14—Preserving with chemicals not covered by groups A23B4/02 or A23B4/12
- A23B4/18—Preserving with chemicals not covered by groups A23B4/02 or A23B4/12 in the form of liquids or solids
- A23B4/20—Organic compounds; Microorganisms; Enzymes
- A23B4/22—Microorganisms; Enzymes; Antibiotics
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
- A23C9/00—Milk preparations; Milk powder or milk powder preparations
- A23C9/152—Milk preparations; Milk powder or milk powder preparations containing additives
- A23C9/1526—Amino acids; Peptides; Protein hydrolysates; Nucleic acids; Derivatives thereof
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/142—Amino acids; Derivatives thereof
- A23K20/147—Polymeric derivatives, e.g. peptides or proteins
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K20/00—Accessory food factors for animal feeding-stuffs
- A23K20/10—Organic substances
- A23K20/195—Antibiotics
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
- A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
- A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
- A23L3/3463—Organic compounds; Microorganisms; Enzymes
- A23L3/34635—Antibiotics
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
- A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
- A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
- A23L3/3463—Organic compounds; Microorganisms; Enzymes
- A23L3/3526—Organic compounds containing nitrogen
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/26—Selective adsorption, e.g. chromatography characterised by the separation mechanism
- B01D15/32—Bonded phase chromatography
- B01D15/325—Reversed phase
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/26—Selective adsorption, e.g. chromatography characterised by the separation mechanism
- B01D15/34—Size selective separation, e.g. size exclusion chromatography, gel filtration, permeation
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/26—Selective adsorption, e.g. chromatography characterised by the separation mechanism
- B01D15/36—Selective adsorption, e.g. chromatography characterised by the separation mechanism involving ionic interaction
- B01D15/361—Ion-exchange
- B01D15/363—Anion-exchange
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D61/00—Processes of separation using semi-permeable membranes, e.g. dialysis, osmosis or ultrafiltration; Apparatus, accessories or auxiliary operations specially adapted therefor
- B01D61/14—Ultrafiltration; Microfiltration
- B01D61/145—Ultrafiltration
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D61/00—Processes of separation using semi-permeable membranes, e.g. dialysis, osmosis or ultrafiltration; Apparatus, accessories or auxiliary operations specially adapted therefor
- B01D61/58—Multistep processes
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2400/00—Lactic or propionic acid bacteria
- A23V2400/41—Pediococcus
- A23V2400/413—Acidilactici
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Health & Medical Sciences (AREA)
- Polymers & Plastics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Zoology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Food Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Wood Science & Technology (AREA)
- Analytical Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Dentistry (AREA)
- Plant Pathology (AREA)
- Pest Control & Pesticides (AREA)
- Agronomy & Crop Science (AREA)
- Environmental Sciences (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Nutrition Science (AREA)
- Water Supply & Treatment (AREA)
- Animal Husbandry (AREA)
- Medicinal Chemistry (AREA)
- Genetics & Genomics (AREA)
- Biophysics (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Fodder In General (AREA)
Abstract
本发明公开了一种乳酸片球菌来源的抗菌六肽及其制备方法,属于生物工程领域。本发明的抗菌肽ENGEEE是首次被报道。本发明的抗菌肽乳酸菌素R16的pH稳定性和热稳定性好,对大肠杆菌、单增李斯特菌、金黄色葡萄球菌均有抑制作用,能够有效降低豆粕中大肠杆菌的数量。其次,抗菌肽乳酸菌素R16对于酿酒酵母和植物乳杆菌具有一定的增殖作用,对过氧化氢、羟自由基、DPPH·自由基和超氧阴离子具有一定的清除能力。本发明的抗菌肽乳酸菌素R16可作为生物防治使用,可作为饲料添加剂使用,在将来替代抗生素和解决饲料安全问题中发挥重要作用。
Description
技术领域
本发明涉及一种乳酸片球菌来源的抗菌六肽及其制备方法,属于生物工程领域。
背景技术
目前,亚治疗剂量的抗生素作为饲料添加剂被广泛应用于禽畜饲料中,其在促进动物生长、预防某些疾病发生方面发挥着重要作用,但是,随着抗生素滥用事件频繁发生,让人们对抗生素的副作用有了进一步的认识。滥用抗生素不仅会引起动物内源性感染,产生耐药菌株,而且会使畜禽免疫功能下降,更严重的是,残留于畜禽产品中的抗生素会通过不同渠道而流入人体,并对人体造成危害。
欧盟监管委员会决定,自2006年1月起在动物养殖业中禁用抗生素生长促进剂。从2013年12月开始,美国FDA发布了《兽医饲料指令》,要求有执照的兽医监督抗生素的使用,计划从2014年起,用3年时间禁止在牲畜饲料中使用预防性抗生素,最大限度地减少食用牲畜带给消费者抗生素耐药性问题。韩国也将在2018年7月全面禁止饲用抗生素的使用。
随着细菌对抗生素耐药性和抗生素的残留问题日益严重,研究和开发绿色饲料添加剂产品已经成为世界性的研究课题,大量研究表明,中草药提取物、微生物制剂、酶制剂、益生元、抗菌肽、酸化剂等新型饲料添加剂能有效减少或者替代饲用抗生素的使用。其中,抗菌肽由于具有较好的抗菌和免疫调节的活性,且不易残留、没有副作用、没有抗药性,对环境不产生不良影响等优点,成为了抗生素潜在的有效替代物。
发明内容
为了解决上述问题,本发明从能够抑制大肠杆菌生长的乳酸菌的发酵液中分离得到一种对细菌具有抑制作用的乳酸菌素,通过液相色谱-质谱联用技术(liquidchromatography-mass spectrometry,LC-MS)初步鉴定该乳酸菌素的分子量大小为705.63Da,等电点为3.58,氨基酸序列为ENGEEE。本发明还通过化学合成的方法确认该乳酸菌素对多种革兰氏阳性菌具有抑制作用,将其命名为乳酸菌素R16。
本发明的第一个目的是提供一种抗菌肽,所述抗菌肽的氨基酸序列为:ENGEEE(如SEQ ID NO.1所示),命名为乳酸菌素R16。
所述抗菌肽,是从能够抑制大肠杆菌生长的乳酸菌的发酵液中分离得到的。
本发明的第二个目的是提供编码所述抗菌肽的核苷酸片段。
本发明的第三个目的是提供一种能够表达所述抗菌肽的重组载体或者基因工程菌。比如,可以将若干(如20)个该抗菌肽的编码基因首尾连接,在每两段基因中间添加凝血酶酶切位点,整段基因与载体连接得到重组载体,转化入宿主菌表达。表达产物经凝血酶酶切即可得到该抗菌肽。
在本发明的一种实施方式中,所述载体可以是pPIC9K。
在本发明的一种实施方式中,所述宿主菌可以是毕赤酵母GS115。
本发明的第四个目的是提供含有所述抗菌肽的组合物。
在本发明的一种实施方式中,所述组合物为饲料添加剂、饲料、生物防腐剂。
本发明的第五个目的是提供所述抗菌肽、编码所述抗菌肽的核苷酸片段、能够表达所述抗菌肽的重组载体或者能够表达所述抗菌肽的基因工程菌的应用。
在本发明的一种实施方式中,所述应用,是作为饲料添加剂或者生物防腐剂。
本发明的第六个目的是提供一种抑制细菌的方法,是使用所述的抗菌肽。
在本发明的一种实施方式中,所述细菌为革兰氏阳性菌或革兰氏阴性菌。
在本发明的一种实施方式中,所述细菌包括:大肠杆菌、单增李斯特菌、金黄色葡萄球菌等。
本发明还提供一种所述抗菌肽的制备方法,是将乳酸片球菌发酵得到的发酵上清液先由超滤进行粗提,然后经阴离子交换色谱柱Hitrap Q FF和凝胶过滤色谱SuperduxPeptide 10/300GL分离,最后用反相柱Phenomenex Luna/C18分离纯化得到抗菌六肽。
在本发明的一种实施方式中,所述超滤是超滤(截留分子量3000Da)浓缩8~10倍,得到粗提液。
在本发明的一种实施方式中,所述制备方法具体是:
(1)超滤粗提:超滤(截留分子量3000Da)浓缩8~10倍,得到粗提液;
(2)经阴离子交换色谱Hitrap Q FF的分离:粗提液上样于Hitrap Q FF色谱柱分离得到2个洗脱峰,收集对大肠杆菌有抑菌作用的活性峰L2,透析后冷冻干燥;
(3)经凝胶过滤色谱Superdux Peptide 10/300GL的分离:将阴离子交换色谱的洗脱峰L2上样于Superdux Peptide 10/300GL凝胶过滤层析柱,分离得到2个洗脱峰,收集对大肠杆菌有抑菌作用的活性峰N2,透析后冷冻干燥;
(4)反相C18柱分离:将上一步冷冻干燥的样品用5%的乙腈(含0.1%TFA,下同)溶解,并经5%的乙腈平衡的Phenomenex Luna/C18柱,用浓度为5-50%的线性梯度递增的乙腈洗脱,收集活性洗脱峰Ⅱ,并经旋转蒸发和冷冻干燥后,得到最终产品。
本发明的有益效果:
(1)本发明的抗菌肽ENGEEE,与抗菌肽数据库(TheAntimicrobial PeptideDatabase)等数据库比对,此种抗菌肽是首次被报道。
(2)本发明利用超滤和色谱等技术,制备对革兰氏阳性菌具有明显抗性的活性肽乳酸菌素R16。本发明的制备工艺简单、分离效果好、操作条件温和、快速高效、产品纯度高,经MS鉴定其纯度可达95%以上。
(3)本发明中的获得的抗菌肽乳酸菌素R16的pH稳定性和热稳定性好,对大肠杆菌和单增李斯特菌的最小抑菌浓度为6.4mg·mL-1,对金黄色葡萄球菌的最小抑菌浓度为3.4mg·mL-1。
(4)本发明的抗菌肽乳酸菌素R16对于酿酒酵母和植物乳杆菌具有一定的增殖作用,对其他对益生菌的影响很小。
(5)本发明的抗菌肽乳酸菌素R16对过氧化氢、羟自由基、DPPH·自由基和超氧阴离子具有一定的清除能力,清除率分别为25.65%、70.13%、79.42%和10.11%。
(6)本发明的抗菌肽乳酸菌素R16可作为生物防治使用,是对已测氨基酸序列的乳酸菌素库的有效补充。可作为饲料添加剂使用,在将来替代抗生素和解决饲料安全问题中发挥重要作用。
附图说明
图1:六肽的质谱鉴定图;
图2:不同pH值对乳酸菌素R16的影响;
图3:温度对乳酸菌素R16的影响;
图4:乳酸菌素R16对益生菌生长的影响;
图5:抑菌活性物质对羟自由基、DPPH自由基和超氧阴离子自由基的清除能力。
具体实施方式
本发明所涉及的各种缓冲液配方均可以在实验手册中查到。
实施例1:乳酸菌素R16的制备
按照下面的方法制备抗菌六肽乳酸菌素R16:
1.抗菌六肽经乳酸片球菌发酵而来,发酵上清液经离心之后,收集上清液超滤(截留分子量10000Da)浓缩8~10倍,得到粗提液。
2.经阴离子交换色谱Hitrap Q FF的分离:粗提液上样于Hitrap Q FF色谱柱分离得到2个洗脱峰,收集对大肠杆菌有抑菌作用的活性峰,并将收集的活性峰L2经Mill-Q水充分透析后冷冻干燥。
3.经凝胶过滤色谱Superdux Peptide 10/300GL的分离:将阴离子交换色谱的洗脱峰L2上样于Superdux Peptide 10/300GL凝胶过滤层析柱,分离得到2个洗脱峰,收集对大肠杆菌有抑菌作用的活性峰,并将收集的活性峰N2经Mill-Q水充分透析后冷冻干燥。
4.反相C18柱的分离:将上一步冷冻干燥的样品用5%的乙腈(含0.1%TFA,下同)溶解,并经5%的乙腈平衡的C18柱,用浓度为5-50%的线性梯度递增的乙腈洗脱,收集活性洗脱峰H3,并经旋转蒸发和冷冻干燥后,最终得到产品。
5.LC-MS的鉴定:将上述步骤得到的具有抑菌活性的冷冻干燥样品,用Mill-Q水溶解后经LC-MS鉴定,流动相为A(乙腈:水:甲酸=30:970:1(V:V:V))和B(乙腈:水:甲酸=700:300:1(V:V:V));洗脱程序为0-10min,100%A,0%B;20min,70%A,30%B;30min,0%A,100%B;35min,100%A,0%B;flow rate,1mL/min;柱温30℃。质谱条件:毛细管电压,3.88kV;圆锥体电压,20V;离子源温度,120℃;去溶温度,300℃;流速,1mL/min;分流比,50:1。结果用软件MassLynx 4.1分析。
质谱鉴定此抗菌肽的分子量为705.17Da,其氨基酸序列为E-N-G-E-E-E(序列如SEQ ID NO.1所示)。
经与抗菌肽数据库(The Antimicrobial Peptide Database)等数据库比对,此种抗菌肽并未见报道。
实施例2:乳酸菌素R16的生物学特性
(1)乳酸菌素R16在pH中的稳定性
取10份浓度为100mg·mL-1纯化过后的活性物质乳酸菌素R16,用1.0mol·L-1NaOH和1.0mol·L-1HCl将其pH分别调成1.0~10.0,置于37℃的水浴锅处理2h,后将其pH调至4.5[62],做抑菌实验,观察其对大肠杆菌的抑制情况。结果如图2所示。
图2显示,在pH 3.0~5.0时,乳酸菌素R16的抑菌活性达到最大。在pH 6.0~8.0时,乳酸菌素R16的抑菌活性逐步降低,且降低趋势缓慢,残留活性维持在75%左右,说明乳酸菌素R16在此pH条件下稳定性有一定降低,但此pH对乳酸菌素R16的稳定性影响不大。
(2)乳酸菌素R16的热稳定性
取8份浓度为100mg·mL-1纯化过后的活性物质乳酸菌素R16,分别置于40、50、60、70、80、90、100℃中处理30min,121℃高压灭菌15min,做抑菌实验,观察其对大肠杆菌的抑制情况。结果如图3所示。
从图3可以看出,乳酸菌素R16具有一定的热稳定性,在40、50、60、70、80℃处理30min,乳酸菌素R16的抑菌活性无明显变化,在90℃和100℃处理30min后,乳酸菌素R16的抑菌活性仅仅分别损失了21.95%和49.47%。121℃处理15min,乳酸菌素R16的抑菌活性损失了59.98%。在温度为75~85℃,持续时间为1~2min的制粒过程中,其活性能够达到100%的保留。
(3)乳酸菌素R16的抑菌谱
取8份浓度为100mg·mL-1纯化过后的活性物质乳酸菌素R16,运用牛津杯法,检测其对大肠杆菌、单增李斯特菌、金黄色葡萄球菌的抑制作用。
饲料在运输、储存过程中容易携带大肠杆菌、单增李斯特菌等病原微生物,这些病原微生物随着饲料进入动物体内,会引起动物肠道传染病,对动物的健康构成严重威胁。乳酸菌素R16对常见动物致病菌的抑制作用如表1所示。
表1乳酸菌素R16的抑菌谱
注:“+++”为强抑制;“++”为较强抑制;“+”为抑制;“-”为无抑制
从表1可知,乳酸菌素R16对金黄色葡萄球菌、大肠杆菌K99,大肠杆菌JM109和单增李斯特菌表现出较强的抑菌活性,其中对金黄色葡萄球菌的抑菌效果最强。
(4)乳酸菌素R16的最低抑菌浓度
采用最小抑菌浓度法(minimum inhibitory concentration,MIC)检测抗菌活性,操作如下:
(a)将菌种按1%接种量接种于LB液体培养基中,37℃,200r·min-1摇床培养过夜。
(b)将上述培养液按1%接种量接种于20mL LB液体培养基中,37℃,200r·min-1摇床培养至OD600=0.5左右。
(c)将OD600=0.5的菌悬液按1‰接种量接种于10mL LB液体培养基中,震荡摇匀,使细菌密度在1×105~5×105CFU·mL-1,测定MIC。
(d)将抑菌活性物质进行梯度稀释,使其终浓度为256、128、64、32、16、8、4、2和1mg·mL-1。
(e)向无菌组织培养板中加入450μL制备好的菌悬液,再分别加入50μL相应浓度的抑菌活性物质稀释液,待测抑菌活性物质的浓度分别为25.6、12.8、6.4、3.2、1.6、0.8、0.4、0.2和0.1mg·mL-1。以500μL菌悬液为阳性对照,以500μL LB液体培养基为阴性对照,37℃恒温培养箱培养18~24h。
(f)观察各个孔底部是否有细菌沉淀,无肉眼可见细菌沉淀的最小浓度为该抑菌活性物质的MIC。
乳酸菌素R16对大肠杆菌,单增李斯特菌和金黄色葡萄球菌的最小抑菌浓度如表2所示,从表2可以看出抑菌活性物质对大肠杆菌和单增李斯特菌的最小抑菌浓度为6.4mg·mL-1,对金黄色葡萄球菌的最小抑菌浓度为3.2mg·mL-1。
表2乳酸菌素R16对菌种的最低抑菌浓度
(5)乳酸菌素R16对益生菌的影响
将各益生菌进行二级活化,按1%的接种量接种于含有抑菌活性物质为6.4mg·mL-1的MRS培养基中,以接种于MRS培养基中的益生菌为对照,37℃恒温培养24h。采用活菌计数法进行涂板计数。结果如图4所示。
微生态制剂作为绿色饲料添加剂的一种,被广泛应用于饲料中,其中以酵母菌,乳酸菌和芽孢菌构成的复合型微生态制剂更为常见,其不仅能够降解和软化粗纤维,提高蛋白含量,还能够改善饲料的适口性,调节动物肠道微生态平衡,促进消化吸收和生长。本发明选取酿酒酵母、戊糖片球菌、屎肠球菌,植物乳酸菌和枯草芽孢杆菌作为研究对象,研究了乳酸菌素R16对其的影响,结果见图4所示。
由图4可知,乳酸菌素R16对酿酒酵母和植物乳杆菌有一定的增殖作用,其中对酿酒酵母的增殖作用更为明显,酿酒酵母的数量与对照相比,增加了24.50%。乳酸菌素R16对屎肠球菌和枯草芽孢杆菌有一定的抑制作用,但抑制幅度较低,屎肠球菌和枯草芽孢杆菌与对照相比,其细菌总数分别减少了6.92%和4.03%,且在生长24h之后,其细菌总数都能够达到108CFU·mL-1左右。与对照相比,戊糖片球菌的细菌总数无明显的变化,可见乳酸菌素R16对戊糖片球菌无影响。
(6)乳酸菌素R16的抗氧化能力
饲料中存在一定量的不饱和脂肪、脂肪酸、脂溶性维生素、类胡萝卜素及其他脂溶性物质,在饲料存放过程中极易被氧化,特别是脂肪和脂肪酸,其会被氧化酸败产生醛、醇、脂、酸等不同化合物,这些物质可能对产生各种异味,严重影响饲料的适口性,有些化合物本身具有一定的毒性,对动物会产生不良影响。乳酸菌素R16对过氧化氢、羟自由基,DPPH·自由基和超氧阴离子的抗氧化能力如图5所示。
从图5可见,乳酸菌素R16对过氧化氢、羟自由基,DPPH·自由基和超氧阴离子都有一定的清除能力,其中对DPPH·自由基的清除的能力最强,清除率达到了79.42%,其次是羟自由基,清除率为70.13%,对超氧阴离子和过氧化氢的清除能力最差,清除率仅为10.11%和25.65%。
实施例3:乳酸菌素R16的应用
应用方法:
(A)将豆粕用刀片粉粹机进行粉粹,取4份,每份40g加入500mL发酵瓶中,编号为组5,组6,组7,组8,于105℃灭菌10min。
(B)发酵条件见表3。
表3含乳酸菌素R16的豆粕初始发酵条件
(C)采用饲料中大肠菌群的测定国标测定大肠杆菌的数量。
通过向灭菌过后的豆粕中添加大肠杆菌,研究了乳酸菌素R16对豆粕中大肠杆菌的抑制效果。结果显示,在没有添加乳酸菌素R16时,大肠杆菌的数量由发酵前1380个·100g-1增加到了发酵后4×106个·100g-1,在添加了乳酸菌素R16之后,大肠杆菌的数量由发酵前1380个·100g-1降低到了123个·100g-1,说明,乳酸菌素R16在有效降低大肠杆菌数量的同时,能够大幅度的去除豆粕中的大肠杆菌,其去除率达到了33.44%。
考虑到工业乳酸菌发酵豆粕与实验室有一定区别,本发明在原有的实验基础上,将1:0.8的料水比改为1:0.12,其他实验条件保持不变,进行了豆粕发酵实验,结果显示乳酸菌素R16对豆粕中大肠杆菌的去除率由33.44%增加到了56.64%,这说明工业条件下,乳酸菌素R16更能够充分作用于大肠杆菌。
实施例4:乳酸菌素R16的应用
乳酸菌素作为目前被唯一允许使用在食品防腐的细菌素,可以被添加到牛奶、肉制品等食品中抑制杂菌生长,延长货架期等应用。
虽然本发明已以较佳实施例公开如上,但其并非用以限定本发明,任何熟悉此技术的人,在不脱离本发明的精神和范围内,都可做各种的改动与修饰,因此本发明的保护范围应该以权利要求书所界定的为准。
Claims (7)
1.一种抗菌肽,其特征在于,所述抗菌肽的氨基酸序列如SEQ ID NO.1所示。
2.一种组合物,其特征在于,所述组合物为含有权利要求1所述抗菌肽的饲料添加剂、饲料或者生物防腐剂。
3.编码权利要求1所述的抗菌肽的多核苷酸。
4.一种能够表达权利要求1所述的抗菌肽的重组载体或者基因工程菌。
5.权利要求1所述的抗菌肽、权利要求3所述的多核苷酸、权利要求4所述的重组载体或者基因工程菌在制备饲料添加剂或者生物防腐剂中的应用。
6.一种不以疾病的诊断或治疗为目的的抑制细菌的方法,其特征在于,所述方法是使用权利要求1所述的抗菌肽。
7.根据权利要求6所述的方法,其特征在于,所述细菌包括大肠杆菌、单增李斯特菌、金黄色葡萄球菌。
Priority Applications (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201610728511.4A CN106432419B (zh) | 2016-08-25 | 2016-08-25 | 一种乳酸片球菌来源的抗菌六肽及其制备方法 |
US15/746,596 US10487112B2 (en) | 2016-08-25 | 2016-09-08 | Pediococcus acidilactici derived antimicrobial hexapeptide and preparation method thereof |
PCT/CN2016/098420 WO2018035892A1 (zh) | 2016-08-25 | 2016-09-08 | 一种乳酸片球菌来源的抗菌六肽及其制备方法 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201610728511.4A CN106432419B (zh) | 2016-08-25 | 2016-08-25 | 一种乳酸片球菌来源的抗菌六肽及其制备方法 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN106432419A CN106432419A (zh) | 2017-02-22 |
CN106432419B true CN106432419B (zh) | 2019-02-22 |
Family
ID=58182256
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN201610728511.4A Active CN106432419B (zh) | 2016-08-25 | 2016-08-25 | 一种乳酸片球菌来源的抗菌六肽及其制备方法 |
Country Status (3)
Country | Link |
---|---|
US (1) | US10487112B2 (zh) |
CN (1) | CN106432419B (zh) |
WO (1) | WO2018035892A1 (zh) |
Families Citing this family (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN107043769A (zh) * | 2017-05-11 | 2017-08-15 | 福建农林大学 | 一种提高乳酸菌素plnJ的产量的基因工程方法 |
CN108277179B (zh) * | 2018-03-07 | 2021-08-03 | 武汉生物样本库有限公司 | 一种枯草芽孢杆菌的高密度发酵及喷雾干燥方法 |
KR102421280B1 (ko) * | 2019-12-23 | 2022-07-15 | 이금남 | 반추동물용 해바라기 조사료, 이의 제조방법 및 해바라기 조사료를 이용한 반추동물 사육방법 |
CN110982745B (zh) * | 2019-12-24 | 2021-06-25 | 扬州大学 | 一种戊糖片球菌z-1、戊糖片球菌细菌素z-1及戊糖片球菌细菌素z-1的生产方法 |
TW202210628A (zh) * | 2020-06-02 | 2022-03-16 | 日商朝日集團控股股份有限公司 | 乳酸菌增殖促進劑 |
CN112851751B (zh) * | 2020-12-09 | 2024-01-16 | 江苏海洋大学 | 抗菌色天光肽及其制备方法与用途 |
CN113861280A (zh) * | 2021-08-24 | 2021-12-31 | 晨光生物科技集团股份有限公司 | 一种棉籽球蛋白正电荷抗菌肽及其应用 |
CN114516900B (zh) * | 2022-01-27 | 2023-06-16 | 中国海洋大学 | 抗菌肽yhx-4及其应用 |
CN115093991B (zh) * | 2022-06-08 | 2023-05-26 | 中国科学院西北生态环境资源研究院 | 一种抗氧化节杆菌及其次级代谢产物的制备和应用 |
CN117160254B (zh) * | 2023-10-31 | 2024-01-23 | 湖南沁森高科新材料有限公司 | 一种反渗透复合膜及其制备方法 |
Family Cites Families (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20040002591A1 (en) * | 1997-09-05 | 2004-01-01 | Human Genome Sciences, Inc. | 50 human secreted proteins |
US7280923B2 (en) * | 2002-10-18 | 2007-10-09 | Regents Of The University Of California | Methods for crystallographic structure determination employing hydrogen exchange analysis |
US9353359B2 (en) * | 2012-12-26 | 2016-05-31 | Yeda Research And Development Co. Ltd. | Bacterial anti-phage defense systems |
WO2016044683A1 (en) * | 2014-09-19 | 2016-03-24 | Tensive Controls, Inc. | Anti-microbial peptides |
CN105816912A (zh) * | 2016-03-09 | 2016-08-03 | 上海交通大学医学院附属第九人民医院 | 一种抗感染和促进成骨分化的抗菌肽修饰钛合金假体及其制备方法 |
-
2016
- 2016-08-25 CN CN201610728511.4A patent/CN106432419B/zh active Active
- 2016-09-08 US US15/746,596 patent/US10487112B2/en not_active Expired - Fee Related
- 2016-09-08 WO PCT/CN2016/098420 patent/WO2018035892A1/zh active Application Filing
Non-Patent Citations (2)
Title |
---|
Pediocin SA-1,an antimicrobial peptide from Pediococcus acidilactici NRRL B5627:Production conditions,purification and characterization;Sofia Anatasiadou等;《Bioresource Technology》;20080930;第99卷(第13期);第5384-5390页 |
乳酸菌抗菌肽(细菌素)抗菌机理的研究进展;任璐雅等;《中国食品添加剂》;20151231(第1期);第143-149页 |
Also Published As
Publication number | Publication date |
---|---|
US20190092811A1 (en) | 2019-03-28 |
CN106432419A (zh) | 2017-02-22 |
US10487112B2 (en) | 2019-11-26 |
WO2018035892A1 (zh) | 2018-03-01 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN106432419B (zh) | 一种乳酸片球菌来源的抗菌六肽及其制备方法 | |
CN105087423A (zh) | 新型芽孢杆菌cjbv及含有其的抗菌组合物 | |
JP5931064B2 (ja) | 新規乳酸菌及びこれを用いるサイレージ又は発酵飼料の調製方法 | |
Fernandez et al. | Enrichment and preservation of a vegetable smoothie with an antioxidant and antimicrobial extract obtained from beet by-products | |
JP6595761B2 (ja) | バチルスアミロリキファシエンス菌株及び該菌株を含む抗菌組成物 | |
CN105176874A (zh) | 凝结芽孢杆菌fm603及其应用 | |
CN105175518A (zh) | 凝结芽孢杆菌fm603产生的细菌素及其制备方法 | |
US10820612B2 (en) | Method for producing fermented bamboo extract and immunostimulating agent | |
CN105613707A (zh) | 枯草芽孢杆菌生物保鲜剂及在大黄鱼保鲜中的应用 | |
KR20160136718A (ko) | 황기 발효산물 및 그것의 제조 방법 | |
KR101374696B1 (ko) | 스타필로코코스 파스트리 rsp―1 및 이의 용도 | |
CN110547384B (zh) | 一种小黄鱼骨胶原抗菌肽及其应用 | |
US11077170B2 (en) | Antimicrobial peptide from skate skin and uses thereof | |
KR100862649B1 (ko) | 기능성 김과 그의 제조방법 | |
CN105104712A (zh) | 一种微生物饲料添加剂及其制备方法 | |
KR20180120660A (ko) | 탱자 발효 추출물을 포함하는 항진균 및 항균 조성물 및 이의 제조방법 | |
CN108968040A (zh) | 一种番茄酵素饮料及其制备方法 | |
CN101849608B (zh) | 油菜饼抗菌肽 | |
CN112375799A (zh) | 一种提高鲍鱼生物制品抗氧化能力的方法 | |
CN105494625B (zh) | 一种抑菌保护剂以及制备方法和应用 | |
KR102677601B1 (ko) | 장대 품종의 케나프 종자 추출물을 포함하는 항균 조성물 | |
KR102191432B1 (ko) | 유해세균에 대해 항균 활성을 갖는 락토바실러스 타이완엔시스 균주 및 이의 용도 | |
CN105901336A (zh) | 从细菌发酵液中富集抗菌肽的方法及一种抗菌肽产品 | |
KR102287391B1 (ko) | 모링가 지하부 추출물을 포함하는 항균 조성물 | |
KR20230077110A (ko) | 표고버섯 수확 후의 버섯폐배지 추출물을 포함하는 항균 조성물 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
C06 | Publication | ||
PB01 | Publication | ||
C10 | Entry into substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
GR01 | Patent grant | ||
GR01 | Patent grant |