CA3001389A1 - Feed additives for aquatic animals comprising essential oils and lysozyme - Google Patents
Feed additives for aquatic animals comprising essential oils and lysozyme Download PDFInfo
- Publication number
- CA3001389A1 CA3001389A1 CA3001389A CA3001389A CA3001389A1 CA 3001389 A1 CA3001389 A1 CA 3001389A1 CA 3001389 A CA3001389 A CA 3001389A CA 3001389 A CA3001389 A CA 3001389A CA 3001389 A1 CA3001389 A1 CA 3001389A1
- Authority
- CA
- Canada
- Prior art keywords
- amino acids
- lysozyme
- alpha
- feed
- polypeptide
- 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.)
- Abandoned
Links
- 235000010335 lysozyme Nutrition 0.000 title claims abstract description 100
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 title claims abstract description 96
- 108010014251 Muramidase Proteins 0.000 title claims abstract description 95
- 239000004325 lysozyme Substances 0.000 title claims abstract description 95
- 229960000274 lysozyme Drugs 0.000 title claims abstract description 95
- 241001465754 Metazoa Species 0.000 title claims abstract description 54
- 102000016943 Muramidase Human genes 0.000 title claims abstract 17
- 239000003674 animal food additive Substances 0.000 title claims description 6
- 239000000341 volatile oil Substances 0.000 title description 22
- 235000019688 fish Nutrition 0.000 claims abstract description 75
- 241000251468 Actinopterygii Species 0.000 claims abstract description 74
- 239000000203 mixture Substances 0.000 claims abstract description 59
- GRWFGVWFFZKLTI-UHFFFAOYSA-N α-pinene Chemical compound CC1=CCC2C(C)(C)C1C2 GRWFGVWFFZKLTI-UHFFFAOYSA-N 0.000 claims abstract description 52
- 230000000694 effects Effects 0.000 claims abstract description 49
- PXIKRTCSSLJURC-UHFFFAOYSA-N Dihydroeugenol Chemical compound CCCC1=CC=C(O)C(OC)=C1 PXIKRTCSSLJURC-UHFFFAOYSA-N 0.000 claims abstract description 41
- 108090000765 processed proteins & peptides Proteins 0.000 claims abstract description 39
- 229920001184 polypeptide Polymers 0.000 claims abstract description 36
- 102000004196 processed proteins & peptides Human genes 0.000 claims abstract description 36
- RRAFCDWBNXTKKO-UHFFFAOYSA-N eugenol Chemical compound COC1=CC(CC=C)=CC=C1O RRAFCDWBNXTKKO-UHFFFAOYSA-N 0.000 claims abstract description 33
- KJPRLNWUNMBNBZ-QPJJXVBHSA-N (E)-cinnamaldehyde Chemical compound O=C\C=C\C1=CC=CC=C1 KJPRLNWUNMBNBZ-QPJJXVBHSA-N 0.000 claims abstract description 31
- KJPRLNWUNMBNBZ-UHFFFAOYSA-N cinnamic aldehyde Natural products O=CC=CC1=CC=CC=C1 KJPRLNWUNMBNBZ-UHFFFAOYSA-N 0.000 claims abstract description 29
- 229940117916 cinnamic aldehyde Drugs 0.000 claims abstract description 29
- MOYAFQVGZZPNRA-UHFFFAOYSA-N Terpinolene Chemical compound CC(C)=C1CCC(C)=CC1 MOYAFQVGZZPNRA-UHFFFAOYSA-N 0.000 claims abstract description 28
- GRWFGVWFFZKLTI-IUCAKERBSA-N 1S,5S-(-)-alpha-Pinene Natural products CC1=CC[C@@H]2C(C)(C)[C@H]1C2 GRWFGVWFFZKLTI-IUCAKERBSA-N 0.000 claims abstract description 26
- MVNCAPSFBDBCGF-UHFFFAOYSA-N alpha-pinene Natural products CC1=CCC23C1CC2C3(C)C MVNCAPSFBDBCGF-UHFFFAOYSA-N 0.000 claims abstract description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 24
- RLSSMJSEOOYNOY-UHFFFAOYSA-N m-cresol Chemical compound CC1=CC=CC(O)=C1 RLSSMJSEOOYNOY-UHFFFAOYSA-N 0.000 claims abstract description 23
- 229940100630 metacresol Drugs 0.000 claims abstract description 22
- NPBVQXIMTZKSBA-UHFFFAOYSA-N Chavibetol Natural products COC1=CC=C(CC=C)C=C1O NPBVQXIMTZKSBA-UHFFFAOYSA-N 0.000 claims abstract description 16
- 239000005770 Eugenol Substances 0.000 claims abstract description 16
- UVMRYBDEERADNV-UHFFFAOYSA-N Pseudoeugenol Natural products COC1=CC(C(C)=C)=CC=C1O UVMRYBDEERADNV-UHFFFAOYSA-N 0.000 claims abstract description 16
- 229960002217 eugenol Drugs 0.000 claims abstract description 16
- WUOACPNHFRMFPN-SECBINFHSA-N (S)-(-)-alpha-terpineol Chemical compound CC1=CC[C@@H](C(C)(C)O)CC1 WUOACPNHFRMFPN-SECBINFHSA-N 0.000 claims abstract description 14
- OVKDFILSBMEKLT-UHFFFAOYSA-N alpha-Terpineol Natural products CC(=C)C1(O)CCC(C)=CC1 OVKDFILSBMEKLT-UHFFFAOYSA-N 0.000 claims abstract description 14
- 229940088601 alpha-terpineol Drugs 0.000 claims abstract description 14
- 238000006243 chemical reaction Methods 0.000 claims abstract description 10
- 241000972773 Aulopiformes Species 0.000 claims abstract description 6
- 241001519451 Abramis brama Species 0.000 claims abstract description 5
- 241000252233 Cyprinus carpio Species 0.000 claims abstract description 5
- 241000276707 Tilapia Species 0.000 claims abstract description 5
- 241001233037 catfish Species 0.000 claims abstract description 5
- 244000000010 microbial pathogen Species 0.000 claims abstract description 5
- 244000005706 microflora Species 0.000 claims abstract description 5
- 235000019515 salmon Nutrition 0.000 claims abstract description 5
- 235000021051 daily weight gain Nutrition 0.000 claims abstract description 3
- 230000001105 regulatory effect Effects 0.000 claims abstract description 3
- 239000013543 active substance Substances 0.000 claims abstract 6
- 150000001413 amino acids Chemical class 0.000 claims description 78
- 230000000813 microbial effect Effects 0.000 claims description 40
- 238000006467 substitution reaction Methods 0.000 claims description 24
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 17
- 238000012217 deletion Methods 0.000 claims description 13
- 230000037430 deletion Effects 0.000 claims description 13
- 239000012634 fragment Substances 0.000 claims description 13
- 238000003780 insertion Methods 0.000 claims description 13
- 230000037431 insertion Effects 0.000 claims description 13
- 241000277331 Salmonidae Species 0.000 claims description 7
- 241000235349 Ascomycota Species 0.000 claims description 4
- 241001326562 Pezizomycotina Species 0.000 claims description 4
- 230000002538 fungal effect Effects 0.000 claims description 3
- 230000006806 disease prevention Effects 0.000 claims 1
- 239000002075 main ingredient Substances 0.000 claims 1
- 239000000126 substance Substances 0.000 abstract description 20
- 238000004519 manufacturing process Methods 0.000 abstract description 9
- 244000005700 microbiome Species 0.000 abstract description 8
- 230000006872 improvement Effects 0.000 abstract description 5
- 208000015181 infectious disease Diseases 0.000 abstract description 5
- 102100033468 Lysozyme C Human genes 0.000 description 79
- 235000001014 amino acid Nutrition 0.000 description 63
- 229940024606 amino acid Drugs 0.000 description 58
- 150000001875 compounds Chemical class 0.000 description 32
- 102000004190 Enzymes Human genes 0.000 description 18
- 108090000790 Enzymes Proteins 0.000 description 18
- 239000003921 oil Substances 0.000 description 17
- 235000019198 oils Nutrition 0.000 description 17
- 238000000034 method Methods 0.000 description 16
- 235000002639 sodium chloride Nutrition 0.000 description 16
- 201000010099 disease Diseases 0.000 description 15
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 15
- 229940088598 enzyme Drugs 0.000 description 15
- 235000012054 meals Nutrition 0.000 description 14
- 108090000623 proteins and genes Proteins 0.000 description 13
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 12
- 241000894006 Bacteria Species 0.000 description 11
- 239000008188 pellet Substances 0.000 description 11
- VTYYLEPIZMXCLO-UHFFFAOYSA-L Calcium carbonate Chemical compound [Ca+2].[O-]C([O-])=O VTYYLEPIZMXCLO-UHFFFAOYSA-L 0.000 description 10
- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical class CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 description 10
- 230000000845 anti-microbial effect Effects 0.000 description 10
- 239000000306 component Substances 0.000 description 10
- 230000012010 growth Effects 0.000 description 10
- 230000002401 inhibitory effect Effects 0.000 description 10
- 102000004169 proteins and genes Human genes 0.000 description 10
- 150000003839 salts Chemical class 0.000 description 10
- 239000004375 Dextrin Substances 0.000 description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 9
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 9
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 9
- 229920002472 Starch Polymers 0.000 description 9
- 235000019425 dextrin Nutrition 0.000 description 9
- XPFVYQJUAUNWIW-UHFFFAOYSA-N furfuryl alcohol Chemical compound OCC1=CC=CO1 XPFVYQJUAUNWIW-UHFFFAOYSA-N 0.000 description 9
- 230000001717 pathogenic effect Effects 0.000 description 9
- 235000018102 proteins Nutrition 0.000 description 9
- 239000008107 starch Substances 0.000 description 9
- 235000019698 starch Nutrition 0.000 description 9
- 235000000346 sugar Nutrition 0.000 description 9
- 229920001353 Dextrin Polymers 0.000 description 8
- 230000001580 bacterial effect Effects 0.000 description 8
- 239000003795 chemical substances by application Substances 0.000 description 8
- 210000001035 gastrointestinal tract Anatomy 0.000 description 8
- 239000004615 ingredient Substances 0.000 description 8
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 7
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 7
- 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 7
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 7
- 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 description 7
- 229930006000 Sucrose Natural products 0.000 description 7
- 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 7
- 229940000425 combination drug Drugs 0.000 description 7
- 239000008103 glucose Substances 0.000 description 7
- 238000002360 preparation method Methods 0.000 description 7
- 229910052938 sodium sulfate Inorganic materials 0.000 description 7
- 235000011152 sodium sulphate Nutrition 0.000 description 7
- 239000000600 sorbitol Substances 0.000 description 7
- 239000005720 sucrose Substances 0.000 description 7
- 235000015112 vegetable and seed oil Nutrition 0.000 description 7
- 239000008158 vegetable oil Substances 0.000 description 7
- 235000019733 Fish meal Nutrition 0.000 description 6
- 125000003412 L-alanyl group Chemical group [H]N([H])[C@@](C([H])([H])[H])(C(=O)[*])[H] 0.000 description 6
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 6
- 238000000576 coating method Methods 0.000 description 6
- 235000021323 fish oil Nutrition 0.000 description 6
- 239000004467 fishmeal Substances 0.000 description 6
- 244000052769 pathogen Species 0.000 description 6
- YPFDHNVEDLHUCE-UHFFFAOYSA-N propane-1,3-diol Chemical compound OCCCO YPFDHNVEDLHUCE-UHFFFAOYSA-N 0.000 description 6
- WXMKPNITSTVMEF-UHFFFAOYSA-M sodium benzoate Chemical compound [Na+].[O-]C(=O)C1=CC=CC=C1 WXMKPNITSTVMEF-UHFFFAOYSA-M 0.000 description 6
- 239000004299 sodium benzoate Substances 0.000 description 6
- 235000010234 sodium benzoate Nutrition 0.000 description 6
- 229960003885 sodium benzoate Drugs 0.000 description 6
- 239000011780 sodium chloride Substances 0.000 description 6
- 229960002668 sodium chloride Drugs 0.000 description 6
- 229960003010 sodium sulfate Drugs 0.000 description 6
- GETQZCLCWQTVFV-UHFFFAOYSA-N trimethylamine Chemical compound CN(C)C GETQZCLCWQTVFV-UHFFFAOYSA-N 0.000 description 6
- CHHHXKFHOYLYRE-UHFFFAOYSA-M 2,4-Hexadienoic acid, potassium salt (1:1), (2E,4E)- Chemical compound [K+].CC=CC=CC([O-])=O CHHHXKFHOYLYRE-UHFFFAOYSA-M 0.000 description 5
- 108010011619 6-Phytase Proteins 0.000 description 5
- 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 5
- 241000238557 Decapoda Species 0.000 description 5
- 108010068370 Glutens Proteins 0.000 description 5
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 5
- 241000194040 Lactococcus garvieae Species 0.000 description 5
- 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
- OVRNDRQMDRJTHS-FMDGEEDCSA-N N-acetyl-beta-D-glucosamine Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O OVRNDRQMDRJTHS-FMDGEEDCSA-N 0.000 description 5
- 241000194056 Streptococcus iniae Species 0.000 description 5
- 238000009360 aquaculture Methods 0.000 description 5
- 244000144974 aquaculture Species 0.000 description 5
- 235000010216 calcium carbonate Nutrition 0.000 description 5
- 229960003563 calcium carbonate Drugs 0.000 description 5
- 229910000019 calcium carbonate Inorganic materials 0.000 description 5
- 239000001913 cellulose Substances 0.000 description 5
- 235000010980 cellulose Nutrition 0.000 description 5
- 238000010790 dilution Methods 0.000 description 5
- 239000012895 dilution Substances 0.000 description 5
- 235000021312 gluten Nutrition 0.000 description 5
- 229910052500 inorganic mineral Inorganic materials 0.000 description 5
- 239000008101 lactose Substances 0.000 description 5
- 235000010755 mineral Nutrition 0.000 description 5
- 239000011707 mineral Substances 0.000 description 5
- 229940085127 phytase Drugs 0.000 description 5
- 239000004302 potassium sorbate Substances 0.000 description 5
- 235000010241 potassium sorbate Nutrition 0.000 description 5
- 229940069338 potassium sorbate Drugs 0.000 description 5
- OTYBMLCTZGSZBG-UHFFFAOYSA-L potassium sulfate Chemical compound [K+].[K+].[O-]S([O-])(=O)=O OTYBMLCTZGSZBG-UHFFFAOYSA-L 0.000 description 5
- 229910052939 potassium sulfate Inorganic materials 0.000 description 5
- 239000001509 sodium citrate Substances 0.000 description 5
- 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 5
- 235000011083 sodium citrates Nutrition 0.000 description 5
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 description 4
- 102000002322 Egg Proteins Human genes 0.000 description 4
- 108010000912 Egg Proteins Proteins 0.000 description 4
- 235000010469 Glycine max Nutrition 0.000 description 4
- 102100037611 Lysophospholipase Human genes 0.000 description 4
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 4
- 241000277263 Salmo Species 0.000 description 4
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 4
- 239000004480 active ingredient Substances 0.000 description 4
- 210000004899 c-terminal region Anatomy 0.000 description 4
- OSGAYBCDTDRGGQ-UHFFFAOYSA-L calcium sulfate Chemical compound [Ca+2].[O-]S([O-])(=O)=O OSGAYBCDTDRGGQ-UHFFFAOYSA-L 0.000 description 4
- 210000002421 cell wall Anatomy 0.000 description 4
- 235000013339 cereals Nutrition 0.000 description 4
- 239000011248 coating agent Substances 0.000 description 4
- 239000012141 concentrate Substances 0.000 description 4
- 239000007771 core particle Substances 0.000 description 4
- 235000014103 egg white Nutrition 0.000 description 4
- 210000000969 egg white Anatomy 0.000 description 4
- 239000000284 extract Substances 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 244000005709 gut microbiome Species 0.000 description 4
- 230000036541 health Effects 0.000 description 4
- 150000002632 lipids Chemical class 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- BWHMMNNQKKPAPP-UHFFFAOYSA-L potassium carbonate Chemical compound [K+].[K+].[O-]C([O-])=O BWHMMNNQKKPAPP-UHFFFAOYSA-L 0.000 description 4
- 229940093914 potassium sulfate Drugs 0.000 description 4
- 235000011151 potassium sulphates Nutrition 0.000 description 4
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 229960004063 propylene glycol Drugs 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 238000003307 slaughter Methods 0.000 description 4
- AKHNMLFCWUSKQB-UHFFFAOYSA-L sodium thiosulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=S AKHNMLFCWUSKQB-UHFFFAOYSA-L 0.000 description 4
- 229940001474 sodium thiosulfate Drugs 0.000 description 4
- 235000019345 sodium thiosulphate Nutrition 0.000 description 4
- 239000008247 solid mixture Substances 0.000 description 4
- 238000012360 testing method Methods 0.000 description 4
- 229940088594 vitamin Drugs 0.000 description 4
- 235000013343 vitamin Nutrition 0.000 description 4
- 229930003231 vitamin Natural products 0.000 description 4
- 239000011782 vitamin Substances 0.000 description 4
- 235000019786 weight gain Nutrition 0.000 description 4
- 230000004584 weight gain Effects 0.000 description 4
- JIAARYAFYJHUJI-UHFFFAOYSA-L zinc dichloride Chemical compound [Cl-].[Cl-].[Zn+2] JIAARYAFYJHUJI-UHFFFAOYSA-L 0.000 description 4
- MTCFGRXMJLQNBG-REOHCLBHSA-N (2S)-2-Amino-3-hydroxypropansäure Chemical compound OC[C@H](N)C(O)=O MTCFGRXMJLQNBG-REOHCLBHSA-N 0.000 description 3
- 241000932614 Acremonium alcalophilum Species 0.000 description 3
- 241000233866 Fungi Species 0.000 description 3
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 description 3
- 125000000570 L-alpha-aspartyl group Chemical group [H]OC(=O)C([H])([H])[C@]([H])(N([H])[H])C(*)=O 0.000 description 3
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 description 3
- MSFSPUZXLOGKHJ-UHFFFAOYSA-N Muraminsaeure Natural products OC(=O)C(C)OC1C(N)C(O)OC(CO)C1O MSFSPUZXLOGKHJ-UHFFFAOYSA-N 0.000 description 3
- MNLRQHMNZILYPY-MDMHTWEWSA-N N-acetyl-alpha-D-muramic acid Chemical compound OC(=O)[C@@H](C)O[C@H]1[C@H](O)[C@@H](CO)O[C@H](O)[C@@H]1NC(C)=O MNLRQHMNZILYPY-MDMHTWEWSA-N 0.000 description 3
- 108010013639 Peptidoglycan Proteins 0.000 description 3
- 235000019484 Rapeseed oil Nutrition 0.000 description 3
- 240000008042 Zea mays Species 0.000 description 3
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 3
- 230000004075 alteration Effects 0.000 description 3
- 125000000539 amino acid group Chemical group 0.000 description 3
- 229920002678 cellulose Polymers 0.000 description 3
- 238000009313 farming Methods 0.000 description 3
- 239000008187 granular material Substances 0.000 description 3
- 210000000987 immune system Anatomy 0.000 description 3
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 3
- 235000019341 magnesium sulphate Nutrition 0.000 description 3
- 239000005445 natural material Substances 0.000 description 3
- MXXWOMGUGJBKIW-YPCIICBESA-N piperine Chemical compound C=1C=C2OCOC2=CC=1/C=C/C=C/C(=O)N1CCCCC1 MXXWOMGUGJBKIW-YPCIICBESA-N 0.000 description 3
- WVWHRXVVAYXKDE-UHFFFAOYSA-N piperine Natural products O=C(C=CC=Cc1ccc2OCOc2c1)C3CCCCN3 WVWHRXVVAYXKDE-UHFFFAOYSA-N 0.000 description 3
- 235000020777 polyunsaturated fatty acids Nutrition 0.000 description 3
- 244000144977 poultry Species 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 229960001790 sodium citrate Drugs 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 230000007306 turnover Effects 0.000 description 3
- -1 variant Proteins 0.000 description 3
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 2
- 244000105624 Arachis hypogaea Species 0.000 description 2
- 235000010777 Arachis hypogaea Nutrition 0.000 description 2
- 241000473391 Archosargus rhomboidalis Species 0.000 description 2
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical class OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 description 2
- 239000005996 Blood meal Substances 0.000 description 2
- 235000004977 Brassica sinapistrum Nutrition 0.000 description 2
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 102100028717 Cytosolic 5'-nucleotidase 3A Human genes 0.000 description 2
- 241000723298 Dicentrarchus labrax Species 0.000 description 2
- 241000276438 Gadus morhua Species 0.000 description 2
- 241000287828 Gallus gallus Species 0.000 description 2
- 229920001503 Glucan Polymers 0.000 description 2
- 244000068988 Glycine max Species 0.000 description 2
- 102000005744 Glycoside Hydrolases Human genes 0.000 description 2
- 108010031186 Glycoside Hydrolases Proteins 0.000 description 2
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 2
- 125000003440 L-leucyl group Chemical group O=C([*])[C@](N([H])[H])([H])C([H])([H])C(C([H])([H])[H])([H])C([H])([H])[H] 0.000 description 2
- 125000002842 L-seryl group Chemical group O=C([*])[C@](N([H])[H])([H])C([H])([H])O[H] 0.000 description 2
- AYFVYJQAPQTCCC-GBXIJSLDSA-N L-threonine Chemical compound C[C@@H](O)[C@H](N)C(O)=O AYFVYJQAPQTCCC-GBXIJSLDSA-N 0.000 description 2
- 241000219745 Lupinus Species 0.000 description 2
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 description 2
- 239000004472 Lysine Substances 0.000 description 2
- 108020002496 Lysophospholipase Proteins 0.000 description 2
- 241000124008 Mammalia Species 0.000 description 2
- OVRNDRQMDRJTHS-UHFFFAOYSA-N N-acelyl-D-glucosamine Natural products CC(=O)NC1C(O)OC(CO)C(O)C1O OVRNDRQMDRJTHS-UHFFFAOYSA-N 0.000 description 2
- 244000046052 Phaseolus vulgaris Species 0.000 description 2
- 235000010627 Phaseolus vulgaris Nutrition 0.000 description 2
- 108090000553 Phospholipase D Proteins 0.000 description 2
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 2
- 240000006394 Sorghum bicolor Species 0.000 description 2
- 235000019764 Soybean Meal Nutrition 0.000 description 2
- 235000019772 Sunflower meal Nutrition 0.000 description 2
- 241000259813 Trichophaea saccata Species 0.000 description 2
- 241000209140 Triticum Species 0.000 description 2
- 235000021307 Triticum Nutrition 0.000 description 2
- 208000025865 Ulcer Diseases 0.000 description 2
- 206010047400 Vibrio infections Diseases 0.000 description 2
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 2
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 2
- FMRLDPWIRHBCCC-UHFFFAOYSA-L Zinc carbonate Chemical compound [Zn+2].[O-]C([O-])=O FMRLDPWIRHBCCC-UHFFFAOYSA-L 0.000 description 2
- ZOIORXHNWRGPMV-UHFFFAOYSA-N acetic acid;zinc Chemical compound [Zn].CC(O)=O.CC(O)=O ZOIORXHNWRGPMV-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 239000008272 agar Substances 0.000 description 2
- 235000004279 alanine Nutrition 0.000 description 2
- 108090000637 alpha-Amylases Proteins 0.000 description 2
- 108010030291 alpha-Galactosidase Proteins 0.000 description 2
- 235000019730 animal feed additive Nutrition 0.000 description 2
- 239000003242 anti bacterial agent Substances 0.000 description 2
- 229940088710 antibiotic agent Drugs 0.000 description 2
- 239000004599 antimicrobial Substances 0.000 description 2
- YZXBAPSDXZZRGB-DOFZRALJSA-N arachidonic acid Chemical compound CCCCC\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O YZXBAPSDXZZRGB-DOFZRALJSA-N 0.000 description 2
- VSGNNIFQASZAOI-UHFFFAOYSA-L calcium acetate Chemical compound [Ca+2].CC([O-])=O.CC([O-])=O VSGNNIFQASZAOI-UHFFFAOYSA-L 0.000 description 2
- 239000001639 calcium acetate Substances 0.000 description 2
- 235000011092 calcium acetate Nutrition 0.000 description 2
- 229960005147 calcium acetate Drugs 0.000 description 2
- 239000004301 calcium benzoate Substances 0.000 description 2
- 235000010237 calcium benzoate Nutrition 0.000 description 2
- 239000001110 calcium chloride Substances 0.000 description 2
- 229910001628 calcium chloride Inorganic materials 0.000 description 2
- 229960002713 calcium chloride Drugs 0.000 description 2
- 235000011148 calcium chloride Nutrition 0.000 description 2
- FNAQSUUGMSOBHW-UHFFFAOYSA-H calcium citrate Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O.[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O FNAQSUUGMSOBHW-UHFFFAOYSA-H 0.000 description 2
- 239000001354 calcium citrate Substances 0.000 description 2
- 229960004256 calcium citrate Drugs 0.000 description 2
- MCFVRESNTICQSJ-RJNTXXOISA-L calcium sorbate Chemical compound [Ca+2].C\C=C\C=C\C([O-])=O.C\C=C\C=C\C([O-])=O MCFVRESNTICQSJ-RJNTXXOISA-L 0.000 description 2
- 239000004303 calcium sorbate Substances 0.000 description 2
- 235000010244 calcium sorbate Nutrition 0.000 description 2
- 235000011132 calcium sulphate Nutrition 0.000 description 2
- HZQXCUSDXIKLGS-UHFFFAOYSA-L calcium;dibenzoate;trihydrate Chemical compound O.O.O.[Ca+2].[O-]C(=O)C1=CC=CC=C1.[O-]C(=O)C1=CC=CC=C1 HZQXCUSDXIKLGS-UHFFFAOYSA-L 0.000 description 2
- YKPUWZUDDOIDPM-SOFGYWHQSA-N capsaicin Chemical compound COC1=CC(CNC(=O)CCCC\C=C\C(C)C)=CC=C1O YKPUWZUDDOIDPM-SOFGYWHQSA-N 0.000 description 2
- 150000001720 carbohydrates Chemical class 0.000 description 2
- 235000014633 carbohydrates Nutrition 0.000 description 2
- 229910052799 carbon Inorganic materials 0.000 description 2
- 230000003197 catalytic effect Effects 0.000 description 2
- 229940106135 cellulose Drugs 0.000 description 2
- 235000008504 concentrate Nutrition 0.000 description 2
- 230000001143 conditioned effect Effects 0.000 description 2
- 238000010411 cooking Methods 0.000 description 2
- 235000005822 corn Nutrition 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 235000005911 diet Nutrition 0.000 description 2
- 235000014113 dietary fatty acids Nutrition 0.000 description 2
- 235000019621 digestibility Nutrition 0.000 description 2
- 231100000676 disease causative agent Toxicity 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 239000000194 fatty acid Substances 0.000 description 2
- 229930195729 fatty acid Natural products 0.000 description 2
- 150000004665 fatty acids Chemical class 0.000 description 2
- 210000003746 feather Anatomy 0.000 description 2
- 208000010824 fish disease Diseases 0.000 description 2
- 235000013312 flour Nutrition 0.000 description 2
- 238000009472 formulation Methods 0.000 description 2
- 239000013505 freshwater Substances 0.000 description 2
- 230000007062 hydrolysis Effects 0.000 description 2
- 238000006460 hydrolysis reaction Methods 0.000 description 2
- 238000000338 in vitro Methods 0.000 description 2
- 238000010348 incorporation Methods 0.000 description 2
- XMGQYMWWDOXHJM-UHFFFAOYSA-N limonene Chemical compound CC(=C)C1CCC(C)=CC1 XMGQYMWWDOXHJM-UHFFFAOYSA-N 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 239000012669 liquid formulation Substances 0.000 description 2
- 230000002101 lytic effect Effects 0.000 description 2
- 229960003390 magnesium sulfate Drugs 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000035772 mutation Effects 0.000 description 2
- 229950006780 n-acetylglucosamine Drugs 0.000 description 2
- 229920000136 polysorbate Polymers 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- SCVFZCLFOSHCOH-UHFFFAOYSA-M potassium acetate Chemical compound [K+].CC([O-])=O SCVFZCLFOSHCOH-UHFFFAOYSA-M 0.000 description 2
- XAEFZNCEHLXOMS-UHFFFAOYSA-M potassium benzoate Chemical compound [K+].[O-]C(=O)C1=CC=CC=C1 XAEFZNCEHLXOMS-UHFFFAOYSA-M 0.000 description 2
- 239000004300 potassium benzoate Substances 0.000 description 2
- 235000010235 potassium benzoate Nutrition 0.000 description 2
- 229940103091 potassium benzoate Drugs 0.000 description 2
- 229910000027 potassium carbonate Inorganic materials 0.000 description 2
- 229940093956 potassium carbonate Drugs 0.000 description 2
- 235000011181 potassium carbonates Nutrition 0.000 description 2
- 239000001103 potassium chloride Substances 0.000 description 2
- 235000011164 potassium chloride Nutrition 0.000 description 2
- 229960002816 potassium chloride Drugs 0.000 description 2
- 239000001508 potassium citrate Substances 0.000 description 2
- 229960002635 potassium citrate Drugs 0.000 description 2
- QEEAPRPFLLJWCF-UHFFFAOYSA-K potassium citrate (anhydrous) Chemical compound [K+].[K+].[K+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O QEEAPRPFLLJWCF-UHFFFAOYSA-K 0.000 description 2
- 235000011082 potassium citrates Nutrition 0.000 description 2
- 230000003405 preventing effect Effects 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 239000004456 rapeseed meal Substances 0.000 description 2
- 239000010822 slaughterhouse waste Substances 0.000 description 2
- 239000001632 sodium acetate Substances 0.000 description 2
- 235000017281 sodium acetate Nutrition 0.000 description 2
- 229960004249 sodium acetate Drugs 0.000 description 2
- 229910000029 sodium carbonate Inorganic materials 0.000 description 2
- 229940001593 sodium carbonate Drugs 0.000 description 2
- 235000017550 sodium carbonate Nutrition 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 239000004455 soybean meal Substances 0.000 description 2
- 235000013337 tricalcium citrate Nutrition 0.000 description 2
- WGIWBXUNRXCYRA-UHFFFAOYSA-H trizinc;2-hydroxypropane-1,2,3-tricarboxylate Chemical compound [Zn+2].[Zn+2].[Zn+2].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O.[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O WGIWBXUNRXCYRA-UHFFFAOYSA-H 0.000 description 2
- 231100000397 ulcer Toxicity 0.000 description 2
- 229960005486 vaccine Drugs 0.000 description 2
- 239000012138 yeast extract Substances 0.000 description 2
- 239000004246 zinc acetate Substances 0.000 description 2
- 229960000314 zinc acetate Drugs 0.000 description 2
- 235000013904 zinc acetate Nutrition 0.000 description 2
- 239000011667 zinc carbonate Substances 0.000 description 2
- 235000004416 zinc carbonate Nutrition 0.000 description 2
- 229910000010 zinc carbonate Inorganic materials 0.000 description 2
- 229940043825 zinc carbonate Drugs 0.000 description 2
- 239000011592 zinc chloride Substances 0.000 description 2
- 235000005074 zinc chloride Nutrition 0.000 description 2
- 229960001939 zinc chloride Drugs 0.000 description 2
- 239000011746 zinc citrate Substances 0.000 description 2
- 235000006076 zinc citrate Nutrition 0.000 description 2
- 229940068475 zinc citrate Drugs 0.000 description 2
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 2
- 229960001763 zinc sulfate Drugs 0.000 description 2
- 229910000368 zinc sulfate Inorganic materials 0.000 description 2
- YZSUHNYMDVSXAS-RJNTXXOISA-L zinc;(2e,4e)-hexa-2,4-dienoate Chemical compound [Zn+2].C\C=C\C=C\C([O-])=O.C\C=C\C=C\C([O-])=O YZSUHNYMDVSXAS-RJNTXXOISA-L 0.000 description 2
- JDLYKQWJXAQNNS-UHFFFAOYSA-L zinc;dibenzoate Chemical compound [Zn+2].[O-]C(=O)C1=CC=CC=C1.[O-]C(=O)C1=CC=CC=C1 JDLYKQWJXAQNNS-UHFFFAOYSA-L 0.000 description 2
- OCUSNPIJIZCRSZ-ZTZWCFDHSA-N (2s)-2-amino-3-methylbutanoic acid;(2s)-2-amino-4-methylpentanoic acid;(2s,3s)-2-amino-3-methylpentanoic acid Chemical compound CC(C)[C@H](N)C(O)=O.CC[C@H](C)[C@H](N)C(O)=O.CC(C)C[C@H](N)C(O)=O OCUSNPIJIZCRSZ-ZTZWCFDHSA-N 0.000 description 1
- WSWCOQWTEOXDQX-MQQKCMAXSA-M (E,E)-sorbate Chemical compound C\C=C\C=C\C([O-])=O WSWCOQWTEOXDQX-MQQKCMAXSA-M 0.000 description 1
- VBICKXHEKHSIBG-UHFFFAOYSA-N 1-monostearoylglycerol Chemical compound CCCCCCCCCCCCCCCCCC(=O)OCC(O)CO VBICKXHEKHSIBG-UHFFFAOYSA-N 0.000 description 1
- ZIIUUSVHCHPIQD-UHFFFAOYSA-N 2,4,6-trimethyl-N-[3-(trifluoromethyl)phenyl]benzenesulfonamide Chemical compound CC1=CC(C)=CC(C)=C1S(=O)(=O)NC1=CC=CC(C(F)(F)F)=C1 ZIIUUSVHCHPIQD-UHFFFAOYSA-N 0.000 description 1
- DVSDEXBFPJVTGH-UHFFFAOYSA-N 6-methoxy-4-prop-2-enylcyclohexa-2,4-dien-1-ol Chemical compound COC1C=C(CC=C)C=CC1O DVSDEXBFPJVTGH-UHFFFAOYSA-N 0.000 description 1
- 244000215068 Acacia senegal Species 0.000 description 1
- QTBSBXVTEAMEQO-UHFFFAOYSA-M Acetate Chemical compound CC([O-])=O QTBSBXVTEAMEQO-UHFFFAOYSA-M 0.000 description 1
- 241000607528 Aeromonas hydrophila Species 0.000 description 1
- 241000607525 Aeromonas salmonicida Species 0.000 description 1
- 241000122170 Aliivibrio salmonicida Species 0.000 description 1
- 239000004382 Amylase Substances 0.000 description 1
- 102000013142 Amylases Human genes 0.000 description 1
- 108010065511 Amylases Proteins 0.000 description 1
- 235000019737 Animal fat Nutrition 0.000 description 1
- 241000272814 Anser sp. Species 0.000 description 1
- 102000044503 Antimicrobial Peptides Human genes 0.000 description 1
- 108700042778 Antimicrobial Peptides Proteins 0.000 description 1
- 235000003276 Apios tuberosa Nutrition 0.000 description 1
- 101710152845 Arabinogalactan endo-beta-1,4-galactanase Proteins 0.000 description 1
- 235000017060 Arachis glabrata Nutrition 0.000 description 1
- 235000018262 Arachis monticola Nutrition 0.000 description 1
- 235000010744 Arachis villosulicarpa Nutrition 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- 108010024976 Asparaginase Proteins 0.000 description 1
- DCXYFEDJOCDNAF-UHFFFAOYSA-N Asparagine Natural products OC(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-N 0.000 description 1
- JEBFVOLFMLUKLF-IFPLVEIFSA-N Astaxanthin Natural products CC(=C/C=C/C(=C/C=C/C1=C(C)C(=O)C(O)CC1(C)C)/C)C=CC=C(/C)C=CC=C(/C)C=CC2=C(C)C(=O)C(O)CC2(C)C JEBFVOLFMLUKLF-IFPLVEIFSA-N 0.000 description 1
- 235000007319 Avena orientalis Nutrition 0.000 description 1
- 244000075850 Avena orientalis Species 0.000 description 1
- 241000271566 Aves Species 0.000 description 1
- 108010077805 Bacterial Proteins Proteins 0.000 description 1
- 231100000699 Bacterial toxin Toxicity 0.000 description 1
- 101710130006 Beta-glucanase Proteins 0.000 description 1
- 235000014698 Brassica juncea var multisecta Nutrition 0.000 description 1
- 240000002791 Brassica napus Species 0.000 description 1
- 235000006008 Brassica napus var napus Nutrition 0.000 description 1
- 235000006618 Brassica rapa subsp oleifera Nutrition 0.000 description 1
- 244000188595 Brassica sinapistrum Species 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 244000020518 Carthamus tinctorius Species 0.000 description 1
- 235000003255 Carthamus tinctorius Nutrition 0.000 description 1
- 241000626614 Chalaropsis Species 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- 241000195493 Cryptophyta Species 0.000 description 1
- 235000014375 Curcuma Nutrition 0.000 description 1
- 244000164480 Curcuma aromatica Species 0.000 description 1
- ZZZCUOFIHGPKAK-UHFFFAOYSA-N D-erythro-ascorbic acid Natural products OCC1OC(=O)C(O)=C1O ZZZCUOFIHGPKAK-UHFFFAOYSA-N 0.000 description 1
- 108010082495 Dietary Plant Proteins Proteins 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 101710121765 Endo-1,4-beta-xylanase Proteins 0.000 description 1
- 101710147028 Endo-beta-1,4-galactanase Proteins 0.000 description 1
- 101000925646 Enterobacteria phage T4 Endolysin Proteins 0.000 description 1
- 241001656472 Epinephelus morio Species 0.000 description 1
- 241000701533 Escherichia virus T4 Species 0.000 description 1
- 241000192125 Firmicutes Species 0.000 description 1
- 206010017553 Furuncle Diseases 0.000 description 1
- 241000287826 Gallus Species 0.000 description 1
- DCXXMTOCNZCJGO-UHFFFAOYSA-N Glycerol trioctadecanoate Natural products CCCCCCCCCCCCCCCCCC(=O)OCC(OC(=O)CCCCCCCCCCCCCCCCC)COC(=O)CCCCCCCCCCCCCCCCC DCXXMTOCNZCJGO-UHFFFAOYSA-N 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 229920000084 Gum arabic Polymers 0.000 description 1
- 208000032969 Hemorrhagic Septicemia Diseases 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 240000005979 Hordeum vulgare Species 0.000 description 1
- 235000007340 Hordeum vulgare Nutrition 0.000 description 1
- 102000004157 Hydrolases Human genes 0.000 description 1
- 108090000604 Hydrolases Proteins 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 description 1
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 description 1
- HNDVDQJCIGZPNO-YFKPBYRVSA-N L-histidine Chemical compound OC(=O)[C@@H](N)CC1=CN=CN1 HNDVDQJCIGZPNO-YFKPBYRVSA-N 0.000 description 1
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 description 1
- 125000001176 L-lysyl group Chemical group [H]N([H])[C@]([H])(C(=O)[*])C([H])([H])C([H])([H])C([H])([H])C(N([H])[H])([H])[H] 0.000 description 1
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 1
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 1
- 125000000769 L-threonyl group Chemical group [H]N([H])[C@]([H])(C(=O)[*])[C@](O[H])(C([H])([H])[H])[H] 0.000 description 1
- 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 1
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 1
- 125000003798 L-tyrosyl group Chemical group [H]N([H])[C@]([H])(C(=O)[*])C([H])([H])C1=C([H])C([H])=C(O[H])C([H])=C1[H] 0.000 description 1
- 125000003580 L-valyl group Chemical group [H]N([H])[C@]([H])(C(=O)[*])C(C([H])([H])[H])(C([H])([H])[H])[H] 0.000 description 1
- 241000270322 Lepidosauria Species 0.000 description 1
- 235000019738 Limestone Nutrition 0.000 description 1
- 241001153358 Micrococcus luteus NCTC 2665 Species 0.000 description 1
- 241001600139 Moritella viscosa Species 0.000 description 1
- 241000202240 Morone americana Species 0.000 description 1
- 241001481825 Morone saxatilis Species 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 239000007832 Na2SO4 Substances 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 108700022034 Opsonin Proteins Proteins 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 102100026367 Pancreatic alpha-amylase Human genes 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102100035200 Phospholipase A and acyltransferase 4 Human genes 0.000 description 1
- 102000011420 Phospholipase D Human genes 0.000 description 1
- 102100032967 Phospholipase D1 Human genes 0.000 description 1
- 108010064785 Phospholipases Proteins 0.000 description 1
- 102000015439 Phospholipases Human genes 0.000 description 1
- 108010058864 Phospholipases A2 Proteins 0.000 description 1
- 241001517016 Photobacterium damselae Species 0.000 description 1
- 241001517024 Photobacterium damselae subsp. piscicida Species 0.000 description 1
- 241000192126 Piscirickettsia salmonis Species 0.000 description 1
- 208000037113 Piscirickettsiaceae Infections Diseases 0.000 description 1
- 241001600434 Plectroglyphidodon lacrymatus Species 0.000 description 1
- 241000269980 Pleuronectidae Species 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 241000157468 Reinhardtius hippoglossoides Species 0.000 description 1
- 102100037486 Reverse transcriptase/ribonuclease H Human genes 0.000 description 1
- 241000606651 Rickettsiales Species 0.000 description 1
- 244000178231 Rosmarinus officinalis Species 0.000 description 1
- 241000282849 Ruminantia Species 0.000 description 1
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 1
- 241000277289 Salmo salar Species 0.000 description 1
- 235000007238 Secale cereale Nutrition 0.000 description 1
- 244000082988 Secale cereale Species 0.000 description 1
- 206010040047 Sepsis Diseases 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- 241000276699 Seriola Species 0.000 description 1
- 235000011684 Sorghum saccharatum Nutrition 0.000 description 1
- 241000736131 Sphingomonas Species 0.000 description 1
- 241000191940 Staphylococcus Species 0.000 description 1
- 241000282898 Sus scrofa 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
- 241000499912 Trichoderma reesei Species 0.000 description 1
- 240000000359 Triticum dicoccon Species 0.000 description 1
- QIVBCDIJIAJPQS-UHFFFAOYSA-N Tryptophan Natural products C1=CC=C2C(CC(N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-UHFFFAOYSA-N 0.000 description 1
- 102000014384 Type C Phospholipases Human genes 0.000 description 1
- 108010079194 Type C Phospholipases Proteins 0.000 description 1
- 241000607618 Vibrio harveyi Species 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 229930003268 Vitamin C Natural products 0.000 description 1
- 229920002522 Wood fibre Polymers 0.000 description 1
- 241001148129 Yersinia ruckeri Species 0.000 description 1
- 235000016383 Zea mays subsp huehuetenangensis Nutrition 0.000 description 1
- 239000000205 acacia gum Substances 0.000 description 1
- 235000010489 acacia gum Nutrition 0.000 description 1
- 229940022663 acetate Drugs 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 230000002730 additional effect Effects 0.000 description 1
- 238000012867 alanine scanning Methods 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- JAZBEHYOTPTENJ-JLNKQSITSA-N all-cis-5,8,11,14,17-icosapentaenoic acid Chemical compound CC\C=C/C\C=C/C\C=C/C\C=C/C\C=C/CCCC(O)=O JAZBEHYOTPTENJ-JLNKQSITSA-N 0.000 description 1
- OENHQHLEOONYIE-UKMVMLAPSA-N all-trans beta-carotene Natural products CC=1CCCC(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C OENHQHLEOONYIE-UKMVMLAPSA-N 0.000 description 1
- 102000004139 alpha-Amylases Human genes 0.000 description 1
- 102000005840 alpha-Galactosidase Human genes 0.000 description 1
- 229940024171 alpha-amylase Drugs 0.000 description 1
- WUOACPNHFRMFPN-UHFFFAOYSA-N alpha-terpineol Chemical compound CC1=CCC(C(C)(C)O)CC1 WUOACPNHFRMFPN-UHFFFAOYSA-N 0.000 description 1
- 235000019418 amylase Nutrition 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 235000021120 animal protein Nutrition 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 238000002802 antimicrobial activity assay Methods 0.000 description 1
- 229940114079 arachidonic acid Drugs 0.000 description 1
- 235000021342 arachidonic acid Nutrition 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 235000009582 asparagine Nutrition 0.000 description 1
- 229960001230 asparagine Drugs 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 235000013793 astaxanthin Nutrition 0.000 description 1
- 239000001168 astaxanthin Substances 0.000 description 1
- MQZIGYBFDRPAKN-ZWAPEEGVSA-N astaxanthin Chemical compound C([C@H](O)C(=O)C=1C)C(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)C(=O)[C@@H](O)CC1(C)C MQZIGYBFDRPAKN-ZWAPEEGVSA-N 0.000 description 1
- 229940022405 astaxanthin Drugs 0.000 description 1
- 239000000688 bacterial toxin Substances 0.000 description 1
- 230000003385 bacteriostatic effect Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 235000013734 beta-carotene Nutrition 0.000 description 1
- 239000011648 beta-carotene Substances 0.000 description 1
- TUPZEYHYWIEDIH-WAIFQNFQSA-N beta-carotene Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CCCC1(C)C)C=CC=C(/C)C=CC2=CCCCC2(C)C TUPZEYHYWIEDIH-WAIFQNFQSA-N 0.000 description 1
- 229960002747 betacarotene Drugs 0.000 description 1
- 230000009141 biological interaction Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000002374 bone meal Substances 0.000 description 1
- 229940036811 bone meal Drugs 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229960005069 calcium Drugs 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 235000001465 calcium Nutrition 0.000 description 1
- 159000000007 calcium salts Chemical class 0.000 description 1
- 229940095672 calcium sulfate Drugs 0.000 description 1
- 229940041514 candida albicans extract Drugs 0.000 description 1
- 229960002504 capsaicin Drugs 0.000 description 1
- 235000017663 capsaicin Nutrition 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 235000021466 carotenoid Nutrition 0.000 description 1
- 150000001747 carotenoids Chemical class 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000000460 chlorine Substances 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 235000012343 cottonseed oil Nutrition 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- VFLDPWHFBUODDF-FCXRPNKRSA-N curcumin Chemical compound C1=C(O)C(OC)=CC(\C=C\C(=O)CC(=O)\C=C\C=2C=C(OC)C(O)=CC=2)=C1 VFLDPWHFBUODDF-FCXRPNKRSA-N 0.000 description 1
- FYGDTMLNYKFZSV-MRCIVHHJSA-N dextrin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)OC1O[C@@H]1[C@@H](CO)OC(O[C@@H]2[C@H](O[C@H](O)[C@H](O)[C@H]2O)CO)[C@H](O)[C@H]1O FYGDTMLNYKFZSV-MRCIVHHJSA-N 0.000 description 1
- 230000037213 diet Effects 0.000 description 1
- 230000000378 dietary effect Effects 0.000 description 1
- 235000013325 dietary fiber Nutrition 0.000 description 1
- 238000002050 diffraction method Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 238000007598 dipping method Methods 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- JAZBEHYOTPTENJ-UHFFFAOYSA-N eicosapentaenoic acid Natural products CCC=CCC=CCC=CCC=CCC=CCCCC(O)=O JAZBEHYOTPTENJ-UHFFFAOYSA-N 0.000 description 1
- 229960005135 eicosapentaenoic acid Drugs 0.000 description 1
- 235000020673 eicosapentaenoic acid Nutrition 0.000 description 1
- 238000002003 electron diffraction Methods 0.000 description 1
- 239000003995 emulsifying agent Substances 0.000 description 1
- 230000001804 emulsifying effect Effects 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 235000020776 essential amino acid Nutrition 0.000 description 1
- 239000003797 essential amino acid Substances 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000003925 fat Substances 0.000 description 1
- 235000019197 fats Nutrition 0.000 description 1
- 235000021050 feed intake Nutrition 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 235000013373 food additive Nutrition 0.000 description 1
- 239000002778 food additive Substances 0.000 description 1
- 239000004459 forage Substances 0.000 description 1
- 208000003512 furunculosis Diseases 0.000 description 1
- 229940098330 gamma linoleic acid Drugs 0.000 description 1
- VZCCETWTMQHEPK-UHFFFAOYSA-N gamma-Linolensaeure Natural products CCCCCC=CCC=CCC=CCCCCC(O)=O VZCCETWTMQHEPK-UHFFFAOYSA-N 0.000 description 1
- VZCCETWTMQHEPK-QNEBEIHSSA-N gamma-linolenic acid Chemical compound CCCCC\C=C/C\C=C/C\C=C/CCCCC(O)=O VZCCETWTMQHEPK-QNEBEIHSSA-N 0.000 description 1
- 230000002496 gastric effect Effects 0.000 description 1
- 235000001727 glucose Nutrition 0.000 description 1
- 235000013922 glutamic acid Nutrition 0.000 description 1
- 239000004220 glutamic acid Substances 0.000 description 1
- ZDXPYRJPNDTMRX-UHFFFAOYSA-N glutamine Natural products OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 230000013595 glycosylation Effects 0.000 description 1
- 238000006206 glycosylation reaction Methods 0.000 description 1
- 230000036449 good health Effects 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 description 1
- 244000052637 human pathogen Species 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 210000000936 intestine Anatomy 0.000 description 1
- 230000002147 killing effect Effects 0.000 description 1
- 239000000787 lecithin Substances 0.000 description 1
- 235000010445 lecithin Nutrition 0.000 description 1
- 239000006028 limestone Substances 0.000 description 1
- 229940087305 limonene Drugs 0.000 description 1
- 235000001510 limonene Nutrition 0.000 description 1
- 235000012680 lutein Nutrition 0.000 description 1
- 239000001656 lutein Substances 0.000 description 1
- KBPHJBAIARWVSC-RGZFRNHPSA-N lutein Chemical compound C([C@H](O)CC=1C)C(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\[C@H]1C(C)=C[C@H](O)CC1(C)C KBPHJBAIARWVSC-RGZFRNHPSA-N 0.000 description 1
- 229960005375 lutein Drugs 0.000 description 1
- ORAKUVXRZWMARG-WZLJTJAWSA-N lutein Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CCCC1(C)C)C=CC=C(/C)C=CC2C(=CC(O)CC2(C)C)C ORAKUVXRZWMARG-WZLJTJAWSA-N 0.000 description 1
- 235000009973 maize Nutrition 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 229930182817 methionine Natural products 0.000 description 1
- 235000006109 methionine Nutrition 0.000 description 1
- 229960004452 methionine Drugs 0.000 description 1
- 125000001360 methionine group Chemical group N[C@@H](CCSC)C(=O)* 0.000 description 1
- 238000009629 microbiological culture Methods 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 238000003801 milling Methods 0.000 description 1
- 235000013379 molasses Nutrition 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 229920001542 oligosaccharide Polymers 0.000 description 1
- 150000002482 oligosaccharides Chemical class 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 244000045947 parasite Species 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 238000009931 pascalization Methods 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 235000020232 peanut Nutrition 0.000 description 1
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 description 1
- 230000026731 phosphorylation Effects 0.000 description 1
- 238000006366 phosphorylation reaction Methods 0.000 description 1
- 238000005222 photoaffinity labeling Methods 0.000 description 1
- 239000000419 plant extract Substances 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 229920000223 polyglycerol Polymers 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 229920005862 polyol Polymers 0.000 description 1
- 150000003077 polyols Chemical class 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 230000004481 post-translational protein modification Effects 0.000 description 1
- 235000007686 potassium Nutrition 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- 229960003975 potassium Drugs 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 235000011056 potassium acetate Nutrition 0.000 description 1
- 229960004109 potassium acetate Drugs 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000000069 prophylactic effect Effects 0.000 description 1
- 229940070376 protein Drugs 0.000 description 1
- 244000000040 protozoan parasite Species 0.000 description 1
- 238000010188 recombinant method Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- WBHHMMIMDMUBKC-XLNAKTSKSA-N ricinelaidic acid Chemical compound CCCCCC[C@@H](O)C\C=C\CCCCCCCC(O)=O WBHHMMIMDMUBKC-XLNAKTSKSA-N 0.000 description 1
- FEUQNCSVHBHROZ-UHFFFAOYSA-N ricinoleic acid Natural products CCCCCCC(O[Si](C)(C)C)CC=CCCCCCCCC(=O)OC FEUQNCSVHBHROZ-UHFFFAOYSA-N 0.000 description 1
- 229960003656 ricinoleic acid Drugs 0.000 description 1
- 210000003296 saliva Anatomy 0.000 description 1
- 239000013535 sea water Substances 0.000 description 1
- 230000028327 secretion Effects 0.000 description 1
- 235000004400 serine Nutrition 0.000 description 1
- 239000004460 silage Substances 0.000 description 1
- 238000002741 site-directed mutagenesis Methods 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 235000015424 sodium Nutrition 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 229940083542 sodium Drugs 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 229940075554 sorbate Drugs 0.000 description 1
- 235000012424 soybean oil Nutrition 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
- 238000001256 steam distillation Methods 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000009885 systemic effect Effects 0.000 description 1
- 235000018553 tannin Nutrition 0.000 description 1
- 239000001648 tannin Substances 0.000 description 1
- 229920001864 tannin Polymers 0.000 description 1
- 210000001138 tear Anatomy 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 235000008521 threonine Nutrition 0.000 description 1
- KBPHJBAIARWVSC-XQIHNALSSA-N trans-lutein Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2C(=CC(O)CC2(C)C)C KBPHJBAIARWVSC-XQIHNALSSA-N 0.000 description 1
- 238000013519 translation Methods 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 241001515965 unidentified phage Species 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 238000001771 vacuum deposition Methods 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 235000019154 vitamin C Nutrition 0.000 description 1
- 239000011718 vitamin C Substances 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
- 239000002025 wood fiber Substances 0.000 description 1
- FJHBOVDFOQMZRV-XQIHNALSSA-N xanthophyll Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1=C(C)CC(O)CC1(C)C)C=CC=C(/C)C=CC2C=C(C)C(O)CC2(C)C FJHBOVDFOQMZRV-XQIHNALSSA-N 0.000 description 1
- 125000002256 xylenyl group Chemical class C1(C(C=CC=C1)C)(C)* 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
- 235000016804 zinc Nutrition 0.000 description 1
- OENHQHLEOONYIE-JLTXGRSLSA-N β-Carotene Chemical compound CC=1CCCC(C)(C)C=1\C=C\C(\C)=C\C=C\C(\C)=C\C=C\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C OENHQHLEOONYIE-JLTXGRSLSA-N 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K50/00—Feeding-stuffs specially adapted for particular animals
- A23K50/80—Feeding-stuffs specially adapted for particular animals for aquatic animals, e.g. fish, crustaceans or molluscs
-
- 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
- A23K20/105—Aliphatic or alicyclic compounds
-
- 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
- A23K20/189—Enzymes
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K40/00—Shaping or working-up of animal feeding-stuffs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/01—Hydrocarbons
- A61K31/015—Hydrocarbons carbocyclic
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/045—Hydroxy compounds, e.g. alcohols; Salts thereof, e.g. alcoholates
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/075—Ethers or acetals
- A61K31/085—Ethers or acetals having an ether linkage to aromatic ring nuclear carbon
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/43—Enzymes; Proenzymes; Derivatives thereof
- A61K38/46—Hydrolases (3)
- A61K38/47—Hydrolases (3) acting on glycosyl compounds (3.2), e.g. cellulases, lactases
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/24—Hydrolases (3) acting on glycosyl compounds (3.2)
- C12N9/2402—Hydrolases (3) acting on glycosyl compounds (3.2) hydrolysing O- and S- glycosyl compounds (3.2.1)
- C12N9/2462—Lysozyme (3.2.1.17)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y302/00—Hydrolases acting on glycosyl compounds, i.e. glycosylases (3.2)
- C12Y302/01—Glycosidases, i.e. enzymes hydrolysing O- and S-glycosyl compounds (3.2.1)
- C12Y302/01017—Lysozyme (3.2.1.17)
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2200/00—Function of food ingredients
- A23V2200/30—Foods, ingredients or supplements having a functional effect on health
- A23V2200/32—Foods, ingredients or supplements having a functional effect on health having an effect on the health of the digestive tract
Abstract
The present invention relates to the use of natural active substances selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with a polypeptide having lysozyme activity in the manufacture of a feed composition for aquatic animals, especially for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish. More particular, this invention relates to the use of a substance as defined above for the improvement of the feed conversion ratio and/or daily weight gain in fish, for regulating the micro flora of the gut and for protecting the animal against infections caused by pathogenic microorganisms, in particular gram+ microorganisms.
Description
2 PCT/EP2016/074407 FEED ADDITIVES FOR AQUATIC ANIMALS COMPRISING ESSENTIAL OILS AND LYSOZYME
The present invention relates to the use of natural essential oil actives in combi-nation with at least one polypeptide having lysozyme activity in the manufacture of feed for aquatic animals including fish and shrimp, especially for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish.
REFERENCE TO A SEQUENCE LISTING
This application contains a Sequence Listing in computer readable form, which is incorporated herein by reference.
SEQ ID NO: 1 is the mature amino acid sequence of a wild type GH25 lysozyme from Acremonium alcalophilum as described in WO 2013/076253.
SEQ ID NO: 2 is the gene sequence of the GH24 lysozyme as isolated from Tri-chophaea saccata.
SEQ ID NO: 3 is the amino acid sequence as deduced from SEQ ID NO: 2.
SEQ ID NO: 4 is the mature amino acid sequence of a wild type GH24 lysozyme from Trichophaea saccata.
SEQ ID NO: 5 is the mature amino acid sequence of a wild type GH22 lysozyme from Gallus gal/us (hen egg white lysozyme).
SEQ ID NO: 6 is primer F-80470.
SEQ ID NO: 7 is primer R-80470.
SEQ ID NO: 8 is primer 8643.
SEQ ID NO: 9 is primer 8654.
INTRODUCTION
The present invention relates to the use of natural essential oil actives selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, di-hydroeugenol, eugenol, meta-cresol and terpinolene in the manufacture of feed for aquatic animals including fish and shrimp, especially for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish.
More particular, this invention relates to the use of at least one, preferably at least two essential oil actives as defined above in combination with a polypeptide hav-ing lysozyme activity for the improvement of the feed conversion ratio and/or daily weight gain in fish, for reducing mortality by regulating the micro flora of the gut and/or by protecting the animal against infections caused by pathogenic microor-ganisms.
Lysozyme is an 0-glycosyl hydrolase produced as a defensive mechanism against bacteria by many organisms. The enzyme causes the hydrolysis of bacte-rial cell walls by cleaving the glycosidic bonds of peptidoglycan; an important structural molecule in bacteria. After having their cell walls weakened by lysozyme action, bacterial cells lyse resulting from osmotic pressure.
Lysozyme occurs in many organisms such as viruses, plants, insects, birds, rep-tiles and mammals. In mammals, Lysozyme has been isolated from nasal secre-tions, saliva, tears, intestines, urine and milk. The enzyme cleaves the glycosidic bond between carbon number 1 of N-acetylmuramic acid and carbon number 4 of N-acetyl-D-glucosamine. In vivo, these two carbohydrates are polymerized to form the cell wall polysaccharide.
Lysozyme has been classified into five different glycoside hydrolase (GH) families (CAZy, www.cazy.org): hen egg-white lysozyme (GH22), goose egg-white lyso-zyme (GH23), bacteriophage T4 lysozyme (GH24), Sphingomonas flagellar pro-tein (GH73) and Chalaropsis lysozymes (GH25). Lysozymes from the families GH23 and GH24 are primarily known from bacteriophages and have only recently been identified in fungi. The lysozyme family GH25 has been found to be struc-turally unrelated to the other lysozyme families.
Hen egg white lysozyme is the primary product available on the commercial mar-ket, but does not cleave N,6-0-diacetylmuramic acid in e.g. Staphylococcus aure-us cell walls and is thus unable to lyse this important human pathogen among others (Masschalck B, Deckers D, Michiels OW (2002), "Lytic and nonlytic mech-anism of inactivation of gram-positive bacteria by lysozyme under atmospheric and high hydrostatic pressure", J Food Prot. 65(12):1916-23).
The present invention relates to the use of natural essential oil actives in combi-nation with at least one polypeptide having lysozyme activity in the manufacture of feed for aquatic animals including fish and shrimp, especially for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish.
REFERENCE TO A SEQUENCE LISTING
This application contains a Sequence Listing in computer readable form, which is incorporated herein by reference.
SEQ ID NO: 1 is the mature amino acid sequence of a wild type GH25 lysozyme from Acremonium alcalophilum as described in WO 2013/076253.
SEQ ID NO: 2 is the gene sequence of the GH24 lysozyme as isolated from Tri-chophaea saccata.
SEQ ID NO: 3 is the amino acid sequence as deduced from SEQ ID NO: 2.
SEQ ID NO: 4 is the mature amino acid sequence of a wild type GH24 lysozyme from Trichophaea saccata.
SEQ ID NO: 5 is the mature amino acid sequence of a wild type GH22 lysozyme from Gallus gal/us (hen egg white lysozyme).
SEQ ID NO: 6 is primer F-80470.
SEQ ID NO: 7 is primer R-80470.
SEQ ID NO: 8 is primer 8643.
SEQ ID NO: 9 is primer 8654.
INTRODUCTION
The present invention relates to the use of natural essential oil actives selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, di-hydroeugenol, eugenol, meta-cresol and terpinolene in the manufacture of feed for aquatic animals including fish and shrimp, especially for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish.
More particular, this invention relates to the use of at least one, preferably at least two essential oil actives as defined above in combination with a polypeptide hav-ing lysozyme activity for the improvement of the feed conversion ratio and/or daily weight gain in fish, for reducing mortality by regulating the micro flora of the gut and/or by protecting the animal against infections caused by pathogenic microor-ganisms.
Lysozyme is an 0-glycosyl hydrolase produced as a defensive mechanism against bacteria by many organisms. The enzyme causes the hydrolysis of bacte-rial cell walls by cleaving the glycosidic bonds of peptidoglycan; an important structural molecule in bacteria. After having their cell walls weakened by lysozyme action, bacterial cells lyse resulting from osmotic pressure.
Lysozyme occurs in many organisms such as viruses, plants, insects, birds, rep-tiles and mammals. In mammals, Lysozyme has been isolated from nasal secre-tions, saliva, tears, intestines, urine and milk. The enzyme cleaves the glycosidic bond between carbon number 1 of N-acetylmuramic acid and carbon number 4 of N-acetyl-D-glucosamine. In vivo, these two carbohydrates are polymerized to form the cell wall polysaccharide.
Lysozyme has been classified into five different glycoside hydrolase (GH) families (CAZy, www.cazy.org): hen egg-white lysozyme (GH22), goose egg-white lyso-zyme (GH23), bacteriophage T4 lysozyme (GH24), Sphingomonas flagellar pro-tein (GH73) and Chalaropsis lysozymes (GH25). Lysozymes from the families GH23 and GH24 are primarily known from bacteriophages and have only recently been identified in fungi. The lysozyme family GH25 has been found to be struc-turally unrelated to the other lysozyme families.
Hen egg white lysozyme is the primary product available on the commercial mar-ket, but does not cleave N,6-0-diacetylmuramic acid in e.g. Staphylococcus aure-us cell walls and is thus unable to lyse this important human pathogen among others (Masschalck B, Deckers D, Michiels OW (2002), "Lytic and nonlytic mech-anism of inactivation of gram-positive bacteria by lysozyme under atmospheric and high hydrostatic pressure", J Food Prot. 65(12):1916-23).
- 3 - PCT/EP2016/074407 Furthermore, the present invention relates to a novel fish feed composition com-prising as active ingredients at least one, preferably at least two or three active compound(s) selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combi-nation with at least one microbial polypeptide having lysozyme activity.
One important factor in aquaculture is the turnover rate. Turnover rate is deter-mined by how fast the fish grow to a harvestable size. As an example, it takes from 12 to 18 months to raise Atlantic salmon from smolt (the physiological stage when the Atlantic salmon can first be transferred from fresh water to sea water) to harvestable size. A fast turnover has several positive results. First, it helps cash flow. Second, it improves risk management. Especially, a high mortality rate is a substantial risk for fish farmers.
It is generally known that mortality rate increases by an unbalanced microflora and/or by infections caused by pathogenic microbes. Fish diseases are common, and the likelihood of an outbreak is higher over a long growing period. There is also a risk that fish will escape due to accidents, e.g. when shifting nets, or due to bad weather causing wrecked fish pens.
For other farm animals it is well known to use antibiotics and vaccines to prevent the development of diseases. In aquaculture, antibiotics are not so much used -at least in cold water aquaculture - due to the fact that disease spread very quickly, diseased fish do not eat much and also due to the negative impact on the envi-ronment of the wasted medicated feed. Vaccines are widely used when available but they are not developed for all diseases.
As an alternative to synthetic drugs, the use of plant extracts and essential oils in animal feed is described in the literature. For example, patent W02011/006993 describes the use of essential oils in fish feed. W02011/006993 is however silent with regard to the protection against infections caused by gram+
microorganisms.
Moreover, the application of essential oils has in the disclosed case above only limited efficiency.
It therefore remains a need in aquaculture to prevent the development of diseas-es, thereby reducing mortality by any prophylactic means including antimicrobial activity against gram+ microorganisms.
The inventors of the present application surprisingly found that the combination of substances as defined above have a great potential for use in fish feed, e.g.
for
One important factor in aquaculture is the turnover rate. Turnover rate is deter-mined by how fast the fish grow to a harvestable size. As an example, it takes from 12 to 18 months to raise Atlantic salmon from smolt (the physiological stage when the Atlantic salmon can first be transferred from fresh water to sea water) to harvestable size. A fast turnover has several positive results. First, it helps cash flow. Second, it improves risk management. Especially, a high mortality rate is a substantial risk for fish farmers.
It is generally known that mortality rate increases by an unbalanced microflora and/or by infections caused by pathogenic microbes. Fish diseases are common, and the likelihood of an outbreak is higher over a long growing period. There is also a risk that fish will escape due to accidents, e.g. when shifting nets, or due to bad weather causing wrecked fish pens.
For other farm animals it is well known to use antibiotics and vaccines to prevent the development of diseases. In aquaculture, antibiotics are not so much used -at least in cold water aquaculture - due to the fact that disease spread very quickly, diseased fish do not eat much and also due to the negative impact on the envi-ronment of the wasted medicated feed. Vaccines are widely used when available but they are not developed for all diseases.
As an alternative to synthetic drugs, the use of plant extracts and essential oils in animal feed is described in the literature. For example, patent W02011/006993 describes the use of essential oils in fish feed. W02011/006993 is however silent with regard to the protection against infections caused by gram+
microorganisms.
Moreover, the application of essential oils has in the disclosed case above only limited efficiency.
It therefore remains a need in aquaculture to prevent the development of diseas-es, thereby reducing mortality by any prophylactic means including antimicrobial activity against gram+ microorganisms.
The inventors of the present application surprisingly found that the combination of substances as defined above have a great potential for use in fish feed, e.g.
for
- 4 - PCT/EP2016/074407 improving the feed conversion ratio (FOR) and/or weight gain and/or for the modu-lation of the gut flora. Further, the inventors surprisingly found that the novel fish feed compositions have also antimicrobial activity against gram+
microorganisms resulting in a reduced mortality. The unique selection of active compounds of the present invention allows for the first time controlling a number of fish diseases caused by a number of different pathogens.
Therefore, in a first particular embodiment, the invention relates to methods for using at least one, preferably at least two active compounds selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeuge-nol, eugenol, meta-cresol and terpinolene in combination with a microbial lyso-zyme in fish feed for improving the Feed Conversion Ratio (FOR) and/or weight gain and/or for reducing mortality by modulation of the gut microflora and/or by preventing diseases caused by pathogenic microorganisms, preferably caused by gram+ microorgansims.
For example, it has been shown that selected compounds of the invention (eg.:
cinnamaldehyde and alpha-pinene in combination with a bacterial lysozyme) ex-hibit excellent effects in inhibiting the growth of Lactococcus garvieae a disease found especially in cold water fish. The combination according to the present in-vention also exhibits excellent effects against Streptococcus iniae, a disease found especially in warm water fish.
In alternative embodiments, alpha-pinene and/or alpha-terpineol and/or cinnamal-dehyde and/or dihydroeugenol and/or eugenol and/or meta-cresol and/or terpino-lene is/are used in combination with at least one microbial polypeptide having ly-sozyme activity to improve animal feed digestibility and/or maintain animal health by supporting immune system function.
In another embodiment, the invention relates to methods for using at least three active compounds selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with at least one microbial polypeptide having lysozyme activity in fish feed for improving the Feed Conversion Ratio (FOR) and/or weight gain and/or for reducing mortality by modulation of the gut microflora and/or by pre-venting diseases caused by pathogenic microorganisms.
microorganisms resulting in a reduced mortality. The unique selection of active compounds of the present invention allows for the first time controlling a number of fish diseases caused by a number of different pathogens.
Therefore, in a first particular embodiment, the invention relates to methods for using at least one, preferably at least two active compounds selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeuge-nol, eugenol, meta-cresol and terpinolene in combination with a microbial lyso-zyme in fish feed for improving the Feed Conversion Ratio (FOR) and/or weight gain and/or for reducing mortality by modulation of the gut microflora and/or by preventing diseases caused by pathogenic microorganisms, preferably caused by gram+ microorgansims.
For example, it has been shown that selected compounds of the invention (eg.:
cinnamaldehyde and alpha-pinene in combination with a bacterial lysozyme) ex-hibit excellent effects in inhibiting the growth of Lactococcus garvieae a disease found especially in cold water fish. The combination according to the present in-vention also exhibits excellent effects against Streptococcus iniae, a disease found especially in warm water fish.
In alternative embodiments, alpha-pinene and/or alpha-terpineol and/or cinnamal-dehyde and/or dihydroeugenol and/or eugenol and/or meta-cresol and/or terpino-lene is/are used in combination with at least one microbial polypeptide having ly-sozyme activity to improve animal feed digestibility and/or maintain animal health by supporting immune system function.
In another embodiment, the invention relates to methods for using at least three active compounds selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with at least one microbial polypeptide having lysozyme activity in fish feed for improving the Feed Conversion Ratio (FOR) and/or weight gain and/or for reducing mortality by modulation of the gut microflora and/or by pre-venting diseases caused by pathogenic microorganisms.
- 5 - PCT/EP2016/074407 In a preferred embodiment, alpha-pinene and cinnamaldehyde are used in combi-nation with a bacterial lysozyme to improve animal feed digestibility and/or main-tain animal health by supporting immune system function.
DEFINITIONS
The term feed or feed composition means any compound, preparation, mixture, or composition suitable for, or intended for intake by an animal.
The FOR may be determined on the basis of a fish growth trial comprising a first treatment in which a mixture of at least two compounds according to the invention is added to the animal feed in a suitable concentration per kg feed, and a second treatment (control) with no addition of the compound(s) to the animal feed.
As it is generally known, an improved FOR is lower than the control FOR. In particular embodiments, the FOR is improved (i.e., reduced) as compared to the control by at least 1.0 (Yo, preferably at least 1.5 (Yo, 1.6 (Yo, 1.7 (Yo, 1.8 (Yo, 1.9 (Yo, 2.0 (Yo, 2.1 (Yo, 2.2 (Yo, 2.3 (Yo, 2.4 (Yo, or at least 2.5 (Yo.
The term "gut" as used herein designates the gastrointestinal or digestive tract (also referred to as the alimentary canal) and it refers to the system of organs within multi-cellular animals which takes in food, digests it to extract energy and nutrients, and expels the remaining waste.
The term gut "microflora" as used herein refers to the natural microbial cultures residing in the gut and maintaining health by aiding in proper digestion.
The term "modulate" as used herein in connection with the gut microflora generally means to change, manipulate, alter, or adjust the function or status thereof in a healthy and normally functioning animal, i.e. a non-therapeutic use.
The term "supporting immune system function" as used herein refers to the immune stimulation effect obtained by the compounds.
The term "mortality" as used herein refers to the ratio of life animals at the end of the growth phase versus the number of animals originally included into the pond.
It may be determined on the basis of a fish challenge trial comprising two groups of fish challenged by a particular fish pathogen with the aim to provoke a mortality of 40 to 80 % of the animals in the untreated group. However, in the challenge group fed with a suitable concentration per Kg of feed of a mixture of at least two compounds according to the invention, the mortality is reduced compared to the
DEFINITIONS
The term feed or feed composition means any compound, preparation, mixture, or composition suitable for, or intended for intake by an animal.
The FOR may be determined on the basis of a fish growth trial comprising a first treatment in which a mixture of at least two compounds according to the invention is added to the animal feed in a suitable concentration per kg feed, and a second treatment (control) with no addition of the compound(s) to the animal feed.
As it is generally known, an improved FOR is lower than the control FOR. In particular embodiments, the FOR is improved (i.e., reduced) as compared to the control by at least 1.0 (Yo, preferably at least 1.5 (Yo, 1.6 (Yo, 1.7 (Yo, 1.8 (Yo, 1.9 (Yo, 2.0 (Yo, 2.1 (Yo, 2.2 (Yo, 2.3 (Yo, 2.4 (Yo, or at least 2.5 (Yo.
The term "gut" as used herein designates the gastrointestinal or digestive tract (also referred to as the alimentary canal) and it refers to the system of organs within multi-cellular animals which takes in food, digests it to extract energy and nutrients, and expels the remaining waste.
The term gut "microflora" as used herein refers to the natural microbial cultures residing in the gut and maintaining health by aiding in proper digestion.
The term "modulate" as used herein in connection with the gut microflora generally means to change, manipulate, alter, or adjust the function or status thereof in a healthy and normally functioning animal, i.e. a non-therapeutic use.
The term "supporting immune system function" as used herein refers to the immune stimulation effect obtained by the compounds.
The term "mortality" as used herein refers to the ratio of life animals at the end of the growth phase versus the number of animals originally included into the pond.
It may be determined on the basis of a fish challenge trial comprising two groups of fish challenged by a particular fish pathogen with the aim to provoke a mortality of 40 to 80 % of the animals in the untreated group. However, in the challenge group fed with a suitable concentration per Kg of feed of a mixture of at least two compounds according to the invention, the mortality is reduced compared to the
- 6 - PCT/EP2016/074407 untreated group by at least 5 "Yo, preferably at least, 10 "Yo, 15 "Yo, 20 "Yo, 25 "Yo, 30 "Yo, 35 "Yo, 40 "Yo, 45 "Yo, or at least 50 %.
Animal feed: The term "animal feed" refers to any compound, preparation, or mix-ture suitable for, or intended for intake by an animal. Animal feed for a monogas-tric animal typically comprises concentrates as well as vitamins, minerals, en-zymes, direct fed microbial, amino acids and/or other feed ingredients (such as in a premix) whereas animal feed for ruminants generally comprises forage (includ-ing roughage and silage) and may further comprise concentrates as well as vita-mins, minerals, enzymes direct fed microbial, amino acid and/or other feed ingre-dients (such as in a premix).
Antimicrobial activity: The term "antimicrobial activity" is defined herein as an activity that kills or inhibits the growth of microorganisms, such as, algae, archea, bacteria, fungi and/or protozoans. The antimicrobial activity can for example be bactericidal meaning the killing of bacteria or bacteriostatic meaning the preven-tion of bacterial growth. The antimicrobial activity can include catalyzing the hy-drolysis of 1,4-beta-linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine residues in a peptidoglycan and between N-acetyl-D-glucosamine residues in chitodextrins. Antimicrobial activity can also include the lysozyme binding to the surface of the microorganism and inhibiting its growth. The antimi-crobial effect can also include the use of the lysozymes of the present invention for activation of bacterial autolysins, as an immunostimulator, by inhibiting or re-ducing bacterial toxins and by an opsonin effect.
For the purpose of the present invention, antimicrobial activity is determined ac-cording to the antimicrobial assay described in Example 6 ("Determination of an-timicrobial activity"). Antimicrobial activity is determined by the diameter of the clearing zone and is evaluated as significant if the clearing zone is more than 2 mm using 50% Mueller¨Hinton broth, pH 6.
Concentrates: The term "concentrates" means feed with high protein and energy concentrations, such as fish meal, molasses, oligosaccharides, sorghum, seeds and grains (either whole or prepared by crushing, milling, etc. from e.g.
corn, oats, rye, barley, wheat), oilseed press cake (e.g. from cottonseed, safflower, sunflow-er, soybean (such as soybean meal), rapeseed/canola, peanut or groundnut), palm kernel cake, yeast derived material and distillers grains (such as wet distill-ers grains (WDS) and dried distillers grains with solubles (DDGS)).
Animal feed: The term "animal feed" refers to any compound, preparation, or mix-ture suitable for, or intended for intake by an animal. Animal feed for a monogas-tric animal typically comprises concentrates as well as vitamins, minerals, en-zymes, direct fed microbial, amino acids and/or other feed ingredients (such as in a premix) whereas animal feed for ruminants generally comprises forage (includ-ing roughage and silage) and may further comprise concentrates as well as vita-mins, minerals, enzymes direct fed microbial, amino acid and/or other feed ingre-dients (such as in a premix).
Antimicrobial activity: The term "antimicrobial activity" is defined herein as an activity that kills or inhibits the growth of microorganisms, such as, algae, archea, bacteria, fungi and/or protozoans. The antimicrobial activity can for example be bactericidal meaning the killing of bacteria or bacteriostatic meaning the preven-tion of bacterial growth. The antimicrobial activity can include catalyzing the hy-drolysis of 1,4-beta-linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine residues in a peptidoglycan and between N-acetyl-D-glucosamine residues in chitodextrins. Antimicrobial activity can also include the lysozyme binding to the surface of the microorganism and inhibiting its growth. The antimi-crobial effect can also include the use of the lysozymes of the present invention for activation of bacterial autolysins, as an immunostimulator, by inhibiting or re-ducing bacterial toxins and by an opsonin effect.
For the purpose of the present invention, antimicrobial activity is determined ac-cording to the antimicrobial assay described in Example 6 ("Determination of an-timicrobial activity"). Antimicrobial activity is determined by the diameter of the clearing zone and is evaluated as significant if the clearing zone is more than 2 mm using 50% Mueller¨Hinton broth, pH 6.
Concentrates: The term "concentrates" means feed with high protein and energy concentrations, such as fish meal, molasses, oligosaccharides, sorghum, seeds and grains (either whole or prepared by crushing, milling, etc. from e.g.
corn, oats, rye, barley, wheat), oilseed press cake (e.g. from cottonseed, safflower, sunflow-er, soybean (such as soybean meal), rapeseed/canola, peanut or groundnut), palm kernel cake, yeast derived material and distillers grains (such as wet distill-ers grains (WDS) and dried distillers grains with solubles (DDGS)).
- 7 - PCT/EP2016/074407 European Production Efficacy Factor (EPEF): The European Production Effica-cy Factor is a way of comparing the performance of animals. This single-figure fa-cilitates comparison of performance within and among farms and can be used to assess environmental, climatic and managemental variables. The EPEF is calcu-lated as [(liveability CYO x Liveweight (kg)) / (Age at depletion (days) x FOR)] x 100, wherein livability is the percentage of animals alive at slaughter, Liveweight is the average weight of the animals at slaughter, age of depletion is the age of the animals at slaughter and FOR is the feed conversion ratio at slaughter.
Feed Conversion Ratio (FCR): FOR is a measure of an animal's efficiency in converting feed mass into increases of the desired output. Animals raised for meat ¨ such as swine, poultry and fish ¨ the output is the mass gained by the an-imal. Specifically FOR is calculated as feed intake divided by weight gain, all over a specified period. Improvement in FOR means reduction of the FOR value. A
FOR improvement of 2% means that the FOR was reduced by 2%.
Fragment: The term "fragment" means a polypeptide or a catalytic domain having one or more (e.g., several) amino acids absent from the amino and/or carboxyl terminus of a mature polypeptide or domain; wherein the fragment has lysozyme or phytase activity. In one aspect, a fragment comprises at least 170 amino acids, such as at least 175 amino acids, at least 177 amino acids, at least 180 amino ac-ids, at least 185 amino acids, at least 190 amino acids, at least 195 amino acids or at least 200 amino acids of SEQ ID NO: 1 and has lysozyme activity.
In another aspect, a fragment comprises at least 210 amino acids, such as at least 215 amino acids, at least 220 amino acids, at least 225 amino acids, at least 230 amino acids, at least 235 amino acids or at least 240 amino acids of SEQ
ID
NO: 4 and has lysozyme activity.
Isolated: The term "isolated" means a substance in a form or environment that does not occur in nature. Non-limiting examples of isolated substances include (1) any non-naturally occurring substance, (2) any substance including, but not lim-ited to, any enzyme, variant, nucleic acid, protein, peptide or cofactor, that is at least partially removed from one or more or all of the naturally occurring constitu-ents with which it is associated in nature; (3) any substance modified by the hand of man relative to that substance found in nature; or (4) any substance modified by increasing the amount of the substance relative to other components with which it is naturally associated (e.g., multiple copies of a gene encoding the sub-stance; use of a stronger promoter than the promoter naturally associated with the
Feed Conversion Ratio (FCR): FOR is a measure of an animal's efficiency in converting feed mass into increases of the desired output. Animals raised for meat ¨ such as swine, poultry and fish ¨ the output is the mass gained by the an-imal. Specifically FOR is calculated as feed intake divided by weight gain, all over a specified period. Improvement in FOR means reduction of the FOR value. A
FOR improvement of 2% means that the FOR was reduced by 2%.
Fragment: The term "fragment" means a polypeptide or a catalytic domain having one or more (e.g., several) amino acids absent from the amino and/or carboxyl terminus of a mature polypeptide or domain; wherein the fragment has lysozyme or phytase activity. In one aspect, a fragment comprises at least 170 amino acids, such as at least 175 amino acids, at least 177 amino acids, at least 180 amino ac-ids, at least 185 amino acids, at least 190 amino acids, at least 195 amino acids or at least 200 amino acids of SEQ ID NO: 1 and has lysozyme activity.
In another aspect, a fragment comprises at least 210 amino acids, such as at least 215 amino acids, at least 220 amino acids, at least 225 amino acids, at least 230 amino acids, at least 235 amino acids or at least 240 amino acids of SEQ
ID
NO: 4 and has lysozyme activity.
Isolated: The term "isolated" means a substance in a form or environment that does not occur in nature. Non-limiting examples of isolated substances include (1) any non-naturally occurring substance, (2) any substance including, but not lim-ited to, any enzyme, variant, nucleic acid, protein, peptide or cofactor, that is at least partially removed from one or more or all of the naturally occurring constitu-ents with which it is associated in nature; (3) any substance modified by the hand of man relative to that substance found in nature; or (4) any substance modified by increasing the amount of the substance relative to other components with which it is naturally associated (e.g., multiple copies of a gene encoding the sub-stance; use of a stronger promoter than the promoter naturally associated with the
- 8 - PCT/EP2016/074407 gene encoding the substance). An isolated substance may be present in a fer-mentation broth sample.
Lysozyme activity: The term "lysozyme activity" means the hydrolysis of the 1,4-beta-linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine resi-dues in a peptidoglycan or between N-acetyl-D-glucosamine residues in chitodex-trins, resulting in bacteriolysis. Lysozyme belongs to the enzyme class EC
3.2.1.17. Lysozyme activity is typically measured by turbidimetric determination.
The method is based on the changes in turbidity of a suspension of Micrococcus luteus ATCC 4698 induced by the lytic action of lysozyme. In appropriate experi-mental conditions these changes are proportional to the amount of lysozyme in the medium (c.f. INS 1105 of the Combined Compendium of Food Additive Speci-fications of the Food and Agriculture Organisation of the UN (www.fao.org)).
For the purpose of the present invention, lysozyme activity is determined according to the turbidity assay described in example 5 ("Determination of Lysozyme Activity").
In one aspect, the polypeptides of the present invention have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the lysozyme activity of SEQ ID NO: 1. In anoth-er aspect, the polypeptides of the present invention have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the lysozyme activity of SEQ ID NO: 4.
Mature polypeptide: The term "mature polypeptide" means a polypeptide in its final form following translation and any post-translational modifications, such as N-terminal processing, C-terminal truncation, glycosylation, phosphorylation, etc.
Microbial lysozyme: The term "microbial lysozyme" means a polypeptide having lysozyme activity which is obtained or obtainable from a microbial source.
Exam-ples of microbial sources are fungi. In particular, the the microbial lysozyme is ob-tained or obtainable from the phylum Ascomycota, such as the sub-phylum Pezizomycotina.
Sequence identity: The relatedness between two amino acid sequences or be-tween two nucleotide sequences is described by the parameter "sequence identi-ty".
For purposes of the present invention, the sequence identity between two amino acid sequences is determined using the Needleman-Wunsch algorithm (Needle-man and Wunsch, 1970, J. Mol. Biol. 48: 443-453) as implemented in the Needle
Lysozyme activity: The term "lysozyme activity" means the hydrolysis of the 1,4-beta-linkages between N-acetylmuramic acid and N-acetyl-D-glucosamine resi-dues in a peptidoglycan or between N-acetyl-D-glucosamine residues in chitodex-trins, resulting in bacteriolysis. Lysozyme belongs to the enzyme class EC
3.2.1.17. Lysozyme activity is typically measured by turbidimetric determination.
The method is based on the changes in turbidity of a suspension of Micrococcus luteus ATCC 4698 induced by the lytic action of lysozyme. In appropriate experi-mental conditions these changes are proportional to the amount of lysozyme in the medium (c.f. INS 1105 of the Combined Compendium of Food Additive Speci-fications of the Food and Agriculture Organisation of the UN (www.fao.org)).
For the purpose of the present invention, lysozyme activity is determined according to the turbidity assay described in example 5 ("Determination of Lysozyme Activity").
In one aspect, the polypeptides of the present invention have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the lysozyme activity of SEQ ID NO: 1. In anoth-er aspect, the polypeptides of the present invention have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the lysozyme activity of SEQ ID NO: 4.
Mature polypeptide: The term "mature polypeptide" means a polypeptide in its final form following translation and any post-translational modifications, such as N-terminal processing, C-terminal truncation, glycosylation, phosphorylation, etc.
Microbial lysozyme: The term "microbial lysozyme" means a polypeptide having lysozyme activity which is obtained or obtainable from a microbial source.
Exam-ples of microbial sources are fungi. In particular, the the microbial lysozyme is ob-tained or obtainable from the phylum Ascomycota, such as the sub-phylum Pezizomycotina.
Sequence identity: The relatedness between two amino acid sequences or be-tween two nucleotide sequences is described by the parameter "sequence identi-ty".
For purposes of the present invention, the sequence identity between two amino acid sequences is determined using the Needleman-Wunsch algorithm (Needle-man and Wunsch, 1970, J. Mol. Biol. 48: 443-453) as implemented in the Needle
- 9 - PCT/EP2016/074407 program of the EMBOSS package (EMBOSS: The European Molecular Biology Open Software Suite, Rice et al., 2000, Trends Genet. 16: 276-277), preferably version 5Ø0 or later. The parameters used are gap open penalty of 10, gap ex-tension penalty of 0.5, and the EBLOSUM62 (EMBOSS version of BLOSUM62) substitution matrix. The output of Needle labeled "longest identity" (obtained using the -nobrief option) is used as the percent identity and is calculated as follows:
(Identical Residues x 100)/(Length of Alignment - Total Number of Gaps in Alignment) Substantially pure polypeptide: The term "substantially pure polypeptide"
means a preparation that contains at most 10%, at most 8%, at most 6%, at most 5%, at most 4%, at most 3%, at most 2%, at most 1`)/0, and at most 0.5% by weight of other polypeptide material with which it is natively or recombinantly associated.
Preferably, the polypeptide is at least 92% pure, e.g., at least 94% pure, at least 95% pure, at least 96% pure, at least 97% pure, at least 98% pure, at least 99%, at least 99.5% pure, and 100% pure by weight of the total polypeptide material present in the preparation. The polypeptides of the present invention are prefera-bly in a substantially pure form. This can be accomplished, for example, by pre-paring the polypeptide by well known recombinant methods or by classical purifi-cation methods.
Variant: The term "variant" means a polypeptide having lysozyme or phytase ac-tivity comprising an alteration, i.e., a substitution, insertion, and/or deletion, of one or more (several) amino acid residues at one or more (e.g., several) positions. A
substitution means replacement of the amino acid occupying a position with a dif-ferent amino acid; a deletion means removal of the amino acid occupying a posi-tion; and an insertion means adding 1, 2, or 3 amino acids adjacent to and imme-diately following the amino acid occupying the position.
In one aspect, a lysozyme variant according to the invention may comprise from to 5; from 1 to 10; from 1 to 15; from 1 to 20; from 1 to 25; from 1 to 30;
from 1 to 35; from 1 to 40; from 1 to 45; or from 1-50, i.e. 1, 2, 3, 4, 5, 6, 7, 8, 9,
(Identical Residues x 100)/(Length of Alignment - Total Number of Gaps in Alignment) Substantially pure polypeptide: The term "substantially pure polypeptide"
means a preparation that contains at most 10%, at most 8%, at most 6%, at most 5%, at most 4%, at most 3%, at most 2%, at most 1`)/0, and at most 0.5% by weight of other polypeptide material with which it is natively or recombinantly associated.
Preferably, the polypeptide is at least 92% pure, e.g., at least 94% pure, at least 95% pure, at least 96% pure, at least 97% pure, at least 98% pure, at least 99%, at least 99.5% pure, and 100% pure by weight of the total polypeptide material present in the preparation. The polypeptides of the present invention are prefera-bly in a substantially pure form. This can be accomplished, for example, by pre-paring the polypeptide by well known recombinant methods or by classical purifi-cation methods.
Variant: The term "variant" means a polypeptide having lysozyme or phytase ac-tivity comprising an alteration, i.e., a substitution, insertion, and/or deletion, of one or more (several) amino acid residues at one or more (e.g., several) positions. A
substitution means replacement of the amino acid occupying a position with a dif-ferent amino acid; a deletion means removal of the amino acid occupying a posi-tion; and an insertion means adding 1, 2, or 3 amino acids adjacent to and imme-diately following the amino acid occupying the position.
In one aspect, a lysozyme variant according to the invention may comprise from to 5; from 1 to 10; from 1 to 15; from 1 to 20; from 1 to 25; from 1 to 30;
from 1 to 35; from 1 to 40; from 1 to 45; or from 1-50, i.e. 1, 2, 3, 4, 5, 6, 7, 8, 9,
10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 alterations and have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the phytase activity of the parent phytase, such as SEQ ID NO: 1. In another aspect, a lysozyme variant according to the invention may comprise from 1 to 5; from 1 to 10; from 1 to 15;
from 1 to 20; from 1 to 25; from 1 to 30; from 1 to 35; from 1 to 40; from 1 to 45; or from 1-50, i.e. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 alterations and have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the lysozyme activity of the parent lysozyme, such as SEQ ID NO: 4.
DETAILED DESCRIPTION OF THE INVENTION
In particular, the inventors of the present application surprisingly found that the compound combinations according to the invention are effective against a number pathogenic gram+ microorganisms of cold and warm water fish. Compositions ac-cording to the invention were shown to exhibit inhibitory effect against Lactococ-cus garvieae a disease found especially in cold water fish and against Strepto-coccus iniae, a disease found especially in warm water fish.
Furthermore, the use of at least one microbial polypeptide having lysozyme activi-ty in combination as defined above further improves existing inhibitory effects of natural essential oil actives.
Known inhibitory effects of essential oils are:
Alpha-pinene, cinnamaldehyde, dihydroeugenol, eugenol, limonene, and meta-cresol were shown to exhibit inhibitory effect against Aeromonas salmonicida which is the pathogen causing a disease known as furunculosis.
Alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene were shown to exhibit inhibitory effect against Edwardsellia tarda causing systemic infection in fish.
Alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, and terpinolene were shown to exhibit inhibitory effect against Lactococcus garvieae which is the etiological agent of Latococcosis, an emergent disease which affects many fish species and causes important economic losses both in marine and freshwater aquaculture when water temperature increases over 16 C in summer months.
Cinnamaldehyde, dihydroeugenol, eugenol, and meta-cresol exhibit excellent in-hibitory effects on the growth of Yersinia ruckeri, a pathogenic microorganism which causes Enteric Redmouth (ERM), a disease found especially in salmonids.
from 1 to 20; from 1 to 25; from 1 to 30; from 1 to 35; from 1 to 40; from 1 to 45; or from 1-50, i.e. 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 alterations and have at least 20%, e.g., at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 100% of the lysozyme activity of the parent lysozyme, such as SEQ ID NO: 4.
DETAILED DESCRIPTION OF THE INVENTION
In particular, the inventors of the present application surprisingly found that the compound combinations according to the invention are effective against a number pathogenic gram+ microorganisms of cold and warm water fish. Compositions ac-cording to the invention were shown to exhibit inhibitory effect against Lactococ-cus garvieae a disease found especially in cold water fish and against Strepto-coccus iniae, a disease found especially in warm water fish.
Furthermore, the use of at least one microbial polypeptide having lysozyme activi-ty in combination as defined above further improves existing inhibitory effects of natural essential oil actives.
Known inhibitory effects of essential oils are:
Alpha-pinene, cinnamaldehyde, dihydroeugenol, eugenol, limonene, and meta-cresol were shown to exhibit inhibitory effect against Aeromonas salmonicida which is the pathogen causing a disease known as furunculosis.
Alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene were shown to exhibit inhibitory effect against Edwardsellia tarda causing systemic infection in fish.
Alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, and terpinolene were shown to exhibit inhibitory effect against Lactococcus garvieae which is the etiological agent of Latococcosis, an emergent disease which affects many fish species and causes important economic losses both in marine and freshwater aquaculture when water temperature increases over 16 C in summer months.
Cinnamaldehyde, dihydroeugenol, eugenol, and meta-cresol exhibit excellent in-hibitory effects on the growth of Yersinia ruckeri, a pathogenic microorganism which causes Enteric Redmouth (ERM), a disease found especially in salmonids.
- 11 - PCT/EP2016/074407 Cinnamaldehyde and meta-cresol were shown to exhibit inhibitory effect against:
- Vibrio salmonicida which is a psychrophilic bacterium that is the causative agent of cold-water vibriosis in Atlantic salmon.
- Aeromonas hydrophila causing ulcers and hemorrhagic septicaemia. This pathogen is very resistant to conventional simple antimicrobials like chlorine.
- Photobacterium damselae formerly Pasteurella piscicida: a pathogen causing high losses in the culture industry of economically important marine fishes such as seriola and red grouper in Japan and striped bass and white perch in the United States.
- Streptococcus iniae which is highly pathogenic in marine fish and is highly le-thal: outbreaks may be associated with 30-50 (:)/0 mortality.
Other aquatic pathogens such as - Piscirickettsia salmonis the causative agent of piscirickettsiosis or salmonid rickettsial septicaemia (SRS), - Vibrio viscosus recently renamed Monte/la viscosa etiologically responsible for the disease referred to as "winter ulcer", - lch (parasite) one of the most prevalent protozoan parasites of fish, - Vibrio harveyi, responsible for luminous vibriosis, a disease that affects com-mercially-farmed prawns will also be inhibited by the compound mixture described in the present invention.
In a second aspect, the present invention provides a fish feed composition com-prising at least one microbial peptide having lysozyme activity in combination with at least two active compounds selected from the group consisting of alpha-pinene (CAS 99-86-5), alpha-terpineol (CAS 98-55-5), cinnamaldehyde (CAS 14371-10-9 /104-55-2), dihydroeugenol (CAS 2785-87-7), eugenol (CAS 97-53-0), meta-cresol (CAS 108-39-4) and terpinolene (CAS 554-61-0). The compounds accord-ing to the invention are commercially available or can easily be prepared by a skilled person using processes and methods well-known in the prior art.
- Vibrio salmonicida which is a psychrophilic bacterium that is the causative agent of cold-water vibriosis in Atlantic salmon.
- Aeromonas hydrophila causing ulcers and hemorrhagic septicaemia. This pathogen is very resistant to conventional simple antimicrobials like chlorine.
- Photobacterium damselae formerly Pasteurella piscicida: a pathogen causing high losses in the culture industry of economically important marine fishes such as seriola and red grouper in Japan and striped bass and white perch in the United States.
- Streptococcus iniae which is highly pathogenic in marine fish and is highly le-thal: outbreaks may be associated with 30-50 (:)/0 mortality.
Other aquatic pathogens such as - Piscirickettsia salmonis the causative agent of piscirickettsiosis or salmonid rickettsial septicaemia (SRS), - Vibrio viscosus recently renamed Monte/la viscosa etiologically responsible for the disease referred to as "winter ulcer", - lch (parasite) one of the most prevalent protozoan parasites of fish, - Vibrio harveyi, responsible for luminous vibriosis, a disease that affects com-mercially-farmed prawns will also be inhibited by the compound mixture described in the present invention.
In a second aspect, the present invention provides a fish feed composition com-prising at least one microbial peptide having lysozyme activity in combination with at least two active compounds selected from the group consisting of alpha-pinene (CAS 99-86-5), alpha-terpineol (CAS 98-55-5), cinnamaldehyde (CAS 14371-10-9 /104-55-2), dihydroeugenol (CAS 2785-87-7), eugenol (CAS 97-53-0), meta-cresol (CAS 108-39-4) and terpinolene (CAS 554-61-0). The compounds accord-ing to the invention are commercially available or can easily be prepared by a skilled person using processes and methods well-known in the prior art.
- 12 - PCT/EP2016/074407 As fish feed composition, the compounds of the invention can be used alone or in mixtures thereof, in the form of natural available extracts or extract-mixtures or in the form of a natural substance.
The term "extract" as used herein includes compositions obtained by solvent ex-traction (which are also known as "extracted oils"), steam distillation (which are also known as "essential oils") or other methods known to the skilled person.
Suitable extraction solvents include alcohols such as ethanol.
The term "natural" is in this context understood a substance which consists of compounds occurring in nature and obtained from natural products or through synthesis.
To the active essential oil compound(s) or natural substance or extract further in-gredients may be added in minor amounts. Examples of such ingredients are:
capsaicin, tannin, piperin, trimethylamine, 3,4,xylenol, furfuryl alcohol and mix-tures thereof.
If a mixture of at least two essential oil compounds as specified above is pre-ferred, cinnamaldehyde and/or meta-cresol are used as a major component of the mixture. Suitably the mixture contains 10-90 "Yo by weight of meta-cresol and/or cinnamaldehyde, 1-50 "Yo by weight of alpha-pinene, 1-50 "Yo by weight of dihydro-eugenol, wherein the amounts being calculated on the total amount of said com-ponents. The total amount of these active ingredients may vary within wide limits but is finally used in the fish feed from 10 to 5000 ppm, preferably between and 1000 ppm, calculated on the dry weight of the fish feed.
The most preferred mixture of at least two essential oil compounds as specified above comprises, cinnamaldehyde and alpha-pinene as a major component of the mixture. Suitably the mixture contains 30-70 (:)/0 by weight of alpha-pinene, "Yo by weight cinnamaldehyde, 1-20 (:)/0 by weight of metacresol, 1-20 (:)/0 by weight of dihydro-eugenol, wherein the amounts being calculated on the total amount of said components. The total amount of these active ingredients may vary within wide limits but is finally used in the fish feed from 10 to 5000 ppm, preferably be-tween 100 and 1000 ppm, calculated on the dry weight of the fish feed.
All essential oil compounds defined herein above (active compounds and addi-tional ingredients) may be used in combination with an emulsifying surfactant.
The emulsifying agent can be selected advantageously from those of a rather hydro-philic nature, for example among polyglycerol esters of fatty acids such as esteri-
The term "extract" as used herein includes compositions obtained by solvent ex-traction (which are also known as "extracted oils"), steam distillation (which are also known as "essential oils") or other methods known to the skilled person.
Suitable extraction solvents include alcohols such as ethanol.
The term "natural" is in this context understood a substance which consists of compounds occurring in nature and obtained from natural products or through synthesis.
To the active essential oil compound(s) or natural substance or extract further in-gredients may be added in minor amounts. Examples of such ingredients are:
capsaicin, tannin, piperin, trimethylamine, 3,4,xylenol, furfuryl alcohol and mix-tures thereof.
If a mixture of at least two essential oil compounds as specified above is pre-ferred, cinnamaldehyde and/or meta-cresol are used as a major component of the mixture. Suitably the mixture contains 10-90 "Yo by weight of meta-cresol and/or cinnamaldehyde, 1-50 "Yo by weight of alpha-pinene, 1-50 "Yo by weight of dihydro-eugenol, wherein the amounts being calculated on the total amount of said com-ponents. The total amount of these active ingredients may vary within wide limits but is finally used in the fish feed from 10 to 5000 ppm, preferably between and 1000 ppm, calculated on the dry weight of the fish feed.
The most preferred mixture of at least two essential oil compounds as specified above comprises, cinnamaldehyde and alpha-pinene as a major component of the mixture. Suitably the mixture contains 30-70 (:)/0 by weight of alpha-pinene, "Yo by weight cinnamaldehyde, 1-20 (:)/0 by weight of metacresol, 1-20 (:)/0 by weight of dihydro-eugenol, wherein the amounts being calculated on the total amount of said components. The total amount of these active ingredients may vary within wide limits but is finally used in the fish feed from 10 to 5000 ppm, preferably be-tween 100 and 1000 ppm, calculated on the dry weight of the fish feed.
All essential oil compounds defined herein above (active compounds and addi-tional ingredients) may be used in combination with an emulsifying surfactant.
The emulsifying agent can be selected advantageously from those of a rather hydro-philic nature, for example among polyglycerol esters of fatty acids such as esteri-
- 13 - PCT/EP2016/074407 fied ricinoleic acid or propylene glycol esters of fatty acids, saccharo-esters or saccharo-glycerides, polyethylene glycol, lecithins etc.
In a preferred embodiment of the invention, a mixture of essential oils may contain 20 % by weight of cinnamaldehyde, 20 % by weight of meta-cresol, 20 % by weight of dihydro-eugenol, 20 % by weight of alpha-pinene, 3 % by weight of tri-methylamine, 1.8% by weight of piperin and 4% by weight of furfuryl alcohol.
In another preferred embodiment of the invention, a mixture of essential oil active compounds contains 40 to 60 wt.-% alpha-pinene, 40 to 60 wt.-% cinnamalde-hyde, and may further contain 1 to 5 wt.-% trimethylamine, 1 to 5 % piperin, and 3 to 8 wt.-% furfuryl alcohol.
As mentioned above, it has been surprisingly found that supplementing an animal feed with a microbial lysozyme results in a significant performance benefit in aquatic animals including fish and shrimp. This is surprising since improved ani-mal performance using a microbial lysozyme has never previously been demon-strated.
Thus the invention also relates to a method of improving the European Production Efficiency Factor (EPEF) comprising administering an animal feed or animal feed additive comprising at least two essential oil compounds and one or more micro-bial lysozymes to the animal.
In a preferred embodiment, the improvement is compared to an animal feed or an-imal feed additive wherein the microbial lysozyme is not present (herein referred to as the control).
In one embodiment, the EPEF is improved by at least 1`)/0, such as by at least 1.5%, at least 2.0%, at least 2.5%, at least 3%, at least 3.5%, at least 4% or at least 5% compared to the control. In another embodiment, the EPEF is improved by between 1% and 15%, such as between 1% and 12%, between 1% and 10%, 1.5% and 8%, 2.0% and 7% compared to the control, or any combination of these intervals.
In one embodiment, the microbial lysozyme is dosed at a level of 8 to 250 ppm enzyme protein per kg animal feed, such as 9 to 200 ppm, 10 to 150 ppm, 11 to 125 ppm, 12 to 100 ppm, 13 to 75 ppm, 15 to 50 ppm, 17.5 to 40 ppm, 25 to 75 ppm or 30 to 60 ppm enzyme protein per kg animal feed, or any combination of these intervals.
In a preferred embodiment of the invention, a mixture of essential oils may contain 20 % by weight of cinnamaldehyde, 20 % by weight of meta-cresol, 20 % by weight of dihydro-eugenol, 20 % by weight of alpha-pinene, 3 % by weight of tri-methylamine, 1.8% by weight of piperin and 4% by weight of furfuryl alcohol.
In another preferred embodiment of the invention, a mixture of essential oil active compounds contains 40 to 60 wt.-% alpha-pinene, 40 to 60 wt.-% cinnamalde-hyde, and may further contain 1 to 5 wt.-% trimethylamine, 1 to 5 % piperin, and 3 to 8 wt.-% furfuryl alcohol.
As mentioned above, it has been surprisingly found that supplementing an animal feed with a microbial lysozyme results in a significant performance benefit in aquatic animals including fish and shrimp. This is surprising since improved ani-mal performance using a microbial lysozyme has never previously been demon-strated.
Thus the invention also relates to a method of improving the European Production Efficiency Factor (EPEF) comprising administering an animal feed or animal feed additive comprising at least two essential oil compounds and one or more micro-bial lysozymes to the animal.
In a preferred embodiment, the improvement is compared to an animal feed or an-imal feed additive wherein the microbial lysozyme is not present (herein referred to as the control).
In one embodiment, the EPEF is improved by at least 1`)/0, such as by at least 1.5%, at least 2.0%, at least 2.5%, at least 3%, at least 3.5%, at least 4% or at least 5% compared to the control. In another embodiment, the EPEF is improved by between 1% and 15%, such as between 1% and 12%, between 1% and 10%, 1.5% and 8%, 2.0% and 7% compared to the control, or any combination of these intervals.
In one embodiment, the microbial lysozyme is dosed at a level of 8 to 250 ppm enzyme protein per kg animal feed, such as 9 to 200 ppm, 10 to 150 ppm, 11 to 125 ppm, 12 to 100 ppm, 13 to 75 ppm, 15 to 50 ppm, 17.5 to 40 ppm, 25 to 75 ppm or 30 to 60 ppm enzyme protein per kg animal feed, or any combination of these intervals.
- 14 - PCT/EP2016/074407 In one embodiment, the microbial lysozyme is of fungal origin. In an embodiment, the microbial lysozyme is obtained or obtainable from the phylum Ascomycota, such as the sub-phylum Pezizomycotina.
In one embodiment, the microbial lysozyme comprises one or more domains se-lected from the list consisting of GH24 and GH25.
In a preferred embodiment, the invention relates to a method of improving the Eu-ropean Production Efficiency Factor (EPEF) and/or feed conversion ratio (FOR) comprising administering an animal feed or animal feed additive comprising one or more microbial lysozymes to the fish, wherein:
(a) the microbial lysozyme is a microbial lysozyme comprising one or more do-mains selected from the list consisting of GH24 and GH25, is dosed at a level of 10 to 150 ppm enzyme protein per kg animal feed;
(b) the aquatic animal is a selected from shrimp, for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish and;
(c) European Production Efficiency Factor (EPEF) and/or feed conversion ratio (FOR) is improved by at least 1% compared to control.
In one embodiment, the microbial lysozyme is of fungal origin. In an embodiment, the microbial lysozyme is obtained or obtainable from the phylum Ascomycota, such as the sub-phylum Pezizomycotina.
In one embodiment, the microbial lysozyme has at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91`)/0, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 1.
In one embodiment, the microbial lysozyme comprises or consists of the amino acid sequence of SEQ ID NO: 1 or an allelic variant thereof; or is a fragment thereof having lysozyme activity, wherein the fragment comprises at least 170 amino acids, such as at least 175 amino acids, at least 177 amino acids, at least 180 amino acids, at least 185 amino acids, at least 190 amino acids, at least amino acids or at least 200 amino acids. In another embodiment, the microbial ly-sozyme comprises or consists of the amino acid sequence of SEQ ID NO: 1 or an
In one embodiment, the microbial lysozyme comprises one or more domains se-lected from the list consisting of GH24 and GH25.
In a preferred embodiment, the invention relates to a method of improving the Eu-ropean Production Efficiency Factor (EPEF) and/or feed conversion ratio (FOR) comprising administering an animal feed or animal feed additive comprising one or more microbial lysozymes to the fish, wherein:
(a) the microbial lysozyme is a microbial lysozyme comprising one or more do-mains selected from the list consisting of GH24 and GH25, is dosed at a level of 10 to 150 ppm enzyme protein per kg animal feed;
(b) the aquatic animal is a selected from shrimp, for cold water fish as for example salmon, bream, bass and for warm water fish as for example carp, tilapia, catfish and;
(c) European Production Efficiency Factor (EPEF) and/or feed conversion ratio (FOR) is improved by at least 1% compared to control.
In one embodiment, the microbial lysozyme is of fungal origin. In an embodiment, the microbial lysozyme is obtained or obtainable from the phylum Ascomycota, such as the sub-phylum Pezizomycotina.
In one embodiment, the microbial lysozyme has at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91`)/0, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 1.
In one embodiment, the microbial lysozyme comprises or consists of the amino acid sequence of SEQ ID NO: 1 or an allelic variant thereof; or is a fragment thereof having lysozyme activity, wherein the fragment comprises at least 170 amino acids, such as at least 175 amino acids, at least 177 amino acids, at least 180 amino acids, at least 185 amino acids, at least 190 amino acids, at least amino acids or at least 200 amino acids. In another embodiment, the microbial ly-sozyme comprises or consists of the amino acid sequence of SEQ ID NO: 1 or an
- 15 - PCT/EP2016/074407 allelic variant thereof and a N-terminal and/or C-terminal His-tag and/or HQ-tag.
In another aspect, the polypeptide comprises or consists of amino acids 1 to of SEQ ID NO: 1.
In another embodiment, the microbial lysozyme is a variant of SEQ ID NO: 1 wherein the variant has lysozyme activity and comprises one or more substitu-tions, and/or one or more deletions, and/or one or more insertions or any combi-nation thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions. In another embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combina-tion thereof in SEQ ID NO: 1 is between 1 and 45, such as 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ
ID
NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodi-ment, the number of substitutions, preferably conservative substitutions, in SEQ
ID NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further em-bodiment, the number of conservative substitutions in SEQ ID NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.
The amino acid changes may be of a minor nature, that is conservative amino ac-id substitutions or insertions that do not significantly affect the folding and/or activ-ity of the protein; small deletions, typically of 1-30 amino acids; small amino- or carboxyl-terminal extensions, such as an amino-terminal methionine residue; a small linker peptide of up to 20-25 residues; or a small extension that facilitates purification by changing net charge or another function, such as a poly-histidine tract, an antigenic epitope or a binding domain.
Examples of conservative substitutions are within the groups of basic amino acids (arginine, lysine and histidine), acidic amino acids (glutamic acid and aspartic ac-id), polar amino acids (glutamine and asparagine), hydrophobic amino acids (leu-cine, isoleucine and valine), aromatic amino acids (phenylalanine, tryptophan and tyrosine), and small amino acids (glycine, alanine, serine, threonine and methio-nine). Amino acid substitutions that do not generally alter specific activity are known in the art and are described, for example, by H. Neurath and R.L. Hill,
In another aspect, the polypeptide comprises or consists of amino acids 1 to of SEQ ID NO: 1.
In another embodiment, the microbial lysozyme is a variant of SEQ ID NO: 1 wherein the variant has lysozyme activity and comprises one or more substitu-tions, and/or one or more deletions, and/or one or more insertions or any combi-nation thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions. In another embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combina-tion thereof in SEQ ID NO: 1 is between 1 and 45, such as 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ
ID
NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodi-ment, the number of substitutions, preferably conservative substitutions, in SEQ
ID NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further em-bodiment, the number of conservative substitutions in SEQ ID NO: 1 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.
The amino acid changes may be of a minor nature, that is conservative amino ac-id substitutions or insertions that do not significantly affect the folding and/or activ-ity of the protein; small deletions, typically of 1-30 amino acids; small amino- or carboxyl-terminal extensions, such as an amino-terminal methionine residue; a small linker peptide of up to 20-25 residues; or a small extension that facilitates purification by changing net charge or another function, such as a poly-histidine tract, an antigenic epitope or a binding domain.
Examples of conservative substitutions are within the groups of basic amino acids (arginine, lysine and histidine), acidic amino acids (glutamic acid and aspartic ac-id), polar amino acids (glutamine and asparagine), hydrophobic amino acids (leu-cine, isoleucine and valine), aromatic amino acids (phenylalanine, tryptophan and tyrosine), and small amino acids (glycine, alanine, serine, threonine and methio-nine). Amino acid substitutions that do not generally alter specific activity are known in the art and are described, for example, by H. Neurath and R.L. Hill,
- 16 - PCT/EP2016/074407 1979, In, The Proteins, Academic Press, New York. Common substitutions are Ala/Ser, Val/Ile, Asp/Glu, Thr/Ser, Ala/Gly, Ala/Thr, Ser/Asn, Ala/Val, Ser/Gly, Tyr/Phe, Ala/Pro, Lys/Arg, Asp/Asn, Leu/Ile, Leu/Val, Ala/Glu, and Asp/Gly.
Essential amino acids in a polypeptide can be identified according to procedures known in the art, such as site-directed mutagenesis or alanine-scanning muta-genesis (Cunningham and Wells, 1989, Science 244: 1081-1085). In the latter technique, single alanine mutations are introduced at every residue in the mole-cule, and the resultant mutant molecules are tested for lysozyme activity to identi-fy amino acid residues that are critical to the activity of the molecule. See also, Hilton et al., 1996, J. Biol. Chem. 271: 4699-4708. The active site of the enzyme or other biological interaction can also be determined by physical analysis of structure, as determined by such techniques as nuclear magnetic resonance, crystallography, electron diffraction, or photoaffinity labeling, in conjunction with mutation of putative contact site amino acids. See, for example, de Vos et al., 1992, Science 255: 306-312; Smith et al., 1992, J. Mol. Biol. 224: 899-904;
Wlodaver et al., 1992, FEBS Lett. 309: 59-64. The identity of essential amino ac-ids can also be inferred from an alignment with a related polypeptide.
The crystal structure of the Acremonium alcalophilum CBS114.92 lysozyme was solved at a resolution of 1.3 A as disclosed in WO 2013/076253. These atomic coordinates can be used to generate a three dimensional model depicting the structure of the Acremonium alcalophilum CBS114.92 lysozyme or homologous structures (such as the variants of the present invention). Using the x/ray struc-ture, amino acid residues D95 and E97 (using SEQ ID NO: 1 for numbering) were identified as catalytic residues.
In one embodiment, the microbial lysozyme has at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91`)/0, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 4.
In one embodiment, the microbial lysozyme comprises or consists of the amino acid sequence of SEQ ID NO: 4 or an allelic variant thereof; or is a fragment thereof having lysozyme activity, wherein the fragment comprises at least 210 amino acids, such as at least 215 amino acids, at least 220 amino acids, at least 225 amino acids, at least 230 amino acids, at least 235 amino acids or at least 240 amino acids. In another embodiment, the microbial lysozyme comprises or
Essential amino acids in a polypeptide can be identified according to procedures known in the art, such as site-directed mutagenesis or alanine-scanning muta-genesis (Cunningham and Wells, 1989, Science 244: 1081-1085). In the latter technique, single alanine mutations are introduced at every residue in the mole-cule, and the resultant mutant molecules are tested for lysozyme activity to identi-fy amino acid residues that are critical to the activity of the molecule. See also, Hilton et al., 1996, J. Biol. Chem. 271: 4699-4708. The active site of the enzyme or other biological interaction can also be determined by physical analysis of structure, as determined by such techniques as nuclear magnetic resonance, crystallography, electron diffraction, or photoaffinity labeling, in conjunction with mutation of putative contact site amino acids. See, for example, de Vos et al., 1992, Science 255: 306-312; Smith et al., 1992, J. Mol. Biol. 224: 899-904;
Wlodaver et al., 1992, FEBS Lett. 309: 59-64. The identity of essential amino ac-ids can also be inferred from an alignment with a related polypeptide.
The crystal structure of the Acremonium alcalophilum CBS114.92 lysozyme was solved at a resolution of 1.3 A as disclosed in WO 2013/076253. These atomic coordinates can be used to generate a three dimensional model depicting the structure of the Acremonium alcalophilum CBS114.92 lysozyme or homologous structures (such as the variants of the present invention). Using the x/ray struc-ture, amino acid residues D95 and E97 (using SEQ ID NO: 1 for numbering) were identified as catalytic residues.
In one embodiment, the microbial lysozyme has at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91`)/0, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 4.
In one embodiment, the microbial lysozyme comprises or consists of the amino acid sequence of SEQ ID NO: 4 or an allelic variant thereof; or is a fragment thereof having lysozyme activity, wherein the fragment comprises at least 210 amino acids, such as at least 215 amino acids, at least 220 amino acids, at least 225 amino acids, at least 230 amino acids, at least 235 amino acids or at least 240 amino acids. In another embodiment, the microbial lysozyme comprises or
- 17 - PCT/EP2016/074407 consists of the amino acid sequence of SEQ ID NO: 4 or an allelic variant thereof and a N-terminal and/or C-terminal His-tag and/or HQ-tag. In another aspect, the polypeptide comprises or consists of amino acids 1 to 245 of SEQ ID NO: 4.
In another embodiment, the microbial lysozyme is a variant of SEQ ID NO: 4 wherein the variant has lysozyme activity and comprises one or more substitu-tions, and/or one or more deletions, and/or one or more insertions or any combi-nation thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,
In another embodiment, the microbial lysozyme is a variant of SEQ ID NO: 4 wherein the variant has lysozyme activity and comprises one or more substitu-tions, and/or one or more deletions, and/or one or more insertions or any combi-nation thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17,
18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions. In another embodiment, the number to of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combina-tion thereof in SEQ ID NO: 4 is between 1 and 45, such as 1-40, 1-35, 1-30, 1-25, 1-20, 1-15, 1-10 or 1-5 positions. In an embodiment, the number of positions comprising one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in SEQ ID NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In another embodiment, the number of substitutions, deletions, and/or insertions in SEQ
ID
NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodi-ment, the number of substitutions, preferably conservative substitutions, in SEQ
ID NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further em-bodiment, the number of conservative substitutions in SEQ ID NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.
Examples of amino acid changes, conservative substitutions and N- and/or C-terminal linkers are described above.
The enzyme of the invention may be formulated as a liquid or a solid. For a liquid formulation, the formulating agent may comprise a polyol (such as e.g.
glycerol, ethylene glycol or propylene glycol), a salt (such as e.g. sodium chloride, sodium benzoate, potassium sorbate) or a sugar or sugar derivative (such as e.g.
dextrin, glucose, sucrose, and sorbitol). Thus in one embodiment, the composition is a liquid composition comprising the enzyme(s) of the invention and one or more formulating agents selected from the list consisting of glycerol, ethylene glycol, 1,2-propylene glycol, 1,3-propylene glycol, sodium chloride, sodium benzoate, po-tassium sorbate, dextrin, glucose, sucrose, and sorbitol. The liquid formulation may be sprayed onto the feed after it has been pelleted or may be added to drink-ing water given to the animals.
For a solid formulation, the formulation may be for example as a granule, spray dried powder or agglomerate. The formulating agent may comprise a salt (organic or inorganic zinc, sodium, potassium or calcium salts such as e.g. such as calcium acetate, calcium benzoate, calcium carbonate, calcium chloride, calcium citrate, calcium sorbate, calcium sulfate, potassium acetate, potassium benzoate, potas-sium carbonate, potassium chloride, potassium citrate, potassium sorbate, potas-sium sulfate, sodium acetate, sodium benzoate, sodium carbonate, sodium chlo-ride, sodium citrate, sodium sulfate, zinc acetate, zinc benzoate, zinc carbonate, zinc chloride, zinc citrate, zinc sorbate, zinc sulfate), starch or a sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sorbitol).
In an embodiment, the solid composition is in granulated form. The granule may have a matrix structure where the components are mixed homogeneously. How-ever, the granule typically comprises a core particle and one or more coatings, which typically are salt and/or wax coatings. The core particle can either be a homogeneous blend of the enzyme(s) of the invention optionally combined with one or more additional enzymes and optionally together with one or more salts or an inert particle with the enzyme(s) of the invention optionally combined with one or more additional enzymes applied onto it.
In an embodiment, the material of the core particles are selected from the group consisting of inorganic salts (such as calcium acetate, calcium benzoate, calcium carbonate, calcium chloride, calcium citrate, calcium sorbate, calcium sulfate, po-tassium acetate, potassium benzoate, potassium carbonate, potassium chloride, potassium citrate, potassium sorbate, potassium sulfate, sodium acetate, sodium benzoate, sodium carbonate, sodium chloride, sodium citrate, sodium sulfate, zinc acetate, zinc benzoate, zinc carbonate, zinc chloride, zinc citrate, zinc sorbate, zinc sulfate), starch or a sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sorbitol), sugar or sugar derivative (such as e.g. sucrose, dex-trin, glucose, lactose, sorbitol), small organic molecules, starch, flour, cellulose and minerals.
The salt coating is typically at least 1 pm thick and can either be one particular salt or a mixture of salts, such as Na2SO4, K2SO4, MgSO4 and/or sodium citrate.
Other examples are those described in e.g. WO 2008/017659, WO 2006/034710, WO 1997/05245, WO 1998/54980, WO 1998/55599, WO 2000/70034 or polymer coating such as described in WO 2001/00042.
ID
NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further embodi-ment, the number of substitutions, preferably conservative substitutions, in SEQ
ID NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10. In a further em-bodiment, the number of conservative substitutions in SEQ ID NO: 4 is not more than 10, e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10.
Examples of amino acid changes, conservative substitutions and N- and/or C-terminal linkers are described above.
The enzyme of the invention may be formulated as a liquid or a solid. For a liquid formulation, the formulating agent may comprise a polyol (such as e.g.
glycerol, ethylene glycol or propylene glycol), a salt (such as e.g. sodium chloride, sodium benzoate, potassium sorbate) or a sugar or sugar derivative (such as e.g.
dextrin, glucose, sucrose, and sorbitol). Thus in one embodiment, the composition is a liquid composition comprising the enzyme(s) of the invention and one or more formulating agents selected from the list consisting of glycerol, ethylene glycol, 1,2-propylene glycol, 1,3-propylene glycol, sodium chloride, sodium benzoate, po-tassium sorbate, dextrin, glucose, sucrose, and sorbitol. The liquid formulation may be sprayed onto the feed after it has been pelleted or may be added to drink-ing water given to the animals.
For a solid formulation, the formulation may be for example as a granule, spray dried powder or agglomerate. The formulating agent may comprise a salt (organic or inorganic zinc, sodium, potassium or calcium salts such as e.g. such as calcium acetate, calcium benzoate, calcium carbonate, calcium chloride, calcium citrate, calcium sorbate, calcium sulfate, potassium acetate, potassium benzoate, potas-sium carbonate, potassium chloride, potassium citrate, potassium sorbate, potas-sium sulfate, sodium acetate, sodium benzoate, sodium carbonate, sodium chlo-ride, sodium citrate, sodium sulfate, zinc acetate, zinc benzoate, zinc carbonate, zinc chloride, zinc citrate, zinc sorbate, zinc sulfate), starch or a sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sorbitol).
In an embodiment, the solid composition is in granulated form. The granule may have a matrix structure where the components are mixed homogeneously. How-ever, the granule typically comprises a core particle and one or more coatings, which typically are salt and/or wax coatings. The core particle can either be a homogeneous blend of the enzyme(s) of the invention optionally combined with one or more additional enzymes and optionally together with one or more salts or an inert particle with the enzyme(s) of the invention optionally combined with one or more additional enzymes applied onto it.
In an embodiment, the material of the core particles are selected from the group consisting of inorganic salts (such as calcium acetate, calcium benzoate, calcium carbonate, calcium chloride, calcium citrate, calcium sorbate, calcium sulfate, po-tassium acetate, potassium benzoate, potassium carbonate, potassium chloride, potassium citrate, potassium sorbate, potassium sulfate, sodium acetate, sodium benzoate, sodium carbonate, sodium chloride, sodium citrate, sodium sulfate, zinc acetate, zinc benzoate, zinc carbonate, zinc chloride, zinc citrate, zinc sorbate, zinc sulfate), starch or a sugar or sugar derivative (such as e.g. sucrose, dextrin, glucose, lactose, sorbitol), sugar or sugar derivative (such as e.g. sucrose, dex-trin, glucose, lactose, sorbitol), small organic molecules, starch, flour, cellulose and minerals.
The salt coating is typically at least 1 pm thick and can either be one particular salt or a mixture of salts, such as Na2SO4, K2SO4, MgSO4 and/or sodium citrate.
Other examples are those described in e.g. WO 2008/017659, WO 2006/034710, WO 1997/05245, WO 1998/54980, WO 1998/55599, WO 2000/70034 or polymer coating such as described in WO 2001/00042.
- 19 - PCT/EP2016/074407 In another embodiment, the composition is a solid composition comprising the en-zyme(s) of the invention and one or more formulating agents selected from the list consisting of sodium chloride, sodium benzoate, potassium sorbate, sodium sul-fate, potassium sulfate, magnesium sulfate, sodium thiosulfate, calcium car-s bonate, sodium citrate, dextrin, glucose, sucrose, sorbitol, lactose, starch and cel-lulose. In a preferred embodiment, the formulating agent is selected from one or more of the following compounds: sodium sulfate, dextrin, cellulose, sodium thio-sulfate and calcium carbonate. In a preferred embodiment, the solid composition is in granulated form. In an embodiment, the solid composition is in granulated form and comprises a core particle, an enzyme layer comprising the enzyme(s) of the invention and a salt coating.
In a further embodiment, the formulating agent is selected from one or more of the following compounds: glycerol, ethylene glycol, 1, 2-propylene glycol or 1, 3-propylene glycol, sodium chloride, sodium benzoate, potassium sorbate, sodium sulfate, potassium sulfate, magnesium sulfate, sodium thiosulfate, calcium car-bonate, sodium citrate, dextrin, glucose, sucrose, sorbitol, lactose, starch and cel-lulose. In a preferred embodiment, the formulating agent is selected from one or more of the following compounds: 1, 2-propylene glycol, 1, 3-propylene glycol, sodium sulfate, dextrin, cellulose, sodium thiosulfate and calcium carbonate.
The incorporation of the fish feed composition containing the active compound(s) into the fish feed may be performed as described in example 1. The mixture of ac-tive compounds is then prepared directly as an oil which is then mixed with the oil sprayed onto the feed pellets as described in example 1.
The incorporation of the fish feed composition containing the active compound(s) into the fish feed may alternatively be carried out by preparing a premix of the ac-tive ingredients and other suitable additives. Such a premix may comprise 2-10 (:)/0 by weight of the active mixture or natural substance or extract, 0-40 (:)/0 by weight of other conventional additives, such as flavorings, and 50-98 (:)/0 by weight of any conventional absorbing support.
The support may contain, for example, 40-50 (:)/0 by weight of wood fibers, 8-10 (:)/0 by weight of stearin, 4-5 (:)/0 by weight of curcuma powder, 4-5 (:)/0 by weight of rosemary powder, 22-28 (:)/0 by weight of limestone, 1-3 (:)/0 by weight of a gum, such as gum Arabic, 5-50 (:)/0 by weight of sugar and/or starch and 5-15 (:)/0 by weight of water.
In a further embodiment, the formulating agent is selected from one or more of the following compounds: glycerol, ethylene glycol, 1, 2-propylene glycol or 1, 3-propylene glycol, sodium chloride, sodium benzoate, potassium sorbate, sodium sulfate, potassium sulfate, magnesium sulfate, sodium thiosulfate, calcium car-bonate, sodium citrate, dextrin, glucose, sucrose, sorbitol, lactose, starch and cel-lulose. In a preferred embodiment, the formulating agent is selected from one or more of the following compounds: 1, 2-propylene glycol, 1, 3-propylene glycol, sodium sulfate, dextrin, cellulose, sodium thiosulfate and calcium carbonate.
The incorporation of the fish feed composition containing the active compound(s) into the fish feed may be performed as described in example 1. The mixture of ac-tive compounds is then prepared directly as an oil which is then mixed with the oil sprayed onto the feed pellets as described in example 1.
The incorporation of the fish feed composition containing the active compound(s) into the fish feed may alternatively be carried out by preparing a premix of the ac-tive ingredients and other suitable additives. Such a premix may comprise 2-10 (:)/0 by weight of the active mixture or natural substance or extract, 0-40 (:)/0 by weight of other conventional additives, such as flavorings, and 50-98 (:)/0 by weight of any conventional absorbing support.
The support may contain, for example, 40-50 (:)/0 by weight of wood fibers, 8-10 (:)/0 by weight of stearin, 4-5 (:)/0 by weight of curcuma powder, 4-5 (:)/0 by weight of rosemary powder, 22-28 (:)/0 by weight of limestone, 1-3 (:)/0 by weight of a gum, such as gum Arabic, 5-50 (:)/0 by weight of sugar and/or starch and 5-15 (:)/0 by weight of water.
- 20 - PCT/EP2016/074407 This premix is then mixed with vitamins, as for example vitamin C, mineral salts and other feed additive ingredients, as for example yeast extracts containing nu-cleotides, glucan and other gut microflora modulators and then finally added to the feed in such quantities that the feed will contain 10-5000 ppm, preferably 1000 ppm or 100-500 ppm of the active ingredients according to the invention.
Moreover, the composition of the present invention will be preferably used togeth-er with yeast extract containing nucleotides, and glucan.
Further, optional, feed-additive ingredients are coloring agents, e.g.
carotenoids such as beta-carotene, astaxanthin, and lutein; aroma compounds; stabilisers;
antimicrobial peptides; polyunsaturated fatty acids; and/or at least one enzyme selected from amongst phytase (EC 3.1.3.8 or 3.1.3.26); xylanase (EC 3.2.1.8);
galactanase (EC 3.2.1.89); alpha-galactosidase (EC 3.2.1.22); protease (EC
3.4.), phospholipase Al (EC 3.1.1.32); phospholipase A2 (EC 3.1.1.4);
lysophospholipase (EC 3.1.1.5); phospholipase C (EC 3.1.4.3); phospholipase D
(EC 3.1.4.4); amylase such as, for example, alpha-amylase (EC 3.2.1.1); and/or beta-glucanase (EC 3.2.1.4 or EC 3.2.1.6).
Examples of polyunsaturated fatty acids are 018, 020 and 022 polyunsaturated fatty acids, such as arachidonic acid, docosohexaenoic acid, eicosapentaenoic acid and gamma-linoleic acid.
The fish feed as described herein has a proximate composition of 20-60 wt.-%
protein, and 1-45 wt.-% moisture and lipid.
In some specific examples, the fish feed comprises one or more of sources of:
- protein, carbohydrate and lipid (for example, fish meal, fish oil, blood meal, feather meal, poultry meal, chicken meal and/or other types of meal produced from other slaughterhouse waste), - animal fat (for example poultry oil), - vegetable meal (e.g. soya meal, lupin meal, pea meal, bean meal, rape meal and/or sunflower meal), - vegetable oil (e.g. rapeseed oil, soya oil), - gluten (e.g. wheat gluten or corn gluten) and - added amino acids (e.g. lysine) The term "fish feed" as used herein includes a fish feed composition according to the invention and components as described above. Typically, fish feed includes
Moreover, the composition of the present invention will be preferably used togeth-er with yeast extract containing nucleotides, and glucan.
Further, optional, feed-additive ingredients are coloring agents, e.g.
carotenoids such as beta-carotene, astaxanthin, and lutein; aroma compounds; stabilisers;
antimicrobial peptides; polyunsaturated fatty acids; and/or at least one enzyme selected from amongst phytase (EC 3.1.3.8 or 3.1.3.26); xylanase (EC 3.2.1.8);
galactanase (EC 3.2.1.89); alpha-galactosidase (EC 3.2.1.22); protease (EC
3.4.), phospholipase Al (EC 3.1.1.32); phospholipase A2 (EC 3.1.1.4);
lysophospholipase (EC 3.1.1.5); phospholipase C (EC 3.1.4.3); phospholipase D
(EC 3.1.4.4); amylase such as, for example, alpha-amylase (EC 3.2.1.1); and/or beta-glucanase (EC 3.2.1.4 or EC 3.2.1.6).
Examples of polyunsaturated fatty acids are 018, 020 and 022 polyunsaturated fatty acids, such as arachidonic acid, docosohexaenoic acid, eicosapentaenoic acid and gamma-linoleic acid.
The fish feed as described herein has a proximate composition of 20-60 wt.-%
protein, and 1-45 wt.-% moisture and lipid.
In some specific examples, the fish feed comprises one or more of sources of:
- protein, carbohydrate and lipid (for example, fish meal, fish oil, blood meal, feather meal, poultry meal, chicken meal and/or other types of meal produced from other slaughterhouse waste), - animal fat (for example poultry oil), - vegetable meal (e.g. soya meal, lupin meal, pea meal, bean meal, rape meal and/or sunflower meal), - vegetable oil (e.g. rapeseed oil, soya oil), - gluten (e.g. wheat gluten or corn gluten) and - added amino acids (e.g. lysine) The term "fish feed" as used herein includes a fish feed composition according to the invention and components as described above. Typically, fish feed includes
- 21 - PCT/EP2016/074407 fish meal as a component. Suitably, fish feed is in the form of flakes or pellets, for example extruded pellets.
In a third aspect, the invention relates to a feed composition for aquatic animals and to the use of this composition for feeding fish. The feed is particularly suitable for feeding salmonids, including Atlantic salmon (Salmo salar), other salmon spe-cies and trout, and non-salmonids such as cod, sea bass, sea bream and eel.
However, it can be fed to all types of fish, for example turbot, halibut, yellow tail and tuna.
The invention described and claimed herein is not to be limited in scope by the specific embodiments herein disclosed, since these embodiments are intended as illustrations of several aspects of the invention. Any equivalent embodiments are intended to be within the scope of this invention. Indeed, various modifications of the invention in addition to those shown and described herein will become appar-ent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims.
Example 1: Preparation of Pressed Fish Feed The main raw materials are ground and mixed. Microingredients are then added to the mixer and the homogenous mix is conditioned by adding water and steam to the mass in a preconditioner. This starts a cooking process in the starch fraction (the binding component). The mass is fed into a pellet mill. The mass is forced through the mill's die and the strings are broken into pellets on the outside of the die. The moisture content is low and drying of the feed is not necessary.
Additional oil including a fish feed composition according to the present invention is then sprayed onto the surface of pellets, but as the pellets are rather compact, the total lipid content rarely exceeds 24 (Yo. The added oil may be fish oil or vegetable oils, for example rape seed oil or soy oil, or a mixture of vegetable oils or a mixture of fish oil and vegetable oils. After oil coating, the pellets are cooled in a cooler and bagged. The final pressed fish feed contains 10 to 5000 ppm of the composition as described in the invention.
Example 2: Method for Preparation of Extruded Fish Feed The main raw materials are ground and mixed. Micro ingredients incl. a fish feed composition according to the invention are added to the mixer. The homogenous mix is conditioned by adding water and steam to the mass in a preconditioner.
In a third aspect, the invention relates to a feed composition for aquatic animals and to the use of this composition for feeding fish. The feed is particularly suitable for feeding salmonids, including Atlantic salmon (Salmo salar), other salmon spe-cies and trout, and non-salmonids such as cod, sea bass, sea bream and eel.
However, it can be fed to all types of fish, for example turbot, halibut, yellow tail and tuna.
The invention described and claimed herein is not to be limited in scope by the specific embodiments herein disclosed, since these embodiments are intended as illustrations of several aspects of the invention. Any equivalent embodiments are intended to be within the scope of this invention. Indeed, various modifications of the invention in addition to those shown and described herein will become appar-ent to those skilled in the art from the foregoing description. Such modifications are also intended to fall within the scope of the appended claims.
Example 1: Preparation of Pressed Fish Feed The main raw materials are ground and mixed. Microingredients are then added to the mixer and the homogenous mix is conditioned by adding water and steam to the mass in a preconditioner. This starts a cooking process in the starch fraction (the binding component). The mass is fed into a pellet mill. The mass is forced through the mill's die and the strings are broken into pellets on the outside of the die. The moisture content is low and drying of the feed is not necessary.
Additional oil including a fish feed composition according to the present invention is then sprayed onto the surface of pellets, but as the pellets are rather compact, the total lipid content rarely exceeds 24 (Yo. The added oil may be fish oil or vegetable oils, for example rape seed oil or soy oil, or a mixture of vegetable oils or a mixture of fish oil and vegetable oils. After oil coating, the pellets are cooled in a cooler and bagged. The final pressed fish feed contains 10 to 5000 ppm of the composition as described in the invention.
Example 2: Method for Preparation of Extruded Fish Feed The main raw materials are ground and mixed. Micro ingredients incl. a fish feed composition according to the invention are added to the mixer. The homogenous mix is conditioned by adding water and steam to the mass in a preconditioner.
- 22 - PCT/EP2016/074407 Additional oil may also be added to the mass at this stage. This starts a cooking process in the starch fraction (the binding component). The mass is fed into an extruder. The extruder may be of the single screw or the twin-screw type. Due to the rotational movement of the mass in the extruder, the mass is further mixed.
Additional oil, water and steam may be added to the mass in the extruder. At the end of the extruder, the mass has a temperature above 100 C and a pressure above ambient pressure. The mass is forced through the openings in the extruder's die plate. Due to the relief in temperature and pressure, some of the moisture will evaporate immediately (flash off) and the extruded mass becomes porous. The strings are cut into pellets by a rotating knife. The water content is rather high (18-28 %) and the pellets are therefore immediately dried to approximately 10 % water content in a dryer.
After the dryer, more oil may be added to the feed by spraying oil onto the surface of the feed, or by dipping the feed in oil. It is advantageous to add the oil to the feed in a closed vessel where the air pressure is below ambient (vacuum coating) so that the porous feed pellets absorb more oil. Feed containing more than 40 %
lipid may be produced this way. After the coater, the feed is cooled and bagged.
Oil may be added at several places in the process as explained above, and may be fish oil or vegetable oils, by example rape seed oil or soy oil, or a mixture of vegetable oils or a mixture of fish oil and vegetable oils.
Fish need protein, fat, minerals and vitamins in order to grow and to be in good health. The diet of carnivorous fish is particularly important. Originally in the farming of carnivorous fish, whole fish or ground fish were used to meet the nutritional requirements of the farmed fish. Ground fish mixed with dry raw materials of various kinds, such as fish meal and starch, was termed soft or semi-moist feed. As farming became industrialized, soft or semi-moist feed was replaced by pressed dry feed. This was itself gradually replaced by extruded dry feed.
Today, extruded feed is nearly universal in the farming of a number of fish species such as various types of salmonid, cod, sea bass and sea bream.
The dominant protein source in dry feed for fish has been fish meal of different qualities. Other animal protein sources are also used for dry fish feed. Thus, it is known to use blood meal, bone meal, feather meal and other types of meal produced from other slaughterhouse waste, for example chicken meal. These are typically cheaper than fish meal and fish oil. However, in some geographic
Additional oil, water and steam may be added to the mass in the extruder. At the end of the extruder, the mass has a temperature above 100 C and a pressure above ambient pressure. The mass is forced through the openings in the extruder's die plate. Due to the relief in temperature and pressure, some of the moisture will evaporate immediately (flash off) and the extruded mass becomes porous. The strings are cut into pellets by a rotating knife. The water content is rather high (18-28 %) and the pellets are therefore immediately dried to approximately 10 % water content in a dryer.
After the dryer, more oil may be added to the feed by spraying oil onto the surface of the feed, or by dipping the feed in oil. It is advantageous to add the oil to the feed in a closed vessel where the air pressure is below ambient (vacuum coating) so that the porous feed pellets absorb more oil. Feed containing more than 40 %
lipid may be produced this way. After the coater, the feed is cooled and bagged.
Oil may be added at several places in the process as explained above, and may be fish oil or vegetable oils, by example rape seed oil or soy oil, or a mixture of vegetable oils or a mixture of fish oil and vegetable oils.
Fish need protein, fat, minerals and vitamins in order to grow and to be in good health. The diet of carnivorous fish is particularly important. Originally in the farming of carnivorous fish, whole fish or ground fish were used to meet the nutritional requirements of the farmed fish. Ground fish mixed with dry raw materials of various kinds, such as fish meal and starch, was termed soft or semi-moist feed. As farming became industrialized, soft or semi-moist feed was replaced by pressed dry feed. This was itself gradually replaced by extruded dry feed.
Today, extruded feed is nearly universal in the farming of a number of fish species such as various types of salmonid, cod, sea bass and sea bream.
The dominant protein source in dry feed for fish has been fish meal of different qualities. Other animal protein sources are also used for dry fish feed. Thus, it is known to use blood meal, bone meal, feather meal and other types of meal produced from other slaughterhouse waste, for example chicken meal. These are typically cheaper than fish meal and fish oil. However, in some geographic
- 23 - PCT/EP2016/074407 regions, there has been a prohibition against using such raw materials in the production of feeds for food-producing animals and fish.
It is also known to use vegetable protein such as wheat gluten, maize (corn) gluten, soya protein, lupin meal, pea meal, bean meal, rape meal, sunflower meal and rice flour.
Example 3: Evaluation of the effect of the active compounds according to the invention on survival of Lactococcus garvieae and Streptococcus iniae.
The antimicrobial activity of the composition of the invention towards of Lactococ-cus garvieae and Streptococcus iniae were determined in vitro.
Feeding regime:
1. lysozyme alone for 48h 2. lysozyme for 24 hours and essential oils added on top for the next 24h 3. Lysozyme and essential oils mixed together and added for 48h ¨ Lysozyme: LBF 007 batch 9 PPL 38353 T.reesei at 52 g EP/kg ¨ Essential oil combination at 1/2:
= 50% alpha pinene = 50% cinnamaldehyde In the tests the following organisms, growth media, culture conditions and evalua-tion method were used:
Bacteria: All tested pathogenic strains belong to the strain collection of the Cen-tre for fish and wildlife health, Institute of Animal Pathology, University of Bern (Switzerland).
Determination of suitable bacteria dilution: From a 24 hour old subculture of bacteria on blood sheep agar (Biomerieux, Geneva) a small amount was trans-ferred to sterile NaCI until a McFarland value of 0.5 was obtained. From this solu-tion 3, 1.5 and 0.75 (:)/0 dilutions in TSB were made on 96 well plates with round bottoms. Each well received a total volume of 100 pl. After 24 hours at 22 C
the growth of bacteria was assessed. The dilution resulting in a well demarcated spot covering half of the round bottom well was selected for the experiments.
Determination of solvent effect: To solve test substances in TSB agar, alcohol (ETOH) was used. To determine a possible effect of alcohol, the calculated final concentrations of alcohol in the test wells (0.1, 0.05 and 0.025 %) were tested
It is also known to use vegetable protein such as wheat gluten, maize (corn) gluten, soya protein, lupin meal, pea meal, bean meal, rape meal, sunflower meal and rice flour.
Example 3: Evaluation of the effect of the active compounds according to the invention on survival of Lactococcus garvieae and Streptococcus iniae.
The antimicrobial activity of the composition of the invention towards of Lactococ-cus garvieae and Streptococcus iniae were determined in vitro.
Feeding regime:
1. lysozyme alone for 48h 2. lysozyme for 24 hours and essential oils added on top for the next 24h 3. Lysozyme and essential oils mixed together and added for 48h ¨ Lysozyme: LBF 007 batch 9 PPL 38353 T.reesei at 52 g EP/kg ¨ Essential oil combination at 1/2:
= 50% alpha pinene = 50% cinnamaldehyde In the tests the following organisms, growth media, culture conditions and evalua-tion method were used:
Bacteria: All tested pathogenic strains belong to the strain collection of the Cen-tre for fish and wildlife health, Institute of Animal Pathology, University of Bern (Switzerland).
Determination of suitable bacteria dilution: From a 24 hour old subculture of bacteria on blood sheep agar (Biomerieux, Geneva) a small amount was trans-ferred to sterile NaCI until a McFarland value of 0.5 was obtained. From this solu-tion 3, 1.5 and 0.75 (:)/0 dilutions in TSB were made on 96 well plates with round bottoms. Each well received a total volume of 100 pl. After 24 hours at 22 C
the growth of bacteria was assessed. The dilution resulting in a well demarcated spot covering half of the round bottom well was selected for the experiments.
Determination of solvent effect: To solve test substances in TSB agar, alcohol (ETOH) was used. To determine a possible effect of alcohol, the calculated final concentrations of alcohol in the test wells (0.1, 0.05 and 0.025 %) were tested
- 24 - PCT/EP2016/074407 with different concentrations of bacteria, and did not show any effect on bacterial growth.
Concentration of test substances: The in vitro dose range was estimated con-sidering a probable dietary concentration of at least 1000 ppm and a daily feeding rate of 2 %. The potential concentration in the gut was established at maximum 0.1 p1/100 pl. A serial 2 dilution was then tested leading to final concentrations of the substance of 0.85 pg/ml; 0.42 pg/ml and 0.21 pg/ml when adjusted to average essential oil density.
From each substance a stock solution consisting of 2 pl substance, 18 pl ETOH
and 180 pl PBS was prepared.
Preparation of Plates: Triplicates of three concentrations (0.21, 0.42 and 0.85 pg/ml) of each substance were tested. On each plate a positive control consisting of bacteria in TSB and a blank control (PBS) was included. Further Triplicates of three dilutions of ETOH were also included on each plate.
Reading of plates: After an incubation of 24 hours at 22 C the plates were read using a score of 0 (no bacterial growth = no dot on the bottom of the well) to (normal growth = size of dot comparable to dot of positive control).
The following results were obtained and are summarized in Figures 1 and 2.
Concentration of test substances: The in vitro dose range was estimated con-sidering a probable dietary concentration of at least 1000 ppm and a daily feeding rate of 2 %. The potential concentration in the gut was established at maximum 0.1 p1/100 pl. A serial 2 dilution was then tested leading to final concentrations of the substance of 0.85 pg/ml; 0.42 pg/ml and 0.21 pg/ml when adjusted to average essential oil density.
From each substance a stock solution consisting of 2 pl substance, 18 pl ETOH
and 180 pl PBS was prepared.
Preparation of Plates: Triplicates of three concentrations (0.21, 0.42 and 0.85 pg/ml) of each substance were tested. On each plate a positive control consisting of bacteria in TSB and a blank control (PBS) was included. Further Triplicates of three dilutions of ETOH were also included on each plate.
Reading of plates: After an incubation of 24 hours at 22 C the plates were read using a score of 0 (no bacterial growth = no dot on the bottom of the well) to (normal growth = size of dot comparable to dot of positive control).
The following results were obtained and are summarized in Figures 1 and 2.
Claims (16)
1. Use of at least one natural active substances selected from the group con-sisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with a polypeptide hav-ing lysozyme activity in a feed composition for improving feed conversion ra-tio and/or daily weight gain in aquatic animals.
2. Use of at least two natural active substances selected from the group con-sisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with a polypeptide hav-ing lysozyme activity in a feed composition for regulating the micro flora of the gut in aquatic animals.
3. Use of at least two natural active substances selected from the group con-sisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with a polypeptide hav-ing lysozyme activity in a feed composition for reducing mortality in aquatic animals.
4. Use according to any of claims 1 to 3, wherein the aquatic animal is a cold water fish as for example salmon, trout, bream or bass.
5. Use according to any of claims 1 or 3, wherein the aquatic animal is a warm water fish as for example carp, tilapia or catfish.
6. Use according to any of claims 1 to 5, wherein the feed composition contains alpha-pinene and cinnamaldehyde.
7. Use according to claims 6, wherein the feed composition contains alpha-pinene, cinnamaldehyde, dihydro-eugenol and meta-cresol each in a con-centration between 10 mg and 5 g per Kg of feed.
8. Use according to claim 6, wherein the feed composition contains alpha-pinene, cinnamaldehyde, dihydro-eugenol and meta-cresol each in a con-centration between 0.1 g and 1 g per Kg of feed.
9. Use of any of claims 1 to 8, wherein the microbial lysozyme is of fungal origin.
10. Use of any of claims 1 to 9, wherein the microbial lysozyme is obtained or obtainable from the phylum Ascomycota.
11. Use of any of claims 1 to 10, wherein the microbial lysozyme is obtained or obtainable from the subphylum Pezizomycotina.
12. Use of any of claims 1 to 11, wherein the microbial lysozyme comprises one or more domains selected from the list consisting of GH24 and GH25.
13. Use of any of claims 1 to 12, wherein the microbial lysozyme is selected from the group consisting of:
(a) a polypeptide having at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 1;
(b) a variant of SEQ ID NO: 1 wherein the variant has lysozyme activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;
(c) a fragment of the polypeptide of (a) or (b) that has lysozyme activity wherein the fragment comprises at least 170 amino acids, such as at least 175 amino acids, at least 177 amino acids, at least 180 amino acids, at least 185 amino acids, at least 190 amino acids, at least 195 amino acids or at least 200 amino acids;
(d) a polypeptide having at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 4;
(e) a variant of SEQ ID NO: 4 wherein the variant has lysozyme activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions; and (f) a fragment of the polypeptide of (d) or (e) that has lysozyme activity wherein the fragment comprises at least 210 amino acids, such as at least 215 amino acids, at least 220 amino acids, at least 225 amino acids, at least 230 amino acids, at least 235 amino acids or at least 240 amino acids.
(a) a polypeptide having at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 1;
(b) a variant of SEQ ID NO: 1 wherein the variant has lysozyme activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions;
(c) a fragment of the polypeptide of (a) or (b) that has lysozyme activity wherein the fragment comprises at least 170 amino acids, such as at least 175 amino acids, at least 177 amino acids, at least 180 amino acids, at least 185 amino acids, at least 190 amino acids, at least 195 amino acids or at least 200 amino acids;
(d) a polypeptide having at least 50%, e.g., at least 60%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 4;
(e) a variant of SEQ ID NO: 4 wherein the variant has lysozyme activity and comprises one or more amino acid substitutions, and/or one or more amino acid deletions, and/or one or more amino acid insertions or any combination thereof in 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 ,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 positions; and (f) a fragment of the polypeptide of (d) or (e) that has lysozyme activity wherein the fragment comprises at least 210 amino acids, such as at least 215 amino acids, at least 220 amino acids, at least 225 amino acids, at least 230 amino acids, at least 235 amino acids or at least 240 amino acids.
14. Use of any of claims 1 to 13, wherein the microbial lysozyme is selected from the group consisting of amino acids 1 to 208 of SEQ ID NO: 1 and amino acids 1 to 245 of SEQ ID NO: 4.
15. Use of at least two natural active substances selected from the group con-sisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with a polypeptide hav-ing lysozyme activity according to any of claims 9 - 14 for treatment and prevention of diseases caused by pathogenic microorganisms in aquatic an-imals.
16. A feed composition or a premix composition, or a feed additive for aquatic animals thereof, comprising at least two natural active substances selected from the group consisting of alpha-pinene, alpha-terpineol, cinnamaldehyde, dihydroeugenol, eugenol, meta-cresol and terpinolene in combination with a polypeptide having lysozyme activity according to any of claims 9 ¨ 14 as main ingredients.
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP15189495 | 2015-10-13 | ||
EP15189495.3 | 2015-10-13 | ||
PCT/EP2016/074407 WO2017064092A1 (en) | 2015-10-13 | 2016-10-12 | Feed additives for aquatic animals comprising essential oils and lysozyme |
Publications (1)
Publication Number | Publication Date |
---|---|
CA3001389A1 true CA3001389A1 (en) | 2017-04-20 |
Family
ID=54293139
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CA3001389A Abandoned CA3001389A1 (en) | 2015-10-13 | 2016-10-12 | Feed additives for aquatic animals comprising essential oils and lysozyme |
Country Status (5)
Country | Link |
---|---|
EP (1) | EP3361882A1 (en) |
CN (1) | CN108289478A (en) |
CA (1) | CA3001389A1 (en) |
CL (1) | CL2018000925A1 (en) |
WO (1) | WO2017064092A1 (en) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109105665A (en) * | 2018-09-05 | 2019-01-01 | 山东大学 | A kind of protectant method of screening turbot feed |
Families Citing this family (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US10945449B2 (en) | 2015-07-02 | 2021-03-16 | Novozymes A/S | Animal feed compositions and uses thereof |
JP7275048B2 (en) * | 2017-05-12 | 2023-05-17 | ノボザイムス アクティーゼルスカブ | Polypeptide having lysozyme activity, polynucleotide encoding same, use and composition thereof |
WO2019121930A1 (en) * | 2017-12-20 | 2019-06-27 | Dsm Ip Assets B.V. | Animal feed compositions and uses thereof |
MX2020006588A (en) * | 2017-12-20 | 2020-12-10 | Dsm Ip Assets Bv | Animal feed compositions comprising muramidase and uses thereof. |
BR112020021365A2 (en) * | 2018-04-25 | 2021-01-19 | Novozymes A/S | ANIMAL FEED, METHOD FOR IMPROVING THE EUROPEAN PRODUCTION EFFICIENCY FACTOR, BODY WEIGHT GAIN AND / OR RATION CONVERSION REASON FOR A MONOGRAPHIC ANIMAL, USE OF AT LEAST ONE PROBIOTIC IN COMBINATION WITH A POLYPEPTIOUS ACTIVITY OF BEING ACTIVELY ACTIVE IN HIS ACTIVITY. FEED, AND, FEED COMPOSITION OR PRE-MIXTURE COMPOSITION, OR ANIMAL FEED ADDITIVE. |
BR112021004814A2 (en) * | 2018-09-17 | 2021-06-22 | Dsm Ip Assets B.V. | composition of animal feed and its use |
BR112021004812A2 (en) * | 2018-09-17 | 2021-06-08 | Dsm Ip Assets B.V. | animal feed compositions and their uses |
CN109170369A (en) * | 2018-10-10 | 2019-01-11 | 徐州市宏翔蛋白饲料有限公司 | A kind of formula of the bone powder fodder containing duck |
Family Cites Families (15)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US7504490B1 (en) * | 1998-10-16 | 2009-03-17 | Oscient Pharmaceuticals Corporation | Nucleic acid and amino acid sequences relating to Apergillus fumigatus for diagnostics and therapeutics |
JP2002316909A (en) * | 2000-12-22 | 2002-10-31 | Erubu:Kk | Functional material |
DE102004019751A1 (en) * | 2004-04-23 | 2005-11-17 | Henkel Kgaa | Novel Alkaline Proteases and Detergents Containing These Novel Alkaline Proteases |
US7354888B2 (en) * | 2004-11-10 | 2008-04-08 | Danisco A/S | Antibacterial composition and methods thereof comprising a ternary builder mixture |
US8088976B2 (en) * | 2005-02-24 | 2012-01-03 | Monsanto Technology Llc | Methods for genetic control of plant pest infestation and compositions thereof |
ES2523667T3 (en) * | 2007-06-29 | 2014-11-28 | Dsm Ip Assets B.V. | Use of benzoic acid and thymol, eugenol and piperidine in animal feed |
HU229104B1 (en) * | 2007-11-22 | 2013-07-29 | Pharmateka Bt | Use of edta and its salts for the prevention and treatment of bacterial intestinal swine diseases caused by brachyspira hyodysenteriae and for enhancement of efficiency of antibiotics in curing such illnesses |
CN101773513A (en) * | 2009-01-14 | 2010-07-14 | 邢建慧 | Water soluble body lubricant with natural antibacterial components and with pH value consistence with pH value of vagina |
JP5881175B2 (en) * | 2009-07-17 | 2016-03-09 | ディーエスエム アイピー アセッツ ビー.ブイ. | Use of natural substances as feed additives for aquatic animals |
US20130101634A1 (en) * | 2011-10-12 | 2013-04-25 | Deerland Enzymes, Inc. | Compositions and methods for reducing microbial overgrowth in the small intestines |
US9663775B2 (en) * | 2011-11-25 | 2017-05-30 | Novozymes A/S | Polypeptides having lysozyme activity and polynucleotides encoding same |
CN103571613A (en) * | 2012-08-06 | 2014-02-12 | 湖北大自然农业实业有限公司 | Preparation method of vitex negundo linn volatile oil |
CN104673558A (en) * | 2013-11-30 | 2015-06-03 | 楚乐然 | Acarus killing perfumed soap |
CN104256194B (en) * | 2014-10-14 | 2016-07-06 | 湖南百宜饲料科技有限公司 | A kind of functional type is saved one's life pig milk powder creep feed |
CN104286420A (en) * | 2014-10-27 | 2015-01-21 | 天津市世昌科技发展有限公司 | Feed additive capable of improving farrowing and milk production performances of breeding fur-bearing animals and preparation method of feed additive |
-
2016
- 2016-10-12 CA CA3001389A patent/CA3001389A1/en not_active Abandoned
- 2016-10-12 EP EP16784817.5A patent/EP3361882A1/en not_active Withdrawn
- 2016-10-12 WO PCT/EP2016/074407 patent/WO2017064092A1/en active Application Filing
- 2016-10-12 CN CN201680058693.7A patent/CN108289478A/en active Pending
-
2018
- 2018-04-11 CL CL2018000925A patent/CL2018000925A1/en unknown
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109105665A (en) * | 2018-09-05 | 2019-01-01 | 山东大学 | A kind of protectant method of screening turbot feed |
Also Published As
Publication number | Publication date |
---|---|
CL2018000925A1 (en) | 2018-08-03 |
CN108289478A (en) | 2018-07-17 |
EP3361882A1 (en) | 2018-08-22 |
WO2017064092A1 (en) | 2017-04-20 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CA3001389A1 (en) | Feed additives for aquatic animals comprising essential oils and lysozyme | |
AU2016288468B2 (en) | Animal feed compositions and uses thereof | |
AU2016288515B2 (en) | Methods of improving animal performance | |
CA2767873C (en) | Use of natural substances as feed additives for aquatic animals | |
WO2016060934A1 (en) | Bacillus strains with fast germination and antimicrobial activity against clostridium perfringens | |
WO2019207053A1 (en) | Animal feed compositions and uses thereof | |
WO2017147130A2 (en) | Direct fed microbial for prevention of shrimp disease | |
BR112020012498A2 (en) | animal feed compositions and uses thereof | |
BR112021004519A2 (en) | composition of animal feed and its use | |
BR112021004826A2 (en) | animal feed compositions and their uses | |
BR112021004812A2 (en) | animal feed compositions and their uses | |
BR112021004501A2 (en) | composition of animal feed and its use | |
KR100857771B1 (en) | Compositions for addition to feed for fish comprising Bacillus polyfermenticus, Bacillus licheniformis and Saccharomyces serevisiae | |
BR112021004464A2 (en) | composition of animal feed and its use | |
KR20210057773A (en) | Animal feed composition and use thereof | |
BR112021004817A2 (en) | animal feed compositions and their uses | |
BR112019018381A2 (en) | fungal fucosidases and their use in the prevention and / or treatment of a pathogenic infection in an animal | |
US20240122209A1 (en) | Animal feed compositions and uses thereof | |
EP4045642A1 (en) | Animal feed compositions and uses thereof | |
CN116887689A (en) | Method for improving the digestibility of carbohydrases in animal feed by using serine proteases |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
FZDE | Discontinued |
Effective date: 20230104 |
|
FZDE | Discontinued |
Effective date: 20230104 |