Dental care composition for animals

Abstract

The present invention relates to a composition comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase; the use of the composition as a pharmaceutical agent; the use of the composition in the treatment of oral diseases; a treatment method comprising administering the composition to an animal; a method for preventing or removing oral biofilms, comprising contacting the oral biofilm with the composition; a method for reducing the risk of oral biofilm formation; a method for removing formed oral biofilms; and a kit of parts comprising the composition.

Description

Technical Field

[0001] Reference to Sequence Listing The present application contains a Sequence Listing in computer-readable form. Said computer-readable form is incorporated herein by reference.

[0002] The present invention relates to a composition comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase, and lipase and / or phosphodiesterase, use of the composition as a medicament for animals, use of the composition in the treatment of dental care for animals, a treatment method comprising administering the composition to an animal, a method for preventing or removing oral biofilm comprising contacting oral biofilm with the composition, a method for reducing the risk of oral biofilm formation, and a kit of parts comprising the composition.

Background Art

[0003] Many animal owners focus on dental care for their animals, especially companion animals such as dogs and cats, to avoid dental diseases that cause undesirable dental caries, bleeding from gums, and even tooth loss. Furthermore, when bacteria residing on teeth enter the body of an animal, they can adversely affect the heart, lungs, kidneys and other organs of the animal and cause other undesirable diseases. It is estimated that over 80 percent of all dogs have severe oral problems. When a veterinarian examines a dog for vaccination, the examination often ends with an investigation of the dog's general health. Dental hygiene is the most common cause requiring immediate action. In order to solve such problems, dental care products dedicated to pet animals, especially pet dogs, have been used in recent years to prevent dental diseases in animals. However, there is a need for dental care products that, along with prevention, remove any bacterial biofilm on the teeth of animals to avoid dental disease or reduce the severity of dental disease.

Summary of the Invention

Problem to be Solved by the Invention

[0004] Therefore, there is a need for improved dental care products for animals to prevent and mitigate dental diseases in animals. [Means for solving the problem]

[0005] The present invention provides a composition comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase, which is useful for preventing and / or removing dental biofilm formation.

[0006] In a first aspect, the present invention relates to a composition for animal dental care, comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase.

[0007] In a second aspect, the present invention relates to a dental care composition comprising a combination of enzymes and at least one dental care component. This combination of enzymes includes alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase.

[0008] In a third aspect, the present invention relates to a composition according to the first or second aspect for use as a pharmaceutical agent.

[0009] In a fourth aspect, the present invention relates to a method for preventing or removing an oral biofilm in an animal, comprising contacting the oral biofilm with a composition according to the first or second aspect.

[0010] In a fifth aspect, the present invention relates to a method for reducing the risk of oral biofilm formation in animals, comprising contacting the oral biofilm with a composition according to the first or second aspect.

[0011] In a sixth aspect, the present invention relates to a method for preventing or removing plaque, reducing the likelihood of caries, or reducing gingivitis in animals, comprising bringing a composition according to the first or second aspect into contact with the teeth of the animal. Related aspects relate to the use of the composition or dental care composition of the present invention for preventing or removing plaque, reducing plaque formation, reducing the likelihood of caries, or reducing gingivitis in animals.

[0012] A further aspect of the present invention is a dental care composition comprising a combination of enzymes and at least one dental care component, wherein the combination of enzymes is a. One or more polypeptides having alpha-amylase activity, wherein one or more polypeptides have at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2. b. One or more polypeptides having glucanase activity, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4. ii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6. iii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8. iv. Polypeptides having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10, and v. Polypeptides having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12. One or more polypeptides selected from the group consisting of, c. One or more polypeptides having xylanase activity, wherein one or more polypeptides have at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16. d. One or more polypeptides having lipase activity, wherein one or more polypeptides have at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18. This applies to dental care compositions that include [the specified ingredient / method].

[0013] Further aspects of the present invention relate to compositions of the present invention for use in preventing or removing biofilm formation in animal dentition. Related aspects of the present invention relate to compositions of the present invention for use in preventing or removing plaque formation in animals. One aspect of the present invention relates to a method for removing oral biofilm formed in an animal, comprising contacting the oral biofilm with a composition of the present invention or a dental care composition. One aspect of the present invention relates to a method for removing oral biofilm formed in an animal, comprising contacting the oral biofilm with a composition of the present invention or a dental care composition.

[0014] Array Overview Sequence ID 1 is the amino acid sequence (including the signal peptide) of GH13_5 alpha-amylase obtained from Bacillus amyloliquefaciens.

[0015] Sequence ID 2 is the amino acid sequence (mature polypeptide) of GH13_5 alpha-amylase obtained from Bacillus amyloliquefaciens.

[0016] SEQ ID NO: 3 is the amino acid sequence (including the signal peptide) of GH5 beta-glucanase obtained from Trichoderma reesei.

[0017] SEQ ID NO: 4 is the amino acid sequence (mature polypeptide) of GH5 beta-glucanase obtained from Trichoderma reesei.

[0018] SEQ ID NO: 5 is the amino acid sequence (including the signal peptide) of GH6 glucanase obtained from Trichoderma reesei.

[0019] SEQ ID NO: 6 is the amino acid sequence (mature polypeptide) of GH6 glucanase obtained from Trichoderma reesei.

[0020] SEQ ID NO: 7 is the amino acid sequence (including the signal peptide) of GH7 endo-glucanase obtained from Trichoderma reesei.

[0021] SEQ ID NO: 8 is the amino acid sequence (mature polypeptide) of GH7 endo-glucanase obtained from Trichoderma reesei.

[0022] SEQ ID NO: 9 is the amino acid sequence (including the signal peptide) of GH7 exo-glucanase obtained from Trichoderma reesei.

[0023] SEQ ID NO: 10 is the amino acid sequence (mature polypeptide) of GH7 exo-glucanase obtained from Trichoderma reesei.

[0024] SEQ ID NO: 11 is the amino acid sequence (including the signal peptide) of GH7 beta-glucanase obtained from Humicola insolens.

[0025] SEQ ID NO: 12 is the amino acid sequence (mature polypeptide) of GH7 beta-glucanase obtained from Humicola insolens.

[0026] SEQ ID NO: 13 is the amino acid sequence (including the signal peptide) of GH16 beta-glucanase obtained from Bacillus amyloliquefaciens.

[0027] SEQ ID NO: 14 is the amino acid sequence (mature polypeptide) of GH16 beta-glucanase obtained from Bacillus amyloliquefaciens.

[0028] SEQ ID NO: 15 is the amino acid sequence (including the signal peptide) of GH10 xylanase obtained from Humicola insolens.

[0029] SEQ ID NO: 16 is the amino acid sequence (mature polypeptide) of GH10 xylanase obtained from Humicola insolens.

[0030] SEQ ID NO: 17 is the amino acid sequence (including the signal peptide) of GH10 xylanase obtained from Aspergillus aculeatus.

[0031] SEQ ID NO: 18 is the amino acid sequence (mature polypeptide) of GH10 xylanase obtained from Aspergillus aculeatus.

[0032] Sequence ID 19 is the amino acid sequence (including the signal peptide) of lipase obtained from Thermomyces lanuginosus.

[0033] Sequence ID 20 is the amino acid sequence (mature polypeptide) of lipase obtained from Thermomyces lanuginosus.

[0034] Sequence ID 21 is the amino acid sequence (including the signal peptide) of a phosphodiesterase obtained from Metabacillus indicus.

[0035] Sequence ID 22 is the amino acid sequence (mature polypeptide) of phosphodiesterase obtained from Metabacillus indicus.

[0036] Sequence ID 23 is the nucleic acid sequence of GH13_5 alpha-amylase isolated from Bacillus amyloliquefaciens.

[0037] Sequence ID 24 is the nucleic acid sequence of GH5 beta-glucanase obtained from Trichoderma reesei.

[0038] Sequence ID 25 is the nucleic acid sequence of GH6 glucanase obtained from Trichoderma reesei.

[0039] Sequence ID 26 is the nucleic acid sequence of GH7 endoglucanase obtained from Trichoderma reesei.

[0040] Sequence ID 27 is the nucleic acid sequence of GH7 exoglucanase obtained from Trichoderma reesei.

[0041] Sequence ID 28 is the nucleic acid sequence of GH16 beta-glucanase obtained from Bacillus amyloliquefaciens.

[0042] Sequence ID 29 is the nucleic acid sequence of GH10 xylanase obtained from Humicola insolens.

[0043] Sequence ID 30 is the nucleic acid sequence of GH10 xylanase obtained from Aspergillus aculeatus.

[0044] Sequence ID 31 is the nucleic acid sequence of lipase obtained from Thermomyces lanuginosus.

[0045] Sequence ID 32 is the nucleic acid sequence of phosphodiesterase obtained from Metabacillus indicus. [Modes for carrying out the invention]

[0046] The present invention relates to a composition for animal dental care, comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase. As shown in the examples of this application, the inventors have identified alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase that are effective in preventing and / or reducing the risk of oral biofilm formation in animals. The inventors have further demonstrated that the combination of alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase is capable of potently removing biofilm already present in the oral cavity of animals.

[0047] Therefore, in the first aspect, the present invention relates to a dental care composition comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase, and at least one dental care component.

[0048] definition In accordance with this detailed explanation, the following definitions apply. Note that the singular forms "a," "an," and "that" refer to multiple objects unless otherwise clearly indicated by the context.

[0049] Unless otherwise defined or clearly indicated by the context, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art to which this invention pertains.

[0050] Alpha-amylase: The term "alpha-amylase" refers to a polypeptide having alpha-amylase activity that catalyzes the internal hydrolysis of (1→4)-α-D-glucosidic bonds in polysaccharides containing three or more (1→4)-α-linked D-glucose units. The systematic name for alpha-amylase is 4-α-D-glucan-glucanohydrolase (EC 3.2.1.1). The terms "alpha-amylase" and "α-amylase," as well as the expression "polypeptide having alpha-amylase activity," are used synonymously throughout this application. For the purposes of the present invention, alpha-amylase activity can be measured according to alpha-amylase activity assay I or alpha-amylase activity assay II, as described in Example 2 below.

[0051] Glucanase: The term "glucanase" refers to any enzyme capable of hydrolyzing glycosidic bonds.

[0052] Xylanase: The term "xylanase" refers to a xylanase enzyme, as defined according to those known in the art as endo-1,4-0-xylanase or 4-0-D-xylanxylanohydrolase, which is known to catalyze the internal hydrolysis of the (1→4)-0-D-xyloside bond in xylene.

[0053] Lipase: The term "lipase" refers to a subset of carboxylic acid ester hydrolases that catalyze the hydrolysis of ester bonds in lipids such as triglycerides, phospholipids, and galactolipids.

[0054] Phosphodiesterase: The term "phosphodiesterase (PDE)" refers to a type of phosphatase that breaks down phosphodiester bonds, such as cyclic adenosine monophosphate (cAMP), cyclic guanosine monophosphate (cGMP), and similar compounds. These enzymes catalyze reactions that produce phosphomonoesters and alcohols through hydrolysis, and are classified into a total of 12 subtypes. The term "phosphodiesterase," used synonymously with "PDE," refers to members of the cyclic nucleotide phosphodiesterase family.

[0055] Animals: The term "animals" means organisms that make up the biological animal kingdom, such as domesticated animals, such as dogs, cats, cows, horses, and pigs, as well as undomesticated animals, such as deer, moose, alpacas, llamas, and moose; carnivores, such as bears, wolves, lions, and tigers; rodents, such as rats, mice, and beavers; bats; marsupials, such as kangaroos and opossums; and cetaceans, such as whales, dolphins, and porpoises.

[0056] Biofilm: In this context, biofilm formation is the process by which microorganisms adhere to and proliferate on the surface of the oral cavity, producing extracellular polymers that promote adhesion and matrix formation. Biofilm formation leads to plaque formation on teeth. Oral biofilms vary depending on the animal species and their microbial composition differs.

[0057] Biofilm prevention: The term "biofilm prevention" refers to the ability of a polypeptide to reduce the amount of biofilm that grows under defined conditions. For the purposes of the present invention, biofilm prevention can be determined according to Example 1 of this specification.

[0058] Fragments: The term "fragment" refers to a polypeptide having one or more amino acids that are not present at the amino group and / or carboxyl terminus of a mature polypeptide, and this fragment has the enzymatic activity of the mature polypeptide. A glucanase fragment has glucanase activity, a xylanase fragment has xylanase activity, a lipase fragment has lipase activity, a phosphodiesterase fragment has phosphodiesterase activity, and an alpha-amylase fragment has alpha-amylase activity.

[0059] Mature polypeptide: The term "mature polypeptide" refers to the mature form of a polypeptide after N-terminal and / or C-terminal processing (e.g., removal of signal peptides).

[0060] Sequence identity: The relationship between two amino acid sequences or two nucleotide sequences is described by a parameter called "sequence identity."

[0061] For the purposes of this invention, sequence identity between two amino acid sequences is determined as the “longest identity” output using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, J.Mol.Biol.48:443-453), preferably implemented in the Needle program of the EMBOSS package (EMBOSS: The European Molecular Biology Open Software Suite, Rice et al., 2000, Trends Genet. 16:276-277), version 6.6.0 or later. The parameters used are a gap-open penalty of 10, a gap-extension penalty of 0.5, and an EBLOSUM62 (EMBOSS version of BLOSUM62) substitution matrix. The -no brief option must be specified on the command line for the Needle program to report the longest identity. The Needle output labeled “longest identity” is calculated as follows: (Identical residues × 100) / (Length of alignment - Total number of gaps in the alignment).

[0062] For the purposes of this invention, sequence identity between two polynucleotide sequences is determined as the “longest identity” output using the Needleman-Wunsch algorithm (Needleman and Wunsch, 1970, cited above), preferably version 6.6.0 or later, implemented in the Needle program of the EMBOSS package (EMBOSS: The European Molecular Biology Open Software Suite, Rice et al., 2000, cited above). The parameters used are a gap-open penalty of 10, a gap-extension penalty of 0.5, and an EDNAFULL (EMBOSS version of NCBI NUC4.4) substitution matrix. The no brief option must be specified on the command line for the Needle program to report the longest identity. The Needle output labeled “longest identity” is calculated as follows: (100 identical deoxyribonucleotides) / (length of alignment - total number of gaps in the alignment).

[0063] Variant: The term "variant" refers to an alpha-amylase, glucanase, xylanase, lipase, or phosphodiesterase that contains artificial mutations (i.e., substitutions, insertions (including extensions), and / or deletions (e.g., truncations)) at one or more locations. Substitution means replacing an amino acid occupying a certain position with a different amino acid, deletion means removing an amino acid occupying a certain position, and insertion means adding 1 to 5 amino acids (e.g., 1 to 3 amino acids, 1 amino acid) adjacent to and immediately following an amino acid occupying a certain position.

[0064] alpha-amylase Alpha-amylase can be obtained from any organism. Preferably, the alpha-amylase is of microbial origin, and most preferably, it is of bacterial or fungal origin. In a preferred embodiment, the alpha-amylase is obtained from Bacillus amyloliquefaciens. The alpha-amylase is GH13 alpha-amylase. In a preferred embodiment, the alpha-amylase is GH13_5 alpha-amylase.

[0065] In one embodiment, alpha-amylase is (a) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2. (b) A polypeptide encoded by a polynucleotide having at least 80%, for example, at least 85%, 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 with respect to the mature polypeptide coding sequence of SEQ ID NO: 1 or its cDNA sequence. (c) A polypeptide derived from SEQ ID NO: 1, which is a mature polypeptide of SEQ ID NO: 1 obtained by substitution, deletion or addition of one or more amino acids. (d) Polypeptides derived from the polypeptides of (a), (b), and (c) (the N-terminus and / or C-terminus are extended by the addition of one or more amino acids), and (e) A polypeptide fragment of (a), (b), or (c) (this polypeptide has alpha-amylase activity) It is selected from the group consisting of the following.

[0066] In a preferred embodiment, the alpha-amylase comprises, is essentially, or consists of the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2.

[0067] glucanase Glucanase can be obtained from any organism. Preferably, the glucanase is of microbial origin, and most preferably, it is of bacterial or fungal origin. In a preferred embodiment, the glucanase is obtained from Trichoderma reesei. The glucanase is selected from the group consisting of GH5 beta-glucanase, GH6 glucanase, GH7 endo-glucanase, GH7 exo-glucanase, and GH16 beta-glucanase.

[0068] In one embodiment, glucanase, (a) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4. (b) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6. (c) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8. (d) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10. (e) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12. (f) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 13 or the polypeptide of SEQ ID NO: 14. (g) Polypeptides encoded by polynucleotides having at least 80%, for example, at least 85%, 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 with respect to the mature polypeptide coding sequence of SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, or SEQ ID NO: 13, or its cDNA sequence, (h) A mature polypeptide of SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11 or SEQ ID NO: 13, obtained by substitution, deletion or addition of one or more amino acids, a polypeptide derived from SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9 or SEQ ID NO: 11 (i) Polypeptides derived from polypeptides of (a), (b), (c), (d), (e), (f), (g), or (h) (the N-terminus and / or C-terminus are extended by the addition of one or more amino acids), and (g) A polypeptide fragment of (a), (b), (c), (d), (e), (f), (g), or (h) (this polypeptide has glucanase activity) It is selected from the group consisting of the following.

[0069] In a preferred embodiment, the glucanase comprises, is essentially, or consists of the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4.

[0070] In a preferred embodiment, the glucanase comprises, essentially comprises, or consists of the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6.

[0071] In a preferred embodiment, the glucanase comprises, is essentially, or consists of the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8.

[0072] In a preferred embodiment, the glucanase comprises, is essentially, or consists of the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10.

[0073] In a preferred embodiment, the glucanase comprises, essentially comprises, or consists of the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12.

[0074] In a preferred embodiment, the glucanase comprises, essentially comprises, or consists of the mature polypeptide of SEQ ID NO: 13 or the polypeptide of SEQ ID NO: 14.

[0075] Xylanase Xylanase can be obtained from any organism. Preferably, the xylanase is of microbial origin, and most preferably, the glucanase is of bacterial or fungal origin. In a preferred embodiment, the xylanase is obtained from Humicola insolens. The xylanase is GH10 xylanase.

[0076] In one embodiment, xylanase is (a) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16. (b) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18. (c) A polypeptide encoded by a polynucleotide having at least 80%, for example, at least 85%, 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 with respect to the mature polypeptide coding sequence of SEQ ID NO: 15 or SEQ ID NO: 17 or its cDNA sequence. (d) A mature polypeptide of SEQ ID NO: 13 or SEQ ID NO: 15, obtained by substitution, deletion or addition of one or more amino acids, derived from SEQ ID NO: 15 or SEQ ID NO: 17 (e) Polypeptides derived from polypeptides of (a), (b), (c), or (d) (the N-terminus and / or C-terminus are extended by the addition of one or more amino acids), and (f) A fragment of the polypeptide of (a), (b), (c), or (d) (this polypeptide has xylanase activity) It is selected from the group consisting of the following.

[0077] In a preferred embodiment, the glucanase comprises, essentially comprises, or consists of the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16.

[0078] In a preferred embodiment, the glucanase comprises, essentially comprises, or consists of the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18.

[0079] Lipase Lipase can be obtained from any organism. Preferably, the lipase is of microbial origin, and most preferably, it is of bacterial or fungal origin. In a preferred embodiment, the lipase is obtained from Thermomyces lanuginosus.

[0080] In one embodiment, lipase, (f) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 19 or the polypeptide of SEQ ID NO: 20. (g) A polypeptide encoded by a polynucleotide having at least 80%, for example, at least 85%, 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 with respect to the mature polypeptide coding sequence of SEQ ID NO: 19 or its cDNA sequence. (h) A mature polypeptide of SEQ ID NO: 19, which is a polypeptide derived from SEQ ID NO: 19, which is a mature polypeptide of SEQ ID NO: 19 obtained by substitution, deletion or addition of one or more amino acids. (i) Polypeptides derived from the polypeptides of (a), (b), and (c) (the N-terminus and / or C-terminus are extended by the addition of one or more amino acids), and (j) A fragment of the polypeptide of (a), (b), or (c) (this polypeptide has lipase activity) It is selected from the group consisting of the following.

[0081] In a preferred embodiment, the alpha-amylase comprises, essentially comprises, or consists of the mature polypeptide of SEQ ID NO: 19 or the polypeptide of SEQ ID NO: 20.

[0082] Phosphodiesterase Phosphodiesterases can be obtained from any organism. Preferably, the phosphodiesterase is of microbial origin, and most preferably, it is of bacterial or fungal origin. In a preferred embodiment, the phosphodiesterase is obtained from Metabacillus indicus.

[0083] In one embodiment, phosphodiesterase is (a) A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 21 or the polypeptide of SEQ ID NO: 22. (b) A polypeptide encoded by a polynucleotide having at least 80%, for example, at least 85%, 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 with respect to the mature polypeptide coding sequence of SEQ ID NO: 21 or its cDNA sequence. (c) A mature polypeptide of SEQ ID NO: 21, which is a polypeptide derived from SEQ ID NO: 21, which is a mature polypeptide of SEQ ID NO: 21 obtained by substitution, deletion or addition of one or more amino acids. (d) Polypeptides derived from the polypeptides of (a), (b), and (c) (the N-terminus and / or C-terminus are extended by the addition of one or more amino acids), and (e) A fragment of the polypeptide of (a), (b), or (c) (this polypeptide has lipase activity) It is selected from the group consisting of the following.

[0084] In a preferred embodiment, the alpha-amylase has at least 80%, for example, at least 85%, 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 with respect to the mature polypeptide of SEQ ID NO: 21 or the polypeptide of SEQ ID NO: 22.

[0085] Dental care ingredients The dental care composition of the present invention comprises a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase, and at least one dental care component.

[0086] Dental care ingredients for animals may be modified according to various types of dental care ingredient compositions and the desired properties and / or activity of such dental care ingredient compositions. For the purposes of this invention, the terms “ingredient” and “constituent” are used synonymously with respect to dental care ingredient compositions.

[0087] The dental care composition of the present invention may be a composition such as toothpaste, toothpaste gel, toothpaste cream, toothpaste powder, toothpaste agent, oral spray, chewable product, oral freshener, mouthwash, mouth rinse, dental pack, and the like, and this composition is designed to remove oral biofilm (for example, biofilm present on teeth, oral soft tissues, and dentures present in the oral cavity).

[0088] In preferred embodiments, the dental care composition is at least one dental care component selected from the group consisting of abrasives, wetting agents, solvents, thickeners, binders, buffers, foaming agents, foam regulators, sweeteners, softeners, plasticizers, flavoring agents, colorants, therapeutic agents, antibacterial agents, tartar inhibitors, fluoride sources, preservatives, detergents, surfactants, colorants, buffers, softeners, plasticizers, whitening agents, bleaching agents, gum base components, and fillers.

[0089] This dental care composition may further contain various colorants and flavoring additives, binders, emulsifiers, preservatives and antioxidants, and may be formulated as an aqueous solution, non-aqueous solution, suspension, gel, foam, paste, powder, fine powder, solid or emulsion.

[0090] This dental care composition can also be applied to various types of animal feed and edible treats.

[0091] This dental care composition may further contain additives, including but not limited to: sorbitol, dextrin, maltodextrin, collagen, glycerin, hexose, glucosamine, and mixtures thereof.

[0092] This dental care composition may further contain natural polymers, which include, but are not limited to, the following: starches, such as potato starch, corn starch, wheat starch, tapioca starch, and pea starch; pectin, gum arabic, acacia gum, guar gum, xanthan gum; proteins, such as egg white, pea protein, and gluten; and oils, such as coconut oil, rapeseed oil, or corn oil.

[0093] This dental care composition may further contain sugar, edible toothpaste, nutritional supplements, water-soluble thickeners, or a combination thereof.

[0094] The following are possible but not limited to sugars: agave nectar, barley malt syrup, barley sugar, kaba xylose, brown sugar, caramel, coconut sugar, corn syrup, date sugar, dextrose, sucrose, lactose, maltose, fructose, galactose, glucose, golden syrup, high-fructose corn syrup, high-maltose corn syrup, honey, confectionery sugar, inositol, invert sugar syrup, jaggery, mannose, maple sugar, maple syrup, sugar syrup (derived from sugar beets), sugar syrup (derived from sugarcane), palm sugar, ribose, rhamnose, toffee, molasses, trehalose, xylose, and mixtures thereof.

[0095] Examples of edible toothpastes include, but are not limited to, dental-grade silica abrasives, dicalcium phosphate dihydrate, pumice, polyphosphates, microcrystalline cellulose, magnesium stearate, and mixtures thereof.

[0096] Nutritional supplements may include, but are not limited to, brewer's yeast, Echinacea purpurea, Ascophyllum nodosum, flaxseed, polyunsaturated fatty acids, calcium, vitamins and minerals, and combinations thereof.

[0097] Examples of water-soluble thickeners include, but are not limited to, gelatin, carboxymethylcellulose, agar, starch, carrageenan, and mixtures thereof.

[0098] This dental care composition may further contain coloring additives, flavoring additives (parathant enhancers), binders, emulsifiers, preservatives, antioxidants, or combinations thereof.

[0099] This dental care composition can be formulated as an aqueous solution, a non-aqueous solution, a suspension, a gel, a foam, a paste, a powder, a fine powder, a solid, or an emulsion.

[0100] The dental care ingredients referred to herein are classified by general headings according to their functionality, but this should not be construed as limiting, as ingredients may include additional functionality as will be understood by those skilled in the art.

[0101] Uses of dental care compositions The dental care composition of the present invention is suitable for use in the treatment of oral diseases in animals where prevention or removal of oral biofilm is desired. The composition of the present invention is particularly suitable for the prevention or removal of plaque, reduction of the possibility of caries, or reduction of gingivitis in animals.

[0102] This biofilm is typically a biofilm in the oral cavity of a companion animal such as a dog, cat, horse, or pig, more typically a dog, cat, or horse, and even more typically a dog or cat. In a typical embodiment, this animal is a companion animal such as a dog, cat, horse, or pig, more typically a dog, cat, or horse, and even more typically a dog or cat.

[0103] In a preferred embodiment, the dental care composition is administered into the oral cavity of an animal. In a more typical embodiment, the dental care composition is applied to the teeth or dentition of an animal. The composition may be applied by any various means, such as using a toothbrush, or by brushing it onto the teeth using another instrument or finger, or as a formulation that adheres to the teeth (adhesive formulation) and temporarily adheres to the dentition, such as a fluid formulation (e.g., a fluid formulation applied to animal feed), or integrated into a chew toy.

[0104] The dental care composition of the present invention may be added to and incorporated into a chew toy that releases the dental care composition while the dog chews it. In addition, the chew toy has a mechanical effect on established tooth biofilms. The chew toy may then be replenished with gel before being given to the dog. Several different types of chew toys exist, such as rubber, rawhide, wood, paper, and minerals. While chew toys are generally associated with puppies and adult dogs, this chew toy can also be used for cats, horses, pigs, birds, rodents, and rabbits. In one embodiment, the present invention relates to a dental care composition comprising a combination of enzymes, including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase, for use as a pharmaceutical agent, and at least one dental care component.

[0105] In one embodiment, the present invention relates to a dental care composition for use in the treatment of oral diseases, comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase, and at least one dental care component.

[0106] In a preferred embodiment, the present invention relates to a dental care composition comprising a combination of enzymes including alpha-amylase, glucanase and / or xylanase and lipase and / or phosphodiesterase, and at least one dental care component, for use in preventing or removing plaque, reducing the likelihood of caries or reducing gingivitis in animals.

[0107] As is known to those skilled in the art, the concentration of each enzyme is determined by its enzymatic activity. The compositions of the present invention contain enzymes at concentrations suitable for reducing the rate of biofilm formation in animals, or at concentrations suitable for removing biofilm or plaque that develops after repeated application. Furthermore, the enzyme concentration is determined by the method of application of the composition to animals, in particular, since the frequency of application of the composition may differ between methods of application, various concentrations may be required when the method of application is toothpaste, compared to the concentration when the method of application is by means of chewing. In typical embodiments, each enzyme class is concentrated at concentrations of 1 to 100 ppm, for example, 5 to 80 ppm, and more typically 10 to 60 ppm.

[0108] Preferred Embodiment item: 1. A composition for animal dental care, comprising a combination of enzymes, the combination of enzymes is i. One or more types of alpha-amylase, ii. One or more glucanases and / or one or more xylanases, iii. One or more lipases and / or one or more phosphodiesterases A composition containing the following:

[0109] 2. The composition according to item 1, further comprising at least one dental care ingredient.

[0110] 3. The composition according to paragraph 1, wherein the alpha-amylase is GH13 alpha-amylase.

[0111] 4. The composition according to item 1 or 3, wherein the alpha-amylase has at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2.

[0112] 5. The composition according to item 4, wherein the alpha-amylase comprises, is essentially derived from, or consists of the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2.

[0113] 6. The composition according to item 1, wherein the glucanase is selected from the group consisting of GH5 beta-glucanase, GH6 glucanase, GH7 endoglucanase, GH7 exo-glucanase, GH7 beta-glucanase, and GH16 beta-glucanase.

[0114] 7. Glucanase is, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4. ii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6. iii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8. iv. Polypeptides having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10, and v. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12. vi. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 13 or the polypeptide of SEQ ID NO: 14. A composition according to item 1 or 6, selected from the group consisting of the following.

[0115] 8. The composition according to paragraph 7, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4, the glucanase.

[0116] 9. The composition according to paragraph 7, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6.

[0117] 10. The composition according to item 7, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8.

[0118] 11. The composition according to paragraph 7, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10, the glucanase.

[0119] 12. The composition according to paragraph 7, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12, the glucanase.

[0120] 13. The composition according to paragraph 7, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 13 or the polypeptide of SEQ ID NO: 14, the glucanase.

[0121] 14. The composition according to item 1, wherein the xylanase is GH10 xylanase.

[0122] 15. Xylanase is, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16. ii. Polypeptides having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18. A composition according to item 1 or 14, selected from the group consisting of the following.

[0123] 16. The composition according to item 14, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16, the xylanase.

[0124] 17. The composition according to item 14, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18, the xylanase.

[0125] 18. The composition according to item 1, wherein the lipase has sequence identity of at least 80%, for example, at least 85%, 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%, and at least 100% with respect to the mature polypeptide of SEQ ID NO: 19 or the polypeptide of SEQ ID NO: 20.

[0126] 19. A composition according to item 1 or 18, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 19 or the polypeptide of SEQ ID NO: 20, wherein the lipase is...

[0127] 20. The composition according to item 1, wherein the phosphodiesterase has at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 21 or the polypeptide of SEQ ID NO: 22.

[0128] 21. The composition according to item 1 or 20, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 21 or the polypeptide of SEQ ID NO: 22, the phosphodiesterase.

[0129] 22. The composition according to paragraph 2, wherein the dental care component is selected from the group consisting of benzoates (preferably sodium benzoate), EDTA, ethanol, fluoride (preferably sodium fluoride), glycerol, hydrogen peroxide, mannitol, phosphates (preferably sodium phosphate), SDS, sorbates (preferably potassium sorbate), and sorbitol.

[0130] 23. The combination of enzymes comprises one or more alpha-amylases, one or more glucanases, and one or more phosphodiesterases, as described in any one of claims 1 to 22.

[0131] 24. The enzyme combination comprises one or more alpha-amylases, one or more glucanases, and one or more lipases, as described in any one of claims 1 to 22.

[0132] 25. The combination of enzymes comprises one or more alpha-amylases, one or more xylanases, and one or more lipases, as described in any one of claims 1 to 22.

[0133] 26. The combination of enzymes comprises one or more alpha-amylases, one or more xylanases, and one or more phosphodiesterases, as described in any one of claims 1 to 22.

[0134] 27. The combination of enzymes comprises one or more alpha-amylases, one or more glucanases, one or more xylanases, and one or more lipases, as described in any one of claims 1 to 22.

[0135] 28. The combination of enzymes comprises one or more alpha-amylases, one or more glucanases, one or more xylanases, and one or more phosphodiesterases, as described in any one of claims 1 to 22.

[0136] 29. The combination of enzymes comprises one or more alpha-amylases, one or more glucanases, one or more xylanases, one or more lipases, and one or more phosphodiesterases, according to any one of claims 1 to 22.

[0137] 30. The composition according to any one of the headings 1 to 29, wherein the animal is a companion animal selected from the group consisting of dogs, cats, horses, or llamas.

[0138] 31. The companion animal is a dog, according to any one of the claims 1 to 30.

[0139] 32. A dental care composition comprising a combination of enzymes and at least one dental care ingredient, wherein the combination of enzymes is i. One or more types of alpha-amylase, ii. One or more glucanases and / or one or more xylanases, iii. One or more lipases and / or one or more phosphodiesterases A dental care composition containing [the specified ingredient].

[0140] 33. The dental care composition described in paragraph 32, wherein the alpha-amylase is GH13 alpha-amylase.

[0141] 34. The dental care composition according to claim 32 or 33, wherein the alpha-amylase has at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2.

[0142] 35. The dental care composition according to paragraph 34, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2.

[0143] 36. The dental care composition according to item 32, wherein the glucanase is selected from the group consisting of GH5 beta-glucanase, GH6 glucanase, GH7 endo-glucanase, GH7 exo-glucanase, GH7 beta-glucanase, and GH16 beta-glucanase.

[0144] 37. Glucanase is, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4. ii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6. iii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8. iv. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10. v. Polypeptides having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12, and vi. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 13 or the polypeptide of SEQ ID NO: 14. A dental care composition according to item 32 or 36, selected from the group consisting of the following.

[0145] 38. The dental care composition according to paragraph 37, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4, glucanase.

[0146] 39. The dental care composition according to paragraph 37, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6.

[0147] 40. The dental care composition according to paragraph 37, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8.

[0148] 41. The dental care composition according to paragraph 37, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10, glucanase.

[0149] 42. The dental care composition according to paragraph 37, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12.

[0150] 43. The dental care composition according to paragraph 37, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 13 or the polypeptide of SEQ ID NO: 14.

[0151] 44. The dental care composition described in paragraph 32, wherein the xylanase is GH10 xylanase.

[0152] 45. Xylanase is, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16. ii. Polypeptides having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18. A dental care composition according to item 32 or 44, selected from the group consisting of the following.

[0153] 46. ​​The dental care composition according to paragraph 45, comprising, essentially, or consisting of xylanase, the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16.

[0154] 47. The dental care composition according to paragraph 45, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18, xylanase.

[0155] 48. The dental care composition according to claim 32, wherein the lipase has at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 19 or the polypeptide of SEQ ID NO: 20.

[0156] 49. The dental care composition according to paragraph 48, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 19 or the polypeptide of SEQ ID NO: 20.

[0157] 50. The dental care composition according to item 32, wherein the phosphodiesterase has at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 21 or the polypeptide of SEQ ID NO: 22.

[0158] 51. The dental care composition according to paragraph 50, comprising, essentially, or consisting of the mature polypeptide of SEQ ID NO: 21 or the polypeptide of SEQ ID NO: 22, the phosphodiesterase.

[0159] 52. The dental care composition according to item 32, wherein the dental care component is selected from the group consisting of benzoates (preferably sodium benzoate), EDTA, ethanol, fluoride (preferably sodium fluoride), glycerol, hydrogen peroxide, mannitol, phosphates (preferably sodium phosphate), SDS, sorbates (preferably potassium sorbate), and sorbitol.

[0160] 53. The combination of enzymes comprises one or more alpha-amylases, one or more glucanases, and one or more phosphodiesterases, as described in any one of paragraphs 32 to 52.

[0161] 54. The combination of enzymes comprises one or more alpha-amylases, one or more glucanases, and one or more lipases, as described in any one of the items 32 to 52.

[0162] 55. The combination of enzymes comprises one or more alpha-amylases, one or more xylanases, and one or more lipases, as described in any one of the items 32 to 52.

[0163] 56. The combination of enzymes comprises one or more alpha-amylases, one or more xylanases, and one or more phosphodiesterases, as described in any one of claims 32 to 52.

[0164] 57. A dental care composition according to any one of claims 32 to 52, wherein the combination of enzymes comprises one or more alpha-amylases, one or more glucanases, one or more xylanases, and one or more lipases.

[0165] 58. A dental care composition according to any one of claims 32 to 52, wherein the combination of enzymes comprises one or more alpha-amylases, one or more glucanases, one or more xylanases, and one or more phosphodiesterases.

[0166] 59. A dental care composition according to any one of claims 32 to 52, wherein the combination of enzymes comprises one or more alpha-amylases, one or more glucanases, one or more xylanases, one or more lipases, and one or more phosphodiesterases.

[0167] 60. A dental care composition comprising a combination of enzymes and at least one dental care ingredient, wherein the combination of enzymes is b. Alpha-amylase having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2. c. A group of glucanases, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO: 4. ii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO: 6. iii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO: 8. iv. Polypeptides having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10, and v. Polypeptides having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO: 12. A group of glucanases consisting of d. Xylanase having at least 80%, for example at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16. e. A lipase having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18. A dental care composition containing [the specified ingredient].

[0168] 61.a. Alpha-amylase containing, essentially derived from, or consisting of the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2 b. A glucanase comprising, essentially comprising, or consisting of the mature polypeptide of SEQ ID NOs. 3, 5, 7, 9, and 11, or the polypeptide of SEQ ID NOs. 4, 6, 8, 10, and 12. c. A xylanase comprising, essentially having, or consisting of the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO: 16. d. Lipases containing, essentially derived from, or consisting of the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO: 18. A dental care composition as described in paragraph 60, comprising:

[0169] 62. The dental care composition according to paragraph 60, wherein the dental care component is selected from the group consisting of benzoates (preferably sodium benzoate), EDTA, ethanol, fluoride (preferably sodium fluoride), glycerol, hydrogen peroxide, mannitol, phosphates (preferably sodium phosphate), SDS, sorbates (preferably potassium sorbate), and sorbitol.

[0170] 63. A dental care composition according to any one of the items 32 to 62, used as a drug for dental hygiene in animals.

[0171] 64. The dental care composition described in paragraph 63, wherein the agent is toothpaste, toothpaste gel, toothpaste cream, toothpaste powder, toothpaste agent, oral spray, chewable product, oral freshener, mouthwash, mouth rinse and dental pack.

[0172] 65. The animal is a companion animal, as described in paragraph 63, for the dental care composition.

[0173] 66. The companion animal is selected from the group consisting of dogs, cats, horses, and llamas, for the dental care composition according to item 65.

[0174] 67. The companion animal is a dog, the dental care composition as described in subheading 65 or 66.

[0175] 68. A method for preventing or removing an oral biofilm in an animal, comprising contacting the oral biofilm with a composition described in any one of the headings 1 to 32 or a dental care composition described in any one of the headings 32 to 67.

[0176] 69. A method for reducing the risk of oral biofilm formation in animals, comprising contacting the oral biofilm with the composition described in any one of the items 1 to 32 or the dental care composition described in any one of the items 32 to 67.

[0177] 70. A method for removing an oral biofilm formed in an animal, comprising contacting the oral biofilm with a composition described in any one of the items 1 to 31 or a dental care composition described in any one of the items 32 to 67.

[0178] 71. A method for preventing or removing plaque, reducing the likelihood of caries, or reducing gingivitis in an animal, comprising bringing a composition described in any one of paragraphs 1 to 67 into contact with the teeth of the animal.

[0179] 72. Use of any one of the compositions described in any of the headings 1 to 31 or any one of the dental care compositions described in any one of the headings 33 to 67 to prevent or remove plaque, reduce the likelihood of caries, or reduce gingivitis in animals.

[0180] 73. A composition according to any one of the items 1 to 67, for use in preventing or removing biofilm formation in the dentition of animals.

[0181] 74. A composition according to any one of the paragraphs 1 to 67, for use in preventing plaque formation in animals or for use in removing plaque in animals.

[0182] 75.i. A dental care composition as described in any one of paragraphs 32 to 67, ii. Instructions for use and A kit of parts including parts. [Examples]

[0183] [Table 1]

[0184] [Table 2]

[0185] Example 1: Preventive assay for multiple biofilms in canids The biofilm-preventive effects of the various enzyme blends listed above were tested on multiple biofilms grown in 96-well microtiter plates containing the following five canine dental pathogens: Neisseria zoodegmatis (DSM21643), Pasteurella canis (DSM22968), Pseudomonas fragi (DSM3456), Capnocytophaga canis (DSM101831), and Enterococcus faecalis (DSM20478) (see references below). A 96-well microtiter plate (Nunclon Delta surface ThermoScientific #167008) was filled with 75 μl of Tripticase Soy Broth (TSB) + 2% glucose containing 1 × 10⁷ CFU / ml bacterial inoculum of each of the five strains described above, and 25 μl of enzyme solution in citrate-phosphate buffer pH 6 (McIIvaine buffer), to a final concentration of 60 ppm. For the control samples, the enzyme solution was replaced with McIIvaine buffer pH 6. The plates were incubated at 30°C for 72 hours without shaking in a Thermo Scientific® Rectangular AnaeroBox® Container (ThermoScientific AnaeroGen 2.5L #AN0025A) under microaerophilic conditions. The enzyme samples and control samples were subjected to eight repeated tests.

[0186] A citrate-phosphate buffer solution with a pH of 6 (McIIvaine buffer) was prepared by mixing 12.63 ml of 0.2 M Na2HPO4 with 7.37 ml of 0.1 M citrate.

[0187] After incubation, the plates were gently washed twice with 100 μL of 0.9% sodium chloride to remove suspended bacteria, and the biofilm was stained with 0.095% crystal violet solution at room temperature for 15 minutes. The plates were rinsed twice with 100 μL of 0.9% sodium chloride, and the remaining dye was dissolved in a solution of 96% ethanol and 0.1% acetic acid in water. Absorbance was measured at 600 nm using a SpectraMax M3, Molecular Devices microplate reader.

[0188] For data processing purposes, absorbance was assumed to be proportional to the degree of biofilm remaining after enzyme or buffer treatment. The results were expressed as a percentage of biofilm prevention and calculated as follows: 100 - ((A600nm of enzyme-treated sample) / (A600nm of buffer-controlled sample) × 100) (wherein A600nm refers to the average of 8 absorbance measurements taken at 600nm for either the enzyme-treated or buffer-treated sample).

[0189] The results are listed in Table 1.

[0190] [Table 3]

[0191] [Table 4]

[0192] The results showed that the enzyme blends listed above have a preventive effect against the formation of multiple biofilms in canids.

[0193] Repeating this experiment demonstrated once again that a high level of prevention is possible with combinations within the scope of the present invention.

[0194] [Table 5]

[0195] Example 2: Assay for the removal of multiple biofilms in canids The biofilm removal effects of the various enzyme blends listed above were tested on multiple biofilms grown in 96-well microtiter plates from the following five canine dental pathogens: Neisseria zoodegmatis DSM21643, Pasteurella canis DSM22968, Pseudomonas fragi DSM3456, Capnocytophaga canis DSM101831, and Enterococcus faecalis DSM20478 (see references below). Each of the five strains was measured in 1 × 10⁶ wells of the above-mentioned bacteria on a 96-well microtiter plate (Nunclon Delta surface ThermoScientific #163320). 7 The plates were filled with 200 μl of Tripticase Soy Broth (TSB) + 2% glucose containing a bacterial inoculum of CFU / ml. The plates were incubated at 30°C for 72 hours without shaking in a Thermo Scientific® Rectangular AnaeroBox® Container (ThermoScientific AnaeroGen 2.5L #AN0025A) under microaerophilic conditions.

[0196] After incubation, the supernatant containing suspended bacteria was taken from each well, and the sample was treated with either 200 μL of citrate-phosphate buffer pH 8 (McIlvaine buffer) as a control, or the 60 ppm enzyme blend listed above in McIlvaine buffer pH 8, for 30 minutes at 37°C with 50 rpm shaking. The enzyme samples and control samples were subjected to eight repeated tests.

[0197] A citrate-phosphate buffer solution with a pH of 8 (McIIvaine buffer) was prepared by mixing 19.45 ml of 0.2 M Na2HPO4 with 0.55 ml of 0.1 M citrate.

[0198] After enzymatic treatment, the samples were rinsed twice with 200 μL of 0.9% sodium chloride and stained with 0.095% crystal violet solution at room temperature for 15 minutes. The samples were then rinsed three times with 200 μL of 0.9% sodium chloride, the dye was extracted with 33% acetic acid solution, and the absorbance at 600 nm was measured.

[0199] For data processing purposes, absorbance was assumed to be proportional to the amount of biofilm remaining after enzyme or buffer treatment. The percentage of biofilm removal was calculated as follows: 100 - ((A600nm of enzyme-treated sample) / (A600nm of buffer-treated control sample) × 100) (wherein A600nm refers to the average of 8 absorbance measurements taken at 600nm using SpectraMax M3, Molecular Devices for either the enzyme-treated or buffer-treated sample). The results are listed in Table 2.

[0200] [Table 6]

[0201] The results showed that the enzyme blends listed above have a destructive effect on the formation of multiple biofilms in canids.

[0202] References 1.Cunha et al.BMC Veterinary Research(2020)16:469.A polymicrobial biofilm model for testing the antimicrobial potential of a nisin-biogel for canine periodontal disease control 2.Holcombe et al.,2014,PLOS ONE | DOI:10.1371 / journal.pone.0113744.Early Canine Plaque Biofilms:Characterization of Key Bacterial Interactions Involved in Initial Colonization of Enamel 3.A.Manuel,J.V.Rao,K.John&J.M.Aranjani.2014.Biofilm Production and Antibiotic Susceptibility of Planktonic and Biofilm Bacteria of Canine Dental Tartar Isolates.Acta Scientiae Veterinariae.42:1202.

Claims

1. A composition for animal dental care, comprising a combination of enzymes, the combination of enzymes is i. One or more types of alpha-amylase, ii. One or more glucanases and / or one or more xylanases, iii. One or more lipases and / or one or more phosphodiesterases A composition containing the following:

2. The composition according to claim 1, wherein the alpha-amylase is GH13 alpha-amylase.

3. The composition according to claim 1, wherein the glucanase is selected from the group consisting of GH5 beta-glucanase, GH6 glucanase, GH7 endo-glucanase, GH7 exo-glucanase, GH7 beta-glucanase, and GH16 beta-glucanase.

4. The composition according to claim 1, wherein the xylanase is GH10 xylanase.

5. The composition according to claim 1, wherein the combination of enzymes comprises one or more alpha-amylases, one or more glucanases, and one or more phosphodiesterases.

6. The composition according to claim 1, wherein the combination of enzymes comprises one or more alpha-amylases, one or more glucanases, and one or more lipases.

7. The composition according to claim 1, wherein the combination of enzymes comprises one or more alpha-amylases, one or more xylanases, and one or more lipases.

8. A dental care composition comprising a combination of enzymes and at least one dental care component, wherein the combination of enzymes is i. One or more types of alpha-amylase, ii. One or more glucanases and / or one or more xylanases, iii. One or more lipases and / or one or more phosphodiesterases A dental care composition containing [the specified ingredient].

9. A dental care composition comprising a combination of enzymes and at least one dental care component, wherein the combination of enzymes is a. One or more polypeptides having alpha-amylase activity, having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 1 or the polypeptide of SEQ ID NO: 2, b. One or more polypeptides having glucanase activity, i. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 3 or the polypeptide of SEQ ID NO:

4. ii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 5 or the polypeptide of SEQ ID NO:

6. iii. A polypeptide having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 7 or the polypeptide of SEQ ID NO:

8. iv. Polypeptides having at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 9 or the polypeptide of SEQ ID NO: 10, and v. Polypeptides having at least 80%, for example, at least 85%, 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 at least 100% sequence identity with the mature polypeptide of SEQ ID NO: 11 or the polypeptide of SEQ ID NO:

12. One or more polypeptides selected from the group consisting of, c. One or more polypeptides having xylanase activity, wherein one or more polypeptides have at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 15 or the polypeptide of SEQ ID NO:

16. d. One or more polypeptides having lipase activity, wherein one or more polypeptides have at least 80%, for example, at least 85%, 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%, and at least 100% sequence identity with respect to the mature polypeptide of SEQ ID NO: 17 or the polypeptide of SEQ ID NO:

18. A dental care composition containing [the specified ingredient].

10. A method for preventing or removing an oral biofilm in an animal, comprising contacting the oral biofilm with the composition described in any one of claims 1 to 7 or the dental care composition described in claim 8 or 9.

11. A method for reducing the risk of oral biofilm formation in animals, comprising contacting the oral biofilm with the composition according to any one of claims 1 to 7 or the dental care composition according to claim 8 or 9.

12. A method for removing an oral biofilm formed in an animal, comprising contacting the oral biofilm with the composition described in any one of claims 1 to 7 or the dental care composition described in claim 8 or 9.

13. A method for preventing or removing plaque in an animal, reducing the likelihood of caries, or reducing gingivitis, comprising bringing the composition according to any one of claims 1 to 9 into contact with the teeth of the animal.

14. Use of the composition according to any one of claims 1 to 7 or the dental care composition according to claim 8 or 9 for preventing or removing plaque, reducing the likelihood of caries, or reducing gingivitis in animals.

15. A composition according to any one of claims 1 to 7 or a dental care composition according to claim 8 or 9, for use in preventing biofilm formation in the dentition of animals or for use in removing biofilm in the dentition of animals.

Images