TW201705932A - Peracid-generating compositions - Google Patents

Abstract

Described herein are tooth whitening strips comprising an hydratable adhesive film having a granular bleaching ingredient attached thereto, wherein upon hydration, the granular bleaching ingredient releases hydrogen peroxide which is used by an enzyme catalyst having perhydrolytic activity to enzymatically produce an effective amount of peracid bleaching agent from an acyl donor substrate. Methods of making and use the tooth whitening strips are also provided.

Description

Acid-producing composition

Illustrated herein is a tooth whitening toothpaste comprising a hydratable adhesive film to which a particulate bleaching component is attached, wherein upon hydration, the particulate bleaching component releases hydrogen peroxide, the system An enzymatic catalyst having perhydrolysis activity is used to enzymatically produce an effective amount of peracid bleach from the thiol donor matrix. The method and use of the tooth whitening tooth paste are also provided.

There is a need for a whitening toothpaste suitable for home use which has a reduced total peroxide concentration but provides improved whitening activity.

The present invention provides a whitening toothpaste comprising a granular bleaching component combined with an enzyme having perhydrolysis activity ("perhydrolase"), which comprises a conservative structural motif of the carbohydrate esterase family 7 and a mercapto donor. In the use, the peroxide released from the granular bleaching component reacts with the sulfhydryl donor in the presence of a perhydrolase to form a peroxyacid, which is in direct proximity to the teeth without substantial formulation. Dilution in the excipients thus allows for a greater total amount of peroxide to enhance tooth whitening.

The dental patch comprises an adhesive film which may be a single layer or a plurality of layers (e.g., two layers) which become sufficiently viscous to adhere to the teeth when hydrated with water or saliva. The particulate bleaching component is attached to the side of the film that will contact the teeth. When used, the bleaching component is placed directly on the teeth (i.e., between the teeth and the adhesive layer). Then, the granules release peroxidation by rapidly dissolving in water. Things. The bleaching component can optionally be embedded in a rapidly dissolving material such as sodium sulfate, corn starch or gum arabic. Optionally, the second layer provided by the tooth in the film is for extended exposure time. The second layer may be insoluble in water, the user needs to remove the dental patch after treatment, or may be eroded in water, which will cause the dental patch to dissolve after sufficient treatment. The dental patch further includes a perhydrolase (an enzyme capable of catalyzing the reaction of a carboxylic acid ester and hydrogen peroxide to form a peroxyacid), which may also be provided in a particle form on the surface of the membrane, and a thiol donor, for example, And a carboxylic acid and a mercapto compound, for example, triacetin or sorbitol hexaacetate, wherein the mercapto donor system reacts with the peroxide in the dental patch from the presence of a perhydrolase Peroxyacid is formed which enhances the bleaching action of the dental prosthesis.

Some specific embodiments of the present invention provide a tooth whitening toothpaste comprising a hydratable adhesive film comprising a first side and a second side, the first side comprising a particulate bleaching component attached thereto, wherein the tooth The whitening toothpaste further comprises the following attached to the first side of the film in a film or on a film or in a particulate form: a) a hydrolyzed enzyme having a carbohydrate ester aligned with the reference sequence SEQ ID NO: Enzyme Family 7 (CE-7) tag motif comprising: i) a RGQ motif at a position corresponding to positions 118-120 of SEQ ID NO: 1; ii) a GXSQG motif corresponding to SEQ ID NO: position 1 position 186-190; and iii) HE motif at a position corresponding to positions 303-304 of SEQ ID NO: 1; and b) at least one thiol donor matrix selected from the group consisting of : i) an ester of the formula: [X] _m R ₅ wherein X = an ester group of the formula R ₆ C(O)O; R ₆ = a C1 to C7 linear, branched or cyclic hydrocarbon moiety, optionally is a hydroxyl group or a C1 to C4 substituted alkoxy group, wherein R ₆ = C2 to C7 to time, R ₆ include any one or more ether bonds linked; R ₅ = C1 C6 straight chain, branched or cyclic hydrocarbon moiety or a cyclic heteroaryl five or six-membered ring aryl or heteroaryl moieties, any hydroxy group is substituted; R ₅ is in which each of the carbon atoms based Independently comprising no more than one hydroxyl group or no more than one ester group or a carboxylic acid group; wherein R ₅ optionally includes one or more ether linkages; m is the number of carbon atoms from 1 to R ₅ An integer, and wherein the ester has a water solubility of 5 ppm at 25 ° C; ii) a glyceride having the following structural formula Wherein R ₁ = C1 to C7 is a straight or branched alkyl group, any of which is substituted by a hydroxy group or a C1 to C4 alkoxy group and R ₃ and R ₄ are independently H or R ₁ C(O); iii) one or a variety of esters of the following formula Wherein R ₁ is a C1 to C7 straight or branched alkyl group, which is optionally substituted by a hydroxy or C1 to C4 alkoxy group and R ₂ is a C1 to C10 straight or branched alkyl group, an alkenyl group, an alkynyl group, Aryl, alkylaryl, alkylheteroaryl, heteroaryl, (CH ₂ CH ₂ O) _n , or (CH ₂ CH(CH ₃ )-O) _n H and n is from 1 to 10; An acetylated saccharide selected from the group consisting of acetylated monosaccharides, acetylated disaccharides, and acetylated polysaccharides; wherein upon hydration of the hydratable adhesive film, the hydrogen peroxide is composed of granular bleaching The fraction is released and the enzyme is an enzymatically effective amount of peroxyacid.

A further embodiment of the present invention provides a method of whitening teeth comprising providing a packaging system comprising a tooth whitening toothpaste according to any of the preceding claims; removing the tooth whitening toothpaste from the packaging system; The tooth whitening tooth paste directly contacts the tooth for a period of time sufficient to whiten the tooth; wherein the tooth whitening tooth paste is hydrated by the presence of water in the mouth or on the surface of the tooth or after step (b) but in step (c) Before hydration.

Further areas of applicability of the present invention will become apparent from the Detailed Description. The detailed description and specific examples are intended to be illustrative of the preferred embodiments of the invention

A brief description of the biological sequence

The following sequences are in accordance with 37 CFR §§ 1.821-1.825 ("Disclosure Requirements for Nucleotide Sequences and/or Amino Acid Sequences - Sequence Rules") and in accordance with World Intellectual Property Organization (WIPO) Guidelines ST.25 (2009) and Europe Sequence Listing Requirements in the Patent Convention (EPC) and Patent Cooperation Treaty (PCT) Rules 5.2 and 49.5 (a-bis), and Chapter 208 and Appendix C of the Administrative Instructions. The symbols and formats used in the nucleotide and amino acid sequence data are in accordance with the rules set forth in 37 C.F.R.§ 1.822.

SEQ ID NO: 1 is the amino acid sequence of the Thermotoga maritima C277S variant perhydrolase.

SEQ ID NO: 2 is the amino acid sequence of a fusion protein comprising a sea stagnation C277S variant perhydrolase coupled to a tooth-binding structural region (also known as "EZ-7" in International Patent Application Publication No. WO2012/ 087970A2-Bartrick, etc.).

SEQ ID NO: 3 encoding from Bacillus subtilis (Bacillus subtilis) ATCC ^® 31954 ^TM Bacillus head of cephalosporin C (cephalosporin C) to the nucleic acid sequence of the enzyme acetyl.

SEQ ID NO: 4 is the amino acid sequence from Bacillus subtilis ATCC ^® 31954 ^TM to the head cephalosporin C acetyl Bacillus enzyme.

SEQ ID NO: 5 is the amino acid sequence of Bacillus subtilis subsp.

SEQ ID NO: 6 is the amino acid sequence from Bacillus subtilis ATCC ^® 6633 ^TM to the head cephalosporin C acetyl Bacillus enzyme.

SEQ ID NO: 7 is the head ATCC ^® 14580 ^TM Bacillus cephalosporin C acetyl enzyme to the amino acid sequence derived from Bacillus licheniformis (B. licheniformis).

SEQ ID NO: 8 is the amino acid sequence of the acetyl xylan esterase (acetyl xylan esterase) from B. pumilus PS213 .

SEQ ID NO: 9 is the amino acid sequence of the acetyl xylan esterase from the ATCC ^® 27405 ^TM thermophilic Clostridium (Clostridium thermocellum).

SEQ ID NO: 10 is the amino acid sequence of the beta xylan esterase from the cloned Thermotoga neapolitana .

SEQ ID NO: 11 is the amino acid sequence of the acetyl xylan esterase acetyl xylan esterase from Thermotoga maritima MSB8.

SEQ ID NO: 12 is the amino acid sequence of the acetyl xylan esterase from Thermoanaerobacterium sp . JW/SL YS485.

SEQ ID NO: 13 is the amino acid sequence of the cephalosporin C deacetylase from Bacillus halodurans C-125.

SEQ ID NO: 14 is the amino acid sequence of the cephalosporin C deacetylase from Bacillus clausii KSM-K16.

SEQ ID NO: 15 is the amino acid sequence of a cloned new Apollo Thermotoxin xylan esterase variant from U.S. Patent Application Publication No. 2010-0087529, the entire disclosure of which is incorporated herein by reference. Wherein the Xaa residue at position 277 is Ala, Val, Ser, or Thr.

SEQ ID NO: 16 is the amino acid sequence of the marine robes MSB8 acetyl xylan esterase variant from U.S. Patent Application Publication No. 2010-0087529, wherein the Xaa residue at position 277 is Ala, Val, Ser. , or Thr.

SEQ ID NO: 17 is the amino acid sequence derived from the Thermotoga lettingae xylan esterase variant from US Patent Application Publication No. 2010-0087529, wherein the Xaa residue at position 277 The base is Ala, Val, Ser, or Thr.

SEQ ID NO: 18 is the amino acid sequence of the Thermotoga sinensis xylan esterase variant from U.S. Patent Application Publication No. 2010-0087529, wherein the Xaa residue at position 277 is Ala, Val, Ser. , or Thr.

SEQ ID NO: 19 is the amino acid sequence of the Thermotoga sp. RQ2 acetyl xylan esterase variant derived from "RQ2(a)" of U.S. Patent Application Publication No. 2010-0087529. Wherein the Xaa residue at position 277 is Ala, Val, Ser, or Thr.

SEQ ID NO: 20 is the amino acid sequence of the Thermus sp. RQ2 acetyl xylan esterase variant derived from "RQ2(b)" of U.S. Patent Application Publication No. 2010-0087529, wherein at position 278 The Xaa residue is Ala, Val, Ser, or Thr.

SEQ ID NO: 21 is the amino acid sequence of the Thermotoga xylan esterase of Letinger.

SEQ ID NO: 22 is the amino acid sequence of the Thermotoga sylvestre xylan esterase.

SEQ ID NO: 23 is the amino acid sequence of the first acetyl xylan esterase from the Thermus sp. RQ2, designated "RQ2 (a)".

SEQ ID NO: 24 is the amino acid sequence of the second acetyl xylan esterase from the Thermus sp. RQ2 described as "RQ2 (b)".

SEQ ID NO: 25 is the amino acid sequence of Thermoanearobacterium saccharolyticum cephalosporin C deacetylase.

SEQ ID NO: 26 is the amino acid sequence of the acetyl xylan esterase from Lactococcus lactis (Lactococcus lactis) (GENBANK ^® registration number ABX75634.1) of.

SEQ ID NO: 27 is the amino acid sequence of the acetyl xylan esterase from Mesorhizobium thrips (Mesorhizobium loti) (GENBANK ^® registration number BAB53179.1) of.

SEQ ID NO: 28 is the amino acid sequence of the acetyl xylan esterase from Geobacillus stearothermophilus (Geobacillus stearothermophilus) (GENBANK ^® registration number AAF70202.1) of.

Detailed description

The following description of the preferred embodiments is merely illustrative and is in no way intended to limit the invention, the application, or the application.

"an," and "the" and "the" The articles "a", "an" and "the"

The word "comprising", as used herein, is intended to mean the presence of the features, integers, steps or components described in the claims, but does not exclude the presence or addition of one or more other features, integers, steps, components Or a collection thereof. The word "including" is intended to include the specific examples encompassed by the words "mainly composed" and "composed as". Similarly, the term "main composition" is intended to include the specific examples covered by the word "composed as".

As used herein, the term "about" used to modify the amount of a component or reactant used refers to a change that can occur in numerical quantities, for example, from typical measurements and liquid handling procedures used to make concentrates or Solutions used in the real world; due to unintentional loss in these processes; differences in manufacturing, source, or purification used to make components or to perform methods; The term "about" also encompasses the number of differences due to the different equilibrium conditions of the composition resulting from a particular initial mixture. Whether or not modified by the word "about", the scope of the patent application includes the equivalent of the quantity.

In the current situation, all scopes are inclusive and combinable. For example, when quoting the range of "1 to 5", the recited range should be interpreted to include "1 to 4", "1 to 3", "1-2", "1-2 & 4-5", " 1-3 & 5", etc.

As used herein, the terms "substrate", "suitable substrate", "thiol donor", and "carboxylate matrix" are used interchangeably to mean, in particular, (a) one or more esters having the formula Class [X] _m R ₅ wherein X is an ester group of the formula R ₆ C(O)O; R ₆ is a C1 to C7 straight chain, branched or cyclic hydrocarbon moiety, any of which is a hydroxyl group or a C1 to C4 alkane Substituted by an oxy group, wherein when R ₆ is C2 to C7, R ₆ optionally includes one or more ether linkages; R ₅ is a C1 to C6 straight chain, a branched or cyclic hydrocarbon moiety or a cyclic five member a heteroaryl or a six-membered cyclic aryl or heteroaryl moiety, optionally substituted by a hydroxy group; wherein each of the carbon atoms in R ₅ includes no more than one hydroxy group or no more than one An ester group, and wherein R ₅ optionally includes one or more ether linkages; m is an integer from the number of carbon atoms in 1 to R ₅ , and one or more esters having a water solubility of 5 ppm at 25 ° C; (b) one or more glycerides having the following structural formula Wherein R ₁ is a C1 to C7 straight or branched alkyl group, which is optionally substituted by a hydroxy group or a C1 to C4 alkoxy group and R ₃ and R ₄ are independently H or R ₁ C(O); or (c) One or more esters of the formula Wherein R ₁ is a C1 to C7 straight or branched alkyl group, which is optionally substituted by a hydroxy or C1 to C4 alkoxy group and R ₂ is a C1 to C10 straight or branched alkyl group, an alkenyl group, an alkynyl group, Aryl, alkylaryl, alkylheteroaryl, heteroaryl, (CH ₂ CH ₂ O) _n , or (CH ₂ CH(CH ₃ )-O) _n H and n is from 1 to 10; or d) one or more acetylated monosaccharides, acetylated disaccharides, or acetaminolated polysaccharides; or (e) any combination of (a) to (d).

As used herein, the term "peroxyacid (peroxyacid)" is synonymous with peroxyacid, peroxycarboxylic acid (peroxycarboxylic acid), peroxy acid, percarboxylic acid (peroxycarboxylic acid). Per carboxylic acid) and peroxoic acid.

As used herein, the term "peracetic acid" is abbreviated as "PAA" and is synonymous with peroxyacetic acid, ethane peroxyacid and all CAS registration numbers 79-21-0.

As used herein, the term "monoacetin" is synonymous with glycerol monoacetate, glycerin monoacetate, and glyceryl monoacetate.

As used herein, the term "diacetin" is synonymous with glycerol diacetate; glycerin diacetate, glyceryl diacetate, and all Synonyms of other CAS registration numbers 25395-31-7.

As used herein, the term "triacetin (triacetin)" is synonymous with glycerin triacetate; glycerol triacetate; glyceryl triacetate 1,1,2,3-triethoxypropane; 1,2,3-propanetriol triacetate and all other CAS registration numbers 102-76-1.

As used herein, the terms "sugar" and "saccharide" are used to mean a mono-, di-, and polysaccharide comprising at least one ethyl hydrazide group. Examples include, but are not limited to, glucose pentaacetic acid; xylose tetraacetic acid; acetylated xylan; acetylated xylan fragment; β-D-ribofuranosyl-1,2,3,5-tetraacetic acid; -O-acetamidine-D-galactose; and tri-O-acetamidine-gluconate.

As used herein, the terms "hydrocarbyl", "hydrocarbyl group", and "hydrocarbyl moiety" refer to a straight, branched or cyclic distribution of a carbon atom by means of a carbon-carbon mono, di or triple bond. And/or by ether bonds to replace one with a hydrogen atom. These hydrocarbyl groups can be aliphatic and/or aromatic. Examples of the hydrocarbyl group include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, cyclopropyl, cyclobutyl, pentyl, cyclopentyl, methylcyclopentyl. , hexyl, cyclohexyl, benzyl, and phenyl. In a preferred embodiment, the hydrocarbyl moiety is a linear, branched or cyclic distribution of carbon atoms which is bonded by a carbon-carbon single bond and/or by an ether linkage to be substituted with a hydrogen atom.

As used herein, 1,2-ethanediol; 1,2-propanediol; 1,3-propanediol; 1,2-butanediol; 1,3-butanediol; 2,3-butanediol; 1,4-butanediol; 1,2-pentanediol; 2,5-pentanediol; 1,5-pentanediol; 1,6-pentanediol; 1,2-hexanediol; 5-hexanediol; 1,6-hexanediol; and "monoesters" in mixtures thereof And the term "diester" means that the compound includes at least one ester group of the formula RC(O)O, wherein R is a C1 to C7 straight chain hydrocarbon moiety. In one embodiment, the carboxylate matrix is selected from the group consisting of propylene glycol diacetate (PGDA), ethylene glycol diacetate (EDGA), and mixtures thereof.

As used herein, the term "propylene glycol diacetate" is synonymous with 1,2-diethoxypropane, propylene diacetate, 1,2-propylene glycol diacetate, And synonymous with all other CAS registration numbers 623-84-7.

As used herein, the term "vinyl ethylene glycol diacetate" is synonymous with 1,2-diethoxymethoxyethane, vinyl diacetate, ethylene glycol diacetate, CAS. Synonyms in registration number 111-55-7.

As used herein, "appropriate enzymatic reaction mixture", "component suitable for in situ formation of peroxyacid", "appropriate reaction component", "suitable aqueous reaction mixture", "reaction mixture", and "produced" The "peroxyacid component" means a substance in which the reactant and the perhydrolase catalyst are in contact with each other and water (from the saliva and/or applied by the user to the hydratable adhesive film before use). The peroxyacid-producing component will comprise at least a hydrolytically active enzyme, wherein the perhydrolase is preferably at least one CE-7 perhydrolase (any fusion protein type targeting the body surface), at least one suitable carboxylate The acid ester matrix, the peroxide source, and water (from the saliva and/or applied by the user to the hydratable adhesive film prior to use).

As used herein, the term "perhydrolysis" is defined as the reaction of a selected substrate with a peroxide to form a peroxyacid. Typically, the inorganic peroxide is combined with the selected substrate to form a peroxyacid in the presence of a catalyst. As used herein, the term "chemical perhydrolysis" includes perhydrolysis wherein a substrate (peroxyacid precursor) is combined with a source of hydrogen peroxide, wherein the peroxyacid is formed in the absence of an enzyme catalyst. As used herein, the term "enzymatic perhydrolysis" includes perhydrolysis reactions in which a carboxylate matrix (peroxyacid precursor; the "mercapto donor") is hydrolyzed with hydrogen peroxide and, accordingly, The enzyme catalyst catalyzes the formation of peroxyacids.

As used herein, the term "perhydrolase activity" refers to a protein activity per unit mass (eg, milligrams) of stem cell weight, or a fixed catalyst weight.

As used herein, "one unit of enzyme activity" or "one unit of activity" or "U" is defined as the amount of perhydrolase activity required to produce 1 [mu]mol of peroxyacid product per minute at a particular temperature.

As used herein, the terms "enzymatic catalyst" and "perhydrolase catalyst" refer to a catalyst comprising a perhydrolysis activity enzyme and the type thereof may be whole microbial cells, permeabilized microbial cells, one or A cellular component of a plurality of microbial cell extracts, a partially purified enzyme, or a purified enzyme. The enzyme catalyst can also be chemically modified (e.g., by pegylation or by reaction with a crosslinking reagent). The perhydrolase catalyst can also be immobilized on a soluble or insoluble support; see, for example, enzyme and cell immobilization; Gordon F. Bickstaff, editor; Hamana Publishing, Toto Watts, New Jersey, USA; 1997. In one embodiment, the perhydrolase catalyst can be immobilized non-covalently in or on an oral care patch (eg, a whitening toothpaste) or a crankset. In another embodiment, the non-covalent immobilization of the dental patch or the sprocket can be via the use of a binding structural region of the peptide having a strong affinity for the substance in or on the dental patch or the sprocket (eg A fused protein comprising a perhydrolase coupled to a binding structural region of a peptide via an arbitrary peptide spacer). In another embodiment, the crankset is a deformable crankset. In still another embodiment, the perhydrolase catalyst is immobilized in or on the deformable dental disk after forming the dental impression.

As used herein, "acetyl xylan esterase" refers to an enzyme (E.C. 3.1.1.72; AXEs) which catalyzes the deacetylation of acetylated xylan and other acetylated sugars.

As used herein, the terms "cephalosporin C deacetylase" and "cephalosporin C hydrazine hydrolase" refer to an enzyme (EC 3.1.1.41) which catalyzes the cephalosporin as a head. Deacetylation of sporecin C and 7-aminocephalosporin (Manji et al., (1995) Environmental Microbial Applications 61 (6): 2224-2229). Amino acid sequences of many cephalosporin C deacetylases having significant perhydrolysis activity are provided herein.

As used herein, by "Bacillus subtilis ATCC ^® 31954 ^TM" refers to the words registered in the American Type Culture Center (American Type Culture Collection; ATCC) having international deposition under the registration number ATCC ^® 31954 ^TM of the bacterial cells. As defined herein, derived from Bacillus subtilis ATCC ^® 31954 ^TM having the amino acid sequences are significantly hydrolyzed by an enzyme activity in SEQ ID NO: 4 is provided (see, U.S. Patent Application Publication Number 2010-0041752).

As used herein, the term "Haiqi hot robe MSB8" refers to a bacterial cell that has been reported to have acetyl sterol esterase activity (GENBANK ^® NP_227893.1; see U.S. Patent Application Publication No. 2008-0176299). The amino acid sequence of the enzyme having perhydrolase activity from the sea robes MSB8 is provided as SEQ ID NO: 11. The seaweed robes MSB8 variant perhydrolase were provided as SEQ ID NOs: 1 and 16.

As used herein, "isolated nucleic acid molecule", "isolated polynucleotide", and "isolated nucleic acid fragment" are used interchangeably and refer to either RNA or DNA polymer, which is mono- or double. - Strands, optionally containing synthetic, unnatural or altered nucleotide bases. A DNA polymer type of isolated nucleic acid molecule can include one or more cDNA, genotypic DNA or synthetic DNA fragments.

The term "amino acid" refers to the basic chemical structural unit of a protein or polypeptide. The following abbreviations are used herein to identify specific amino acids:

As used herein, the term "about" is used to modify the amount of the component or reactant used, which refers to a numerical change that can occur, for example, via a typical measurement and liquid treatment process for the preparation of a concentrate or . As used herein, the terms "tag motif" and "identification motif" refer to conserved structures shared between families of enzymes having defined activities. The tag motif can be used in a defined family of matrices to define and/or identify a family of similar enzymes with similar enzymatic activity. The label motif can be a single continuous amino acid sequence or a collection of discrete, conserved motifs that together form a label motif. Typically, the conservative mode system is derived from an amino acid sequence. representative. In one embodiment, the perhydrolase enzyme used in the compositions and methods of the present invention comprises a CE-7 carbohydrate esterase tag motif.

As used herein, the term "sequence analysis software" refers to any computer algorithm or software program used to analyze nucleotide or amino acid sequences. "Sequence Analysis Software" is commercially available or independently developed. Typical sequence analysis software will include, but is not limited to, the GCG suite of programs (Wisconsin Software Pack Version 9.0, Accelrys Software, Inc., San Diego, Calif.), BLASTP, BLASTN, BLASTX (Ochu et al., Molecular Biology 215: 403-410 (1990)), and DNASTAR (DNASTAR, Inc. Madison South Park Road 1228, West Indiana 53715 United States), CLUSTALW (eg, version 1.83; Thompson et al, Nucleic Acids Research, 22(22): 4673-4680 (1994)), and the FASTA program incorporates the Smith Waltman algorithm (WR Pearson, Computer Methods Genomics Research, [Proc. Int. Symp.] (1994), date of the conference 1992, 111-20. Edit: Su Hai, Sandel. Issuer: Pu Nan, New York City, NY), Vector NTI (Informax, Bethesda, MD) and Sequencher v.4.05. In the context of this application, it should be understood that when using sequence analysis software for analysis, the results of the analysis will be based on the "initial value" of the referenced program unless otherwise stated. As used herein, "initial value" means any value or set of parameters set by the software manufacturer, the original loading of the software when first started.

The term "body surface" refers to any surface of the human body that serves as a beneficiary, such as a target for peroxyacid beneficiaries. The methods and compositions of the present invention relate to oral care applications and products. Therefore, the body surface includes the oral substance/surface. In one embodiment, the oral material comprises tooth enamel.

As used herein, the terms "whitening teeth" and "bleaching teeth" are used interchangeably and refer to improving the brightness of a tooth (eg, whitening). The whitening tooth pastes described herein comprise a component suitable for enzymatically producing an effective amount of peroxyacid to whiten the teeth upon hydration.

As used herein, "intrinsic stains" on a tooth refers to the color caused by a color chromophore between the enamel and the associated dentin. The inner color of human teeth tends to be yellower with age, which is due to the thinning of the enamel and the gradual darkening of the yellow dentin. Removal of intrinsic staining typically requires the use of peroxides or other oxidizing agents that can penetrate the enamel and decolorize the inner chromophore.

Compared to intrinsic staining, "exogenous staining" is formed on the surface of the tooth when the exogenous chromogenic material is bound to the enamel, usually attached to the surface. Most people accumulate a degree of unsightly exogenous staining on their teeth over time. This dyeing process is motivated by (1) ingesting foods and beverages containing tannins such as coffee, tea, or red wine; (2) using tobacco products; and/or (3) exposing to certain cationic substances (eg , tin, iron, and chlorhexidine). These substances tend to adhere to the enamel hydroxyapatite structure, which causes discoloration of the teeth and a simultaneous reduction in tooth whiteness. After a few years, exogenous staining can penetrate the enamel layer and cause intrinsic staining.

As used herein, the term "destain" or "destaining" refers to the process of removing staining from the surface of the oral cavity. The staining can be intrinsic staining, exogenous staining, or a combination thereof.

As used herein, "effective amount of perhydrolase" refers to the amount of perhydrolase enzyme that must achieve the enzymatic activity required for a particular use. Such effective amounts are well established by those skilled in the art and are based on a number of factors, such as the particular enzyme variant employed.

As used herein, "peroxide source" and "source of peroxide" refer to a compound that provides hydrogen peroxide at a concentration of about 1 mM or greater in an aqueous solution, including, but not limited to, hydrogen peroxide. Hydrogen peroxide adducts (eg, urea-hydrogen peroxide adducts (per urea peroxide)), perborates, and percarbonates. As defined herein, the peroxide source is in the form of a particulate particle in the whitening toothpaste of the present invention wherein the user hydrolyzes the particulate peroxide particles to release an effective amount of hydrogen peroxide. As defined herein, the concentration of hydrogen peroxide provided by the peroxide compound in the aqueous reaction formulation is initially at least 0.1 mM or greater upon binding of the reactive components. In one embodiment, the hydrogen peroxide concentration in the aqueous reaction formulation is at least 0.5 mM. In one embodiment, the hydrogen peroxide concentration in the aqueous reaction formulation is at least 1 mM. In another embodiment, the hydrogen peroxide concentration in the aqueous reaction formulation is at least 10 mM. In another embodiment, the hydrogen peroxide concentration in the aqueous reaction formulation is at least 100 mM. In another embodiment, the hydrogen peroxide concentration in the aqueous reaction formulation is at least 200 mM. In another embodiment, the hydrogen peroxide concentration in the aqueous reaction formulation is 500 mM or more. There are still other specific examples in which the hydrogen peroxide concentration in the aqueous reaction formulation is 1000 mM or more. In the formulation, the molar ratio of hydrogen peroxide to the enzyme substrate, such as triglyceride, (H ₂ O ₂ :matrix) may be from about 0.002 to 20, preferably from about 0.1 to 10, and most preferably about 0.5 to 5.

As used herein, the term "oligosaccharide" refers to a monosaccharide compound containing between 2 and at least 24 linked by a glycosidic linkage. The term "monosaccharide" refers to a compound of the formula (CH ₂ O) _n wherein n 3. The carbon skeleton is unbranched, except for one, each carbon atom contains a hydroxyl group, and the remaining carbon atoms are aldehydes or ketones at carbon atom 1. The term "monosaccharide" also refers to the form of a cyclic hemiacetal or hemi-ketal in the cell.

As used herein, the term "hydratable adhesive" shall mean a hydratable adhesive material. The hydratable adhesive is substantially dry and non-sticky until hydrated. Upon hydration, the hydratable adhesive becomes sufficiently viscous to attach the tooth whitening patches/film to the tooth surface. The hydratable adhesive film also includes a particulate bleaching component which upon hydration releases an effective amount of hydrogen peroxide for enzymatic formation of a peroxyacid bleach. The whitening patch/film is typically thin (typically less than 2 mm), shaped and sized to fit inside the mouth, and sufficiently flexible to allow the film to be applied and placed in contact with a plurality of teeth whereby the hydration The adhesive helps to attach the film/dental to the tooth surface and provides sufficient time for the peroxyacid bleach to whiten the teeth.

As used herein, the term "effective amount" shall mean the amount of material required to achieve the desired effect.

As used herein, the term "substantially non-adhesive until hydrated" shall mean a lack of adhesion strength sufficient to adhere the tooth whitening film to the surface of the tooth prior to hydration. Thus, the hydratable adhesive film will be easy to handle and handle prior to application and hydration by the user.

By "sequence homology" is meant the amino acid sequence homology using a sequence alignment program, eg, ClustalW or BLAST, as generally described in Ochu SF, Gem W, Miller W, Myers EW, Lippmann DJ, "Basic Local Alignment Search Tool", Journal of Molecular Biology (1990) 215(3): 403-410, and Gu Yi M, Mai William H, Li W, Valentin F, History Sato S, Pylon J, Lopez R, Nucleic Acids Research (2010) 38 Supplement: W695-9.

The thiol donor used in the present invention, for example, in the reaction with a peroxide to form a peroxyacid, is selected from one or more (i) C _2-18 carboxylic acids, such as C _2-6 carboxylic acid. An acid (eg, acetic acid), including a lower linear or branched alkyl carboxylic acid, optionally substituted with a hydroxy group and/or a C _1-4 alkoxy group; (ii) a hydrolyzable and acceptable ester thereof (eg Mono-, di-, and tri-glycerides and deuterated sugars) and (iii) mixtures thereof. For example, sulfhydryl donors include 1,2,3-triethoxypropane (sometimes referred to herein as triacetin or triacetin) and deuterated sugars, such as acetylated saccharides. In a particular embodiment, the ester in this use can be, for example, an ester having a water solubility of at least 5 ppm at 25 °C.

The thiol donor and/or enzyme may be optionally encapsulated. Various methods of encapsulation are well known in the art, both natural and synthetic. Modified starches and gum arabic are particularly suitable because they are food grade, relatively inexpensive, rapidly soluble, and capable of adsorbing a relatively high degree of liquid oil. Any impact on the final viscosity needs to be considered.

In some embodiments, the particles include an anti-allergic agent that can desensitize or block dentinal tubules. In some embodiments, the anti-allergic agent is selected from the group consisting of a potassium ion source, a citrate, and a stannous ion. Source, basic amino acid, clay, and combinations thereof. In some embodiments, the potassium ion source is an orally acceptable potassium salt and is present in an amount effective to reduce tooth sensitivity. In some embodiments, the potassium ion source is selected from the group consisting of potassium chloride, potassium nitrate, and combinations thereof. In some embodiments, the basic amino acid is arginine. In some embodiments, the basic amino acid is selected from the group consisting of arginine phosphate, arginine hydrogencarbonate, and arginine hydrochloride. In some embodiments, the citrate is calcium citrate.

CE-7 perhydrolase

The compositions and methods of the present invention include an enzyme having perhydrolysis activity which is structurally classified as a member of the esterase family 7 (CE-7 family) of a carbohydrate family enzyme (see Coutinho, PM, Hennessa, B). "Carbohydrate Active Enzymes: An Integrated Database Approach" Recent Developments in Carbo Bioengineering, HJ Gilbert, G. Davis, B. Hennessa and B. Svensson, (1999) UK Royal Society of Chemistry, Cambridge, pp. 3-12). The CE-7 family of enzymes has been shown to be particularly effective in producing peroxyacids from a variety of carboxylate matrices when combined with a source of peroxides (U.S. Patent Nos. 7,794,378; 7,951,566; 7,723,083; and 7,964,378 and U.S. Patent Application Publication No. 2008- 0176299, 2010-0087529, 2011-0081693, and 2011-0236335 - Dicosimo, et al; each incorporated herein by reference.

Members of the CE-7 family include cephalosporin C deacetylase (CAHs; E.C. 3.1.1.41) and acetyl xylan esterase (AXEs; E.C. 3.1.1.72). Members of the CE-7 esterase family share a conserved label motif (Wenson et al., J. Mol. Biol. 330: 593-606 (2003)). Perhydrolase enzymes include CE-7 tag motifs ("CE-7 perhydrolase") and/or substantially similar structures suitable for use in the compositions and methods described herein. Methods for identifying substantially similar biomolecules are well known in the art (e.g., sequence alignment protocols, nucleic acid hybridization, and/or the presence of a conserved label motif). In one aspect, the perhydrolase comprises an enzyme comprising a CE-7 tag motif and is at least 20%, preferably at least 30%, more preferably at least 33%, more preferably at least 40%, more preferably at least 42%, more preferably at least 50. %, more preferably at least 60%, more preferably at least 70%, More preferably at least 80%, more preferably at least 90%, and most preferably at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% amino acid sequence Same as one of the sequences provided herein.

As used herein, the phrase "the enzyme is structurally classified as CE-7 enzyme", "CE-7 perhydrolase" or "structurally classified as carbohydrate esterase family 7 enzyme" is used to refer to Hydrolytic activity, an enzyme structurally classified as CE-7 carbohydrate esterase. This family of enzymes can be defined by the presence of a tag motif (Wenson et al., supra). The CE-7 esterase tag motif includes three conserved motifs (numbers of residue positions are related to the reference sequence EQ ID NO: 1; Haiqi hot robe C277S variant perhydrolase). Arg118-Gly119-Gln120; Gly186-Xaa187- Ser188- Gln189-Gly190; and His303- Glu304.

Typically, Xaa at position 187 of the amino acid residue is glycine, alanine, valine, tryptophan, or threonine. Two of the three amino acid residues belonging to the catalytic triad are in bold. In one embodiment, the Xaa at position 187 of the amino acid residue is selected from the group consisting of glycine, alanine, valine, tryptophan, and threonine.

Further analysis of the conserved motif in the CE-7 carbohydrate esterase family revealed the presence of other conserved motifs (LXD at amino acid positions 272-274 of SEQ ID NO: 1), which can be used to further define CE-7 a perhydrolase of the carbohydrate esterase family. In other embodiments, a tag motif as defined above may include other (fourth) conserved motifs as defined below: Leu272-Xaa273- Asp274 . Xaa at position 273 of the amino acid residue is typically isoleucine, valine, or methionine. The fourth region includes an aspartic acid residue (bold) belonging to the catalytic triad ( Ser188-Asp274-His303 ).

The CE-7 perhydrolase may be a fusion protein version having at least one peptide component that has an affinity for at least one body surface. In a specific example, all used to confirm whether the target is The perhydrolase (fusion protein) comprising the CE-7 tag motif will be based on an amino acid sequence of a perhydrolase that does not contain a peptide component that has an affinity for the body surface.

Many world-renowned alignment algorithms (i.e., sequence analysis software) can be used to align two or more amino acid sequences of enzymes representing perhydrolase activity to determine if the enzyme comprises a tag motif of the invention. The aligned sequence is compared to a reference sequence (SEQ ID NO: 1) to determine the presence of the tag motif. In one embodiment, CLUSTAL alignment (eg, CLUSTALW) using a reference amino acid sequence (such as the perhydrolase sequence (SEQ ID NO: 1) used herein) is used to confirm that it belongs to the CE-7 esterase family. Hydrolase. The relevant numbering of the conserved amino acid residues is based on the numbering of the residues numbered in the reference amino acid sequence to identify insertions or deletions in the aligned sequences (e.g., typically 5 amino acids or less).

Examples of other suitable algorithms can be used to confirm that a sequence comprising a tag motif of the invention (when compared to a reference sequence) includes, but is not limited to, Nederman and Vaughan (Journal of Molecular Biology 48, 443-453 (1970) ); a global comparison tool) and Smith-Watman (Journal of Molecular Biology 147:195-197 (1981); local comparison tool). In one embodiment, the Smith-Wattman alignment is implemented using default parameters. Examples of suitable initial parameters include the use of the BLOSUM6 scoring matrix with a GAP open penalty = 10 and a GAP extension penalty = 0.5.

In one embodiment, a suitable perhydrolase, including an enzyme, comprising a CE-7 tag motif and at least 20%, preferably at least 30%, 33%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% amino acid sequence is identical to SEQ ID NO: 1.

Examples of suitable CE-7 carbohydrate esterases having perhydrolysis activity include, but are not limited to, enzymes having amino acid sequences such as SEQ ID NOs: 1, and 4-28. In one embodiment, the enzyme comprises an amino acid sequence selected from the group consisting of 1, 10, 11, 15, and 16.

As used herein, the terms "CE-7 variant", "variant perhydrolase" or "variant" refers to a genetically engineered CE-7 perhydrolase that causes at least one amino acid to increase, decrease, and / or substitution, when the corresponding enzyme (typically wild-type enzyme) is compared to the derived variant; as long as the CE-7 tag motif and associated perhydrolysis activity are maintained. CE-7 variant perhydrolase enzymes can also be used in the compositions and methods of the present invention. Examples of CE-7 variants are provided as SEQ ID NOs: 1, 15, 16, 17, 18, 19, and 20. In one embodiment, the variant can include SEQ ID NOs: 1 and 16.

The skilled artisan will be aware that substantially similar CE-7 perhydrolase sequences (retained label motifs) can also be used in the compositions and methods of the present invention. In one embodiment, substantially similar sequences are defined by their ability to hybridize, under very stringent conditions, to nucleic acid molecules associated with the sequences exemplified herein. In another embodiment, the sequence alignment algorithm can define substantially similar enzymes based on the percentage of DNA or amino acid sequences provided by the present invention.

As used herein, a nucleic acid molecule "can hybridize" to another nucleic acid molecule, such as cDNA, genotype DNA, or RNA, when the single strand of the first molecule can be renatured at an appropriate temperature and ionic strength of the solution. When it comes to another molecule. Hybridization and washing conditions are well known and exemplified in Sambrook, J. and Russell, D., T. Molecular Cloning: A Laboratory Manual, 3rd Edition, Cold Spring Harbor Laboratory, Cold Spring Harbor (2001). The conditions of temperature and ionic strength determine the "stringency" of hybridization. Stringent conditions can be adjusted to screen for moderately similar molecules, such as homologous sequences from organisms that are distantly related, to very similar molecules, such as genes that replicate functional enzymes from closely related organisms. Post-hybrid cleaning typically determines stringent conditions. The preferred conditions are set to use a series of cleanings. Initially, 6X SSC, 0.5% SDS was used at room temperature for 15 minutes, then repeated with 2X SSC, 0.5% SDS at 45 °C for 30 minutes, and then repeated at 50 °C for 30 minutes with 0.2X SSC, 0.5% SDS. The preferred conditions of use are set to a higher temperature, wherein the cleaning is the same as above except that in the last two 30 minutes at 0.2X SSC, the cleaning temperature of 0.5% SDS is increased to 60 °C. Another comparison The extremely stringent hybridization conditions were set to 0.1X SSC, 0.1% SDS, 65 ° C and cleaned with 2X SSC, 0.1% SDS followed by 0.1X SSC, 0.1% SDS, and 65 °C.

Hybridization requires that two nucleic acids contain complementary sequences, although mismatching between bases may occur depending on the stringency of the hybridization. The appropriate stringency of hybrid nucleic acids is based on the length and complementarity of the nucleic acids, and variations thereof are well known to those skilled in the art. The greater the similarity or homology between the two nucleotide sequences, the greater the hybridization Tm value of the nucleic acid having the sequences. The relative stability of nucleic acid hybridization (corresponding to higher Tm) is reduced in the following order: RNA: RNA, DNA: RNA, DNA: DNA. Hybrids of nucleotides greater than 100 in length, derived from the computational system Tm (Sambrook and Russell, as before). When hybridizing to shorter nucleic acids, i.e., oligonucleotides, the location of mismatches becomes more important, and the length of the oligonucleotide determines its specificity (Sambrooke and Russell, as before). In one aspect, the hybridizable nucleic acid is at least about 10 nucleotides in length. Preferably, the hybrid nucleic acid has a minimum length of at least about 15 nucleotides, more preferably at least about 20 nucleotides in length, more preferably at least 30 nucleotides in length, and more preferably at least 300 nucleosides in length. The acid is preferably at least 800 nucleotides in length. Furthermore, those skilled in the art will know that the temperature and the salt concentration of the cleaning solution can be adjusted according to factors such as the length of the probe as needed.

As used herein, the term "percent identity" is the relationship between two or more polypeptide sequences or two or more polynucleotide sequences, as determined by comparing the sequences. In the art, "identity" also refers to the degree of sequence association between polypeptide or polynucleotide sequences, as determined by the match between strings of such sequences. "Identity" and "similarity" can be easily calculated by known methods, including but not limited to those of the following: Computational Molecular Biology (Lisk, AM Editor) Oxford University Press, New York (1988); Biocomputing: Informatics and Genomics Project (Smith, DW, ed.) Academic Publishing, New York (1993); Computer Analysis of Sequence Data, Part I (Griffin, AM, and Griffin, HG, Editor) Shuman Inc., New Jersey (1994); Sequence Analysis of Molecular Biology (Feng Heine, G. Editor) Academic Publication (1987); and Introduction to Sequence Analysis (Jibikov, M. and Dick, J. Editor) Stockton Publishing, New York State (1991). Recognition and similar validation methods are compiled in publicly available computer programs. The alignment of the alignment and the percentage of identity can be calculated using the MegAlign program sequence of the LASERGENE bioinformatics computing group (DNASTAR, Madison, West Indiana), the AlignX program of Vector NTI v.7.0 (Informax, Bay Sesda, Maryland), or EMBOSS Open Software Suite (EMBL-EBI; Reith et al., Genetics Trends 16, (6): 276-277 (2000)). Multiple alignment of sequences can be performed using the following CLUSTAL alignment methods (eg CLUSTALW; eg version 1.83) (Higgins and Sharp, CABIOS, 5: 151-153 (1989); Higgins et al., Nucleic Acids Research. 22: 4673-4680 (1994); and Zinano et al., Nucleic Acids Research 31 (13): 3497-500 (2003)), available from the European Molecular Biology Laboratory via the European Institute for Bioinformatics, with initial parameters. Suitable parameters for CLUSTALW protein alignment include GAP extension penalty = 15, GAP extension = 0.2, matrix = Gonnet (eg, Gonnet 250), protein ENDGAP = -1, protein GAPDIST = 4, and KTUPLE = 1. In one embodiment, the initial setting is used for fast or slow comparisons, with slower alignment being preferred. Alternatively, parameters using the CLUSTALW method (eg, version 1.83) can also be modified to use KTUPLE=1, GAP penalty=10, GAP extension=1, matrix=BLOSUM (eg, BLOSUM64), WINDOW=5, and TOP DIAGONALS SAVED =5.

In one aspect, the suitably isolated nucleic acid molecule encodes at least about 20%, preferably at least 30%, 33%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%. , 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the polypeptide of the amino acid sequence identical to the amino acid sequence reported herein. In another aspect, the suitably isolated nucleic acid molecule encodes at least about 20%, preferably at least 30%, 33%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91. %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the polypeptide of the amino acid sequence identical to the amino acid sequence reported herein. Suitable nucleic acid molecules not only have the above homologs, but also typically encode polypeptides having a length of from about 210 to 340 amino acids, from about 300 to about 340 The amino acid is preferably from about 310 to about 330 amino acids, and most preferably from about 318 to about 325 amino acid lengths wherein each polypeptide is characterized by perhydrolysis activity.

Over-hydrolase

As used herein, the terms "target perhydrolase" and "enzymatically active enzyme" refer to a fusion protein comprising at least one perhydrolase (wild type or variant thereof) fused/coupled to at least one For the target surface, it is advisable to have a peptide component with affinity for the target body surface. The perhydrolase in the target perhydrolase can be any CE-7 carbohydrate esterase having perhydrolysis activity. The CE-7 perhydrolase can be confirmed by the presence of a CE-7 tag motif, which is compared to the reference sequence EQ ID NO: 1, which includes: i) the RGQ motif corresponding to the SEQ ID NO: position 1 position 118-120; ii) position of GXSQG motif corresponding to position 186-190 of SEQ ID NO: 1; and iii) HE motif corresponding to position 303-304 of SEQ ID NO: The location.

In one embodiment, the perhydrolase may be at least about 20%, preferably at least 30%, 33%, 40%, 50%, 60%, 70%, 80%, 85% of those amino acid sequences. , 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% similar to any of the amino acid sequences reported herein (ie, SEQ ID NOs: 1, and 4-28).

In another embodiment, the fusion protein comprises a perhydrolase having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1, and 4-28.

As used herein, "peptide composition", "peptide composition having affinity for the oral surface", "oral binding structure region", and "OCBD" refer to a fusion protein component, which is A portion of a perhydrolase (including at least one polymer of two or more amino acids joined by a peptide bond); wherein the component has an affinity for the surface of the target oral cavity. In a preferred aspect, the OCBD has an affinity for tooth enamel.

In one embodiment, the peptide component having affinity for the body surface can be an antibody, a Fab antibody fragment, a single-chain variable fragment (scFv) antibody, a Camelidae antibody (Mudeman, S., molecule) Bioengineering Review (2001) 74:277-302), a non-antibody scaffold display protein (Hors et al., Protein Science (2006) 15(1): 14-27 and Bins, H. et al. 2005) Natural Biotechnology 23, 1257-1268 for a review of various scaffold-assisted approaches) or a single-chain polypeptide lacking an immunoglobulin population. On the other hand, the peptide component having affinity for oral tissues/surfaces (e.g., tooth enamel) is a single-stranded peptide lacking an immunoglobulin group.

The peptide component having affinity for the oral surface can be separated by a perhydrolase by any peptide linker. The particular peptide linker/spacer region is from 1 to 100 or from 1 to 50 amino acid lengths. In certain embodiments, the peptide spacer sequence region is from about 1 to about 25, from 3 to about 40, or from 3 to about 30 amino acid lengths. In other embodiments, the spacer sequence region is from about 5 to about 20 amino acid lengths. Multiple peptide linkers can be used. In one embodiment, at least one peptide linker is present and can be repeated up to 10 fold.

In one embodiment, the fusion peptide comprises at least one oral surface-binding peptide selected from the group consisting of SEQ ID NOs: 178-197.

In another embodiment, the target surface material is part of a package, such as a whitening or polymeric underlayer (when a polymeric primer layer is used to apply a hydratable adhesive thereto) and/or to a mouth. . The peptide component is selected for use as a substance such as a polymer, a plastic, and a film due to its affinity. The over-hydrolase fusion protein design of the subject allows control of the perhydrolase to be delivered or removed by the user by retaining it on a removable device such as, but not limited to, a dial or a patch.

The peptide component having affinity for the oral surface can be isolated by CE-7 perhydrolase by any peptide linker. Specific peptide linker/spacer sequence range from 1 to 100 Or 1 to 50 amino acid lengths. In certain embodiments, the peptide spacer sequence region is from about 1 to about 25, from 3 to about 40, or from 3 to about 30 amino acid lengths. In other embodiments, the spacer sequence region is from about 5 to about 20 amino acid lengths. Multiple peptide linkers can be used. Examples of peptide linkers are provided as SEQ ID NOs: 164-177.

Thus, examples of the subject CE-7 perhydrolase can include, but are not limited to, any CE-7 perhydrolase having an amino acid sequence selected from the group consisting of SEQ ID NOs 1, and 4-28 to The surface of the oral cavity has a peptide component of affinity. In a preferred embodiment, examples of the subject perhydrolase may include, but are not limited to, any CE-7 perhydrolase having an amino acid sequence selected from the group consisting of SEQ ID NOs 1, 4, 5, and 6. , 7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23,24,25,26,27, and 28, which are coupled to One or more body surface-affinity peptides that have affinity for the oral surface (arbitrary via the peptide spacer sequence region). In a preferred embodiment, the subject perhydrolase comprises a CE-7 perhydrolase having an amino acid sequence selected from the group consisting of SEQ ID NOs: 1 and 16.

In one embodiment, the perhydrolase is a CE-7 perhydrolase in the form of a fusion protein of the general structural formula: PAH-[L]y-OCBD or OCBD-[L]y-PAH wherein PAH is An enzyme having perhydrolysis activity, for example, having a CE-7 tag motif, such as SEQ ID NO: 1, and OCBD is a peptide component having affinity for the oral surface; and L is an arbitrary linker; and y is An integer from 0 to 10. In one embodiment, the linker (L) is present and is a peptide linker ranging from 1 to 100 amino acid lengths.

For example, SEQ ID NO: 2 is a fusion protein having a perhydrolase sequence SEQ ID NO: 1 coupled to the C-terminus of a structural region targeted for oral tissue.

The perhydrolase enzymes used in the products and methods of the invention may be free, protected (e.g., acetamidine), or salted.

In another embodiment, the target surface material is part of the package and/or delivered to the oral cavity. The peptide component is selected for use as a substance such as a polymer, a plastic, and a film due to its affinity. The subject CE-7 perhydrolase fusion protein design allows control of the perhydrolase to be delivered or removed by the user by retaining it on a removable device, such as a dial or a patch.

Affinity

The peptide component having affinity for the oral surface includes a binding affinity of 10 ^-5 moles (M) or less to the oral surface. In a particular embodiment, the peptide component is one or more oral surface-binding peptides and/or binding structural regions having a binding affinity for tooth enamel of 10 ^-5 moles (M) or less. In certain embodiments, the binding peptide or structural region has a binding affinity value of 10 ^-5 M or less in the presence of at least about 50-500 mM salt. The term "binding affinity" refers to the strength of the binding of a peptide to its interaction with a corresponding substrate. Binding affinity of the peptide solution can be defined by measuring the binding or dissociation constant ( "K _D"), or "MB _50" of the word.

"K _D " corresponds to the concentration of the peptide, wherein the binding site on the target is occupied by half, that is, when the concentration of the target containing the peptide bond (bonding target substance) and the peptide-free bond When the target concentration is the same. The smaller the dissociation constant, the tighter the peptide is tied. For example, a peptide with a nanomolar (nM) dissociation constant is tighter than a peptide with a dissociation constant of micromolecules (μM). A specific embodiment of the present invention has a K _D value of 10 ^-5 or less.

Signal "MB _50" refers to the combination of the peptide to which it is given to ELISA- 50% concentration maximum combined analysis of the signals obtained it. See, for example, Example 3 of U.S. Patent Application Publication No. 2005/022683, incorporated herein by reference. The MB ₅₀ line provides an indication of the strength of the binding interaction or affinity of the complex components. The lower the MB ₅₀ value, the stronger the interaction of the peptide with its corresponding substrate, that is, "the better." For example, a nanogram (nM) MB ₅₀ peptide is more tightly bound than a microgram (μM) MB ₅₀ peptide. A specific embodiment of the present invention has an MB ₅₀ value of 10 ^-5 M or less.

In certain embodiments, the binding affinity have affinity for peptides consisting of the parts for the oral surface, such as by ₅₀ MB or K _D measurement values, less than or equal to about 10 ^-5 M, less than or equal About 10 ^-6 M, less than or equal to about 10 ^-7 M, less than or equal to about 10 ^-8 M, less than or equal to about 10 ^-9 M, or less than or equal to about 10 ^-10 M.

In certain embodiments, the oral cavity surface - bound peptides and / or oral cavity surface - binding affinity binding zone structure, such as by ₅₀ MB or K _D measurement values, less than or equal to about 10 ^-5 M, Less than or equal to about 10 ^-6 M, less than or equal to about 10 ^-7 M, less than or equal to about 10 ^-8 M, less than or equal to about 10 ^-9 M, or less than or equal to about 10 ^-10 M .

As used herein, the term "strong affinity" means a binding affinity having a K _D or MB ₅₀ value of less than or equal to about 10 ^-5 M, preferably less than or equal to about 10 ^-6 M, more preferably less than Or equal to about 10 ^-7 M, more preferably less than or equal to about 10 ^-8 M, less than or equal to about 10 ^-9 M, or most preferably less than or equal to about 10 ^-10 M.

Enzyme powder

In some embodiments, the personal care composition can use an enzyme catalyst that stabilizes the enzyme powder form. A method of making and stabilizing a formulation comprising an enzyme powder is described in U.S. Patent Application Publication Nos. 2010-0086534 and 2010-0086535.

In one embodiment, the enzyme may be present in the enzyme powder in a range from about 0.5 weight percent (wt%) to about 75 wt%, such as from 1 wt% to about 60 wt%, based on the dry weight of the enzyme powder. Preferably, the enzyme is present in the enzyme powder/spray dry mixture in a weight percentage ranging from about 10% to 50% by weight, and the enzyme is preferably in an enzyme powder/spray dry mixture in a weight percent range. 20% by weight to 33% by weight.

In one embodiment, the enzyme powder may further comprise an excipient. In one aspect, the amount of excipient is provided in the range of from about 95% to about 25% by weight, based on the dry weight of the enzyme powder. The preferred weight percent of the excipient in the enzyme powder ranges from about 90% to 50% by weight, and more preferably the excipient in the enzyme powder ranges from about 80% to about 67% by weight.

In one embodiment, the excipient used to prepare the enzyme powder can be an oligosaccharide excipient. In one embodiment, the oligosaccharide excipient has an average molecular weight of at least about 1250 and an average molecular weight of at least about 9000. In some embodiments, the oligosaccharide excipient has an average molecular weight of at least about 1700 and an average molecular weight of at least about 15,000. Specific oligosaccharides may include, but are not limited to, maltodextrin, xylan, mannan, fucoidan sulfate, galactomannan, chitosan, raffinose, stachyose, fruit Gum, insulin, fructan, melin fructan, amyloid pectin, sucrose, galactose, lactose, maltose, trehalose, cellobiose, Aspergillus trisaccharide, maltotriose, pine Sugar, maltotriulose, raffinose, canetriose, and mixtures thereof. In a preferred embodiment, the oligosaccharide excipient is maltodextrin. The oligosaccharide-based excipients may also include, but are not limited to, water-soluble nonionic cellulose ethers such as hydroxymethyl-cellulose and hydroxypropylmethylcellulose, and mixtures thereof. In still other embodiments, the excipient may be selected from, but not limited to, one or more of the following compounds: trehalose, lactose, sucrose, mannitol, sorbitol, glucose, cellobiose, alpha-cyclodextrin Fine, and carboxymethyl cellulose.

Appropriate ester matrix / sulfhydryl donor

Suitable carboxylate matrices may include esters of the formula: (a) one or more esters of the formula [X] _m R ₅ wherein X is an ester group of the formula R ₆ C(O)O; R ₆ is a C1 to C7 linear, branched or cyclic hydrocarbyl moiety, optionally substituted by a hydroxy group or a C1 to C4 alkoxy group, wherein R ₆ optionally includes one or more when R ₆ is C2 to C7 Ether-bonded; R ₅ is a C1 to C6 linear, branched, or cyclic hydrocarbyl moiety or a five-membered cyclic heteroaryl moiety or a six-membered cyclic aryl or heteroaryl moiety, optionally substituted by a hydroxy group Substituent; wherein each of the carbon atoms in R ₅ includes no more than one hydroxyl group or no more than one ester group or a carboxylic acid group, and wherein R ₅ optionally includes one or more ether linkages; m is an integer from 1 to R ₅ in the number of carbon atoms, the one or more esters having a water solubility of at least 5 ppm at 25 ° C; or (b) one or more glycerides having the following structural formula Wherein R ₁ is a C1 to C7 straight or branched alkyl group, which is optionally substituted by a hydroxy group or a C1 to C4 alkoxy group and R ₃ and R ₄ are independently H or R ₁ C(O); or (c One or more esters of the formula Wherein R ₁ is a C1 to C7 straight or branched alkyl group, which is optionally substituted by a hydroxy group or a C1 to C4 alkoxy group and R ₂ is a C1 to C10 linear or branched alkyl group, an alkenyl group, an alkynyl group, and an aromatic group. Alkyl, alkylaryl, alkylheteroaryl, heteroaryl, (CH ₂ CH ₂ O) _n , or (CH ₂ CH(CH ₃ )-O) _n H and n is from 1 to 10; or (d One or more acetylated monosaccharides, acetylated disaccharides, or acetaminophen; or (e) any combination of (a) to (d).

Suitable matrices may also include one or more deuterated sugars selected from the group consisting of acetylated mono-, di-, and polysaccharides. In another embodiment, the deuterated sugar is selected from the group consisting of: acetylated xylan; acetylated xylan fragment; acetylated xylose (such as xylose tetraacetic acid); Glucose (such as α-D-glucose pentaacetate; β-D-glucose pentaacetate; 1-thio-β-D-glucose-2,3,4,6-tetraacetate); β-D-half Lactose pentaacetate; sorbitol pentaacetate; sucrose octaacetate; β-D-ribofuranosyl-1,2,3,5-tetraacetate; β-D-ribofuranosyl-1,2,3,4 -tetraacetate; tris-O-acetamidine-D-galactose; tris-O-acetamidine-D-glucoenose; β-D-furanosyltetraacetate, β-D-glucopyranose pentaacetic acid Ester; β-D-glucopyranose-1,2,3,4-tetraacetate; β-D-glucopyranose-2,3,4,6-tetraacetate; 2-acetamido-2 -deoxy-1,3,4,6-tetraacetyl-β-D-glucopyranose; 2-acetamido-2-deoxy-3,4,6-triethyl fluorene-1-chloro- α-D-glucopyranose; β-D-mannopyranomannose pentaacetate, and acetylated cellulose. In a preferred embodiment, the acetylated sugar is selected from the group consisting of β-D-ribofuranosyl-1,2,3,5-tetraacetate; tris-O-acetyl-D-galactose; Tri-O-acetam-D-glucoenose; sucrose octaacetate; and acetylated cellulose.

In another embodiment, other suitable substrates may also include 5-ethoxymethyl-2-furaldehyde; 3,4-diethoxy-1-butene; 4-ethyloxybenzoic acid Vanillin acetic acid; propylene glycol methyl ether acetate; methyl lactic acid; ethyl lactic acid; methyl alcohol; ethyl alcohol; methyl methoxy acetate; ethyl methoxy acetate; 3-hydroxybutyrate; ethyl 3-hydroxybutyrate; and triethyl 2-acetamidine citrate.

In another embodiment, a suitable matrix is selected from the group consisting of: glyceryl monoacetate; glyceryl diacetate; glyceryl triacetate; glycerol monopropionate; glyceryl dipropionate; glyceryl tripropionate; Glyceryl butyrate; glyceryl dibutyrate; glyceryl tributate; glucose pentaacetate; xylose tetraacetic acid; acetylated xylan; acetylated xylan fragment; β-D-ribofuranose-1 , 2,3,5-tetraacetate; tris-O-acetamidine-D-galactose; tris-O-acetyl-D-glucoenose; 1,2-ethanediol; 1,2-propane Alcohol; 1,3-propanediol; 1,2-butanediol; 1,3-butanediol; 2,3-butanediol; 1,4-butanediol; 1,2-pentanediol; 5-pentanediol; 1,5-pentanediol; 1,6-pentanediol; 1,2-hexanediol; 2,5-hexanediol; monoester of 1,6-hexanediol or Diesters; and mixtures thereof. In another embodiment, the substrate is a C1 to C6 polyol comprising one or more ester groups. In a preferred embodiment, one or more of the hydroxyl groups on the C1 to C6 polyol are one or more ethoxylated groups (eg, 1,3-propanediol diacetate; 1,2-propanediol). Diacetate; 1,4-butanediol diacetate; 1,5-pentanediol diacetate, etc., substituted. In another embodiment, the substrate is propylene glycol diacetate (PGDA), vinyl glycol diacetate (EGDA), or a mixture thereof.

In another embodiment, a suitable matrix is selected from the group consisting of: monoacetin, diacetin, triacetin, glycerol monopropionate, dipropionate, tripropionate, single Butyric acid glyceride, dibutyric acid glyceride, and tributyrin. In still another aspect, the matrix is selected from the group consisting of diacetin and triacetin. In a preferred embodiment, the suitable substrate comprises triacetin.

In the case of an enzyme-catalyzed perhydrolysis reaction, the carboxylic acid ester is present in a concentration sufficient to produce the desired concentration of peroxyacid. The carboxylic acid ester does not need to be completely dissolved in the reaction formulation, but the solubility must be sufficient to allow the conversion of the ester to the corresponding peroxyacid by the perhydrolase catalyst. The carboxylic acid ester is present in the reaction formulation at a concentration of from 0.05% by weight to 40% by weight of the reaction formulation, preferably from 0.1% to 20% by weight of the reaction formulation, and more preferably from 0.5% by weight of the reaction formulation to A concentration of 10% by weight.

The peroxide source is provided as particles deposited in or on the hydratable adhesive film and may include a hydrogen peroxide adduct (eg, urea (urea)-hydrogen peroxide adduct (carbamide peroxidation) ()) perborate, percarbonate and peroxide salts. The concentration of the peroxide compound may range from 0.0033 wt% to about 50 wt%, more preferably from 0.033 wt% to about 40 wt%, and more preferably from 0.1 wt% to about 30 wt%, of the reaction formulation. %.

Many perhydrolase catalysts (whole cells, permeabilized whole cells, and partially purified whole cell extracts) have been reported to have catalase activity (EC 1.11.1.6). Catalase is a reaction that catalyzes the conversion of hydrogen peroxide to oxygen and water. In one aspect, the perhydrolysis catalyst lacks catalase activity. On the other hand, a catalase inhibitor can be added to the reaction formulation. In this regard, the skilled artisan can adjust the concentration of the catalase inhibitor as needed. The catalase inhibitor concentration typically ranges from 0.1 mM to about 1 M; preferably from about 1 mM to about 50 mM; more preferably from about 1 mM to about 20 mM.

In another embodiment, the enzyme catalyst lacks significant catalase activity or can be designed to reduce or eliminate catalase activity. The catalase activity in the host cell can be down-regulated or eliminated by disrupting gene expression in response to catalase activity using well-known techniques including, but not limited to, transposon mutations, RNA Antisense expression, target mutations, and random mutations.

The peroxyacid concentration (e.g., peracetic acid) produced by the perhydrolysis reaction of at least one carboxylic acid ester is at least about 0.1 ppm, preferably at least 0.5 ppm, 1 ppm, 5 ppm, 10 ppm, 20 ppm, 100 ppm, 200 ppm, 300 ppm, 500 ppm, 700 ppm, 1000 ppm, 2000 ppm, 5000 ppm or 10,000 ppm of peroxyacid is preferably within 5 minutes from the start of the hydrolysis reaction within 10 minutes. It will be apparent that those skilled in the art can adjust the composition of the reaction to achieve the desired peroxyacid concentration.

In one aspect, the reaction time required to produce the desired peroxyacid concentration is no more than about 2 hours, preferably no more than about 30 minutes, more preferably no more than about 10 minutes, and most preferably about 5 minutes or less. In other aspects, the oral surface is contacted with a peroxycarboxylic acid formed by combining water with the reactive components in 5 minutes according to the methods described herein. In one embodiment, the dental tannin is contacted with a peroxycarboxylic acid formed according to the methods and compositions described herein, at about 5 minutes. The clock is combined for about 24 hours or about 5 minutes to 2 hours (via hydration by the user) when the reaction component appears on or in the whitening toothpaste/film.

HPLC analysis of peroxyacid and hydrogen peroxide concentrations

A variety of analytical methods can be used to analyze the reactants and products, including, but not limited to, titration, high performance liquid chromatography (HPLC), gas chromatography (GC), mass spectrometry (MS), Capillary Electrophoresis (CE), Analytical Processes are described in U.K. Kenny et al. (Analytical Chemistry, 69(17): 3623-3627 (1997)), and 2,2'-azide-bis (3- Analysis of ethylbenzothiazoline-6-sulfonate (ABTS) (U.F. Kenny et al., 122:567-571 (1997) and Danu et al., 20:392-398 ( 2010)), as explained in the example of this case.

Specific examples

Specific Example 1. A tooth whitening toothpaste (dental patch 1) comprising a hydratable adhesive film having a first side and a second side, the first side comprising a particulate bleaching component attached thereto, wherein the dental patch comprises a film The particles are attached to the first side of the membrane or thereon, (i) a protein having perhydrolase activity, which contains a tag motif of a member of the carbohydrate esterase family 7; (ii) a thiol donor, eg, It is selected from the group consisting of carboxylates and mercapto compounds.

Specific Example 2. Dental patch 1, wherein the protein having perhydrolase activity comprises an amino acid sequence selected from the group consisting of: a) (SEQ ID NO: 1),

b) an amino acid sequence having i) a RGQ motif corresponding to position 118-120 of SEQ ID NO: 1; ii) a GXSQG motif corresponding to position 186-190 of SEQ ID NO: And iii) the HE motif at a position corresponding to positions 303-304 of SEQ ID NO: 1; and c) an amino acid sequence having at least 80% sequence identical to SEQ ID NO: 1.

Specific Example 3. Specific example 1 or 2, wherein the hydrolyzed protein comprises an amino acid sequence which has affinity for the oral surface, such as in combination with or complex with it, or for the composition of one or more whitening patches Parts have affinity, such as in combination with or complex with them.

Specific Example 4. Any of the foregoing dental patches comprising a protein having perhydrolase activity which binds to or complexes with the surface of the oral cavity (e.g., a dental film or enamel) comprising an amino acid sequence selected from the group consisting of a) (SEQ ID NO: 2), and b) an amino acid sequence having at least 80% sequence identical to SEQ ID NO: 2.

Specific Example 5. Any of the foregoing dental patches, wherein the hydrolytically active protein is provided in a particle form on the first surface of the film.

Specific Example 6. Any of the foregoing dental patches, wherein the second side of the hydratable adhesive film is attached to a layer which inhibits dissolution of the hydratable adhesive film.

Specific Example 7. Any of the foregoing dental patches wherein the particulate bleaching component is embedded with a rapidly dissolvable material such as corn starch, sodium alginate, and combinations thereof.

Specific Example 8. Any of the foregoing dental patches, wherein the particulate bleaching component is selected from the group consisting of organic and/or inorganic oxidizing agent particles, for example, selected from the group consisting of hydrogen peroxide, urea peroxide, percarbonate, perborate, and peroxymonophosphate Ester, peroxodisulfate, peroxyacid, and peracetic acid.

Specific Example 9. Any of the foregoing dental patches, wherein the particulate bleaching component is selected from the group consisting of solid peroxides and solid peroxide donors, for example, selected from peroxide salts or complexes (eg, such as peroxyphosphate). Oxidized carbonate, perborate, peroxypoxide, or persulphate; for example, calcium perphosphate, sodium perborate, sodium percarbonate, sodium peroxyphosphate, and potassium persulfate; hypochlorite; Urea peroxide; hydrogen peroxide polymer complex such as hydrogen peroxide-polyvinylpyrrolidone polymer complex; metal peroxide such as zinc peroxide and calcium peroxide; peroxyacid and mixtures thereof.

Specific Example 10. Any of the foregoing dental patches, wherein the particulate bleaching component comprises urea peroxide, hydrogen peroxide-polyvinylpyrrolidone polymer complex, sodium percarbonate, or a combination of two or more thereof.

Specific Example 11. Any of the foregoing dental patches, wherein the particle size (D ₅₀ ) on the first surface of the film, such as a granular bleaching component in the form of a particle, or a perhydrolase or a thiol donor, is from 0.1 to 300 microns, such as 10 -275 microns, eg, 100-250 microns.

Specific Example 12. Any of the foregoing dental patches wherein the particulate bleaching component comprises greater than 0.01%, such as from 0.01% to 0.1%, such as from 0.02% to 0.08%, the total weight of the hydratable adhesive film and the particulate bleaching component attached thereto.

Specific Example 13. Any of the foregoing dental patches, wherein the amount of particulate bleaching agent on the first side of the hydratable adhesive film is from 0.001 to 10 mg/cm ² , such as from 0.001 to 1 mg/cm ² , for example from 0.005 to 0.015 mg. /cm ² .

Specific Example 14. Any of the foregoing dental patches, wherein the thiol-based system is selected from the group consisting of (i) one or more C _2-18 carboxylic acid esters, such as C _2-6 carboxylic acid esters (such as acetamyl ester), comprising lower linear or branched alkyl groups. a carboxylic acid ester, optionally substituted by a hydroxy group and/or a C _1-4 alkoxy group; (ii) one or more acetyl glycerides (such as mono-, di-, or triglycerides), (iii) Acetyl saccharides, and (iv) mixtures thereof.

Specific Example 15. In any of the foregoing dental patches, the thiol based system is selected from the group consisting of 1,2,3-triethoxypropane (sometimes referred to herein as triacetin or triacetin) and deuterated sugars such as B. Deuterated sugar.

Specific Example 16. Any of the foregoing dental patches comprising a thiol donor comprising an ester compound having a water solubility of at least 5 ppm at 25 °C.

Specific Example 17. Any of the foregoing dental patches comprising a peroxyacid or which upon use produce a peroxyacid.

Specific Example 18. Any of the foregoing dental patches, wherein the component is present in a dosage sufficient to provide a bleaching dose and concentration effective to whiten the teeth upon mixing.

Specific Example 19. Any of the foregoing dental patches, wherein the hydratable adhesive film comprises one or more water soluble polymers selected from the group consisting of hydrophilic cellulose ethers (eg, carboxymethyl cellulose, hydroxypropyl cellulose, hydroxypropyl). Methyl cellulose), polyvinyl acetate, carbomer (eg, CARBOPOL ^® 971P), polysaccharide gum (such as xanthan gum), modified food starch, gelatin (such as animal or fish-based gelatin), linked carboxyvinyl copolymers, cross-linked polyvinyl pyrrolidone, polyethylene oxide (e.g. POLYOX ^TM), polyacrylic acid and polyacrylates, polyvinyl alcohol, sodium alginate, casein, pullulan, and a combination thereof .

Specific Example 20. Any of the foregoing dental patches, wherein the hydratable adhesive film comprises one or more water soluble polymers selected from the group consisting of hydrophilic cellulose ethers (such as hydroxypropylmethylcellulose or hydroxypropylcellulose), Ethylene oxide, polyvinyl acetate, and carbomer (eg, CARBOPOL ^® 971P); and combinations thereof.

Specific Example 21. Any of the foregoing dental patches, wherein the hydratable adhesive film comprises hydroxypropylmethylcellulose, polyvinyl acetate, and, for example, a dry weight ratio of 10-20 HPMC: 2-10 PVAc: 1 carbomer card Pom.

Specific Example 22. Any of the foregoing dental patches, wherein the hydratable adhesive film further comprises a plasticizer such as propylene glycol, polyethylene glycol or triacetin.

Specific Example 23. Any of the foregoing dental patches, wherein the first side of the hydratable adhesive film is covered by a protective layer prior to use.

Specific Example 24. Any of the foregoing dental patches, wherein the hydratable adhesive film has a viscosity of at least 100,000 cps upon activation, such as a viscosity of from 100,000 to 200,000 cps.

Specific Example 25. Any of the foregoing dental patches, wherein the hydratable adhesive film is substantially dry prior to use.

Specific Example 26. Any of the foregoing dental patches, wherein the hydratable adhesive film has a thickness of from 0.1 to 5 mm, such as from 0.5 to 5 mm.

Specific Example 27. Any of the foregoing dental patches, wherein the approximate overall size is 2-10 cm long x 0.5-2 cm wide x 0.1-10 mm thick, such as 1-10 mm thick, such as a dental patch, wherein the surface area of one side For 5-20 cm ² , for example, about 5-15 cm ² , for example, about 10 cm ² .

Specific Example 28. Any of the foregoing dental patches comprising a hydrogen peroxide-polyvinylpyrrolidone polymer complex embedding particles, urea peroxide and/or sodium percarbonate and perhydrolase particles on the first surface of the membrane, and dispersed in the membrane Triacetin.

Specific Example 29. Any of the foregoing dental patches further comprising anti-allergic agent particles such as potassium nitrate or arginine.

Specific Example 30. A method of whitening teeth (method 2) comprising applying a first side of a dental patch as previously described, such as a dental patch 1 and below, directly to the tooth and leaving it for a sufficient time, such as at least 5 minutes, for example 10 - 60 minutes, for example, 10-30 minutes to whiten the teeth.

Specific Example 31. A method for preparing a tooth whitening tooth paste (method 3), such as a tooth patch as described above, according to the tooth patch 1 or below, comprising providing a half dry hydratable adhesive film, as described above, for example, the film system It is detached from the water and is not completely dried, or the film is already wet, and is added to one surface of the film particles of the particulate bleaching component, as described above, and the film having the added particles to a surface is dried.

For example, the dental prosthesis can be manufactured by first preparing a hydratable adhesive film using conventional methods and then adding a particulate whitening component to a surface. The hydratable adhesive film dental patch can be detached from the water by a variety of means known in the art, such as by extrusion, or by detachment from an aqueous suspension (e.g., at a solids level of 10-30%) to a heating belt. Water evaporates from it. Alternatively, the film is dry but then wet. The particles can be added to the surface of the film with the film being semi-dry, i.e., only wet and sticky, thereby allowing the particles to adhere to the surface of the film. Once the film is sufficiently dried and cooled to room temperature, the particles continue to adhere to the surface of the film. Prior to use, the integral hydratable adhesive film and the dental patch are thus substantially dry. Since the peroxide is only on the surface of the membrane, only a relatively small amount of particles are required to provide an effective concentration on the surface.

Preferred specific examples:

When exposed to saliva or other sources of water, such as tap water, the particles dissolve and release the reactive components to enzymatically produce the desired peroxyacid. Hydrating the hydratable adhesive layer increases the adhesiveness of the film, allowing the whitening film/dental to bond to the target surface (i.e., tooth enamel).

The hydratable adhesive film comprises one or more water-soluble, orally acceptable polymers selected from the group consisting of hydrophilic cellulose ethers (eg, carboxymethyl cellulose, hydroxypropyl cellulose, hydroxypropyl A) Cellulose), polyvinyl acetate, carbomer (eg, CARBOPOL ^® 971P), polysaccharide gum (eg, xanthan gum), modified food starch, gelatin (eg, animal or fish-based gelatin), cross-linked carboxyvinyl copolymers, cross-linked polyvinyl pyrrolidone, polyethylene oxide (e.g., POLYOX ^TM), polyacrylic acid and polyacrylates, polyvinyl alcohol, sodium alginate, casein, pullulan, and Its combination. The formation of an adhesive containing a tooth whitening agent is known in the art, for example, as described in U.S. Patent Nos. 7,862,801, 5,746,598, 6, 730, 316, and 7,128, 899 each incorporated herein by reference. The adhesive film allows the peroxyacid bleach to stay in contact with the teeth for a prolonged period of time and to protect the soft tissue, and thus provides a high viscosity, for example, a viscosity of at least 100,000 centipoise (cps) when used (about 100 Pascal-second) (Pa.s)), preferably from 100,000 to 200,000 cps (100 to 200 Pa.s).

Wherein, the second film layer is used to protect the hydratable adhesive film from rapid degradation or dissolution, and the carrier or backing material can be made of the following: textile, cloth, synthetic wood board, resin, elastic material, paper, insoluble or insoluble. cellulose derivatives such as ethyl cellulose and cellulose acetate, polyvinyl chloride, waxes, PARAFILM ^TM, polyethylene, polyvinyl alcohol, TEFLON ^TM, polyvinyl chloride, vinyl esters and other derivatives of polyvinyl acetate.

The particulate bleaching component can be a solid peroxide or a solid peroxide donor selected from the group consisting of peroxide salts or complexes (such as peroxy phosphate, peroxycarbonate, perborate, Oxidized citrate, or persulphate; for example, calcium perphosphate, sodium perborate, sodium percarbonate, sodium peroxyphosphate, and potassium persulfate), hypochlorite; urea peroxide; hydrogen peroxide polymerization Complexes such as hydrogen peroxide-polyvinylpyrrolidone polymer complexes, and metal peroxides, for example, zinc peroxide and calcium peroxide; solid peroxyacids; and combinations thereof. In a particular embodiment, the particulate bleaching component is urea peroxide or a hydrogen peroxide polyvinylpyrrolidone polymer complex. The particulate bleach component can be optionally embedded to provide enhanced storage stability (e.g., entrapped with sodium sulfate, corn starch or gum arabic).

Preferred specific examples are listed:

Preferred specific example 1. A tooth whitening toothpaste comprising a hydratable adhesive film comprising a first side and a second side, the first side comprising a particulate bleaching component attached thereto, wherein the tooth whitening toothpaste further comprises the following in the film or Attached to the first side of the membrane on a membrane or in a particulate form; a) a hydrolytically active enzyme having a carbohydrate esterase family 7 (CE-7) tag motif aligned with the reference sequence SEQ ID NO: 1. The label motif comprises: i) a RGQ motif at a position corresponding to positions 118-120 of SEQ ID NO: 1; ii) a GXSQG motif at a position corresponding to positions 186-190 of SEQ ID NO: 1; And iii) a HE motif at a position corresponding to positions 303-304 of SEQ ID NO: 1; and b) at least one mercapto donor matrix selected from the group consisting of: i) an ester having the formula Class [X] _m R ₅ wherein X = an ester group of the formula R ₆ C(O)O; R ₆ = a C1 to C7 straight chain, branched or cyclic hydrocarbyl moiety, any of which is a hydroxyl group or a C1 to C4 alkane radicals substituted, in which when R ₆ = C2 to C7, R ₆ include any one or more ether bonds linked; R ₅ = C1 to C6 straight, branched or cyclic hydrocarbon portion Or cyclic heteroaryl five or six-membered ring aryl or heteroaryl moiety, any of which is substituted with a hydroxyl group; wherein each of the carbon atoms R ₅ is based on independent includes no more than one hydroxy group or Not more than one ester group or carboxylic acid group; and wherein R ₅ optionally includes one or more ether linkages; m is an integer from 1 to R ₅ in the number of carbon atoms, and wherein the ester is at 25 The water solubility of °C is 5ppm; ii) the glyceride with the following structural formula Wherein R ₁ = C1 to C7 is a straight or branched alkyl group, any of which is substituted by a hydroxy group or a C1 to C4 alkoxy group and R ₃ and R ₄ are independently H or R ₁ C(O); iii) one or a variety of esters of the following formula Wherein R ₁ is a C1 to C7 straight or branched alkyl group, which is optionally substituted by a hydroxy or C1 to C4 alkoxy group and R ₂ is a C1 to C10 straight or branched alkyl group, an alkenyl group, an alkynyl group, Aryl, alkylaryl, alkylheteroaryl, heteroaryl, (CH ₂ CH ₂ O) _n , or (CH ₂ CH(CH ₃ )-O) _n H and n is from 1 to 10; An acetylated saccharide selected from the group consisting of acetylated monosaccharides, acetylated disaccharides, and acetaminophen; wherein upon hydration of the hydratable adhesive film, the hydrogen peroxide is composed of granular bleaching The fraction is released and the enzyme is an enzymatically effective amount of peroxyacid.

Preferred specific example 2. The tooth whitening toothpaste according to the preferred embodiment 1, wherein the perhydrolysis-active enzyme comprises an amino acid sequence selected from the group consisting of: a) SEQ ID NO: 1; and b) having at least 80% amino acid sequence The amino acid sequence identical to SEQ ID NO: 1.

Preferred specific example 3. The tooth whitening toothpaste according to the preferred embodiment 1, wherein the hydrolyzable enzyme further comprises a binding structural region fused to the N- or C-terminus of the enzyme, the binding structural region for oral tissue or for tooth whitening Dental patches have affinity.

Preferred embodiment 4 is preferred. A tooth whitening toothpaste according to preferred embodiment 3, wherein the binding structural region has an affinity for oral tissues and comprises an amino acid sequence selected from the group consisting of SEQ D NOs: 178-197.

Preferred embodiment 5 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the hydrolyzed enzyme has affinity for oral tissues and comprises an amino acid sequence selected from the group consisting of a) SEQ ID NO: 2, and b An amino acid sequence having at least 80% amino acid sequence identical to SEQ ID NO: 2.

Preferred embodiment 6 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, further comprising a backing layer attached to the second side of the hydratable adhesive film, the backing layer inhibiting dissolution of the hydratable adhesive film.

Preferred embodiment 7 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the particulate bleaching component is embedded in a water soluble coating which is soluble when hydrated.

Preferred specific example 8. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the particulate bleaching component is selected from the group consisting of solid peroxides and solid peroxide donors.

Preferred specific example 9. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the particulate bleaching component is selected from the group consisting of a peroxide salt, a peroxide complex, a peroxy phosphate, a peroxycarbonate, and a perboric acid. Salt, peroxy citrate, persulfate, calcium peroxide, Sodium perborate, sodium carbonate peroxide, sodium peroxyphosphate, potassium persulfate, hypochlorite, urea peroxide, hydrogen peroxide polymer complex, hydrogen peroxide-polyvinylpyrrolidone polymer complex, Metal peroxides, zinc peroxide, calcium peroxide, and combinations thereof.

Preferred embodiment 10 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the particulate bleaching component comprises urea peroxide. In some embodiments, the particulate bleaching component comprises a hydrogen peroxide-polyvinylpyrrolidone polymer complex. In some embodiments, the hydrogen peroxide-polyvinylpyrrolidone polymer complex is a hydrogen peroxide-crosslinked polyvinylpyrrolidone polymer complex.

Preferred embodiment 11 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the particulate bleaching component has a particle size average diameter (D50) ranging from 10 micrometers to 300 micrometers, for example, 10 micrometers to 200 micrometers.

Preferred embodiment 12 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the tooth whitening toothpaste comprises from about 0.01% by weight to about 0.1% by weight peroxyacid. In some embodiments, the particulate bleaching component comprises from about 0.1% to about 30% by weight of a peroxide source.

Preferred embodiment 13 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the amount of particulate bleaching agent on the first side of the hydratable adhesive film ranges from 0.001 mg/cm ² to 10 mg/cm ² , such as 0.001 mg/cm ² to 1 mg/cm ² .

Preferred embodiment 14 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the thiol donor matrix is 1,2,3-triethoxypropane.

Preferred embodiment 15 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the hydratable adhesive film comprises one or more water soluble polymers selected from the group consisting of hydrophilic cellulose ethers, carboxymethyl cellulose, hydroxypropyl cellulose , hydroxypropyl methylcellulose, polyvinyl acetate, carbomer, polysaccharide gum, xanthan gum, modified food starch, gelatin, animal or fish-based gelatin, cross-linked carboxyethylene copolymer, cross-linking Polyvinylpyrrolidone, polyethylene oxide, polyacrylic acid, poly Acrylate, polyvinyl alcohol, sodium alginate, casein, pullulan, and combinations of two or more thereof.

Preferred embodiment 16 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the hydratable adhesive film comprises one or more water soluble polymers selected from the group consisting of hydrophilic cellulose ethers, polyvinyl acetate, carbomer And a combination of two or more thereof.

Preferred embodiment 17 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the hydratable adhesive film comprises hydroxypropylmethylcellulose (HPMC), polyvinyl acetate (PVAc), and HMPC: PVAc: Carbomer The carbomer with a dry weight ratio of 10-20:2-10:1.

Preferred embodiment 18 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, wherein the hydratable adhesive film further comprises a plasticizer.

Preferred embodiment 19 is preferred. A tooth whitening toothpaste according to any of the above preferred embodiments, further comprising propylene glycol.

Preferred embodiment 20 is preferred. A method of whitening teeth comprising: a) providing a packaging system comprising a tooth whitening toothpaste according to any of the above preferred embodiments; b) removing the tooth whitening toothpaste from the packaging system; and c) whitening the teeth The time when the dental patch is in direct contact with the tooth to whiten the tooth; wherein the tooth whitening tooth is hydrated by the presence of water in the mouth or on the surface of the tooth or after step (b) but before step (c) .

Preferred embodiment 21 is preferred. The method of the preferred embodiment 20, wherein the whitening toothpaste further comprises a backing layer attached to the second side of the hydratable adhesive film, the backing layer inhibiting dissolution of the hydratable adhesive film.

Preferred embodiment 22 is preferred. A method according to any of the above preferred embodiments, wherein the particulate bleaching component has a particle size average diameter (D50) ranging from 10 micrometers to 200 micrometers.

Preferred embodiment 23 is preferred. A method according to any of the above preferred embodiments, wherein the particulate bleaching component comprises greater than 0.01% by weight of the hydratable adhesive film and the total weight of the particulate bleaching component attached thereto. In some embodiments, the particulate bleaching component comprises greater than 0.05% by weight of the hydratable adhesive film and the total weight of the particulate bleaching component attached thereto. In some embodiments, the particulate bleaching component comprises from about 0.01% to about 0.1% by weight of the hydratable adhesive film and the total weight of the particulate bleaching component attached thereto.

Preferred embodiment 24 is preferred. A method according to any of the above preferred embodiments, wherein the amount of particulate bleach on the first side of the hydratable adhesive film ranges from 0.001 mg/cm ² to 1 mg/cm ² .

Preferred embodiment 25 is preferred. A method according to any of the above preferred embodiments, wherein the thiol donor substrate is 1,2,3-triethoxypropane.

Preferred embodiment 26 is preferred. A method according to any of the above preferred embodiments, wherein the hydratable adhesive film comprises one or more water soluble polymers selected from the group consisting of hydrophilic cellulose ethers, carboxymethyl cellulose, hydroxypropyl cellulose , hydroxypropyl methylcellulose, polyvinyl acetate, carbomer, polysaccharide gum, xanthan gum, modified food starch, gelatin, animal or fish-based gelatin, cross-linked carboxyethylene copolymer, cross-linking Polyvinylpyrrolidone, polyethylene oxide, polyacrylic acid, polyacrylate, polyvinyl alcohol, sodium alginate, casein, pullulan, and combinations of two or more thereof.

Preferred embodiment 27 is preferred. A method according to any of the above preferred embodiments, wherein the hydratable adhesive film comprises one or more water soluble polymers selected from the group consisting of hydrophilic cellulose ethers, polyvinyl acetate, carbomer, and A combination of two or more.

Preferred embodiment 28 is preferred. The method according to any of the above preferred embodiments, wherein the hydratable adhesive film comprises hydroxypropylmethylcellulose (HPMC), polyvinyl acetate (PVAc), and HMPC:PVAc: carbomer dry weight ratio It is a carbomer of 10-20:2-10:1.

Preferred embodiment 29 is preferred. A method according to any of the above preferred embodiments, wherein the hydratable adhesive film further comprises a plasticizer.

Preferred embodiment 30 is preferred. A method according to any of the above preferred embodiments, wherein the hydratable adhesive film further comprises propylene glycol.

Preferred embodiment 31 is preferred. A method according to any of the above preferred embodiments, wherein the particulate bleaching component is embedded in a water soluble coating which is soluble upon hydration.

Preferred embodiment 32 is preferred. A method of preparing a tooth whitening toothpaste comprising: a) providing a half dry hydratable adhesive film, b) applying a granule of a granular bleaching component to a surface of the film, whereby the particle adheres to the surface, And c) drying the film.

In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of from 10 microns to 300 microns. In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of from 25 microns to 200 microns. In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of from 35 microns to 150 microns. In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of from 50 microns to 125 microns. In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of from 60 microns to 100 microns. In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of about 64 microns. In some embodiments, the particulate bleaching component has a particle size average diameter (D50) of about 94 microns.

In some embodiments, the perhydrolysis-active enzyme has a particle size average diameter (D50) of from 100 micrometers to 300 micrometers. In some embodiments, the perhydrolysis-active enzyme has a particle size average diameter (D50) of from 150 microns to 275 microns. In some embodiments, the perhydrolysis-active enzyme has a particle size average diameter (D50) of from 175 microns to 250 microns.

All of the components used in the dental patches described herein should be orally acceptable. The term "orally acceptable" as used herein means that the amount and form of the components present in the illustrated dental patch does not render the dental patch unsuitable for use in the oral cavity.

Throughout the text, ranges are used in abbreviated form to describe values in each and every range. Any range of values can be selected as the terminal of the range. In addition, all of the references cited herein are hereby incorporated by reference in their entirety. In the event that the disclosure of the present disclosure conflicts with the cited references, the invention is subject to the teachings.

Unless otherwise stated, it is to be understood that all percentages and quantities in this specification and the specification are by weight. The amount given is based on the active weight of the substance.

Instance

The following examples are provided to demonstrate preferred aspects of the invention. It will be appreciated by those skilled in the art that the techniques disclosed in the examples are in accordance with the teachings of the present invention, and thus may be considered as a preferred form of construction. However, those skilled in the art can recognize that many changes can be made in the specific embodiments disclosed and still obtain the same or similar results without departing from the methods and examples disclosed herein. The spirit and scope.

Unless otherwise indicated, all reagents and materials were obtained from DIFCO Laboratories (Detroit, Michigan), GIBCO/BRL (Gaithersburg, Maryland), TCI America (Portland, Oregon), Roche Diagnostic Company (Indiana, Indiana), Thermo Scientific (Pierce Protein Research Products) (Rockford, Ill.) or Sigma/Aldrich Chemical Company (St. Louis, Missouri).

Abbreviations in the following descriptions relate to measurements, techniques, features, or units of compounds: "sec" or "s" means seconds, "min" means minutes, "h" or "hr" means hours, "μL" Refers to microliters, "mL" refers to milliliters, "L" refers to liters, "mM" refers to millimoles, "M" refers to moles, "mmol" refers to milliliters Mohr, "ppm" means "wt" means weight per million, "wt%" means weight percentage, "g" means gram, "mg" means milligram, and "μg" means microgram. And "ng" means nanograms.

Example 1

A tooth paste was prepared as described above to form a hydratable adhesive film and then, when the film was still tacky, a particulate whitening agent and a granular enzyme were added to the surface of one side, and the components in Table 1 were used. The tooth paste will slowly dissolve in the mouth when used, and therefore does not need to be removed.

The dental patch of Example 1 was detached from the water by extrusion, or detached from the aqueous suspension (e.g., at a solids level of 10-30%) to a heating zone from which water was evaporated. The particles are added to the surface of the film when the film is semi-dry or fully dry but still tacky. Once cooled to room temperature, the particles adhere to the surface of the film. When the area of the dental patch is 10 cm ² and the weight is 10 mg/cm ² , this formulation provides 5 mg of triacetin. Assuming that 1.4 mg of bleach particles are distributed over the toothpaste, together with 0.3 mg of enzyme granules, this dose is sufficient to produce sufficient peracetic acid directly on the tooth surface to significantly exceed the peroxide-containing whitening toothpaste. (Peroxide-only dental patches typically contain about 3-10 mg of total peroxide in a typical dose.)

When exposed to saliva or other sources of water, such as tap water, the particles dissolve and activate immediately. The adhesive layer is also activated and effectively adheres to the teeth. Example 1 is designed to slowly erode in the mouth over time so the user does not need to remove it.

Example 2

A dental patch is prepared as described above to form the hydratable adhesive film and then, when the dental patch is still tacky, the granulating agent is added to one side and the protective backing layer is applied to the other side. The components in Table 2. Since the backing layer does not dissolve, the user will remove it after allowing the whitening to occur, typically about 10-30 minutes. The two layers may also be formed by extrusion or solvent casting, and then a particulate whitening agent may be added to the surface of the hydratable adhesive film.

Example 3

The ability of the hydrogen peroxide-polyvinylpyrrolidone complex of various particle sizes and the enzyme having perhydrolysis activity ("enzyme") to uniformly produce peracetic acid on the surface of the hydratable adhesive containing triacetin was evaluated.

To evaluate the formation of peracetic acid from this product, a 3/8" wafer was cut from the membrane. Each disc was incubated with 20 μL of 50 mM sodium phosphate buffer pH 7.2 water for 15 minutes at 37 ° C. 380 μL of 0.1 M phosphoric acid Add to the membrane to stop the enzyme reaction and dilute the sample for testing. Analyze the The peracetic acid of the solution was analyzed by HPLC for peracetic acid using the method previously described in U.S. Patent No. 7,829,315 - DiCosimo et al. At least three samples were cut from the 24 inch long film on the dental patch product in each evaluation.

A dental patch sample containing an enzyme and a hydrogen peroxide-polyvinylpyrrolidone coating was prepared by first adding an enzyme to a pullulan polymer film. The dried enzyme-containing membrane was ground and sieved at a particle size of 60 to 80 mesh (177 μm to 250 μm). The particles were then type of enzyme and hydrogen peroxide - mixing polyvinylpyrrolidone (PEROXYDONE ^TM XL-10, and even blue alkylene group, Wilmington, Delaware). The mixture is deposited onto a two layer film structure of a hydratable adhesive layer comprising primarily polyethylene oxide and triacetin and a polyvinyl alcohol backing layer to provide an insoluble support layer. The results of the evaluation of two consecutive processes for producing peracetic acid are shown in Table 3. The samples were initially and finally evaluated at the production process and confirmed the poor consistency of peracetic acid.

The PEROXYDONE ^TM XL-10 is further treated with ethanol to a high shear granulation process to form larger particles. After granulation, the sample was sieved at a particle size of 60 to 200 mesh (75 μm to 250 μm). Described in Table 3 to manufacture samples of PEROXYDONE ^TM XL-10, and sieved and the particle size distribution based PEROXYDONE ^TM XL-10 based on high shear granulation method described in Table 4. The samples were measured on a Beckman Coulter LS13320 equipped with a Tornado dry feeder. The powder was analyzed directly without being dispersed in a liquid.

The preparation process of another dental patch was first completed by loading the pullulan matrix to twice the concentration of the sample of Table 3 to prepare an enzyme sample. The polysaccharide pullulan film polishing and is of 60 to 80 mesh sieve (177 m to 250 m) and then combined with a particulate PEROXYDONE ^TM XL-10. The mixture was then embedded as described above onto a structure similar to a two-layer film. The evaluation of this sample for the formation of peracetic acid is provided in Table 4 and demonstrates a higher and more consistent production of peracetic acid throughout the production process.

<110> American Palm Company

<120> Acid-producing composition

<130> 9357-00-WO-OC

<150> 61/577,499

<151> 2011-12-19

<160> 197

<170> PatentIn version 3.5

<210> 1

<211> 325

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<400> 1

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<220>

<221> Other important biological functional areas

<222> (1)..(325)

<223> Haiqi hot robe C277S variant hydrolysis?

<220>

<221> Other important biological functional areas

<222> (326)..(375)

<223> Oral surface targeting domain

<400> 2

<210> 3

<211> 960

<212> DNA

<213> Bacillus subtilis

<220>

<221> CDS

<222> (1)..(960)

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<213> Bacillus pumilus

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<213> Clostridium thermophilum

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<213> Cloning the new Apollo Thermotoga

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<400> 11

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<213> Anaerobic thermophilus

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<213> Bacillus licheniformis

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<213> Cloning the new Apollo Thermotoga

<220>

<221> Other important biological functional areas

<222> (277)..(277)

<223> Xaa is Ala, Val, Ser, or Thr.

<400> 15

<210> 16

<211> 325

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<220>

<221> Other important biological functional areas

<222> (277)..(277)

<223> Xaa is Ala, Val, Ser, or Thr.

<400> 16

<210> 17

<211> 326

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<213> Laitinger Thermotoga

<220>

<221> Other important biological functional areas

<222> (277)..(277)

<223> Xaa is Ala, Val, Ser, or Thr.

<400> 17

<210> 18

<211> 325

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<213> Thermotoga

<220>

<221> Other important biological functional areas

<222> (277)..(277)

<223> Xaa is Ala, Val, Ser, or Thr.

<400> 18

<210> 19

<211> 325

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<213> Thermotoxin RQ2a

<220>

<221> Other important biological functional areas

<222> (277)..(277)

<223> Xaa is Ala, Val, Ser, or Thr.

<400> 19

<210> 20

<211> 329

<212> PRT

<213> Thermotoxin RQ2b

<220>

<221> Other important biological functional areas

<222> (278)..(278)

<223> Xaa is Ala, Val, Ser, or Thr.

<400> 20

<210> 21

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<213> Laitinger Thermotoga

<400> 21

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<213> Thermotoga

<400> 22

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<400> 24

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<213> Habitat anaerobic saccharifying bacteria

<400> 25

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<213> Lactococcus lactis

<400> 26

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<220>

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<220>

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<220>

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<400> 42

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<213> Artificial sequence

<220>

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<400> 45

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<213> Artificial sequence

<220>

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<220>

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<400> 47

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<220>

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<220>

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<220>

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<400> 50

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<220>

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<400> 51

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<220>

<223> Synthetic structure

<400> 52

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<212> PRT

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<400> 53

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<220>

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<400> 54

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<220>

<223> Synthetic structure

<400> 55

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<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic structure

<400> 56

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<211> 14

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<220>

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<400> 57

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<220>

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<400> 58

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<220>

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<400> 59

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<220>

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<400> 60

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<211> 14

<212> PRT

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<220>

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<400> 61

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<212> PRT

<213> Artificial sequence

<220>

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<400> 62

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<220>

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<400> 63

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<211> 14

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic structure

<400> 64

<210> 65

<211> 14

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic structure

<400> 65

<210> 66

<211> 20

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic structure

<400> 66

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<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic structure

<400> 67

<210> 68

<211> 20

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic structure

<400> 68

<210> 69

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<212> PRT

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<220>

<223> Synthetic oral surface bonding?

<400> 69

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<211> 20

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<213> Artificial sequence

<220>

<223> Synthetic oral surface bonding?

<400> 70

<210> 71

<211> 20

<212> PRT

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<220>

<223> Synthetic oral surface bonding?

<400> 71

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<220>

<223> Synthetic oral surface bonding?

<400> 72

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<212> PRT

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<220>

<223> Synthetic oral surface bonding?

<400> 73

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<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic oral surface bonding?

<400> 74

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<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 75

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<211> 18

<212> PRT

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<220>

<223> Synthetic oral surface bonding?

<400> 76

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<220>

<223> Synthetic oral surface bonding?

<400> 77

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<211> 15

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<220>

<223> Synthetic oral surface bonding?

<400> 78

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<220>

<223> Synthetic oral surface bonding?

<400> 79

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<211> 15

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<220>

<223> Synthetic oral surface bonding?

<400> 80

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<211> 15

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<220>

<223> Synthetic oral surface bonding?

<400> 81

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<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 82

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<220>

<223> Synthetic oral surface bonding?

<400> 83

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<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 84

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<211> 20

<212> PRT

<213> Artificial sequence

<220>

<223> Synthetic oral surface bonding?

<400> 85

<210> 86

<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 86

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<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 87

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<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 88

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<211> 20

<212> PRT

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<220>

<223> Synthetic oral surface bonding?

<400> 89

<210> 90

<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 90

<210> 91

<211> 20

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<220>

<223> Synthetic oral surface bonding?

<400> 91

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<211> 15

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<220>

<223> Synthetic oral surface bonding?

<400> 92

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<220>

<223> Synthetic oral surface bonding?

<400> 93

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<211> 15

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<400> 94

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<220>

<223> Synthetic oral surface bonding?

<400> 95

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<211> 20

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<220>

<223> Synthetic oral surface bonding?

<400> 96

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<220>

<223> Synthetic oral surface bonding?

<400> 97

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<220>

<223> Synthetic oral surface bonding?

<400> 98

<210> 99

<211> 18

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<220>

<223> Synthetic oral surface bonding?

<400> 99

<210> 100

<211> 20

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<220>

<223> Synthetic oral surface bonding?

<400> 100

<210> 101

<211> 20

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<220>

<223> Synthetic oral surface bonding?

<400> 101

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<220>

<223> Synthetic oral surface bonding?

<400> 102

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<400> 103

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<400> 104

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<400> 105

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<400> 106

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<400> 107

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<400> 108

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<400> 109

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<400> 110

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<400> 111

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<400> 112

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<400> 113

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<400> 114

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<400> 115

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<400> 116

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<400> 117

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<400> 118

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<400> 119

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<400> 120

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<400> 121

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<400> 125

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<400> 128

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<400> 130

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Claims (16)

A tooth whitening toothpaste comprising a hydratable adhesive film having a first side and a second side, the first side comprising a particulate bleaching component attached thereto, wherein the tooth whitening toothpaste further comprises; a) A hydrolytically active enzyme having a carbohydrate esterase family 7 (CE-7) tag motif aligned with the reference sequence SEQ ID NO: 1, the tag motif comprising: i) RGQ motif corresponding to To the position of position 118-120 of SEQ ID NO: 1; ii) the position of GXSQG motif corresponding to position 186-190 of SEQ ID NO: 1; and iii) the position of HE motif corresponding to SEQ ID NO: a position of 303-304; and (b) a ruthenium-based donor substrate, the matrix being 1,2,3-triethoxypropane; wherein the particulate bleaching component is selected from the group consisting of peroxide salts, Peroxide complex, peroxyphosphate, percarbonate, perborate, peroxyphenate, persulfate, calcium perphosphate, sodium perborate, sodium percarbonate, sodium peroxyphosphate, Potassium sulphate, hypochlorite, urea peroxide, hydrogen peroxide polymer complex, hydrogen peroxide-polyvinylpyrrolidine a polymer complex, a metal peroxide, zinc peroxide, calcium peroxide, and combinations thereof; wherein the hydrolytically active enzyme and the thiol donor matrix are attached to the first side of the film in a particulate form or In the film; wherein, upon hydration of the hydratable adhesive film, the hydrogen peroxide is released from the particulate bleaching component and the enzyme is capable of producing an enzymatically effective amount of peroxyacid; and wherein the hydratable adhesive film is The amount of particulate bleach on the first side ranges from 0.001 mg/cm ² to 1 mg/cm ² . A tooth whitening toothpaste according to the first aspect of the invention, wherein the perhydrolysis-active enzyme comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 1. A tooth whitening toothpaste according to the first aspect of the invention, wherein the hydrolytically active enzyme has affinity for oral tissues and comprises an amino acid sequence selected from the group consisting of SEQ ID NO: 2. The tooth whitening toothpaste of claim 1, further comprising a backing layer attached to the second side of the hydratable adhesive film, the backing layer inhibiting dissolution of the hydratable adhesive film. A tooth whitening toothpaste according to the first aspect of the patent application, wherein the granular bleaching component is embedded in a water-soluble paint which is soluble in hydration. A tooth whitening toothpaste according to the first aspect of the invention, wherein the particulate bleaching component is selected from the group consisting of a solid peroxide and a solid peroxide donor. The tooth whitening toothpaste according to the first aspect of the invention, wherein the granular bleaching component is selected from the group consisting of urea peroxide, sodium percarbonate, hydrogen peroxide-polyvinylpyrrolidone polymer complex, and two or A variety of compositions. A tooth whitening toothpaste according to the first aspect of the invention, wherein the granular bleaching component has a particle size average diameter (D50) of from 100 micrometers to 250 micrometers. a tooth whitening toothpaste according to item 1 of the patent application scope, wherein the hydrolyzable enzyme The average particle size (D50) is from 100 microns to 250 microns. A tooth whitening toothpaste according to the first aspect of the invention, wherein the dental patch produces at least about 0.1% by weight of peroxyacid. The tooth whitening toothpaste according to claim 1, wherein the hydratable adhesive film comprises one or more water-soluble polymers selected from the group consisting of hydrophilic cellulose ether, carboxymethyl cellulose, and hydroxypropyl group. Cellulose, hydroxypropyl methylcellulose, polyvinyl acetate, carbomer, polysaccharide gum, xanthan gum, modified food starch, gelatin, animal or fish-based gelatin, cross-linked carboxyethylene copolymer, Crosslinked polyvinylpyrrolidone, polyethylene oxide, polyacrylic acid, polyacrylate, polyvinyl alcohol, sodium alginate, casein, pullulan, and combinations of two or more thereof. The tooth whitening toothpaste according to claim 1, wherein the hydratable adhesive film comprises one or more water-soluble polymers selected from the group consisting of hydrophilic cellulose ethers, polyvinyl alcohol, carbomers, and A combination of two or more thereof. A tooth whitening toothpaste according to the first aspect of the invention, wherein the hydratable adhesive film further comprises a plasticizer. A tooth whitening toothpaste according to the first aspect of the invention, wherein the hydratable adhesive film further comprises propylene glycol. A tooth whitening toothpaste according to the first aspect of the patent application, wherein the granular bleaching component is embedded in a water-soluble paint which is soluble in hydration. A tooth whitening toothpaste according to the first aspect of the invention, wherein an effective amount of peroxyacid is formed across the surface of the tooth whitening tooth at a substantially uniform concentration.