Enzyme composition for treatment of plaque and plaque bacterial biofilm and for natural whitening of enamel of teeth

Abstract

One aspect of the invention relates to an oral health enzyme complex for the treatment of plaque and plaque bacterial biofilms and for the natural whitening of tooth enamel, comprising the combination of:-a sucrase enzyme, in particular a beta-2, 6 and / or beta-2, 1 bond capable of hydrolyzing sucrose; -a mixture of glucanohydrolases comprising a mutanase having excision activity and obtained from fungi, and a mutanase having excision activity and obtained from bacteria, in particular capable of hydrolyzing (1-3)-alpha-D-glucan and (1-6)-alpha-D-glucan; -a glycoside hydrolase enzyme, in particular a (1-6)-alpha-D glycosidic bond capable of hydrolyzing monosaccharide dimers and trimers; an oxide reductase, in particular capable of forming hydrogen peroxide from glucose and monosaccharide monomers; -a peroxidase, in particular capable of converting thiocyanates into hypothiocyanates in the presence of hydrogen peroxide.

Description

Technical Field

[0001] The technical field of the present invention is the technical field of oral-dental hygiene and beauty products.

[0002] The present invention relates to a novel enzyme complex that can prevent the formation of dental plaque, caries and tartar while making tooth enamel naturally whiter. The present invention also relates to a new way to prevent and treat dental bacterial biofilm by using the enzyme complex. Background Art

[0003] Sugars in foods, over 90% of which are sucrose, are broken down in the mouth by cariogenic bacteria into glucans, which form bacterial biofilms on the tooth enamel. Bacterial biofilms are organized, structured communities of bacterial cells enclosed in an extracellular matrix of varying density and composition. The primary component of the dental biofilm matrix is ​​composed of polysaccharides. In fact, up to 40% of the dry weight of dental biofilm is composed of polysaccharides. In the presence of sucrose, this bacterial biofilm grows and thickens on the teeth, initially forming soft dental plaque composed of soluble glucans (called (1-6)-α-D-glucans or dextran). These dextran are then converted to water-insoluble glucans (called (1-3)-α-D-glucans or their variants) to form hard dental plaque.

[0004] If the diet is mainly composed of sugar, the pH value of dental plaque will be acidic, which will promote the formation of tooth decay; if the diet is mainly composed of meat, the pH value will be alkaline, and the hard dental plaque will calcify to form tartar, which is extremely difficult to remove.

[0005] According to a 2021 report from the World Health Organization, 3.5 billion people worldwide suffered from dental problems related to caries and / or tartar buildup in 2020. Similarly, according to the 2018 U.S. National Health and Nutrition Examination Survey, 59% of adolescents aged 12-19 and 92% of adults aged 20-64 had caries in their permanent teeth. Therefore, there is a continued need to reduce the incidence of these conditions.

[0006] Current methods for reducing dental plaque include mechanical cleaning, chemical agents, fluoride application, and broad-spectrum antiseptics and antimicrobials such as chlorhexidine. Although these therapies are effective in the short term, they all have side effects. In fact, mechanical cleaning is known to cause tooth sensitivity and gingival bleeding; chemical agents pose health risks; the use of fluoride may have significant adverse effects on human health and has limited effectiveness in preventing dental caries; and chlorhexidine may cause tooth staining and aggravate dental caries. In addition, the use of antiseptics and chemical agents has been shown to disrupt commensal microbiota, enhance the pathogenicity and cytotoxicity of host cells, and have other adverse effects on oral health.

[0007] Therefore, these solutions are not satisfactory.

[0008] There is a growing trend towards more preventive treatments that are more respectful of the consumer's overall health and have minimal side effects, such as the use of probiotics, prebiotics or natural substances, but their therapeutic effectiveness is clearly limited.

[0009] Among oral pathogens, Streptococcus mutans is the primary cause of bacterial biofilm formation, a major cause of dental caries. It utilizes dietary sucrose to produce lactic acid and synthesize extracellular polysaccharides, particularly (1-3)- and (1-6)-α-D-glucans, which promote biofilm formation and enhance the resistance of microorganisms within the biofilm. Lactic acid demineralizes the tooth surface, promoting the formation of dental plaque and periodontitis. The acidic biofilm formed by S. mutans is highly resistant to environmental stress, host immunity, and antimicrobial therapy, particularly lysozyme. Studies in dental pathology have demonstrated that these streptococci, protected by the bacterial biofilm matrix, are difficult to eliminate.

[0010] Therefore, there is a need to prevent the proliferation of S. mutans.

[0011] Lactoperoxidase is known to be an enzyme with a bactericidal effect when hydrogen peroxide and thiocyanate are present, and is particularly effective against Streptococcus mutans. It is known that these three substances can be provided by supplying exogenous lactoperoxidase, glucose oxidase and potassium thiocyanate. However, in this type of composition, the formation of hydrogen peroxide is not enough to make the bactericidal effect of lactoperoxidase have a significant impact on the Streptococcus mutans biofilm in terms of reducing caries and / or tartar formation. In addition, in the field of oral hygiene, it is not recommended to add exogenous glucose to this type of composition. Therefore, this solution is not satisfactory in preventing the proliferation of Streptococcus mutans.

[0012] Overall, therefore, a key challenge in combating caries and tartar, and maintaining healthy and aesthetically pleasing teeth, is to maintain the pH of dental plaque as close to physiological neutrality as possible in daily life. Recent research on dental diseases indicates that the best way to prevent these conditions is to eradicate bacterial biofilms. To this end, it is particularly important to prevent the formation of Streptococcus mutans biofilms and to disintegrate existing biofilms so that they can be replaced by physiological plaque—the natural protective biofilm composed of saliva and oral microbiota that surrounds healthy teeth.

[0013] In addition to having healthy teeth, people also want their teeth to be as aesthetically pleasing as possible. Therefore, there is a constant need to whiten the enamel of the teeth in order to combat yellowing, which is caused in particular by aging and the formation of insoluble dental plaque.

[0014] Documents FR 2651433, FR 2822700, FR 3020758, FR 2803199 and WO 2021 / 144211 all disclose enzyme complexes for oral health, but none of these documents disclose a glucan hydrolase mixture comprising a mutanase with exo-activity and derived from fungi and a mutanase with endo-activity and derived from bacteria. Summary of the Invention

[0015] The present invention provides a solution to the above-discussed problems by providing an enzyme complex whose composition is based, inter alia, on the synergistic effect of its different components, resulting from extensive research by the applicant. This enzyme complex is of natural origin, respects the overall health of the consumer, and has no side effects.

[0016] One aspect of the present invention relates to an enzyme complex for use in the treatment of dental plaque and bacterial biofilm of dental plaque and for the natural whitening of tooth enamel, comprising the following compounds:

[0017] - sucrase;

[0018] - a glucan hydrolase mixture comprising a glucanase, a mutanase with exo-active activity and obtained from fungi, and a mutanase with endo-active activity and obtained from bacteria;

[0019] -glycoside hydrolases;

[0020] -oxidoreductases;

[0021] -Peroxidase.

[0022] According to one aspect of the present invention, the glucan hydrolase mixture comprises a mixture of chimeric glucanases comprising a glucanase and a mutanase linked by genetic engineering.

[0023] According to another aspect of the present invention, the mutanase having exo-activity is obtained from the fungus Trichoderma Harzianum.

[0024] According to another aspect of the present invention, the mutanase having endo-activation activity is obtained from Paracoccus Mutanolyticus, Paenibacillus or Streptomyces bacteria.

[0025] According to one aspect of the invention, the sucrase is isomaltase or invertase.

[0026] According to another aspect of the present invention, the glycoside hydrolase is an amyloglucosidase, preferably an amyloglucosidase.

[0027] According to yet another aspect of the present invention, the oxidoreductase is glucose oxidase.

[0028] According to one aspect of the invention, the peroxidase is lactoperoxidase.

[0029] According to another aspect of the present invention, the enzyme complex comprises the following compounds:

[0030] - sucrase, preferably invertase or isomaltase;

[0031] - glucanase;

[0032] a mutanase mixture comprising an exoactive mutanase obtained from a fungus and an endoactive mutanase obtained from a bacterium;

[0033] - amyloglucosidase, preferably amyloglucosidase-α-1,6-glucosidase;

[0034] - glucose oxidase; and

[0035] -Lactoperoxidase.

[0036] According to another aspect of the present invention, the enzyme complex comprises the following compounds, wherein the indicated percentages are by volume:

[0037] - 10%-30%, preferably 15%-25% sucrase;

[0038] - 1%-10%, preferably 4%-7% glucanase;

[0039] - 10% to 40%, preferably 15% to 30%, of a mixture comprising a mutanase having exo-activity and obtained from a fungus and a mutanase having endo-activity and obtained from a bacterium;

[0040] - 20%-60%, preferably 30%-50% amyloglucosidase, preferably amylo-α-1,6-glucosidase;

[0041] - 5%-20%, preferably 7%-15% glucose oxidase; and

[0042] - 1%-10%, preferably 4%-7% lactoperoxidase.

[0043] According to one aspect of the present invention, the enzyme complex further comprises lysozyme.

[0044] Another aspect of the present invention relates to an oral composition for the treatment of dental plaque and bacterial biofilm of dental plaque and for the natural whitening of tooth enamel, comprising the above enzyme complex.

[0045] According to one aspect of the invention, the oral composition comprises 1.4% to 3%, preferably 2.0% to 2.5% of the enzyme complex.

[0046] According to another aspect of the present invention, the oral composition further comprises lactoferrin.

[0047] According to yet another aspect of the present invention, the oral composition further comprises:

[0048] - 20%-60%, preferably 30%-50% sorbitol;

[0049] -1%-20%, preferably 5%-10% xylitol;

[0050] - 0-3%, preferably 1%-2% polysorbate 80;

[0051] - 0-3%, preferably 1%-2% polysorbate 20;

[0052] - 0-0.5%, preferably 0.1%-0.2% saccharin sodium;

[0053] - 0-5%, preferably 1%-2% colloidal silicon oxide;

[0054] - 0-3%, preferably 1%-2% dimethicone;

[0055] - 0-5%, preferably 1%-2% mint flavor and / or mint essential oil;

[0056] - 0.1%-0.5%, preferably 0.1%-0.3% zinc citrate;

[0057] - 0-0.5%, preferably 0.1%-0.2% potassium thiocyanate;

[0058] - 0-0.5%, preferably 0.1%-0.2% of preservatives; and

[0059] - Water, in an amount sufficient to make up to 100%.

[0060] According to one aspect of the present invention, the oral composition is in the form of a paste, liquid, gel, mouthwash, mist, spray, capsule, tablet or chewing gum.

[0061] As described in detail below, the glycolytic enzyme system of the present invention significantly reduces the toxicity of Streptococcus mutans by combining the multiple antibacterial and bactericidal effects of anti-caries Streptococci. This enzyme complex acts on dietary sucrose to deprive the bacteria of their substrate while directly degrading the cell walls of these Streptococci.

[0062] Since sucrase, preferably invertase or isomaltase, hydrolyzes the β-2,6 and / or β-2,1 bonds of sucrose, the enzyme complex of the present invention advantageously reduces the amount of sucrose available in the oral cavity by converting sucrose into monomers of monosaccharides (glucose) and fructose, thereby depriving cariogenic bacteria of metabolic substrates, significantly reducing the production of soluble and insoluble glucans, and thus helping to reduce bacterial biofilm.

[0063] Because the glucanohydrolase mixture hydrolyzes (1-3) and (1-6)-α-D-glucan bonds, which are key structural and functional components of the S. mutans biofilm matrix, the enzyme complex of the present invention advantageously prevents the formation of bacterial biofilms and breaks down existing biofilms. Disruption of bacterial biofilms further reduces the formation of dental plaque, which has a foul odor and can yellow teeth in the presence of colorants such as tobacco or coffee and tea. Soluble glucans are converted into monosaccharide dimers or trimers by the glucanohydrolase mixture and then converted into monosaccharide monomers by the action of a glycoside hydrolase, preferably an amyloglucosidase, such as an amyloglucosidase, which hydrolyzes the (1-6)-α-D-glucosidic bonds of the monosaccharide dimers and trimers.

[0064] The preferred combination of a glucanase and a mutanase, more preferably derived from different fungi or bacteria, or a chimeric glucanase (glucanase-mutanase), has shown very satisfactory results in preventing mutans S. mutans biofilm formation and breaking down existing biofilms within a few minutes at nanomolar concentrations. This combination significantly increases the sensitivity of biofilms to antimicrobial agents and antibacterial agents, such as lysozyme, highlighting its great potential in combating dental caries associated with acidic biofilms.

[0065] The residual glycoside derivatives produced by the degradation of glucans in dental plaque are subsequently converted into monosaccharide monomers by glycoside hydrolases that hydrolyze (1-6)-α-D-glucosides. These monosaccharide monomers are then converted into D-glucono-δ-lactone and, more importantly, hydrogen peroxide by oxidoreductases. In fact, oxidoreductases form hydrogen peroxide from monosaccharide monomers. The formation of hydrogen peroxide is beneficial because it has a bactericidal effect, actively participating in the elimination of Streptococcus mutans, and has a natural whitening effect on tooth enamel, contributing to a more aesthetically pleasing tooth appearance.

[0066] These beneficial effects are advantageously enhanced by the fact that the enzyme complex of the present invention provides a multiple source of hydrogen peroxide in the oral cavity by in situ conversion of sucrose, monosaccharide dimers and trimers, soluble glucans, and insoluble glucans into monosaccharide monomers, namely glucose. This regularly available glucose is converted by the action of glucose oxidase to produce hydrogen peroxide that directly contacts the teeth in a regular, gentle, and sustained manner, thereby achieving optimal results in terms of oral hygiene and enamel whitening. Due to the real-time conversion of glucose, the amount of glucose present in the oral cavity at any one time is advantageously kept at a much lower level, thus being much less harmful than when exogenous glucose is added.

[0067] The in situ generated hydrogen peroxide by the enzyme complex of the present invention uses much lower amounts of hydrogen peroxide than is typically supplied exogenously for tooth whitening, but remains in the oral cavity for a longer period of time, thereby enabling satisfactory, gradual whitening without the common problems associated with large amounts of exogenous hydrogen peroxide supply, namely: demineralization of tooth hard tissues, mucosal irritation, reaction with sealing materials, premature wear of tooth enamel with tooth hypersensitivity, and weakening of teeth.

[0068] Furthermore, in the presence of hydrogen peroxide, peroxidase advantageously converts thiocyanate naturally present in saliva into hypothiocyanate, a potent bactericidal agent, further enhancing the effect of the enzyme complex of the present invention on dental health. To enhance this effect, exogenous thiocyanate may also be added.

[0069] The addition of a sucrase, preferably an invertase or an isomaltase, to the enzyme complex of the invention is advantageous because this enzyme breaks down any starch and glycogen present in the mouth into monosaccharide dimers (maltose), which are then converted into monosaccharide monomers by the action of α-amylase naturally present in saliva.

[0070] The addition of lysozyme to the enzyme complex of the present invention is advantageous because lysozyme hydrolyzes the polysaccharide cell walls of bacteria and exhibits potent bactericidal activity even after the bacterial biofilm produced by S. mutans has disappeared, actively contributing to oral hygiene. In fact, lysozyme enhances the bactericidal activity of the enzyme complex of the present invention, making it more effective "after" the bacterial biofilm has disappeared.

[0071] In summary, through the synergistic action of its ingredients, the enzyme complex of the present invention provides the following beneficial effects while respecting the overall health of the consumer and without side effects:

[0072] - It has an anti-caries effect by inhibiting the acidic conversion of sugars in the diet, thus having a caries-preventing effect;

[0073] - It has a destructive effect on the bacterial biofilm of dental plaque, thus having the effect of preventing periodontal disease and tartar;

[0074] - Under the stimulation of the salivary lactoperoxidase system, it reduces pathogenic bacteria and has a purifying effect on the oral microbiome;

[0075] - Provides a gentle and long-lasting whitening effect on tooth enamel through the natural production of hydrogen peroxide.

[0076] The addition of lactoferrin to oral compositions containing the enzyme complex advantageously leads to improved eradication of mutans Streptococcus biofilms by sequestering the iron necessary for the survival of cariogenic Streptococci, which is bound by polysaccharides in the microbial envelope. This contributes to the aforementioned synergistic effect. Indeed, lactoferrin can lyse the bacterial membranes of many organisms, including acidogenic Streptococci, thereby significantly enhancing the body's immune defenses.

[0077] The following description will provide a better understanding of the invention and its various applications. DETAILED DESCRIPTION

[0078] Streptococcus mutans utilizes dietary sucrose to synthesize extracellular polysaccharides (EPS) to promote bacterial biofilm formation. Recent studies have demonstrated that the EPS matrix, composed of glucose polymers linked by branches of α-(1-6) linkages (dextrose) and α-(1-3) linkages (mutans), plays a key role in regulating the formation and virulence of cariogenic biofilms by influencing the physical and biochemical properties of the biofilm. Cariogenic biofilms promote microbial aggregation and adhesion, accelerate bacterial cell-to-cell binding and binding to apatite surfaces, thereby regulating the initial steps of cariogenic biofilm development and proliferation and promoting the formation of mature dental plaque. Furthermore, as a reserve energy source, the EPS matrix protects microorganisms from adverse influences, influences the diffusion of substances into and out of the biofilm, and helps concentrate metal ions and other physiological nutrients in the microenvironment. Acting as a physical barrier, cariogenic biofilms hinder the diffusion of antibiotics and host-induced antimicrobial factors into the deepest layers of the biofilm, thereby enhancing the resistance of microorganisms within the biofilm. To this end, Streptococcus mutans produces the glucanase DexA, which hydrolyzes the α-1,6 linkage of dextran and produces isomaltooligosaccharides of varying sizes. DexA is crucial in bacterial biofilm formation and is thought to be involved in the pathogenic dental plaque ecosystem: DexA hydrolyzes glucans, which serve as potential storage polysaccharides, providing nutrients for bacterial metabolism and controlling the amount and content of extracellular glucans, making them more adherent. However, the effects of DexA glucanase can be counteracted by exogenously supplied glucanases.

[0079] In the research conducted to select suitable enzyme complexes for the present invention, it was specifically attempted to identify enzymes that selectively target and degrade EPS. In this study, it was noted that when glucanase was supplied exogenously in combination with mutanase, it showed higher activity in inhibiting biofilm formation and destroying pre-formed biofilms. But even more interesting was the use of mutanases from different sources—one from an exo-acting fungal source and another from an endo-acting bacterial source, such as Streptococcus, or a chimeric glucanase (glucanase-mutanase)—which prevented the formation of S. mutans biofilms in a very satisfactory manner and broke down existing biofilms at nanomolar concentrations within a few minutes. This combination significantly increased the sensitivity of biofilms to antimicrobials and lysozymes, highlighting its great potential in combating caries associated with acidic biofilms.

[0080] This innovative effect on bacterial biofilms, combined with invertases, isomaltase, or other enzymes from the sucrase family that degrade sucrose, thereby depriving caries-causing bacteria of their metabolic substrate, significantly reduces glucan production, which in turn contributes to a significant reduction in bacterial biofilms. The resulting glycoside derivatives generate monosaccharide monomers that, under the action of oxidoreductases, produce hydrogen peroxide, which triggers the highly bactericidal salivary lactoperoxidase reaction, leading to the disappearance of bacterial biofilms.

[0081] Furthermore, this small but continuous production of hydrogen peroxide naturally and gradually whitens the tooth enamel, thus avoiding any tooth sensitivity issues.

[0082] Hereinafter, the enzyme complex of the present invention will be described in detail.

[0083] In this article, enzyme complex refers to a stable combination of multiple different proteins. These proteins, based on a specific structure, have various enzymatic activities that act on the target substrate in an orderly and coordinated manner, resulting in synergistic activity.

[0084] The enzyme complex of the present invention is intended for use in the oral cavity to improve oral health and whiten tooth enamel.

[0085] It contains a combination of the following ingredients:

[0086] - sucrase, in particular capable of hydrolyzing β-2,6 and / or β-2,1 bonds of sucrose;

[0087] - a glucanohydrolase mixture, particularly capable of hydrolyzing (1-3)-α-D-glucan and (1-6)-α-D-glucan;

[0088] - glycoside hydrolases, particularly capable of hydrolyzing the (1-6)-α-D-glycosidic bonds of monosaccharide dimers and trimers;

[0089] - oxidoreductases, in particular capable of breaking down monosaccharide monomers to form hydrogen peroxide;

[0090] - Peroxidase, in particular capable of converting thiocyanate into hypothiocyanate in the presence of hydrogen peroxide.

[0091] The sucrase capable of hydrolyzing the β-2,6 and / or β-2,1 bonds of sucrose is preferably an invertase or an isomaltase, which degrades sucrose into glucose and fructose.

[0092] The glucan hydrolase mixture degrades dental plaque into dimers or trimers of monosaccharides. This is preferably a mixture of mutanase and glucanase. Glucanase degrades soft dental plaque by converting soluble dextran into 1,6 monosaccharide dimers or trimers of glycoside polymers, while mutanase degrades hard dental plaque by converting insoluble mutan into 1,3 monosaccharide dimers or trimers of glycoside polymers.

[0093] In the glucan hydrolase mixture, preferred are mutanases with exo- and endo-activity. As mutanases with exo-activity, mutanases obtained from fungi, in particular Trichoderma Harzianum, can be used. As mutanases with endo-activity, mutanases obtained from bacteria, in particular Paracoccus Mutanolyticus, Paenibacillus, or Streptomyces can be used.

[0094] In the glucan hydrolase mixture, a chimeric glucanase mixture comprising a glucanase and a mutanase linked by genetic engineering may also be used instead of a mixture of a mutanase and a glucanase.

[0095] Glycoside hydrolases are enzymes that reduce dimers or trimers of monosaccharides to glycoside monomers, i.e., glucose. They are preferably amyloglucosidase, more preferably amyloglucosidase-α-1,6-glucosidase. Amyloglucosidase converts glycoside derivatives into glucose.

[0096] Oxidoreductases are enzymes that convert glucose into hydrogen peroxide. Preferably, they are glucose oxidases (also known as glucose-oxidases). Glucose-oxidases convert glucose into D-glucono-delta-lactone and hydrogen peroxide. D-glucono-delta-lactone is an acidity regulator. In the oral cavity, it is partially converted into gluconic acid.

[0097] In the presence of hydrogen peroxide, peroxidase converts thiocyanate into hypothiocyanate according to the following reaction:

[0098] SCN-+H2O2→OSCN-+H2O

[0099] The peroxidase is preferably lactoperoxidase.

[0100] Hypothiocyanate is a powerful antiseptic that limits the growth of bacteria in the mouth.

[0101] The enzyme complex of the present invention preferably has the following components (the percentages indicated are by volume):

[0102] - Invertase: 10%-30%, preferably 15%-25%;

[0103] - Mutantase: 10%-40%, preferably 15%-30%;

[0104] - Glucanase: 1%-10%, preferably 4%-7%;

[0105] - Amyloglucosidase: 20%-60%, preferably 30%-50%;

[0106] - Glucose oxidase: 5%-20%, preferably 7%-15%; and

[0107] - Lactoperoxidase: 1%-10%, preferably 4%-7%.

[0108] The table below shows a control formulation CET of an enzyme complex with a high amyloglucosidase content and without mutanase, and three exemplary formulations CE1-3 of the enzyme complex of the present invention. The percentages indicated are percentages by volume.

[0109] Table 1

[0110]

[0111] In a six-month clinical trial conducted by the applicant, with the participation of a research laboratory led by a medical doctor, the results of the formula CET were unsatisfactory. In fact, the presence of dental plaque and a very slight whitening of the enamel of the teeth (<1 shade) were observed in the subjects who participated in the six-month study and used the formula, compared to the control subjects.

[0112] The results of formula CE1 were more favorable than those of formula CET. In fact, in the subjects who participated in the six-month study and used this formula, a lower amount of dental plaque was observed in the mouth, as well as a significant whitening of the enamel of the teeth (+1 shade whiter) compared to the control subjects.

[0113] The results of formula CE2 were more favorable than those of formula CET. In fact, a reduction in the amount of dental plaque in the mouth and a significant whitening of the enamel of the teeth (+1 shade whiter) were observed in the subjects who used this formula for six months compared to the control subjects.

[0114] The results of formula CE3 were even more favorable than those of formulas CE1 and CE2. In fact, in the subjects who participated in the six-month study and used this formula, a near-plaque-free oral cavity and a significant whitening of the enamel of the teeth (+2 shades whiter) were observed compared to the control subjects.

[0115] The enzyme complex of the present invention may further comprise lysozyme.

[0116] The enzyme complex preferably comprises 0.1%-0.4% by weight, more preferably 0.2% by weight of lysozyme. It may also preferably comprise 0-0.4% by weight of sucrase, preferably invertase or isomaltase, more preferably 0.2% by weight.

[0117] The enzyme complexes of the present invention can be used in oral compositions to promote oral health and whiten tooth enamel.

[0118] Such oral compositions may be in the form of, for example, a paste, liquid, gel, mouthwash, mist, spray, capsule, tablet or chewing gum.

[0119] The oral composition of the present invention preferably has the following components (the enzyme complex is the enzyme complex described above, and the percentages indicated are by weight):

[0120] - Enzyme complex: 1.4%-3%, preferably 2.0%-2.5%;

[0121] - Sorbitol: 20%-60%, preferably 30%-50%;

[0122] - Xylitol: 1%-20%, preferably 5%-10%;

[0123] - Polysorbate 80: 0-3%, preferably 1%-2%;

[0124] - Polysorbate 20: 0-3%, preferably 1%-2%;

[0125] - saccharin sodium: 0-0.5%, preferably 0.1%-0.2%;

[0126] - Colloidal silicon dioxide: 0-5%, preferably 1%-2%;

[0127] - dimethicone: 0-3%, preferably 1%-2%;

[0128] - mint flavor and / or mint essential oil: 0-5%, preferably 1%-2%;

[0129] - Zinc citrate: 0.1%-0.5%, preferably 0.1%-0.3%;

[0130] - Potassium thiocyanate: 0-0.5%, preferably 0.1%-0.2%;

[0131] - Preservatives: 0-0.5%, preferably 0.1%-0.2%;

[0132] - Water: Use enough to make up to 100%.

[0133] The preservative is preferably citric acid.

[0134] The oral composition of the present invention may further comprise lactoferrin, preferably 0-0.4% by weight, more preferably 0.1% by weight.

[0135] It may also contain hydrogen peroxide, such as 0-0.1%, and a purple colorant for a more immediate whitening effect.

[0136] The following table shows an example of a control formulation COT of an oral composition for oral-dental use without an enzyme complex, and three example formulations CO1-3 of oral compositions according to the invention containing the enzyme complex CE3. The percentages indicated are percentages by weight.

[0137] Table 2

[0138]

[0139]

[0140] The efficacy of these various formulations was tested on Streptococcus salivarius, a Gram-positive coccus from the same genus Viridans as Streptococcus mutans that also forms biofilms in the presence of sucrose and is known to have the same susceptibility to antiseptics and antimicrobials as Streptococcus mutans. The efficacy of these formulations was compared to a placebo containing no enzyme complex of the present invention and to a mouthwash solution containing 0.12% chlorhexidine.

[0141] In an in vitro clinical trial conducted by the applicant, involving another analytical laboratory led by a doctor of pharmacy, using the enzyme complex CE3, the results for the COT formulation were unsatisfactory. In fact, compared with placebo, the COT formulation, and chlorhexidine, a high number of S. salivarius was observed in the placebo, a high number of S. salivarius was still observed in the COT formulation, and no S. salivarius was observed in the chlorhexidine-based formulation.

[0142] The results for formulation CO1 were more favorable than those for formulation COT. Indeed, compared with placebo, formulation CO1, and chlorhexidine, a high number of S. salivarius was observed with placebo, a low number with formulation CO1, and no S. salivarius was observed with the chlorhexidine-based formulation.

[0143] The results with formulation CO2 were more promising than those with formulations COT and CO1. Indeed, compared with placebo, formulation CO2, and chlorhexidine, a high number of S. salivarius was observed with placebo, no S. salivarius was observed with formulation CO2, and no S. salivarius was observed with the chlorhexidine-based formulation.

[0144] The results with formulation CO3 were equally promising as those with formulation CO2. Indeed, compared with placebo, formulation CO3, and chlorhexidine, a high number of S. salivarius were observed with placebo, no S. salivarius were observed with formulation CO3, and no S. salivarius were observed with the chlorhexidine-based formulation.

[0145] Therefore, for the oral composition for oral-dental use comprising the enzyme complex according to enzyme formula CE3, although clinical effects were observed at an enzyme complex dosage below 2%, the bactericidal effect against Streptococci was fully effective at an enzyme complex dosage of 2% or more.

Claims

1. An enzyme complex for the treatment of dental plaque and bacterial biofilm of dental plaque and for the natural whitening of tooth enamel, characterized in that The enzyme complex contains the following compounds: - sucrase; a glucan hydrolase mixture comprising a glucanase, a mutanase having exo-activity and obtained from a fungus, and a mutanase having endo-activity and obtained from a bacterium; -glycoside hydrolases; -oxidoreductases; -Peroxidase.

2. The enzyme complex according to claim 1, characterized in that The glucan hydrolase mixture comprises a mixture of chimeric glucanases comprising a glucanase and a mutanase linked by genetic engineering.

3. The enzyme complex according to claim 1 or 2, characterized in that The exo-active mutanase is obtained from the fungus Trichoderma Harzianum.

4. The enzyme complex according to any one of the preceding claims, characterized in that The mutanase with endo-activation activity is obtained from Paracoccus Mutanolyticus, Paenibacillus or Streptomyces bacteria.

5. The enzyme complex according to any one of the preceding claims, characterized in that The sucrase is isomaltase or invertase.

6. The enzyme complex according to any one of the preceding claims, characterized in that The glycoside hydrolase is amyloglucosidase, preferably amyloglucosidase.

7. The enzyme complex according to any one of the preceding claims, characterized in that The oxidoreductase is glucose oxidase.

8. The enzyme complex according to any one of the preceding claims, characterized in that The peroxidase is lactoperoxidase.

9. The enzyme complex according to any one of the preceding claims, characterized in that The enzyme complex contains the following compounds: - a sucrase, preferably an invertase or an isomaltase; - glucanase; a mutanase mixture comprising a mutanase having exo-activity and obtained from a fungus and a mutanase having endo-activity and obtained from a bacterium; - amyloglucosidase, preferably amyloglucosidase-α-1,6-glucosidase; - glucose oxidase; and -Lactoperoxidase.

10. The enzyme complex according to claim 8, characterized in that The enzyme complex comprises the following compounds, the percentages indicated are by volume: - 10%-30%, preferably 15%-25% sucrase; - 1%-10%, preferably 4%-7% glucanase; - 10% to 40%, preferably 15% to 30%, of a mixture comprising a mutanase having exo-activity and obtained from a fungus and a mutanase having endo-activity and obtained from a bacterium; - 20%-60%, preferably 30%-50% amyloglucosidase, preferably amylo-α-1,6-glucosidase; - 5%-20%, preferably 7%-15% glucose oxidase; and - 1%-10%, preferably 4%-7% lactoperoxidase.

11. An oral composition for the treatment of dental plaque and dental plaque bacterial biofilm and for the natural whitening of tooth enamel, characterized in that It comprises an enzyme complex according to any one of the preceding claims.

12. The oral composition according to claim 11, characterized in that It contains 1.4%-3%, preferably 2.0%-2.5% of the enzyme complex.

13. The oral composition according to claim 11 or 12, characterized in that It also contains lactoferrin.

14. The oral composition according to any one of claims 11 to 13, characterized in that Also includes: - 20%-60%, preferably 30%-50% sorbitol; -1%-20%, preferably 5%-10% xylitol; - 0-3%, preferably 1%-2% polysorbate 80; - 0-3%, preferably 1%-2% polysorbate 20; - 0-0.5%, preferably 0.1%-0.2% saccharin sodium; - 0-5%, preferably 1%-2% colloidal silicon dioxide; - 0-3%, preferably 1%-2% dimethicone; - 0-5%, preferably 1%-2% mint flavor and / or mint essential oil; - 0.1%-0.5%, preferably 0.1%-0.3% zinc citrate; - 0-0.5%, preferably 0.1%-0.2% potassium thiocyanate; - 0-0.5%, preferably 0.1%-0.2% preservatives; - Water, in an amount sufficient to make up to 100%.

15. The oral composition according to any one of claims 11 to 14, characterized in that It comes in the form of a paste, liquid, gel, mouthwash, mist, spray, capsule, tablet, or chewing gum.

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