Reduction of pathogenic bacteria using arginine

Abstract

Methods of reducing pathogenic bacteria using arginine are disclosed. The present invention relates to the use of an oral care composition for improving the growth of beneficial oral bacteria in the oral cavity of a subject in need thereof and for reducing the presence of pathogenic oral bacteria in the oral cavity of a subject in need thereof, the oral care composition comprises: arginine in an effective amount to obtain an arginine concentration of 10-25 micromole / ml in the oral cavity; zinc oxide; and zinc citrate.

Description

[0001] This application is a divisional application of a Chinese patent application filed on December 17, 2019, with application number "201980085773.5" and invention name "Using arginine to reduce pathogenic bacteria". Technical Field

[0002] The present invention relates to the field of reducing pathogenic bacteria, and in particular to reducing pathogenic bacteria using arginine. Background Art

[0003] Current research in the intestinal microbiome has reflected the potential benefits of beneficial bacteria to overall health. It has been observed that promoting the growth of selected bacteria (such as Bifidobacteria and Lactobacilli species) has important benefits for gastrointestinal health and the overall health of people. A method currently being sought in the field of intestinal microbiology is prebiotics. Prebiotics are defined as indigestible substances that induce growth or bacteria that can help improve host health. These compounds serve as important selective nutritional tools that can effectively change the composition of microorganisms, improve the overall health of intestinal microbial groups and intestinal health. This shows that influencing the microbial community within the host may be of great significance in promoting better health. Interestingly, this method has not yet been applied to the oral microbiome to improve oral health.

[0004] The oral microbiome is composed of ~700 different microbial species, including both beneficial and pathogenic oral bacterial species.

[0005] Beneficial bacteria within the oral microbial community appear to play an important role in health, producing factors associated with oral health. Beneficial bacteria may cause one or more of the following effects due to their presence or metabolic activity: reducing the number or proportion of pathogenic oral bacteria; reducing inflammation and inflammatory processes; reducing the metabolic activity of pathogenic oral bacterial species; reducing the production of pathogenic oral bacterial species or inhibiting the virulence factors produced by oral pathogenic bacterial species; reducing or inhibiting biofilm formation; occupying niches that may otherwise be colonized by pathogens; limiting the ability of pathogens to adhere to oral surfaces; affecting the vitality, metabolic activity or growth of pathogens; reducing the ability of pathogens to produce virulence factors; degrading virulence factors produced by pathogens or oral microbiota; and / or weakening the host's response to pathogens. Certain species of oral bacteria can be beneficial in maintaining periodontal health. Without being bound by any theory, it is believed that beneficial oral bacteria can interfere with the colonization of pathogenic oral bacteria in biofilms through the oral epithelium and the oral cavity. For example, studies have shown that Streptococcus sanguinis, Streptococcus mitis, and Streptococcus salivarius have an inhibitory effect on the colonization of epithelial cells with A. actinomycetemcomitans in vitro (W. Teughels et al., J Dent Res 86(7), 611-617, 2007). It has also been shown using a canine model that the application of beneficial bacteria to the periodontal pocket after delayed root planing reduces the recolonization of the periodontal pocket by pathogenic bacteria (W. Teughels et al., J Dent Res, 86(11), 1078-1082, 2007). The beneficial bacteria S. sanguinis, S. mitis, and S. salivarius have also been shown to inhibit the production of the inflammatory cytokine interleukin-8 (IL-8) induced by Aggregatibacter actinomycetemcomitans by the human oral keratinocyte cell line HOK-18A, an inflammatory response that is associated with tissue destruction associated with periodontitis (I. Sliepen et al., J Dent Res 88(11), 1026-1030, 2009).

[0006] Pathogenic bacterial species are associated with diseases and conditions. Some oral pathogenic bacterial species (e.g., Porphyromonas gingivalis, Tannerella forsythia, and A. actinomycetemcomitans) are associated with the development of periodontal disease, such as periodontitis, gingivitis, necrotizing periodontitis, necrotizing gingivitis, and peri-implantitis. Certain oral pathogenic bacterial species are associated with tooth decay (e.g., Streptococcus mutans). The onset of such oral diseases and conditions is mainly due to an increase in the population of pathogenic bacteria on the tooth surface (caries bacteria) or in the gums (periodontal pathogens).

[0007] Current methods of controlling such oral diseases and conditions are to improve gum health by reducing dental plaque or to use chemical agents (e.g., fluoride) to promote enamel remineralization and avoid the harmful effects of cariogenic bacteria. Compositions and methods including the use of standard antimicrobial treatments are designed to promote good oral health by eliminating pathogenic bacteria. Since the microbial flora in the oral cavity also contains beneficial bacteria, it is also possible to eliminate beneficial bacteria by using standard antimicrobial treatments. Therefore, non-selective methods of reducing oral dental plaque may not be the best way to promote oral health.

[0008] Selectively promoting the growth of beneficial oral bacteria can provide an effective preventive method for oral health, such as preventing periodontitis and / or preventing dental caries. The method of promoting beneficial bacteria in the oral cavity changes the composition of dental plaque to a healthier state with fewer pathogens. Relative to the growth of pathogenic bacteria in the oral cavity (e.g., in a biofilm in the oral cavity), the composition and method of selectively promoting the growth of beneficial oral bacteria can reduce the total pathogenic oral bacteria load in the oral cavity. Relative to the growth of pathogenic bacteria in the oral cavity (e.g., in a biofilm in the oral cavity), the composition and method of selectively promoting the growth of beneficial oral bacteria can promote good oral health, and prevent or reduce the severity of diseases and conditions caused by pathogenic bacteria. In addition, identifying the method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic bacteria in the oral cavity (e.g., in a biofilm in the oral cavity) and identifying the method of promoting the composition of dental plaque to change to a healthier state with fewer pathogens is a useful method for identifying compounds and compositions that promote good oral health. Summary of the invention

[0009] Methods of selectively promoting beneficial oral bacteria and reducing the overall pathogenic oral bacterial load within an individual's oral cavity can be used to alter the dental plaque composition to a healthier state with fewer pathogens.

[0010] A method of reducing the total pathogenic oral bacterial load in an individual's oral cavity is provided. The method comprises the step of administering 1-50 mg of arginine to the oral cavity.

[0011] A method of reducing the total pathogenic oral bacterial load in an individual's oral cavity is provided. The method comprises the step of administering to the oral cavity an amount of arginine sufficient to achieve a concentration of at least 20-25 micromoles / ml or more in the oral cavity.

[0012] Also provided is a method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in an individual's oral cavity. The method comprises the step of administering 1-50 mg of arginine to the oral cavity.

[0013] Also provided is a method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm in the oral cavity of an individual. The method comprises the step of administering 1-50 mg of arginine to the oral cavity.

[0014] Also provided is a method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in an individual's oral cavity. The method comprises the step of applying to the oral cavity an amount of arginine sufficient to achieve a concentration of at least 20-25 micromoles / ml or more in the oral cavity.

[0015] Also provided is a method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm in the oral cavity of an individual. The method comprises the step of applying to the oral cavity an amount of arginine sufficient to achieve a concentration of at least 20-25 micromoles / ml or more in the oral cavity.

[0016] Also provided is a method for selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a bacterial population comprising beneficial oral bacteria and pathogenic oral bacteria. The method comprises the step of contacting the bacterial population with arginine at a concentration of at least 20-25 micromoles / ml or more.

[0017] Also provided is a method for selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm comprising both beneficial oral bacteria and pathogenic oral bacteria. The method comprises the step of contacting the biofilm comprising both beneficial oral bacteria and pathogenic oral bacteria with arginine at a concentration of at least 20-25 micromoles / ml or more.

[0018] A method for identifying a compound or composition that enhances the selective promotion of arginine on the growth of beneficial oral bacteria relative to pathogenic oral bacteria is provided. The method comprises the following steps: performing: a beneficial oral bacteria growth test assay; a pathogenic oral bacteria growth test assay; a beneficial oral bacteria growth control assay; and a pathogenic oral bacteria growth control assay. The beneficial oral bacteria growth test assay comprises culturing a beneficial oral bacteria species in a culture medium comprising arginine and a test compound or composition, and measuring the growth of the beneficial oral bacteria in the beneficial oral bacteria growth test assay. The pathogenic oral bacteria growth test assay comprises culturing a pathogenic oral bacteria species in a culture medium comprising arginine and a test compound or composition, and measuring the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth test assay. The beneficial oral bacteria growth control assay comprises culturing a beneficial oral bacteria species in a culture medium comprising arginine but not containing a test compound or composition, and measuring the growth of the beneficial oral bacteria in the beneficial oral bacteria growth control assay. The pathogenic oral bacteria growth control assay includes culturing pathogenic oral bacteria species in a medium containing arginine without a test compound or composition, and measuring the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay. The ratio of the growth of the beneficial oral bacteria in the beneficial oral bacteria growth test assay to the growth of the beneficial oral bacteria in the beneficial oral bacteria growth control assay is calculated, and the ratio of the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth test assay to the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay is calculated. The two ratios are compared. A ratio of the growth of the beneficial oral bacteria in the beneficial oral bacteria growth test assay to the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay that is higher than the ratio of the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth test assay to the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay indicates that the test compound or composition enhances the selective promotion of arginine on the growth of beneficial oral bacteria relative to pathogenic oral bacteria.

[0019] A method for identifying a compound or composition that enhances the selective promotion of the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm by arginine is provided. The method includes the steps of performing a dual-species biofilm test assay and a dual-species biofilm control assay. The dual-species biofilm test assay includes: co-culturing beneficial oral bacterial species and pathogenic oral bacterial species in a dual-species test biofilm containing arginine and a test compound or composition; quantifying the beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm; and comparing the number of beneficial oral bacteria in the dual-species test biofilm with the number of pathogenic oral bacteria to determine the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm. The dual-species biofilm control assay includes: co-culturing beneficial oral bacterial species and pathogenic oral bacterial species in a dual-species control biofilm containing arginine without a test compound or composition; quantifying the beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm; and comparing the number of beneficial oral bacteria in the dual-species control biofilm with the number of pathogenic oral bacteria to determine the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm. The beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm is compared to the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm. A higher beneficial oral bacteria-pathogenic oral bacteria load in the dual-species test biofilm compared to the beneficial oral bacteria-pathogenic oral bacteria load in the dual-species control biofilm indicates that the test compound or composition enhances the selective promotion of the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in the biofilm by arginine. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 Results of qPCR assays are shown. Beneficial and pathogenic bacteria increased fold when cultured with increasing arginine concentrations. DETAILED DESCRIPTION

[0021] The following description of the preferred embodiment is merely exemplary in nature and is in no way intended to limit the invention, its application, or uses.

[0022] As used herein, the expression "oral cavity" includes not only the cavity itself, but also the saliva, teeth, gums, periodontal pockets, cheeks, tongue, mucosa, tonsils, any implants, and any device or structure placed into the oral cavity. The tonsils provide a reservoir for the growth of anaerobic bacteria that can produce bad breath (tonsil stones).

[0023] As used herein, the phrase "healthy oral microbiota" refers to the population of microorganisms in the oral cavity when the oral cavity is in a non-disease state (e.g., when there is no periodontal disease, such as gingivitis, periodontitis, caries, peri-implantitis, peri-implant mucositis, necrotizing gingivitis and / or necrotizing periodontitis), i.e., a health-associated oral microbiota.

[0024] The term "beneficial oral bacteria" encompasses those bacteria that are present in the oral cavity in higher numbers or proportions in a healthy oral cavity, but are present in lower numbers or proportions in oral disease conditions (e.g., gingivitis, periodontitis, caries, peri-implantitis, peri-implant mucositis, necrotizing gingivitis, and / or necrotizing periodontitis). The term also includes bacteria from oral or non-oral sources that have a proven beneficial effect on oral health by preventing or treating oral diseases, which may already be present in the oral cavity or may be intentionally introduced into the oral cavity (e.g., as probiotics). Beneficial bacteria may, by virtue of their presence or metabolic activity: reduce the number or proportion of pathogenic oral bacteria; reduce inflammation and inflammatory processes; reduce the metabolic activity of pathogenic species; reduce the production of or inhibit virulence factors produced by pathogenic bacteria; reduce or inhibit biofilm formation; occupy niches that might otherwise be colonized by pathogens; limit the ability of pathogens to adhere to oral surfaces; affect the viability, metabolic activity, or growth of pathogens; reduce the ability of pathogens to produce virulence factors; degrade virulence factors produced by pathogens or the oral microbiota; and / or attenuate the host response to pathogenic species. Examples of beneficial oral bacteria include Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, Streptococcus sanguinis, Veillonella parvula, Capnocytophagasputigena, and Actinomyces naeslundii.

[0025] Examples of pathogenic oral bacteria include Streptococcus mutans, Prevotella intermedia, Porphyromonas gingivalis, Fusobacterium nucleatum, Tannerella forsythia, Aggregatibacter actinomycetemcomitans, and Streptococcus sobrinus.

[0026] As used herein, the term "oral care composition" refers to a composition that is delivered to the oral cavity surface. The composition may be a product that is not intended for systemic administration of a specific therapeutic agent, is not intended to be swallowed, but is maintained in the oral cavity for a sufficient time to contact substantially all of the tooth surfaces and / or oral tissues for oral active purposes during the normal course of use. Examples of such compositions include, but are not limited to, toothpaste or dentifrices, mouthwashes or rinses, topical oral gels, denture cleansers, and the like.

[0027] As used herein, unless otherwise specified, the term "dentifrices" refers to pastes, gels, or liquid formulations. In some embodiments, the dentifrice composition can be a combination of multiple pastes, multiple gels, or a paste and a gel. In some embodiments, the dentifrice composition can be a combination of multiple pastes, multiple gels, or a paste and a gel.

[0028] Bacteria require certain substances to enable them to grow, proliferate, be metabolically active and colonize. Certain substances can be selectively used by certain microorganisms, such as certain bacteria, to promote their growth, metabolic activity, proliferation and colonization, thereby directly or indirectly inhibiting the growth of other microorganisms.

[0029] Arginine has been found to have a greater effect of promoting the growth of beneficial oral bacteria relative to pathogenic bacteria, including in terms of bacterial growth, biofilm formation, and biofilm competitive growth. Arginine can be used in methods to reduce the total pathogenic oral bacterial load in the oral cavity. Arginine can be used in methods to change the composition of dental plaque to a healthier state with fewer pathogens.

[0030] Evaluation of the effect of arginine on the growth of beneficial and pathogenic oral bacteria demonstrated that, although both beneficial and pathogenic oral bacteria can metabolize arginine, arginine stimulates the growth of beneficial oral bacteria at an increased growth rate, whereas no corresponding effect is observed in pathogenic oral bacteria. In some embodiments, arginine present at a concentration of 20-25 micromoles / ml preferentially promotes the growth of beneficial oral bacteria relative to pathogenic oral bacteria. Arginine present at a concentration of 20-25 micromoles / ml is particularly effective in preferentially promoting the growth of A. viscosus, S. sanguinis, and S. gordonii.

[0031] Evaluation of the effects of arginine on biofilm formation by beneficial oral bacteria and by pathogenic oral bacteria demonstrated that the presence of arginine promoted total biofilm growth by A. actinomycetemcomitans.

[0032] Studies using dual-species biofilms in which beneficial bacterial species are co-cultured with pathogenic bacteria have demonstrated that the presence of arginine results in an increase in beneficial bacteria relative to pathogenic bacteria in the biofilm. In some embodiments, arginine promotes the growth of beneficial bacteria while reducing the number of pathogenic bacteria such as Porphyromonas gingivalis (P. gingivalis), Prevotella intermedia (P. intermedia), Streptococcus mutans (S. mutans), and Streptococcus sobrinus (S. sobrinus).

[0033] Arginine as a selective nutrient has an important impact on altering the composition of the oral microbiota by providing a competitive advantage to beneficial bacteria, helping them to grow and outcompete pathogenic microorganisms in the oral cavity. Reducing the overall pathogen load in the oral cavity can have important oral health implications.

[0034] Arginine can be provided in oral care compositions to improve overall oral health. Arginine can be provided in oral care compositions, including but not limited to dentifrice and mouthwash formulations. Oral care compositions comprising arginine can be used to selectively promote in the oral cavity a) the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria, b) biofilm formation by beneficial oral bacteria relative to biofilm formation by pathogenic oral bacteria, and c) the growth of beneficial oral bacteria in biofilms relative to the growth of pathogenic oral bacteria in biofilms. Oral care compositions comprising arginine can be used to maintain and / or reestablish a healthy oral microbiome.

[0035] Methods for reducing the total pathogenic oral bacterial load in the oral cavity are provided. Methods for selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a bacterial population comprising beneficial oral bacteria and pathogenic oral bacteria are provided. In some embodiments, such methods selectively promote the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in such a bacterial population in the oral cavity of an individual. Methods for selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm comprising beneficial oral bacteria and pathogenic oral bacteria are provided. In some embodiments, such methods selectively promote the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm in the oral cavity of an individual. Methods for maintaining and / or reestablishing a healthy oral microbiome are provided. Methods for selectively promoting biofilm formation by beneficial oral bacteria relative to biofilm formation by pathogenic oral bacteria are provided in the oral cavity.

[0036] The method may include a step of treating or supplementing the oral cavity of an individual by applying an effective amount of arginine, preferably an oral care composition comprising arginine, or contacting the oral cavity of the individual with an effective amount of arginine, preferably an oral care composition comprising arginine, such as a dentifrice comprising arginine, a toothpaste comprising arginine, a gel comprising arginine, a tooth powder comprising arginine, a mouthwash comprising arginine, a mouthwash comprising arginine, a lozenge comprising arginine, a tablet comprising arginine, a spray comprising arginine, a chewing gum comprising arginine, or a film that can be completely or partially dissolved or insoluble and comprises arginine. In certain embodiments, the contact of the oral cavity with the oral care composition comprising arginine comprises the following steps: applying the oral care composition to the oral cavity using a brush, rinsing the oral cavity with the oral care composition in the form of a mouthwash, or spraying the oral care composition into the oral cavity using, for example, a sprayer. The individual or subject may be a mammal. In some embodiments, the individual or subject is a human. In some embodiments, the individual or subject is an animal, such as a companion animal (e.g., a cat or dog).

[0037] The oral cavity of the individual is contacted with an effective amount of arginine. The effective amount of arginine selectively promotes the growth of beneficial oral bacteria over the growth of pathogenic oral bacteria; and / or selectively promotes biofilm formation by beneficial oral bacteria over biofilm formation by pathogenic oral bacteria; and / or selectively promotes the growth of beneficial oral bacteria in a biofilm over the growth of pathogenic oral bacteria in a biofilm; and / or maintains and / or reestablishes a healthy oral microbiome.

[0038] In some embodiments, the oral cavity of an individual is contacted with arginine in an amount sufficient to provide arginine in the oral cavity at a concentration of at least 5 micromoles / ml, in some embodiments at least 10 micromoles / ml, in some embodiments at least 20 micromoles / ml, in some embodiments at least 25 micromoles / ml or more, in some embodiments about 20-25 micromoles / ml, in some embodiments about 0.05-0.1 mg / ml, in some embodiments about 0.1-1.0 mg / ml or more, in some embodiments about 1.0-5.0 mg / ml, in some embodiments about 5-10 mg / ml, in some embodiments about 10-15 mg / ml, in some embodiments about 1 mg / ml, about 2 mg / ml, about 3 mg / ml, about 4 mg / ml, about 5 mg / ml, about 6 mg / ml, about 7 mg / ml, about 8 mg / ml, about 9 mg / ml, about 10 mg / ml, about 11 mg / ml, about 12 mg / ml, about 13 mg / ml, about 14 mg / ml, or about 15 mg / ml.

[0039] In some embodiments, the oral cavity of the individual is contacted with 0.1-50 mg arginine, in some embodiments, 0.25-50 mg arginine, in some embodiments, 0.5-50 mg arginine, in some embodiments, 1-50 mg arginine, in some embodiments, 5-50 mg arginine, in some embodiments, 10-40 mg arginine, in some embodiments, 1 mg, 2 mg, 3 mg, 4 mg, 5 mg, 6 mg, 7 mg, 8 mg, 9 mg, 10 mg, 11 mg, 12 mg, 1 3mg, 14mg, 15mg, 16mg, 17mg, 18mg, 19mg, 20mg, 21mg, 22mg, 23mg, 24mg, 25mg, 26mg, 27mg, 28mg, 29mg, 30mg, 31mg, 32mg, 33mg, 34mg, 35mg, 36mg, 37mg, 38mg, 39mg, 40mg, 41mg, 42mg, 43mg, 44mg, 45mg, 46mg, 47mg, 48mg, 49mg or 50mg arginine.

[0040] In some embodiments, the pathogenic oral bacteria include one or more pathogenic oral bacterial species selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus.

[0041] In some embodiments, the beneficial oral bacteria include one or more beneficial oral bacteria species selected from the group consisting of Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis.

[0042] In some embodiments, one or more pathogenic oral bacterial species selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus are identified as being present in the oral cavity of the individual.

[0043] In some embodiments, one or more beneficial oral bacterial species selected from the group consisting of Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis are identified as being present in the oral cavity of the individual.

[0044] The methods provided herein can be used as a method of preventing one or more of gingivitis, periodontitis, peri-implantitis, peri-implant mucositis, necrotizing gingivitis, necrotizing periodontitis, and dental caries.

[0045] In some embodiments, the method selectively promotes biofilm formation by beneficial oral bacteria relative to biofilm formation by pathogenic oral bacteria after incubation with the beneficial oral bacteria and pathogenic oral bacteria for 48 hours.

[0046] Some embodiments provide methods comprising administering an effective amount of arginine to the oral cavity of an individual. An oral composition comprising arginine is used to apply arginine to the oral cavity. In some embodiments, the oral composition comprises arginine and a zinc ion source. In some embodiments, the oral composition comprises arginine, a zinc ion source, and a fluoride ion source. The zinc ion source can include zinc oxide particles, such as zinc oxide particles of 1 to 7 microns. The zinc ion source can be selected from zinc citrate, zinc sulfate, zinc silicate, zinc lactate, zinc phosphate, zinc oxide, or a combination thereof. The zinc ion source can be a combination of zinc oxide and zinc citrate. The fluoride source can be stannous fluoride.

[0047] The oral care composition comprises arginine or a salt thereof. In some embodiments, arginine is L-arginine or a salt thereof. Suitable salts include salts known in the art, which are pharmaceutically acceptable salts, generally considered to be physiologically acceptable in the amount and concentration provided. Physiologically acceptable salts include salts derived from pharmaceutically acceptable inorganic or organic acids or bases, such as acid addition salts formed by acids forming physiologically acceptable anions, such as hydrochlorides or bromide salts, and base addition salts formed by bases forming physiologically acceptable cations, such as base addition salts derived from alkali metals (such as potassium and sodium) or alkaline earth metals (such as calcium and magnesium). Physiologically acceptable salts can be obtained using standard procedures known in the art, such as by reacting a compound (such as an amine) with a suitable acid providing a physiologically acceptable anion. In some embodiments, arginine is partially or entirely in salt form, such as arginine phosphate, arginine hydrochloride or arginine bicarbonate. In some embodiments, arginine is present in an amount corresponding to 0.1% to 15%, such as 0.1% to 10% by weight, such as 0.1 to 5% by weight, such as 0.5% to 3% by weight, about, for example, 1%, 1.5%, 2%, 3%, 4%, 5% or 8% of the total weight of the composition, wherein the weight of the arginine is calculated in free form. In some embodiments, arginine is present in an amount corresponding to about 0.5% to about 20% by weight of the total weight of the composition, about 0.5% to about 10% by weight of the total weight of the composition, such as about 1.5%, about 3.75%, about 5% or about 7.5% by weight, wherein the weight of the arginine is calculated in free form. In some embodiments, arginine is present in an amount of 0.5% to 10% by weight, or 0.5% to 3% by weight, or 1% to 2.85% by weight, or 1.17% to 2.25% by weight, or 1.4% to 1.6% by weight, or 0.75% to 2.9% by weight, or 1.3% to 2% by weight, or about 1.5% by weight, based on the total weight of the composition. Typically, arginine is present in an amount of up to 5% by weight, further optionally 0.5% to 5% by weight, and further optionally 2.5% to 4.5% by weight, based on the total weight of the oral care composition. In some embodiments, arginine is present in an amount of 0.1% to 6.0% by weight (e.g., about 1.5% by weight), or about 4.5% to 8.5% by weight (e.g., 5.0%), or 3.5% to 9% by weight, or 8.0% by weight. In some embodiments, arginine is present in a dentifrice, for example, at about 0.5% to 2% by weight, such as at about 1% in a mouthwash.

[0048] In some embodiments, the oral care composition comprises a combination of arginine and ZnO. Such oral care compositions may optionally further comprise a fluoride source. In some embodiments, the ZnO particles may have an average particle size of 1 to 7 microns. In some embodiments, the ZnO particles have an average particle size of 5 microns or less. In some embodiments, in addition to zinc oxide, the composition may also comprise other metal oxides, such as stannous oxide, titanium oxide, calcium oxide, copper oxide and iron oxide or a mixture thereof. In some embodiments, suitable zinc oxide particles for oral care compositions have a particle size distribution of, for example, 3 to 4 microns, or alternatively, a particle size distribution of 5 to 7 microns, alternatively, a particle size distribution of 3 to 5 microns, alternatively, a particle size distribution of 2 to 5 microns, or alternatively, a particle size distribution of 2 to 4 microns. Zinc oxide may have a particle size of a median particle size. Suitable particles may have, for example, a median particle size of 8 microns or less, alternatively, a median particle size of 3 to 4 microns, alternatively, a median particle size of 5 to 7 microns, alternatively, a median particle size of 3 to 5 microns, alternatively, a median particle size of 2 to 5 microns, or alternatively, a median particle size of 2 to 4 microns. In another aspect, the particle size is an average (mean) particle size. In one embodiment, the average particle accounts for at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, or at least 40% of the total metal oxide particles in the oral care composition of the present invention. The particles may be present in an amount of up to 5% by weight based on the total weight of the oral care composition, for example, in an amount of 0.5% to 5% by weight, preferably up to 2% by weight, more preferably 0.5% to 2% by weight, more preferably 1% to 2% by weight, or in some embodiments 2.5% to 4.5% by weight. In some embodiments, the source of the zinc oxide particles and / or the form in which they can be incorporated into the oral care composition is selected from one or more of a powder, a nanoparticle solution or suspension, or encapsulated in a polymer or bead. The zinc oxide particles can be selected to achieve entrapment of dentin particles. The particle size distribution can be measured using a Malvern particle size analyzer model Mastersizer 2000 (or a comparable model) (Malvern Instruments, Inc., Southborough, Mass.), in which a helium-neon gas laser beam is projected through a transparent cell containing silica (such as, for example, silica hydrogel particles suspended in an aqueous solution). The light that hits the particles is scattered at an angle inversely proportional to the particle size. A photodetector array measures the amount of light at several predetermined angles. An electrical signal that is positively correlated with the measured light flux value is then processed by a microcomputer system relative to a scattering pattern predicted from theoretical particles defined by the refractive index of the sample and the aqueous dispersant to determine the particle size distribution of the metal oxide.It should be understood that other methods of measuring particle size are known in the art, and based on the teachings set forth herein, the skilled artisan will understand how to calculate the median particle size, average particle size, and / or particle size distribution of the metal oxide particles.

[0049] In some embodiments, the oral care composition comprises arginine in combination with a zinc ion source selected from zinc oxide, zinc citrate, zinc lactate, zinc chloride, zinc acetate, zinc gluconate, zinc glycinate, zinc sulfate, sodium zinc citrate, zinc silicate, zinc phosphate, and combinations thereof. Such oral care compositions may optionally further comprise a fluoride source. Examples of effective amounts of zinc ions are amounts of zinc effective to inhibit erosion, such as 0.005%-5% zinc, e.g., 0.01%-0.05% for a mouthwash, or 0.1%-3% for a dentifrice, e.g., a dentifrice comprising 1% to 3% zinc citrate.

[0050] In some embodiments, the oral care composition comprises a combination of arginine and two or more zinc salts, at least one of which is zinc oxide and at least one of which is zinc citrate ... Such oral care compositions may optionally further comprise a fluoride source. In some embodiments, the weight ratio of zinc oxide to zinc citrate is 1.5: 1 to 4.5: 1, 1.5: 1 to 4: 1, 1.7: 1 to 2.3: 1, 1.9: 1 to 2.1: 1, or about 2: 1. In addition, corresponding molar ratios based on these weight ratios may be used. In some embodiments, the total concentration of zinc salts in the composition is 0.2 wt % to 5 wt %, or 0.5 wt % to 2.5 wt %, or 0.8 wt % to 2 wt %, or about 1.5 wt %, based on the total weight of the composition. In some embodiments, the molar ratio of arginine to total zinc salts is 0.05: 1 to 10: 1. In some embodiments, based on the total weight of the composition, the composition includes zinc oxide in an amount of 0.5% to 1.5% by weight and zinc citrate in an amount of 0.25% to 0.75% by weight. In some embodiments, based on the total weight of the composition, the composition may include zinc oxide in an amount of 0.75% to 1.25% by weight and zinc citrate in an amount of 0.4% to 0.6% by weight. In some embodiments, based on the total weight of the composition, the composition includes zinc oxide in an amount of about 1% by weight and zinc citrate in an amount of about 0.5% by weight. In some embodiments, based on the weight of the oral care composition, zinc oxide can be present in an amount of 0.75% to 1.25% by weight (e.g., 1.0% by weight), and zinc citrate is an amount of 0.25% to 1.0% by weight (e.g., 0.25% to 0.75% by weight, or 0.5% by weight). In some embodiments, zinc citrate is about 0.5% by weight. In some embodiments, zinc oxide is about 1.0% by weight.

[0051] One or more fluoride ion sources are optionally present in an amount that provides a clinically effective amount of soluble fluoride ions to the oral care composition. Fluoride ion sources can be used as anticaries agents, for example. Any oral acceptable granular fluoride ion source can be used, including stannous fluoride, sodium fluoride, potassium fluoride, potassium monofluorophosphate, sodium monofluorophosphate, ammonium monofluorophosphate, sodium fluorosilicate, ammonium fluorosilicate, indium fluoride, fluorinated amines such as olafluro (N'-octadecyl trimethylene diamine-N, N, N'-tri (2-ethanol)-dihydrofluoride), ammonium fluoride, titanium fluoride, hexafluorosulfate and combinations thereof. When present, fluoride can be, for example, about 25 to about 25,000ppm, for example, for consumer toothpaste (e.g., 1000-1500ppm, for example, about 1000ppm, for example, about 1450ppm) products, with a level of about 50 to about 5000ppm, about 750 to about 2,000ppm. In some embodiments, fluoride exists about 100 to about 1000, about 200 to about 500 or about 250ppm, 500 to 3000ppm fluoride ions. In some embodiments, the fluoride source provides 50 to 25,000ppm (e.g., 750-7000ppm, for example, 1000-5500ppm, for example, about 500ppm, 1000ppm, 1100ppm, 2800ppm, 5000ppm or 25000ppm) of fluoride ions. In some embodiments, the fluoride source is stannous fluoride. In some embodiments, the fluoride source is stannous fluoride, which provides an amount of 750-7000ppm (e.g., about 1000ppm, 1100ppm, 2800ppm, 5000ppm) of fluoride. In some embodiments, the fluoride source is stannous fluoride, which provides an amount of about 5000ppm of fluoride. In some embodiments, the fluoride source is sodium fluoride, which provides 750–2000ppm (e.g., about 1450ppm) of fluoride. In some embodiments, the fluoride source is selected from sodium fluoride and sodium monofluorophosphate, and it provides 1000ppm-1500ppm of fluoride. In some embodiments, the fluoride source is sodium fluoride or sodium monofluorophosphate, and it provides about 1450ppm of fluoride. In some embodiments, stannous fluoride is the only fluoride source. Fluoride ion sources can be added to the composition at a level of about 0.001% by weight to about 10% by weight, for example, about 0.003% by weight to about 5% by weight, 0.01% by weight to about 1% by weight, or about 0.05% by weight. % is added to the composition of the present invention at a level of. However, it should be understood that the weight of the fluoride salt providing an appropriate level of fluoride ions will obviously vary based on the weight of the counterions in the salt, and such an amount can be easily determined by a person skilled in the art.In some embodiments, the fluoride source is a fluoride salt (e.g., sodium fluoride (e.g., about 0.32 wt %) or sodium monofluorophosphate) present in an amount of 0.1 wt % to 2 wt % (0.1 wt %-0.6 wt %) of the total composition weight, e.g., 0.3%-0.4%, e.g., about 0.32% sodium fluoride.

[0052] The oral care compositions described herein may also include one or more other agents, such as those generally selected from the group consisting of abrasives, antiplaque agents, whitening agents, antibacterial agents, cleaning agents, flavoring agents, sweeteners, binders, surfactants, foam regulators, pH regulators, humectants, mouthfeel agents, colorants, tartar control (anticalculus) agents, polymers, saliva stimulants, nutrients, viscosity regulators, anti-sensitivity agents, antioxidants, and combinations thereof.

[0053] In some embodiments, the oral care composition comprises one or more abrasive particles, such as those that can be used as, for example, polishing agents. Any oral acceptable abrasive can be used, but the type, fineness (granularity) and amount of the abrasive should be selected so that tooth enamel is not excessively worn in the normal use of the composition. The example of usable abrasive particles includes abrasives, such as sodium bicarbonate, insoluble phosphates (such as orthophosphates, polymetaphosphates and pyrophosphates, including dicalcium orthophosphate dihydrate, calcium pyrophosphate, tricalcium phosphate, calcium polymetaphosphate and insoluble sodium polymetaphosphate), calcium phosphate (for example, dicalcium phosphate dihydrate), calcium sulfate, natural calcium carbonate (CC), precipitated calcium carbonate (PCC), silicon dioxide (for example, hydrated silicon dioxide or silica gel or in the form of precipitated silicon dioxide or mixed with aluminum oxide), iron oxide, aluminum oxide, aluminum silicate, calcined aluminum oxide, bentonite, other siliceous materials, perlite, plastic particles (for example, polyethylene), and combinations thereof. Natural calcium carbonate abrasive is usually ground limestone, which can be optionally refined or partially refined to remove impurities. The material preferably has an average particle size of less than 10 microns, such as 3-7 microns, such as about 5.5 microns. For example, small particle silicon dioxide can have an average particle size (D50) of 2.5–4.5 microns. Because natural calcium carbonate can contain a high proportion of relatively large particles that are not carefully controlled, this can unacceptably increase abrasiveness, so preferably no more than 0.01% by weight, preferably no more than 0.004% by weight of particles will not pass through a 325 mesh sieve. The material has a strong crystalline structure, and is therefore much harder and more abrasive than precipitated calcium carbonate. The tap density of natural calcium carbonate is, for example, between 1 and 1.5 g / cc, such as about 1.2, such as about 1.19 g / cc. There are different polymorphs in natural calcium carbonate, such as calcite, aragonite and vaterite, and calcite is preferred for the purpose of the present invention. Examples of commercially available products suitable for the present invention include Vicron? 25-11FG. Precipitated calcium carbonate has a different crystal structure than natural calcium carbonate. It is generally more brittle and more porous and therefore has lower abrasiveness and higher water absorption. For use in the present invention, the particles are relatively small, for example having an average particle size of 1-5 microns, and for example no more than 0.1% by weight, preferably no more than 0.05% by weight of the particles will not pass through a 325 mesh sieve. The particles may, for example, have a D50 of 3-6 microns, for example 3.8-4.9, for example about 4.3; a D50 of 1-4 microns, for example 2.2-2.6 microns, for example about 2.4 microns; and a D10 of 1-2 microns, for example 1.2-1.4, for example about 1.3 microns. The particles have a relatively high water absorption, for example at least 25g / 100g, for example 30-70g / 100g. Examples of suitable commercially available products include, for example, Quimica® from Lagos Industria Quimica 15Plus. In some embodiments, additional calcium-containing abrasives are used, such as calcium phosphate abrasives, such as tricalcium phosphate, hydroxyapatite, or dicalcium phosphate dihydrate or calcium pyrophosphate; and / or silica abrasives, sodium metaphosphate, potassium metaphosphate, aluminum silicate, calcined alumina, bentonite or other siliceous materials, or combinations thereof. Examples of silica abrasives include, but are not limited to, precipitated or hydrated silicas having an average particle size of up to about 20 microns, such as Zeodent 105 and Zeodent 1 14 (sold by JM Huber Chemicals Division, Havre de Grace, Md. 21078); Sylodent 783 (sold by Davison Chemical Division of WR Grace & Company); or Sorbosil AC 43 (obtained from PQ Corporation). In some embodiments, the effective amount of silica abrasive is about 10%-30%, for example, about 20%. In some embodiments, the acidic silica abrasive Sylodent is included at a concentration of about 2 wt % to about 35 wt %, about 3 wt % to about 20 wt %, about 3 wt % to about 15 wt %, about 10 wt % to about 15 wt %. For example, the acidic silica abrasive may be present in an amount selected from the following: 2 wt %, 3 wt %, 4 wt %, 5 wt %, 6 wt %, 7 wt %, 8 wt %, 9 wt %, 10 wt %, 11 wt %, 12 wt %, 13 wt %, 14 wt %, 15 wt %, 16 wt %, 17 wt %, 18 wt %, 19 wt %, 20 wt %. When measured as a 5 wt % aqueous slurry, Sylodent 783 has a pH of 3.4-4.2, and the silica material has an average particle size of less than 10 microns, such as 3-7 microns, such as about 5.5 microns. In some embodiments, the silica is a synthetic amorphous silica (e.g., 1 wt%-28 wt%) (e.g., 8 wt%-25 wt%). In some embodiments, the silica abrasive is a silica gel or a precipitated amorphous silica, such as a silica having an average particle size in the range of 2.5 microns to 12 microns. Some embodiments also include small-particle silica (e.g., about 5 wt% of Sorbosil AC43 from PQ CorporationWarrington, United Kingdom) with a median particle size (d50) of 1-5 microns (e.g., 3-4 microns). The composition may include 5 to 20 wt% of small-particle silica, or, for example, 10–15 wt%, or, for example, 5 wt%, 10 wt%, 15 wt%, or 20 wt% of small-particle silica.In some embodiments, 20-30% by weight of the total silica in the composition is small particle silica (e.g., having a median particle size (d50) of 3-4 microns), and wherein the small particle silica is about 5% by weight of the oral care composition. In some embodiments, silica is used as a thickener, such as particulate silica. In some embodiments, the composition comprises calcium carbonate, such as highly absorbent precipitated calcium carbonate (e.g., 20% to 30% by weight of the composition, or 25% of highly absorbent precipitated calcium carbonate), or light precipitated calcium carbonate (e.g., about 10% of light precipitated calcium carbonate) or about 10% natural calcium carbonate.

[0054] In some embodiments, the oral care composition comprises a whitening agent, such as a whitening agent selected from the group consisting of peroxides, metal chlorites, perborates, percarbonates, peroxyacids, hypochlorites, hydroxyapatite, and combinations thereof. The oral care composition may comprise hydrogen peroxide or a source of hydrogen peroxide, such as urea peroxide or a peroxide salt or complex (e.g., such as peroxyphosphates, peroxycarbonates, perborates, peroxysilicates, or persulfates; e.g., calcium peroxyphosphate, sodium perborate, sodium peroxycarbonate, sodium peroxyphosphate, and potassium persulfate) or a hydrogen peroxide polymer complex (such as a hydrogen peroxide-polyvinylpyrrolidone polymer complex).

[0055] In some embodiments, the oral care composition comprises an effective amount of one or more antibacterial agents, for example, an antibacterial agent selected from the group consisting of halogenated diphenyl ethers (e.g., triclosan), triclosan monophosphate, herbal extracts and essential oils (e.g., rosemary extract, tea extract, magnolia extract, thymol, menthol, eucalyptol, geraniol, carvacrol, citral, hibiscus phenol, magnolol, ursolic acid, ursodeoxycholic acid (ursic acid), and ursolic acid. acid), morin, catechol, methyl salicylate, epigallocatechin gallate, epigallocatechin, gallic acid, miswak extract, sea buckthorn extract), biguanide preservatives (e.g., chlorhexidine, alexidin or octenidine), quaternary ammonium compounds (e.g., cetylpyridinium chloride (CPC), benzalkonium chloride, tetradecylpyridinium chloride (TPC), N-tetradecyl-4-ethylpyridinium chloride (TDEPC)), phenolic preservatives, hexetidine, furanone, bacteriocin, ethyl lauroyl arginate, arginine bicarbonate, camellia extract, flavonoids, flavan, halogenated diphenyl ethers, myosin Acid, sanguinarine, povidone iodine, delmopinol, salifluor, metal ions (e.g., zinc salts, stannous salts, copper salts, iron salts), propolis and oxidants (e.g., hydrogen peroxide, buffered sodium peroxyborate or sodium peroxycarbonate), phthalic acid and its salts, monoperoxyphthalic acid and its salts and esters, ascorbyl stearate, oleyl sarcosine, alkyl sulfates, dioctyl sulfosuccinate, salicylanilide, domiphen bromide, delmopinol, octopirox and other piperidinyl derivatives, nicotinic acid preparations, chlorite; parabens such as methylparaben or propylparaben and mixtures of any of the foregoing. One or more additional antibacterial agents or preservatives may optionally be present in the composition in a total amount of about 0.01% to about 0.5%, optionally about 0.05% to about 0.1% or about 0.3% by weight, based on the total weight of the composition.

[0056] In some embodiments, the oral care composition may include at least one bicarbonate salt, which can be used to impart a "clean feel" to the teeth and gums due to effervescence and release of carbon dioxide. Any orally acceptable bicarbonate salt can be used, including but not limited to alkali metal bicarbonates such as sodium and potassium bicarbonates, ammonium bicarbonate, etc. One or more additional bicarbonates are optionally present in a total amount of about 0.1% to about 50% by weight, for example, about 1% to about 20% by weight, based on the total weight of the composition.

[0057] In some embodiments, the oral care composition also includes at least one flavoring agent, which can be used to enhance the taste of the composition, for example. Any oral acceptable natural or synthetic flavoring agent can be used, including but not limited to essential oils and various flavoring aldehydes, esters, alcohols and similar substances, tea flavoring agents, vanillin, sage, marjoram, parsley oil, spearmint oil, cinnamon oil, wintergreen oil, peppermint oil, clove oil, bay oil, anise oil, eucalyptus oil, citrus oil, fruit oil, sassafras and fruit flavors (including those derived from lemons, oranges, limes, grapefruits, apricots, bananas, grapes, apples, strawberries, cherries, pineapples, etc.), flavoring agents derived from beans and nuts (such as coffee, cocoa, cola nuts, peanuts, almonds), etc., adsorbed and encapsulated flavoring agents, etc. Also included in the flavoring agent herein are ingredients that provide fragrance and / or other sensory effects (including cool or warming effects) in the mouth. Such ingredients illustratively include menthol, carvone, methyl acetate, menthyl lactate, camphor, eucalyptus oil, eucalyptol, anethole, eugenol, cinnamon, hydroxyphenyl butanone, alpha-ionone, propenyl guaiacol, thymol, linalool, benzaldehyde, cinnamaldehyde, N-ethyl-p-menthane-3-carboxamide, N,2,3-trimethyl-2-isopropylbutanamide, 3-(1-menthoxy)-propane-1,2-diol, cinnamaldehyde glycerol acetal (CGA), menthone glycerol acetal (MGA), and the like. Based on the total weight of the composition, one or more flavoring agents are optionally present in a total amount of about 0.01% to about 5% by weight, for example, about 0.03% to about 2.5% by weight, optionally about 0.05% to about 1.5% by weight, further optionally about 0.1% to about 0.3% by weight, and in some embodiments, about 0.01% to about 1% by weight, about 0.05% to about 2% by weight, about 0.1% to about 2.5% by weight, and about 0.1% to about 0.5% by weight.

[0058] In some embodiments, the oral care composition comprises at least one sweetener, which can be used, for example, to enhance the taste of the composition. Sweeteners useful herein include glucose, polydextrose, sucrose, maltose, dextrin, dried invert sugar, mannose, xylose, ribose, fructose, levulose, galactose, corn syrup, partially hydrolyzed starch, hydrogenated starch hydrolysate, ethanol, sorbitol, mannitol, xylitol, maltitol, isomalt, aspartame, neotame, saccharin and its salts (e.g., saccharin sodium), sucralose, strong sweeteners based on dipeptides, sodium cyclamate, dihydrochalcone, glycerol, propylene glycol, polyethylene glycol, poloxamer polymers such as POLOXOMER 407, PLURONIC F108 (both from BASF Corporation), alkyl glycosides (APG), polysorbates, PEG40, castor oil, menthol, and mixtures thereof. One or more sweeteners are optionally present in a total amount that depends closely on the particular sweetener chosen, but is typically 0.005 wt % to 5 wt %, optionally 0.005 wt % to 0.2 wt %, and also optionally 0.05 wt % to 0.1 wt %, based on the total weight of the composition.

[0059] In some embodiments, the oral care composition further comprises an agent that interferes with or prevents bacterial attachment, for example, ethyl lauroylarginine (ELA), methylparaben (solbrol), or chitosan, and a plaque dispersant such as an enzyme (papain, glucoamylase, etc.).

[0060] In some embodiments, the oral care composition also includes at least one surfactant. Any oral acceptable surfactant can be used, most of which are anionic, cationic, zwitterionic, nonionic and mixtures thereof. Examples of suitable surfactants include water-soluble salts of monosulfate esters of higher fatty acid monoglycerides, such as sodium salts of monosulfate monoglycerides of hydrogenated coconut oil fatty acids; higher alkyl sulfates, such as sodium lauryl sulfate, sodium coconut glycerol monoester sulfonate, sodium lauryl sarcosinate, sodium lauryl isothiocyanate, sodium lauryl ether carboxylate and sodium dodecylbenzene sulfonate; alkyl aryl sulfonates, such as sodium dodecylbenzene sulfonate; higher alkyl sulfoacetates, such as sodium lauryl sulfoacetate; higher fatty acid esters of 1,2-dihydroxypropane sulfonic acid; and substantially saturated higher aliphatic acylamides of lower aliphatic aminocarboxylic acid compounds, such as those having 12 to 16 carbons in fatty acids, alkyl or acyl groups; and the like. Examples of amides include sodium, potassium and ethanolamine salts of N-lauryl sarcosine and N-lauryl, N-myristoyl or N-palmitoyl sarcosine. Examples of cationic surfactants include derivatives of aliphatic quaternary ammonium compounds having a long alkyl chain containing 8 to 18 carbon atoms, such as lauryl trimethyl ammonium chloride, cetyl pyridinium chloride, cetyl trimethyl ammonium bromide, diisobutyl phenoxyethyl dimethyl benzyl ammonium chloride, coconut alkyl trimethyl ammonium nitrite, cetyl pyridinium fluoride and mixtures thereof. Suitable nonionic surfactants include, but are not limited to, poloxamers, polyoxyethylene sorbitan esters, fatty alcohol ethoxylates, alkylphenol ethoxylates, tertiary amine oxides, tertiary phosphine oxides, dialkyl sulfoxides and the like. Other examples include, for example, non-anionic polyoxyethylene surfactants such as poloxamer 407, stearyl polyoxyethylene 30, polysorbate 20, and castor oil; and amphoteric surfactants such as derivatives of aliphatic secondary and tertiary amines with anionic groups such as carboxylates, sulfates, sulfonates, phosphates, or phosphonates, such as cocamidopropyl betaine (tegobaine) and cocamidopropyl betaine lauryl glucoside, condensation products of ethylene oxide with various hydrogen-containing compounds that can react with ethylene oxide and have a hydrophobic long chain (e.g., an aliphatic chain of 12 to 20 carbon atoms), the condensation products (ethoxamers) containing hydrophilic polyoxyethylene moieties, such as condensation products of poly(ethylene oxide) with fatty acids, fatty alcohols, fatty amides, and other fatty moieties, and condensation products with propylene oxide and polypropylene oxide. In some embodiments, the oral composition comprises a surfactant system of sodium lauryl sulfate (SLS) and cocamidopropyl betaine. One or more surfactants are optionally present in a total amount of about 0.01 wt % to about 10 wt %, such as about 0.05 wt % to about 5 wt % or about 0.1 wt % to about 2 wt %, such as about 1.5 wt %, based on the total weight of the composition.In some embodiments, the oral composition comprises an anionic surfactant, such as a surfactant selected from sodium lauryl sulfate, ether sodium lauryl sulfate, and mixtures thereof, such as sodium lauryl sulfate (SLS) in an amount of about 0.3% to about 4.5% by weight, such as 1% to 2%; and / or a zwitterionic surfactant, such as a betaine surfactant, such as cocoamidopropyl betaine, such as cocoamidopropyl betaine in an amount of about 0.1% to about 4.5% by weight, such as 0.5% to 2%. Some embodiments comprise a nonionic surfactant in an amount of 0.5%-5% (e.g., 1%-2%) selected from poloxamers (poloxamer 407), polysorbates (e.g., polysorbate 20), polyoxyethylene hydrogenated castor oil (e.g., polyoxyethylene 40 hydrogenated castor oil), and mixtures thereof. In some embodiments, the poloxamer nonionic surfactant has a polyoxypropylene molecular weight of 3000 to 5000 g / mol and a polyoxyethylene content of 60 to 80 mol%, for example, the poloxamer nonionic surfactant comprises poloxamer 407. Any of the aforementioned compositions may further comprise sorbitol, wherein the total amount of sorbitol is 10-40% (e.g., about 23%).

[0061] In some embodiments, the oral care composition comprises at least one foam regulator, which can be used, for example, to increase the amount, density or stability of the foam produced by the composition when stirring. Any oral acceptable foam regulator can be used, including but not limited to polyethylene glycol (PEG), also referred to as polyoxyethylene. High molecular weight PEG is suitable, including average molecular weight of 200,000 to 7,000,000, such as 500,000 to 5,000,000, or 1,000,000 to 2,500,000, by the gross weight of the composition, one or more PEGs are optionally present in a total amount of about 0.1 % by weight to about 10 % by weight, such as about 0.2 % by weight to about 5 % by weight, or about 0.25 % by weight to about 2 % by weight.

[0062] In some embodiments, the oral care composition comprises at least one pH adjusting agent. Such agents include acidifiers that lower the pH, alkalizers that increase the pH, and buffers that control the pH within a desired range. For example, one or more compounds selected from acidifiers, alkalizers, and buffers may be included to provide a pH of 2 to 10, or 2 to 8, 3 to 9, 4 to 8, 5 to 7, 6 to 10, 7 to 9, etc. in various illustrative embodiments. Any orally acceptable pH adjusting agent may be used, including but not limited to carboxylic acids; phosphoric and sulfonic acids; acid salts (e.g., monosodium citrate, disodium citrate, monosodium malate, etc.); alkali metal hydroxides, such as sodium hydroxide; carbonates, such as sodium carbonate; bicarbonates, such as sodium bicarbonate; sesquicarbonates; borates; silicates; bisulfates; phosphates (e.g., monosodium phosphate, trisodium phosphate, monopotassium phosphate, dipotassium hydrogen phosphate, trisodium phosphate, sodium tripolyphosphate, phosphoric acid); imidazole; sodium phosphate buffer (e.g., monosodium phosphate and disodium phosphate); citrates (e.g., citric acid, trisodium citrate dehydrate); pyrophosphates (sodium and potassium salts), etc. and mixtures thereof. One or more pH adjusting agents are optionally present in a total amount effective to maintain the composition within an orally acceptable pH range. The composition may have an acidic or alkaline pH, for example, pH 4 to pH 5.5 or pH 8 to pH 10. In some embodiments, the amount of buffer is sufficient to provide a pH of about 5 to about 9, preferably about 6 to about 8, and more preferably about 7, when the composition is dissolved in water, a mouthwash base, or a toothpaste base. Typical amounts of buffer are from about 5% to about 35%, in one embodiment from about 10% to about 30%, and in another embodiment from about 15% to about 25%, based on the total weight of the composition.

[0063] In some embodiments, the oral care composition also includes at least one wetting agent. Any oral acceptable wetting agent can be used, including but not limited to polyols such as glycerol, sorbitol (optionally as an aqueous solution of 70 wt %), xylitol or low molecular weight polyethylene glycol (PEG) and their mixtures. Most of wetting agents also serve as sweeteners. In some embodiments, the composition includes a wetting agent of 15 wt % to 70 wt % or 30 wt % to 65 wt %. Suitable wetting agents include edible polyhydroxy alcohols, such as glycerol, sorbitol, xylitol, propylene glycol and other polyols and mixtures of these wetting agents. The mixture of glycerol and sorbitol can be used in some embodiments as the wetting agent component of the composition herein. By the gross weight of the composition, one or more wetting agents are optionally present in an amount of about 1 wt % to about 70 wt %, for example, about 1 wt % to about 50 wt %, about 2 wt % to about 25 wt %, or about 5 wt % to about 15 wt %. In some embodiments, a humectant such as glycerol is present in an amount of at least >20%, such as 20%-40%, such as 25%-35%.

[0064] Mouthfeel agents are included in materials that impart desired texture or other sensations during use of the composition. In some embodiments, the oral care composition includes at least one thickener suitable for, for example, imparting the desired consistency and / or mouthfeel of the composition. Any oral acceptable thickener can be used, including but not limited to carbomer, also known as carboxyvinyl polymer; carrageenan, also known as Irish moss, and more specifically i-carrageenan (Ι-carrageenan); cellulose polymers, such as hydroxyethylcellulose; and water-soluble salts of cellulose ethers (e.g., sodium carboxymethylcellulose and sodium carboxymethyl hydroxyethylcellulose); carboxymethylcellulose (CMC) and its salts, for example, sodium CMC; natural gums, such as karaya gum, flavin, gum arabic and tragacanth; colloidal magnesium aluminum silicate; colloidal silicon dioxide; starch; polyvinyl pyrrolidone; hydroxyethyl propyl cellulose; hydroxybutyl methylcellulose; hydroxypropyl methylcellulose and hydroxyethyl cellulose and amorphous silicon dioxide, etc. One preferred class of thickeners or gelling agents includes a class of acrylic acid homopolymers crosslinked with pentaerythritol alkyl ethers or sucrose alkyl ethers, or carbomers. Carbomers can be used as Series is commercially available from BFGoodrich. Particularly preferred Carbopol includes Carbopol 934, 940, 941, 956, 974P and mixtures thereof. Silica thickeners may also be used, such as DT267 (derived from PPG Industries). By the total weight of the composition, one or more thickeners are optionally present in a total amount of about 0.01 wt % to 15 wt %, for example, about 0.1 wt % to about 10 wt % or about 0.2 wt % to about 5 wt %. Some embodiments include sodium carboxymethylcellulose (e.g., 0.5 wt %-1.5 wt %). In certain embodiments, a thickener of about 0.5% to about 5.0% of the amount is used by weight of the total composition. The thickener may be present in an amount of 1 wt % to 15 wt %, 3 wt % to 10 wt %, 4 wt % to 9 wt %, 5 wt % to 8 wt %, for example, 5 wt %, 6 wt %, 7 wt % or 8 wt %.

[0065] In some embodiments, the oral care composition comprises at least one coloring agent. Coloring agents herein include pigments, dyes, lakes, and agents such as pearlescent agents that impart specific gloss or reflectivity. In various embodiments, coloring agents are operable to provide white or light-colored coatings on tooth surfaces, serve as indications of positions effectively contacted by the composition on tooth surfaces, and / or modify the appearance of the composition, particularly color and / or opacity to enhance the attractiveness to consumers. Any acceptable coloring agent for oral cavity can be used, including FD&C dyes and pigments, talc, mica, magnesium carbonate, calcium carbonate, magnesium silicate, magnesium aluminum silicate, silicon dioxide, titanium dioxide, zinc oxide, red, yellow, brown and black iron oxide, ammonium ferrocyanide, manganese violet, ultramarine, titanium mica, bismuth oxychloride, and mixtures thereof. By the gross weight of the composition, one or more coloring agents are optionally present in an amount of about 0.001% to about 20%, for example, about 0.01% to about 10% or about 0.1% to about 5%.

[0066] In some embodiments, oral care compositions also include anticalculus (tartar control) agents. Suitable anticalculus agents include, but are not limited to: phosphates and polyphosphates, polyaminopropane sulfonic acid (AMPS), polyolefin sulfonates, polyolefin phosphates, diphosphates such as azacycloalkane-2,2-diphosphates (e.g., azacycloheptane-2,2-diphosphoric acid), N-methylazacyclopentane-2,3-diphosphoric acid, ethane-1-hydroxy-1,1-diphosphoric acid (EHDP) and ethane-1-amino-1,1-diphosphate, phosphoryl alkane carboxylic acid. Available inorganic phosphates and polyphosphates include sodium dihydrogen phosphate, disodium hydrogen phosphate and trisodium phosphate. Soluble pyrophosphate is an available anticalculus agent. Pyrophosphate can be any alkali metal pyrophosphate. In certain embodiments, salts include tetraalkali metal pyrophosphates, dialkali metal diacid pyrophosphates, trialkali metal monoacid pyrophosphates and mixtures thereof, wherein the alkali metal is sodium or potassium. Pyrophosphates also promote the preservation of compositions by reducing water activity, tetrasodium pyrophosphate (TSPP), tetrapotassium pyrophosphate, sodium tripolyphosphate, tetrapolyphosphate, sodium tripolyphosphate, sodium hexametaphosphate, and mixtures thereof. Both hydrated and unhydrated salts are suitable. The effective amount of pyrophosphates that can be used in the compositions of the present invention is generally sufficient to provide at least 0.1% by weight of pyrophosphate ions, such as 0.1% to 3% by weight, such as 0.1% to 2% by weight, such as 0.1% to 1% by weight, such as 0.2% to 0.5% by weight.

[0067] Other useful tartar control agents include polymers and copolymers. In some embodiments, the oral care composition comprises one or more polymers such as polyethylene glycol, polyvinyl methyl ether maleic acid copolymer, a polysaccharide (e.g., a cellulose derivative such as carboxymethyl cellulose; or a polysaccharide gum such as xanthan gum or carrageenan). The acidic polymer (e.g., polyacrylate gel) can be provided in the form of its free acid or a partially or fully neutralized water-soluble alkali metal (e.g., potassium and sodium) or ammonium salt. Certain embodiments comprise a 1:4 to 4:1 copolymer of maleic anhydride or maleic acid with another polymerizable ethylenically unsaturated monomer such as methyl vinyl ether (methoxyethylene) having a molecular weight (MW) of about 30,000 to about 1,000,000, a polyvinyl methyl ether / maleic anhydride (PVM / MA) copolymer such as (For example, S-97 polymer). In some embodiments, the PVM / MA copolymer comprises a copolymer of methyl vinyl ether / maleic anhydride, wherein the anhydride is hydrolyzed after copolymerization to obtain the corresponding acid. In some embodiments, the PVM / MA copolymer has an average molecular weight (MW) of about 30,000 to about 1,000,000, such as about 300,000 to about 800,000, such as wherein the anionic polymer is about 1%-5%, such as about 2%, of the weight of the composition. In some embodiments, the anticalculus agent is present in the composition in an amount of 0.2% to 0.8%, 0.3% to 0.7%, 0.4% to 0.6%, or about 0.5% by weight, based on the total weight of the composition. The copolymer can be obtained, for example, as Gantrez AN 139 (MW 500,000), AN 119 (MW 250,000) and S-97 pharmaceutical grade (MW 70,000) from GAF Chemicals. Other polymers that work include those such as 1:1 copolymers of maleic anhydride with ethyl acrylate, hydroxyethyl methacrylate, N-vinyl-2-pyrrolidone or ethylene, the latter being available, for example, as Monsanto EMA No. 1 103, MW 10,000 and EMA grade 61; and 1:1 copolymers of acrylic acid with methyl methacrylate or hydroxyethyl methacrylate, methyl acrylate or ethyl acrylate, isobutyl vinyl ether or N-vinyl-2-pyrrolidone. Generally, suitable are polymerized olefinic or ethylenically unsaturated carboxylic acids containing an activated carbon-carbon olefinic double bond and at least one carboxyl group, i.e., acids containing an olefinic double bond that readily works in polymerization because it is present in the monomer molecule in the alpha-beta position relative to the carboxyl group or as part of a terminal methylene group. Illustrative of such acids are acrylic acid, methacrylic acid, ethacrylic acid, α-chloroacrylic acid, crotonic acid, β-acryloyloxypropionic acid, sorbic acid, α-chlorosorbic acid, cinnamic acid, β-styryl acrylic acid, mucofuric acid, itaconic acid, citraconic acid, methyl fumaric acid, glutaconic acid, aconitic acid, α-phenylacrylic acid, 2-benzylacrylic acid, 2-cyclohexylacrylic acid, angelic acid, umbelliferous acid, fumaric acid, maleic acid and anhydrides. Other different olefinic monomers that can be copolymerized with such carboxylic acid monomers include vinyl acetate, vinyl chloride, dimethyl maleate, etc. The copolymer contains sufficient carboxylate groups for water solubility. Another class of polymeric agents includes compositions containing homopolymers of substituted acrylamides and / or homopolymers of unsaturated sulfonic acids and their salts, particularly where the polymer is based on an unsaturated sulfonic acid selected from acrylamidoalkanesulfonic acid such as 2-acrylamido-2-methylpropanesulfonic acid, with a molecular weight of about 1,000 to about 2,000,000. Another class of useful polymerizing agents includes polymeric amino acids, especially those containing proportions of anionic surface active amino acids such as aspartic acid, glutamic acid and phosphoserine.

[0068] In some embodiments, the oral care composition comprises a saliva stimulant that can be used, for example, to improve dry mouth. Any orally acceptable saliva stimulant can be used, including but not limited to food acids such as citric acid, lactic acid, malic acid, succinic acid, ascorbic acid, adipic acid, fumaric acid, and tartaric acid, and mixtures thereof. One or more saliva stimulants are optionally present in a total amount effective to stimulate saliva.

[0069] In some embodiments, the oral care composition comprises nutrients. Suitable nutrients include vitamins, minerals, amino acids and mixtures thereof. Vitamins include vitamins C and D, chloramine, riboflavin, calcium pantothenate, niacin, folic acid, niacinamide, pyridoxine, cyanocobalamin, p-aminobenzoic acid, bioflavonoids and mixtures thereof. Nutritional supplements include amino acids (such as L-tryptophan, L-lysine, methionine, threonine, L-carnitine and L-carnitine), lipophilic substances (such as choline, inositol, betaine and linoleic acid) and mixtures thereof.

[0070] In some embodiments, the oral care composition comprises at least one viscosity modifier, which can be used, for example, to help inhibit sedimentation or separation of ingredients or to promote redispersibility of liquid compositions when stirred. Any orally acceptable viscosity modifier can be used, including but not limited to mineral oil, petrolatum, clay and organically modified clay, silica, etc. One or more viscosity modifiers are optionally present in a total amount of about 0.01 wt % to about 10 wt %, such as about 0.1 wt % to about 5 wt %, based on the total weight of the composition.

[0071] In some embodiments, the oral care composition comprises an anti-sensitizing agent, for example, potassium salts such as potassium nitrate, potassium bicarbonate, potassium chloride, potassium citrate, and potassium oxalate; capsaicin; eugenol; strontium salts; chloride salts; and combinations thereof. Depending on the agent selected, such agents may be added in an effective amount, for example, from about 1% to about 20% by weight based on the total weight of the composition.

[0072] In some embodiments, the oral care composition comprises an antioxidant. Any orally acceptable antioxidant may be used, including butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), vitamin A, carotenoids, coenzyme Q10, PQQ, vitamin A, vitamin C, vitamin E, anethole-dithiothione, flavonoids, polyphenols, ascorbic acid, herbal antioxidants, chlorophyll, melatonin, and mixtures thereof.

[0073] In some embodiments, the oral care composition is composed of one or more alkali metal phosphates, such as sodium, potassium or calcium salts, such as selected from dialkali metal hydrogen phosphate and alkali metal pyrophosphate, such as selected from the following alkali metal phosphates: disodium hydrogen phosphate, dipotassium hydrogen phosphate, dicalcium phosphate dihydrate, calcium pyrophosphate, tetrasodium pyrophosphate, tetrapotassium pyrophosphate, sodium tripolyphosphate, disodium hydrogen orthophosphate, sodium dihydrogen phosphate, pentapotassium triphosphate and mixtures of two or more thereof, for example, by weight of the composition, in an amount of 0.01-20%, such as 0.1-8%, such as 0.1 to 5%, such as 0.3 to 2%, such as 0.3 to 1%, such as about 0.01%, about 0.1%, about 0.5%, about 1%, about 2%, about 5%, about 6%. In some embodiments, the composition comprises tetrapotassium pyrophosphate, disodium hydrogen orthophosphate, sodium dihydrogen phosphate and pentapotassium triphosphate. In some embodiments, the composition comprises 0.1 wt% - 1.0 wt% (eg, about .5 wt%) tetrasodium pyrophosphate.

[0074] In some embodiments, the oral care composition comprises a calcium and phosphate source selected from the group consisting of: (i) calcium-glass complexes, such as sodium calcium phosphosilicate, and (ii) calcium-protein complexes, such as casein phosphopeptide-amorphous calcium phosphate. Any of the foregoing compositions further comprises a soluble calcium salt, such as selected from calcium sulfate, calcium chloride, calcium nitrate, calcium acetate, calcium lactate, and combinations thereof.

[0075] In some embodiments, the oral care composition comprises an additional ingredient selected from the group consisting of benzyl alcohol, methylisothiazolinone ("MIT"), sodium bicarbonate, sodium methyl cocoyl tauranol, lauryl alcohol, and polyphosphates. Some embodiments comprise benzyl alcohol present at 0.1% to 0.8%, or 0.2% to 0.7%, or 0.3% to 0.6%, or 0.4% to 0.5%, such as about 0.1%, about 0.2%, about 0.3%, about 0.4%, about 0.5%, about 0.6%, about 0.7%, or about 0.8%.

[0076] In some embodiments, the oral care composition comprises 5% - 40%, such as 10% - 35%, such as about 15%, 25%, 30%, and 35% or more water.

[0077] Provided is a method for determining the effect of a combination of a compound or composition and arginine on the growth of beneficial oral bacteria and on the growth of pathogenic oral bacteria. The method can be used to identify a compound or composition that enhances the selective promotion of arginine to the growth of beneficial oral bacteria relative to pathogenic oral bacteria. In some embodiments, the method includes performing one or more beneficial oral bacteria growth test assays, one or more pathogenic oral bacteria growth test assays, one or more beneficial oral bacteria growth control assays, and one or more beneficial oral bacteria growth control assays. The beneficial oral bacteria growth test assay includes culturing beneficial oral bacteria species in a culture medium containing arginine and a test compound or composition, and measuring the growth of beneficial oral bacteria in the beneficial oral bacteria growth test assay. The pathogenic oral bacteria growth test assay includes culturing pathogenic oral bacteria species in a culture medium containing arginine and a test compound or composition, and measuring the growth of pathogenic oral bacteria in the pathogenic oral bacteria growth test assay. The beneficial oral bacteria growth control assay includes culturing beneficial oral bacteria species in a culture medium containing arginine without a test compound or composition, and measuring the growth of beneficial oral bacteria in the beneficial oral bacteria growth control assay. The pathogenic oral bacteria growth control assay includes culturing pathogenic oral bacteria species in a medium containing arginine without a test compound or composition, and measuring the growth of pathogenic oral bacteria in the pathogenic oral bacteria growth control assay. The ratio of the growth of beneficial oral bacteria in the beneficial oral bacteria growth test assay to the growth of beneficial oral bacteria in the beneficial oral bacteria growth control assay (the beneficial ratio) is calculated, and the ratio of the growth of pathogenic oral bacteria in the pathogenic oral bacteria growth test assay to the growth of pathogenic oral bacteria in the pathogenic oral bacteria growth control assay (the pathogenic ratio) is also calculated. The beneficial ratio is then compared to the pathogenic ratio, and a higher beneficial ratio indicates that the test compound or composition enhances the selective promotion of arginine to the growth of beneficial oral bacteria relative to pathogenic oral bacteria. Alternatively, the ratio of the growth of beneficial oral bacteria in the beneficial oral bacteria growth test assay to the growth of pathogenic oral bacteria in the pathogenic oral bacteria growth test assay (test ratio) is calculated, and the ratio of the growth of beneficial oral bacteria in the beneficial oral bacteria growth control assay to the growth of pathogenic oral bacteria in the pathogenic oral bacteria growth control assay (control ratio) is calculated. The test ratio is then compared to the control ratio, and a higher test ratio as compared to the control ratio indicates that the test compound or composition enhances the selective promotion of the growth of beneficial oral bacteria relative to pathogenic oral bacteria by arginine.

[0078] In some embodiments, the test assays (both the beneficial oral bacteria growth test assay and the pathogenic oral bacteria growth test assay) include a series of such test assays using culture media supplemented with 5, 10, 20, and 25 micromoles / ml of arginine. In some embodiments, the test assays (both the beneficial oral bacteria growth test assay and the pathogenic oral bacteria growth test assay) use culture media supplemented with at least 5 micromoles / ml, at least 10 micromoles / ml, at least 20 micromoles / ml, at least 25 micromoles / ml, about 0.05-0.1 mg / ml, about 0.1-1.0 mg / ml, about 1 mg / ml, about 2 mg / ml, about 3 mg / ml, about 4 mg / ml, about 5 mg / ml, about 6 mg / ml, about 7 mg / ml, about 8 mg / ml, about 9 mg / ml, about 10 mg / ml, about 11 mg / ml, about 12 mg / ml, about 13 mg / ml, about 14 mg / ml, and / or about 15 mg / ml of arginine. In some embodiments, such test determinations include a series of such test determinations using a culture medium supplemented with a test compound or composition present at different concentrations. In some embodiments, a 96-well polystyrene microtiter plate is used for the test determination. The bacterial culture is incubated under conditions that promote growth. In some embodiments, the bacterial culture is incubated under anaerobic conditions or under an environment containing 5% carbon dioxide. In some embodiments, bacterial growth is monitored at one or more time points, such as daily monitoring, for 1-4 days (i.e., at 24, 48, 72 and 96 hours). Growth can be measured using various known methods, such as qPCR or optical density at a wavelength of 630nm. In some embodiments, control determinations (beneficial oral bacteria growth control determinations and pathogenic oral bacteria growth control determinations) are included. Such control determinations are the same determinations as the corresponding test determinations performed, except that the test compound or composition is not included in the control determinations. The bacterial growth results in the test determinations are compared with the bacterial growth results in the control determinations. In some embodiments, instead of performing control determinations, the bacterial growth results in the test determinations are compared with the reference results of the control determinations of the known growth rates. In some embodiments, the beneficial oral bacteria used may be selected from Streptococcus garneri, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis and Streptococcus sanguinis. In some embodiments, the pathogenic oral bacteria used may be selected from Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans and Streptococcus sobrinus. The method may be used to identify such compounds and compositions that may be used in combination with arginine to reduce the pathogenic load in the oral cavity and change the dental plaque composition to a healthier state with fewer pathogenic bacteria, thereby producing less destructive dental plaque.The methods can be used to identify compounds and compositions that can be used in combination with arginine to prevent one or more of gingivitis, periodontitis, peri-implantitis, peri-implant mucositis, necrotizing gingivitis, necrotizing periodontitis, and dental caries.

[0079] Methods are provided for determining the effect of a combination of a compound or composition with arginine on the selective growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm. The method can be used to identify a compound or composition that enhances the selective promotion of the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm by arginine. The method includes performing a dual-species biofilm test assay, performing a dual-species biofilm control assay, and comparing the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm with the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm. The dual-species biofilm test assay includes co-culturing beneficial oral bacteria species and pathogenic oral bacteria species in a dual-species test biofilm comprising arginine and a test compound or composition; quantifying the beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm; and comparing the number of beneficial oral bacteria in the dual-species test biofilm with the number of pathogenic oral bacteria to determine the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm. The dual-species biofilm control assay includes co-culturing beneficial oral bacteria species and pathogenic oral bacteria species in a dual-species control biofilm containing arginine but without a test compound or composition; quantifying the beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm; and comparing the number of beneficial oral bacteria in the dual-species control biofilm with the number of pathogenic oral bacteria to determine the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm. The beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm is then compared to the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm. A higher beneficial oral bacteria-pathogenic oral bacteria load in the dual-species test biofilm compared to the beneficial oral bacteria-pathogenic oral bacteria load in the dual-species control biofilm indicates that the test compound or composition enhances the selective promotion of the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in the biofilm by arginine.

[0080] In some embodiments, dual species biofilm test determinations include a series of such test determinations using culture media supplemented with 5, 10, 20 and 25 micromoles / ml of arginine. In some embodiments, dual species biofilm test determinations use culture media supplemented with at least 5 micromoles / ml, at least 10 micromoles / ml, at least 20 micromoles / ml, at least 25 micromoles / ml, about 0.05-0.1 mg / ml, about 0.1-1.0 mg / ml, about 1 mg / ml, about 2 mg / ml, about 3 mg / ml, about 4 mg / ml, about 5 mg / ml, about 6 mg / ml, about 7 mg / ml, about 8 mg / ml, about 9 mg / ml, about 10 mg / ml, about 11 mg / ml, about 12 mg / ml, about 13 mg / ml, about 14 mg / ml and / or about 15 mg / ml of arginine. In some embodiments, such test determinations include a series of such test determinations using culture media supplemented with test compounds or compositions present at different concentrations. Bacterial cultures are incubated under conditions that promote growth. In some embodiments, bacterial cultures are incubated under anaerobic conditions or in an environment containing 5% carbon dioxide. In some embodiments, the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm and the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm are measured at one or more time points, such as daily, for 1-4 days (i.e., at 24, 48, 72 and 96 hours). In some embodiments, the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm and the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm are measured by qPCR. In some embodiments, instead of performing a dual-species control biofilm assay, the bacterial growth results in the dual-species test biofilm assay are compared with the reference results of a control assay of a known growth rate. In some embodiments, the beneficial oral bacteria used can be selected from Streptococcus grisea, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis and Streptococcus sanguinis. In some embodiments, the pathogenic oral bacteria used can be selected from Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus. The method can be used to identify compounds and compositions that can be used with arginine to reduce the pathogenic load in the oral cavity and change the composition of dental plaque to a healthier state with fewer pathogenic bacteria, thereby producing less destructive dental plaque. The method can be used to identify compounds and compositions that can be used in combination with arginine to prevent one or more of gingivitis, periodontitis, peri-implantitis, peri-implant mucositis, necrotizing gingivitis, necrotizing periodontitis, and dental caries.

[0081] Other methods include determining the effect of a combination of a compound or composition with arginine on the growth of biofilms produced by beneficial oral bacteria and the growth of biofilms produced by pathogenic oral bacteria. The method includes performing one or more beneficial oral bacteria biofilm growth test assays, one or more pathogenic oral bacteria biofilm growth test assays, one or more beneficial oral bacteria biofilm growth control assays, and one or more beneficial oral bacteria biofilm growth control assays. The beneficial oral bacteria biofilm growth test assay includes culturing a beneficial oral bacteria species in a culture medium containing arginine and a test compound or composition, and measuring the growth of a biofilm produced by beneficial oral bacteria in the beneficial oral bacteria growth test assay. The pathogenic oral bacteria biofilm growth test assay includes culturing a pathogenic oral bacteria species in a culture medium containing arginine and a test compound or composition, and measuring the growth of a biofilm produced by pathogenic oral bacteria in the pathogenic oral bacteria growth test assay. The beneficial oral bacteria biofilm growth control assay includes culturing a beneficial oral bacteria species in a culture medium containing arginine without a test compound or composition, and measuring the growth of a biofilm produced by beneficial oral bacteria in a beneficial oral bacteria growth control assay. The pathogenic oral bacteria biofilm growth control assay includes culturing pathogenic oral bacteria species in a medium containing arginine without a test compound or composition, and measuring the growth of biofilm produced by pathogenic oral bacteria in the pathogenic oral bacteria growth control assay. The total biofilm growth in each assay is assessed using crystal violet staining. Ratios are calculated as in the beneficial oral bacteria growth assay and the pathogenic oral bacteria growth assay described above.

[0082] In some embodiments, the beneficial oral bacteria biofilm growth test assay and the pathogenic oral bacteria biofilm growth test assay include a series of such test assays using culture media supplemented with 5, 10, 20, and 25 micromoles / ml of arginine. In some embodiments, the test assay uses culture media supplemented with at least 5 micromoles / ml, at least 10 micromoles / ml, at least 20 micromoles / ml, at least 25 micromoles / ml, about 0.05-0.1 mg / ml, about 0.1-1.0 mg / ml, about 1 mg / ml, about 2 mg / ml, about 3 mg / ml, about 4 mg / ml, about 5 mg / ml, about 6 mg / ml, about 7 mg / ml, about 8 mg / ml, about 9 mg / ml, about 10 mg / ml, about 11 mg / ml, about 12 mg / ml, about 13 mg / ml, about 14 mg / ml, and / or about 15 mg / ml of arginine. In some embodiments, such test determinations include a series of such test determinations using a culture medium supplemented with a test compound or composition present in different concentrations. The bacterial culture is incubated under conditions that promote biofilm growth. In some embodiments, the bacterial culture is incubated under anaerobic conditions or under an environment containing 5% carbon dioxide. In some embodiments, total biofilm growth is measured at one or more time points, such as daily, for 1-4 days (i.e., at 24, 48, 72 and 96 hours). In some embodiments, the beneficial oral bacteria used can be selected from Streptococcus griseus, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis and Streptococcus sanguinis. In some embodiments, the pathogenic oral bacteria used can be selected from Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans and Streptococcus sobrinus.

[0083] Example

[0084] Example 1

[0085] To evaluate the effects of the amino acid arginine on beneficial and pathogenic oral bacteria, representative bacterial species were grown in 96-well polystyrene microtiter plates in medium supplemented with 5, 10, 20, and 25 micromol / mL arginine. Bacterial cultures were incubated under anaerobic conditions or in an atmosphere containing 5% carbon dioxide, and growth was monitored at 24 and 48 hours by optical density at a wavelength of 630 nm.

[0086] Although both pathogenic and beneficial bacteria have the ability to metabolize arginine, the amino acid appears to have selective activity, preferentially promoting the growth of beneficial bacteria relative to pathogens (Table 1).

[0087] Table 1. Fold increase in beneficial and pathogenic bacteria when cultured with increasing arginine concentrations, as determined by qPCR (see Figure 1 ).

[0088] *Arginine stimulated single species growth and biofilm formation compared to the reference control set to 1 (p<0.05). The cutoff for growth was set to 1.2 (upper 5% of all data), while the cutoff for biofilm formation was set to 1.45 (upper 20% of all data). Aa: Actinobacillus actinomycetemcomitans, Fn: Fusobacterium nucleatum, Ssob: Streptococcus sobrinus, Avisc: Actinomyces viscosus, Ssal: Streptococcus salivarius, Smitis: Streptococcus mitis, Ssang: Streptococcus sanguinis, Sgord: Streptococcus gordonii, Soralis: Streptococcus oralis.

[0089] Under the conditions described, the growth of beneficial microorganisms (e.g., Actinomyces viscosus, Streptococcus mitis and Streptococcus garinii) was stimulated by 1.2-1.3-fold increase with 10-25 μmol / ml arginine. Interestingly, no significant increase in the growth of pathogenic bacteria was observed.

[0090] Example 2

[0091] Total biofilm growth in the presence of arginine was assessed by crystal violet staining. An increase of 1.2-fold or more compared to the no substrate control was considered significant (p<0.05), as calculated using Student's t-test.

[0092] Evaluation of biofilm formation showed that although an increase in A. actinomycetemcomitans biofilm was observed, no significant increase in biofilm formation was observed for either pathogenic or beneficial microorganisms.

[0093] Example 3

[0094] The effect of arginine was evaluated in dual-species biofilms by co-culturing a beneficial bacterial species with a pathogenic microorganism in medium containing 25 μmol / mL arginine. The total beneficial-pathogen load in the biofilm was determined by qPCR using a DNA probe.

[0095] The role of arginine as a selective nutrient was evaluated using a two-species biofilm model using a competitive biofilm assay (Table 2).

[0096] Table 2. The proportion of pathogenic species in dual-species biofilms decreased after arginine treatment.

[0097]

[0098] *% reduction calculated from beneficial and pathogenic CFU recovered after biofilm treatment with 25 μmol / ml arginine. Reduction in pathogenic bacteria is indicated by negative values ​​expressed as a percentage.

[0099] Evaluation of arginine-cultured biofilms indicated an increase in the growth of beneficial bacteria, while a concurrent decrease in the numbers of specific pathogenic bacteria was observed, particularly Porphyromonas gingivalis and Prevotella intermedia (bacteria associated with gum disease) and Streptococcus mutans and Streptococcus sobrinus (bacteria associated with dental caries).

[0100] Example 4

[0101] Oral compositions comprising arginine are disclosed in WO / 2011 / 123123, which corresponds to US 8,652,495, both of which are incorporated herein by reference. In some embodiments, the oral care composition comprises an orally acceptable vehicle, zinc oxide particles, and arginine. In some such embodiments, the zinc oxide particles have a median particle size of 1 to 7 microns; in some such embodiments, the zinc oxide particles have a particle size distribution of 3 to 4 microns, a particle size distribution of 5 to 7 microns, a particle size distribution of 3 to 5 microns, a particle size distribution of 2 to 5 microns, or a particle size distribution of 2 to 4 microns. In some of these various described embodiments, the zinc oxide particles are present in an amount of up to 5% by weight, such as 0.5% to 2% by weight, based on the total weight of the oral care composition. In some of these various described embodiments, the zinc oxide particles of the zinc oxide particles are selected from powders, nanoparticle solutions, nanoparticle suspensions, capsules, and beads. In some of these various embodiments of description, oral care composition also includes at least one other metal oxide selected from stannous oxide, titanium oxide, calcium oxide, cupric oxide and ferric oxide or their mixture. In some of these various embodiments of description, arginine is L-arginine. In some of these various embodiments of description, as based on the gross weight of oral care composition, arginine is at most 5 weight %, such as 0.5 weight % to 5 weight %, such as 2.5 weight % to 4.5 weight % amount exists. In some of various embodiments, oral care composition also includes at least one or more amino acids except arginine, and described at least one or more amino acids are selected from cysteine, leucine, isoleucine, lysine, L-lysine, alanine, asparagine, aspartic acid, phenylalanine, glutamate, glutamic acid, threonine, glutamine, tryptophan, glycine, valine, proline, serine, tyrosine and histidine, and two or more combinations thereof. In some of these various embodiments of description, oral care composition also includes polymer adhesive material. In some of these various described embodiments, the source of zinc oxide particles is a capsule, and the capsule comprises a polymeric adhesive material, and in some such embodiments, the polymeric adhesive material comprises one or more cellulosic polymers, such as embodiments wherein at least one of the one or more cellulosic polymers is a hydroxyalkyl cellulose polymer selected from hydroxypropyl methylcellulose (HPMC), hydroxyethyl propyl cellulose (HEPC), hydroxybutyl methyl cellulose (HBMC), and carboxymethyl cellulose (CMC). In some of these various described embodiments, the polymeric adhesive material comprises a mixture of two hydroxyalkyl cellulose polymers having different molecular weights and zinc oxide encapsulated in the mixture of the two hydroxyalkyl cellulose polymers.In some of these various described embodiments, the polymer adhesive material comprises one or more polymers selected from poly (ethylene oxide) polymers, polyvinyl pyrrolidone (PVP), polyethylene glycol (PEG) / polypropylene glycol (PPG) copolymers, ethylene oxide (EO)-propylene oxide (PO) block copolymers, ester glue, shellac, pressure-sensitive silicone adhesives, methacrylates or their mixtures. In some of these various described embodiments, the oral care composition is a dentifrice composition, such as toothpaste or gel. In some of these various described embodiments, the oral care composition is formulated into a form selected from mouthwash, chewing gum, soluble lozenges and soluble films.

[0102] Example 5

[0103] Oral compositions comprising arginine are disclosed in WO 2014 / 088572, which corresponds to US2015 / 0313813, both of which are incorporated herein by reference. In some embodiments, the oral care composition comprises: about 0.05% by weight to about 5% by weight of a zinc ion source; a fluoride ion source that effectively delivers about 500ppm to about 5,000ppm of fluoride; and about 0.1% by weight to about 10% by weight of arginine. In some such embodiments, the zinc ion source is selected from, for example, zinc citrate, zinc sulfate, zinc silicate, zinc lactate, zinc phosphate, zinc oxide, and combinations thereof, in an amount that is effective in inhibiting erosion. In some such embodiments, the oral composition is in the form of a dentifrice comprising an abrasive. In some such embodiments, the amount of zinc is 0.5% by weight to 4% by weight. In some such embodiments, the composition may also include one or more abrasives, one or more wetting agents, and one or more surfactants. In some such embodiments, the composition may also include an effective amount of one or more alkali metal phosphates and / or an effective amount of one or more antibacterial agents and / or whitening agents. In some such embodiments, the composition comprises zinc phosphate and one or more other sources of zinc ions. In some such embodiments, the pH of the composition is alkaline. In some such embodiments, the composition may comprise, in a silica abrasive dentifrice matrix: 1% to 3% zinc citrate; 1% to 8% arginine; 700ppm to 2000ppm fluoride; and 2% to 8% alkali metal phosphate selected from disodium hydrogen phosphate, dipotassium hydrogen phosphate, calcium hydrogen phosphate dihydrate, tetrasodium pyrophosphate, tetrapotassium pyrophosphate, calcium pyrophosphate, sodium tripolyphosphate, and a combination of two or more thereof.

[0104] Example 6

[0105] The oral composition comprising arginine is disclosed in WO 2015 / 094849, which corresponds to US 2016 / 0338921, both of which are incorporated herein by reference. In some embodiments, the oral care composition comprises: arginine in free or salt form; and zinc oxide and zinc citrate. In some embodiments, based on the total weight of the composition, arginine is present in an amount of 0.5% by weight to 3% by weight, such as 1% by weight to 2.85% by weight, such as 1.17% by weight to 2.25% by weight, such as 1.4% by weight to 1.6% by weight, such as about 1.5% by weight. In some embodiments set forth above, based on the total weight of the composition, the total concentration of zinc salt in the composition is 0.2% by weight to 5% by weight. In some embodiments set forth above, the molar ratio of arginine to total zinc salt is 0.05:1 to 10:1. In some embodiments set forth above, the composition comprises zinc oxide in an amount of 0.5% to 1.5% by weight, such as 1% by weight, and zinc citrate in an amount of 0.25% to 0.75% by weight, such as 0.5% by weight, based on the total weight of the composition. In some embodiments set forth above, the weight ratio of zinc oxide to zinc citrate is 1.5:1 to 4.5:1, optionally 1.5:1 to 4:1, 1.7:1 to 2.3:1, 1.9:1 to 2.1:1, or about 2:1.

[0106] Example 7

[0107] The oral composition comprising arginine is disclosed in WO 2017 / 003844, which corresponds to US 2018 / 0021234, both of which are incorporated herein by reference. In some embodiments, the oral care composition comprises: arginine, zinc oxide and zinc citrate and a fluoride source. In some embodiments, arginine has an L-configuration. In some embodiments, based on the gross weight of the composition, arginine is present in an amount corresponding to 0.1% to 15%, or 0.1% to 8%, or about 5.0% by weight, or about 8.0% by weight, or about 1.5% by weight, and the weight of arginine is calculated as a free form. In some embodiments, arginine is in a free form or in a partial or complete salt form. In some embodiments set forth above, the ratio of the amount of zinc oxide (by weight %) to zinc citrate (by weight %) is 2: 1, 2.5: 1, 3: 1, 3.5: 1 or 4: 1, wherein the ratio is by the weight of the overall composition. In some embodiments, based on the gross weight of the composition, the amount of zinc citrate is 0.25 wt % to 1.0 wt %, and zinc oxide can be present in an amount of 0.75 wt % to 1.25 wt %, or the amount of zinc citrate is 0.5 wt %, and zinc oxide is present in an amount of about 1.0%. In some embodiments set forth above, the fluoride source is sodium fluoride or sodium monofluorophosphate. In some such embodiments, based on the gross weight of the composition, sodium fluoride or sodium monofluorophosphate is 0.1 wt %-2 wt %. In some embodiments, sodium fluoride or sodium monofluorophosphate is a soluble fluoride salt, which provides a soluble fluoride of the amount of 50ppm to 25,000ppm fluoride, such as an amount of about 1000ppm-1500ppm, for example, an amount of about 1450ppm. In some embodiments, based on the gross weight of the composition, the fluoride source is sodium fluoride in an amount of about 0.32 wt %. In some embodiments, the fluoride source is stannous fluoride. Some embodiments set forth above further comprise a preservative selected from the group consisting of benzyl alcohol, methylisothiazolinone ("MIT"), sodium bicarbonate, sodium methyl cocoyl tauranol, lauryl alcohol, and polyphosphates. Some embodiments set forth above further comprise benzyl alcohol in an amount of 0.1 wt%-0.8 wt%, or 0.3 wt%-0.5 wt%, or about 0.4 wt%, based on the total weight of the composition. In some embodiments, the oral care composition comprises about 1.0% zinc oxide, about 0.5% zinc citrate, about 1.5% L-arginine, about 1450 ppm sodium fluoride, and optionally about 0.1 wt% benzyl alcohol and / or about 5% small particle silica (e.g., AC43), based on the total weight of the composition. In some embodiments, the oral care composition comprises about 1.0% zinc oxide, about 0.5% zinc citrate, about 5% L-arginine, about 1450 ppm sodium fluoride, and optionally about 0.1 wt % benzyl alcohol and / or about 5% small particle silica (e.g., AC43), based on the total weight of the composition.In some embodiments set forth above, the oral care composition may include about 1.0% zinc oxide, about 0.5% zinc citrate, about 1.5% L-arginine, about 0.22%-0.32% sodium fluoride, about 0.5% tetrasodium pyrophosphate, and optionally about 0.1% by weight benzyl alcohol, based on the total weight of the composition. In some embodiments set forth above, the oral care composition may be any one of the following oral care compositions selected from the group consisting of a toothpaste or dentifrice, a mouthwash or rinse, a topical oral gel, and a denture cleanser.

[0108] Example 8

[0109] The oral composition comprising arginine is disclosed in WO 2017 / 003856, which is incorporated herein by reference. In some embodiments, the oral care composition comprises: arginine, zinc oxide and zinc citrate in free or salt form and a fluoride source comprising stannous fluoride. In some embodiments, arginine has an L-configuration. In some embodiments, based on the gross weight of the composition, arginine is present in an amount corresponding to 0.1% to 15%, or 0.1% to 8%, or about 5.0% by weight, or about 8.0% by weight, or about 1.5% by weight, and the weight of arginine is calculated as a free form. In some embodiments, arginine is in free form or partially or completely in salt form. In some embodiments set forth above, the ratio of the amount of zinc oxide (by weight %) to zinc citrate (by weight %) is 2: 1, 2.5: 1, 3: 1, 3.5: 1 or 4: 1, wherein the ratio is by the weight of the overall composition. In some embodiments set forth above, based on the gross weight of the composition, the amount of zinc citrate is 0.25% by weight to 1.0% by weight, and zinc oxide can be present in an amount of 0.75% by weight to 1.25% by weight, or the amount of zinc citrate is 0.5% by weight, and zinc oxide is present in an amount of about 1.0% by weight. In some embodiments set forth above, the fluoride source also includes at least one member selected from the following: sodium fluoride, potassium fluoride, sodium monofluorophosphate, sodium fluorosilicate, ammonium fluorosilicate, fluorinated amines (e.g., N'-octadecyl trimethylenediamine-N, N, N'-tri(2-ethanol)-dihydrofluoride), ammonium fluoride, titanium fluoride, hexafluorosulfate, and combinations thereof. In some embodiments set forth above, based on the gross weight of the composition, stannous fluoride is present in an amount of 0.1% by weight to 2% by weight. In some embodiments set forth above, stannous fluoride is a soluble fluoride salt providing 50 to 25,000 ppm fluoride, or about 750-7000 ppm, or about 1000-5500 ppm, or about 5000 ppm of soluble fluoride. In some embodiments, based on the total weight of the composition, the oral care composition comprises about 1.0% zinc oxide, about 0.5% zinc citrate, about 1.5% L-arginine, about 750-7000 ppm fluoride ions, and optionally about 5% small particle silica (e.g., AC43). In some embodiments, based on the total weight of the composition, the oral care composition comprises about 1.0% zinc oxide, about 0.5% zinc citrate, about 750-7000 ppm stannous fluoride, and optionally about 39.2% glycerol. In some embodiments set forth above, the oral care composition can include about 1.0% zinc oxide, about 0.5% zinc citrate, about 1.5% L-arginine, stannous fluoride, and optionally about 0.1% by weight benzyl alcohol, based on the total weight of the composition.In some embodiments set forth above, the oral care composition can be any of the following oral compositions selected from the group consisting of a toothpaste or dentifrice, a mouthwash or rinse, a topical oral gel, and a denture cleanser.

[0110] Example 9

[0111] Dentifrice compositions having the formulation of Table A were prepared. The compositions had varying amounts of zinc oxide (0 wt % to 2 wt % variations) and amino acids (0 wt % to 5 wt % variations). An example of a dentifrice composition comprises 1 wt % zinc oxide powder, the ZnO being encapsulated into a polymer film at a loading of 50 wt %, particularly a polymer film comprising a combination of two HPMC materials of different molecular weights, the polymer film comprising Methocel E5 and Methocel E50. The combined zinc oxide / polymer film comprises 1 wt % ZnO and 1 wt % polymer film, each weight being based on the total weight of the composition. The illustrated dentifrice composition comprises 4.3 wt % L-arginine.

[0112] Table A

[0113]

[0114] Example 10

[0115] Test dentifrices containing 1% and 2% zinc citrate in combination with 5% arginine, 1450 ppm fluoride and phosphate were prepared as shown in Table B:

[0116] Table B

[0117]

[0118] Embodiment 11

[0119] As shown in Table CG, a test dentifrice containing arginine, zinc oxide, zinc citrate and a fluoride source was prepared: Table C

[0120] Element Compound I Moisturizer 20.0-25.0 Nonionic surfactant 1.0-2.0 Amphoteric surfactant 3.0-4.0 Flavoring / fragrance / coloring agent 2.0-3.0 polymer 10.0-15.0 pH Adjusters 1.5-3.0 Precipitated calcium carbonate 35 Zinc Citrate Trihydrate 0.5 Zinc Oxide 1.0 Sodium Fluoride-USP,EP 0.32 Arginine Bicarbonate 13.86 Demineralized water Moderate

[0121] Table D

[0122] Element Compound A Compound B Compound C Compound D Moisturizer 25.0-40.0 25.0-40.0 25.0-40.0 25.0-40.0 Anionic surfactants 1.0-3.0 1.0-3.0 1.0-3.0 1.0-3.0 Flavoring / fragrance / coloring agent 2.5-4.0 2.5-4.0 2.5-4.0 2.5-4.0 polymer 4.0-6.0 4.0-6.0 4.0-6.0 4.0-6.0 pH Adjusters 5.0-6.0 5.0-6.0 5.0-6.0 5.0-6.0 Synthetic Amorphous Precipitated Silica 16.00 21.37 17.92 7.81 Alumina 0.02 0.01 0.01 0.01 Silicon dioxide - - - 15.0 Lauryl Alcohol 0.02 0.02 0.02 0.02 Zinc Citrate 0.5 0.5 0.5 0.5 Zinc Oxide 1.0 1.0 1.0 1.0 Sodium Fluoride-USP,EP 0.32 0.32 0.32 0.32 L-Arginine Bicarbonate 5.0 5.0 5.0 5.0 Demineralized water Moderate Moderate Moderate Moderate

[0123] Table E

[0124] Element Compound E Compound F Compound G Moisturizer 25.0-40.0 25.0-40.0 25.0-40.0 Anionic surfactants 1.0-3.0 1.0-3.0 1.0-3.0 Nonionic surfactant 0.1-1.0 0.1-1.0 0.1-1.0 Amphoteric surfactant 0.1-1.0 0.1-1.0 0.1-1.0 Flavoring / fragrance / coloring agent 4.0-6.0 4.0-6.0 4.0-6.0 polymer 0.1-2.0 0.1-2.0 0.1-2.0 pH Adjusters 5.0-6.0 5.0-6.0 5.0-6.0 Thickener 6.0 6.5 7.0 Alumina 0.1 0.1 0.1 Synthetic Amorphous Precipitated Silica 17.6 8.8 22.4 Silicon dioxide - 15.0 - Benzyl alcohol 0.1 0.1 0.1 Synthetic Amorphous Silica 5.0 5.0 5.0 Zinc Citrate 0.5 0.5 0.5 Zinc Oxide 1.0 1.0 1.0 Sodium Fluoride-USP,EP 0.32 0.32 0.32 L-Arginine Bicarbonate 1.5 1.5 1.5 Demineralized water Moderate Moderate Moderate

[0125] Table F

[0126] Element Compound H Compound I Moisturizer 45.0-55.0 35.0-45.0 Abrasive 14.0-16.0 9.0-11.0 Anionic surfactants 1.0-3.0 1.0-3.0 Nonionic surfactant 0.1-1.0 - Amphoteric surfactant 1.0-2.0 - Flavoring / fragrance / coloring agent 1.0-3.0 2.0-4.0 polymer 0.1-2.0 3.0-8.0 pH Adjusters 0.1-2.0 4.0-8.0 Silica thickener 5.0 5.0-10.0 Benzyl alcohol 0.1 - Zinc Citrate Trihydrate 0.5 0.5 Zinc Oxide 1.0 1.0 Sodium Fluoride-USP,EP 0.32 0.32 L-Arginine 1.5 5.0 Demineralized water Moderate Moderate

[0127] Table G

[0128] Element Compound J Compound K Compound L Moisturizer 20.0-50.0 20.0-50.0 20.0-50.0 Abrasive 5.0-20.0 5.0-20.0 5.0-20.0 Anionic surfactants 1.0-3.0 1.0-3.0 1.0-3.0 Nonionic surfactant 0.1-1.0 0.1-1.0 0.1-1.0 Amphoteric surfactant 0.1-2.0 0.1-2.0 0.1-2.0 Flavoring / fragrance / coloring agent 1.0-5.0 1.0-5.0 1.0-5.0 polymer 0.1-2.0 0.1-2.0 0.1-2.0 pH Adjusters 0.1-2.0 0.1-2.0 0.1-2.0 Thickener 6.0 6.5 7.0 Dental Silica - - 15.0 High cleaning silica - 15.0 - Synthetic Abrasive 10.0 - - Synthetic Amorphous Silica 5.0 5.0 5.0 Benzyl alcohol 0.4 0.4 0.4 Zinc Citrate Trihydrate 0.5 0.5 0.5 Zinc Oxide 1.0 1.0 1.0 Sodium Fluoride-USP,EP 0.32 0.32 0.32 L-Arginine 1.5 1.5 1.5 Demineralized water Moderate Moderate Moderate

[0129] Example 12

[0130] Test dentifrices containing arginine, zinc oxide, zinc citrate, and stannous fluoride were prepared as shown in Table H:

[0131] Table H

[0132] Element Moisturizer 20.0-60.0 20.0-50.0 20.0-50.0 Abrasive 10.0-40.0 5.0-20.0 5.0-20.0 Anionic surfactants 1.0-3.0 1.0-3.0 1.0-3.0 Amphoteric surfactant 0.5-1.5 0.1-2.0 0.1-2.0 Flavoring / fragrance / coloring agent 0.5-5.0 1.0-5.0 1.0-5.0 polymer 1.0-10.0 0.1-2.0 0.1-2.0 pH Adjusters 1.0-10.0 0.1-2.0 0.1-2.0 Zinc Citrate 0.25-1.0 0.5 0.5 Zinc Oxide 0.75-1.25 1.0 1.0 Stannous fluoride 0.1-1.0 0.32 0.32 L-Arginine 0.1-10.0 1.5 1.5 Demineralized water Moderate Moderate Moderate

[0133] The present invention also relates to, for example, the following technical solutions:

[0134] 1. A method of reducing the total pathogenic oral bacterial load in an individual's oral cavity, the method comprising the steps of: administering 1-50 mg of arginine to the oral cavity.

[0135] 2. The method of claim 1 , wherein the pathogenic oral bacteria in the individual's oral cavity include one or more pathogenic oral bacterial species selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus.

[0136] 3. The method of claim 1 or 2, wherein the beneficial oral bacteria in the individual's oral cavity include one or more beneficial oral bacterial species selected from the group consisting of: Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis.

[0137] 4. The method of any one of items 1-3, wherein the arginine is applied to the oral cavity by using an oral care composition comprising arginine.

[0138] 5. The method of any one of items 1-3, wherein the arginine is applied to the oral cavity by using an oral care composition comprising arginine, wherein the oral care composition is selected from the group consisting of a toothpaste containing arginine and a mouthwash containing arginine.

[0139] 6. A method as described in any one of items 1-6, comprising the step of supplementing the oral cavity with an amount of arginine sufficient to obtain a concentration of at least 20-25 μmol / ml in the biofilm in the oral cavity.

[0140] 7. A method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in an individual's oral cavity, the method comprising the step of: administering 1-50 mg of arginine to the oral cavity.

[0141] 8. A method of selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm in the oral cavity of an individual, the method comprising the step of: administering 1-50 mg arginine to the oral cavity.

[0142] 9. The method of items 7-8, wherein the pathogenic oral bacteria comprises one or more pathogenic oral bacterial species selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus.

[0143] 10. The method of any one of items 7-9, wherein the beneficial oral bacteria comprises one or more beneficial oral bacterial species selected from the group consisting of: Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis.

[0144] 11. The method of any one of items 7-10, wherein the arginine is applied to the oral cavity by using an oral care composition comprising arginine.

[0145] 12. The method of any one of items 7-10, wherein the arginine is applied to the oral cavity by using an oral care composition comprising arginine, wherein the oral care composition is selected from the group consisting of a toothpaste containing arginine and a mouthwash containing arginine.

[0146] 13. A method for selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a bacterial population comprising beneficial oral bacteria and pathogenic oral bacteria, the method comprising the steps of: contacting the bacterial population with arginine at a concentration of at least 20-25 micromoles / ml or more.

[0147] 14. A method for selectively promoting the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria in a biofilm comprising the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria, the method comprising the steps of: contacting the biofilm comprising the growth of beneficial oral bacteria relative to the growth of pathogenic oral bacteria with arginine at a concentration of at least 20-25 micromoles / ml or more.

[0148] 15. A method as described in claim 13 or 14, wherein the bacterial population is contacted with arginine at a concentration of at least 0.05-0.1 mg / ml or higher.

[0149] 16. A method as described in claim 13 or 14, wherein the bacterial population is contacted with arginine at a concentration of at least 0.1-1.0 mg / ml or higher.

[0150] 17. A method as described in claim 13 or 14, wherein the bacterial population is contacted with arginine at a concentration of at least 1-15 mg / ml or higher.

[0151] 18. A method as described in any of items 13-17, wherein the pathogenic oral bacteria include one or more pathogenic oral bacteria species selected from the group consisting of: Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus.

[0152] 19. The method of any one of items 13-18, wherein the beneficial oral bacteria comprises one or more beneficial oral bacteria species selected from the group consisting of: Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis.

[0153] 20. A method of identifying a compound or composition that enhances the selective promotion of growth of beneficial oral bacteria relative to pathogenic oral bacteria by arginine, the method comprising the steps of:

[0154] a) performing a beneficial oral bacteria growth test assay, said beneficial oral bacteria growth test assay comprising culturing a beneficial oral bacteria species in a culture medium comprising arginine and a test compound or composition, and measuring said growth of said beneficial oral bacteria in said beneficial oral bacteria growth test assay;

[0155] b) performing a pathogenic oral bacteria growth test assay comprising culturing a pathogenic oral bacteria species in a culture medium comprising arginine and said test compound or composition, and measuring said growth of said pathogenic oral bacteria in said pathogenic oral bacteria growth test assay;

[0156] c) performing a beneficial oral bacteria growth control assay, said beneficial oral bacteria growth control assay comprising culturing a beneficial oral bacteria species in a medium comprising arginine without said test compound or composition, and measuring said growth of said beneficial oral bacteria in said beneficial oral bacteria growth control assay;

[0157] d) performing a pathogenic oral bacteria growth control assay comprising culturing a pathogenic oral bacteria species in a medium comprising arginine without a test compound or composition, and measuring said growth of said pathogenic oral bacteria in said pathogenic oral bacteria growth control assay;

[0158] e) calculating a ratio of said growth of said beneficial oral bacteria in said beneficial oral bacteria growth test assay to said growth of said beneficial oral bacteria in said beneficial oral bacteria growth control assay;

[0159] f) calculating a ratio of said growth of said pathogenic oral bacteria in said pathogenic oral bacteria growth test assay to said growth of said pathogenic oral bacteria in said pathogenic oral bacteria growth control assay;

[0160] g) comparing the ratio in e) with the ratio in f);

[0161] wherein the ratio in e) is higher relative to the ratio in f) indicates that the test compound or composition enhances the selective promotion of the growth of beneficial oral bacteria over pathogenic oral bacteria by arginine.

[0162] 21. A method as described in claim 20, wherein the culture medium contains 20-25 μmol / ml arginine.

[0163] 22. A method as described in claim 20, wherein the growth of the beneficial oral bacteria in the beneficial oral bacteria growth test assay, the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth test assay, the growth of the beneficial oral bacteria in the beneficial oral bacteria growth control assay, and the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay are measured at 24 hours.

[0164] 23. A method as described in claim 20, wherein the growth of the beneficial oral bacteria in the beneficial oral bacteria growth test assay, the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth test assay, the growth of the beneficial oral bacteria in the beneficial oral bacteria growth control assay, and the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay are measured at 48 hours.

[0165] 24. A method as described in claim 20, wherein the growth of the beneficial oral bacteria in the beneficial oral bacteria growth test assay, the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth test assay, the growth of the beneficial oral bacteria in the beneficial oral bacteria growth control assay, and the growth of the pathogenic oral bacteria in the pathogenic oral bacteria growth control assay are measured by optical density at a wavelength of 630nm.

[0166] 25. The method of any one of items 20-24, wherein the pathogenic oral bacteria is selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus.

[0167] 26. The method of any one of items 20-25, wherein the beneficial oral bacteria is selected from the group consisting of: Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis.

[0168] 27. A method of identifying a compound or composition that enhances the selective promotion of growth of beneficial oral bacteria relative to pathogenic oral bacteria in a biofilm by arginine, the method comprising the steps of:

[0169] a) performing a dual species biofilm test assay, the dual species biofilm test assay comprising

[0170] i) co-culturing a beneficial oral bacterial species and a pathogenic oral bacterial species in a dual-species test biofilm comprising arginine and a test compound or composition,

[0171] ii) quantifying beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm; and

[0172] iii) comparing the number of beneficial oral bacteria to the number of pathogenic oral bacteria in the dual-species test biofilm to determine the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm;

[0173] b) performing a dual-species biofilm control assay comprising

[0174] i) co-culturing a beneficial oral bacterial species and a pathogenic oral bacterial species in a two-species control biofilm comprising arginine but without the test compound or composition,

[0175] ii) quantifying beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm; and

[0176] iii) comparing the number of beneficial oral bacteria to the number of pathogenic oral bacteria in the dual-species control biofilm to determine the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm;

[0177] c) comparing the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species test biofilm to the beneficial oral bacteria-pathogenic oral bacteria load of the dual-species control biofilm;

[0178] wherein a higher beneficial oral bacteria-pathogenic oral bacteria load in the dual-species test biofilm compared to the beneficial oral bacteria-pathogenic oral bacteria load in the dual-species control biofilm indicates that the test compound or composition enhances the selective promotion of arginine on the growth of beneficial oral bacteria relative to pathogenic oral bacteria in a biofilm.

[0179] 28. The method of claim 27, wherein the dual-species test biofilm comprises 20-25 micromoles / ml arginine and the dual-species control biofilm comprises 20-25 micromoles / ml arginine.

[0180] 29. The method of item 27, wherein the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm and the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm are determined at 24 hours.

[0181] 30. The method of item 27, wherein the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm and the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm are determined at 48 hours.

[0182] 31. The method of claim 27, wherein the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species test biofilm and the number of beneficial oral bacteria and pathogenic oral bacteria in the dual-species control biofilm are determined by qPCR.

[0183] 32. The method of any one of items 27-31, wherein the pathogenic oral bacteria is selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, and Streptococcus sobrinus.

[0184] 33. The method of any one of items 27-32, wherein the beneficial oral bacteria is selected from the group consisting of: Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, and Streptococcus sanguinis.

Claims

1. A use of an oral care composition for increasing the growth of beneficial oral bacteria and reducing the presence of pathogenic oral bacteria in the oral cavity of a subject in need thereof, the oral care composition comprising: arginine in an amount effective to obtain an arginine concentration of 10-25 micromoles / ml in the oral cavity; zinc oxide; and zinc citrate.

2. The use according to claim 1, wherein the pathogenic oral bacteria is selected from the group consisting of Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Streptococcus mutans, Streptococcus sobrinus, and a combination of two or more thereof.

3. The use according to claim 1 or 2, wherein the beneficial oral bacteria is selected from the group consisting of: Streptococcus gordonii, Actinomyces viscosus, Streptococcus salivarius, Streptococcus oralis, Streptococcus mitis, Streptococcus sanguis, and a combination of two or more thereof.

4. Use according to any one of the preceding claims, wherein the beneficial oral bacteria are increased by at least 1.2 times.

5. Use according to any one of the preceding claims, wherein the bacterial population is contacted with arginine at a concentration of at least 0.05-0.1 mg / ml or more.

6. Use according to any one of the preceding claims, wherein the bacterial population is contacted with arginine at a concentration of at least 0.1-1.0 mg / ml or more.

7. Use according to any one of the preceding claims, wherein the bacterial population is contacted with arginine at a concentration of at least 1-15 mg / ml or more.

8. Use according to any one of the preceding claims, wherein the oral care composition is selected from toothpaste and mouthwash.

9. Use according to any one of the preceding claims, wherein the weight ratio of zinc oxide to zinc citrate is from 1.7:1 to 2.3:

1.

10. Use according to any one of the preceding claims, wherein the weight ratio of the zinc oxide to the zinc citrate is about 2:1.

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