Dental paste

Abstract

The present invention relates to a gel or paste having a composition comprising at least 2 to 4% w / w of a thickener, optionally agar, at least 50 to 95% w / w of glycerol, and at least 0.03 to 3% w / w of an epithelial-adherent protein complex. The gel or paste of the present invention is formulated to deliver and maintain a local and substantial effect from the activity of the protein complex on the oral mucosa, thereby enabling the restoration and maintenance of oral health in mammals or animals, such as dogs.

Description

Background Technology

[0001] Delivering bioactive components to maintain and / or improve the health of humans or animals, such as companion animals, presents various challenges to formulators, which vary particularly depending on the type of formulation to be delivered and the target site.

[0002] Many bioactive agents are susceptible to degradation, and exposure to oxygen, heat, light, and / or moisture can substantially eliminate the health benefits of their bioactivity. Since formulations used to deliver bioactive ingredients require inherent protection of the activity, they may include solid dosage forms such as capsules or tablets.

[0003] Unfortunately, traditional encapsulation or other processing methods that provide a physical barrier to help successfully reach or access the target site are not always useful. For example, the mouth, which defines the oral cavity of mammals, humans, or animals such as pets, includes the oral mucosa (gums, lips, cheeks, palate, and floor of the mouth), tongue, and teeth. When the target site is located within the structures of the mouth, the formulation must be maintained proximal for the minimum possible duration and localized to deliver the required efficacy of the bioactive ingredient.

[0004] However, delivery of many oral health products formulated for use in the mouth, such as mouthwashes, lozenges, and pastes, is unreliable, / unavailable, or inconsistent, and a sufficient amount of bioactivity to produce a measurable effect (e.g., on the mucosal membrane or tongue) never reaches.

[0005] This is particularly true in cases involving the administration of oral health products to dependent subjects, such as children, the elderly, or animals, such as companion animals. Furthermore, due to the risk of involuntary ingestion of the active substance, it is difficult to overcome undesirable effects associated with the specific task mentioned above; this includes reliably controlling efficacy and toxicity and preventing related side effects, such as gastrointestinal sensitivity and nausea, that may result therefrom.

[0006] Common oral diseases often arise from changes in the balance of the natural flora within the biofilm (plaque) covering the teeth and gums. The bacteria contained within plaque are generally benign, and the major components of the biofilm are typically commensal bacteria, such as Lactobacillus However, if pathogenic bacteria become the dominant bacteria, oral health can deteriorate. As a result, dental caries and / or gum disease may begin to develop in the mouths of mammals, especially if, for example, the teeth of pets are not cleaned regularly to remove plaque buildup.

[0007] Colonization of pathogenic species is known to weaken and dissolve dental enamel, causing caries or lesions on the teeth, and exacerbates the problem by the formation of calcified deposits, which provide the colonies with a local base where they can continue to thrive on the teeth. This process contributes to frequently recorded symptoms of poor oral health, such as bad breath, inflammation of gingival tissue, and bleeding of the gums.

[0008] To prevent oral disease and / or maintain oral health, oral care products, including those used in the veterinary field, often contain fungicides to reduce pathogen levels. However, since these agents are generally non-selective, they serve to destroy all natural biofilms, including commensal species. This generally does not provide a useful therapy for mammals, and especially companion animals, because restoring the balance of the flora is necessary to restore and / or maintain long-term oral health.

[0009] EP3424516 is sensuously (literally [sic]) neutral and Streptococci We propose a composition containing binder lactic acid bacterium microorganisms that reduce colonies. This composition can be added to pet food, and does not cause taste or odor issues that generally hinder pet compliance when using the effective disinfectant proposed by the authors. However, rather than effectively restoring the natural balance of the entire mucosal flora, this solution StreptococciThe goal is to reduce only. The described solution is formulated and administered as part of a feed that must be chewed for application. The problem to be solved

[0010] The invention of the present invention originated from the desire to overcome the aforementioned problems, and the present invention provides a dental composition that effectively restores and / or maintains the oral health of mammals and, in particular, companion animals. means of solving the problem

[0011] The invention disclosed herein relates to a dental paste or gel comprising at least 2 to 4% w / w of a thickener, preferably agar; 62 to 95% w / w of glycerol; and 0.03 to 3% w / w of an epithelial-adhering protein complex (EAPC).

[0012] The epithelial-adhesion protein complex may comprise one or more of the following: 1) a microbiome regulatory protein or "protein fraction" and 2) a modified whey protein isolate (WPI) from milk.

[0013] Microbiome regulatory protein(s) may be selected from lactoperoxidase, lactoferrin, lysomaal alpha-mannosidase, immunoglobulin G, angiogenin, ribonuclease 4, or quesine sulfhydryl oxidase and / or combinations thereof. Modified WPI is formed from soluble native protein, protein aggregate, and protein fibrils prepared from whey protein isolate (WPI).

[0014] Dental pastes or gels can be effectively applied directly to the structures of the oral mucosa by brushing or coating. The novel form of the dental paste ensures that the complex remains at the application site, namely near relevant structures such as gums and teeth. The composition is not immediately washed away or swallowed and remains stable during delivery; when the active ingredient is exposed to the target site, the selective antibacterial properties are effective in situ. In particular, bioactive ingredients such as microbiome-regulating proteins are generally unstable and may lose their activity prematurely relative to the shelf life of the paste product. Since the dental paste or gel protects microbiome-regulating proteins by reducing or preventing the absorption of oxygen, moisture, and / or light, efficacy is not affected during storage or delivery. This formulation acts as a temporary barrier to protect the protein complex and its inherently microbiome-supporting selective antibacterial properties. Dental pastes or gels specifically stabilize the complex for a longer period so that the bioactivity maintains efficacy and effect when delivered to site-specific structures within the oral mucosa.

[0015] Nevertheless, the gel or paste formulated according to the present invention can protect and maintain selective antimicrobial activity and deliver a bioactive protein complex locally within the oral mucosa for an effective period, thereby enabling the substantive effects observed from the bioactivity regarding the recovery and maintenance of oral health in mammals or animals, such as dogs.

[0016] In addition, the manufactured dental paste or gel is soft, malleable, and has a moist structure, so it can be easily applied to the teeth and gums of pets, such as cats and / or dogs, for example, as an instant fluid gel coating applied to the surface of a brush or toy. Advantageously, compared to traditional delivery vehicles such as tablets or capsules, this formulation is palatable. The invention extends to chew articles or animal toys coated with the dental paste or gel strip of the present invention.

[0017] In one embodiment, the thickener component, preferably agar, is 2 to 3% w / w, and in some preferred embodiments, the thickener or agar is approximately 2.6% w / w. A paste formulation having a combination of glycerol and at least one thickener, such as agar, specifically maintains low water activity in the final dental paste. This is important to ensure that all functions of other essential components within the epithelial-adhesion protein complex of the paste, namely stability and bioactivity, are maintained. If any of these are lost, the final product paste may not be suitable for use as a dental paste.

[0018] A thickener, preferably agar, serves to control the viscosity of glycerol in the final paste. In one embodiment, other thickeners, for example, including carrageenan and / or xanthan gum, may also be used in the formulation. However, agar is preferred. Additionally, the texture of the paste may be further controlled by adding one or more other textural agents that may also have a thickening effect, such as rice flour, chitosan and / or silica.

[0019] In one embodiment, the glycerol component is 80 to 95% w / w. In a further embodiment, the glycerol component comprises at least 90 to 95% w / w.

[0020] In one embodiment, the composition comprises at least 0.03% w / w of an epithelial-adhesion protein complex, optionally this component is present in a range of at least 0.1 to 3%, 0.5 to 3%, 1 to 3%, 2 to 3%, or 2.5 to 3% w / w.

[0021] The composition of the present invention may include up to 32% w / w of other / additional excipients designed to improve the texture of the composition, including additional thickeners or stabilizers, such as chitosan or rice flour.

[0022] In one embodiment, other gums and excipients may be used to improve the consistency of the composition, including but not limited to arable gum, xanthan gum, alginate, chitin, aloe extract, guar gum, hydroxypropyl cellulose, sodium casein, corn starch, carboxymethylcellulose, gelatin, agar, dextrin, methylcellulose, polyvinyl alcohol, microfiber cellulose, microcrystalline cellulose, seaweed cellulose, sodium polyacrylate, sodium polyphosphate, carrageenan, or yeast cell wall.

[0023] The present invention extends to a dental paste or gel according to any prior claim for use as a pharmaceutical, optionally as a veterinary medicament.

[0024] In one embodiment, the paste or gel is intended for use in the treatment or prevention of oral diseases, particularly oral diseases of canines mediated by an excess of pathogenic bacteria. The oral disease may be selected from, but is not limited to, dental caries, gingivitis, and / or tartar.

[0025] The paste or gel composition may also be used for veterinary treatment to restore the oral health of animals, particularly canids or felines.

[0026] Dental paste is useful for combating chronic and harmful oral health conditions in animals, such as bacterial infections (especially when the disease may be caused by an increase in pathogenic bacteria).

[0027] As previously described, an increase in pathogenic bacteria generally causes harmful oral health conditions in animals, including one or more of halitosis, plaque, and / or gum disease. In one embodiment, an increase in pathogenic bacteria causing these harmful conditions Psychrobacter, Porphyromanas, Pasteurella and / or Actinomyces It may contain bacteria inside.

[0028] Poor health in animals can be attributed to a decrease in symbiotic bacteria, which leads to an imbalance in the microbiota of the oral mucosa or, in particular, plaque. Since this imbalance can result in one or more conditions selected from halitosis, plaque, and / or gum disease, the use of the paste can help restore this balance, particularly in dogs, as has been adequately demonstrated herein. It may also be suitable for other animals, particularly cats, that regularly suffer from periodontal disease and tooth loss due to caries.

[0029] The paste is gentle on the digestive system even if partially swallowed, and therefore can be used as part of and / or frequently used as a long-term viable oral solution to maintain the oral health of animals over a reasonable period of months or years.

[0030] In one embodiment, when the dental paste is used for the prevention, treatment, and / or improvement of oral health, and more specifically dental health, an appropriate dosage regimen may be followed. To reduce the number of pathogens and significantly improve oral health, this composition may be most effective when the final product is used repeatedly on a daily basis. When applied as a single daily dose, the product delivers a dose of at least 5, 15, or 25 to 50 mg of epithelial-adhesion protein complex.

[0031] In some embodiments, where a smaller volume of paste product is required to deliver a particularly effective daily dose (e.g., a single small paste ball (suitable for brushing or direct application)) to the tooth, the epithelial-adhesion protein complex is provided in a higher range, e.g., 1.5 to 3% w / w. In other embodiments, for example, where a larger volume of gel product is required to cover a much larger surface area for toy coating, the daily dose required for effect can be achieved with a lower % composition, e.g., as low as 0.03%.

[0032] Depending on the final formulation and product form, the paste or gel required to provide an effective amount of active ingredient for a given single application may be about 1 g to 50 g by weight according to the scope of the invention and may have a relevant % EAPC.

[0033] Preferably, the product is applied daily for at least 5 consecutive days of 7 days per week, and more preferably for at least 4 weeks.

[0034] Once a rebalanced microbiome is established, the same dose can be used at a reduced frequency, such as every other day or twice a week, to maintain health through a properly rebalanced microbiome of the oral mucosa. However, regular administration will more effectively prevent the recurrence of pathogen accumulation and thus maintain a properly rebalanced oral mucosa.

[0035] In an embodiment, the present invention further relates to a method for improving or maintaining oral health, comprising administering the dental gel or paste composition described above to a human or animal in need thereof. In an embodiment, the present invention further comprises a method for preventing or treating oral diseases, comprising administering the composition described herein to a mammal and / or animal in need thereof.

[0036] The present invention is further extended to a method for manufacturing a dental paste or gel, which comprises at least the following steps:

[0037] A step of providing glycerol at least 50%, optionally 62% to 95% w / w, and optionally heating to up to about 95°C;

[0038] A step of adding and dissolving a thickener, preferably agar, in an amount of 2 to 4% w / w to the glycerol;

[0039] Optionally, adding one or more excipients in an amount of 0 to 32% w / w to a combination of glycerol and a thickener, and cooling, optionally cooling to 40°C or lower;

[0040] A step of adding and dissolving an epithelial-adhesion protein complex of 0.03 to 3% w / w; and

[0041] A step of cooling, optionally to approximately 18°C ​​or lower, to form a dental paste or gel.

[0042] Consequently, the manufactured dental paste is formulated to deliver and maintain a local and substantial effect from the activity of the protein complex. As described in more detail below, the applicant's research showed that the novelly formulated dental paste composition maintained significant bioactivity at surprisingly low concentrations. This suggests that the form formulated according to the embodiments of the invention further enhances the effect of EAPC beyond what could be expected. Subsequent animal studies also provided evidence of the translation of bioactivity to mammals, leading to the inference that the restoration and / or maintenance of oral and dental health is achieved when the dental paste of the invention is applied to the oral mucosa of mammals.

[0043] Furthermore, the texture of the paste or gel produced by these means is also more suitable for easier oral administration to certain mammalian groups, such as infants, the elderly, and companion animals, such as dogs. Once accepted in the mouth, the recipient is more likely to accept the composition on, for example, a brush or other surface applicator, rather than automatically rejecting a capsule or tablet with a hard or unpleasant taste. The dental paste or gel composition may additionally include one or more excipients to optimize and improve the recipient's palatability. Brief explanation of the drawing

[0044] Further aspects of the present invention are provided as examples and will become clear from the following description with reference to the accompanying drawings: Figure 1 shows commensal bacteria. L. acidophilus This is a graph showing the nil effect of the EAPC used in the paste of the present invention on the growth; Figure 2 shows pathogenic bacteria E. coli This is a graph showing the sterilization effect of EAPC used in the paste; Figure 3 shows that the EAPC used in the paste adheres well to the epithelial surface and pathogenic bacteria S. aureus This is a graph showing that it maintains bactericidal activity against; FIG. 4 is a graph representation of data showing that the activity of lactoperoxidase, a key microbiome regulatory protein, is sufficiently maintained in the dental paste of the present invention after preparation (according to the disclosures of the present invention and subsequent storage); Fig. 5 shows the period of 12 hours and 40 minutes (760 minutes) under laboratory conditions. B. subtilis This is a graph showing the effect of each paste sample on the growth rate; Fig. 6 is B. subtilis Graphing the concentration of active microbiome regulatory proteins in each type of paste sample required to inhibit growth (relative level 0% in the case of complete inhibition); FIG. 7 compares the abundance of six types of key pathogenic bacteria associated with dental disease in canines after treatment with the test dental paste of the present invention, using a graph against a placebo in a canine microbiome test; Figure 8 is a graph showing the statistical correlation between the use of the test dental paste of the present invention and the effect compared to a placebo paste in a microbiome test of canine animals. Specific details for implementing the invention

[0045] For the purposes of this specification, the terms “about” or “approximately” and their grammatical variations mean a quantity, level, degree, value, number, frequency, percentage, dimension, size, amount, weight, or length that varies by up to 0.1% with respect to a reference quantity, level, degree, value, number, frequency, percentage, dimension, size, amount, weight, or length. The term “substantially” or its grammatical variations mean at least about 50%, e.g., 75%, 85%, 95%, or 98%.

[0046] The term 'comprise(s)' and its grammatical variations must have an inclusive meaning; that is, they should be interpreted to include not only the listed components that are directly referenced but also other unspecified components or elements.

[0047] The applicant's work stems from further research, investigation, and improvement of recently developed formulations, and is particularly related to the recovery, improvement, and maintenance of oral health in companion animals, especially canines.

[0048] Microbiome regulatory proteins

[0049] The enzyme lactoperoxidase is a protein involved in microbiome regulatory proteins isolated from bovine milk (NZ547859). This publication describes a combination of protein fractions from bovine dairy milk, for example, Lactobacillus Without affecting the growth of symbiotic bacteria of the family E. coli and S. aureus It is explained that it may be effective in preventing the growth of pathogenic bacteria such as [specific bacteria]. Formulations based on milk bioactive proteins extracted from milk have been disclosed in this field of technology. These ingredients are naturally produced by cows through immune defense responses against infection and inflammation. These anti-inflammatory, antioxidant, and antibacterial actions support the 'good' bacterial flora and kill 'bad' bacteria, and have been shown to be safe in some clinical applications. However, at the time, practical use for dental or oral applications in mammals or companion animals was not possible. This is because these proteins alone cannot deliver their beneficial properties, and formulations encompassing these agents were not identified as effective.

[0050] Prior to the author's own research and development regarding these components, there was no means to successfully utilize the microbiome-modulating activity of these proteins and / or their fractions (or very similar types of functional agents) for effects on the oral mucosa.

[0051] Nevertheless, the applicant has determined a method for successfully complexing microbiome-regulating proteins, which introduces novel physical functions to stabilize the proteins and enable the complexed proteins to adhere to epithelial surfaces upon topical application. This invention will enable a product that not only maintains excellent protein functionality (selective bactericidal quality) but also possesses additional properties necessary to overcome unmet challenges (e.g., topical adhesion, which may be suitable for oral mucosa). The successful results of these protein complexes are attributed in part to the critical chemical and physical conditions under which the two components react together to form the complex in the applicant's process, which have been diligently studied and controlled to optimize the functional activity of the two components.

[0052] This protein complex or "epithelial adhesion protein complex" EAPC is one of the essential components of the dental paste or gel of the present invention disclosed herein. However, at the time of the disclosure of the present invention, the protein complex component was not publicly disclosed.

[0053] Accordingly, a complete description of the component and how it is obtained is provided herein, and for the first time, how EAPC is effectively used in the formulation and manufacture of the new dental paste of the present invention and in embodiments thereof is described.

[0054] Epithelial-Adhesion Protein Complex "EAPC"

[0055] The present invention discloses a protein complex of the applicant comprising a microbiome regulatory protein complexed with a modified whey protein isolate (WPI) from milk, wherein the complexed microbiome regulatory protein is configured for topical application and to adhere to an epithelial surface upon application. The microbiome regulatory protein is selectively toxic to pathogenic bacteria and minimizes any reduction of the commensal bacteria population.

[0056] The microbiome regulatory proteins in the complex may include at least one of lactoperoxidase, lactoferrin, lysomal alpha-mannosidase, immunoglobulin G, angiogenin, ribonuclease 4, quercetin sulfhydryl oxidase, and combinations thereof.

[0057] Microbiome regulatory proteins

[0058] For the sake of brevity, the term 'protein' or its grammatical variations as used herein are intended to encompass a single protein, multiple proteins, a single protein fraction, or protein fractions, and references to a singular protein should not be considered limiting.

[0059] Microbiome regulatory proteins may be a blend of protein fractions from milk that is selectively toxic to pathogenic bacteria but minimizes any reduction in symbiotic bacterial populations.

[0060] Microbiome regulatory proteins may include lactoperoxidase, lactoferrin, lysomaal alpha-mannosidase, immunoglobulin G, angiogenin, ribonuclease 4, quercetin sulfhydryl oxidase, and combinations thereof. These proteins may include a selection or all of some of the proteins listed above. The microbiome regulatory proteins selected for the method may initially be in the form of a dry powder.

[0061] Modified whey protein isolate (WPI)

[0062] As used herein, the term 'modified whey protein isolate' or its grammatical variations refer to soluble native proteins, protein aggregates, and protein fibrils prepared from whey protein isolate (WPI). The modified WPI selected for the above method may initially be in the form of an aqueous gel. The selected modified WPI may be derived from the milk of bovine species, but may also be sourced from modified WPI obtained from other animals such as goats and sheep.

[0063] This gel enables not only the binding of active ingredients but also the controlled release of the formulation. It possesses shear-thinning rheology (thixotropic) and provides excellent stability and a positive skin texture. It consists of a semi-solid material composed of soluble native proteins, protein aggregates, and protein fibrils prepared from whey protein isolate (WPI), forming a modified WPI. The modified WPI gel used herein generally has a viscosity of 0.25 to 4.5 Poise.

[0064] Method for preparing epithelial-adhesion protein complex (EAPC)

[0065] A method for manufacturing EPAC comprises the steps of: selecting a microbiome regulatory protein in powder form; selecting a modified whey protein isolate (WPI) from milk in gel form; dissolving the microbiome regulatory protein in an aqueous salt solution having an ionic strength of 25–200 mM NaCl; blending the dissolved microbiome regulatory protein and the modified WPI together; adjusting the pH to 2.0–6.0; optionally adjusting the temperature to 18–37°C; and maintaining the blend at a pH of 2.0–6.0 and a temperature of 18–37°C for at least 30 minutes.

[0066] The following flowchart explains the EAPC manufacturing method:

[0067]

[0068]

[0069] The resulting EAPC can be a semi-solid aqueous gel with a viscosity of approximately 0.25 to 4.5 poise. The viscosity of the modified whey protein isolate is important for facilitating complexation, and the optimal temperature for the most suitable viscosity is in the range of 18–37°C. The ionic strength of the mixture is important for the proper interaction of protein groups in both components, and the optimal ionic environment is maintained by 25–200 mM NaCl at pH 2.0–6.0.

[0070] The ratio of microbiome regulatory protein to modified WPI present in the complexed protein used in and / or disclosed in the above method may be 1:1, or 1:2, or 1:3, or 1:4, or 1:5, or 1:6, or 1:7, or 1:8, or 1:9, or 1:10. For example, the ratio may be 1:1 to 1:10, i.e., 1 part of microbiome regulatory protein to 1 to 10 parts of modified whey protein isolate. The ratio may be 1:1.

[0071] To ensure that the required degree of complexation is achieved, it may be necessary to have at least that much modified WPI for the protein. A lower ratio may be used, but at lower ratios, waste or non-complexed protein may be generated in the above method due to the presence of insufficient modified WPI.

[0072] Salt

[0073] The salt solution may contain a mixture of water and a chemical salt. The chemical salt may be NaCl, but other salts such as MgCl may also be used. The salt may be added in an amount necessary to provide the indicated ionic strength. The ionic strength of the salt solution may be approximately 25, or 50, or 75, or 100, or 125, or 150, or 175, or 200 mM NaCl. As mentioned above, the ionic strength may vary between 25 and 200 mM NaCl. The ionic strength may be approximately 75 mM NaCl. This ionic strength is maintained in the complexed blend of protein and modified WPI during and after holding.

[0074] This range of ionic strength appears desirable for the dissolution or complexation of microbiome-regulating proteins into modified WPI, but some variation may occur depending on the pH, temperature, and holding time used. The salt solution is important for complex formation by establishing the correct ionic nature of the solution, ensuring the dissolution of powdered proteins, and allowing the proteins to carry the correct charge.

[0075] Blending

[0076] Blending is performed gently to ensure that foaming of the mixture is absent or minimized during blending. Blending can be completed, for example, using a planetary mixer. Blending may continue during pH adjustment, temperature adjustment, and / or holding. Gentle or slow blending of proteins can be important to minimize or prevent denaturation or loss of function.

[0077] pH

[0078] The pH of the blend before pH adjustment may be approximately 6.5 to 7.5 or approximately neutral pH 7.0. This starting point of pH may vary depending on the pH of the water used to form the aqueous salt solution. The pH of the blend after pH adjustment may be approximately 2.0, or 2.5, or 3.0, or 3.5, or 4.0, or 4.5, or 5.0, or 5.5, or 6.0. As mentioned above, in one embodiment, the pH may be 2.0 to 6.0. In one embodiment, the pH may be approximately 4.0. While this pH range and value appear to be optimal for the dissolution or complexation of the protein into the modified WPI, some variation may occur depending on the ionic strength, temperature, and holding time used. Acidic conditions after adjustment appear to maintain an appropriate charge on the protein to ensure the binding of the complex. The pH may be adjusted using an acid. The acid may be hydrochloric acid, but other acids may also be used. According to the inventor's experience, a buffer is not required to maintain the reduced pH once adjusted.

[0079] temperature

[0080] After any adjustment (if necessary), the temperature of the mixture may be approximately 18, or 19, or 20, or 21, or 22, or 23, or 24, or 25, or 26, or 27, or 28, or 29, or 30, or 31, or 32, or 33, or 34, or 35, or 36, or 37°C. As mentioned above, after any adjustment, the temperature may be 18–37°C. In one embodiment, the temperature after adjustment may be approximately 25°C. The temperature may be adjusted to maintain the modified WPI at a desired viscosity that enables optimal complexation. This desired viscosity may be in the range of approximately 0.25 to 4.5 poise. While the described temperature ranges and values ​​appear to be optimal for the dissolution or complexation of the protein into the modified WPI, some variation may occur depending on the ionic strength, pH, and holding time used. The temperature can be adjusted from ambient conditions (or maintained from ambient conditions if the ambient temperature is within the desired range).

[0081] Temperature adjustment can be made up or down depending on the difference between the desired temperature and the ambient temperature.

[0082] Maintenance time

[0083] The retention time may be at least 30 minutes. The retention time may be from 30 minutes to 24 hours. In one embodiment, the retention time may be approximately 60 minutes. In the inventor's experience, most complexation occurs within 30 to 60 minutes based on selected parameters. An extended period may be used, and based on the applicant's experience; there is no harm or risk of loss of function in retaining the mixture for a longer period. This timing appears to be optimal for the dissolution or complexation of the protein into modified WPI, but some variation may occur depending on the ionic strength, temperature, and pH used.

[0084] dry

[0085] Optionally, drying may be completed prior to subsequent formulation. In this embodiment, the dried complex may be solid. It may be dried such that the water activity is less than 0.6. In a selected embodiment, the water activity may be as low as 0.2 to 0.3. The drying of the complex may be performed under specific conditions of low temperature and low pressure over a time course that ensures the epithelial adhesion properties and the selective bactericidal activity of the complex components are maintained. Pressure during drying is important because, at low pressure, water boils off from the gel at lower temperatures, allowing drying to occur at low temperatures. Additionally, very gradually reducing the pressure may be important to prevent the complexed protein solution in the dryer from foaming. Drying may be completed by freeze-drying or any other gentle drying process using non-denaturing temperature / pressure (e.g., vacuum below 30°C). Complexed proteins are functionally maintained when rehydrated after drying.

[0086] Beneficial effects in the resulting protein complex could only be obtained through such careful experimentation and the balancing of parameters. Indeed, the degree of protein adhesion to the epithelial surface after complexation exceeded twice that of each component alone; based on the inventors' experience, this suggests that an inherently synergistic effect, or at least one that far exceeded expectations, was possible, indicating that previously unforeseen or predictable synergistic effects were possible.

[0087] It was observed that the selective bactericidal activity of the protein described herein may be negatively affected if the temperature of any step of the process exceeds 37°C and the pH is less than 2.0 or exceeds 9.0.

[0088] Attachment of EAPC to the epithelial surface

[0089] To determine whether the complex adheres to the epithelial layer, a portion of pig skin was immersed in a 50 mg / mL solution of the complex (containing 50 mg / mL protein), washed twice with a pH 7.4 phosphate buffer, and stained with Coomassie Blue, which stains proteins. This resulted in skin that was more strongly stained compared to pig skin not immersed in the complex (Fig. 1). When pig skin was treated with either the 50 mg / mL modified whey protein isolate fraction (containing 50 mg / mL protein) or the microbiome regulatory protein from bovine milk (containing 50 mg / mL protein) alone, followed by washing, it was found that these components bound to the skin to some extent, but not to the same degree as the 50 mg / mL complex of the two components.

[0090] Pig skin samples treated with or not treated with EAPC, and pig skin samples treated with individual components of EAPC, were washed twice, and proteins attached to the epidermal layer of the pig skin were stained.

[0091]

[0092] The epithelial binding activity of the complex was significantly greater (more than double) than that of modified whey protein isolate alone or microbiome regulatory protein alone. The modified whey protein isolate was specially prepared to attach to epithelial surfaces. Therefore, the binding amount of this component to porcine skin is expected to be similar to that of the complex. However, the binding of the complex was 44% greater than that of the modified whey protein isolate alone, suggesting a synergistic effect of the components or that the process used to generate the complex increased the adhesion ability of the protein complex.

[0093] Retention of enzyme bioactivity

[0094] Lactoperoxidase enzyme is a protein involved in microbiome regulation proteins isolated from the milk of bovine animals (NZ547859 and PCT / NZ2017 / 050043). Lactoperoxidase activity was measured by adding the complex or its components individually to a hydrogen peroxide solution in 100 mM phosphate buffer (pH 5.5) and monitoring the oxidation of 2,2-azino-bis(3-ethylbenzthiazolin-6-sulfonic acid) (ABTS) at 436 nm using a spectrophotometer.

[0095] The lactoperoxidase activity of a combination of microbiome regulatory proteins from bovine milk was maintained when the microbiome regulatory proteins were complexed with modified whey protein isolate to form EAPC. As expected, the modified whey protein isolate had no peroxidase activity.

[0096] Lactoperoxidase activity of EAPC compared to complex components:

[0097]

[0098] Specific antibacterial activity

[0099] Symbiotic bacteria

[0100] Microbiome regulatory proteins isolated from bovine milk Lactobacillus It was shown that it does not affect the growth of symbiotic bacteria such as those derived from the same family (PCT / NZ2017 / 050043). EAPC containing these microbiome regulatory proteins is symbiotic bacteria Lactobacillus acidophilus To confirm the effect on [it], bacteria were cultured anaerobically, the complex was added to the culture, and the bacterial count after 24 hours was determined by optical density at 650 nm. The complex of modified whey protein and microbiome regulatory proteins isolated from bovine milk was symbiont bacteria Lactobacillus acidophilus It did not affect the growth of ( Fig. 1 ).

[0101] Pathogenic bacteria

[0102] Microbiome regulatory proteins isolated from bovine milk E. coli It was shown to have a bactericidal effect against pathogenic bacteria such as (PCT / NZ2017 / 050043). EAPC containing microbiome regulatory proteins E. coli To determine the effect on [it], these bacteria were cultured, EAPC was added to the culture, and the bacterial count after 24 hours was determined as optical density at 650 nm. EAPC [is] pathogenic bacteria to a degree similar to that of microbiome regulatory proteins from bovine milk alone. E. coli inhibited the growth of ( Fig. 2 The square data points in Figure 2 represent the results when microbiome regulatory proteins are added, and the circular data points represent the results when EAPC is added.

[0103] Effects of skin-bound EAPC on the growth of pathogenic bacteria

[0104] S. aureus is a pathogenic bacterium often associated with skin diseases. Microbiome regulatory proteins isolated from bovine milk were grown by culture S. aureus It was shown to have a sterilizing effect (PCT / NZ2017 / 050043). Pig skin samples washed after treatment with a microbiome-regulating protein from bovine milk were S. aureus It showed gradual inhibition of growth. However, porcine skin samples washed after treatment with EAPC were S. aureus It showed much greater inhibition of growth ( Fig. 3 This confirms the results of Table 1 and confirms that EAPC binds well to epithelial surfaces such as skin and maintains the bactericidal activity of microbiome regulatory proteins from cow's milk.

[0105] Dental Paste - Formulation and Manufacturing Method

[0106] A method for manufacturing a dental paste or gel according to an embodiment of the present invention comprises at least the following steps:

[0107] A step of heating 62 to 95% w / w of glycerol, optionally at approximately 95°C;

[0108] A step of blending 2 to 4% w / w of a thickener, optionally agar, with the glycerol;

[0109] A step of optionally adding one or more excipients in the range of 0 to 45%, optionally 0 to 32% w / w, to a combination of glycerol and a thickener, and cooling, optionally cooling to 40℃ or lower;

[0110] A step of adding and dissolving an epithelial-adhesion protein complex (EAPC) in an amount of 0.03 to 3% w / w; and

[0111] A step of cooling, optionally to approximately 18°C ​​or lower, to form a dental paste or gel.

[0112] For example, thickener blending is performed in a step-wise manner, preferably at approximately 90 to 100°C. For example, cooling is performed in a step-wise manner at 40°C to 18°C.

[0113] working range

[0114] During additional formulation studies, additional in vitro and in vivo Before conducting the activity study, it was evident that some ingredients and ingredient ranges could be slightly modified.

[0115] The following working range was additionally determined in the test:

[0116]

[0117] Accordingly, in addition to the specific examples provided below, the agar component may be replaced with at least one alternative thickener that achieves the same function. To enhance the function of controlling the viscosity of glycerol, other additional textural agents may be included to provide a higher total thickener.

[0118] The table below illustrates the composition of the dental paste provided in some possible embodiments:

[0119]

[0120] Dental Paste Manufacturing and Effects / Applications

[0121] Accordingly, the applicant has developed a new process and a dental gel or paste composition that enables selective bactericidal activity, which, when applied to the teeth of companion animals, such as canines, is specifically delivered to the oral mucosa for the first time and remains for a sufficiently long time to continuously deliver an effective amount of EAPC.

[0122] By doing so, oral health can be restored and symptomatic signs of oral disease in canids and companion animals can be reduced. However, to ensure these effects, it was important to confirm that an appropriate level of bioactivity is maintained in the product claimed and manufactured by the process described herein. The following tests were performed to verify these claims.

[0123] Bioactivity - Preliminary Study

[0124] The paste / gel of the present invention was tested using lactoperoxidase as an indicator of microbiome regulatory activity. As previously discussed, the lactoperoxidase enzyme is a protein implicated with a microbiome regulatory protein isolated from bovine milk.

[0125] Therefore, lactoperoxidase activity was measured by adding the dental paste provided in Example 3 of the table above to a hydrogen peroxide solution in 100 mM phosphate buffer (pH 5.5) and monitoring the oxidation of 2,2-azino-bis(3-ethylbenzthiazolin-6-sulfonic acid) (ABTS) at 436 nm using spectrophotometry.

[0126] The lactoperoxidase activity level was 56 U / g immediately after preparation and 75 U / g 10 days after preparation, and this Fig. 4 It is also represented in the form of a graph.

[0127] The lactoperoxidase activity of the combination of microbiome-regulating proteins formulated in the complex in the dental paste was maintained, and this level is nearly consistent with what is expected in the original complexed form, indicating that further formulation of EAPC into the new dental paste does not reduce the biological activity of the antimicrobial component.

[0128] Bioactivity - Confirmation Study

[0129] Additional tests were performed to confirm the post-formulation bioactivity of the bioactive component, namely the microbiome regulatory protein within the epithelial-adhesion protein complex, when combined with the mentioned component and provided in the final formulation of the dental paste of the present invention.

[0130] Preparation of EAPC (Epidermal-Adhesion Protein Complex)

[0131] Two samples of EAPC were prepared using two batches of microbiome regulatory proteins having superior or low activity to compare the expected bioactivity of the two pastes:

[0132] 1. 50mg of EAPC (contains 25mg Microbiome Regulating Protein Batch 18713 - excellent activity)

[0133] 2. 50mg EAPC (contains 25mg Microbiome Regulating Protein Batch 18317 - Very Low Active)

[0134] Preparation of comparative paste

[0135] A formulation to exemplify the functional characteristics of the composition was provided by the placebo composition below.

[0136] Pastes (3g) were prepared from each comparative bioactive form using a placebo formulation according to the details of the table below, namely: test dental paste (a placebo paste containing 50mg EAPC 1 containing 25mg of active microbiome regulatory protein batch 18713), active microbiome regulatory paste (a placebo paste containing only 25mg of microbiome regulatory protein batch 18713 and without EAPC), and low-activity microbiome regulatory protein paste (a placebo paste containing 50mg EAPC 2 containing 25mg / ml of low-activity microbiome regulatory protein batch 18317).

[0137] Paste formulation table

[0138]

[0139] Where applicable, the paste components of the formulation were within the previously specified operating range. For the test dental paste, the components followed the various embodiments of the present invention described above.

[0140] The water component was included solely for the consistency of the formulation, particularly to prevent heating of glycerol, ensuring that there was no potential denaturation of bioactivity during the formulation and testing process.

[0141] In vitro comparative activity of the paste ( in vitro ) check

[0142] Subsequently, the initial indicator that activity can be maintained in the formulated dental paste was additional in vitro ( in vitro ) was confirmed by testing.

[0143] To confirm this activity, two related indicators were tested. Lactoperoxidase is one of the enzyme proteins that is selectively fractionated as a microbiome-regulating component of the epithelial-adhesion protein complex in the formulation. Therefore, reconfirming the activity of lactoperoxidase in the final formulated paste will indicate the presence of the activity required for the intended application. Furthermore, the ability to kill pathogenic bacteria is generally cultured bacteria Bacillus subtilis It can be determined from the antimicrobial activity expressed for. Therefore, dental paste B. subtilis The impact on growth was also tested.

[0144] The table below shows the lactoperoxidase activity of the three comparison pastes:

[0145]

[0146] A paste expected to be of little activity, formulated with a batch of microbiome regulatory proteins known to have low activity, was found to have very little (immaterial) lactoperoxidase activity in the formulation. The enzymatic activity of the paste using a batch of active microbiome regulatory proteins provided an active paste as expected. The test dental paste was much greater than the low-activity paste, but some activity was lost when compared to an active paste containing only the same batch of microbiome regulatory proteins. The decrease in lactoperoxidase activity may have occurred during the EAPC manufacturing process.

[0147] Next, for 12 hours and 40 minutes (760 minutes) under laboratory conditions B. subtilis The effects of three pastes on the growth of bacteria were determined. The dosage of each paste delivered an equal amount (0.08 mg / ml) of microbiome-regulating protein components to the bacterial culture solution samples.

[0148] If there is no paste B. subtilis The growth rate (not shown) was determined to be similar to the growth rate in the case of low-activity paste, and Fig. 5 As shown in [figure], it grew to a maximum OD of approximately 0.25 after 760 minutes. As expected, the paste containing only active microbiome regulatory proteins was compared to the low-activity paste. B. subtilis It suppressed the growth rate of more.

[0149] With respect to the test paste, in vitro (shown in previous comparable lactoperoxidase activity) in vitro Based on ) activity B. subtilis It can be expected that the inhibition will be slightly reduced. However, the dental paste of the present invention during the irradiation period B. subtilis Growth was completely inhibited. This suggests that when a biological challenge is presented, the activity of a bioactive component at a comparable concentration can be enhanced when provided in the form according to the present invention.

[0150] B. subtilis To establish the concentration of microbiome regulatory proteins required to completely inhibit growth, three pastes were progressively diluted in a bacterial culture solution, and bacterial growth was measured after 500 minutes, at which point the bacteria were still in a state of rapid growth.

[0151] The result is Fig. 6 This is visually represented in the graph, and the minimum inhibitory concentration (MIC) of the paste is as follows:

[0152]

[0153] As expected from previous data, microbiome regulatory proteins from 0.15 mg / ml B. subtilisIt was effective in rapidly reducing the growth rate, but it did not completely inhibit growth until the concentration reached 0.93 mg / ml. The low-activity paste required a much higher concentration to inhibit growth and failed to inhibit relative growth to less than 70% (i.e., did not inhibit it completely). However, the test dental paste, despite being prepared in the same amount and batch (18713) as the paste containing only active microbiome regulatory proteins, was more effective than either of the two other pastes B. subtilis It was much more effective in preventing the growth of.

[0154] In conclusion, the tested dental paste appears to exhibit a significantly higher level of antimicrobial activity than expected from its microbiome-regulating protein content.

[0155] The applicant proposes that the positive interaction between the formulated components in the composition of the present invention yields an effect that goes beyond a mere additive effect. When the necessary components are complexed and formulated according to the composition and embodiments of the present invention described herein, a synergistic effect occurs in which the bioactivity is enhanced compared to what is expected in a relative composition containing an equivalent amount of bioactivity.

[0156] Dental paste test

[0157] After it was confirmed that useful bioactive effects could be maintained and enhanced by the composition of the embodiment of the present invention, evidence was required to clarify that the bioactivity could lead to actual veterinary applications. Specifically, an inactive placebo paste and Two major functional components within the biocomplex (EAPC)A new active formulation containing a combination of [the components] was compared. Test dental pastes were prepared according to the previously presented table, containing the components of glycerol, thickener(s), and EAPC 1, and having the described working tolerance and working range. The placebo paste was the same as the one without EAPC.

[0158] By providing the test dental paste and relative comparison placebo paste described herein in a designed 14-day trial, the actual use of the dental paste in vivo ( in vivo We confirmed whether it brings benefits to the microbiome of canids in ).

[0159] Animal Sample Pool and Tooth Condition:

[0160] A total of 20 dogs were recruited for the experiment and divided into a test paste group and a placebo paste group. The dogs varied in age and breed.

[0161] The dog had to have a significant dental condition at the start of the test according to the following grading agency:

[0162] Wisconsin Department of Agriculture, Trade and Consumer Protection and Division of Animal - Canine Tooth Condition Grading Criteria: https: / / datcp.wi.gov / Documents2 / CanineDentalGradeChart.pdf .

[0163] To be included in the test, all dogs were required to have a dental condition of at least Grade 1 but less than Grade 3 as follows:

[0164] Minor / Grade 1

[0165] Soft plaque has accumulated on the surface of the teeth. A small amount of tartar is also present. Tartar is formed when mineral deposits from saliva are deposited into the plaque. Bacteria grow and multiply, invading the gum tissue surrounding the teeth and causing early gingivitis and red, inflamed gums.

[0166] Moderate / Grade 2

[0167] Moderate plaque buildup is present. This acts as a wedge, pushing the gums away from the teeth, allowing food, bacteria, and debris to accumulate there. The gums are red and swollen, painful to the touch, and bleeding. At this stage, proper dental care is crucial to reverse the progression of the dental disease.

[0168] Major / Grade 3

[0169] Plaque and tartar formation are severe. Pockets of bacteria and debris have accumulated, forming additional pockets of infection along the teeth. The infection has destroyed the tissues surrounding the teeth that hold them in place, and some have become loose. The gums appear bright red and bleed freely.

[0170] Severe / Grade 4

[0171] Severe tartar formation and gum disease are present. Toxic debris and inflammation have caused extensive tissue necrosis. The roots have become infected, abscessed, and decayed. The thin walls of the bone surrounding the teeth have deteriorated, and many teeth have become loose. Bacteria have spread throughout the body via the bloodstream, potentially harming vital organs. Pain can also be severe at this stage.

[0172] Test Protocol

[0173] Participants received one tube of dental paste (test dental paste or placebo paste) and two oral swab collection kits. All participants were instructed to follow the test protocol:

[0174] Day 1

[0175] Baseline oral swab collection: Roll the tip of the swab along the upper right gum line and wipe for 15 seconds. Place the swab in the collection tube and prepare the sample for transport according to the instructions provided with the swab kit. Label the swab with the dog's name, the date, and a number (1) indicating that it is the first (baseline) collection swab.

[0176] Day 2 - Day 13

[0177] Apply the paste to the dog's mouth once a day between meals (i.e., at least one hour before or after feeding the dog). Using your finger, apply 3 ml of the paste to the upper arch of the gum line inside the dog's mouth (i.e., the tooth / gum boundary).

[0178] Day 14

[0179] For the final application, apply the paste to the dog's mouth. Do this at least one hour after the dog has eaten anything. Do not feed the dog between the final paste application and the second swab collection.

[0180] Collect a second (final) swab 30-45 minutes after applying the final paste: roll the tip of the swab along the upper right gum line and wipe for 15 seconds. Place the swab in a collection tube and prepare the sample for transport according to the standard instructions provided with the swab kit. Label the swab with the dog's name, the date, and a number (2) indicating that it is the second (final) collection swab.

[0181] Results and Discussion

[0182] At completion, 16 dogs successfully completed the test, and swabs were sent to the laboratory for long-read 16S mRNA sequencing for analysis.

[0183] There were 4 dogs in the placebo paste group and 12 dogs in the test paste group.

[0184] The abundance of bacteria in the oral cavity of dogs was determined by long-term 16S mRNA sequencing of material from swabs taken from the oral cavities of dogs in the treatment group (administered with a test paste prepared according to an embodiment of the present invention containing the EAPC (epithelial-adhesion protein complex) described above) and dogs in the placebo paste group (paste not containing EAPC). Bacterial species were determined from the 16S mRNA sequences.

[0185] Key pathogenic species typically associated with oral health in canids have been identified in the literature. For example, the species is [mentioned in] "Assessment of changes in the oral microbiome that occurs in dogs with periodontal disease (2021) Santibanez R et al . Veterinary Sciences , 8, 291.", "The bacteriome of the oral cavity in healthy dogs and dogs with periodontal disease (2021). Niemiec B et al . American Journal of Veterinary Science It is described in ", 83, 1." and "Molecular identification of bacteria associated with canine periodontal disease (2011). Riggio MP" et al . Veterinary Microbiology , 150, 3-4.", ( Porphyromanas gulae, Pasteurella dagmatis, Actinomyces sp., Tannerella forsythia, Treponema denticola and Corynebacterium spWe measured and compared and identified any statistically significant differences in the levels of these six pathogenic bacteria in the microbiome of canids after treatment with the test paste and comparison with the placebo paste group.

[0186] Fig. 7 As illustrated by direct comparison, the abundance of six pathogenic bacterial species associated with dental disease in canids was significantly reduced using a test paste containing EAPC. As shown in the graph, Porphyromanas gulae , Pasteurella dagmatis , Actinomyces sp. , Tannerella forsythia , Treponema denticola and Corynebacterium sp. All were reduced in the test group using the test dental paste of the present invention containing EAPC.

[0187] In contrast, in the placebo paste group, the abundance of four of these species increased, and the abundance of two of these species decreased, but to a lesser extent than in the group administered the test paste.

[0188] Fig. 8 As clearly shown in [the text], when the microbiome of canids was treated with the test paste of the present invention, the total abundance of six key pathogenic bacterial species decreased by 6.63%, whereas the placebo paste did not have a statistically significant effect on the total abundance of the same species.

[0189] This strongly suggests that the test dental paste or gel according to the embodiment described above has the potential to be useful as a veterinary medicine. Additionally, by applying it daily, the paste can be included in an effective treatment protocol.

[0190] Therefore, such dental paste has the potential to help prevent oral diseases mediated by an excess of pathogenic bacteria in mammals and / or animals, particularly canids. The dental paste of the present invention may be particularly useful when the oral disease involves one or more of dental caries, gingivitis, calculus, periodontal disease, and bad breath.

[0191] In addition, chewing articles (chews) or animal toys that are at least partially coated with the dental paste or gel described above, or contain the dental paste or gel described above, may be beneficial for maintaining the oral health of animals.

[0192] Although specific representative embodiments and details have been presented to describe the invention, it will be apparent to those skilled in the art that various modifications may be made without departing from the scope of the disclosure, as further explained in the claims appended below.

[0193] It may also be said that the embodiments described above may be broadly composed of parts, components, and features mentioned or indicated in the application specification, individually or collectively, and any or all combinations of any two or more of said parts, components, or features.

Claims

Claim 1 A dental paste or gel composition comprising at least 2 to 4% w / w of a thickener, optionally agar; at least 50 to 95% w / w of glycerol; and at least 0.03 to 3% w / w of an epithelial-adherent protein complex. Claim 2 A composition according to claim 1, wherein the epithelial-adhesion protein complex comprises a microbiome regulatory protein and a modified whey protein isolate from milk. Claim 3 In paragraph 2, the microbiome regulatory protein(s) are selected from lactoperoxidase, lactoferrin, lysomaal alpha-mannosidase, immunoglobulin G, angiogenin, ribonuclease 4 or quesine sulfhydryl oxidase and / or combinations thereof, and / or the modified WPI is formed from soluble native protein, protein aggregates and protein fibrils produced from whey protein isolate, a composition. Claim 4 A composition according to any one of claims 1 to 3, wherein the thickener, optionally agar, is in a range of at least 2 to 3% w / w, preferably 2.6 to 3% w / w. Claim 5 A composition according to any one of claims 1 to 4, wherein the epithelial-adhesion protein complex component of the composition is at least 0.03%, 0.5%, 2.5%, 2%, or 1% w / w and / or 3% or less. Claim 6 A composition according to any one of claims 1 to 5, wherein the glycerol is in the range of 80 to 95% w / w, optionally in the range of 90 to 95% w / w. Claim 7 A composition according to any one of claims 1 to 6, wherein the composition further comprises, up to 45% w / w, optionally up to 32% w / w, one or more excipients selected from arable gum, xanthan gum, alginate, chitin, aloe extract, guar gum, hydroxypropyl cellulose, sodium casein, corn starch, carboxymethylcellulose, gelatin, agar, dextrin, methylcellulose, polyvinyl alcohol, microfiber cellulose, microcrystalline cellulose, seaweed cellulose, sodium polyacrylate, sodium polyphosphate, carrageenan, or yeast cell wall. Claim 8 A composition for use as a pharmaceutical product, optionally for use as a veterinary pharmaceutical product, in any one of paragraphs 1 to 7. Claim 9 In claim 8, the composition, wherein use as a veterinary medicine comprises daily topical application of the composition to the oral mucosa. Claim 10 A composition according to claim 8 or 9 for use in the treatment or prevention of oral diseases of mammals and / or animals, preferably canines, mediated by an excess of pathogenic bacteria. Claim 11 A composition according to claim 10, wherein the above oral disease comprises one or more of the following conditions: dental caries, gingivitis, tartar, periodontal disease, and bad breath. Claim 12 A food supplement comprising a composition according to any one of claims 1 to 11 used for maintaining the health of canids or felines. Claim 13 A chew article or animal toy comprising a composition according to any one of claims 1 to 12, wherein the chew article or animal toy is optionally coated at least partially with said composition. Claim 14 A method for preventing or treating oral or dental diseases and / or improving or maintaining oral health, comprising administering a composition according to any one of claims 1 to 7 to a mammal or animal in need thereof, preferably a canine. Claim 15 A method for preparing a dental paste or gel composition, comprising: a step of providing glycerol in an amount of at least 50 to 95% w / w and optionally heating, preferably to a maximum of 95°C; a step of adding and dissolving a thickener in an amount of 2 to 4% w / w, optionally agar, to the glycerol; Optionally, a step of adding one or more excipients of 0 to 45% w / w to the glycerol and thickener combination; a step of cooling the combination, optionally to 40°C or lower; a step of adding and dissolving an epithelial-adhesion protein complex of 0.03 to 3% w / w, wherein optionally the epithelial-adhesion protein complex comprises a microbiome regulatory protein or "protein fraction" and a modified whey protein isolate (WPI) from milk, and optionally the microbiome regulatory protein(s) are selected from lactoperoxidase, lactoferrin, lysomaal alpha-mannosidase, immunoglobulin G, angiogenin, ribonuclease 4 or quesine sulfhydryl oxidase and / or combinations thereof, and / or the modified WPI is formed from soluble native proteins, protein aggregates and protein fibrils generated from the whey protein isolate; A method comprising the step of cooling to form a dental paste or gel composition, optionally cooling to 18°C ​​or lower.

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