AU721548B3 - Protective peptide formulation - Google Patents

Description

S&F Ref: 492892

AUSTRALIA

PATENTS ACT 1990 PETTY PATENT SPECIFICATION

ORIGINAL

Name and Address of Applicant: Actual Inventor(s): Address for Service: Invention Title: Pacific Biolink Pty Limited 7/42 Leighton Place Hornsby NSW 2077 Dennis James Bannister Spruson Ferguson St Martins Tower 31 Market Street Sydney NSW 2000 Protective Peptide Formulation The following statement is a full description of this invention, including the best method of performing it known to me/us:- 5843c Protective Peptide Formulation Technical Field The invention relates to formulations designed to provide protection to biological interfaces, and to deliver bioactive constituents to biological surfaces.

Background Art A number of peptides, isolated from bovine milk, have been shown to be advantageous in dental health. These peptides have been available for a number of years, and have been shown to have beneficial properties when complexed with calcium and phosphate. Notable advantageous properties of these peptides include the remineralisation of enamel and dentine in human and animal io dentition. A suitable example of such peptides is illustrated in US 5 015 628. These peptides may be prepared using the method described in AU 9216468.

However, it has been shown in recent times that the present formulations in which these peptides reside also suffer from a number of significant disadvantages that preclude application.

These disadvantages include: present peptide formulations suffer from a limited shelf life at room temperature before the occurrence of odorous bacterial breakdown and/or the appearance of a precipitate of peptide resulting from the addition of incompatible components, and the viscosity of present formulations is highly variable.

The present invention provides a stabilised protective peptide formulation having an extended shelf life, and a formulation which produces a reliable gel, without the loss of activity of the protective peptide and other bioactive constituents, whilst maintaining the activity of the peptide and associated bioactive constituents.

Object of the Invention An object of the invention is to provide an improved protective peptide formulation having an ease of application, and providing a delivery system for bioactive substances.

Disclosure of the Invention According to a first embodiment of the invention, there is provided a protective peptide formulation including a suspension or solution of a peptide in water, and a suitable amount of a thickening agent to form a thixotropic gel, wherein said phosphopeptide is selected from the group consisting of: T1. Gly-Met-Glu-Ala-Glu-Pse-lle-Pse-Pse-Pse-Glu-Glu-lle-Val-Pro-Asn-Pse-Val-Glu-Gln-Lys, T2. Glu-Leu-Glu-Glu-Leu-Asn-Val-Pro-Gly-Glu- le-Val-Glu-Pse-Leu-Pse-Pse-Pse-Glu-Glu- Ser-lle-Thr-Arg, T3. Asn-Thr-Met-Glu-His-Val-Pse-Pse-Pse-Glu-Glu-Ser-lle-lle-Pse-Gln-Glu-Thr-Tyr-Lys, Cf~ 0 L I :\DayLib\LiBH]285808doc:BAV I a 14. Asn-Ala-Asn-Glu-Glu-Glu-Tyr-Ser-Ile-Gly-Pse-Pse-Pse-Glu-Glu-Pse-Ala-Glu-Val-Ala-Thr-Glu- Glu-Val-Lys, and Glu-Gln-Leu-Pse-Pth-Pse-Glu-Glu-Asn-Ser-Lys.

Typically, the peptide may comprise a phosphopeptide as disclosed in US 5 015 628, the sdisclosure of which is incorporated herein by reference.

US 5 015 628 described peptides may include: a phosphopeptide or a salt thereof, the phosphopeptide having from 5 to 40 amino acids including the sequence: A-B-C-D-E Where A, B, C, D and E are independently phosphoserine, phosphothreonine, phosphotyrosine, phosphohistidine, glutamate and aspartate.

ID Typical phosphopeptides are those wherein A, B and C are independently phosphoserine, phosphothreonine, phosphotyrosine and phosphohistidien and D and E are independently phosphoserine, phosphothreonine, glutamate and aspartate.

More typically, the peptide may be chosen from the group consisting of: 11. Gly-Met-G Iu -Ala-G u-Pse- Ile-Pse-Pse-Pse-G lu-G lu- le-Val-Pro-Asn-Pse-Val-G Iu-GIn -Lys 1 2. Glu-Leu-Glu-Glu-Leu-Asn-Val-Pro-Gly-Glu-le-Val-Glu-Pse-Leu-Pse-Pse-Pse-Glu-Glu-Ser-le- Thr-Arg, I I\ayi\1_1311I128580.doc:BAV T3. Asn-Thr-Met-Glu-His-Val-Pse-Pse-Pse-Glu-Glu-Ser-Ile-Ile-Pse-Gln-Glu- Thr-Tyr-Lys, T4. Asn-Ala-Asn-Glu-Glu-Glu-Tyr-Ser-Ile-Gly-Pse-Pse-Pse-Glu-Glu-Pse- Ala-Glu-Val-Ala-Thr-Glu-Glu-Val-Lys, and T5. Glu-Gln-Leu-Pse-Pth-Pse-Glu-Glu-Asn-Ser-Lys, Wherein, the amino acid symbols are as follows: Pse-phosphoserine, Ser-serine, Pthphosphothreonine, Thr-threonine, Glu-glutamate, Asp-aspartate, Ala-alanine, Asnasparagine, Gln-glutamine, Gly-glycine, Arg-arginine, His-histidine, le-isoleucine, Lyslysine, Met-methionine, Pro-proline, Tyr-tyrosine and Val-valine.

Typically, the peptide may be obtained from bovine, ovine or caprine milk.

Note that unless otherwise stated, all percentages of components of the protective peptide formulation are by weight, based on the total weight of the protective peptide formulation.

Typically, the amount of peptide present in the formulation is between 0.01 and More typically, the amount of peptide present in the formulation is between and Even more typically, the amount of peptide present in the formulation is between 1 and The thickening agent is selected on the basis that it does not deactivate the active peptide component of the protective peptide formulation. Essentially, thickening agents of the present invention do not cause flocculation or precipitation of the peptide or any other active constituents within the protective peptide formulation.

Typically, the thickening agent is selected from the group consisting of: a a synthetic or natural polymer, or a combination of any of these.

Even more typically, the thickening agent comprises a clay chosen from the group consisting of: synthetic hectorite including laponite D (and all other types of laponite such as DF, CP,SP 2002), calcium montmorillonite, sodium montmorillonite (bentonite), sodium exchanged montmorillonite, acid activated bleaching earth and palygorskite.

The thickening agent may be selected from the natural polymers: alginate, cellulose and cellulose derivatives.

The thickening agent may be synthetic polymer: carboxymethylcellulose.

Other suitable thickening agents include irish moss, gum tragacanth, starch, polyvinylpyrrolidone, hydroxyethylpropylcellulose, hydroxybutylmethylcellulose, hydroxypropylmethylcellulose, hydroxyethylcellulose (eg. available as Natrasol), sodium carboxymethylcellulose, and colloidal silica such as finely ground Syloid.

Typically, the amount of thickening agent present in the formulation is between 0.05 and More typically, the amount of thickening agent present in the formulation is between 0.1 and Even more typically, the amount of thickening agent present in the formulation is between 0.5 and 6%.

The protective peptide formulation may also be subject to bacterial contamination and odorous breakdown. Therefore, the protective peptide formulation may also further include at least one anti-microbial agent, wherein any anti-microbial agent used in commerce is also suitable for use in the protective peptide formulation.

Typically, the anti-microbial agent may include an organic anti-microbial agent, wherein the organic anti-microbial agent is typically water-soluble.

More typically, the anti-microbial agent may include organic anti-microbial agents as disclosed in US 5 368 844, the disclosure of which is incorporated herein by reference.

As described in US 5 368 844 the anti-microbial agent may include: halogenated diphenyl ethers, such as: 2',4,4'-trichloro-2-hydroxy-diphenyl ether (Triclosan); phenolic compounds, including phenol and its homologues, such as: 2-methyl-phenol, 3-methylphenol, 4-methyl-phenol, 4-ethyl-phenol, 2,4-dimethyl-phenol, 3,4-dimethyl-phenol, 2,6dimethyl-phenol, 2,2'-methylene bis (4-chloro-6-bromo-phenol); mono- and poly-alkyl and aromatic halophenols, including p chlorophenols such as: methyl-p-chlorophenol, ethyl-p-chlorophenol, n-propyl-p-chlorophenol, n-butyl-chlorophenol; -o-chlorophenols; p-bromophenols; -o-bromophenols; resorcinol and its derivatives, such as: n-methyl hexyl resorcinol; bisphenolic compounds and halogenated carbanilides.

Still more typically, the anti-microbial agent may be selected from the group consisting of: ethanol, sorbitol, mannitol, sodium benzoate, methy-p-hydroxybenzoate, ethyl-p-hydroxy benzoate, N-propyl p-hydroxybenzoate, butyl-p-hydroxybenzoate, phenoxyethanol and quaternary ammonium salts, such as benzethonium chloride, and dissobutyl-phenoxyethoxyethyl dimethyl benzyl ammonium chloride.

Other types of anti-microbial agents may include amidines, such as substituted guanidine, including, chlorhexidine, and other known bis-biguanidines; and cationic tertiary amines.

Typically, the amount of sodium benzoate present in the formulation is between 0.0001 and More typically, the amount of sodium benzoate present in the formulation is between 0.005 and 0.3%.

Even more typically, the amount of sodium benzoate present in the formulation is between 0.0025 and 0.2%.

Yet even more typically, the amount of sodium benzoate present in the formulation is between 0.001 and 0.1%.

Typically, the amount of ethanol present in the formulation is between 0.05 and More typically, the amount of ethanol present in the formulation is between and Even more typically, the amount of ethanol present in the formulation is between 1 and 8%.

Yet even more typically, the amount of ethanol present in the formulation is between 2 and 6%.

Typical oral formulations of the present invention are in the form of toothpaste creams or gels, or mouthwashes. Suitable ingredients which may typically be included in the toothpaste and gels in accordance with the invention include: abrasive polishing materials, sudsing agents, flavouring agents, humectants, binders, sweetening agents, and water. Abrasives which may be used in the formulations of the invention include alumina and hydrates thereof, such as amorphous silica, alpha alumina trihydrate, magnesium trisilicate, dicalcium phosphate, magnesium carbonate, aluminosilicate, such as calcined aluminium silicate and aluminium silicate, calcium carbonate, zirconium silicate, poly(methyl methacrylate), powdered poly(ethylene), poly(propylene), silica xerogels, hydrogels and aerogels and the like. Also suitable as abrasive agents are calcium pyrophosphate, insoluble sodium metaphosphate, dicalcium orthophosphate, particulate hydroxy apatite and the like. Depending on the form which the oral formulation is to take, the abrasive may be present in an amount of from 0 to 70% by weight, typically 1 to 70% by weight, more typically from 10 to 70% by weight, particularly for toothpastes.

Humectants contemplated for use in the formulations of the present invention include: glycerol, polyol, sorbitol, polyethylene glycols, propylene glycol, hydrogenated partially hydrolysed polysaccharides and the like. The humectants are generally present in amounts of from 0 to 80%, typically 5 to 70% by weight, particularly for toothpastes.

Various other materials may be incorporated in the oral formulations of the present invention, such as whitening agents, preservatives, silicones, chlorophyll compounds, other anticalculus agents, and/or ammoniated material such as urea, diammonium phosphate, and mixtures thereof. These adjuvants, where present, are incorporated in the preparations in amounts which do not substantially adversely affect the properties and characteristics desired.

Any suitable favouring or sweetening material may also be employed. Examples of suitable flavouring constituents are flavouring oils. For example, oil of spearmint, peppermint, wintergreen, sassafras, clove, sage, eucalyptus, marjoram, cinnamon, lemon, and orange, and methyl salicylate. Suitable sweetening agents include sucrose, lactose, maltose, dextrose, laevulose, sorbitol, xylitol, d-tryptophan, dihydrochalcones, sodium cyclamate, perillartine, APM (aspartylphenylalanine, methyl ester), saccharine and the like. Suitably, flavouring and sweetening agents may together comprise from about 0.1% to 10% by weight or more of the preparation, and more typically, from about 0.1% to by weight or more of the preparation.

Typically, an oral formulation according to the present invention such as a dentifrice is preferably applied by brushing regularly to dental enamel, such as every second or third day or preferably from 1 to 3 times daily, at a pH of about 4.5 to generally about 5.5 to 9, preferably about 6 to 8 for at least 2 weeks up to 8 weeks or more up to lifetime. The dentifrice is typically removed by rinsing with water after each application.

According to a second embodiment of the invention there is provided a protective peptide formulation in accordance with the first embodiment of the invention, wherein the peptide formulation associates with bioactive constituents, thereby providing a delivery system for the bioactive constituents.

Typically, the protective peptide formulation may optionally contain bioactive inorganic or organic constituents which bind to the peptide, yet do not affect its biological activity.

Typically, the peptide and at least one bioactive constituent are present in association within the formulation in accordance with the present invention. More typically, the association between the peptide and the bioactive constituent is by virtue of the presence of both negative and positive groups associated along the length of the peptide molecules, wherein these groups can ionically interact with a range of bioactive constituents to form a complex with the peptide or be soluble in water. Even more typically, wherein the bioactive constituents include constituents insoluble in water, the constituents may be stabilised in an emulsion or suspension within the peptide formulation of the present invention. Thus, the formulation of the present invention comprising a suspension or solution of at least one isolated and purified casein protein, together with at least one bioactive constitiuent, provides a delivery system for a range of bioactive constituents.

In particular, the phosphopeptide has an affinity for biological surfaces, and as a consequence, when the formulation in accordance with the present invention is applied to a biological surface, by virtue of this affinity of the peptide resident in the formulation, the concentration of the associated bioactive constituent at the biological surface is increased, thereby providing a delivery system wherein the bioactive constituent is presented to the biological surface.

Typically, inorganic bioactive constituents are selected from the group consisting of: calcium, phosphate, fluorophosphate, fluoride, zirconia, magnesium, barium, zinc, iron, copper, aluminium, tin, silver, and salts of said bioactive constituents, selected from the group consisting of Calcium phosphate, sodium fluoride, calcium fluoride, calcium fluorophosphate, stannous fluoride, titanium dioxide, and calcium oxide. In a typical form of the invention, the phosphopeptide acts to sequester the bioactive constituents, calcium and phosphate to form an amorphous complex, which can then be used as a soluble source of calcium and phosphate ions.

Generally, inorganic constituents of the protective peptide formulation are insoluble, and are held in a suspension within the thixotropic gel, and not necessarily through interaction with the resident peptide. However, the protective peptide formulation may also comprise soluble inorganic constituents, such as fluorides, chlorides, bromides, and iodides.

Typically, fluoride sources used in the formulation of the present invention include: sodium fluoride, stannous fluoride, sodium monofluorophosphate, zinc ammonium fluoride, tin ammonium fluoride, calcium fluoride or cobalt ammonium fluoride.

Fluoride ions are typically provided at a level of from about Oppm to 6000ppm, more typically, 0 to 3000ppm, even more typically, 0 to 1500ppm and still more typically, 10 to 1500ppm.

Typically, surfactants, such as soap, anionic, nonionic, cationic, amphoteric and/or zwitterionic surfactants, may also be present in the formulation as a bioactive constituent, or as an additional agent within the formulation. More typically, the surfactants are present within the range of 0 to 15%, even more typically 0.1 to 15%, still more typically 0.25 to 10% by weight. Anionic surfactants are most preferred, such as sodium dodecyl sulfate, sodium lauryl sarcosinate and sodium dodecylbenzene sulphonate.

Typically, the amount of inorganic bioactive constituents present in the formulation is between about 0.005 and 50%. More typically, the amount of inorganic constituents present in the formulation is between about 0.2 and about 35%. Even more typically, the amount of bioactive inorganic constituents present in the formulation is between about 0.5 and about 15%. Yet even more typically, the amount of inorganic constituents present in the formulation is between 0.5 and Typically, the protective peptide formulation may optionally contain organic constituents which bind to the peptide, yet do not affect its biological activity. Organic constituents may safeguard the protective peptide formulation against ultra-violet or visual light radiolytic degredation or may even be applied to the skin for the prevention of sunburn.

Typically, the organic constituents are selected from the group consisting of octyl methoxycinnamate, butyl methoxydibenzoylmethane and other commercially available ultra-violet or visual light absorbing compounds.

Typically, the amount of organic constituents present in the formulation is between 0.05 and 30%. More typically, the amount of organic constituents present in the formulation is between 1 and 20%. Even more typically, the amount of organic constituents present in the formulation is between 1 and 15%. Still more typically, the amount of organic constituents present in the formulation is between 1 and 6%.

According to a third embodiment of the invention, there is provided a protective peptide formulation including a suspension or a solution of a peptide in water, together with a suitable amount of an anti-microbial agent to form a mouthwash.

Typically, the peptide includes those peptides defined in relation to the first embodiment of the invention.

Typically, the anti-microbial agent includes those agents defined above.

Typically, mouthwashes comprise a water/alcohol solution, flavour, humectant, sweetener, sudsing agent, and colorant. The corresponding compounds mentioned above which are used in toothpastes, are generally suitable within the ranges above for mouthwashes as well. The mouthwash can include ethanol at a level of from 0 to preferably from 4 to 30% by weight.

According to a fourth embodiment of the invention, there is provided a method of treating or preventing dental caries in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention, wherein said formulation is capable of controlling or preventing dental caries in humans or animals.

Typically, treatment or prevention of dental caries involves the release of bioactive constituents from the peptide formulation.

More typically, the bioactive constituents of the peptide formulation in accordance with the fourth embodiment of the invention include: polyphosphates, generally employed in the form of their wholly or partially dehydrated polyphosphate water-soluble alkali metal, or ammonium salt, such as, tetrasodium pyrophosphate; Nmethylpyrrolidone or pyrrolidone-5,5-diphosphonic acids, such as, 2-pyrrolidone-5,5diethyl phosphonic acid.

Typically, the amount ofbioactive constituents present in accordance with the fourth embodiment of the invention is between 0.01 to 40% of the peptide formulation.

More typically, between 0.01 to 25% of the peptide formulation.

According to a fifth embodiment of the invention, there is provided a method of treating or preventing dental sensitivity in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention, wherein said formulation is capable of controlling or preventing dental sensitivity in humans or animals.

Typically, treatment or prevention of dental sensitivity involves release of bioactive constituents from the peptide formulation.

More typically, the bioactive constituents of the peptide formulation in accordance with the fifth embodiment of the invention include: glycerine, strontium chloride, sodium citrate, potassium nitrate or dicalcium phosphate.

Typically, the amount of bioactive constituents present in accordance with the fifth embodiment of the invention is between 0.001 to 60% of the peptide formulation.

More typically, between 0.01 to 50% of the peptide formulation.

According to a sixth embodiment of the invention, there is provided a method of treating or preventing gingivitis in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention, wherein said formulation is capable of controlling or preventing gingivitis in humans or animals.

According to a seventh embodiment of the invention, there is provided a method of treating or preventing mouth odour in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention, wherein said formulation is capable of controlling or preventing mouth odour in humans or animals.

Typically, treatment or prevention of gingivitis and/or mouth odour involves release of bioactive constituents from the peptide formulation.

More typically, the bioactive constituents of the peptide formulation in accordance with the sixth or seventh embodiments of the invention include an antimicrobial agent.

Typically, the peptide acts to increase the concentration of anti-microbial agent at biological surfaces.

Typically, the anti-microbial agent includes those anti-microbial agents described above in relation to the first embodiment of the invention.

Typically, the anti-microbial agent may include an organic anti-microbial agent, wherein the organic anti-microbial agent is typically water-soluble. More typically, the anti-microbial agent may include organic anti-microbial agents as disclosed in US 5 368 844, the disclosure of which is incorporated herein by reference. Even more typically, the anti-microbial agent may be selected from the group consisting of: ethanol, sorbitol, mannitol, sodium benzoate, methyl-p-hydroxybenzoate, methyl-p-hydroxybenzoate, Npropyl p-hydroxybenzoate, butyl-p-hydroxybenzoate, phenoxy ethanol and quaternary ammonium salts.

Typically, the amount of sodium benzoate present in the formulation is between 0.0001 and More typically, the amount of sodium benzoate present in the formulation is between 0.005 and Even more typically, the amount of sodium benzoate in the formulation is between 0.0025 and Yet even more typically, the amount of sodium benzoate present in the f6rmulation is between 0.001 and 0.1%.

Typically, the amount of ethanol present in the formulation is between 0.05 and 20%. More typically, the amount of ethanol present in the formulation is between and 10%. Even more typically, the amount of ethanol present in the formulation is between 1 and Yet even more typically, the amount of ethanol present in the formulation is between 2 and 6%.

According to an eighth embodiment of the invention, there is provided a method of recrystallising and remineralising enamel and/or dentine in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first second or third embodiments of the invention, wherein said formulation is capable of recrystallising and remineralising enamel and/or dentine in humans or animals.

Typically, recrystallising and remineralising enamel and/or dentine involves release of bioactive constituents from the peptide formulation.

More typically, the bioactive constituents of the peptide formulation include; fluoride, a calcium phosphate complex or a calcium fluorophosphate complex, wherein said calcium phosphate/fluorophosphate complex provides a soluble (bioavailable) source of calcium and/or phosphate and/or fluorophosphate.

Typically, the amount of fluoride present in the formulation is in the range of lppm to 6000ppm.

Typically, the amount of soluble calcium or phosphate so provided is in the range of 0.01mg/ml to 25mg/ml. More typically, the amount of soluble calcium or phosphate is in the range 0.1mg/ml to According to a ninth embodiment of the invention, there is provided a method of buffering plaque against a decrease in pH in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second and third embodiments of the invention, wherein said formulation is capable of buffering plaque against a fall in pH in humans or animals.

Typically, the peptide within the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention will provide a buffering action in relation to the plaque.

Typically, the buffering action in relation to the plaque involves release of bioactive constituents from the peptide formulation.

More typically, the peptide itself acts as a bioactive component in this regard, acting to buffer plaque against a reduction in pH.

According to a tenth embodiment of the invention, there is provided a method of treating or preventing osteoporosis in humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention, wherein said formulation is capable of controlling or preventing osteoporosis in humans or animals.

Typically, the method of treating or preventing osteoporosis in humans or animals may involve delivery of the bioactive constituents of the peptide formulation of the second embodiment of the invention to the human or animal undergoing said treatment or prevention.

More typically, the bioactive constituents of the peptide formulation include: Vitamin D, or a calcium phosphate and/or calcium fluorophosphate complex, wherein said calcium phosphate complex and/or calcium fluorophosphate complex provides a soluble (bioavailable) source of calcium and phosphate and/or fluorophosphate.

Typically, the amount of soluble calcium provided is in the range of 0.1mg/ml to Typically, the amount of Vitamin D provided is in the range 0.1mg/ml to 100mg/ml.

According to an eleventh embodiment of the invention, there is provided a method of treating or preventing calculus formation in the oral cavity of humans or animals, said method includes administering an effective amount of the protective peptide formulation in accordance with any one of the first, second or third embodiments of the invention, wherein said formulation is capable of controlling or preventing calculus formation in the oral cavity of humans or animals.

Typically, treatment or prevention of calculus formation involves release of bioactive constituents from the peptide formulation.

More typically, the bioactive constituents of the peptide formulation in accordance with the eleventh embodiment of the invention include the phosphopeptide itself Even more typically, the bioactive constituents of the peptide formulation in accordance with the eleventh embodiment of the invention include polyphosphates, such as tetrasodium pyrophosphate, tetrapotassium pyrophosphate, and mixtures thereof Typically, the amount of phosphopeptide present in accordance with the eleventh embodiment of the invention is between 0.01 to 50% of the peptide formulation. More typically, the amount ofphosphopeptide present in the formulation is between 0.5 and 25%. Even more typically, the amount of phosphopeptide present in the formulation is between 1 and Typically, the amount of polyphosphate present in accordance with the eleventh embodiment of the invention is between 0.01 to 50% of the peptide formulation. More typically, the amount of phosphopeptide present in the formulation is between 0.5 and 25%. Even more typically, the amount ofpolyphosphate present in the formulation is between 1 to According to a twelfth embodiment of the invention, there is provided a glass ionomer cement, wherein said cement comprises the formulation in accordance with the first embodiment of the invention, together with liquid and powdered precursors of said glass ionomer cement capable of reacting to form said glass ionomer cement.

Typically, the glass ionomer cement is obtainable by curing a composition comprising a mixture of a liquid precursor of a glass ionomer cement, comprising polymerisable monomers, a carboxylic acid polymer, a solvent, a free radical initiator; and a powdered precursor of a glass ionomer cement, wherein said liquid precursor of a glass ionomer cement, said powdered precursor of a glass ionomer cement are in a ratio typically from between about 2.5:1 to about 1:1 by weight, and wherein said curing is achieved by a free radical polymerisation reaction.

The polymerisable acid or non-acidic monomer may be present in an amount up to 90% by weight based on the total of the liquid precursor of the glass ionomer cement.

Typically, the polymerisable acid or non-acidic monomer is present in a range of between about 2 to about 50% by weight, more typically in a range between about 2 to about about 2 to about 40%, about 2 to about 35%, about 5 to about 35%, about 8 to about about 10 to about 35%, or about 12 to about 35% by weight, and even more typically in a range between about 15 to about 35% by weight.

Typically, the carboxylic acid polymer may be present in an amount up to about by weight based on the total of the liquid precursor of the glass ionomer cement.

Typically, the carboxylic acid polymer is present in a range of between about 5 to about by weight, more typically in a range between about 5 to about 45%, about 5 to about about 5 to about 25%, about 10 to about 45%, or about 10 to about 40% by weight, even more typically in a range between about 15 to 40% by weight.

Similarly, the aqueous solvent may be present in an amount up to about 80% by weight based on the total of the liquid precursor of the glass ionomer cement. Typically, the aqueous solvent is present in a range of between about 10 to about 75% by weight, and more typically in a range between about 30 to about 50% by weight.

Where the material is obtained through a free radical polymaerisation curing process, a free radical initiator may be present in an amount up to about 5% by weight based on the total of the liquid precursor of the glass ionomer cement. Typically, the free radical initiator is present in a range of between about 0.01 to about 2% by weight, and more typically in a range between about 0.1 to about 0.5% by weight.

An activator for the free radical initiator may be present in an amount up to about by weight, based on the total of the liquid precursor of the glass ionomer cement.

Typically, the activator is present in a range of between about 0.01 to about 2% by weight, and more typically in a range between about 0.1 to about 0.5% by weight.

The liquid precursor of the glass ionomer cement may be comprised of a variety of polymerisable acid or non-acidic monomers, including any acidic or non-acidic monomers that will take part in a free radical polymerisation reaction. Acid monomers are those acids that contain carbon-carbon double bonds. These include methacrylic acid, acrylic acid, itaconic acid, maleic acid, and maleic anhydride. The polymerisable nonacidic monomers may include such monomers as: 2-hydroxy ethyl methacrylate, acrylamide, methacrylamide, or tetrahydrofurfuryl methacrylate. Further, these polymerisable monomers may also be combined with acidic or neutral monomers containing more than one carbon-carbon doulble bond such as: 1,5-diallyl-2,4-benzene dicarboxylic acid, triethylene glycol dimethacrylate, triallyl-1,3,5-triazine- 2,4,6(lH,3H,5H)-trione or diallyoxy acetic acid.

The liquid precursor of the glass ionomer cement may be comprised of a variety ofcarboxylic acid polymers including any homopolymers with a single type of unit along their side chain, such as poly(acrylic acid), poly(methacrylic acid), and Poly(itaconic acid). The carboxylic acid polymers may also include any copolymers, such as poly(vinyl mehtyl ether co-maleic acid), poly(methacrylic acid co-acrylic acid), poly(styrene co-acrylic acid co-methacrylic acid). Furthermore, the polymer may have double bonds along the side chain, making the polymer capable of taking part in a free radical reaction.

The liquid precursor of the glass ionomer cement will contain an amount of an aqueous solvent. Suitable aqueous solvents include water, but may also include a mixture such as water and a water miscible liquid such as ethanol or isopropanol.

Both the liquid and powdered precursors of the glass ionomer cement may also contain a free radical initiator such as camphoquinone, azobisisobutyronitrile or riboflavin.

The liquid precursor of the glass ionomer cement may also contain a free radical inhibitor such as butylated hydroxytoluene, hydroquinone and methyl ethyl hydroquinone.

Suitable powdered precursors of the glass ionomer cement include any powder containing any amount of divalent or trivalent metal ions. Examples of these include calcium aluminium fluorosilicate glass, phosphates of zinc and calcium, oxides and hydroxides of calcium, zinc, barium, strontium and aluminium.

The powdered precursor may be comprised of a solid that will generate an acid in the presence of water or an acidic solution. Such a solid may be phosphorous pentoxide, disodium tartrate or disodium maleate.

The powdered precursor may also contain a peroxide initiator so that the powderliquid mixture will undergo a free radical polymerisation in the absence of light. Suitable examples of such a peroxide initiator include: benzoyl peroxide or methyl ethyl ketone peroxide or potassium persulphate.

When preparing the glass ionomer cement, part, or all of the poly(carboxylic acid) component of the glass ionomer cement may be added to the powdered precursor, also containing a spray-dried formulation in accordance with the present invention, so that the dry poly(carboxylic acid) polymer swells or dissolves in the liquid component, when the powdered and liquid precursors of the glass ionomer cement are mixed together.

Typicall, the free radical polymerisation reaction is light activated and is brought about by adding to the liquid formulation, a small amount of an initiator such as camphorquinone and an activator such as a tetramethyl amine. A suitable example of such an amine is N,N-3,5-tetramethyl aniline. More typically, aniline may be present in a range between 0.1% to 5% by weight based on the total weight of the liquid precursor of the glass ionomer cement. More typically, tetramethyl aniline may be present in a range of between about 0.1 to about 0.7% by weight, and even more typically in a range between about 0.2 to about 0.5% by weight.

The acid-base reaction that occurs to form a cured glass ionomer cement involves the neutralising of the acid groups in the polymer network by multivalent metal ions such as calcium ions and aluminium ions as provided by the powdered precursor. The acidbase reaction is slow, and the rate of the reaction is limited by the diffusion of metal ions out of the glass powder into the polymer network, and subsequent ionic crosslinking.

The glass ionomer cement may also contain an amount of a heavy metal that would render the material opaque to X-rays, ie radio-opaque. Examples of such heavy metals include barium, bismuth, gold, silver, tin, lead, cadmium, antimony, palladium, platinum, tungsten or iridium. The heavy metals should be in a form sufficiently bound such that undesirable heavy metals are unable to be leached in vivo.

Typically, the glass ionomer cement may be used as dental restorative material.

Typically, the amount of formulation in accordance with the first embodiment of the invention, typically in a spray-dried form, present in the glass ionomer cement is betweenabout 0.05 to about 50% of the cement. More typically, the amount of the formulation is betweenabout 0.05 and about 25%. Even more typically, the amount of formulation is between about 0.1 and about Typically, the peptide present in the formulation is a phosphopeptide-calcium phosphate complex.

Definitions By the phrase "suitable amount of thickening agent" we mean that the formulation includes a sufficient amount of a thickening agent to form a thixotropic gel, but not so much of the thickening agent that the viscosity of the formulation is so great as to render the formulation unusable.

In the context of this specification, the term "comprising" means "including principally, but not necessarily solely". Further, variations of the word "comprising" such as "comprise" and "comprises" have correspondingly varied meanings.

Best Modes and Other Modes of Carrying Out the Invention Typically, the preferred formulation of the present invention falls within the following ranges, wherein all proportions are expressed by weight: Peptide 0.01 to Laponite 0.1 to 4% Ethanol 0.05 to Water Up to 100% A further formulation of the present invention falls within the following ranges: Peptide 0.5 to Bentonite 0.1 to 8% Sodium benzoate 0.001 to 0.1% Water Up to 100% A more preferred formulation of the present invention falls within the following ranges: Peptide 0.5 to Laponite 0.1 to 4% Ethanol 0.5 to Water Up to 100% Typically, a formulation of the present invention may also contain inorganic constituents associated with the protective pe tide, within the following ranges: Peptide 1 to Laponite 0.5 to 3% Ethanol 1 to 8% Calcium phosphate 0.1 to Water Up to 100% Typically, a formulation of the present invention may also contain inorganic constituents associated with the protective peptide, within the following ranges: Peptide 1 to Laponite DF 0.5 to 3% Ethanol 1 to 8% Sodium fluoride 1ppm to 500ppm Water Up to 100% Typically, a formulation of the present invention may also contain organic constituents associated with the protective pe tide, falls within the following ranges: Peptide 1 to Laponite 0.5 to 3% Ethanol 2 to Octyl methoxycinnamate 0.1 to 0.8% Water Up to 100% Typically, a further formulation of the present invention, wherein the formulation also contains organic constituents associated with the protective peptide, falls within the following ranges: Peptide 1 to Laponite 0.5 to 3% Ethanol 2 to Butyl methoxydibenzoylmethane I 0.1 to 0.8% Water Up to 100% A specific mouthwash formulation in accordance with the present invention includes: Phosphopeptide-calcium phosphate Sodium benzoate 0.03% Sodium fluoride 0.01% Flavour 0.15% Sodium saccharine 0.08% Ethanol Water Up to 100% A specific thixotropic gel for use in a mouthguard in accordance with the present inventinn includes: Phosphopeptide-calcium phosphate Sodium fluoride 0.01% Sodium benzoate 0.03% Sodium saccharine 1.6% Flavour 3.1% Ethanol Water Up to 100% Titanium dioxide Laponite DF (synthetic clay) A specific toothpaste in accordance with the present invention includes: Phosphopeptide-calcium phosphate Sodium fluoride 0.01% Sodium benzoate 0.03% Sodium saccharine 1.2% Flavour 2.3% Ethanol 4.1% Water Up to 100% Titanium dioxide 0.75% Laponite DF Silica abrasive The invention will now be described in greater detail by reference to specific Examples, which should not be construed as limiting on the scope thereof: Examples The phosphopeptide used in some of the following examples was prepared using the method described in US 5 015 628. The casein phosphopeptide-calcium phosphate complex used in some of the following examples, was prepared by first isolating calcium phosphopeptide using the method disclosed in AU9216468, and then slowly adding an aqueous solution of disodium phosphate to a aqueous solution of the calcium phosphopeptide, followed by spray drying. Resulting in a powder with about equal weights of phosphopeptide and calcium phosphate.

Example 1 A stabilised 2.5% by weight protective peptide formulation was prepared according to the following: 1) Part A: add 2.85% by weight of a synthetic smectic clay (Laponite) to 92.15% by weight water and 5% by weight ethanol. Stir with an overhead stirrer for two hours. The clay must be fully hydrated before further processing. When the stirrer is turned off, the mixture will form a gel.

2) Part B: prepare a 20% by weight casein phosphopeptide suspension by weight water and 5% by weight ethanol and 55% by weight water.

3) Mix 12.5% by weight of Part B with 87.5% by weight of Part A to form a stable gel.

Example 2 A stabilised 2.5% by weight protective peptide formulation was prepared Saccording to the following: 1) Part A: add 2.85% by weight of a synthetic smectic clay (Laponite) to 92.15% by weight water and 5% by weight ethanol. Stir with an overhead stirrer for two hours. The clay must be fully hydrated before further processing. When the stirrer is turned off, the mixture will form a gel.

2) Part B: prepare a 40% by weight casein phosphopeptide-calcium phosphate suspension by mixing together the following: 40% by weight casein phosphopeptidecalcium phosphate complex, 5% by weight ethanol and 55% by weight water.

3) Mix 12.5% by weight of Part B with 87.5% by weight of Part A to form a stable gel.

Example 3 A stabilised 16% by weight protective peptide formulation was prepared according to the following: 1) Take 60% by weight of Part A, as described above in Example 2, and stir with an overhead stirrer.

2) Add 40% by weight of casein phosphopeptide-calcium phosphate complex, in by weight gradations, allowing approximately 20 minutes stirring time between each addition.

Example 4 A stabilised protective peptide formulation was prepared according to Example 1, wherein the formulation also comprises: 1% by weight of titanium dioxide. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example A stabilised protective peptide formulation was prepared according to Example 1, wherein the formulation also comprises: 1% by weight of zinc oxide. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example 6 A stabilised protective peptide formulation was prepared according to Example 1, wherein the formulation also comprises: 1% by weight ofzirconia. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example 7 A stabilised protective peptide formulation was prepared according to Example 1, wherein the formulation also comprises: 500ppm of sodium fluoride. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example 8 A stabilised protective peptide formulation was prepared according to Example 1, wherein the formulation also comprises: 0.2% by weight methoxy dibenzoylmethane.

Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example 9 A stabilised protective peptide formulation was prepared according to Example 1, wherein the formulation also comprises: 0.8% by weight octyl methoxycinnamate. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example A stabilised protective peptide formulation was prepared according to Example 1- 9, wherein the formulation also comprises: 0.1% by weight flavourings. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example 11 A stabilised protective peptide formulation was prepared according to Example 1- 9, wherein the formulation also comprises: 0.1% by weight colourings. Other metal containing compounds and inorganic compounds could also be added to form complexes with segments of the peptide, or to be simply supported in the suspension by the gel structure provided by the clay.

Example 12 A 2.5% protective peptide formulation was prepared in the form of a gel according to Example 2 above, and its stability was compared with the high viscosity formulation described in US 5 015 628.

In the formulation described in US 5 015 628, a precipitate was immediately observed to form. However, there was no evidence of a precipitate with the peptide formulation of the present invention.

Similarly, a mouthwash was prepared according to Example 14 described below, and its stability compared with the mouthwash formulation outlined in US 5 015 628.

Odorous bacterial breakdown was evident in the mouthwash formulation outlined in US 5 015 628 after two days. However, after a period of 12 months, there was no evidence of odorous bacterial breakdown within the mouthwash prepared in accordance with Example 13 of the present invention.

Example 13 A stabilised mouthwash was prepared according to the following: Adding 2% phosphopeptide-calcium phosphate complex and 0.1% peppermint flavour to 5% ethanol and 92.9% water, and stirring with an overhead stirrer until the phosphopeptide-calcium phosphate complex was thoroughly dispersed.

Example 14 A stabilised mouthwash was prepared according to the following: Adding 2% phosphopeptide-calcium phosphate complex, 0.03% sodium benzoate, 0.1% sodium saccharin and 0.1% peppermint flavour to 5% ethanol and 92.77% water, and stirring with an overhead stirrer until the phosphopeptide-calcium phosphate complex was thoroughly dispersed.

Example A formulation for the treatment of dental sensitivity was prepared according to the following: 1) Part A: add 2.85% by weight of a synthetic smectic clay (Laponite) to 92.15% by weight water and 5% by weight ethanol. Stir with an overhead stirrer for two hours. The clay must be fully hydrated before further processing. When the stirrer is turned off, the mixture will form a gel.

2) Part B: prepare a 40% by weight casein phosphopeptide-calcium phosphate suspension by mixing together the following: 40% by weight casein phosphopeptidecalcium phosphate complex, 5% by weight ethanol and 55% by weight water.

3) Mix 12.5% by weight of Part B with 87.5% by weight of Part A to form a stable gel.

Example 16 A formulation for the treatment or prevention of gingivitis was prepared according to the following: Adding 2% phosphopeptide-calcium phosphate complex, 0.03% sodium benzoate, 0.1% sodium saccharin and 0.1% peppermint flavour to ethanol and 92.77% water, and stirring with an overhead stirrer until the phosphopeptidecalcium phosphate complex was thoroughly dispersed.

Example 17 A formulation for the treatment or prevention of mouth odour was prepared according to the following: Adding 2% phosphopeptide-calcium phosphate complex, 0.03% sodium benzoate, 0.1% sodium saccharin and 0.1% peppermint flavour to ethanol and 92.77% water, and stirring with an overhead stirrer until the phosphopeptidecalcium phosphate complex was thoroughly dispersed.

Example 18 A formulation for the remineralisation of tooth structure was prepared according to the following: 1) Part A: add 2.85% by weight of a synthetic smectic clay (Laponite) to 92.15% by weight water and 5% by weight ethanol. Stir with an overhead stirrer for two hours. The clay must be fully hydrated before further processing. When the stirrer is turned off, the mixture will form a gel.

2) Part B: prepare a 40% by weight casein phosphopeptide-calcium phosphate suspension by mixing together the following: 40% by weight casein phosphopeptidecalcium phosphate complex, 5% by weight ethanol and 55% by weight water.

3) Mix 12.5% by weight of Part B with 87.5% by weight of Part A to form a stable gel.

Example 19 A formulation for the modification of plaque to act as a buffer was prepared according to the following: Adding 2% phosphopeptide and 0.1% peppermint flavour to ethanol and 92.9% water, and stirring with an overhead stirrer until the phosphopeptide was thoroughly dispersed.

Example A formulation for the treatment or prevention of osteoporosis was prepared according to the following: Adding 5% phosphopeptide-calcium phosphate complex and 0.1% peppermint flavour to 5% ethanol and 89.9% water, and stirring with an overhead stirrer until the phosphopeptide-calcium phosphate complex was thoroughly dispersed.

Example 21 A formulation for the treatment or prevention of calculus was prepared according to the following: Adding 2% phosphopeptide and 0.1% peppermint flavour to 5% ethanol and 92.9% water, and stirring with an overhead stirrer until the phosphopeptide was thoroughly dispersed.

Example 22 The formulation outlined in Example 14 may be spray-dried to form a powder, and then added to a glass ionomer cement.

Industrial Applicability The stabilised protective peptide formulation of the present invention can be used in dental restorations, or as a general delivery system for bioactive substances.

Claims (2)

1. 2. Thle formulation of claim 1, wherein said thickening agent is selected from the 13 grouLp consisting of: a clay chosen from the group consisting of: synthetic hectorite including laponite D, laponite DF, laponite CP, and laponite SP 2002, calcium miontmorilIlonite, sodium montmorillonite (bentonite), sodium exchanged montmori Ilonite, acid activated bleaching earth and palygorskite, alginate, cellulose and cellulose derivatives, ca rboxymethylIcelIlu lose, irish moss, gum tragacanth, starch, polyvinylpyrrol idone, hydroxyethylpropylcel lulose, hydroxybutylmethylcell ulose, hydroxypropy Imethylcellulose, hydrox yethlIcel lu lose sodium carboxymethylcellulose, and colloidal silica.
2. 3. The formulation of claim I or 2, wherein said formulation further comprises at least one anti-microbial agent. Dated 16 May, 2000 2i Pacific Biolink Pty Limited Patent Attorneys for the Applicant/Nominated Person SPRUSON FERGUSON I 1DayLib\L~I BI-1285 78.doc: BAV