AU2022215313A1 - A METHOD OF USING A SILVER SALT PRECURSOR DENTAL SOLUTION, FOR SYNTHESISING SILVER NANO PARTICLES (AgNPs) ON ANY TOOTH SURFACE IN SITU FOR HUMAN AND VETERINARY ATRAUMATIC, NON-INVASIVE DENTAL TREATMENT. - Google Patents

Abstract

Provided are methods of generating silver nanoparticles (AgNPs) in situ on a surface of an animal or human such as on the surface of teeth. The AgNPs are synthesised by first applying a silver salt precursor solution (eg. AgF) to the surface of the teeth followed by the application of a reducing agent such as a solution of stannous fluoride, which thereby forms the AgNPs on said surface.

Description

A METHOD OF USING A SILVER SALT PRECURSOR DENTAL SOLUTION, FOR SYNTHESISING SILVER NANO PARTICLES (AgNPs) ON ANY TOOTH SURFACE IN SITU FOR HUMAN AND VETERINARY ATRAUMATIC, NON INVASIVE DENTAL TREATMENT.

FIELD OF THE INVENTION

[01] The present invention relates to a tooth surface treatment that uses a dental solution containing a metal precursor, such as silver fluoride, which is applied to a tooth surface, followed by an application of an organic or an improved stabilised inorganic reducing agent such as stannous fluoride. Part of the function of the reducing agent is to synthesise nano silver particles (AgNPs) in vivo at body temperature.

BACKGROUND

[02] Dental disease affects almost 3.5 billion people around the globe. Stannous fluoride has long been established as a topical fluoride and antimicrobial agent, with numerous benefits including anticaries, antiplaque, and anti-hypersensitivity properties. However, it has had a challenging history due to stabilising problems. Despite clinical effectiveness, there can be some stability problems especially with storage in challenging climate conditions of humidity and elevated temperatures. Currently, the main use of stannous fluoride is in dentifrices and gels but not in a dental solution form. A common concentration is around 0.42% stannous fluoride, used for daily follow-up home oral care, in contrast to Creighton's dental methodology, used by dental professionals in the dental surgery/dental office, the therapeutic efficacy of stannous fluoride depends on its stability. When free divalent stannous ions (Sn )in solution are oxidised to form tetravalent stannic ions (Sn4 )they no longer provide protective properties. Stannous ions disassociate in water to form tin (stannous) hydroxide, a process known as hydrolysis. All these chemical changes affect the bioavailability of the stannous ion resulting in deactivation of an essential therapeutic ingredient.

[03] Presently, with the state of the art, this problem has been lessened with limited success. One way to prevent the premature hydrolysis of the stannous ion in preparing 1% - 50% (w/v) SnF2(the preferred concentration for topical application) is by not using high speed mechanical mixing and instead, manually mixing the stannous fluoride to limit the incorporation of oxygen.

Accordingly, one method when preparing 330 ml of 10% stannous fluoride (SnF2), 33 g of the SnF2, powder is dissolved in minimum of ultra-pure water that being: 47 ml at 100 degrees Celsius (°C) and making up to volume with glycerol: sorbitol mixture at a ratio of glycerol between 1%-60%w/v: sorbitol between 1%- 40% w/v at 100 °C, and stirring manually with a plastic spoon, until the solution is completely clear.

[04] Nonetheless, inherent instability of stannous fluoride (SnF2) can become evident resulting in the formation of a white precipitate described in the preceding paragraphs especially in areas of high humidity and temperatures. Typical examples are the high humid and elevated temperature climates as in Australia's Northern Territory (N.T) and Queensland (Qld).

[05] Australian Patent No. 2021105210 (Aug 2021) prior art, discloses a carious lesion staining dental solution, known as dental Caries Status Disclosing Solution (CSDS). Stabilised aqueous silver fluoride is applied first to a carious lesion, followed by an inorganic metallic salt (stannous fluoride, being preferred metal salt) or an organic reducing agent. The intended use of this procedure is to stain a carious lesion black. If the lesion remains black over a period it is an indication that caries is not progressing (i.e., caries is arrested). However, if the lesion starts to lighten it is an indication caries is progressing. The existing best method of mitigating the stannous fluoride dental solution's inherent instability is that described in[03]

[06] US Patent No. 4,418,057, published on 29 November 1983, discloses a method of forming a stable gel of stannous fluoride. The method involved in the production of commercial-scale size batches (307 litres) of stable gel containing 0.38%-0.42% stannous fluoride, 96% to 98% anhydrous glycerine (glycerol), 1.8% to 2.2% hydroxyethyl cellulose, stable against deterioration over a prolonged period, in toothpaste, sufficiently viscous to be applied with a toothbrush as follow-up preventive home treatment. However, this prior art is unable to disclose a dental solution other than in gel form.

[07] As such the need exists for a method of stabilising stannous ions in the stannous fluoride reducing solution, to assist in forming nano silver (AgNPs) particles in an oral environment at body temperature.

SUMMARY

[08] It is an object of the present invention to provide a method, to stabilise the

highly active stannous ions in a reducing dental solution, to form a clear solution, with no sign of the characteristic white precipitate which identifies deactivated stannous

ions. The absence of this white powder precipitate is an accurate visual signal of an active stannous ion, the therapeutic ingredients', bioavailability, for at least 24 months

in high humidity and temperature environments.

[09] It is another object of the present invention to provide a method of using a metal precursor such as a stable silver salt dental solution to be used for people's and

veterinary dental treatment. Suitable silver salts include a) silver fluoride (AgF), b) silver diammine fluoride (SDF), c) silver nitrate ((AgNO 3), and d) silver diammine

nitrate (SDN). One of the above silver salts is applied to a tooth surface, followed by an inorganic or organic dental reducing agent solution. This method synthesises silver

nano particles (AgNPs) on the surface of a tooth, in situ, in the mouth (in vivo) at body temperature; AgNPs act as a reservoir of silver ions, the active therapeutic

component. A reservoir of silver ions can continuously replenish the spent silver ions,

diminishing the probability that caries will re-emerge (BMC Oral Health volume 13,

Article number: 73 (2013).

[010] Other objectives and advantages will become apparent when taken into consideration with the following specification.

[011] It is also an object of the present invention to overcome or ameliorate at least one of the disadvantages of the prior art, or to provide a useful alternative.

[012] In one aspect of the present invention, there is provided a method of synthesising silver nano particles (AgNPs) on a surface in situ for human or animal, comprising the step of: applying a silver salt precursor solution on the surface, followed by applying an inorganic or organic reducing agent, wherein the reducing agent reduces the silver ions forming AgNPs on the surface.

[013] Preferably, the AgNPs act as a reservoir of silver ions, and the reservoir of silver ions continuously replenishes the spent silver ions on the surface.

[014] Preferably, the silver salt precursor solution comprises any one or more of: aqueous silver fluoride stabilised with nitic acid (AgF), silver diammine fluoride (SDF) stabilised with ammonia, a silver nitrate (AgN03) and silver diammine nitrate stabilised with ammonia (SDN).

[015] Preferably, the aqueous silver fluoride comprises between 1%-50% w/v silver fluoride solution.

[016] Preferably, the aqueous silver fluoride is made by dissolving silver fluoride powder in ultrapure water.

[017] Preferably, the aqueous silver fluoride is made by dissolving silver fluoride powder in ultrapure water, which can be purged with argon gas.

[018] Preferably, the silver diammine fluoride comprises between 1%-50% w/v silver fluoride solution.

[019] Preferably, the silver diammine fluoride is made by dissolving silver fluoride powder in water with the pH adjusted with ammonia between1%-28% w/v.

[020] Preferably, the inorganic reducing agent is a metallic salt.

[021] Preferably, the metallic salt is between 1%-50% w/v stannous fluoride.

[022] Preferably, the metallic salt is at least one of ferrous fluoride or potassium iodide.

[023] Preferably, the metallic salt comprises between 1%-50% w/v of the reducing agent.

[024] Preferably, the metallic salt reducing solution is made with the complete elimination water, however it may have added glycerol.

[025] Preferably, the glycerol comprises between 1%-100% w/v and between 1% to 20% sorbitol w/v of the reducing agent solution.

[026] Preferably, the glycerol is at a temperature between 25-185 °C.

[027] Preferably, the reducing agent solution is mechanically mixed.

[028] Preferably, the organic reducing agent comprises at least one of tannic acid, a polyphenol, eugenol, a phenylpropanoid, an oligosaccharide, a polysaccharide, a monosaccharide and a disaccharide, a polyethylene glycol, and lactic acid bacteria (Lactobacillus genus).

[029] Preferably, the monosaccharide comprises at least one of aldoses and ketoses classes of organic chemical compounds, and the disaccharide may comprise at least one of lactose, maltose, and calcium sucrose phosphate.

[030] Preferably, the monosaccharide comprises dietary monosaccharides with at least one of galactose, glucose, or fructose,

[031] Preferably, the oligosaccharide comprises at least one of starch or starch derivatives, and wherein at least one of starch and starch-derivatives comprise at least one of glucose syrup, maltodextrin, or dextrin.

[032] Preferably, the polysaccharide(non-sugars) comprises at least one of cellulose, starch, glycogen, or chitin.

[033] Preferably, the organic reducing agent is an artificial sweetener/sugar substitute which can be made in the laboratory,

[034] Preferably, the sugar substitute/artificial sweetener is based on at least one of dextrose, maltodextrin, or sucralose.

[035] Preferably, the reducing agent is a flowing solution which can be applied with a microbrush in volumes as low as 0.01 ml and contains at least one suitable flavouring agent.

DESCRIPTION OF THE INVENTION

[036] Description of the embodiments is for the purposes of promoting the understanding of the principles in accordance with the disclosures, reference will now be made to the embodiments described herein.

[037] It will nevertheless be understood that no limitation of the scope of the disclosures is thereby intended. Any alterations, and further modifications of the inventive features illustrated herein, and any additional applications of the principles of the disclosure as illustrated herein, which would normally occur to one, skilled in the relevant art and having possession of this disclosure, are to be considered within the scope of the disclosure.

[038] One problem of using silver fluoride dental solutions followed by application of inorganic metallic silver salts such as stabilised stannous fluoride for treating dental lesions is that over time the efficacy of the treatment can diminish, and caries are re activated.

[039] To solve this problem, in one embodiment of the present invention, there is provided a use of a dental solution of an aqueous silver fluoride stabilised with nitric

acid or a silver fluoride solution stabilised by ammonia or a silver nitrate solution (AgO 3 ), or a silver diammine nitrate (SDN) stabilised with ammonia,

[040] The method may be followed by an application of an inorganic or organic reducing agent.

[041] In one method of the embodiment, the following steps were taken; preparing improved stabilised stannous fluoride by dissolving 10% (w/v) of stannous fluoride powder with between 1-100% (w/v) glycerol (glycerine), and sorbitol between 1%-20% (w/v)at a temperature of between 25-185 °C, resulted in there being no observed change in stannous ion levels after aging at 55°C for 80 days, (the equivalent of 24 months in a high humidity and .temperature environment) i.e., the solution was clear with no evidence of a white precipitate formed by oxidation of stannous ions.

[042] In another embodiment of the invention, 40% (w/v) silver fluoride was reduced with an equal volume of 10% (w/v) improved stabilised stannous fluoride. The precipitate was washed and dried and subjected to EDX analysis. The analysis confirmed the existence of nano silver particles, confirming the reduction of a silver fluoride dental solution, the preferred metal salt precursor, by improved stabilised stannous fluoride, the preferred metal salt reducing agent, producing nano silver (AgNPs) particles.

[043] In another embodiment according to the present invention, a tooth was treated with aqueous silver fluoride followed by improved stabilised stannous fluoride, formed a black matte on the tooth's treated surface. Scanning electron microscope (SEM) examination (with a capability of nano size resolution) . of the black matte, showed nano silver particles (AgNPs) appeared on the surface following the application of the SnF2 reductant.

[044] As such, in one embodiment of the present invention confirms that:

a) the using a stannous fluoride solution made by dissolving stannous fluoride powder comprising 10% (w/v), between 1 - 100% (w/v) glycerol and sorbitol between 1% - 20% (w/v) (heated to between 25-185 C)), increased the longevity of stannous fluoride reducing solutions in humid and elevated temperature environments.

b) the reduction of a silver fluoride dental solution, the preferred metal precursor, with improved stabilised stannous fluoride, the preferred metal reducing agent, reduces silver ions (Ag*) as well as producing nano silver (AgNPs) particles in an oral environment at body temperature, helping maintain silver ions at therapeutic levels.

[045] The preferred embodiment is a metal fluoride solution comprising improved stannous fluoride, although there are diverse types of suitable reducing agent solutions. The preferred quantity of the reducing agent in solution is between 1% and 50% (w/v).

[046] The improved stabilised stannous fluoride works as reducing solution described in [041], wherein stannous fluoride powder is added to the stannous fluoride as reducing solution at between 25-185 °C.

[047] The purpose of stabilising the stannous fluoride reducing solution using the method described in [041] is to stabilise the highly active stannous ions to form a clear solution with no sign of the characteristic white precipitate which identifies deactivated stannous ion. The absence of this white powder precipitate is an accurate visual signal of stannous ions, the active therapeutic ingredients, bioavailability, for at least 24 months in high humidity and temperature environments.

[048] Specifically, stannous fluoride manufactured in the way described in [041] is stable in high humidity and temperature environments.

[049] Silver fluoride solution

[050] The preferred silver salt precursors are a) silver fluoride (AgF), b) silver diammine fluoride (SDF), c) silver nitrate ((AgNO 3 ), and d) silver diammine nitrate (SDN). Silver fluoride in a), wherein nitric acid is used to bring the pH of the dental solution down to between 5.0-6, and the silver fluoride described in: b), and d) wherein the pH of silver diammine ammonia fluoride, and silver diammine nitrate solutions are adjusted with ammonia between 1% - 28% w/v, and c) silver nitrate (powder) is dissolved in water.

[051] As such, applying any of the silver fluoride dental solution specified in [050] a), [050] b), [050] c) and [050] d) to a tooth surface carious lesion for 1-3 minutes, subsequently followed by, but not exclusively, an application of improved stabilised stannous fluoride immediately, and covered with a medium to exclude saliva allowing the reaction to go to completion, formulated to be stable in places of high humidity and elevated temperatures as specified in [041]. This method can produce nano silver particles AgNPs on the tooth surface after the application of improved stable stannous fluoride reducing agent; AgNPs act as a reservoir of silver ions.

[052] In one embodiment of the present invention, it provides a novel and new method for addressing the need for a stabilised stannous fluoride dental solution by;

a) not dissolving stannous fluoride 10% (w/v) (w/v) powder in water, excluding any water to make a liquid stannous fluoride dental solution, instead dissolving stannous fluoride powder in, between 1%-100% w/v anhydrous glycerol and sorbitol in between 1%-20% (w/v), at a temperature of;

b) between 25 °C-185 °C

c) allows the mixture to be mechanically mixed.

d) the preferred concentration of SnF2 is between 1% to 50% w/v.

[053] A liquid stannous fluoride dental solution made using this method (described in [041] was subjected to accelerated age testing at 55 0 C for 80 days in high humidity and temperature environments, the equivalent of 24 months real time testing. A positive result i.e., the absence of the characteristic white precipitate, indicated the stannous ion, the active therapeutic ingredient was stable, showed this method of producing an improved stable stannous fluoride dental solution was successful.

[054] In a prior art embodiment, a stabilised silver fluoride is applied to a tooth surface for 1-3 minutes, followed by an inorganic or organic reducing agent. Intended to stain the carious lesion; the colour, if black, is a reliable visual signal caries had not progressed; while if the resulting black colouration is lightened or returns to the original yellow-brown caries colour is a reliable sign caries is still progressing. It is a Caries Status Disclosing Solution (CSDS). The intended use is to provide a visual signal a universal, uniform standard for diagnosis of caries progressing (re-emergence or non progression)

[055] In one embodiment of the present invention, there is provided a method of synthesising a reservoir of nano silver particles (AgNPs) that release Ag* ions (silver), which serve as a reservoir for anti-microbial action, Ag* ions produced, attack bacterial cell walls, on a tooth surface using any silver metallic precursor described in [050].

dental solution followed by an advanced stabilised (with between 1-% 100% anydrous glycerine/glycerol (w/v) and between 1-20% sorbitol (w/v), at between 25-185 °C), inorganic metal salt, such as improved stabilised stannous fluoride, the preferred inorganic reducing agent, or an organic reducing agent. The preferred silver salt precursors are described in [050] a), b),c), and d) or any other silver salt dental solution, is capable of synthesising AgNPs in situ under under physiological conditions of temperature and pressure on a tooth surface orally.

[056] Prior art, Australian Patent 2021105210 discloses a Caries Status Disclosing solution that by staining caries different colours shows caries is progressing or not progressing. This invention uses an AgF solution followed by a different stabilised stannous fluoride, made as described in [03], its intended use is to stain caries to disclose the caries status of the lesions to be either progressing or not-progressing; unlike the Creighton invention method, intended to synthesise AgNPs.

[057] This is one of the differences between the present invention and that of the prior art. As described in [054, and 56] above.

[058] US Patent No 6.461.161.B1, published on 8 October 2002 by Ngo, Knight, Craig et al, discloses a tooth treatment method for inhibition of progress of dental caries and for immediate relief of dental hypersensitivity in adults over age 21. The intended use of this prior art method is for a tooth surface treatment for immediate dentinal hypersensitivity relief for adults over the age of twenty-one (21) https://www.sdi.com.au/en-us/product/rivastar/

[059] Australian Dental Journal 2012; 57: 308-311doi: 10.1111/j.1834 7819.2012.01700, discloses a clinical evaluation of diamine silver fluoride/potassium iodide as a dentine desensitizing agent. It is a pilot study conducted by G.G Craig, GM Knight, JM McIntyre. https://onlinelibrary.wiley.com/doi/epdf/10.1111/j.1834 7819.2012.01700.x

[060] SDI Riva Aqua is prior art intended for the purpose listed in the following table.

[061]

Riva Star Riva Star Aqua Silver Diammine Fluoride (SDF) Silver Fluoride (AgF) USA • Treatment of dentinal • Treatment of dentinal hypersensitivity hypersensitivity e For use in adults over the age of e For use in adults over the age of 21 21 Brazil / Latin e Desensitising cervical tooth e Desensitising cervical tooth America hypersensitivity hypersensitivity " Arresting caries e Arresting caries " Detecting caries e Detecting caries Australia / e Desensitising cervical tooth e Desensitising cervical tooth Indications Europe hypersensitivity hypersensitivity Canada e Anti-caries • Not currently available in Canada e Prevents, fights, and/or protect against cavities or caries. Effective decay preventive fluoride. e Helps reverse the tooth decay process at the earliest stage before it can become a cavity. e Helps to reduce sensitivity in cervical hypersensitive teeth in adults. Process 2 Steps patented process 2 Steps patented process

Metal salt inorganic reducing agents

[062] In one preferred embodiment of the present invention, the metallic salt reducing agent is at least one of stannous fluoride, (in any form) the preferred dental reducing solution, ferrous fluoride or potassium iodide.

[063] In one preferred embodiment, the inorganic metallic salt has a concentration of between 1% to 50% w/v of the reducing agent solution.

[064] In one preferred embodiment, the inorganic metallic salt consists of stannous fluoride, in the form of a free flowing liquid dental solution.

[065] In one embodiment, the preferred concentration of SnF2is between 1% to 50% w/v.

[066] In one preferred embodiment, the stannous fluoride free flowing liquid dental solution is only available to dental professionals.

[067] In one preferred embodiment, the stannous fluoride free flowing liquid dental solution is capable of being applied with a microbrush, in volumes as low as 0.01 millilitre (ml).

[068] In one preferred embodiment, the stannous fluoride liquid dental solution is made by dissolving 10% (w/v) stannous fluoride powder in 1%-100% w/v in anhydrous glycerol and 1%-20% (w/v) sorbitol, at a temperature of:

a) between 25 °C-185 °C; and

b) the mixture is mechanically mixed.

[069] This method resulted in an improved liquid stannous fluoride dental solution stable for the equivalent to 24 months at high humidity and temperature conditions (using accelerated age testing at 55 0 C for 80 days).

[070] In one preferred embodiment, the liquid stannous fluoride is made under physiological conditions of temperature and pressure without specialised equipment with automated air moisture and temperature control capabilities.

[071] In one preferred embodiment, the liquid stannous fluoride commercial size batched is 6 - 10 litres.

[072] In one preferred embodiment, the liquid stannous fluoride is produced in one stage only, not in a sequence of stages.

[073] In one preferred embodiment, the temperature is not reduced during production by adding more glycerol to a commercial size batch of 6-10 litres.

Organic reducing agents

[074] In one preferred embodiment of the present invention, the concentration of the organic reducing agent is between 1%-50% w/v.

[075] In one preferred embodiment, the organic reducing agent, comprises at least one of eugenol, tannic acid, polyphenol, any type of phenylpropanoids , and organic compounds such as flavonoids biosynthesis associated with phenylpropanoids.

[076] In one preferred embodiment, the organic reducing agent comprises of sugar of at least an oligosaccharide of at least one of starch, starch-derivatives, glucose syrup. maltodextrin and dextrin, polysaccharides, monosaccharide of at least one of aldoses and ketoses class of organic compounds and dietary monosacchirides comprising of at least one of galactose, glucose, fructose, disaccharide at least one of lactose, maltose, sucrose and calcium, sucrose phosphate.

Artificial sweeteners and sugar substitutes

[077] In one preferred embodiment of the present invention, the dental solution comprises one or more ingredients of artificial sweeteners such as maltodextrin, sucrose, saccharine, and sucralose.

[078] In one embodiment, different functional groups in reducing agents such as ketone, aldehyde, hydroxyl, amine(ammonia), and carboxyl groups can be responsible for the reduction of different metal ions to metal atoms. Aldehyde groups and their concentration in reducing sugars play an essential role in nanoparticle formation.

[079] Different types of sugars are used in the presence of ammonia or sodium hydroxide as accelerator reagents to control the pH and increase the rate of reduction and growth of silver AgNPs. (Not necessary with the present invention, because ammonia is inherently present in silver diammine fluoride (SDF), and silver diammine nitrate (SDN), metal precursor used in Creighton's method).

[080] In one embodiment, sugar substitutes and artificial sweeteners that can be made in the laboratory, such as sucralose, saccharin, and aspartame or can be natural products such as xylitol and sorbitol, are used to synthesise AgNPs.

[081] In one embodiment, commercially available sugars (white and brown), sugar substitutes and artificial sweeteners brands, Splenda, Sweet'N Low, Equal Original, Caribou Coffee, Great Value. Stevia and Whole Earth are used as reductants.

[082] In one embodiment, each sugar and sugar substitutes and artificial sweeteners of the commercially available sugars mentioned, consist of different ingredients such as saccharin. aspartame, sorbitol, acesulfame, potassium, sucralose, cyclamate, xylitol, erythritol, neotame and stevia, each with different functional groups as NH-, OH, NH2, OCH3, CH2OH, for reducing metal ions.

[083] In one prior art, the method comprises the step of using one of the sweeteners, an aqueous solution was heated to 50 °C, under magnetic stirring. However, the present invention does not require heating the solution up to 50 °C or magnetic stirring.

[084] In one prior art embodiment, silver nitrate ((AgNO 3) the metal precursor, is heated and added slowly to the mixture. This is time consuming. However, the present invention does not require heating the solution up to 50 °C or adding to the mixture slowly.

[085] In one prior art embodiment, the ((AgNO3) metal precursor was sonicated for 2 3 minutes. However, the present invention does not require the ((AgNO 3) metal precursor being sonicated.

[086] In one embodiment, silver atoms (Ago) produced during metal ion reduction may undergo some transformation through cluster formation.

[087] In one prior art embodiment, a high temperature above 150 °C is required at the beginning of the reaction to convert the reducing agent and the reduction of silver ions to silver atoms. However, the reduction of silver ions to silver atoms can also be done at a low temperature using sugar substitutes at 50 °C can increase the yield of reducing agent. The Creighton method does not require temperatures above body temperature to increase the yield of the reductant.

[088] In one embodiment, Splenda is the stronger of artificial sweeteners compared to Sweet'N Low, Equal Original, Great Value, and NJOY. Most artificial sweeteners (except Splenda)use Polyvinylpyrrolidone (PVP) as a capping agent to regulate the size of AgNPs. Using Splenda as a reducing agent, in the presence of PVP as a capping agent, caused less reduction of silver ions and formation of AgNPs compared to Splenda without PVP. The Invention uses Splenda, the preferred artificial sweetener organic reducing agent avoiding the use of PVP capping agent as it causes less formation of silver ions.

[089] In conclusion, different artificial sweeteners showed different abilities to reduce Ag* the ones that contain sucralose and maltodextrin as the main ingredients could act as a reducing agent in the absence of sodium hydroxide (NaOH). (Creighton uses sucralose and maltodextrin as reductants without NaOH).

Organic reducing agents-Creighton novel method

[090] In one embodiment, the present invention uses one or more of the reducing agents and / or artificial sweeteners in a novel way. This distinguished Creighton's method from the prior art described in Australian Patent 2021105210, Caries Status Disclosing solution (CSDS) use of organic reducing agents which does not include artificial sweeteners/sugar substitutes to stain carious lesions.

[091] In one embodiment, there is provided a novel method that uses one or more of the main ingredients of artifical sweeteners such as maltodextrin, sucrose, saccharine and sucralose, but unlike the state of the art, Creighton's method does not add sodium hydroxide during the synthesis to increase the rate of silver nano particle formation. The rate of AgNPs formed is adequate for the intended use.

[092] In one embodiment, different types of sugars have been used in the presence of ammonia or sodium hydroxide as accelerator reagents to control the pH and increase the rate of reduction and growth of silver AgNPs. Creighton's method does not add NaOH or ammonia to act as an accelerator to control the pH and increase the rate of reduction and growth of AgNPs, as ammonia is inherent in the SDF or SDN as mentioned in the above paragraph [050].

[093] In one embodiment, the methods in the aforementioned paragraphs, described the use of one of the sweeteners, and an aqueous solution heated to 50 °C, under magnetic stirring. An alternative method in one embodiment (Creighton's method) does not require heating the aqueous solution in which the AgNPs are synthesised, AgNPs are synthesised in the mouth in situ (in vivo) under physiological conditions of temperature and pressure without magnetic stirring.

[094] In one embodiment of the present invention, there is provided a method which comprises the step of the reducing agent not being directly heated or controlled; AgNPs are synthesised in vivo under physiological conditions of temperature and pressure.

[095] In one embodiment, the present invention provides a method where the step of, the metal precursor AgF is not heated but remains at body temperature. The method used in the prior art, wherein the metal precursor, silver nitrate (AgNO 3.), is sonicated for 2-3 minutes (sonicated is the process of applying sound energy to agitate particles in a liquid). No sonicated process is used in this invention, the preferred metal precursor (AgF, SDF, AgNO3, and SDN) is applied to a carious tooth surface for 3 minutes, this is followed by an application of a reducing agent immediately, and covered with a medium to exclude saliva, allowing the reaction to go to completion,

[096] In one embodiment, the artificial sweetener, such as SplendaTM, is used in the absence of Polyvinylpyrrolidone (PVP), a capping agent, to avoid reduced production of silver ions and AgNPs resulting in a high yield of AgNPs.

Silver fluoride-state of the art

[097] In one prior art embodiment, uses a stabilised silver fluoride applied to a tooth surface for 1-3 minutes, followed by the application of an inorganic or organic reducing dental solution agent. Intended to stain the carious lesion; the colour, if black, is a reliable visual indication caries had not progressed; while, if the resulting black colouration is lightened or returns to its original caries colour of yellow brown, it is a reliable sign caries is still progressing. The intended use is to provide a visual signal of a universal standard, unambiguous, and uniform diagnosis of caries progression or non progression.

[098] Referring to prior art Australian Patent No. 2021105210, showed no evidence of nano silver particles AgNPs synthesis which produced silver ions that act as a reservoir maintaining their bactericidal potency.

[099] As shown in (Craig, Powell and Price, 2013), the concentration of silver ions diminishes in the prior art product described in Australian Patent Application No. 2021105210 by the lightening of the black stain or re-emergence of yellow brown coluor of caries lesions indicating caries progression. Indication of caries progression as described above is due to the diminished concentration of silver ions as no reservoir of AgNPs is produced to replenish the spent Ag'ions.

[0100] The absence of an AgNPs reservoir [099] using the state of the art SnF2 where stannous fluoride was not made by the improved stability method that eliminates hydrolysis to stabilise stannous ions by using between 1%- 100% (w/v) glycerol, and between 1%-20% (w/v) sorbitol at 25-185 °C [041]

[0101] In the prior art Australian Patent Application No. 2021105210, the Stannous Fluoride is manufactured with a minimum volume of water (47 ml) to dissolve SnF2 powder in ultra-pure water at 100 °C, and then glycerol: sorbitol mixture at a ratio of glycerol between 1%-60%w/v: sorbitol between 1%- 40% w/v at 100 °C), added to the SnF2 powder dissolved in ultra pure water [03]. This was the best method available to mitigate the inherent instability of the stannous ion in high moisture and temperature environments. This method was partially successful.

[0102] In one preferred method of the present invention, produces an improved stabilised SnF2 described in [041]. Using stabilised AgF followed by improved stabilised SnF2 dental reducing solution, an abundance of AgNPs are produced which act as a reservoir of silver ions reducing the possibility of caries re-suregence.

[0103] A medicament embodiment, of the method described above, the medicament comprising the method of synthesising silver nano particles (AgNPs) on a surface in situ for human or animal, the method comprising the step of applying a silver salt precursor solution on the surface, followed by applying an inorganic or organic reducing agent, wherein the reducing agent reduces the silver forming AgNPs on the surface.

[0104] Although the invention has been described with reference to specific examples, it will be appreciated by those skilled in the art that the invention may be embodied in many other forms, in keeping with the broad principles and the spirit of the invention described herein.

[0105] The present invention and the described embodiments specifically include the best method known to the applicant of performing the invention. The present invention and the described preferred embodiments specifically include at least one feature that is industrially applicable.

Claims (22)

CLAIMS:

1. A method of synthesising silver nano particles (AgNPs) on a surface in situ for human or animal, the method comprising the step of applying a silver salt precursor solution on the surface, followed by applying an inorganic or organic reducing agent, wherein the reducing agent reduces the silver forming AgNPs on the surface.

2. The method of Claim 1, wherein the silver salt precursor solution comprises any one or more of: aqueous silver fluoride stabilised with nitic acid (AgF), silver diammine fluoride (SDF) stabilised with ammonia, a silver nitrate (AgNO3), and silver diammine nitrate stabilised with ammonia (SDN).

3. The method of Claim 2, wherein the aqueous silver fluoride, and silver nitrate comprises between 1%-50% w/v silver fluoride solution.

4. The method of Claim 2 or Claim 3, wherein the aqueous silver fluoride is made by dissolving silver fluoride powder in ultrapure water, and silver nitrate is made by dissolving silver nitrate powder in ultrapure water.

5. The method of any one of Claims 2 to 4, wherein the aqueous silver fluoride is made by dissolving silver fluoride powder in ultrapure water, which can be purged with argon gas.

6. The method of Claim 2, wherein the silver diammine fluoride, and silver diammine nitrate comprises between 1%- 5 0% w/v silver fluoride solution.

7. The method of Claim 2, wherein the silver diammine fluoride is made by dissolving silver fluoride powder in water, silver diamine nitrate (SDN) is made by dissolving silver nitrate powder in water, with the pH adjusted with ammonia between 1%- 28%% w/v.

8. The method of any one of Claims 1 to 7, wherein the reducing agent is a flowing solution which can be applied with a microbrush in volumes as low as 0.01 ml and contains at least one suitable flavouring agent.

9. The method of any one of Claims I to 8, wherein the inorganic reducing agent is a metallic salt.

10. The method of Claim 9, wherein the metallic salt is between 1%-50 w/v stannous fluoride.

11. The method of Claim 9, wherein the metallic salt is at least one of ferrous fluoride, or potassium iodide.

12. The method of any one of Claims 9 to 11, wherein the metallic salt comprises between 1%-50% w/v of the reducing agent.

13. The method of any one of Claims 9 to 12, wherein reducing solutions have added glycerol and sorbitol.

14. The method of Claim 13, wherein the glycerol comprises between 1%-100% w/v, and sorbitol between 1% to 20% (w/v) of the reducing agent solution.

15. The method of Claim 14, wherein the glycerol is heated to a temperature between 25-185 °C.

16. The method of Claim 15, wherein the reducing agent solution is mechanically mixed.

17. The method of any one of Claims 1 to 8, wherein the organic reducing agent comprises at least one of tannic acid, a polyphenol, eugenol, any type of phenylpropanoid, and organic compounds such as flavonoids biosynthesis associated with phenylpropanoids, an oligosaccharide, a polysaccharide, a monosaccharide and a disaccharide, a polyethylene glycol, or lactic acid bacteria.

18. The method of Claim 17, wherein the monosaccharide comprises at least one of aldoses and ketoses classes of organic chemical compounds, and the disaccharide comprises calcium sucrose phosphate or wherein the monosaccharide comprises dietary monosaccharides with at least one of galactose, glucose, or fructose,

19. The method of Claim 17, wherein the oligosaccharide comprises at least one of starch or starch-derivatives, and wherein at least one of starch and starch-derivatives comprise at least one of glucose syrup, maltodextrin, or dextrin.

20. The method of Claim 17, wherein the polysaccharide(non-sugars) comprises at least one of cellulose, starch, glycogen, or chitin.

21. The method of any one of Claims 1 to 8, wherein the organic reducing agent is an artificial sweetener/sugar substitute which can be made in the laboratory,

22. The method of claim 21 wherein the sugar substitute/artificial sweetener is based on at least one of dextrose, maltodextrin, or sucralose.