Is Mouthwash Heterogeneous Or Homogeneous
Title
REDUCED ALCOHOL MOUTHWASH
BACKGROUND OF THE INVENTION
Field of the Invention
The invention relates mostly to mouthwashes for the prevention and elimination of bad breath likewise as for the reduction of oral microorganisms responsible for the development of dental plaque and tooth decay.
Dental plaque can pb to the formation of calculus, gingivitis and other related gum diseases. In
detail, the present invention relates to a reduced alcohol mouthwash that is constructive in preventing those bug.
Description of Related Art
Oral rinse and mouthwash compositions take been used past people for many years for the prevention of bad breath and for the elimination of bacteria and other oral microorganisms that are responsible not only for bad breath but also tooth decay, plaque and gum diseases such as gingivitis and periodontitis. To this end, antiseptic mouthwashes in the past have been designed to clean the oral cavity, provide fresh breath and impale these pathogenic microbes.
The leading antiseptic mouthwashes have always
contained alcohol (i.e., ethanol) at adequately loftier levels, ranging from approximately 25 up to about 30% by volume, based on the total mouthwash volume
(hereinafter referred to every bit "% five/v"). Alcohol is used both as a vehicle and as a solvent in which the active ingredients, and additives such as astringents,
fluorides, color additives, flavor oils, and the like, can exist dissolved and and then dispersed into solution.
Alcohol also provides a preservative role for the mouthwash during storage and apply, and enhances the flavor oil organoleptic cues.
However, the utilize of loftier levels of alcohol has been recently challenged from an overall health standpoint. Some people cannot tolerate alcohol in any form, and there is concern that young children in particular may be adversely affected by ingesting or inadvertently swallowing mouthwash compositions containing alcohol. Senior citizens take as well complained well-nigh problems related to gargling with such mouthwashes, and chronic exposure has been found to upshot in gum "burn down" by the booze. It has likewise been reported that alcoholic mouthwashes can result in an unpleasant "dry out mouth" sensation. Notwithstanding, just reducing the levels of alcohol in these mouthwash compositions has significant disadvantages. It has been institute that lower alcohol levels issue not only in a loss in the solubility of the actives and other ingredients in the composition, simply there is also a noticeable decrease in the ability of the composition to kill the oral microorganisms responsible for bad breath, plaque and gum disease. This loss in antimicrobial activeness is not only due to the reduction of alcohol as a vehicle, merely is also related to the bioavailability of the solubilized actives. Thymol, for instance, is a well known antiseptic
compound, as well known equally an essential oil, which is utilized for its antimicrobial activeness in a multifariousness of mouthwash preparations. In particular, thymol can be utilized in oral hygiene compositions such as mouth rinses in sufficient quantities to provide desired benign therapeutic effects. Listerine®-brand mouthwash is a well-known clarified mouthwash that has been used past millions of people for over 1 hundred years and has been proven constructive in killing microbes in the mouth that are responsible for plaque, gingivitis and bad jiff. Thymol, together with other essential oils such every bit methyl salicylate, menthol and eucalyptol, are active ingredients in clarified mouth rinses such as Listerine®. These oils achieve their efficacy although present in small amounts. Without being restricted to any specific theory, it is now believed that the efficacy and taste of antiseptic mouthwashes such equally Listerine® may be due to the dissolution of these iv active ingredients.
Dissolution is also of import from an artful bespeak of view since a clear mouthwash solution is certainly preferred by consumers to one that is cloudy, turbid or heterogeneous. Obviously and then, there is a substantial demand for the development of a reduced alcohol mouthwash in which the essential oils are completely dissolved and that continues to be effective in the prevention of bad breath, the killing of oral microbes and the resultant reduction or elimination of plaque. Several attempts at formulating reduced alcohol and booze-free mouthwash compositions tin be found in the prior art. For example, United States Patent No.
v,292,527 to Konopa discloses a non-alcoholic, aqueous mouthwash limerick which contains a water- insoluble flavoring oil and an effective corporeality of a dispersion system comprising a non-ionic surfactant selected from a hydrogenated brush oil and a polyoxyethylene- polyoxypropylene block copolymer having about fifty% to near 90% ethylene oxide, a humectant and a
disinfecting amount of a cationic antimicrobial amanuensis such as cetylpyridinium chloride. The humectant, which is preferably glycerin, but which may besides be sorbitol or mannitol, is generally present in amounts from nigh 1% to 20% by weight. The composition allegedly
exhibits a homogeneous, uniform advent and a high degree of bactericidal efficacy.
Us Patent No. v,284,648 to White et al.
discloses an booze-gratuitous oral rinse and pre-rinse consisting of emulsion-containing cleaning and coating compositions that are non-irritating and low foaming for maximum plaque disruption and removal. The
cleaning effect is achieved using an aqueous system containing a mouth conditioner such equally
polydimethylsiloxane emulsified in a surfactant such every bit a block copolymer of polyoxyethylene and
polyoxybutylene. U.s. Patent No. 5,185,153 to Pollock discloses a dental rinse formulation that allegedly lyses the bacterial cells responsible for plaque germination and bad jiff. The composition comprises (a) a humectant (such as glycerol, sorbitol and the like) in an amount of nearly 20-80%, preferably about xxx-50%, by weight, based on the entire limerick volume (hereinafter referred to as "% west/v"); (b) from about 0.five to most two% due west/v of a bicarbonate; (c) from virtually 0.05 to most ane% w/v of an anionic detergent; and (d) from about 0.01 to about three% v/5 of a non-ionic detergent such as Tween twenty. Booze may be added upward to a level of fifteen% v/v, with water comprising the remainder upward to 100%. It is postulated that the "high" levels of humectant cause the bacterial prison cell walls to swell causing separation of the jail cell wall and membrane, which enables the
surfactants to enter the prison cell and deregulate autolysin enzymes which attack the cell wall causing holes and eventual jail cell lysis.
United States Patent No. 5,236,699 to Libin teaches an antimicrobial mouthwash limerick comprising two agile agents, triclosan and cetylpyridinium chloride. Ethanol comprises about 15% -twenty% w/v of the
water/alcohol carrier system and sorbitol is added in amounts of about x% -xv% by weight equally a humectant. United States Patent No. 4,945,087 to Talwar et al. discloses an oral antiseptic composition containing thymol, eucalyptol, menthol and methyl salicylate wherein the unpleasant, medicinal taste of thymol is masked by the combined presence of an constructive amount of one or more carbohydrate alcohols and an effective corporeality of anethole. The saccharide alcohol may be present in amounts of nigh 20 to about 55% w/five, preferably almost 25-fifty% w/v, most preferably about 28-32% westward/v. The corporeality of anethole may be about 0.01-0.035% westward/5, most preferably about 0.015-0.025% west/v. The amount of ethanol is about 5-35% westward/5 and a level of 22.viii% w/v is specifically disclosed.
The assignee of the present inventors has, since about mid-1992, sold Cool Mint Listerine®-brand mouthwash product containing thymol and other essential oils, 21.6% 5/v ethanol, 17.v% w/five sorbitol, and 0.16- 0.20% west/v anethole.
United States Patent No. five,256,401 to Duckenfield et al. discloses a reduced alcohol antibacterial mouthwash in which a substantially water-insoluble non-cationic antibacterial/anti-plaque amanuensis such as triclosan is dissolved in a solvent whose water/alcohol ratio is greater than 10:i. Sorbitol, glycerin and other saccharide alcohols may be incorporated every bit humectants but zilch is attributed to their functioning as solubilizers or active enhancement compounds.
United states Patent No. 5,100,650 to Carlin et al. discloses an oral mouthwash containing an antibacterial active such every bit chlorhexidine with not-ionic surfactants and loftier levels of sorbitol which, it is asserted, increase the antibacterial activity of the
chlorhexidine compounds. Sorbitol solution (70%) is present in amounts of 25-75% w/v, preferably fifty-sixty% w/v, and the water/alcohol weight ratio ranges from nearly i:1 to well-nigh 20:1, preferably from about 3:ane to most 10:1. Clearly, at that place is however a need for a reduced alcohol composition that is highly efficacious in the
prevention of bad jiff, plaque and glue disease.
Moreover, there is a demand for such oral compositions that both kill the oral microflora responsible for these problems and make clean the oral cavity leaving a fresh, lubricous mouthfeel.
SUMMARY OF THE INVENTION The present invention is a reduced alcohol,
antimicrobial mouthwash composition with a high level of efficacy in the prevention of plaque, gum disease and bad breath. In improver, the oral mouthwash compositions of this invention are articulate, aesthetically highly-seasoned products. The reduced booze, antimicrobial mouthwash
composition of the present invention comprises (a) an antimicrobially constructive amount of one or more than
essential oils; (b) a reduced level, up to about 22% v/five, of ethanol; (c) at least about 18% westward/v of one or more polyols sufficient to provide enhanced
antimicrobial action of said essential oils at said reduced level of ethanol; and (d) water.
At ethanol levels of nearly 22% v/v and below, it has been found that relatively high levels of one or more polyols, such as the sugar alcohol, sorbitol, or propylene glycol or glycerin, surprisingly raise the antimicrobial activity of the essential oil actives even at these reduced ethanol levels. According to the present invention, the corporeality of polyols employed is generally inversely proportional to the corporeality of ethanol; that is, as the level of ethanol is reduced below about 22% v/5, it is generally necessary to increase the concentration of polyols across about eighteen% due west/5 to maintain good antimicrobial efficacy. The specific level of polyols employed will vary depending upon the reduced level of ethanol used, the particular polyols employed, and the antimicrobial efficacy desired. For example, in comparing to Listerine®- brand antiseptic mouthwashes containing almost 27% five/v ethanol, the mouthwashes of the nowadays invention are at least equally, or more, efficacious fifty-fifty at ethanol levels of 19% five/v, or lower. In one embodiment of the present invention, a reduced booze, antimicrobial mouthwash limerick is provided which comprises (a) an antimicrobially effective amount of one or more essential oils; (b) a reduced level, up to about 22% 5/v, of ethanol; (c) at least nearly 18% w/v of one or more polyols sufficient to provide an enhanced antimicrobial activeness of said essential oils at said reduced level of ethanol, such that the R-Factor (defined below) of said mouthwash composition is less than about 2.0, preferably less than about 1.2, and virtually preferably less than well-nigh 1.0; and (d) water.
Co-ordinate to another embodiment of the present
invention, a reduced booze, antimicrobial mouthwash composition is provided which comprises (a) an
antimicrobially effective amount of one or more
essential oils; (b) a reduced level, from about ix% 5/5 up to about 22% five/v, of ethanol; (c) at least well-nigh 18% w/v of i or more polyols sufficient to provide enhanced antimicrobial action of said essential oils at said reduced level of ethanol; and (d) h2o.
In another embodiment of the present invention, a reduced alcohol, antimicrobial mouthwash composition is provided which comprises (a) antimicrobially effective amounts of thymol and i or more other essential oils; (b) a reduced level, upward to most xix% v/v, of ethanol; (c) at least about 18% west/v of ane or more polyols sufficient to provide enhanced antimicrobial activity of said essential oils at said reduced level of
ethanol; and (d) h2o.
In another apotheosis of the present invention, a reduced booze, antimicrobial mouthwash composition is provided which comprises (a) antimicrobially effective amounts of thymol and one or more than other essential oils; (b) a reduced level, from about 12% five/5 up to about nineteen% v/v, of ethanol; (c) at to the lowest degree well-nigh eighteen% w/v of one or more than polyols selected from the grouping consisting of a sugar alcohol, propylene glycol and glycerin sufficient to provide enhanced antimicrobial action of said essential oils at said reduced level of ethanol; and (d) water.
The mouthwash compositions of this invention may as well contain flavoring oils, buffers, surfactants, colorants and other conventional additives. For example, minor amounts of anethole or other flavorants may be
included. When anethole is present in the compositions of the present invention, it may exist added in a pocket-size amount or it may exist introduced every bit part of a natural flavor alloy. According to another embodiment of the present
invention, a mouthwash production is provided which is substantially complimentary of anethole (i.due east., there is no
detectable level of anethole or at to the lowest degree the corporeality of any anethole present is so small-scale that it performs no substantial gustatory modality or flavor part). This particular embodiment contemplates that the desired taste or flavor attributes of the mouthwash are achieved through the utilise of flavorants and additives other than
anethole.
DETAILED DESCRIPTION OF THE INVENTION
The reduced alcohol mouthwash compositions of the present invention provide an unexpected high degree of clarified efficacy towards oral microorganisms
responsible for oral malodor and the build-upwards of plaque and calculus and the resulting tooth and gum diseases that may follow. The enhanced antimicrobial efficacy of the reduced alcohol mouthwash compositions of the present invention is attributed to the presence of minor amounts of one or more than essential oils (i.e., thymol, eucalyptol, menthol and methyl salicylate) combined with high levels (at least near eighteen% w/v) of one or more than polyols such as sorbitol which synergistically enhances the antimicrobial efficacy of the essential oils.
Thymol, (CH3)2CHCsixHthree(CHiii)OH (isopropyl-m-cresol), is merely slightly soluble in water but is soluble in alcohol, and its presence is one of the reasons booze was necessary in the well-established, high booze
commercial mouthwashes such every bit Listerine® in the past. Methyl salicylate, (Chalf-dozenH4OHCOOCH3), as well known every bit
wintergreen oil, additionally provides flavoring to the mouthwash together with its antimicrobial role.
Eucalyptol (C10H18O; cineol) is a terpene ether and provides a cooling, spicy taste. Eucalyptol may exist used in place of thymol in sure formulations in the aforementioned corporeality if desired. Menthol (CH3C6H9(CiiiH7)OH;
hexahydrothymol) also is only slightly soluble in booze, is fairly volatile, and in add-on to any antiseptic backdrop provides a cooling, tingling awareness.
The essential oils are used in amounts effective to provide antimicrobial activeness in the rima oris.
Generally, the total amount of essential oils present in a composition of this invention can be from about 0.001% to about 0.35% westward/five, with about 0.16% to about 0.28% w/v existence preferred. The compositions of the present invention preferably contain thymol and
additionally eucalyptol, menthol, or methyl salicylate, or mixtures thereof. Most preferably, the limerick contains all four of these essential oils. Thymol is preferably employed in amounts of from about 0.001% to about 0.25% w/5, and most preferably from almost 0.04% to about 0.07% due west/v. Eucalyptol may be employed in amounts of from about 0.001% to about 0.11% w/v, and preferably from about 0.085% to almost 0.10% w/5. Menthol is preferably employed in amounts of from near 0.001% to nearly 0.25% westward/five and well-nigh preferably from about 0.035 to about 0.05% w/five. Methyl salicylate is preferably employed in amounts of from about 0.001% to nearly 0.08% west/5, and almost preferably from about 0.04% to about 0.07% w/v. The carrier for the essential oils (i.due east., the active ingredients) is a h2o-alcohol mixture, generally water- ethanol. In the past, most antiseptic oral mouthwash compositions, such every bit Listerine®-brand mouthwashes, required high ethanol levels of up to nigh 27% v/5. These high levels were necessary to assist the actives in providing the necessary
antimicrobial functionality as well as providing a clear, aesthetically bonny liquid medium. Merely reducing the alcohol levels, without more than, results in a cloudy, less efficacious product.
Without being bound to whatever theory, it is believed that in these loftier alcohol level oral compositions, the booze solubilizes the antimicrobial actives and in so doing acts equally an active enhancement mechanism. The actives are more than readily dispersed throughout the solution and tin attack pathogenic microbes throughout the oral fissure. Reducing the alcohol levels was believed to adversely affect this enhancement
machinery.
In accordance with the present invention, however, information technology was surprisingly and unexpectedly found that the level of alcohol can be reduced without sacrificing
antimicrobial efficacy or clarity if the mouthwash limerick contains an increased corporeality of i or more polyols. By "reduced level" of alcohol is meant an amount of ethanol up to about 22% five/5, such every bit an amount of from virtually 9% v/v to about 22% v/v,
preferably no more about 19% v/v (due east.g., from about 12% to almost 19% v/v), and most preferably near 19% 5/v. Past "increased amount" of 1 or more than polyols is meant a total corporeality of one or more polyols, at least about 18% w/five, sufficient to heighten the antimicrobial activity of the essential oils from that activity which exists at a given reduced alcohol level to an activeness comparable to that exhibited by an clarified mouthwash having loftier booze levels on the order of most 27% v/v. A convenient reference point is the antimicrobial activity exhibited by a high alcohol level Listerine®- make mouthwash; i.e., i which contains well-nigh 27% v/v ethanol. Expressed in terms of an "R-Cistron, " which, as explained beneath, represents the time necessary to effectively impale typical oral cavity microbes in an in vitro biofilm, normalized against the kill time
exhibited in vitro by a 27% five/five Listerine®-brand mouthwash, the mouthwash compositions of the present invention exhibit an R-Factor of less than about two, preferably less than most i.2, and most preferably less than almost 1.0. The amount of polyol(s) employed in the mouthwash compositions of the present invention is direct and inversely proportional to the amount of ethanol in the carrier solution. As the concentration of ethanol is reduced beneath well-nigh 22% v/5, the amount of polyol(s) must exist correspondingly increased above about 18% due west/v to recapture efficacy lost by decreasing the booze level. What is about surprising is that the microbial kill rate of these reduced alcohol mouthwash
compositions is substantially increased at particular levels of polyol(s). The particular amount of
polyol(s) required will depend on a number of factors including the particular reduced ethanol level involved, the item polyol(s) employed, the antimicrobial efficacy required, the presence of other ingredients which may tend to improve efficacy, the effect of polyol add-on on other characteristics of the limerick such as consistency and clarity, etc. In general, the corporeality of whatsoever given polyol, and the total amount, will be selected to provide the desired R-Factor. By manner of example, for a reduced ethanol level of about 19% five/five, sorbitol used in amounts within the range of from well-nigh 25% w/v to nearly 30% w/v will provide a mouthwash limerick with essentially improved antimicrobial activity (i.e., R- Factors as low equally about 0.82). At 19% five/v ethanol, the amount of sorbitol preferably is within the range of from about 18% due west/v to about 30% w/v, most preferably from about 25% w/v to nearly xxx% west/v. Alternatively, at the same xix% five/v ethanol level, when propylene glycol is used as the polyol in amounts inside a range of from about 15% w/v to nigh 25% west/v, the resulting mouthwash
composition has mostly acceptable antimicrobial action (i.e., R-Factors as low every bit virtually 1.35).
The polyol(s) used in the present invention may be selected from those multi-hydroxy-functional compounds that are conventionally used in oral and ingestible products. Past mode of example, the polyol may be
selected from saccharide alcohols such as sorbitol, xylitol, mannitol, maltitol, inositol, allitol, altritol, dulcitol, galactitol, glucitol, hexitol, iditol, pentitol, ribitol, erythritol and mixtures thereof;
hydrogenated starch hydrolysates; (poly) alkylene glycols such equally propylene glycol and polyethylene glycols; diols such every bit ane,3-butanediol; glycerin; and mixtures of them or mixtures containing them. The preferred polyols are the saccharide alcohols, propylene glycol and glycerin. Sorbitol is the most preferred polyol. In general, the polyol(south) should be non- metabolized and non-fermentable polyol(southward).
The ratio of water to ethanol is in the range of from about three:1 to about 25:1, preferably from about 3.ii:one to about 10:1, past volume. The total corporeality of
ethanol/water mixture in a mouthwash preparation according to this invention depends, of course, on the amount of polyol(s) nowadays, but is typically in the range of from virtually sixty% to about 82%, preferably from about 70% to about 82%, by volume of the total
composition.
Surface agile agents (surfactants) may be used if desired in the compositions of the present invention. They are organic materials which aid in the complete dispersion of the ingredients throughout the solution as well as dispersing the training throughout the oral cavity. Preferably, the surfactant used in the compositions of the present invention is a non- ionic surfactant employed in an amount sufficient to help solubilize the actives. The preferred not-ionic surfactants are selected from the grouping known as poly(oxyethylene)-poly(oxypropylene) cake copolymers. Such copolymers are known commercially as poloxamers and are produced in a wide range of structures and molecular weights with varying contents of ethylene oxide. These non-ionic poloxamers are non-toxic and acceptable every bit direct food additives. They are stable and readily dispersible in aqueous systems and are uniform with a wide diversity of formulations and other ingredients for oral preparations. These surfactants should have an HLB (Hydrophilic-Lipophilic Residuum) of between well-nigh ten and about thirty and
preferably between about 10 and almost 25. By way of example, non- ionic surfactants useful in this invention include the following poloxamers:
More often than not these polymers when nowadays should be used in amounts of from well-nigh 0.01% west/v to almost iv.0% due west/v, and preferably from about 0.25% to about 0.75% w/v. A particularly preferred poloxamer is poloxamer 407 which is incorporated in an amount of about 0.5% w/five. The poloxamer or other surfactant is used to help solubilize the essential oils and flavor oils which may otherwise not exist soluble in these aqueous systems due to their reduced ethanol content. The surfactant(s) as well human action to disperse the actives and flavors throughout the solution and enable the compositions to provide a articulate, uniform appearance that is aesthetically more appealing.
In one apotheosis of the nowadays invention, three types of peppermint oil -- natural, Far W (redistilled, terpeneless) and Rose Mitchum -- are combined to provide a triple alloy. This unique alloy of season oils not merely provides a pleasant tasting mouthwash but also serves to gustation mask the bitter tasting essential oil actives discussed above. Each of these peppermint oils is present in substantially the same amount, from about 0.01% w/v to about i.0% w/v, and preferably in an amount of from about 0.2% w/v to nearly 0.3% w/v.
Combined, the triple blend is incorporated in the mouthwash composition in an amount of approximately 0.1% westward/v to almost 2.0 west/v, preferably in an corporeality of from nigh 0.five% w/v to about 0.9% w/v. The essential oil methyl salicylate not just provides antimicrobial action just, being a wintergreen season oil, also adds to the organoleptic flavour tones and complements the gustation masking part of the
peppermint oil blend.
Other flavor oils may besides be added to further modify or magnify the cooling minty taste of the peppermint, or reduce or mask the abrupt "bite" or "burn down" of
ingredients such as thymol and ethanol. Suitable flavors (and typical amounts) include oil of anise (about 0.01% w/five to about 0.2% due west/v) and benzyl alcohol (about 0.001% w/v to about 0.1% westward/v). In one
item embodiment of the present invention, the mouthwash is a articulate, light-green color and further includes spearmint oil (about 0.01% w/five to about 2.0% w/v) as an boosted flavorant. Other flavors such as citrus oils, vanillin and the like may be incorporated to provide further taste variations.
In some other embodiment, the flavorant anethole may exist employed in amounts upward to well-nigh 0.01% westward/v. Withal, it is specifically within the scope of the nowadays
invention to provide a mouthwash composition that is totally complimentary of anethole or at least substantially free of anethole (i.e., trace amounts or amounts which are so small as to not contribute functionally to the gustation or flavor properties of the composition). The particular flavor oils and other taste-improving ingredients employed volition vary depending upon the particular taste and experience desired. Those skilled in the art can select and customize these types of ingredients to provide the desired results.
Additional conventional components may be added as in mouthwashes of the prior art. Whereas some alcohol containing mouthwashes have a pH of about 7.0,
reduction of the alcohol level requires the improver of acidic preservatives, such as sorbic or benzoic acid, which reduce pH levels. Buffer systems are then necessary to control the pH of the limerick at optimal levels. This is by and large accomplished through the addition of a weak acid and its salt or a weak base and its salt. Useful systems accept been establish to be sodium benzoate and benzoic acrid in amounts of from about 0.01% to virtually four.0% w/5, and sodium citrate and citric acid in amounts of from about 0.001% to about 4.0% w/v. Preferably the buffers are incorporated in amounts that maintain the pH at levels of from most 3.5 to about half-dozen.5, and more preferably from about iv.0 to 5.0. Without beingness jump to any theory, it is believed that these pH levels provide the essential oils with an surround that also maximizes their antimicrobial action. Other conventional ingredients may be used in the mouthwash compositions of this invention, including those known and used in the art. For example,
humectants such equally polyethylene glycol may be added as an boosted solubilizer for the flavor oils and to also provide texture to the composition. These are incorporated in amounts of from about 0.3% w/v to about 0.6% w/v, and preferably about 0.5% westward/v. Softeners such as glycerin may be added to heighten the lubricous mouthfeel of the mouthwash as it is used and to provide a refreshing, moist, organoleptic feeling thereafter. Glycerin (when non used as the polyol, as discussed higher up) may be incorporated in amounts of from about 1.0% w/five to most 10.0% w/v, and preferably in an amount of about 7.v% westward/5. Sweeteners such as aspartame or sodium saccharin and the like may be added for better taste in amounts of from about 0.005% w/v to well-nigh 1.0% w/v, and preferably in an corporeality of
approximately 0.05% w/v.
Zinc chloride may exist added as an astringent for an "antiseptic cleaning" feeling in an amount of from near 0.0025% w/five to about 0.0075% w/v. And although the mouthwash formulations of the present invention may be formulated to be essentially clear and colorless, acceptably canonical nutrient dyes are preferably used to provide a pleasing color to the formulations of the invention. These may be selected from the long listing of acceptable nutrient dyes and in detail may exist
incorporated to provide the spearmint green conception discussed infra. Suitable dyes for this purpose include FD&C yellowish #5, FD&C yellow #x and FD&C green #3. These are added in conventional amounts, typically in individual amounts of from about 0.0003% w/v to virtually 0.0005% westward/v, and preferably from about 0.000035% w/v to nigh 0.00045% w/five.
Water is added to q.s. and the formulation may and then be bottled and packaged for aircraft. The mouthwash limerick of the present invention may also be formulated, if desired, equally gels, foams, pastes,
aerosols or tablets using standard formulations known in the fine art as advisable.
Alternatively, the mouthwash compositions of the present invention may be formulated in a dry out powder, chewing gum, semi-solid, solid or liquid concentrate form. In such embodiments, for example, water is added to q.south. the volume to the necessary total for a liquid concentrate, or water may be removed using standard evaporation procedures known in the art to ready a composition in dry out powder class. Both may and then have water added at a subsequently date when fix for use. Such forms are advantageous for storage and shipping. R- Factor
Biofilms of the microorganism Streptococcus mutans
(ATCC #25175) grown on stainless steel wires simulate thick, semipermeable dental plaque. For purposes of the present invention, an "R-Factor" is a convenient measure of the antimicrobial efficacy of the mouthwash compositions of the present invention, as measured past their ability to kill those biofilms. The R-Factor is defined equally the ratio of (i) the time, in minutes, necessary for a mouthwash composition to impale S. mutans microorganism biofilms grown in vitro on stainless steel wires, to (ii) the time, in minutes, necessary for a standard high alcohol mouthwash limerick to kill like biofilms of the same microorganism grown in vitro on other, identical stainless steel wires. Those kill times are obtained past a plaque penetration assay developed by the present inventors. Plaque Penetration Assay
The plaque penetration analysis employed by the nowadays inventors to obtain their biofilm critical impale times is a modification of the well-known process of
Tanzer, et al., described or referenced in, e.g.,
Tanzer, et al., "Structural requirements of Guanide, Biguanide and Bisbiguanide Agents for Antiplaque
Activity," Antimicrobial Agents and Chemotherapy, Dec. 1977, pp. 721-729; and Tanzer, et al., "In Vitro
Evaluation of Seven Cationic Detergents as Antiplaque
Agents," Antimicrobial Agents and Chemotherapy, Mar. 1979, pp. 408-414.
The loftier alcohol mouthwash composition employed by the present inventors as their standard for their plaque penetration assay contains 27% v/five ethanol and has the composition shown in the post-obit Table i:
The media required for the plaque penetration analysis include sterile deionized water; Letheen Goop (DIFCO); and Jordan's complex medium (with and without
bromocresol regal pH indicator) [see Jordan, H.5. et al. J. Dent. Res. 39: 116-123 (1960)]. The equipment requirements for the assay include a large number of sterile drinking glass test tubes (east.g., 13 ten 100 millimeters); examination tube racks to hold multiple rows of sample tubes; an autoclave; and stainless steel Nichrome wires (1.5 10 85 mm). It is convenient to attach each plaque wire to a Morton Closure by whatsoever suitable means, such as
welding.
Jordan's medium may be prepared past blending the
following ingredients with heating as necessary:
The pH is adjusted to seven.1 with 5N HCl followed by cooling to room temperature, if necessary. l ml is removed from the batch and placed in a 100 milliter flask. Jordan'southward medium with pH indicator ("recovery medium") is prepared by adding 1 ml of a 1% bromocresol royal stock solution (i.eastward., 0.one gram bromocresol royal in ten mis distilled water) to 1 liter of
Jordan'due south medium.
The culture for the assay is prepared as follows. Upon receipt, the ATCC civilization is rehydrated and subcultured co-ordinate to the directions supplied by the American Blazon Culture Collection. The subculture is streaked for purity on Brain-Heart Infusion Agar (DIFCO) and inoculated into 100 ml of sterile BHI. The agar plates are examined for purity later on 14-18 hours. If
acceptable, xi ml of sterile glycerin are added to the BHI civilisation, vortexed and then subdivided into 1.8 ml cryogenic tubes. The cultures are then stored at - lxxx°C. Four days prior to an assay, a frozen vial is thawed and added to the pocket-sized, 100 ml flask containing l ml of Jordan'due south medium to start the cultures used for the biofilm assay. After fourteen-18 hours, the contents of the small flask are decanted aseptically into ii liters of Jordan's medium. The resulting inoculated medium is then aseptically dispensed, in five.0 ml portions, into a number of empty sterile test tubes, each tube having a plaque wire-equipped Morton cap. The inoculated tubes are then incubated anaerobically overnight (i.eastward., 14-xvi hours) at 37°C. The number of exam tubes will vary depending upon the number of unlike mouthwash samples being tested, but it will be convenient to describe an assay of a
standard high alcohol mouthwash and iv reduced alcohol mouthwash samples, which requires racks each holding 75 test tubes (i.east. , five rows of fifteen tubes, each row comprising five sets of 3 tubes each). The first set of three tubes in each row is usually reserved for the standard loftier alcohol
mouthwash, with the succeeding four sets of three tubes each in that row being reserved for the four reduced alcohol samples. In that location are three tubes in each gear up considering each assay is performed in triplicate.
After the overnight incubation, the plaque wires are and so transferred into fresh Jordan's medium in 75 tubes (in a 2d rack) and once more incubated anaerobically for 24 hours at 37°C. This procedure is repeated one time more. Thus, the plaque wires are cultured for 3 days, with two transfers subsequently initial inoculation.
On the third twenty-four hour period, only prior to the analysis, 5
additional racks (each containing 75 sterile test tubes) are prepared: a first (assay) rack whose test tubes each incorporate half-dozen ml of the sample mouthwashes; a second (water) rack whose test tubes each contain half-dozen ml of sterile deionized water; tertiary and quaternary racks whose exam tubes each contain 6 ml of Letheen broth rinse; and a fifth rack whose exam tubes each comprise v ml of Jordan's recovery medium. For convenience, the racks may be marked for test series identity and time (past row). Assay Procedure: Each assay volition involve triplicate testing at time points separated by 1 minute
intervals, east.chiliad., at 2, 3, iv, 5 and half-dozen minutes of
mouthwash handling. The first (or bottom) row of each rack corresponds to the first test time and the
succeeding four rows correspond, respectively, to the next iv test times. The exact time of exposure of the plaque wires to the sample mouthwashes tin exist varied according to the thickness of the "plaques;" ideally, the exposure period volition effect in positive microorganism growth in the commencement one or two sampling intervals of the high booze control grouping, (i.due east., the first and 2d row) and no growth thereafter.
Establishing lower and upper limits of exposure
required for complete kill by the control mouthwash permits an accurate comparing of the 4 sample mouthwashes to this control. Mouthwash exposure takes identify in a 37° New Brunswick shaking water bath (milkshake speed 3) and may exist staggered so that the 5 time points are run meantime, but with sufficient time to permit accurate timing and handling.
(ane) To start assay, transfer i row of plaque wires to the first (bottom) row of tubes in the rack containing half dozen ml of sterile h2o. Leave in place 2 minutes. Repeat for the next iv rows of plaque wires.
(2) After the water rinse, transfer each row of plaque wires into the appropriate, corresponding, row of tubes in the rack containing 6 ml of test
mouthwash. Get out each row of plaque wires in place, with shaking in the 37° water bathroom, for its treatment (exposure) menstruum; i.e., remove the rows of plaque wires sequentially at 5 preset time points and then that each succeeding row of plaque wires is exposed to a mouthwash for successively longer periods of time (e.g.. two, three, 4, 5 and 6 minutes; individual timing can vary co-ordinate to estimated "plaque" thickness. (iii) For each row of plaque wires, at the end of its treatment period, immediately remove the row and identify it in the advisable corresponding row of the first rack of vi ml Letheen Broth
neutralization/rinse tubes. Exit each row of wires in that broth for five minutes and then
transfer it to the advisable corresponding row of the second rack of 6 ml Letheen Broth rinse tubes. (4) At the stop of the second Letheen Goop rinse,
remove each row of plaque wires and place information technology in the appropriate respective row of the rack of half dozen ml Jordan's recovery medium (with bromocresol purple). Incubate anaerobically for 48 hours at 37°C.
(five) Read for growth (+) or no growth (-) at 48 hours.
Positive growth is indicated by a color change from imperial to yellow (i.e., if the microorganism is notwithstanding viable, information technology volition produce an acrid which causes the color change); positive growth is often accompanied past an increase in broth turbidity.
Determination of Critical Impale Times and R-Factor:
Since each mouthwash sample is located in the aforementioned ready of iii tubes in each row of the rack, the critical time necessary for the sample to completely kill the microorganism tin be determined by observing the point (front to back or bottom to pinnacle, equally the case may be) at which the Jordan recovery medium color inverse from xanthous to royal. The critical kill fourth dimension for whatever sample, divided by the critical kill time for the control mouthwash in that same rack, gives the R-Factor for that sample.
Tabular array 2 beneath summarizes a statistical scale developed by the nowadays inventors which relates the observed alter from growth (+) to no growth (-) to critical impale times. For case, as shown in the first row of Table 2, where the observed status changes from growth (continuous +'s) to no growth (continuous -'due south) ("no anomaly"), the critical kill time is adamant by adding 0.50 infinitesimal to the fourth dimension at which the final growth ascertainment (+) was made. The balance of Tabular array two sets along how critical impale times are determined for different observed growth/no growth intervals betwixt continuous growth segments and continuous no growth segments.
Past mode of further case, consider the examples of growth/no growth sequences, and their associated critical impale times, in Table 3. In the beginning row of Table 3, there was no anomaly between continuous +'s and continuous -'s; therefore, CKT (per Table two) =
4.0 + 0.5 = 4.5 minutes (i.east., kill occurred somewhere between 4.0 and five.0 minutes). In the second row of Table 3, the interval between continuous +'s and continuous -'due south is -+; therefore, CKT (per Table ii) = 2.0 + 1.v = iii.5 minutes.
In the case of rows 3 and 4 of Tabular array 3, clearly no finish betoken was reached. It is assumed hither that impale volition occur at some point in backlog of 6.5 minutes (>vi.5) or much beneath 2.0 minutes (<two.0), respectively.
Row 5 (Table three) is an case where the kill scale is dependent on the values which are located to the left of the final + and to the correct of the first -. For that particular case, CKT = two.0 + one.one = 3.1 minutes (per Table two).
The following examples are provided both to illustrate the present invention and its advantages and to provide additional comparing data.
Case 1
A mouthwash composition containing 19% 5/v ethanol and xviii% west/five sorbitol was prepared as follows. First, an alcohol solution of active essential oils and season oils was prepared by dissolving the ingredients listed in Tabular array 4 in 200 mls of ethanol (USP):
To the resulting alcohol solution were then added the additional ingredients shown in Table five (along with suitable dyes to obtain the desired colour):
The pH was adjusted to 4.2. Approximately 600 mls of deionized water was then added, and the solution was mixed until all solids were dissolved. Additional deionized water was added with mixing to bring the full solution to one liter. A clear mouthwash solution was obtained.
Case 2 A number of different mouthwash compositions A-J were prepared. The composition ingredients and amounts used (except for the dyes) are shown in Tabular array 6 beneath.
The mouthwashes were prepared every bit follows. An alcohol solution was prepared by dissolving the flavor oils, part of the thymol and part of the methyl salicylate in 227 or 200 milliters of ethanol (USP). Into the resulting solution was and then dissolved the remaining methyl salicylate, the remaining thymol, the benzoic acid and the Pluronic surfactant. 300 mis of the deionized water was then slowly added to the resulting solution, with mixing. The sorbitol (USP) solution, or propylene glycol, was and then added, with continued mixing, followed by the sodium saccharin, the sodium citrate and the citric acid, with mixing until
dissolved. The dyes were and then added, with mixing, followed by the balance of the deionized h2o.
Additional sodium citrate or citric acid was added, if necessary, to adjust the pH of the resulting clear solution within the range of from about iii.ix to most 4.five, preferably four.two. Each of the mouthwash compositions of Table six was evaluated in vitro for its power to impale
microorganisms in a biofilm, using the protocol
described above. The results are shown beneath in Tabular array 7, which as well shows the percentages of ethanol and polyol in those compositions.
The above data support the inverse relationship between the ethanol and polyol amounts. Thus, with lower amounts of ethanol, i should employ even higher concentrations of polyol(s) above 18% w/v to maintain similar efficacy. Example 3
The mouthwash formulations shown in Table eight beneath, containing 21.half dozen% v/five ethanol and a varying amount of sorbitol, were prepared in a fashion similar to the procedure shown in Example 2, and tested for their antimicrobial efficacy as measured past the amount of endotoxin extracted from in vitro cultures of three dissimilar Gram negative leaner known to be plaque- associated microorganisms, Fusobacterium nucleatum
(ATCC #10953), Veillonella parvula (ATCC #10790), and Actinobacillus actinomycetemcomitans (ATCC #33384):
The same proprietary alloy of flavour oils shown in Tabular array vi was used.
Endotoxin is a component of the outer wall of Gram negative bacteria and if shed by the leaner at or near infection sites most the teeth and gums will elicit inflammation and gingivitis. Removal of
endotoxin from the plaque through employ of a mouthwash composition will consequently reduce these problems.
The cultures were treated with successive dilutions of the mouthwash limerick of Table eight with no sorbitol versus those with sorbitol (17.5% w/v), equally the data reflect in Table 9. Plus (+) signs indicate the presence of endotoxin while negative (-) signs signal its absence. Clearly, the presence of sorbitol
enhances the removal of endotoxin. The cultures were also treated with successive
dilutions of the mouthwash composition of Table 8 with varying sorbitol concentrations. The results are shown in Tables 10 and 11 beneath. In both tables, the
"Control" mouthwash was the loftier alcohol (27% v/v) standard described higher up.
Every bit tables 9-11 clearly bear witness, the addition of sorbitol clearly enhances the endotoxin removal ability of the mouthwash composition. A greater degree of endotoxin is extracted from cultures treated with antispectic mouthwash compositions containing sorbitol than those that practise not. Moreover, as Tables 10 and 11
demonstrate, increasing sorbitol levels in the
mouthwash clearly decrease the degree of endotoxin remaining in the culture, supporting its value in preventing inflammation and subsequent gum illness.
Even so, endotoxin measurements are at best a semi- quantitative prescreen, and therefore a more stringent method for assessing plaque and bacterial kill efficacy -- known every bit the plaque penetration analysis (described above) -- was employed in this invention. The results of this assay, reported as "Critical Impale Times" (or "CKT"), are more predictive of the in vivo effect of oral mouthrinses than endotoxin measurements.
Is Mouthwash Heterogeneous Or Homogeneous,
Source: https://patents.google.com/patent/WO1997030685A1/en
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