The present invention relates to a two-part antimicrobial composition comprising at least one chlorine dioxide generating component comprising at least one metal chlorite and at least one acid-forming compound in a solid carrier, and at least one liquid aqueous component. The composition further comprises at least one antimicrobial fatty acid having from about 2 to about 15 carbon atoms, and preferably from about 6 to about 12 carbon atoms. The components, upon mixing, form a composition having a pH in the range of about 5 to about 10.

The present invention relates to a two-part antimicrobial composition comprising at least one chlorine dioxide generating component comprising at least one metal chlorite and at least one acid-forming compound in a solid carrier, and at least one liquid aqueous component. The composition further comprises at least one antimicrobial fatty acid having from about 2 to about 15 carbon atoms, and preferably from about 6 to about 12 carbon atoms. The components, upon mixing, form a composition having a pH in the range of about 5 to about 10, and preferably from about 5.2 to about 8.
Bacterial infection, particularly bovine mastitis, is the most costly and difficult problem that a dairy herdsman will typically have to deal with.
Mastitis is an inflammation of the mammary gland that occurs primarily as a result of a bacterial infection that gains entry into the udder of the mammal via the teat canal. Mastitis decreases the cow""s milk production and reduces the quality of the milk. In more severe cases of mastitis, the milk will have to be discarded, and if the disease resists treatment, the cow may have to be destroyed. A further problem is that a mastitis infection can spread from cow to cow.
Mastitis can occur in two forms that are referred to as clinical and subclinical mastitis. Both decrease milk production, but the milk must be discarded if the cow has clinical mastitis, and in untreatable cases, as stated above, the cow may have to be destroyed. In the case of the clinical disease, attempts at treatment have involved infusion of an antibiotic into the udder of the animal.
There are numerous organisms that cause mastitis, but the most common are streptococcus agalactiae and staphylococcus aureus. Other environmental and contagious microorganisms that cause this disease include coliforms, Klebsiella pneumoniae, Actinomyces pyogenes, Corynebacterium bovis, Listeria monocytogenes, Pseudomonas aeruginosa, Mycoplasma bovis, as well as other species of staph and strep. Treatment is generally started before the species causing the disease is identified, and it is therefore desirable to have an antibiotic that has the greatest efficacy, and that acts on as many species that cause the disease as possible. These microorganisms lurking in the surrounding environment such as plants and bedding (i.e. straw, hay, grass, wood shavings and the like), feces, soil, water, feed, and so forth are opportunistic and are spread from cow to cow during the milking process. The entrance of the microorganism is through the teat orifice and teat canal of the animal.
Dairy herd management focuses on both the treatment and prevention of mastitis infections. As stated above, treatment typically involves infusion of antibiotics into the udder of the infected cow(s), and prevention involves routine hygienic practices. Routine hygienic practices used for the prevention of mastitis (as well as preventing spreading of the disease) involves treating (i.e. dipping) the teats of the udder of a cow immediately after each milking period with an antiseptic xe2x80x9cteat dip.xe2x80x9d This has become a standard practice for dairy herdsmen as it has been proven to be a very effective way to prevent intramammary mastitis in cows.
Teat dips may be divided into several general categories including xe2x80x9cpredipxe2x80x9d teat dips that are used to clean the udder of the cow prior to milking, xe2x80x9cprepostxe2x80x9d teat dips that are applied to the udder of the cow after milking, and xe2x80x9cbarrierxe2x80x9d teat dips that are also applied to the udder after milking and are further designed to remain on the udder of the cow in between the milking periods.
Mastitis causing microorganisms are found in both the barn in which the cow is milked and in the environment the cow enters in between the milking periods. Prepost teat dips are designed to work on those microorganisms that are found in the barn, and that are spread from cow to cow during the milking period.
The barrier teat dips are designed to remain on the cow during the nonmilking periods. Barrier teat dips therefore comprise a film-forming agent that allows the teat dip to form a barrier or phophylaxis over the teats of the cow in order to prevent entrance of microorganisms through the teat orifice from the surrounding environment in the nonmilking periods. These barrier teat dips may or may not contain an antimicrobial agent, although in recent years it has become increasingly popular that the barrier teat dips have both film-forming characteristics and antimicrobial activity because some of the mastitis causing microorganisms have been known to penetrate the barrier and migrate into the teat canal.
There now exists a variety of commercially available predip, prepost and barrier teat dips, including those based on iodophors (i.e. iodine), quaternary ammonium compounds, chlorhexidine salts, chlorine release compounds such as alkali hypochlorites; oxidizing compounds such as hydrogen peroxide and peracids; protonated carboxylic acids (i.e. fatty acids) such as heptanoic, octanoic, nonanoic, decanoic, undecanoic, and so forth; acid anionics such as alkylaryl sulfonic acids; chlorine dioxide; and so forth. Such compositions have varying degrees of effectiveness.
Chlorine dioxide has been found to be an extremely effective chemical for use as an antimicrobial and over several decades has found increasing popularity for disinfecting, sanitizing, sterilizing, deodorizing, and so forth. Further, this chemical has been found to be quite useful in the prevention and treatment of bovine mastitis. This compound is typically generated in solution (i.e. aqueous medium) by mixing a metal chlorite salt with a strong mineral acid or a large amount of a strong organic acid. The stability of such solutions is an issue, however, and mixing of the acid and the metal chlorite salt is not recommended until just prior to use.
A variety of approaches have been taken for using chlorine dioxide. Such approaches are found in U.S. Pat. No. 4,084,747 and its reissue Re. Pat. No. 31,779; U.S. Pat. Nos. 4,585,482; 4,986,990 and 5,185,161; 4,891,216 and 5,651,977; and 5,820,822.
Various problems have been encountered with teat dips that utilize chlorine dioxide generating components. One such problem has been the low pH typically required for sufficient generation of chlorine dioxide. For antimicrobial compositions used on either human or mammalian skin, an acidic pH is of concern due to the irritation caused to tissue. Another problem associated with chlorine dioxide generation is the shelf life. Once chlorine dioxide generation is initiated, the compositions begin to lose efficacy and must typically be used in a matter of days
There remains a need in the art for a simple, yet effective and long-lasting antimicrobial system based on chlorine dioxide, and that may be used for such applications as the prevention and treatment of mastitis.
The present invention relates to a novel and improved two-part antimicrobial composition having longer lasting efficacy against microbes, and is useful as a teat dip comprising at least one chlorine dioxide generating component in a solid carrier and at least one liquid aqueous component. The chlorine dioxide generating component has at least one metal chlorite and at least one acid-forming compound. The acid-forming compound is preferably a protic acid, but includes any type of acid-forming compound. It is necessary that the acid-forming compound have a pKa of less than that of the fatty acid. Chlorine dioxide will not be adequately generated at a pH of about 5-10.
The antimicrobial composition further comprises at least one antimicrobial fatty acid which may be supplied in the solid component, but is preferably supplied in solution in the aqueous component. The fatty acid is characterized as having from about 2 to about 15 carbon atoms, and preferably from about 6 to about 12 carbon atoms. The fatty acid has a pKa of about 5.
The solid component is permeable to water in liquid form, or is sensitive to or dissolvable in water in liquid form. Upon mixing of the components, chlorine dioxide is generated.
The resultant mixed composition has a pH in the range of about 5 to about 10, preferably from about 5 to about 8, and most preferably from about 5.5 to about 8.
The composition has improved antimicrobial effectiveness in that the chlorine dioxide acts immediately to kill microorganisms that cause mastitis, while the fatty acid remains antimicrobially active over a longer period against the mastitis causing microorganisms.
Furthermore, the composition is effectively antimicrobial at a relatively neutral pH of about 5 to about 10, preferably about 5.5 to about 10, and most preferably from about 5.5 to about 8, which results in less problems with irritation to the skin.
The compositions of the present invention may be utilized as predip and prepost teat dips, as well as barrier teat dips upon the addition of a film-forming agent.
The compositions of the present invention further have the benefit that they are simple and easy to use.
The present invention relates to a two-part antimicrobial composition. The first part comprises at least one acid-forming compound and at least one metal chlorite in a solid form, and the second part is an aqueous based solution. The first part may be referred to as the chlorine dioxide generating component. A fatty acid is further provided in the composition in either the first part as a solid, or in the second part as a solution. Preferably, the antimicrobial fatty acid is provided in aqueous solution.
The addition of the fatty acid to the resultant composition, provides the composition with residual, longer-lasting antimicrobial activity. The two parts are blended prior to use, and are particularly useful as a teat dip composition. The first component generates chlorine dioxide, an effective antimicrobial agent, in the presence of the second, aqueous based part.
The chlorine dioxide generating compounds useful herein include metal chlorites. Preferred metal chlorites are water soluble and include alkaline earth metal chlorites and alkali metal chlorites such as potassium chlorite and sodium chlorite. Alkaline earth metal chlorites include barium chlorite, calcium chlorite and magnesium chlorite. Sodium chlorite is a particularly preferred compound. The metal chlorite is preferably provided in a powder (i.e. salt) form. The metal chlorite is preferably present in the resultant antimicrobial formulation at a concentration of about 0.01% to about 20% by weight, and preferably from about 1.0% to about 14%. However, the amount of metal chlorite selected is dependent on the amount of chlorine dioxide generation that is desired.
Consequently, the metal chlorite is added at a concentration sufficient to generate from about 1 to about 500 ppm chlorine dioxide, preferably from about 20 to about 500 ppm chlorine dioxide, and most preferably from about 100 to about 400 ppm chlorine dioxide. For a teat dip, the amount of chlorine dioxide present depends on the amount of organic soil present at the time of use (10% milk will raise the amount of chlorine dioxide required substantially because protein reacts rapidly with ClO2). As an example, in the absence of organic soil, as little as a few ppm of chlorine dioxide is required for an effective microbial kill. However, in the presence of organic soil, an initial amount of about 200 to about 500 ppm of chlorine dioxide is needed, and preferably from about 250 to about 400 ppm of chlorine dioxide.
For hard surface sanitizing applications, the soil demand is not as high and the amount of chlorine dioxide required is substantially less. For example, the amount of chlorine dioxide required may be about 1 to about 50 ppm and preferably about 2 to about 20 ppm.
The acid-forming compounds useful herein include water sensitive or soluble compounds, as well as substantially water-insoluble acid-forming compounds. The acid-forming material is preferably a solid material, and is preferably hydrophilic. The acid-forming compound does not react to any substantial degree with the metal chlorite in the absence of water. The acid-forming compound may itself be protic, or it may produce protons upon contact with water and the metal chlorite. The pKa of the acid-forming compound is less than that of the antimicrobial fatty acid. Antimicrobial fatty acids have a pKa of about 5, typically they are about 4.8. The acid-forming compound consequently has a pKa of less than 4.8, and is preferably 4.0 or less, more preferably about 3.0 or less and even more preferably about 2.0 or less. It is necessary that the acid-forming compound form a localized low pH. The generation of chlorine dioxide will not be adequate at a pH that is close to neutral, or is basic. The pH required in order to produce chlorine dioxide is lower and consequently, the pH of the solution at the point of reaction between the metal chlorite and the acid-forming compound is lower than that of the solution generally.
The acid-forming compounds useful to the present invention are vast in number and include protic acids, non-protic acids, acidic salts, mineral acids, as well as other types of acid-forming compounds, some discussed in more detail below. A primary requirement is that the acid-forming compound be able to generate protons in water.
The acid-forming compounds useful herein include protic acids. Some useful protic acids include carboxylic acids and dicarboxylic acids such as aromatic carboxylic acids, xcex1-hydroxy carboxylic acids, xcex2-hydroxy carboxylic acids, unsaturated carboxylic acids, and so on and so forth. Some specific acids useful herein include but are not limited to benzoic, boric, citric, fumaric, oxalic, tartaric acid, lactic acid, maleic acid, and so forth. The acid-forming compound may also be the anhydride of these protic acids, for instance, maleic anhydride. Preferably, the protic acids are used in the present invention in amounts of about 0.1% to about 5%, and preferably about 0.1% to about 1% by weight of the total antimicrobial composition.
Other acid-forming compounds include water soluble acid salts of calcium, magnesium, barium, lithium, aluminum, sodium, potassium, and so forth. Such compounds include sulfuric and phosphoric acid salts, calcium chloride, magnesium chloride, magnesium nitrate, lithium chloride, magnesium sulfate, aluminum sulfate, sodium acid sulfate, sodium dihydrogen phosphate, potassium acid sulfate, potassium dihydrogen phosphate, and so forth, and mixtures thereof.
Useful mineral acids include sulfuric, hydrochloric, nitric and sulfamic acids.
Other acid-forming compounds include, but are not limited to, acids which are already included in a solid carrier such as synthetic molecular sieves including synthetic zeolite A, X and Y, mordenite and ZSM-5 which have both been converted to the substantially hydrogen ion (H+) exchanged forms; natural zeolites such as chabazite and clinoptilolite; dealuminated zeolite Y; acid ion exchange resins such as Dowex(copyright) HCR-S resins available from the Dow Chemical Co. in Midland, Mich.; insoluble organic acid anhydrides such as phthalic anhydride; and so forth; and mixtures thereof.
Other acid-forming compounds are the clays including acid treated, calcined and hydrous clays, and so forth. Hydrous clays include bentonite, kaolin, attapulgite and halloysite. Calcined clays include spinel phase calcined kaolin, calcined bentonite, calcined halloysite and calcined attapulgite. Acid treated clays are such that the clays are contacted with acidic solutions containing sulfuric acid, hydrochloric acid, nitric acid or other acidic compounds such as lanthanum chloride and include acid treated metalkaolin, acid treated bentonite, acid treated attapulgite and acid treated halloysite.
Typically, the zeolites or clays are treated with acids such as sulfuric, nitric and hydrochloric acids.
Acid-forming compounds that find utility in the present invention are described in detail in WO 98/38865 and in WO 99/24356 both incorporated in their entirety by reference herein. The above list of acid-forming compounds is intended as illustrative, and not as an exclusive list of all the acid-forming compounds useful herein, and as such is not intended as a limitation on the scope of the present invention. One of skill in the art would know how to select such an acid-forming compound.
The amount of the acid-forming compound will depend on how much metal chlorite is present in the compositions. It is desirable to have an excess of acid-forming compound present. Higher rates of reaction require higher concentrations of acid-forming compounds. The concentration of acid required will also depend on the pKa and reactivity of the acid, however.
The amounts of the metal chlorite and the acid-forming component are selected so as to produce a quantity of chlorine dioxide that is sufficient for effective antimicrobial efficacy. Generally, the amount of chlorine dioxide generated in the resultant aqueous solution of the present invention will be from about 0.5 ppm to about 500 ppm, preferably from about 50 to about 450 ppm, and most preferably from about 100 to about 400 ppm.
The metal chlorite and the acid-forming compound of the present invention are included with a solid carrier. The solid carrier is a neutral compound, and may, for instance, be any clay of the aluminum silicate type including bentonite, kaolin, attapulgite, halloysite, polyorganosilicates, and so forth; micas; talcs; zeolites; and so forth.
The acid-forming compound may be found in either a solid or in a liquid form depending on the acid-forming compound utilized. If the acid is a solid, then the metal chlorite and the acid may be mixed together with the solid carrier at one time, for instance clay or zeolite, and supplied preferably in a pressed tablet form. In this instance, the solid carrier is utilized as a diluent in order to prevent the concentration of the metal chlorite and the acid from being undesirably high. At high concentrations the mix can become explosive.
Alternatively, the metal chlorite may be mixed with a solid carrier in solution, and then dried. Separately, the acid may be mixed with a solid carrier and then the subsequently, the two, now carried reactants, may be mixed together.
If, on the other hand, the acid is a liquid, it must be first mixed with a solid carrier so that the acid is now in solid form. In this instance, the metal chlorite, a solid itself, may be added to the acid/carrier mix, or the metal chlorite may be first mixed with a solid carrier itself, and then the metal chlorite/carrier component is mixed with the acid/carrier component. If the metal chlorite is first mixed with a carrier, it is done in solution and then dried prior to mixing with the acid/carrier mix.
Optionally, the fatty acid antimicrobial may be added to the metal chlorite and/or acid-forming compound at any point of the process.
The resultant mix may then be supplied to the end user in any solid form including powders, agglomerates, tablets, pellets, granules, capsules, and so forth. A preferable solid form is a tablet form. The reactants, i.e. the metal chlorite and the acid forming compound react in the presence of water so it is necessary to further package them in some form that will keep them substantially free from the presence of water, either in vapor form, or in liquid form. Otherwise, chlorine dioxide may be prematurely generated. Chlorine dioxide generation may also occur in the presence of water vapor, i.e. that absorbed from the ambient air. The reactants are therefore preferably packaged, transported and stored in a substantially moisture-proof package.
The packaging materials may be insoluble or soluble. Examples of insoluble materials include thermoplastic polymeric materials such as nonwoven polyethylene such as Tyvek(copyright) available from DuPont in Midland, Mich. and expanded polytetrafluoroethylene such as GoreTex(copyright) available from W. L. Gore.
Permeable membranes may be soluble, dissolvable or permeable and include gelatins, perforated materials such as perforated Tyvek(copyright), and so forth.
Some packaging options include membranes, films, sachets, bags, pouches, capsules and so forth, that keeps the tablet free from moisture, but which degrades or dissolves in water, or is permeable to water in liquid form. These packages will allow for the controlled release of chlorine dioxide. In the case of a package that is permeable to water in liquid form, the package must be permeable to chlorine dioxide, and thereby finction by allowing water in, and chlorine dioxide out. Chlorine dioxide generation can be increased by increasing the water solubility of the package, or by increasing the water permeability of the package in which the solid is placed.
The use of membranes which allow the passage of water in liquid and/or vapor to pass into the membrane to the solid reactants from a liquid body of water and subsequently chlorine dioxide out of the membrane is discussed in WO 99/24356 herein incorporated by reference in its entirety.
The tablets may be removed from the package, and the package discarded. Tablets may also be packaged in composite type materials such as a polyolefin film over an aluminum foil.
In one specific embodiment of the present invention, the metal chlorite and the acid-forming compound are in a solid mixture that has been pressed into a tablet form. Optionally, the fatty acid antimicrobial agent may be added to the pressed powder tablet. The tablet is then packaged in a moisture proof barrier. The moisture-proof barrier may be comprised of any substantially water-insoluble thermoplastic material. Each tablet may be individually packaged, the package discarded once the tablet is removed, or several tablets may be packaged in a resealable container, such as those similar to a Zip-loc(copyright), that will provide a moisture proof barrier. The tablet(s) is then added to a specified amount of the second part, i.e. the aqueous solution.
In another embodiment, the metal chlorite and the acid-forming compound are in a solid powder form and are supplied in a solid carrier such as a sachet, pouch, membrane, and so forth that is permeable to water in liquid form. If a sachet, pouch, or membrane is used, the metal chlorite and the acid-forming compound are separated from the rest of the components by the sachet, pound or membrane. In this embodiment, the solid carrier must also allow the passage of chlorine dioxide into the aqueous medium upon its generation. Further to this embodiment, the solid carrier may be divided into chambers that separate the acid-forming compound and the metal chlorite from one another to prevent a reaction should water in vapor form enter the solid carrier.
Optionally, the tablets or the powder, may be provided in a package or membrane that dissolves upon the addition to an aqueous medium, thereby allowing liquid water to penetrate and come into contact with its ingredients.
One example of a membrane material is gelatin. Gelatin is supplied in the form of a capsule under the tradename of Capsugel(copyright), available from Warner Lambert in Morris Plains, N.J.
As stated above, the pH of the solution is about 5-10, preferably about 5.2 to about 10, preferably about 5.2 to about 8, and most preferably about 5.5 to about 8. Chlorine dioxide may not be adequately generated at such high pH""s, however. It is therefore necessary that the acid-forming compound and the metal chlorite be in close proximity to one another, and that the pH at the local of the metal chlorite be lower than that of the solution generally. The pKa of the acid-forming compound is therefore quite low, i.e. 4.5 or less.
The present invention provides for the controlled release of chlorine dioxide by employing solid reactants that dissolve readily upon contact with water. The solid, e.g. a tablet, dissolves in the aqueous solution, the metal chlorite and the acid-forming compound react and chlorine dioxide is released. Close proximity of the reactants is desirable in order to effectively release chlorine dioxide at such a high pH.
These and other embodiments are envisioned by the present inventors. Furthermore, one of skill in the art will understand that combinations of these embodiments will also provide a controlled release of chlorine dioxide by preventing water actuation of the composition until it is so desired, as well as how quickly water will contact the reactants. Controlled chlorine dioxide generation is discussed in detail in WO 99/24356 incorporated by reference herein in its entirety.
Using the methodology of the present invention, chlorine dioxide will typically be generated over a period of about 15-30 minutes with some agitation. However, if the agitation is too great, the efficiency of the reaction can be greatly decreased, and not all of the metal chlorite may be converted to chlorine dioxide. The reason being, a localized lower pH is required at the point of reaction between the metal chlorite and the acid-forming compound. Too much agitation can lead to dispersion of the acid-forming compound and therefore a higher pH at the reaction site. Agitation will therefore not be necessary throughout the whole 15-30 minute period.
Chlorine dioxide provides an immediate and quite effective antimicrobial action against those contagious mastitis causing pathogens found on the teat when the composition of the present invention is applied. Chlorine dioxide rapidly vaporizes from the surface of the teat, however, and therefore does not provide long-lasting antimicrobial activity. Furthermore, should chlorine dioxide remain on the teat for an extended period of time, tissue irritation would occur.
It is therefore desirable, and quite beneficial, to provide a secondary antimicrobial agent for longer lasting residual antimicrobial activity. Such secondary antimicrobial agents useful herein include medium chain fatty acids. These acids typically have a carbon chain from about 2 to about 15 carbon atoms, and preferably from about 6 to about 12 carbon atoms. Such fatty acids include acetic, propionic, octanoic, heptanoic, decanoic, nonanoic (pelargonic), undecanoic, and so forth. Octanoic and heptanoic are preferable fatty acids. These acids are useful in amounts of about 0.1% to about 10% by weight of the total composition, and preferably from about 0.2% to about 2% by weight of the total composition.
Longer chain length fatty acids, e.g. C11 to C15, are typically solid at room temperature, and are more difficult to solubilize in the aqueous system. It may therefore be necessary to use a co-coupling agent or hydrotrope in order to solubilize the longer chain fatty acid. Shorter chain fatty acids can function to increase the solubility of a longer chain fatty acid. For instance, acetic acid and propionic acid may be used for such a purpose.
Heptanoic and octanoic acids have been found to have better residual antimicrobial activity than some other fatty acids. The fatty acids disrupt the metabolism of a cell by entering the cell membrane and interfering with the cell function. The fatty acid must therefore be able to cross the hydrophobic membrane of the cell. If the number of carbon atoms in the fatty acid chain is too low, the fatty acid is not hydrophobic enough. If the number of carbon atoms is too high, the fatty acid is not soluble in water. The fatty acid therefore loses efficacy.
The solid chlorine dioxide generating component of the present invention may, in addition to the acid-forming-forming compound and the metal chlorite, also contain the antimicrobial fatty acid in a solid form. For instance, some fatty acids, in particular those having a carbon chain length of 8 or lower, are in a naturally liquid state at ambient temperatures, whereas fatty acids having a chain length of greater than about 12 are in a naturally solid state at ambient temperatures. Those fatty acids in a naturally solid state are easily packaged with the metal chlorite and the acid-forming compound in the solid component. However, longer chain length fatty acids may require co-coupling agents or hydrotropes in order to solubilize the longer chain fatty acid into the system. Longer chain fatty acids may also be modified to improve the solubility.
If the fatty acid is a liquid, an absorbent may be used in order to provide a tablet with a layered or sandwich type of structure.
The pH of the mixture of the present invention is preferably from about 5 to about 10, more preferably from about 5.5 to about 10, and most preferably from about 5.5 to about 8. The neutral pH of the present invention provides a composition which exhibits less irritation to the skin of animals or humans.
Other ingredients may be optionally added to the compositions of the present invention including wetting agents (i.e. emulsifiers or surfactants), emulsion stabilizers, defoamers, buffers, other antimicrobial agents, fungicides, emollients (skin conditioning or moisturizing agents), hydrotropes, humectants, thickeners or rheology modifiers, preservatives, dyes, plasticizers, vitamin E, insect repellents, perfumes, and so forth. An ingredient such as a dye may be added to the composition at the time of use as a separate tablet form, for instance.
Wetting agents or surfactants are often used to provide wetting properties to insure complete contact between the surface to be disinfected and the use solution. Wetting agents or surfactants useful herein include linear alcohol alkoxylates, i.e. linear alcohol ethoxylate, nonylphenoxypolyethoxy ethanol, sodium dioctylsulfosuccinate such as Aerosol(copyright) TO-75 available from American Cyanamid Co./Industrial Chemical Division. Surfactants are useful up to about 2% by weight of the composition, and preferably up to about 0.5% by weight of the composition.
Emollients useful herein include but are not limited to propylene glycol, glycerin, lanolin or lanolin derivatives, sorbitol, polyethylene glycol, aloe vera, and so forth. These agents are useful to maintain a healthy teat skin and to counteract any potential irritation effect from formulation ingredients or from the environment. Emollients are useful up to about 15% by weight of the composition. The amount of emollient utilized in the compositions of the present invention may be reduced because the of the mildness of the pH of the final composition, i.e. 5-10.
Polymeric thickeners or rheology modifiers may be optionally added to the compositions of the present invention to allow for better adhesion to the surface of the teat. This is important to improve the residence time of the antimicrobial in contact with the pathogens. Furthermore, it decreases the amount of waste lost in terms of the composition xe2x80x9cdrippingxe2x80x9d from the teat. Rheology modifiers may be inorganic or organic, and they may be water soluble or water dispersible. Organic thickeners may be divided into natural and synthetic polymers. Synthetic polymers may be natural based, or they may be petroleum based. These thickeners are useful up to about 1% by weight of the composition, and preferably up to about 0.5% by weight of the composition.
Inorganic thickeners include colloidal magnesium, aluminum silicates such as those sold under the tradename of Veegum(copyright), colloidal clays such as those sold under the tradename of Bentonite(copyright), silicas such as the Cab-o-sil(copyright) line, and so forth. These thickeners, sometimes referred to as fillers, may be fumed or precipitated to create particles with large surface to size ratios.
Natural hydrogel thickeners are those that are primarily derived from vegetable extrudates. For instance tragacanth, karaya and acacia gums; extractives including caragheenan, locust bean gum, guar gum, pectin and so forth; pure cultured fermentation products such as xanthan gum; and so forth.
Specific examples of useful rheology modifiers include xanthan gum, such as Kelzan(copyright) T available from and Keltrol(copyright) available from Kelco (Div. of Monsanto Company), and partially or fully hydrolyzed polyvinyl alcohol compositions. These materials are susceptible to shear stress wherein viscosity is reduced upon the addition of shear, and upon removal of the shear forces, the viscosity recovers. For teat dip compositions, the viscosity increases upon dipping, and upon removal, the viscosity returns, thereby immobilizing the coating on the teat so that little is wasted through drippage.
The rheology modifier may be a film former itself, or it may act cooperatively with a film-forming agent. For instance, xanthan gum may function both as a film former and as a rheology modifier, and partially or fully hydrolyzed polyvinyl alcohol may function both as a film-former, and as a rheology modifier. Film-forming agents are used in the barrier type teat dips.
Film-forming provides the formation of a barrier over the teat orifice in order to prevent access to the teat canal by the mastitis causing microorganisms. This barrier provides protection from the mastitis causing agents during the intermilking periods.
Useful classes of film forming agents include natural hydrogels (i.e. natural gums), gelatin, vinyl polymers, cellulose based film formers including ethylhydroxyethyl cellulose, and so forth. Film formers are useful from 0% to about 10% by weight of the total composition, and preferably from about 0.01% to about 8% by weight of the total composition.
The above ingredients may be included in the aqueous component in solution, or they may be included in the solid component, but it is preferable to include these ingredients in the aqueous solution. However, the present invention may also comprise a system of several tablets, each having various ingredients, that are placed in water at the time of use.
The compositions of the present invention are useful as teat dips including predips, prepost dips and barrier dips.
Other applications in which the compositions of the present invention find use include liquid and solid drain santizers, carcass dips (e.g., those used at rendering facilities), hoof treatment compositions, antimicrobial lubricants, surgical scrubbing compositions and antimicrobial hand products, topical skin treatments, antimicrobial veterinarian products, body fluid spill disinfection compositions, high level disinfectants for invasive and non-invasive surgical instruments, and so forth.

Antimicrobial composition useful for the treatment of bovine mastitis


A longitudinal vibrator assembly comprising at least one piezoceramic, magnetostrictive or electrostrictive transducer (130) having a coaxial housing (200a, 200b or 200c) comprised of at least one slotted or complete cylindrical flexural member vibrating in a circumferential or radial direction and excited by a solid state transduction material.

The invention in general relates to transducers, and more particularly, to an underwater transducer adapted for low frequency sonar use.
A sonar transducer is a device for generating sound and sensing sound in water. A sonar transducer is at heart a resonator which in the case of ceramic sonar transducers, includes an electroded ferroelectric member. The application of electrical potentials to the electrodes excites mechanical motion in the ferroelectric member used to generate sound waves in the water, and mechanical forces exerted upon the ferroelectric member by sound waves in the water is used to generate an electrical potential in the electrodes to sense the sound.
A common form of sonar transducer includes a xe2x80x9cstackxe2x80x9d of ring shaped drivers, electrically connected in parallel, clamped by means of a stress rod between a tail mass, which is relatively heavy, and a head mass, which constitutes a relatively light, water driving piston. The tail mass, ceramic stack, and head mass form a two mass resonator assembly. The arrangement desirably produces small amplitude vibrations in the tail mass and large amplitude vibrations of the head mass which acts as a water driving piston. This type of transducer is commonly referred to as a xe2x80x9cTonpilzxe2x80x9d design transducer or Tonpilz transducer. The Tonpilz transducer assembly is normally housed in an inactive watertight co-axial tube or inactive housing which serves to contain the active Tonpilz assembly and protect it from water intrusion.
Presently there is no known technique to obtain both low frequency (below on kilohertz) and higher 1-5 kHz response other than using parametric techniques which are limited by poor conversion efficiency and beam widths too narrow to be useful for wide area coverage. This is especially important in the areas of transmit transducers and arrays for anti-submarine warfare (ASW), communications and anti-mine warfare for surface, subsurface and air-launched applications as well as geophysical exploration and target simulation, for example. In the oil exploration industry, broadband coherent sources are greatly desired to take over the role of environmentally prohibitive air guns and explosive sources, for example. Greater frequency diversity provided by a single device which has both high and low frequency capability would be of significant benefit to both naval and geophysical applications.
Accordingly, a device that provides co-location of both low-frequency and high frequency or broadband signal processing is highly desired.
The present invention uses the normally inert housing of the Tonpilz projector to produce useful low frequency sound below the band of the Tonpilz element when used with flexural (flextensional) or slotted cylinder projectors as well as above the band of the Tonpilz element when used with complete cylinders.
The invention permits a relatively small Tonpilz or piston type transducer element to have a powerful and efficient (60-90%) low frequency surveillance transmit capability in addition to the normal tactical band capability normally associated with this type of element. In a preferred embodiment, a magnetostrictive, electrostrictive or piezoelectric driven Tonpilz driver mechanism is located within an active flexural structure such as a wall driven inverse flextensional or slotted cylinder projector (SCP) assembly. The wall driven flextensional or SCP projector provides the low frequency response in a weight-and-size efficient manner and the Tonpilz element makes efficient use of the empty space inside the wall driven flextensional or SCP. Another embodiment involves the use of a complete ceramic cylinder (not slotted) to make up part of the active housing and provide source level capabilities above the band of the Tonpilz element. Due to their higher frequency there placement in relation to head mass is more critical than the low frequency SCP due to diffraction effects.
The present invention is embodied in a longitudinal vibrator assembly comprising at least one piezoceramic, magnetostrictive, or electrostrictive transducer having a coaxial housing comprised of at least one wall driven flextensional, slotted or complete cylindrical flexural member vibrating in a circumferential or radial direction and excited by a solid state transduction material.
An underwater Tonpilz or piston assembly operative in a first longitudinal vibrational frequency mode and comprising an active housing operative for radiating sound at a substantially different frequency from the longitudinal vibrational frequency mode.
A transducer device comprising a Tonpilz element having a vibrating housing actuated by ceramic, magnetostrictive alloy or electrostrictive means, the housing having a flexural or circumferential or radial mode for increasing the effective bandwidth and frequency diversity of the device.

Active housing broadband tonpilz transducer


A method for treating priapism in a mammal by administration of a botulinum toxin.

The present invention relates to methods treating priapism. In particular the present invention relates to methods for treating priapism with a neurotoxin.
Priapism
Priapism is a prolonged, persistent (usually for four hours or longer) and often painful penile erection which is not associated with a sexual stimulus. Typically, only the corpora cavernosa of the penis is affected, the corpora spongiosum of the glans penis remaining flaccid. Both low blood flow and high blood flow forms of priapism have been described. Priapism derives its name from Priapus, the son of Aphrodite the ancient Greek goddess of love. Priapus was the Greek god of fertility and is shown in statutes, mosaics and pottery from the period with enormous genitalia and an apparently perpetual erection.
The most common cause of priapism is as a side effect of certain pharmacologicals, such as neuroleptic (i.e. thorazine and chlorpromazine), and high-blood pressure (i.e. prazosin) drugs. Notably, about 42 percent of all sickle-cell adults and 64 percent of all sickle-cell children develop priapism. Priapism also has been observed in association with: use of intracavernosal injections of medications to treat impotence (i.e. papaverine, phentolamine, and prostaglandin E1); leukemia; multiple myeloma; Fabry disease; mycoplasma pneumonia; amyloidosis; carbon monoxide poisoning; malaria; spider bites; citalopram use (a selective serotonin reuptake inhibitor); hydralazine; metoclopramide; omeprazole; hydroxyzine; prazosin (especially when used in patients with renal failure); tamoxifen; testosterone; calcium channel blockers; anticoagulants (both warfarin-induced and during heparin infusions); cocaine; the drug ecstasy; ethanol abuse; androstenedione (for athletic related purposes); marijuana, and; certain cancers which infiltrate the penis and prevent the outflow of blood.
Childhood priapism is known to occur in association with leukemia (white blood cells occluding outflow of blood from the penis), sickle-cell disease, trauma to the penis or to the perineum, and spinal cord injury. Priapism of the clitoris (female priapism) has been rarely described. See e.g. Brodie-Meijer C. C. et al., Nefazodone-induced clitoral priapism, Int Clin Psychopharmacol 1999 July;14(4):257-8.
Priapism can occur as a result of a disturbance to the normal regulatory mechanisms that initiate and maintain penile flaccidity. Thus, it is believed that activation of post ganglionic (cholinergic) parasympathetic autonomic nerves can induce erection of the penis, while sympathetic (adrenergic) innervation of the penis induces penile detumescence and terminate erection.
Thus, parasympathetic relaxation of penile smooth muscle (possibly mediated by nitric oxide induced drop in cytosolic calcium) causes an erection by permitting blood to flow into penile structures and increase penile cavernosal pressure. Contraction of sympathetic (adrenergic) innervated, penile smooth muscle results in detumescence (a flaccid penis) due to increased drainage from the penis via venous outflow channels, thereby decreasing intracavernosal pressure. Compton M. T. et al., Priapism associated with conventional and atypical medications: a review, J Clin Psychiatry 2001;62:362-366; Rochat M. C., Priapism: a review. Theriogenology 2001;56:713-722; Wagner G. et al., Pathophysiology and diagnosis of male erectile dysfunction, BJU International (2001), 88 (Suppl. 3), 3-10; Lue T F. Erectile Dysfunction. New Engl J Med 2001;342:1802-1813.
Priapism with an onset within 4-6 hours can sometimes be treated successfully with a decongestant (i.e. pseudoephedrine or terbutaline) which acts to decrease blood flow to the penis. If the erection is non-responsive, aspiration can be used to remove about 50 to 150 cc of blood out of the penis through a small needle placed directly in the corporal body, thereby allowing detumesce.
If the erection reoccurs, certain vaso-active type drugs including epinephrine may be instilled into the penis, causing the blood vessels to constrict and prevent the priapism. If this is unsuccessful an alpha-adrenergic agonist (e.g. metaraminol bitartrate, phenylephrine, metaraminol) can be tried, Alpha agonists counteract penile smooth muscle relaxation which causes an erection. However, it is known that an alpha-agonist can cause significant systemic hypertension as well as ventricular tachycardia. Another treatment option is a shunting procedure to permit blood to drain from the engorged penis.
Unfortunately, current therapies for priapism, including drugs, aspiration and shunting, have significant drawbacks and disadvantages, including recurrence of the priapism; bleeding from the holes placed in the penis as a part of a shunt procedure; post-surgical infections including infection of the corporal body and infection of the skin around the corporal body; necrosis of the penis; damage to the urethra; holes between the urethra and the skin; loss of the penis, and death.
Loss of the penis can occur due to post surgical infection or because of penile necrosis due to the duration of the priapism before an effective treatment was initiated. Priapism has resulted in death from suicide as well as from a blood clot formed in the penis after a shunting procedure breaking off and causing pulmonary embolism.
Botulinum Toxin
The anaerobic, gram positive bacterium Clostridium botulinum produces a potent polypeptide neurotoxin, botulinum toxin, which causes a neuroparalytic illness in humans and animals referred to as botulism. Clostridium botulinum and its spores are commonly found in soil and the bacterium can grow in improperly sterilized and sealed food containers of home based canneries, which are the cause of many of the cases of botulism. The effects of botulism typically appear 18 to 36 hours after eating the foodstuffs infected with a Clostridium botulinum culture or spores. The botulinum toxin can apparently pass unattenuated through the lining of the gut and poison peripheral motor neurons. Symptoms of botulinum toxin intoxication can progress from difficulty walking, swallowing, and speaking to paralysis of the respiratory muscles and death.
Botulinum toxin type A is the most lethal natural biological agent known to man. About 50 picograms of a botulinum toxin (purified neurotoxin complex) type A from the Hall strain of a Clostridium botulinum is a LD50 in mice. Interestingly, on a molar basis, botulinum toxin type A is 1.8 billion times more lethal than diphtheria, 600 million times more lethal than sodium cyanide, 30 million times more lethal than cobrotoxin and 12 million times more lethal than cholera. Singh, Critical Aspects of Bacterial Protein Toxins, pages 63-84 (chapter 4) of Natural Toxins II, edited by B. R. Singh et al., Plenum Press, New York (1976) (where the stated LD50 of botulinum toxin type A of 0.3 ng equals 1 U is corrected for the fact that about 0.05 ng of BOTOX(copyright) equals 1 unit). One unit (U) of botulinum toxin is defined as the LD50 upon intraperitoneal injection into female Swiss Webster mice weighing 18-20 grams each.
In other words, one unit of botulinum toxin is the amount of botulinum toxin that kills 50% of a group of female Swiss Webster mice. Seven generally immunologically distinct botulinum neurotoxins have been characterized, these being respectively botulinum neurotoxin serotypes A, B, C1, D, E, F, and G, each of which is distinguished by neutralization with type-specific antibodies. The different serotypes of botulinum toxin vary in the animal species that they affect and in the severity and duration of the paralysis they evoke. For example, it has been determined that botulinum toxin type A is 500 times more potent, as measured by the rate of paralysis produced in the rat, than is botulinum toxin type B. Additionally, botulinum toxin type B has been determined to be non-toxic in primates at a dose of 480 U/kg which is about 12 times the primate LD50 for botulinum toxin type A. The botulinum toxins apparently bind with high affinity to cholinergic motor neurons, are translocated into the neuron and block the presynaptic release of acetylcholine.
Botulinum toxins have been used in clinical settings for the treatment of neuromuscular disorders characterized by hyperactive skeletal muscles. Botulinum toxin type A was approved by the U.S. Food and Drug Administration in 1989 for the treatment of essential blepharospasm, strabismus and hemifacial spasm in patients over the age of twelve. In 2000 the FDA approved commercial preparations of type A and type B botulinum toxin serotypes for the treatment of cervical dystonia, and in 2002 the FDA approved a type A botulinum toxin for the cosmetic treatment of certain hyperkinetic (glabellar) facial wrinkles. Clinical effects of peripheral intramuscular botulinum toxin type A are usually seen within one week of injection and sometimes within a few hours. The typical duration of symptomatic relief (i.e. flaccid muscle paralysis) from a single intramuscular injection of botulinum toxin type A can be about three months, although in some cases the effects of a botulinum toxin induced denervation of a gland, such as a salivary gland, have been reported to last for several years.
Although all the botulinum toxins serotypes apparently inhibit release of the neurotransmitter acetylcholine at the neuromuscular junction, they do so by affecting different neurosecretory proteins and/or cleaving these proteins at different sites. Botulinum toxin A is a zinc endopeptidase which can specifically hydrolyze a peptide linkage of the intracellular, vesicle associated protein SNAP-25. Botulinum type E also cleaves the 25 kiloDalton (kD) synaptosomal associated protein (SNAP-25), but targets different amino acid sequences within this protein, as compared to botulinum toxin type A. Botulinum toxin types B, D, F and G act on vesicle-associated protein (VAMP, also called synaptobrevin), with each serotype cleaving the protein at a different site. Finally, botulinum toxin type C1 has been shown to cleave both syntaxin and SNAP-25. These differences in mechanism of action may affect the relative potency and/or duration of action of the various botulinum toxin serotypes.
Regardless of serotype, the molecular mechanism of toxin intoxication appears to be similar and to involve at least three steps or stages. In the first step of the process, the toxin binds to the presynaptic membrane of the target neuron through a specific interaction between the heavy chain (H chain) and a cell surface receptor; the receptor is thought to be different for each serotype of botulinum toxin and for tetanus toxin. The carboxyl end segment of the H chain, Hc, appears to be important for targeting of the toxin to the cell surface.
In the second step, the toxin crosses the plasma membrane of the poisoned cell. The toxin is first engulfed by the cell through receptor-mediated endocytosis, and an endosome containing the toxin is formed. The toxin then escapes the endosome into the cytoplasm of the cell. This last step is thought to be mediated by the amino end segment of the H chain, HN, which triggers a conformational change of the toxin in response to a pH of about 5.5 or lower. Endosomes are known to possess a proton pump which decreases intra endosomal pH. The conformational shift exposes hydrophobic residues in the toxin, which permits the toxin to embed itself in the endosomal membrane. The toxin then translocates through the endosomal membrane into the cytosol.
The last step of the mechanism of botulinum toxin activity appears to involve reduction of the disulfide bond joining the H and L chain. The entire toxic activity of botulinum and tetanus toxins is contained in the L chain of the holotoxin; the L chain is a zinc (Zn++) endopeptidase which selectively cleaves proteins essential for recognition and docking of neurotransmitter-containing vesicles with the cytoplasmic surface of the plasma membrane, and fusion of the vesicles with the plasma membrane. Tetanus neurotoxin, botulinum toxin B, D, F, and G cause degradation of synaptobrevin (also called vesicle-associated membrane protein (VAMP)), a synaptosomal membrane protein. Most of the VAMP present at the cytosolic surface of the synaptic vesicle is removed as a result of any one of these cleavage events. Each toxin specifically cleaves a different bond.
The molecular weight of the botulinum toxin protein molecule, for all seven of the known botulinum toxin serotypes, is about 150 kD. Interestingly, the botulinum toxins are released by Clostridial bacterium as complexes comprising the 150 kD botulinum toxin protein molecule along with associated non-toxin proteins. Thus, the botulinum toxin type A complex can be produced by Clostridial bacterium as 900 kD, 500 kD and 300 kD forms. Botulinum toxin types B and C1 are apparently produced as only a 500 kD complex. Botulinum toxin type D is produced as both 300 kD and 500 kD complexes. Finally, botulinum toxin types E and F are produced as only approximately 300 kD complexes. The complexes (i.e. molecular weight greater than about 150 kD) are believed to contain a non-toxin hemaglutinin protein and a non-toxin and non-toxic nonhemaglutinin protein. These two non-toxin proteins (which along with the botulinum toxin molecule can comprise the relevant neurotoxin complex) may act to provide stability against denaturation to the botulinum toxin molecule and protection against digestive acids when toxin is ingested. Additionally, it is possible that the larger (greater than about 150 kD molecular weight) botulinum toxin complexes may result in a slower rate of diffusion of the botulinum toxin away from a site of intramuscular injection of a botulinum toxin complex. The toxin complexes can be dissociated into toxin protein and hemagglutinin proteins by treating the complex with red blood cells at pH 7.3. The toxin protein has a marked instability upon removal of the hemagglutinin protein.
All the botulinum toxin serotypes are made by Clostridium botulinum bacteria as inactive single chain proteins which must be cleaved or nicked by proteases to become neuroactive. The bacterial strains that make botulinum toxin serotypes A and G possess endogenous proteases and serotypes A and G can therefore be recovered from bacterial cultures in predominantly their active form. In contrast, botulinum toxin serotypes C1, D, and E are synthesized by nonproteolytic strains and are therefore typically unactivated when recovered from culture. Serotypes B and F are produced by both proteolytic and nonproteolytic strains and therefore can be recovered in either the active or inactive form. However, even the proteolytic strains that produce, for example, the botulinum toxin type B serotype only cleave a portion of the toxin produced. The exact proportion of nicked to unnicked molecules depends on the length of incubation and the temperature of the culture. Therefore, a certain percentage of any preparation of, for example, the botulinum toxin type B toxin is likely to be inactive, possibly accounting for a lower potency of botulinum toxin type B as compared to botulinum toxin type A. The presence of inactive botulinum toxin molecules in a clinical preparation will contribute to the overall protein load of the preparation, which has been linked to increased antigenicity, without contributing to its clinical efficacy.
In vitro studies have indicated that botulinum toxin inhibits potassium cation induced release of both acetylcholine and norepinephrine from primary cell cultures of brainstem tissue. Additionally, it has been reported that botulinum toxin inhibits the evoked release of both glycine and glutamate in primary cultures of spinal cord neurons and that in brain synaptosome preparations botulinum toxin inhibits the release of each of the neurotransmitters acetylcholine, dopamine, norepinephrine, CGRP and glutamate.
High quality crystalline botulinum toxin type A can be produced from the Hall A strain of Clostridium botulinum with characteristics of xe2x89xa73xc3x97107 U/mg, an A260/A278 of less than 0.60 and a distinct pattern of banding on gel electrophoresis. The known Shantz process can be used to obtain crystalline botulinum toxin type A, as set forth in Shantz, E. J., et al, Properties and use of Botulinum toxin and Other Microbial Neurotoxins in Medicine, Microbiol Rev. 56: 80-99 (1992). Generally, the botulinum toxin type A complex can be isolated and purified from an anaerobic fermentation by cultivating Clostridium botulinum type A in a suitable medium. Raw toxin can be harvested by precipitation with sulfuric acid and concentrated by ultramicrofiltration. Purification can be carried out by dissolving the acid precipitate in calcium chloride. The toxin can then be precipitated with cold ethanol. The precipitate can be dissolved in sodium phosphate buffer and centrifuged. Upon drying there can then be obtained approximately 900 kD crystalline botulinum toxin type A complex with a specific potency of 3xc3x97107 LD50 U/mg or greater. This known process can also be used, upon separation out of the non-toxin proteins, to obtain pure botulinum toxins, such as for example: purified botulinum toxin type A with an approximately 150 kD molecular weight with a specific potency of 1-2xc3x97108 LD50 U/mg or greater; purified botulinum toxin type B with an approximately 156 kD molecular weight with a specific potency of 1-2xc3x97108 LD50 U/mg or greater, and; purified botulinum toxin type F with an approximately 155 kD molecular weight with a specific potency of 1-2xc3x97107 LD50 U/mg or greater.
Already prepared and purified botulinum toxins and toxin complexes suitable for preparing pharmaceutical formulations can be obtained from List Biological Laboratories, Inc., Campbell, Calif.; the Centre for Applied Microbiology and Research, Porton Down, U.K.; Wako (Osaka, Japan), as well as from Sigma Chemicals of St Louis, Mo.
It has been reported that a botulinum toxin has been used in clinical settings as follows:
(1) about 75-125 units of BOTOX(copyright)1 per intramuscular injection (multiple muscles) to treat cervical dystonia;
1 Available from Allergan, Inc., of Irvine, Calif. under the tradename BOTOX(copyright).
(2) 5-10 units of BOTOX(copyright) per intramuscular injection to treat glabellar lines (brow furrows) (5 units injected intramuscularly into the procerus muscle and 10 units injected intramuscularly into each corrugator supercilii muscle);
(3) about 30-80 units of BOTOX(copyright) to treat constipation by intrasphincter injection of the puborectalis muscle;
(4) about 1-5 units per muscle of intramuscularly injected BOTOX(copyright) to treat blepharospasm by injecting the lateral pre-tarsal orbicularis oculi muscle of the upper lid and the lateral pre-tarsal orbicularis oculi of the lower lid.
(5) to treat strabismus, extraocular muscles have been injected intramuscularly with between about 1-5 units of BOTOX(copyright), the amount injected varying based upon both the size of the muscle to be injected and the extent of muscle paralysis desired (i.e. amount of diopter correction desired).
(6) to treat upper limb spasticity following stroke by intramuscular injections of BOTOX(copyright) into five different upper limb flexor muscles, as follows:
(a) flexor digitorum profundus: 7.5 U to 30 U
(b) flexor digitorum sublimus: 7.5 U to 30 U
(c) flexor carpi ulnaris: 10 U to 40 U
(d) flexor carpi radialis: 15 U to 60 U
(e) biceps brachii: 50 U to 200 U. Each of the five indicated muscles has been injected at the same treatment session, so that the patient receives from 90 U to 360 U of upper limb flexor muscle BOTOX(copyright) by intramuscular injection at each treatment session.
(7) to treat migraine, pericranial injected (injected symmetrically into glabellar, frontalis and temporalis muscles) injection of 25 U of BOTOX(copyright) has showed significant benefit as a prophylactic treatment of migraine compared to vehicle as measured by decreased measures of migraine frequency, maximal severity, associated vomiting and acute medication use over the three month period following the 25 U injection.
Additionally, intramuscular botulinum toxin has been used in the treatment of tremor in patients with Parkinson""s disease, although it has been reported that results have not been impressive. Marjama-Jyons, J., et al., Tremor-Predominant Parkinson""s Disease, Drugs and Aging 16(4);273-278:2000.
Treatment of certain gastrointestinal and smooth muscle disorders with a botulinum toxin are known. See e.g. U.S. Pat. Nos. 5,427,291 and 5,674,205 (Pasricha). Additionally, transurethral injection of a botulinum toxin into a bladder sphincter to treat a urination disorder is known (see e.g. Dykstra, D. D., et al, Treatment of detrusor-sphincter dyssynergia with botulinum A toxin: A double-blind study, Arch Phys Med Rehabil 1990 January;71:24-6), as is injection of a botulinum toxin into the prostate to treat prostatic hyperplasia. See e.g. U.S. Pat. No. 6,365,164 (Schmidt).
U.S. Pat. No. 5,766,605 (Sanders) proposes the treatment of various autonomic disorders, such as hypersalivation and rhinittis, with a botulinum toxin.
Furthermore, various afflictions, such as hyperhydrosis and headache, treatable with a botulinum toxin are discussed in WO 95/17904 (PCT/US94/14717) (Aoki). EP 0 605 501 B1 (Graham) discusses treatment of cerebral palsy with a botulinum toxin and U.S. Pat. No. 6,063,768 (First) discusses treatment of neurogenic inflammation with a botulinum toxin.
Erectile dysfunction has been reported as a symptom of botulism. Jenzer G., et al., Autonomic dysfunction in botulism B: a clinical report, Neurology 1975;25:150-153; Naumann M. et al., Pure autonomic dysfunction in botulism type B, Naunyn Schmiedeberg""s Archives of Pharmacology June 2002 (supp 2); 365 (abstract 89 at R31). This may be a result of circulating botulinum toxin present in a patient with botulism acting to block release of acetylcholine from cholinergic parasympathetic nerve endings in the corpora cavernosa of the penis. This would cause an inhibition of penile smooth muscle relaxation and therefore a reduced flow of blood into penile structures, and hence a flaccid penis. Contrarily, it has been speculated that a botulinum toxin can be used to cause an erection of the penis. Jones D. High performance. Nature 1989;3:348.
It is known that botulinum toxin type A can have an efficacy for up to 12 months (European J Neurology 6 (Supp 4): S111-S1150:1999), and in some circumstances for as long as 27 months. The Laryngoscope 109:1344-1346:1999. However, the usual duration of an intramuscular injection of Botox(copyright) is typically about 3 to 4 months.
The success of botulinum toxin type A to treat a variety of clinical conditions has led to interest in other botulinum toxin serotypes. Additionally, pure botulinum toxin has been used to treat humans. see e.g. Kohl A., et al., Comparison of the effect of botulinum toxin A (Botox (R)) with the highly-purified neurotoxin (NT 201) in the extensor digitorum brevis muscle test, Mov Disord 2000;15(Suppl 3):165. Hence, a pharmaceutical composition can be prepared using a pure botulinum toxin.
The botulinum toxin molecule (about 150 kDa), as well as the botulinum toxin complexes (about 300-900 kDa), such as the toxin type A complex are also extremely susceptible to denaturation due to surface denaturation, heat, and alkaline conditions. Inactivated toxin forms toxoid proteins which may be immunogenic. The resulting antibodies can render a patient refractory to toxin injection.
A commercially available botulinum toxin containing pharmaceutical composition is sold under the trademark BOTOX(copyright) (available from Allergan, Inc., of Irvine, Calif.). BOTOX(copyright) consists of a purified botulinum toxin type A complex, albumin, and sodium chloride packaged in sterile, vacuum-dried form. The botulinum toxin type A is made from a culture of the Hall strain of Clostridium botulinum grown in a medium containing N-Z amine and yeast extract. The botulinum toxin type A complex is purified from the culture solution by a series of acid precipitations to a crystalline complex consisting of the active high molecular weight toxin protein and an associated hemagglutinin protein. The crystalline complex is re-dissolved in a solution containing saline and albumin and sterile filtered (0.2 microns) prior to vacuum-drying. BOTOX(copyright) can be reconstituted with sterile, non-preserved saline prior to intramuscular injection. Each vial of BOTOX(copyright) contains about 100 units (U) of Clostridium botulinum toxin type A complex, 0.5 milligrams of human serum albumin and 0.9 milligrams of sodium chloride in a sterile, vacuum-dried form without a preservative.
To reconstitute vacuum-dried BOTOX(copyright) sterile normal saline without a preservative; 0.9% Sodium Chloride injection is used by drawing up the proper amount of diluent in the appropriate size syringe. Since BOTOX(copyright) is denatured by bubbling or similar violent agitation, the diluent is gently injected into the vial. BOTOX(copyright) should be administered within four hours after reconstitution. During this time period, reconstituted BOTOX(copyright) is stored in a refrigerator (2xc2x0 to 8xc2x0 C.). Reconstituted BOTOX(copyright) is clear, colorless and free of particulate matter. The vacuum-dried product is stored in a freezer at or below xe2x88x925xc2x0 C. BOTOX(copyright) is administered within four hours after the vial is removed from the freezer and reconstituted. During these four hours, reconstituted BOTOX(copyright) can be stored in a refrigerator (20 to 8xc2x0 C.).
Other commercially available botulinum toxin containing pharmaceutical compositions include Dysport(copyright) (Clostridium botulinum type A toxin haemagglutinin complex with albumin and lactose in the formulation, available from Ipsen Limited, Berkshire, U.K. as a powder to be reconstituted with 0.9% sodium chloride before use), and MyoBloc(trademark) (an injectable solution comprising botulinum toxin type B, human serum albumin, sodium succinate, and sodium chloride at about pH 5.6, available from Elan Corporation, Dublin, Ireland).
Acetylcholine
Typically, only a single type of small molecule neurotransmitter is released by each type of neuron in the mammalian nervous system. The neurotransmitter acetylcholine is secreted by neurons in many areas of the brain, but specifically by the large pyramidal cells of the motor cortex, by several different neurons in the basal ganglia, by the motor neurons that innervate the skeletal muscles, by the preganglionic neurons of the autonomic nervous system (both sympathetic and parasympathetic), by the postganglionic neurons of the parasympathetic nervous system, and by some of the postganglionic neurons of the sympathetic nervous system. Essentially, only the postganglionic sympathetic nerve fibers to the sweat glands, the piloerector muscles and a few blood vessels are cholinergic as most of the postganglionic neurons of the sympathetic nervous system secret the neurotransmitter norepinephine. In most instances, acetylcholine has an excitatory effect. However, acetylcholine is known to have inhibitory effects at some of the peripheral parasympathetic nerve endings, such as inhibition of heart rate by the vagal nerve.
The efferent signals of the autonomic nervous system are transmitted to the body through either the sympathetic nervous system or the parasympathetic nervous system. The preganglionic neurons of the sympathetic nervous system extend from preganglionic sympathetic neuron cell bodies located in the intermediolateral horn of the spinal cord. The preganglionic sympathetic nerve fibers, extending from the cell body, synapse with postganglionic neurons located in either a paravertebral sympathetic ganglion or in a prevertebral ganglion. Since, the preganglionic neurons of both the sympathetic and parasympathetic nervous system are cholinergic, application of acetylcholine to the ganglia will excite both sympathetic and parasympathetic postganglionic neurons.
Acetylcholine activates two types of receptors, muscarinic and nicotinic receptors. The muscarinic receptors are found in all effector cells stimulated by the postganglionic, neurons of the parasympathetic nervous system as well as in those stimulated by the postganglionic cholinergic neurons of the sympathetic nervous system. The nicotinic receptors are found in the adrenal medulla, as well as within the autonomic ganglia, that is on the cell surface of the postganglionic neuron at the synapse between the preganglionic and postganglionic neurons of both the sympathetic and parasympathetic systems. Nicotinic receptors are also found in many nonautonomic nerve endings, for example in the membranes of skeletal muscle fibers at the neuromuscular junction.
Acetylcholine is released from cholinergic neurons when small, clear, intracellular vesicles fuse with the presynaptic neuronal cell membrane. A wide variety of non-neuronal secretory cells, such as, adrenal medulla (as well as the PC12 cell line) and pancreatic islet cells release catecholamines and parathyroid hormone, respectively, from large dense-core vesicles. The PC12 cell line is a clone of rat pheochromocytoma cells extensively used as a tissue culture model for studies of sympathoadrenal development. Botulinum toxin inhibits the release of both types of compounds from both types of cells in vitro, permeabilized (as by electroporation) or by direct injection of the toxin into the denervated cell. Botulinum toxin is also known to block release of the neurotransmitter glutamate from cortical synaptosomes cell cultures.
A neuromuscular junction is formed in skeletal muscle by the proximity of axons to muscle cells. A signal transmitted through the nervous system results in an action potential at the terminal axon, with activation of ion channels and resulting release of the neurotransmitter acetylcholine from intraneuronal synaptic vesicles, for example at the motor endplate of the neuromuscular junction. The acetylcholine crosses the extracellular space to bind with acetylcholine receptor proteins on the surface of the muscle end plate. Once sufficient binding has occurred, an action potential of the muscle cell causes specific membrane ion channel changes, resulting in muscle cell contraction. The acetylcholine is then released from the muscle cells and metabolized by cholinesterases in the extracellular space. The metabolites are recycled back into the terminal axon for reprocessing into further acetylcholine.
What is needed therefore is an effective and long lasting treatment of priapism with few if any side effects.
The present invention meets this need and provides an effective and long lasting treatment of priapism with few if any side effects
Definitions
As used herein, the words or terms set forth below have the following definitions.
xe2x80x9cAboutxe2x80x9d means that the item, parameter or term so qualified encompasses a range of plus or minus ten percent above and below the value of the stated item, parameter or term.
xe2x80x9cBiological activityxe2x80x9d with regard to a neurotoxin includes an ability of the neurotoxin to reduce or inhibit presynaptic release of acetylcholine.
xe2x80x9cLocal administrationxe2x80x9d means direct administration by a non-systemic route at or in the vicinity of the site of an affliction or disorder.
Thus, local administration of a pharmaceutical excludes, for example, intravenous or oral administration, but includes, for example, intramuscular or subcutaneous injection or implant placement drug administration. Systemic administration of a botulinum toxin is contraindicated because botulism can result.
xe2x80x9cNeurotoxinxe2x80x9d includes Clostridial neuro toxins both in pure toxin and as complexed with one to more non-toxin, toxin associated proteins, whether made by the native Clostridial bacterium or by recombinant means in a non-Clostridial species.
xe2x80x9cPurified or pure botulinum toxinxe2x80x9d means a botulinum toxin that is isolated, or substantially isolated, from other proteins, including proteins that form a botulinum toxin complex. A purified botulinum toxin may be greater than 95% pure, and preferably is greater than 99% pure.
xe2x80x9cPatientxe2x80x9d means a human or non-human subject receiving medical or veterinary care.
The present invention encompasses a method for treating priapism by local administration of a Clostridial toxin to a mammal with priapism. The Clostridial toxin can be a botulinum toxin, such as a botulinum toxin type A, B, C, D, E, F or G. Preferably, the botulinum toxin is a botulinum toxin type A.
The botulinum toxin can be administered in an amount between about 1 unit and about 10,000 units and the priapism can be substantially alleviated for between about 2 weeks and about 6 months. Thus, for example, about 1 unit of a type A toxin can be to treat clitoral priapism or infantile priapism. About 10,000 units of a type B botulinum toxin can be used to treat a large mammal with a penis of considerable size. Notably, the local administration step is carried out by direct administration of the Clostridial toxin to a penis of the mammal.
A further embodiment of the present invention is a method for treating priapism, by local administration of a botulinum toxin to a penis of a mammal with priapism. A most preferred embodiment of the present invention is a method for treating priapism, the method comprising the step of local administration of a botulinum toxin type A to a penis of a mammal with priapism.
My invention is based upon the discovery that long term alleviation of priapism can be achieved by local administration of a neurotoxin to a mammal afflicted with this condition.
The mammal treated can be a human or another species which exhibits priapism, such as a domesticated mammal, such as a dog or a horse. See e.g. Rochat M. C., Priapism: a review. Theriogenology 2001;56:713-722.
My invention encompasses use of a neurotoxin, such as a Clostridial toxin, such as a botulinum toxin (any of the serotypes A, B, C, D, E, F or G) for the therapeutic treatment of priapism in humans or other mammals. A method within the scope of my invention can be practiced by local administration of a of botulinum toxin to treat a persistent erection of the penis, i.e. priapism.
To reiterate, an embodiment of the present invention can be carried out by injecting a botulinum toxin type into a penis of the mammal to be treated. As set forth above, it is believed that cholinergic parasympathetic innervation induces penile erection of the penis, while sympathetic nerves induce penile detumescence (flaccidity). Without wishing to be bound by theory, it can be hypothesized that a botulinum toxin, by inhibiting release of acetylcholine from cholinergic parasympathetic nerve endings in the corpora cavernosa of the penis, causes an inhibition of penile smooth muscle relaxation. Hence, the unaffected adrenergic sympathetic innervation causes penile smooth muscle contraction and thereby a flaccid penis because of reduced blood inflow to penile structures. Thus, the treatable priapism within the scope of the present invention is a priapism responsive to a downregulation of penile parasympathetic innervation. High flow priapism may be unsuitable for treatment according to the present invention. For example, a patient who suffers injury to the perineum can exhibit traumatic high-flow priapism because a genital artery had ruptured. The blood enters the artery, goes into the corporal body of the penis, thereby creating an erection, and then immediately leaves the penis because the penile veins were not constricted. A continuous (i.e. high flow) of blood form the ruptured artery into the penis maintains the erection. With such a patient an arteriogram of the pudendal arterial system can be carried out to identify the point of the fistula and a small coil of material (a stent) placed into the damaged artery. Injection of a botulinum toxin would probably have been ineffective to treat such a high flow priapism.
Preferably, because of its ready availability and clinical history to successfully treat a number of indications, a method within the scope of the present invention includes local administration of a botulinum type A. A botulinum toxin type B can also be used because a botulinum toxin type B is also commercially available, although it is used with a larger protein load, as compared to type A toxin. A botulinum toxin type A used in a method within the scope of the present invention can be a complex of toxin and non-toxin proteins, which together comprise a total molecular weight of about 900 kiloDaltons and which is used at a dose of between 1 and 100 units per patient, the range being based upon size, age and health of the patient, as well as upon the particular commercial preparation of the type A botulinum toxin used. A botulinum toxin type B used in a method within the scope of the present invention can be a pure toxin or complex of toxin and non-toxin proteins, which together comprise a total molecular weight of about 700 kiloDaltons and which is used at a dose of between about 50 and about 20,000 units per patient treated.
Other botulinum toxin serotypes can be used in proportion to the dosages and concentrations exemplified herein, according to their respective levels of biological activity. The present invention also encompasses methods for concurrent or serial administration of a mixture of two or more of the above neurotoxins to effectively treat a patient with priapism.
A neurotoxin, such as a botulinum toxin can require, according to the methods of the present invention, from about 1 hour to 7 days to achieve it""s desired detumescence effect.
Additionally a neurotoxin, such as a botulinum toxin, according to the present invention is always locally administered in vivo directly to the patient""s (a mammal) penis. Known local drug administration methods suitable for this purpose include a syringe for liquid pharmaceutical injection and insertion of a controlled release implant (See e.g. U.S. Pat. Nos. 6,383,509 and 6,312,708 (Donovan)). Systemic routes of drug administration such as oral or intravenous administration are excluded from the scope of the present invention because systemic distribution of a neurotoxin, such as a botulinum toxin, is not desirable.
In another embodiment, the methods comprise the administration of a neurotoxin, for example a Clostridial neurotoxin, to a patient wherein the neurotoxin differs from a naturally occurring (native) neurotoxin by at least one amino acid. For example, variants of botulinum toxin type A as disclosed in Biochemistry 34;5175-15181:1995 and Eur. J. Biochem, 185;197-203:1989 can be administered. Practice of the present invention can provide a priapism treatment effective for between 2 weeks (i.e. if a type E toxin is used) to up to about 6 months or longer (type A toxin is used).
The amount of the neurotoxin, such as a botulinum toxin, administered can vary widely according to the particular disorder being treated, its severity and other various patient variables including size, weight, age, and responsiveness to therapy. Generally, the dose of neurotoxin to be administered will vary with the age, presenting condition and weight of the mammal to be treated. The potency of the neurotoxin to be administered is also a consideration.
In one embodiment according to this invention, the therapeutically effective doses of a neurotoxin, for example a botulinum toxin type A complex (as Botox), can be between about 1 unit and about 500 units Less than about 1 unit can result in a suboptimal detumescence effect while more than about than about 500 units of a type A preparation can result in undesired systemic effects.
Although examples of routes of administration and dosages are provided, the appropriate route of administration and dosage are generally determined on a case by case basis by the attending physician. Such determinations are routine to one of ordinary skill in the art (see for example, Harrison""s Principles of Internal Medicine (1998), edited by Anthony Fauci et al., 14th edition, published by McGraw Hill). For example, the route and dosage for administration of a neurotoxin according to the present disclosed invention can be selected based upon criteria such as the solubility characteristics of the neurotoxin chosen as well as the age and health of the patient.
A suitable neurotoxin can be obtained by culturing an appropriate bacterial species. For example, botulinum toxins can be obtained by establishing and growing cultures of Clostridium botulinum in a fermenter and then harvesting and purifying the fermented mixture in accordance with known procedures. All the botulinum toxin serotypes are initially synthesized as inactive single chain proteins which must be cleaved or nicked by proteases to become neuroactive. The bacterial strains that make botulinum toxin serotypes A and G possess endogenous proteases and serotypes A and G can therefore be recovered from bacterial cultures in predominantly their active form. In contrast, botulinum toxin serotypes C1, D and E are synthesized by nonproteolytic strains and are therefore typically unactivated when recovered from culture. Serotypes B and F are produced by both proteolytic and nonproteolytic strains and therefore can be recovered in either the active or inactive form. However, even the proteolytic strains that produce, for example, the botulinum toxin type B serotype only cleave a portion of the toxin produced. The exact proportion of nicked to unnicked molecules depends on the length of incubation and the temperature of the culture.
If a modified neurotoxin is to be used according to this invention, recombinant techniques can be used to produce the desired neurotoxins. The technique includes steps of obtaining genetic materials from natural sources, or synthetic sources, which have codes for a neuronal binding moiety, an amino acid sequence effective to translocate the neurotoxin or a part thereof, and an amino acid sequence having therapeutic activity when released into a cytoplasm of a target cell, preferably a neuron.
A method within the scope of the present invention can provide improved patient function. xe2x80x9cImproved patient functionxe2x80x9d can be defined as an improvement measured by factors such as a reduced pain, reduced time spent in bed, increased ambulation, healthier attitude, more varied lifestyle and/or healing permitted by normal muscle tone. Improved patient function is synonymous with an improved quality of life (QOL). QOL can be assesses using, for example, the known SF-12 or SF-36 health survey scoring procedures. SF-36 assesses a patient""s physical and mental health in the eight domains of physical functioning, role limitations due to physical problems, social functioning, bodily pain, general mental health, role limitations due to emotional problems, vitality, and general health perceptions. Scores obtained can be compared to published values available for various general and patient populations.


The present invention provides a small umbrella (xe2x80x9cBeerbrellaxe2x80x9d) which may be removably attached to a beverage container in order to shade the beverage container from the direct rays of the sun. The apparatus comprises a small umbrella approximately five to seven inches in diameter, although other appropriate sizes may be used within the spirit and scope of the present invention. Suitable advertising and/or logos may be applied to the umbrella surface for promotional purposes. The umbrella may be attached to the beverage container by any one of a number of means, including clip, strap, cup, foam insulator, or as a coaster or the like. The umbrella shaft may be provided with a pivot to allow the umbrella to be suitably angled to shield the sun or for aesthetic purposes. In one embodiment, a pivot joint and counterweight may be provided to allow the umbrella to pivot out of the way when the user drinks from the container.

The present invention relates to a novelty accessory for use with beverages. In particular, the present invention is directed toward a small attachable umbrella (xe2x80x9cBeerbrellaxe2x80x9d) which may be attached to a beverage container (e.g., glass or can of beer).
Beverage accessories are known in the art for use with beverage containers, be they glasses, cans, or bottles, and in particular with alcoholic beverages. Many of these accessories provide novelty amusement as well as practical effects, such as increased insulation to keep a beverage cold. Many of these accessories are sold or given away as promotional items for advertising purposes.
For example, the popular insulated beverage sleeve known as a xe2x80x9ccooziexe2x80x9d may be provided, manufactured of soft expanded polyurethane foam. These beverage sleeves are typically provided with an applied graphic advertising a beverage brand or the name of the company giving away the device as a promotion. A can, glass, or bottle may be inserted into the sleeve. The sleeve acts as an insulator to prevent ambient heat as well as heat from the user""s hands, from warming the beverage.
Similar devices are known for use specifically with bottles beverages. In this variation, a tailored expanded polyurethane jacket may be provided, replete with zipper, to encapsulate substantially all of a bottle.
Various devices are known for supporting beverages, such as coasters and the like as well as beverage stands, trays, and supports. One example is illustrated in Foley et al., U.S. Pat. No. 5,823,496, issued Oct. 20, 1998 and incorporated herein by reference. Foley provides an outdoor stand with a stake or pole which may be inserted into the ground to support a beverage container.
Similia, U.S. Pat. No. 4,638,645, issued Jan. 27, 1987 and incorporated herein by reference, discloses a beverage container cooler for receiving a single beverage container (e.g., can) and providing a location for ice or the like to cool the beverage.
One problem with these Prior Art devices is that although they do provide insulation for beverages, they do not shield the beverage from the direct rays of the sun. A beverage left out in the sun, even if insulated or cooled with ice, quickly warms due to the effect of the intense infrared radiation from the sun, particularly on hot, sunny summer days.
Thus, it remains a requirement in the art to provide a means for shielding a beverage from direct sunlight.
The present invention provides a small umbrella (xe2x80x9cBeerbrellaxe2x80x9d) which may be removably attached to a beverage container in order to shade the beverage container from the direct rays of the sun. The apparatus of the present invention has particular application for use in sunny locations (e.g., poolside, at the bear, outdoor bars, and the like). However, the apparatus of the present invention may also be used to prevent rain or other precipitation from contaminating a beverage.
In addition, the apparatus of the present invention may be used in other locations (e.g., indoors) purely as a decorative novelty item or the like.
The apparatus comprises a small umbrella approximately five to seven inches in diameter, although other appropriate sizes may be used within the spirit and scope of the present invention. The umbrella may be made from any one of a number of materials. A small paper-like umbrella may be used, with balsa framing (e.g., in a similar manner to a cocktail umbrella) if low cost and disposability is an objective. A solid plastic umbrella may also be inexpensively made and may be reusable and waterproof. More elaborate umbrella types such as cloth or the like may also be employed without departing from the spirit and scope of the present invention.
Suitable advertising and/or logos may be applied to the umbrella surface for promotional purposes. The umbrella may be attached to the beverage container by any one of a number of means, including clip, strap, cup, foam insulator, or as a coaster or the like. The umbrella shaft may be provided with a pivot to allow the umbrella to be suitably angled to shield the sun or for aesthetic purposes. In one embodiment, a pivot joint and counterweight may be provided to allow the umbrella to pivot out of the way when the user drinks from the container.