US3275511A - Combined antibiotic and mycocide - Google Patents

Description

United States Patent Original application Aug. 23,, 1957, Ser. No. 680,022.-

Divided and this application Mar. 22, 1966, Ser. No. 536,952-

14 Claims. (Cl. 16758) This is a division of co-pending application Serial No. 680,022, filed August 23, 1957, in the name of Charles G. Shaw.

This invention relates to a new and useful class of compounds which chemically combine an antibiotic substance with a mycocidal substance.

Specifically, the invention contemplates the reaction product of such antibiotic substances as Neomycin, Streptomycin, Terramycin, Aureomycin, and others with mycostatic and mycocidal agents such as the halide-substiuted phenols, in order that the spectrum of the resultant antibiotic-mycostatic or antibiotic-mycocidal compound is greatly enhanced to include broader spectrums of mycocidal and bactericidal activity. Aureomycin and Terramycin are trade names for 7-chlortetracycline and S-hydroxytetracycline respectively.

There has been a considerable amount of work accomplished in developing the phenol derivatives for use as fungicidal or nycostatic agents. Some of the phenols have a toxicity which makes them therapeutically valueless, but even of those mycocodal or mycostatic compounds which are non-toxic none appears to have a potent bactericidal spectrum in addition to the mycocidal spectrum.

Similarly, there is an enormous background of literature and information concerning antibiosis. It is stated by McVickar in Therapy of Fungus Diseases, Sternburg and Newcomer, Little-Brown 1956: None of the antibiotics employed so effectively in the treatment of bacterial and rickettsial infection is effective in the treatment of mycotic infections, systemic or superficial. In the same volume, Waksman refers to antibiotics active against fungi but inactive against bacteria, and with respect to their activity against fungi, highly selective in their actions against different fungi. These spectrum deficiencies are notable in all antibiotics, even those having a fringe action against either bacteria or fungi. There appears to be no. prior art relating to the actual combination in one, molecule of a compound effective in a wide spectrum as both an antibiotic and a mycostatic agent.

It is important to emphasize that the compounds contemplated in the present invention are those wherein a mycostatic agent and the antibiotic agent are chemically reacted together to form a single compound effective in a broad spectrum, wherein there is a chemical bond between the two agents. There may be known methods of medical treatment involving the simultaneous application to a certain body area of a mycostatic agent and an antibiotic agent, but this is not within the scope of the invention and as indicated above there is no known,

case where a mycostatic agent and an antibiotic agent have been chemically compounded for medical or other use.

The advantages of a complex compound of a mycostatic and antibiotic agent are many fold. One important advantage is in the control of solubility and diffusion rates. It is a known fact that the effectiveness of a mycostatic and antibiotic agent is partly dependent on the rate at which the compound diffuses into the body 3,275,51 l Patented Sept. 27, 1966 tissues. Depending on the nature of the area being treated with the treatment desired, various means are employed to vary the rate of diffusion of the compound into the tissues. If, for instance, to a certain area one. wishes to apply separately or in mixture form an antibiotic agent and a mycostatic agent, different solubilities of the two agents would affect the diffusion rates of the two agents and the two components would not remain in the infected area. in the desired proportions. There are many physio-chemical properties of they components other than their solubilities which would. similarly affect application. Chemically combining the antibiotic and mycostatic agents would give them common physiochemical properties including a common diffusion rate and would considerably simplify the control of. the application technique.

A second advantage linked with the above: is the ability in the complex compound to control the. relative bactericidal and mycocidal effectiveness. Briefly, if it. were desirable to achieve an application to the infected area of a mycocidal agent and bactericidal agent in the ratio of one to two, this could easily be facilitated by the methodv herein described, by simply chemically combining. the desired components in the desired ratio. By means of. the chemical link, one is assured of the continued maintenance of this ratio to the infected area.

A further advantage may beseen in what is termeda synergistic effect.

A further advantage is apparent in the fact that a new means is available for therapeutically using the highly toxic mycostatic agents such as pentachlorophenol by means of this bond with an antibiotic, a means which permits even greater concentrations. of the mycostatic agent with less toxicity than previously obtained by buffering techniques or like means of toxicity control.

Having briefly outlined the concepts and advantages of the invention, it; isthus a primary object to provide a new chemical compound having an antibiotic component, wherein the compound will have a considerably extended effective mycocidal spectrum.

It is further an object to provide means in a single chemical compound simultaneously effective against both bacterial and mycoorganisms wherein the relative bactericidal and mycocidal potencies or spectrum can be. varied or adjusted depending on the treatment desired.

It is further. an object to provide a means for controlling the pH of the compound, and solubility of the compound, and for controlling and equalizing the diffusion rates of the components and for minimizing toxic effect or sensitivity of certain components of the compound.

It is another object to provide a compound having a more effective control of the pathogenic fungi and the bacteria than would be obtainable by the separate but.

simultaneous use of the individual components of the compound.

The invention has for another object, the provision of a compound for therapeutic application, which would also be effective against food spoilage and plant diseases, and as an animal food supplement.

Further objects and advantages will become apparent from the following description and examples of the invention.

Microbial activity may be divided into many classifications, by the genus of the involved organism; or the a species, or mutation, or pleomorphic form, etc. Various arts ascribe certain organisms into varying categories and all arts related to the use or control of micro-organisms are concerned with the relative activity or control of many genuses and species. To illustrate, but not in a definitive sense, the following classification will suffice to arrange pathogenic organisms from bacteria to costatic properties. I g benzenecan be formulated into new and .useful mycostatic f; chemotherapeutic agents but none of the agents disclose a 3 mentioned antibiotic-mycocide complexes.

I an extremely stable chemical bond, either directly with an fungi in a spectrum serving as a guide in designating bacteria, Gram-positive bacteria, Coccidae, Mycobaciteria, Actinomycetes, Myco-pathogens, Leptothricyia, Algae and Animalculae.

, There are several other borderline species unknown in the literature termed Catathricyia and complex mycotic }tion by Dr. J. W. Papez at Columbia State Hospital, Co- ;lumbus, Ohio, and Dr. Irene Diller, at the Institute of Cancer Research in Philadelphia.- From studies of the 1microbiota of the oral cavity, these borderline species apitity, some apparently pathogenic in form,-in chronically :granulomatous conditions and in other forms of lesions. They include the extremely fine filamentous form which I ihave called Cata-thrix, and which is now under study in dark field and electron microscopy. It is of a fungus like nature and appears to have pleomorphic spore forms and lto segment into virus like free forms. Certain species of a very 'malodorous and disagreeable feature in the syndrome of such diseases as virus hepatitisthese and cer- }tothricia are readily controlled by therapeutics resulting f of the broadest possible action are desirable, as many diseases evidence symbiotic and complicated infections. 1 Those involving bacteria and myco-pathogens are freitive against all pathogenic organisms. None .has been J tibiotics provide a multiplicity of efiective therapeutics '1 over quite broad portions of the pathogenic spectrum.

l The known antibiotics, originating from the metabolism of micro-organisms, which are effective as bactericides and/ or fungicides, are too varied and obscure in chemical constitution to permit complete categorical classification. However, generally they are in the nature of organic acids or bases and/or other complexes capable of combining i with acidic or basic radicals to form salts. All such anti- 3 biotic salts are within the scope of this invention.

M In the art there is also considerable reference to mycocidalorganic chemical compounds such as the halogensubstituted products of phenol and halogen-substituted compounds of benzoquinone, hydroquinone, and phloroglucinol. There are further many other aromatic halidesubstituted compounds which, either as bare acids, or their 3 metallic or organic salts have marked fungicidal or my- Many other. aromatic derivatives of broad spectrum of bactericidal properties. It may be suff ficient herein to emphasize that many of these halogensubstituted compounds of the various phenols are quite i toxic and produce highly objectionable sensory reactions requiring careful solubility control, pH'control and limited i concentration of the effective ingredient. It is important, i however, to note that the strong acidic properties of the f more highly saturated substituted products of the phenols make them highly desirable in the preparation of the above antibiotic base or indirectly with an acidic antibiotic through a dior tri-valent base such as calcium or his- 1 muth. The strength of this bond between the antibiotic l i substance and the mycocide' complex should be such that it cannot be broken under the conditions in which these new complex compounds may be used'and in many in-. stances a very'delicate balance of the pH inherent in its' I solutions, should be provided .without recourse to supple- V mentary buifering. The substituted mycocidal phenol derivatives can also therapeutic control-viruses, Rickettsia, Gram-negativev forms, now undergoing investigation in my laboratory in connection with dental lesions and those under investiga-.

jpearto be many genuses and species of questionable iden-.

Torganisms are associated with the tongue coating which is jltain others which appear to be pleomorphic forms of Lepquently encountered. An ideal therapeutic would be ac-.

j discovered, though a constantly increasing number of an- They provide Efrom this compound. Antibiotics and chemotherapeutics be reacted with the antibiotics through the linkage of a polybasic strong acid, or by forming a mixed crystal .com plex with a polybasic acid. These concepts are disclosed in the examples which follow.

It should merely be emphasized that all mycostatic agents including the halide substituted benzene derivatives, and means for linking. said agents with the antibiotics are within the scope of this invention.

In all of the antibiotic-mycocide compounds disclosed in the examples which follow and in all those within the scope ofthe inventiomthere is a chemical linkage. or .bond between the mycocidal agent, and the antibiotic. .This bond may involve the presence of a polybasic acid or-salt, or a crystalicomplex. valent base or acid, orsalt having an afiinity or ability to react with one or several molecules of the mycocidal agents. By controlling the reaction, one can leave available valences or linkages onthe antibiotic molecule or elsewhere in the resultant molecule for reaction with acidic or basic molecules to controlthe pH of the final compound. For instance, Neomycin has an afiinity or ability to react with fromone to six molecules of pentachloro phenol, replacing the hydrogen ion of the hydroxyl group of the benzene acid. If the reaction is controlled such that I only four of the six valences of'Neomycin are utilized,

two active valences still remain for reaction with acidic radicals to further control or. adjust the. pH- to a desired level.

The following examples illustrate the concepts of the, invention and representative means for compounding the therapeutic agents contemplated.

It should be noted that even though'several of the examples specify carrier agents or means of application, the. invention is not to. be limited to such agents or. means. In these examples, and in examples where no specific car- 1 rier or means of application is indicated, it will beundcra stood that the ant-ibiotic-mycocide chemical. compounds j can be compounded into solutions, ointments, dusting pow ders, dips, sprays, etc., there being a wide choice. of. ye-

hicles and solventswell known in the arts, to accommo-.

date the varying physio-chemical. properties including solubility of the many antibiotic-mycocidal chemical compo-' nents and further to affect and control diffusion rates depending on the desiredapplicationand nature of t'reatment. =Dispersion on kaolin, talc, and other. suitable ve-' hicle and solvents are obvious.

Example I.-Neomycin hexa-pentachlorophenate Pentachlorophenol is a mycostaticagent which has been Neomycin, a known antibiotic, when chemically combined with pentachlorophenol produces a compound according to shown tobe highly effective against many fungi.

the invention which is highly effective against a wide spectrum of myco-organisms and bacteria.v

The highly basic properties of the neomycin base permit it to react with six molecules of hydrochloric acid to 1 produce a Neomycin chloride salt at about 5.2 pH, the

molecular weight of Neomycin being calculated as 774.8

six valences of the Neomycin base mayibe utilized .to chemically combine with only four acidicpentachlorophenol molecules to produce an effective relative proportion.

Two active valences will remain for, more or less acidic acid radicals adjusting the final pH of the ;whole .compound to a desirable level. The possibilities of pH adjustment' and control of the relative mycostatic and bacteriastatic potencies are apparent.

Generally the antibioticvis a multi- Similarly, Neomycin base can be reacted with six It is not Neomycin hem-pentachlorophenate is prepared in the following manner:

Dissolved in 4.0 cc. water. 2 Dissolved in 20.0 cc. 95% ethyl-alcohol.

The Neomycin and pentachlorophenol solutions are mixed by pouring the pentachlorphenol solution into the Neomycin base solution. The two react at room temperature to form Neomycin hexa-pentachlorophenate. To this is added 60 cos. of Eugenol.

The preparation is useful in root canal therapy and the treatment of periodontal pockets with dressings and topical applicaitons. The preparation may also be used in a well-known periodontal pack comprising zinc oxide, rosin, and tannic acid.

Neomycin hem-pentachlorophenate has bactericidal properties corresponding with those of the Neomycin'sulfate or chloride salt, including activity upon Staphylococcus albus' and- Staphylococcus aureus, Streptococcus fecalis and Mycobacterium tuberculosis, and mycocidal properties with Candida albicans and many other fungi, as also with such organisms as Leptothricia and similar species associated with gingival lesions and tongue coatings.

The compound is highly effective against both bacteria and fungi and in addition against borderline species known in the art. It should be noted further that the individual spectra of both of the components is in no Way. adversely affected, and that the toxicity of the pentachlorophenol is practically eliminated.

Example II.-Streptomycin-tri-pentachlorophenate I Pentachlorophenol, M.W. 266.35, combines with 1 mol. NaOH 40.01 gms. to produce the sodium salt, M.W. 288.34. A /2 mol. solution is prepared of sodium pentachlorophenate in water. A /2 mol. solution of silver nitrate, M.W. 169.89, also in. water, is prepared, and

equivalents of the two above solutions are mixed together with stirring. The resulting reaction produces a silverp'entachlorophenol' salt. Due to the nature of the bond of the substituted chlorine in the pentachlorophenol molecule, there is no. interaction between the silver nitrate and the substituted chlorine, but merely a substitution of the silver ions for the sodium ions. The salt is alcohol soluble.

Streptomycin hydrochloride, according to Merck, p. 907,

. Grams Silver pentachlorophenate 1 1119.66

Streptomycin tri-hydrochloride 2 690.003

Dissolved in alcohol.

-'- Dissolved in water.

The reaction between the silver and the chloride occurs at room temperature and progresses quickly such that no heat or catalyst is necessary. The Streptomycin tri-pentachlorophenate (net weight 1380.63 grams) remains in solution. in the alcohol water mixture and thecloudy silver chloride precipitates and is removed. The solution can be dehydrated or concentrated for storage or application. The resultant antibiotic-mycostatic compoundis fully effective against the known spectra of the components, and in many instances the spectrum is enhanced.

Example III'.Di-hyaro Streptomycin tripentachl0r0- phenate This reaction can be carried out with di-hydro Streptomycin sulphate as follows:

BaCl 2H O gms 244.31 Sodium pentachlorophenate ("/2 mol. solution) gms 532.70 The typical Barium reaction results are:

Barium (pentachlorophenateh gms 670.06

The precipitate is insoluble in water but will dissolve in alcohol.

Di-hydrov Streptomycin base (calculated from the sulfate salt) dissolved in water, 1 mol. grns 583.59 Ba (pentachlorophenolh dissolved in alcohol,

1 /2 mols. gms 1005.09

Example 1V.Phlortetracyline mono-pentachlorophenate Aureomycin, termed chlortetracycline hydrochloride, has a chloride ion available for reaction with the acidic mycostatic agent.

Aureomycin hydrochloride 543.000

is capable of reacting with Silver pentachlorophenate 373.22

to produce Aureomycin mono-pentachlorophenate 773.35

The product is not stable in aqueous solution over long periods. It is also bland but very efiective on organisms associated with certain periodontal disease conditions and on dermal mycosis. It has a virus and rickettsial inhibitory capacity valuable in treating infections which fre' quently complicate oral septic conditions.

Example V.Tetracycline mono-pentachlorophenate Similarly tetracycline hydrochloride, M.W. 506.535, is capable of reacting with silver-pentachlorophenate, M.W.

, 373 .22, to produce tetracycline mono-pentachlorophenate,

This product is also bland and very effective on organisms associated with certain periodontal disease conditionsv and on dermal mycosis. The spectrum of neither component is adversely affected by the chemical bond and is probably enhanced in each instance. There is also definitely produced a virus and rickettsial inhibitory capacity, valuable in treating oral infections.

An acid such as pentachlorophenol, considered much weaker than hydrochloric acid, nevertheless reacts with the six valences of Neomycin base in a manner surprisingly similar to the reaction of the base with hydrochloric acid. The pH of many phases of their neutralization curves is practically coincidental. In this sense, the pentachlorophenate ion has an effective. aflinity similar to the sulphate or chloride ion.

Many of the halo-substituted aromatic mycostatic agents less saturated and acidic in nature are incapable of elfecting a permanent bond with an antibiotic base.

For instance, 2,4,6-trichlorophenol with a Ka. of

7 2.6 is incapable of neutralizing the basic properties of Neomycin base, and yet trichlorophenol and other similar mycostatic compounds have highly desirable and effective mycostatic properties.

It has been found, however, that any of the weak acidic halide substituted phenols can be reacted with a highly acidic mineral acid such as sulphuric and phosphoric acid to combine it with the. first hydrogen, radical of the mineral acid,thus leaving the remaining acidic ions of the mineral acid to form a tenacious bond with a basic antibiotic substance such as Neomycin.

This provides an alternative and very important means for linking the highly saturated mycostatic. compounds with an antibiotic. toward the utilization of pentachlorophenol, but it is to be understood that the same principles can be applied as a means for linking the above mentioned less highly saturated acidic mycostatic substances with the antibiotics. Furthermore, the following example, utilized acetylsalicylic acid but it is to be understood that any of the mineral polybasic acids can be used.

The following example is directed.

Example VI.-C0mplex of Neomycin and salicylic acid pentachlorophenolate It was desired to combine pentachlorophenol with Gms.

Pentachlorophenol 26.635 Acetylsalicylic acid 18.015

were dissolved in ethylene di-chloride and hydrochloric acid and heated resulting in a stable clear solution of Salicylic acid pentachlorophenolate grns 40.346

The following reaction is believed to have occurred.

l or

COOH COOH C o c 0 on: o 01 V V c The carboxylic group is probably left free to .react with the Neomycin base. In any event, it is quite certain that the phenol does not substitute into the aromatic ring of the salicylicacid. Accordingly,-it is appropriate to identify the reaction product as a phenolate.

The salicylic acid pentachlorophenolate was then re- In the reaction, pentachlorophenol' acted with Neomycin base in the ratio of 5 mols. to 1 mol.

respectively.

Salicylic acid pentachlorophenolate grns .1606 Dissolved in non-ionic Tween (Merck) gms .15 H O ccs 10 Alcohol (95%) ccs 40 To this solution was added:

Neomycin base "ms .0775 H O ccs 25 g The resultant compound, was diluted in 200 ccs. of

water, giving a pH of 6.21, and was satisfactory for use in a topical dermal dress incorporated into a nasal spray or topical dermal dressing in the following manner.

r Gms. Mixed crystal of aspirin and entachlorophenol. .1339 Menthol .078 1 Non-ionic Tween (Merck) 7.5

Since the aspirin-pentachlorophenolate is tender, it is preferable to pour the base slowly into the acidic solution to prevent any reaction which might rupture the phenolsalicylic bond.

As indicated above, the pH of the final compoundcan be adjusted by further linking of acidic ions to the antibiotic base. It is,desirable, however, that theacidic ions be of not too great relative acidic strength. Forexample,

sulfate and chloride ions would tend to rupture the phenol salicylic bond, but an acid of equivalent strength to the:

salicylic acid can be used. a

One. can appreciate the variety, of compounds obtain: able with the reactions described above. For instance,

acid andterephthalic acid have a sufficient spread with pchlorophenol, in ionization, to form tenable bonds with it. Many similar compound acids and acid salts possessing high fungicidal properties can thus be formedand utilized to fall within the scope of the invention.

Example VII .Bufiered N eomycin tri-pentachlorophenate As an example of the pH bulfering aspect of the invention, Neomycin tri-pentachlorophenate can be formed to leave available threevalences of the Neomycin base for reaction with a buifering acid or acid salt.

Neomycin base gms -..0775 I Pentachlorophenol gms" .0799 Water ccs-- 5 Alcohol ccs '5 The pentachlorophenol is added to the Neomycin base solution resulting inthc tri-pentachlorophenol base with a pH. of about 7.5. To this solution is added .5 .cc. of .5 NHCI which forms a temporary precipitate and .99 cc. of .5 NHCI progressively thereafter to occupy the remaining 3 valences of Neomycin with the chloride ion. The i resultant solution has a pH about, 5.5. Three mols...of

/2 neutralized o-phthalic, acid or.p-phthalic acid could,

be reacted with the above salt to depress the pH' to between .6 and 5.2, equivalent to the hexa-pentachloride of Example I.

Similarly, many other compound variations such as Neomycin mono pentachlorophana-te pentasulfate or pentachloride can be formed.

2 Example VIlI.Non-alcol1olic nasal spray and topical dermal dressing I i Gms.: Acetylsalicylic acid 1.8015 Pentachlorophenol 2.6635

The two above components were dissolved in acetone and the solution heated to from" F. to F. tor, The residualmass was a crys-. talline'mixture of one mol. each of entachlorophenol evaporate the acetone.

and acetyl-salicylic acid (molecular weight446.50), and

was not a molecular compound as illustrated in Example The above was dissolved in a small amount of water,

about 25 cc., into whichthe Neomycin basewas added;

and stirred at room temperature.

98% pure Neomycin base gms .775 Dissolved in water cc The above formula is based on a proportion of 3 mols. each of aspirin and pentachlorophenol to 1 mol. of Neomycin base. It is apparent that all 6 molecules of the aspirin and pentachlorophenol are free to react and do react with the Neomycin. There is in this manner provided a means of simultaneously effecting the mycostaticantibiotic reaction and adjusting the pH of the resultant solution. It should be noted that any number of appropriate acids or salts can be similarly incorporated into the resultant compound for pH control.

The above solution when diluted with 50 to 100 cc. of water to a desire-d concentration, results in medicament having a quite bland sensory reaction in application and a bacteriostatic and mycostatic spectrum broadly equivalent to Neomycin tri-pentachlorophenate.

Example IX The acidic antibotics such as penicillin can be combined with calcium or bismuth or any strong bior trivalent base to form a basic salt. The remaining valences of the base can then be used to link the penicillin with the mycocidal acidic compound.

This can also be done by reacting 1 mol. of calcium with 1 mol. of pentachlorophenol to form a basic salt which can then be reacted with penicillin.

Example X Tetrachloroquinone, being an oxygen donor, will combine with the antibiotic base, but apparently attacks it by oxidation. By using various proportions of tetrachloro hydroquinone with tetrachloroquinone, the well known quinhydrone reaction takes place, thus modifying the oxidation tendency, and a stable \bond can be formed between the base and the mycostatic substance.

I claim:

1. A method of medical treatment comprising topically applying to tissue 'a complex of an antibiotic selected 'from the group consisting of Streptomycin, iii-hydro Streptomycin, Neomycin, tetracycline, oxytetracycline, chlortetracycline, and a basic salt of penicillin complexed with a halogenated phenol having the formula:

wherein R is selected from the group consisting of hydrogen, halogen and hydroxy and wherein :at least one R radical is always halogen.

2. A method according to claim 1 wherein the antibiotic is selected from the group consisting of calcium and bismuth salts of penicillin.

3. A method according to claim 1 wherein the antibiotie is selected from the group consisting of acid salts of tetracycline, oxytetracycline and chlortetracycline.

4. A method according to claim 1 wherein the antibiotic is selected from the group consisting of mineral acid salts of tetracycline, oxytetracycline and chlortetracycline.

5. A method according to claim 1 wherein the antibiotic is selected from the group consisting of Streptomycin and dihydro Streptomycin.

6. A method according to claim 1 wherein the antibiotic is Neomycin.

7. A method according to claim 1 wherein the anti biotic is tetracycline.

8. A method according to claim 1 wherein the antibiotic is oxytetracycline.

9. A method according to claim 1 wherein the antibiotic is chlortetracycline.

10. A method according to claim 1 wherein the complex is Neomycin hexapentacholrophenate.

11. A method according to claim 1 wherein the complex is Neomycin triphentachlorophenate. V

12. A method according to claim 1 wherein the complex is tetracycline monopentachlorophenate.

13. A method according to claim 1 wherein the complex is chlortetracyclinemonopentachlorophenate.

14. A method according to claim 1 wherein the halogenated phenol is pentachlorophenol.

No references cited.

LEWIS GOTTS, Primary Examiner.

SHEP K. ROSE, Assistant Examiner.

Claims (1)

1. A METHOD OF MEDICAL TREATMENT COMPRISING TOPICALLY APPLYING TO TISSUE A COMPLEX OF AN ANTIBIOTIC SELECTED FROM THE GROUP CONSISTING OF STREPTOMYCIN, DI-HYDRO STREPTOMYCIN, NEOMYCIN, TETRACYCLINE, OXYTETRACYCLINE, CHLORTETRACYCLINE, AND A BASIC SALT OF PENICILLIN COMPLEXED WITH A HALOGENATED PHENOL HAVING THE FORMULA: