US3216999A - Carbocyclic cephalosporins - Google Patents

Description

United States Patent 3,216,999 COCYCLIC CEPHALGSPOS Edwin H. Flynn, indianapolis, Ind, assignor to Eli Lilly and Company, Indianapolis, Ind, a corporation of Indiana No Drawing. Filed Aug. 23, 1961, Ser. No. 133,333

4 Claims. (Cl. 260243) This invention relates to novel organic compounds and to methods for their preparation.

The novel compounds of this invention are represented by the following structural formula:

in which R taken alone, is OH, C -C acyloxy, or tertiary-amino, R is OH when R is OH, R is OH when R is C -C acyloxy, R is --O when R is tertiaryamino, R and R when taken together, are O, and R is a carbocyclic radical of the class consisting of cyclobutyl, cyclopentyl, naphthyl, adamantyl, and substitution products thereof.

Thus, R can be acetoxy, propionoxy, butyroxy, capryloXy, or the like; or N-pyridyl, N-pyrimidyl, trimethylamino, triethylamino, tributylamino, or other tertiaryamino group such as those produced by reaction of cephalosporin C with nicotine, nicotinic acid, isonicotinic acid, nicotinamide, 2-aminopyridine, 2-amino-6-methylpyridine, 2,4,6-trimethylpyridine, Z-hydroxymethylpyridine, sulfapyridine, 3-hydroxypyridine, pyridine-2,3-dicarboxylic acid, quinoline, sulfadiazine, sulfathiazole, picolinic acid, and the like.

R can be cyclobutyl, cyclopentyl, a-naphthyl, ,B-naphthyl, or adamantyl, or substitution products thereof, having in each case one or more chlorine, bromine, fluorine, iodine, nitro, trifiuoromethyl, C -C alkyl, or C -C alkoxy substituents upon the ring.

The structure and preparation of the adamantyl moiety are described by Stetter in Angewandte Chemie, 66, 217 (1954).

The novel compounds of the present invention are related to cephalosporin C insofar as they contain the 5,6- dihydro-2H-l,3-thiazine ring with a fused B-lactam ring in the 2,3 position which is characteristic of cephaloporin C. However, unlike cephalosporin C, which contains the 5-amino-N'-adipamyl group in the 7 position, the compounds of the present invention are characterized by a carbocyclic carboxamido group in the 7 position. Moreover, unlike cephalosporin C, which has a relatively low antibacterial action, the compounds of the present invention are highly effective antibacterial agents, capable of inhibiting the growth of numerous types of microorganisms in a variety of environments.

As will be observed from the formulas given above, the invention includes a variety of related compounds 3,215,999 Patented Nov. 9, 1965 having the bicyclic ring structure of cephalosporin C, but with variations in the substituent groups attached thereto. Among such compounds are those having the nuclei of the cephalosporin'type products known as cephalosporin C desacetylcephalosporin C, and cephalosporin C these nuclei being represented by the following formulas, respectively:

where Am represents a tertiary-amino radical, exemplified above. As will be seen from the above formulas, the nucleus of cephalosporin C includes a fused lactone ring, while the nucleus of cephalosporin C forms an inner salt or zwitterion.

As is the case with the penicillins, to which the compounds of this invention are in some degree related, numerous salts, esters, amides, and like derivatives thereof can be prepared by combination with nontoxic pharmaceutically acceptable cations, anions, alcohol residues, ammonia, and amines, and such derivatives are to be regarded as the full equivalents of the compounds disclosed and claimed herein, and accordingly are to be considered as within the scope of this invention.

For purpose of illustration, there can be mentioned several types of cationic salts which can be prepared from compounds containing the cephalosporin C nucleus, including, for example, water-soluble salts such as the sodium, potassium, lithium, ammonium, and substituted ammonium salts, as well as the less water-soluble salts such as the calcium, barium, procaine, quinine, and .dibenzylethylenediamine salts. Those compounds which contain the cephalosporin C nucleus do not form cationic salts but instead form anionic salts, i.e., acid addition salts, with strong acids such as hydrochloric, hydrobromic, phosphoric, sulfuric, and like acids' The following examples, together with the operating examples appearing hereinafter, will illustrate the types of compounds available in accordance with the present invention:

7-cyclobutanecarboxamidocephalosporanic acid 7-cyclopentanecarboxamidocephalosporanic acid 7-(2'-chlorocyclobutanecarboxamido)cephalosporanic acid 7- (2'bromocyclopentanecarboxamido cephalosporanic acid 7- (3 -fluoro cyclopentanecarb oxamido cephalosp oranic acid 7-(2'methylcyclopentanecarboxamido)cephalosporanic acid 7- 3 'methoxycyclopentanecarboxamido) cephalosporanic acid 7-a-nitro-B-naphthamidocephalosporanic acid 7-a-trifluoromethyl-fi-naphthamidocephalosporanic acid 7-rnethyladamantanecarboxamidocephalosporanic acid and the like, including the cephalosporin C and cepha losporin C analogues thereof.

cephalosporin C can be prepared by cultivating a cephalosporin C-producing organism in a suitable nutrient medium, as described in British patent specification 810,196, published March 11, 1959.

cephalosporin C is readily converted into cephalosporin C by heating with water under acid conditions, as described in Belgian Patent 593,777, published November 30, 1960. This removes the acetyl group from its point of attachment through oxygen to the methyl group in the position of the thiazine ring, and lactonization then spontaneously occurs, yielding the fused cyclic lactone.

Cephalosporin C is also readily converted into compounds of the cephalosporin C type by refluxing in aqueous solution with an excess of pyridine, for example, as described in Belgian Patent 593,777. The reaction is applicable in general to the tertiary amines, of which numerous examples are given above, yielding corresponding derivatives of the cephalosporin C type wherein the tertiary amine is attached to the methyl group in the 5 position of the thiazine ring, and forms an inner salt with the carboxyl group in the 4 position.

Desacetylcephalosporin C. is conveniently prepared by treating cephalosporin C With citrus acetylesterase for several-hours in aqueous phosphate buffer at pH 6.5-7 according to the method of Jansen, J ang, and MacDonnell, Archiv. Biochem., (1947),415-31.

From the various cephalosporin C compounds thus available, the corresponding nucleus is readily obtained by cleaving the 5'-amino-N-adipamyl side chain between its amido nitrogen and its amido carbonyl group. Thus, 7-aminocephalosporanic acid can be obtained by digesting cephalosporin C for an extended period in the presence of a mineral acid and in the absence of light, according to the method described in Belgian Patent 593,777.

The compounds of the present invention are prepared by acylation of the appropriate cephalosporin C nucleus, be it the nucleus of cephalosporin C itself or of cephalosporin C or cephalosporin C or other variant. Alternatively, compounds of the cephalosporin C C and desacetylcephalosporin C classes can be obtained by applying to appropriate 7-acylamidocephalosporanic acids the conversion procedures of Belgian Patent 593,777 and of Jansen et al. to produce compounds having the respective nuclei.

For the acylation of the 7-amino group of the cephalosporin nucleus, as defined above, any of the conventional acylation procedures can be employed, utilizing any of the various types of known acylating agents having a composition which yields the desired side chain.

A convenient acylating agent is the appropriate carbocyclic carbacyl chloride or bromide. The acylation is carried out in water or an appropriate organic solvent, preferably under substantially neutral conditions, and preferably at reduced temperature, i.e., above the freezing point of the reaction mixture and up to about C. In a typical procedure, 7-aminocephalosporanic acid or one of its derivatives as defined herein, together with a sufiicient quantity of sodium bicarbonate or other appropriate alkali to promote solution, is dissolved in aqueous 50 volume-percent acetone, the concentration of the 7- aminocephalosporanic acid being about 1 to about 4 percent by weight. The solution is cooled to around 0 to 5 C., and a solution of the acylating agent is added in about 20 percent excess, with stirring and cooling. The pH of the mixture can be maintained, if it tends to vary, around the neutral level by bubbling carbon dioxide therein. After addition of the acylating agent has been completed, stirring of the reaction mixture is continued, and the mixture is allowed to warm to room temperature. The reaction product is then acidified to around pH 2 and extracted with an organic solvent such as ethyl acetate. The ethyl acetate extract is adjusted to around pH 5.5 with a base containing the desired cation of the final product, and is extracted with water. The water solution is separated and evaporated to dryness. The residue is taken up in the minimum quantity of water, and the desired product is precipitated by adding a large excess of acetone and, if necessary, ether. The crystalline prod- 4 not obtained thereby is filtered, washed with acetone, and dried.

Acylation can also be carried out with the corresponding carbocyclic carboxylic acid, employed in conjunction with an equimolar proportion of a carbodiimide such as N,N-diisopropylcarbodiimide, N,N'-dicyclohexylcarbodiimide, N,N bis (p dimethylaminophenyl)carbodiimide, N-ethyl N (4"-ethylmorpholinyl)carbodiimide, or the like, and the acylation proceeds at ordinary temperatures in such cases.

Alternatively, the carbocyclic carboxylic acid can be converted into the corresponding acid anhydride, or into the azide, or into an activated ester, and any of these derivatives can be used to efiect the desired acylation. Other agents can readily be ascertained from the art.

In many cases, the acylating agent may contain one or more asymmetric carbon atoms and thus exist in optically active forms. When prepared by ordinary chemical means, such compounds are ordinarily obtained in racemic formi.e., an equimolar mixture of the optical isomers, having no optical rotation. When the separate optical isomers are desired, the acylating agent can be resolved in a conventional manner such as by reacting the free acid with cinchonine, strychnine, brucine, or the like, then fractionally crystallizing to separate the diestereoisomeric salts, and separately acidifying the solid phase and the liquid phase to liberate the optical isomers. The free acids thus obtained can be employed as such for the acylation, preferably in conjunction with a carbodiimide, or may be converted by conventional means into the corresponding acid halide or into a mixed anhydride, care being exercised to avoid extremes of conditions which might produce racemization.

Many of the acylating agents, together with methods for their preparation, are described in the literature, and a number of them are commercially available. All of them are readily prepared by methods well known in the art.

The invention will be more readily understood from the following operating examples, which are submitted as illustrations only, and not by Way of limitation. The chemical assays reported herein were carried out by the method of Ford, Analytical Chemistry, 19, 1004 (1947), which is based upon the quantitative determination of the B-lactam moiety of the cephalosporin molecule via reaction with hydroxylamine. The antibiotic potencies were determined against Staphylococcus aureus 209 P by an appropriate modification of the paper disc plate methods of Higgens et al., Antibiotics & Chemotherapy, 3, 50-54 (January 1953) and Loo et al., Journal of Bacteriology, 50, 701-709 (1945). The pKa values were determined by titration in aqueous 66 percent dimethylformamide.

Example 1.7-a-naphthamidocephalosporanic acid 7-aminocephalosporanic acid (1.0 g.) and sodium bicarbonate (680 mg.) were dissolved in a mixture of 50 ml. of water and 40 ml. of acetone. The resulting solution was cooled in an ice bath, and to it was added a solution of 670 mg. of a-naphthoyl chloride in 10 ml. of acetone over a period of minutes, after which stirring was continued for 1.5 additional hours. The reaction product mixture was stripped of acetone, layered with ml. of ethyl acetate, and acidified to pH 2.0 with hydrochloric acid. The ethyl acetate layer was separated and back-extracted into water at pH 5.5, the pH adjustment being carried out with dilute aqueous potassium hydroxide solution. The aqueous extract was concentrated to a syrup and solidified by dilution with acetone, and the precipitate was recrystallized from a mixture of methyl and ethyl alcohols. The product was 250 mg. of 7-a-naphthamidocephalosporanic acid in the form of the potassium salt, having a pK'a of 4.85, a chemical assay of 1330 penicillin G units per milligram, and maxima in its ultraviolet absorption spectrum at 219 and 269 my. (e=47,600 and 10,200, respectively).

Example 2.-7-,8-naphthamidocephalosporanic acid 7-aminocephalosporanic acid (1.0 g.) and sodium bicarbonate (1 g.) were dissolved in a mixture of 50 ml. of water and 40 ml. of acetone. The solution was stirred in an ice bath and to it was added a solution of 670 mg. of ,B-naphthoyl chloride in ml. of acetone over a period of about 30 minutes, after which the mixture was stirred between 2 and 3 additional hours in the cold. The reaction product mixture was then stripped of acetone under vacuum, and 100 ml. of ethyl acetate were added, followed by 1 N hydrochloric acid to adjust the mixture to pH 2. The aqueous phase was separated, washed with 50 ml. of ethyl acetate, and discarded. The ethyl acetate layers were combined and washed with 50 ml. of water. The washed ethyl acetate phase was stirred with 100 ml. of water and adjusted to pH 5.5 with aqueous 0.5 N potassium hydroxide solution. The resulting aqueous extract was separated and evaporated to dryness under vacuum. The residue was triturated with aqueous acetone, and the solids were filtered ofi and dried under vacuum. The yield was 440 mg. of 7-fl-naphthamidocephalosporanic acid in the form of the potassium salt, having a pK'a of 4.90 and a maximum in its ultraviolet absorption spectrum at 236 m (e=46,300), with shoulders at 269 and 330 mg.

The product had a chemical assay of 1510 penicillin G units per milligram and a bioassay potency of 43 penicillin G units per milligram. It was effective against resistant staphylococci in broth, and also showed good activity against Staphylococcus albus, Bacillus subtilis, and Sarcina lutea.

Example 3.7-adamantanecarboxamidocephalosporanic acid 7-adarnantanecarb0xamidocephalosporanic acid was prepared in the form of the potassium salt by reacting 7-aminocephalosporanic acid (1.0 g.) with sodium bicarbonate (l.0 g.) and adamantanecarboxy-l chloride (700 mg.) generally according to the procedure of Example 1, the reaction mixture being allowed to stand for 12 hours. The product, recrystallized from methanol ether, weighed 300 mg. and had a pK'a of 4.90 and a maximum in its ultraviolet absorption spectrum 6 at 260 m (e=7,750). It had a chemical assay of 1260 penicillin G units per milligram.

I claim: 1. An antibiotic substance of the class represented by the following formula:

wherein 2. 7-anaphthamidocephalosporanic acid. 3. 7-B-naphthamidocephalosporanic acid. 4. 7-adamantanecarboxamidocephalosporanic acid.

References Cited by the Examiner UNITED STATES PATENTS 6/60 Doyle et a1. 260-239.1

FOREIGN PATENTS 593,777 12/60 Belgium.

OTHER REFERENCES Wertheim, Textbook of Organic Chemistry, pages 301- 302 and pages 763-764 (1945).

NICHOLAS S. RIZZO, Primary Examiner.

Claims (1)

1. AN ANTIBIOTIC SUBSTANCE OF THE CLASS REPRESENTED BY THE FOLLOWING FORMULA: