IE44393B1

IE44393B1 - New chephalosporins

Info

Publication number: IE44393B1
Application number: IE2377/76A
Authority: IE
Original assignee: Smithkline Corp
Priority date: 1975-10-30
Filing date: 1976-10-27
Publication date: 1981-11-18
Also published as: IE44393L; FR2337720A1; GB1570094A; FI60868C; AU505141B2; SE8006413L; FI60868B; FR2351982B1; LU76092A1; GB1570093A; GR61667B; SE437025B; AU1870276A; SE7610722L; GB1570095A; SE452621B; CA1071188A; ES452769A1; FR2337720B1; NL7612010A

Abstract

The cephalosporin derivatives of the formula I in which the symbols have the meanings stated in Claim 1 are antibiotics. They are prepared from the 7-acylaminocephalosporanic acid of the formula II by replacement of the acetoxy group by the appropriately substituted 5-mercaptotetrazole.

Description

This invention relates to cephalosporin compounds which have antibacterial activity, to processes for the preparation of such compounds, and to compositions containing them.

The compounds of this invention are represented by the 5 formula: (I) where W is hydrogen or methoxy; fP is an acyl group selected from A I X —CH -CO—, Y-CHgCO- and Z-S(0)m-CH2-C0-X is thienyl, di hydrophenyl, phenyl or phenyl monosubstituted with hydroxy, hydroxymethyl, formamido, ureido or carboxymethylamino; A is NH2, OH, COOH or SOgH; or formyloxy when X is phenyl; Y is thienyl, tetrazolyl, sydnon-3-yl, cyano or aminomethyl phenyl; Z is methyl, trifluoromethyl, trifluoroethyl, cyanomethyl or pyridyl; and m is zero, one or two and Ιθ· jj is an integer from two to five.

This invention also includes pharmaceutically acceptable salts thereof and esters of the 4-position carboxylic acid group. The 4-position carboxylic acid group can be readily esterified by methods well known in the art. These esters include, for example, simple alkyl and aryl esters as well as esters which are easily cleaved, within the body, to the parent acid such as indanyl, pivaloyloxymethyl, acetoxymethyl, propionyloxymethyl, glycycloxymethyl, phenylglycycloxymethyl and thienylglycyloxymethyl esters. Of course, with A is COOH, this group can also be esterified, and such esters are included within the scope of this invention.

A selected group of compounds of this invention is represented by Formula I where W is hydrogen and jj is two.

Another group of compounds of this invention is represented by Formula I where U is hydrogen, jj is two, X is phenyl, A is NH2 or OH, ---1 443S3 Y is thienyl or tetrazolyl, Z is trifluoromethyl, and m is zero.

Examples of representative 7-acylamino substituents (pJfiH) of the compounds of Formula I are listed below. α-hydroxyphenylacetamido . α-aminophenyl acetamido a-amino-4-hydroxyphenylacetamido trif1uoromethylthioacetamido methylthioacetamido α-carboxythienylacetamido α-carboxyphenylacetamido α-sulfophenylacetamido ; a-anrino-4-carboxymethyl aminophenyl-acetamido 2-aminomethylphenylacetamido syndon-3-ylacetamido tetrazolylacetamido ' thi enyl acetami do Z,2,2-trifluoroethylsulfinylacetamido - cyanoacetaraido methylsulfonylacetamido Cyanomethylthioacetamido 4-pyridylthioacetamido Some examples of the compounds'of Formula I are 7 - D - mandelamido - 3 - Jj - (Z - sulfaminoethyljtetrazol - 5 ylthiomethyfj - 3 - cephem - 4 - carboxylic acid, 7-(2- thienylacetamido) -3-0-(2- sulf ami noethyl) - tetrazol - 5 - ylthiomethyf] - 3 - cephem - 4 - carboxylic acid, 7-(1- tetrazolyl acetamido) - 3 - El - (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid and - trifluoromethylthioacetamido -3-0-(2- sul faminoethyl) tetrazol - 5 - yl thiomethyl] - 3 - cephem - 4 - carboxylic acid.

Cephalosporin derivatives having 7 - acetylamino substituents as defined above are well documented in the prior art. Although substitution by variously substituted S-heterocyclicthiomethyl groups (-CH2Het) at the 3-position of the cephem nucleus is also known, no compounds containing the 3-(sulfaminoalkyl-substituted tetrazolyl) thiomethyl moiety disclosed herein are believed to be known to the art.

The compounds of Formula I where W is hydrogen can be prepared by acylating 7-aminocephalosporanic acid with an appropriately protected acylating agent, and then displacing the 3-acetoxy group of the acid with the desired sulfaminoalkyltetrazolethiol or a salt thereof, with subsequent removal of the protective group(s) when present. The sulfaminoalkyltetrazole-thiols and salts thereof are described and claimed in our Patent Specification No. 1644/80.

The carboxylic acid group of the acylating agent is activated by any of the standard methods such as conversion into the mixed anhydride, acid chloride, acid imidazolide or activated ester. In addition, a reagent such as dicyclohexylcarbodiimide can be used, provided that the carboxyl group on the cephem nucleus is protected with an easilyremovable protecting group, such as benzhydryl, t-butyl, trichloroethyl, benzyl, benzyloxymethyl, jj-methoxymathyl, £-methoxybenzyl or p-nitrobenzyl. Thereafter, the protecting group can be removed by known methods. When A is NHg> the α-amino group of the acylating agent is preferably protected, prior to acylation, with an easily removable protective group such as jt-butoxycarbonyl, trichloroethoxycarbonyl, benzyloxycarbonyl or 1-methoxycarbony!-2propenyl. ' Alternatively, the compounds of Formula I where W is hydrogen can be prepared by acylation of an appropriate 7 - amino - 3 10 sulfaminoalkyltetrazolylthiomethyl cephalosporin described and claimed in our Patent ^>ecification No. 1645/80, and if desired removing the protective ester-forming group, and optionally forming a salt or an ester of the compound of formula I produced.

When the 7 - amino - 3 - sulfaminoalkyltetrazolylthiomethyl cephalosporin used has a protective ester-forming group R thereon, the protective group can be removed according to known methods, such as with trifluoroacetic acid when t-butyl or t-butoxycarbonyl protective groups are used. The resulting salt can be converted into a zwitterionic product or into the free acid by means of an ion exchange resin’ such as polystyrene-amine ion exchange resin (*Amberlite IR-45); or else by basification of an aqueous solution of the salt.

The acylating agents used as starting materials are either known or prepared by known methods.

*Amberlite is a trade mark Certain compounds of this invention are capable of forming salts, for example with alkali metals, such as sodium or potassium, with alkaline earth metals, such as calcium, or with the ammonium cation. When A of Formula I is NH2, the compounds can exist as a zwitterion or as either an acid or base addition salt. These salts can be prepared by standard methods using a wide variety of pharmaceutically acceptable acids and bases known in the art.

It will be recognized that due to the asymmetric α-carbon atom in the 7-acetamido group of Formula I when R1 is II X—CH-C- , 10 1 optical isomers will exist. Racemic or resolved products are obtained depending upon whether a racemic or resolved side-chain acid is used as an acylating agent. The resolved side-chain acids are readily obtained from the racemic compounds by resolution according to well known methods, including fractional crystallization of a salt formed with an optically active acid or base. All of the isomers, including separated isomers and mixtures thereof, are included within the scope of this invention.

The compounds of Formula I have antibacterial activity against 20 both Gram-positive and Gram-negative organisms. Minimum inhibitory concentrations (MIC's) range from 0.2 to >200 ug/ml in in vitro testing. Test results for representative compounds are given in Table I below. In vivo mouse protection data {EDgg's) is given in Table 2. Names corresponding to compound numbers are given in Table 3. 443S3 TABLE Τ MIC (gg/ml) in vitro Compound Number Bacteria 1 2 3 4 S.aureus HH 127 3.1, 3.1 1.6 3.1 1.6 S. aureus SK 23390 0.8, 0.8 0.4 3.1 1.6 S. villa!uz SK 70390 50, 200 25 >200 >200 Strep, faecalis HH 34358 100, 50 12.5 50 100 E. coli SK 12140 0.8, 0.8 3.1 0.8 0.8 E. coli HH 33779 3.1, 1.6 12.5 0.8 1.6 Kleb. pneumo. SK 4200 1.6, 0.8 3.1 0.8 0.8 Kleb. pneumo. SK 1200 0.4, 0.2 0.8 0.4 0.2 Salmonella ATCC 12175 0.2, 1.6 12.5 0.4 0.4 Shigella HH 117 0.4, 0.2 - 0.4 0.8 Pseudo, aerug. HH 53 > 200, >200 >200 >200 >200 Serratia marc. ATCC 13880 100, 100 >200 200 100 Proteus morgani 179 1.6, 3.1 >200 200 >200 Entero. aerog. ATCC 13048 50, 6.3 25 3.1 6.3 Entero. cloacae HH 31254 1.6, 1.6 6.3 0.8 1.6 (The two columns of results for Compound No. 1 are for two separate tests against each organism).

TABLE 2 ED50 (mg/kg) in vivo E. coli SK 12140 Kleb. pneumo. SK 4200 Compound Number s.c. p.o. s.c. p.o. 1 0.46 50 0.46 2 1.02 >50 - - 3 1.56 - - - 4 1.82 50 - TABLE 3 Compound Compound Name Number 7-D-mandel ami do-3- 0-(2-sulfaminoethyl)tetrazol5-ylthiomethylJ -3-cephem-4-carboxylic acid 7-(2-thienylacetamido)-3-£ 1-(2-sulfami noethyl)tetrazol-5-ylthiomethyl.J -3-cephem-4-carboxylic acid 7-(l-tetrazolylacetamido)-3- 0-(2-sulfaminoethyl)tetrazol-5-ylthiomethyf] -3-cephem-4-carboxylic acid 7-trifluoromethylthioacetamido-3- [j-(2-sulfaminoethyl) tetrazol-5-yl thiomethylj-3-cephem-4carboxylic acid 4439 3 The invention also includes pharmaceutical compositions having antibacterial activity which comprises a pharmaceutically acceptable carrier and a compound of Formula I as well as methods of combatting bacterial infections in non-human animals by administering such a composition to an infected host in a non-toxic amount sufficient to combat.such infections. The administration will be by parenteral injection, for example, subcutaneously, intramuscularly or intravenously. The injection of suitably prepared sterile solutions or suspensions containing an effective, non-toxic amount of the new cephalosporin compound is the preferred route of administration.

The compounds of Formula I will in general be formulated and administered in the same manner as other cephalosporins. The dosage regimen is preferably administered by injection of an active but nontoxic quantity of a compound of Formula I selected from the dosage unit range of from 100 to 1000 mg., with the total daily dosage regimen being from 400 mg to 6g. The precise dosages will be dependent upon the age and weight of the subject, and on the infection being treated, and they can be determined by those skilled in the art based on the data disclosed herein compared with that available to the art for previously known cephalosporins.

The invention also includes within formula I 7a-methoxy compounds of the formula: where iJ and £ are as previously defined) and pharmaceutically acceptable salts thereof.

A selected group of the compounds of Formula III are those where 5 n is two.

Representative of the compounds of Formula III are 7a - methoxy - 7β - (2 - thienylacetamido) -3-0-(2- sul fami noethyl )tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid, 7a - methoxy - 7$ - tri fluoromethyl thi oacetami do -3-0-(210 sulfaminoethyl) tetraol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid, 7β - D - mandelamido - 7a - methoxy - 3 - 0-(2- sul fami noethyl) tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid and 7a - methoxy -70-(1- tetrazolyl acetamido) - 3 - 0-(215 sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid.

The compounds of Formula III are preferably prepared by displacing the 3-acetoxy group from a 7a - methoxy - 7β - acylaminocephalospcranic acid or a protected ester or salt thereof with an appropriate tetrezolethiol described and claimed in our Patent Specification No. 1644/80, or a salt thereof, if necessary removing the protective ester group, and conversion of any salts into the corresponding free acids, all as described hereinabove. The 7a - methoxy - 7e - acylaminocephalosporanic acids and salts thereof are either known to the art or can be prepared by known methods.

As with the compounds of Formula I where W is hydrogen, pharmaceutically acceptable salts and all isomers, including separated isomers and mixtures thereof, of the compounds of Formula III are included within the scope of this invention.

The compounds of Formula III have anti-bacterial activity against both Gram-positive and Gram-negative organisms. They can be administered and formulated in the manner described hereinbefore.

The following Examples illustrate the invention but are not to be construed as limiting the scope thereof. Temperatures are in degrees Centigrade (°C) unless otherwise stated.

EXAMPLE 1 7-D - Mandelamido - 3 - [j - (2 - sulfaminoethyl)tetrazol 5 - ylthiomethylj - 3 - cephent - 4 - carboxylic Acid To a mixture of 2.71 g (0.006 mol) of 7-D-mandelamidocephalosporanic acid sodium salt and 1.18 g (0.004 mol) of 1 - (2 - sulfaminoethyl) -tetrazole - 5 - thiol disodium salt in 30 nil Of water was added 10% aqueous sodium hydroxide solution and then 5% aqueous sodium bicarbonate solution to pH 7.3. The reaction mixture was heated at 70°C for 2.66 hours, then it was cooled, covered with ethyl acetate, acidified to pH 2.5 with 3 N hydrochloric acid, and ί extracted twice with ethyl acetate. The aqueous phase was neutralized to pH 7.0 by addition of 10% aqueous sodium hydroxide and then 5% aqueous sodium bicarbonate solutions, and then it was chromatographed on*Amberlite XAD-7 resin with water and methanol as eluants. (The resin was a non-polar polystyrene polymeric absorbent resin having an average pore diameter 50°A). After removing the methanol in vacuo the chromatography fractions were lyophilized to give 7 - D - mandelamido -3-0-(2- sulfaminoethyl)tetrazol - 5 ylthiomethylj - 3 - cephem - 4 - carboxylic acid disodium salt.

C19H19N7°8S3 · 2 Na . 3 H20 Calculated: 34.08% C; 3.76% H; 14.64% N 34.56% C; 3.25% H; 13.96% N An aqueous solution of 7 - D - mandelamido - 3 - 01-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid disodium salt is passed through a column of Amberlite IR-120H ion exchange resin to give the title compound.

*Amberlite is a trade mark.

EXAMPLE 2 7-(2- Thi enyl acetamido) - 3 - (j - (2 - sul fami noethyl )tetrazole - 5 - ylthiomethyl J - 3 - cephem - 4 - carboxylic Acid A mixture of 1.89 g (0.064 mol) of 1 - (2 - sulfaminoethyl)tetrazole 5 - 5 - thiol disodium salt and 2.67 g (0.064 mol) of 7 (2 - thienylacetamido)cepha1osporanic acid sodium salt in 40 ml of water was heated at 69° for 5.5 hours while maintaining the pH at 7.4 by the addition of dilute aqueous sodium bicarbonate solution. After cooling, the mixture was extracted with ethyl acetate. The aqueous phase was neutralized, evaporated to dryness and the residue was passed through a column of Amberlite XAD-4 resin (as described in Example 1) eluting with water and methanol. The methanol was removed by evaporation, and the aqueous residue was lyophilized to give a solid material. The solid was suspended in methanol, the insoluble material was removed by filtration, and the filtrate was evaporated to dryness to give 7-(2- thi enyl acetamido) - 3 - 01-(2- sul fami noethyl) tetrazoi - 5 ylthiomethyl] - 3 - cephem - 4 - carboxylic acid disodium salt.

C17H17Ny07S2 . 2 Na . 1 CH^O Calculated: 33.90% C; 3.31% H; 15.37% N Found: 34.04% C; . 3.57% H; 14.74% N 7-(2- Thi enyl acetamido) -3-0-(2- sulfaminoethyljtetrazol - 5 - ylthiomethyl! - 3 - cephem - 4 - carboxylic acid disodium salt is converted into the title compound as described in Example 1.

EXAMPLE 3 7-(1- Tetrazolylacetamido) - 3 - β - (2 - sulfaminoethyl)tetrazole - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic Acid A mixture of 3.5 g (8.5 mmol of 7 - (1 tetrazolylacetamidojcephalosporanic acid sodium salt and 2.96 g (10 mmol) of 1-(2- sul fami noethyl) tetrazole - 5 - thiol disodium salt in 50 ml of water was stirred at 65° for 6.5 hours while maintaining the pH of the reaction mixture at 7.0 by the addition of 5% aqueous sodium bicarbonate solution. The mixture was cooled to ambient temperature, acidified to pH 1.5 with 3 N hydrochloric acid, filtered, and extracted three times with ethyl acetate. The pH of the aqueous phase was then adjusted to 7.0 by the addition of sodium bicarbonate, the solution was chromatographed on a column of Amberlite XAD-2 resin and the resulting product was freeze-dried to give 7-(1- tetrazolylacetamido) - 3 - β - (2 sulfaminoethyljtetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid disodium salt. (The resin is a polystyrene absorbent having an average pore diameter of 90°A).

C14H15N11°7S3 · 2 Na · 2H2° Calcualted: 26.80% C; 3.37% H; 24.55% N Found: 27.11% C; 3.40% H; 24.18% N 7-(1- Tetrazolylacetamido) -3-0-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid disodium salt is converted into the title compound as described in Example 1.

EXAMPLE 4 - Trifluoromethylthioacetamido - 3 - 0 - (2 sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 cephem - 4 - carboxylic Acid A solution of 3.05 g (0.01 mol) of 1 - (2 - sulfaminoethyl)tetrazole - 5 - thiol - disodium salt and 4.35 g (0.01 mol) of - trifluoromethylthioacetamidocephalosporanic acid sodium salt in 50 ml of water was heated at 70° for 5.5 hours while maintaining the pK at 7.5 with 5% aqueous sodium bicarbonate. The reaction mixture was diluted with 50 ml of water, and it was extracted twice with ethyl acetate. The aqueous phase was acidified to pH 2 and extracted three times with ethyl acetate. The aqueous layer was brought to pH 7.4 by the addition of 5% aqueous sodium bicarbonate, and the solution was passed through an Amberlite XAD-4 resin column while eluting v/ith water, followed by methanol. The methanol solution was evaporated to dryness, and the residue was dissolved in 75 ml of water.

The aqueous solution was extracted twice with ether and petroleum ether, and then lyophilized. The lyophilized material was dissolved in methanol, the solvent was evaporated to dryness, and the residue was triturated with ether to give 7 - trifluoromethylthioacetamido - 3 0-(2- sul faminoethyl )tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid disodium salt.

CuH14F3N7Q7S4 . 2 Na . 2H20 Calculated: 25.49% C; 2.75% H; 14.85% N Found:' 25.85% C; 2.78% H; 14.13% N - Trifluoromethylthioacetamido -3-0 -(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid disodium salt is converted into the title compound as described in Example 1.

EXAMPLE 5 - (D - α - Aminophenyl acetamido) - 3 - [j - (2 sulfaminoethyl)tetrazol - 5 - ylthiomethylj- 3 cephem - 4 - carboxylic Acid A solution of 7.58 g (0.015 mol) of 7 - (D - a - t butoxycarbonylaminophenylacetamido)cephalosporanic acid, 2.96 g (0.01 mol) of 1-(2- sulfaminoethylJtetrazole - 5 - thiol disodium salt and 1.26 g ((0.015 mol) of sodium bicarbonate in 125 ml of water is stirred at 60° for 5 hours while maintaining the pH at 7.0-7.2 by the addition of sodium bicarbonate. The mixture is cooled, and it is then extracted with ethyl acetate. The aqueous phase is acidified to pH 2.5 with 3 N hydrochloric acid, and the acidic solution is extracted again with ethyl acetate. The aqueous phase is brought to pH 7.1 by the addition of 5% sodium carbonate solution, and it is then passed through an Amberlite XAD-4 ion exchange resin column and eluted with water and methanol to give 7 - (D - a - t - butoxycarbonyl aminophenylacetamido) 3 - Ll (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem 4 - carboxylic acid sodium salt.

- (D - a - t; - Butoxycarbonylaminophenylacetamido) - 3 Ll - (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid disodium salt is stirred at 25° with 25 ml of trifluoroacetic acid and 25 ml of 1,3-dimethoxybenzene for 2.25 hours.

The mixture is evaporated to dryness, ether is added to the residue, and the precipitate is collected, washed with ether, stirred in acetonitrile for 2 hours and dried in vacuo to give the title compound as the trifluoroacetic acid salt. 44393 An aqueous solution of the trifluoroacetic acid salt is brought to pH 5.0 by the addition of di lute, aqueous sodiuni hydroxide. After lyophilization, the lyophilized material is dissolved in methanol, and ether is added to precipitate 7 - (D - a - aminophenylacetamido) - 3 5 [j - (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid sodium salt. The sodium salt is dissolved in water, and the aqueous solution is passed through an Amberlite IR-120H ion exchange resin column. Lyophilization of the eluted material gives the title compound. 1θ EXAMPLE 6 Reaction of the N-jt-butoxycarbonyl derivative of the following cephalosporanic acids: - (a - amino -4^ hydroxypheny1acetamido)cephalosporanic acid, - (a - amino - 4 - formamidophenylacetamido)cephalosporanic acid, 7 - (a - amino - 3 - formamidophenylacetamido)cephalosporanic acid, - (a - amino - 4 - ureidophenylacetamido)cephalosporanic acid, - (a - amino - 3 - ureidophenylacetamido)cephalosporanic acid, - (a - amino - 4 - hydroxymethylphenylacetamido)cepha1osporanic acid, - (« - amino - 1,4 - cyc1ohexadienylacetamido)cephalosporanic acid, 7 - (a - amino - 4 - carboxymethylaminophenylacetamido)cephalosporanic acid, with 1-(2- sul fami noethyl) tetrazole - 5 - thiol disodium salt as described in the procedure of Example 5, followed by removal of the protective group and conversion of the trifluoroacetic acid salts into the free acids as described herein, gives the following compounds of this invention: - (α - amino - 4 - hydroxyphenylacutamido) - 3 01-(2- sulfaminoethyl) tetrazol - 5 - yl thi omethylj - 3 - cephem - 4 - carboxylic acid, - (a - amino - 4 - formamidophenylacetamido) - 3 5 D (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid, - (a - amino - 3 - formamidophenylacetamido) - 3 01-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem 4 - carboxylic acid, 7 - (o - amino - 4 - ureidophenylacetamido) - 3 - 01-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid, - (a - amino - 3 - ureidophenylacetamido - 3 - 01-(2sul fami noethyl) tetrazol - 5 - ylthiomethylj] - 3 - cephem - 4 15 carboxylic acid, - (a - amino - 4 - hydroxymethyl phenylacetamido) - 3 0-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem 4 - carboxylic acid, - (« - amino - 1,4 - cyclohexadienylacetamido) - 3 20 0 - (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid, - (a - amino - 4 - carboxymethylaminophenylacetamido) - 3 0-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid. 4,4 39 3 EXAMPLE 7 - (4 - Hydroxymandelamido) - 3 - j_l - (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid is prepared by reaction of 7-(4- hydroxymandelamido)cephalosporanic acid sodium saTt and 1-(2- sulfaminoethyl)tetrazole - 5.- thiol disodium salt, followed bytreatment of the product with Amberlite IR-120H ion exchange resin as described in the procedure of Example 1.

EXAMPLE 8 When the sodium salt of a cephalosporanic acid listed below: 7 - (sydnon - 3 - y1aeetamido)cephalosporanic acid, - (2 - aminomethylpheny1acetamido)cephalosporanic acid, is reacted with 1-(2 - sulfaminoethyl)tetrazo1e - 5 - thiol disodium salt by the procedure described in Example 1, and the product is converted into the free acid as described therein, the following compounds of this invention are obtained, respectively,: - (sydnon - 3 - ylacetamido) - 3 - Ql .-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 -cephem - 4carboxylic acid, - (2 - aminomethylphenylacetamido) -3 - Qj - (2 20 sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 -carboxylic acid.

EXAMPLE 9 Reaction of the sodium salt of a cephalosporanic acid listed below: - (2,2,2 - trifluoroethylthioacetamidojcephalosporanic acid, 7-methylthioacetamidocephalosporanic acid, with 1-(2- sul fami noethyl) tetrazole - 5 - thiol as described in the procedure of Example 4 gives, after conversion of the salts formed into the free acids, the following compounds of this invention as final products: 7 -(2,2,2 - tri fluoroethylthioacetamido) - 3 0-(2- sul fami noethyl) tetrazoi - 5 - ylthiomethyl! - 3 - cephem - 4 - carboxylic acid, - methyl thi oacetami do - 3 - 01-(2- sul faminoethyl) tetrazoi - 5 - ylthiomethyl! - 3 - cephem - 4 - carboxylic acid.

EXAMPLE 10 Reaction of the disodium salt of the following thiols: 1-(3- sulfaminopropyl)tetrazole - 5 - thiol, 1-(4- sul fami nobutyl) tetrazole - 5 - thiol, 1-(5- sul fami nopentyl )tetrazole - 5 - thiol, with 7 - D - mandelamidocephalosporanic acid sodium salt as described in the procedure of Example 1, followed by conversion of the salts formed into the free acids, gives the following compounds of this invention: 7-D - mandelamido - 3 -01 - (3 - sul fami nopropyl) tetrazol - 5 ylthiomethylj - 3 - cephem - 4 - carboxylic acid, 7-D - mandelamido - 3 - 0 -(4- sul fami nobutyl) tetrazol 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid, 7-D - mandelamido -3-0- (5 - sul fami nopentyl) tetrazol 5 - ylthiomethyf] - 3 - cephem - 4 - carboxylic acid.

Likewise, reaction of the substituted tetrazole thiols or the corresponding disodium salt listed above with any of the 7-acylamino cephalosporanic acids mentioned herein or their corresponding salts according to procedures described herein gives the corresponding compounds of this invention.

EXAMPLE 11 Reaction of a cephalosporanic acid listed below or its correspond ing salt: 7 - (a - hydroxy - 2 - thienylacetamido)cephalosporanic acid, - (« - carboxy - 2 - thienylacetamido)cepha1osporanic acid, - (a - sulfophenylacetamido)cephalosporanic acid, with 1 - (2- sul faminoethyl) tetrazole - 5 - thiol disodium salt by procedures described hereinabove gives, after conversion of the product into the free acid, the following compounds of this invention: - (a - hydroxy - 2 - thienylacetamido) - 3 - 0-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid, - (α - carboxy - 2 - thienyl acetami do) - 3 - £.1-(2sulfaminoethyl)tetrazol - 5 - ylthiomethyt] - 3 - cephem - 4 carboxylic acid, - (a - sulfophenyl acetamido) - 3 - 0-(2- sul faminoethyl)5 tetrazol - 5 - ylthiomethyf] - 3 - cephem - 4 - carboxylic acid.

EXAMPLE 12 7-(4- Pyridyl thioacetamido) - 3 - 0 -(2sul faminoethyl )tetrazol - 5 - ylthiomethyl]1 - 3 - cephem - 4 - carboxylic Acid (4 - PyridylthioJacetyl chloride (0.53 g., 2.9 mmol) is added dropwise to a mixture of 1.0 g of 7 - amino -3-01-(2sulfaminoethyl)tetrazol - 5 - ylthiomethyfj - 3 - cephem - 4 - carboxylic acid and 0.9 g (9.0 mmol) of thienylamine in 10 ml of dry dimethylformamide. The reaction mixture is stirred for 1.5 hours at -io°, and then it is warmed to ambient temperature and stirred for 1 hour.

This mixture is filtered, and the filtrate is diluted with 200 ml of ether-petroleum ether. The precipitate is collected by filtration, and it is taken up in methanol. The methanol solution is filtered, and the filtrate is evaporated to dryness to give the title compound as its corresponding sodium salt. 7-(4- Pyridyl thi oacetami do) -3-0-(2- sulfaminoethyljtetrazol - 5 - ylthiomethyf] - 3 - cephem - 4 - carboxylic acid sodium salt is converted into the title compound by procedures described above. 4439 3 EXAMPLE 13 Acylation of 7 - amino - 3 - 01-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethyl! - 3 - cephem - 4 - carboxylic acid with an activated derivative of the following acids: cyanoacetic acid 3-pyridylthioacetic acid cyanomethylthioacetic acid 2j2,2-trif1uoroethy1sulfinylacetic acid methylsulfonylacetic acid as described in the procedure of Example 12 gives the following compounds of this invention: - cyanoacetamido - 3-01-(2- sulfaminoethyl)tetrazol 5 - ylthiomethyl! - 3 - cephem - 4 - carboxylic acid, - (3 - pyridylthioacetamido) -3-0-(2- sul fami noethyl )15 tetrazoi - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid, - cyanomethyl thi oacetami do -3-^1-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethyl]! - 3 - cephem - 4 - carboxylic acid, - (2,2,2 - trifluoroethylsulfinylacetamido) -3-0.1(2 - sulfaminoethyl)tetrazol - 5 - ylthiomethyl! - 3 - cephem - 4 20 carboxylic acid, - methylsulfonyTacetamido - 3 - 0 - (2 sul fami noethyl) tetrazoi - 5 - ylthiomethyl] - 3 - cephem - 4 carboxylic acid.

EXAMPLE 14 - (D - α - Formyloxyphenylacetamido) - 3 - J - (2 sulfjaminoethyl)tetrazol - 5 - ylthiomethylj - 3 cephem - 4 - carboxylic Acid - Amino -3-0-(2- sul fami noethyl) tetrazol - 5 ylthiomethylj - 3 - cephem - 4 - carboxylic acid is reacted with the formate ester of D - mandeloyl chloride according to the procedure of Example 12 to give the title compound.

EXAMPLE 15 7a - Methoxy -76-(2- thienylacetamido) -3-0-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 cephem - 4 - carboxylic Acid A solution of 1.28 g (3 mmol) of 7a - methoxy - 7b (2 - thienylacetamido)cephalosporanic acid sodium salt is dissolved in 50 ml of water, 1.33 g (4.5 mmol) of 1-(2- sulfaminoethyl)tetrazole - 5 - thiol disodium salt is added, and the solution is heated at 70° until thin layer chromatography indicates consumption of the starting material (ca. 5 hours). The reaction mixture is chromatographed on Amberlite XAD-4 ion exchange resin with, after washing with water, methanol as eluant. Evaporation of the methanol solution gives the title compound as the disodium salt. 7a - Methoxy - 7e - (2 - thienylacetamido) -3-0-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj] - 3 - cephem - 4 carboxylic acid disodium salt is converted into the title compound as described above. -14 3 9 3 EXAMPLE 16 7α -Methoxy - 7β - trifluoromethylthioacetamido - 3 - (j - (2 sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic Acid To a cold solution of 5.25 g (0.012 mol) of 7β - amino - 7a methoxycephalosporanic acid benzhydryl ester in 200 ml of methylene chloride containing 1.79 g (0.012 mol) of Ν,Ν-diethylaniline is added dropwise over a 20 minute period a solution of 1.82 g (0.012 mol) of trifluoron.-thylthioaref/i chloride in 50 ml of methylene chloride. After stirring for 30 minutes, the mixture is extracted successively with 5% aqueous sodium bicarbonate, % aqueous hydrochloric acid, and finally with brine. The organic phase is dried (MgSOJ, and the solvent is evaporated to give 7a - methoxy - 7β trifluoromethylthioacetamidocephalosporanic acid benzhydryl ester. 7a - Methoxy - 7β - trifluoromethylthioacetamidocephalosporanic acid benzhydryl ester is dissolved in a cold mixture of trifluoroacetic acidanisole (2:1), and the mixture is stirred for 1.5 hours without external cooling. The solvent is evaporated in vacuo, and the residual product is taken up in ethyl acetate, washed with water, dried (MgSOJ, and concentrated in vacuo to a small volume. This solution is added dropwise to stirred petroleum ether to precipitate 7a - methoxy - 7β trif1uoromethylthioacetamidocephalosporanic acid. 7a - Methoxy - 7β - trifluoromethylthioacetamidocephalosporanic acid (2.2 g, 5 mmol) is suspended in 75 ml of water and 0.4 g of solid sodium bicarbonate is added until dissolution is complete. To this solution is added 2.21 g (7.5 mmol) of 1 - (2 - sulfaminoethyl)tetrazole - 5 - thiol disodium salt, and the mixture is heated at 70° for 7 hours. The pH of the reaction mixture is maintained at 7.5 by the dropwise addition of 3 N hydrochloric acid, as necessary. Progress of the reaction is monitored by thin layer chromatography, and it is judged to be complete when tic indicates disappearance of starting material. The reaction mixture is chromatographed on a column of Amberlite XAD-4 resin, and the product is eluted from the column with methanol. Evaporation of the methanol solution gives 7a-methoxy-7B - trifluoromethylthioacetamido - 3 26 (_1 - (2 - sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem 4 - carboxylic acid disodium salt.

The di sodium salt is converted into the title compounds by procedures described hereinabove.

EXAMPLE 17 7β - D - Mandelamido - 7a - methoxy -3-01-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 cephem - 4 - carboxylic Acid A cold solution of 2.6 g (6 mmol of 7β - amino - 7a methoxycephalosporanic acid benzyhydryl ester in 100 ml of methylene chloride containing 0.9 g (6 mmol) of Ν,Ν-diethylaniline is treated dropwise over a 15 minute period with a solution of 1.7 g (6 mmol) of D - 0 - dichloroacetylmandeloyl chloride in 25 ml of methylene chloride.

The reaction mixture is allowed to come to room temperature with stirring, and then it is extracted successively with 5% aqueous sodium bicarbonate, % hydrochloric acid, and brine. The organic phase is dried and evaporated in vacuo. The residue is dissolve in cold trifluoroacetic acid-anisole (2:1), and the mixture is stirred at ambient temperature for 1 hour. The mixture is evaporated in vacuo, and the residue is dissolved in 5% aqueous sodium bicarbonate. The pH is raised to 9-9.3 by the addition of 5% aqueous sodium carbonate, and the pH is maintained there for 30 minutes to complete cleavage of the dichloroacetyl group. The solution is cooled in ice, layered with ethyl acetate, and it is acidified to pH 2.0 with dilute hydrochloric acid. The layers are separated, and, after a second extraction of the aqueous layer with ethyl acetate. the organic phases are combined, dried and evaporated in vacuo to yield 7β - D - mandelamido - 7a methoxycephalosporanic acid. 7β - D - Mandelamido - 7a - methoxycephalosporanic acid (2.2 g, mmol) is suspended in 75 ml of water, and solid sodium bicarbonate is added until all of the acid has dissolved. To this are added 2.21 g (7.5 mmol) of 1 - (2 - sulfaminoethyl)tetrazole - 5 - thiol disodium salt, and the mixture is heated at 70° for 7 hours. The pH of the reaction mixture is maintained at 7.5 by the addition of 3 N hydrochloric acid. Chromatography of this solution on Amberlite XAD-4 resin while eluting with methanol gives, upon evaporation of the methanol the title compound as its disodium salt. 7g - D - Mandelamido - 7a - methoxy - 3 - rl - (2 sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid disodium salt is converted into the title compound by procedures described hereinabove.

EXAMPLE 18 7a - Methoxy - 7β -(D - a - aminophenylacetamido) - 3 01-(2- sul faminoethyl )tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic Acid To a solution of 5.3 g (0.012 mol) of 70 - amino - 7a methoxycephalosporanic acid £-m'trobsnzyl ester in 200 ml of methylene chloride are added 3.0 g (0.012 mol) of D-a-jtbutoxycarbonylaminophenyl acetic acid and 2.5 g (0.012 mol) of 25 dicyclohexylcarbodiimide. The mixture is stirred for 18 hours at ambient temperature and then filtered. The filtrate is evaporated in vacuo, and the residue is dissolved in methanol-tetrahydrofuran and 4393 hydrogenated over 5% palladium on carbon to give 7p -(D -a - t butoxycarbcnylanrinopheny1acetanrido)-7a - methoxycephalosporanic acid. 7β - (D - a - t - Butoxycarbonyl ami nophenyl acetamido) - 7a methoxycephalosporanic acid (2.58 g., 5 mmol) is dissolved in 75 ml of water by adding 0.5 g of solid sodium bicarbonate. 1-(2- Su 1 faminoethy 1)tetrazole - 5 - thiol disodium salt (2.21 g., 7.5 mmol) is added, and the reaction mixture is heated at 70° until thin layer chromatography indicates that the starting material has disappeared. The reaction mixture is chromatographed on Amberlite XAD-4 resin and eluated with methanol. Evaporation of the methanol solution gives 7a - methoxy -7g-(D-a-j;butoxycarbonyl ami nophenyl acetamido) -3-0-(2sulfaminoethylJtetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid disodium salt.

The disodium salt is suspended in 1:1 trifluoroacetic acidanisole and stirred at ambient temperature for two hours. Excess trifluoroacetic acid is removed by evaporation, the residue is triturated with ether, and the resulting precipitate is collected by filtration and stirred with acetonitrile to give the title compound as its trifluoroacetic acid salt.

An aqueous solution of the trifluoroacetic acid salt is brought to pH 7 by the addition of 5% aqueous sodium bicarbonate, and then it is chromatographed on Amberlite XAD-4 resin with methanol as eluant. The solid material obtained after evaporation of the methanol is dissolved in water and the aqueous solution is passed through a cation exchange column (IR-120H). Lyophilization of the eluted material gives the title compound.

EXAMPLE 19 7α - Methoxy - 7β - (1 - tetrazolylacetamido) - 3 U -(2- sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid Substitution of an equivalent amount of 1-tetrazolylacetyl chloride in the procedure of Example 16 gives 7a - methoxy - >e - (1tetrazolylacetamido)cephalosporanic acid benzhydryl ester which is converted into 7a - methoxy - 7g - (1 tetrazolylacetanndo)cephalosporanic acid as described therein.

Reaction of 7a - methoxy - 7β- (1 tetrazolylacetamido)cephalosporanic acid, 1-(2- sul faminoethyl) tetrazole - 5 - thiol disodium salt and sodium bicarbonate as described in Example 16 gives the title compound, after conversion of the product disodium salt into the free acid as described above.

EXAMPLE 20 An injectable pharmaceutical composition is formed by adding sterile water or sterile saline solution (2 ml) to 500 mg of 7-D - mandelamido - 3 - [l - (2 - sul faminoethyl) tetrazol - 5 ylthiomethylj - 3 - cephem - 4 - carboxylic acid disodium salt.

Pharmaceutical compositions of the other antibacterial compounds disclosed above can be formulated in a similar manner.

Claims

1. A compound of the formula: where W is hydrogen or methoxy; g R^ is an acyl group selected from X —CH -CO—, I A Y-CHg-CO- and Z-S(O) m -CHg-CO- (where X is thienyl, dihydrophenyl, phenyl or phenyl monosubstituted with hydroxy, hydroxymethyl, formamido, ureido or carboxymethyl ami no; 10 A is NHg, OH, COOH or SOgH; or formyloxy when X is phenyl; Y is thienyl, tetrazolyl, sydon-3-yl, cyano or aminomethylphenyl; Z is methyl, trifluoromethyl, trifluoroethyl, cyanomethyl or pyridyl; and m is zero (one or two) and ji is an integer from two to five. 2. 3. A compound according to Claim 1, in which n^ is two. A compound according to Claim 1, in which R is X — CH — CO, A is phenyl, and A is NHg or OH. A compound according to Claim 3, in which W is hydrogen. A compound according to Claim 3, in which W is methoxy. A compound according to Claim 1, in which R^ is Υ-CHg-CO-, and is thienyl or tetrazolyl. A compound according to Claim 2, in which W is methoxy. A compound according to Claim 6, in which W is methoxy. A compound according to Claim 1, in which R^ is Z-S(0) ra -CHg-CO is trifluoromethy!, and m is zero, one or two. A compound according to Claim 9, in which ll is hydrogen. A compound according to Claim 9, in which W is methoxy. 7 - D - Mandelamido -3-0-(2- sul faminoethyl )tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid. 13. 7β - D - Mandelamido - 7a - methoxy -3-0-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxyli acid. 14. 7-(2- Thienylacetamido) -3-01-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethyl] - 3 - cephem - 4 - carboxylic acid. 15. 7-(1- Tetrazolylacetamido) -3-01-(2- sul fami noethyl) tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 - carboxylic acid. 5 16. 7a - Methoxy -78-(2- thienylacetamido) - 3 01-(2- sulfaminoethyl)tetrazo1 - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid. 17. 7a - Methoxy -78-(1- tetrazolylacetamido) - 3 El - (2 - sulfaminoethylJtetrazol - 5 - ylthiomethylj - 3 - cephem - 4 10 carboxylic acid. 18. 7 - Trifluoromethylthioacetamido -3-01-(2sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid. 19. 7a - Methoxy - 7β - trifluoromethyethioacetamido - 3·15 01-(2- sulfaminoethyl)tetrazol - 5 - ylthiomethylj - 3 - cephem - 4 carboxylic acid. 20. A compound according to any of the preceding claims in the form of a pharmaceutically acceptable salt thereof or an ester thereof. 21. A process for preparing a compound according to Claim 1, where 20 W is hydrogen which comprises acylating an appropriate 7 - amino - 3 - sulfaminoalkyltetrazolyl thiomethyl-cephalosporin of formula 4. 43 9 3 (CH 2 ) n -NHS0 3 H (where £ is as defined in Claim 1 and is hydrogen or a protective ester-forming group) with an appropriate acylating agent, and if desired removing the protective ester-forming group, and optionally 5. Forming a salt or an ester of the compound of formula I produced. 22. A process according to Claim 21, substantially as herein described in any of Examples 12 to 14. 23. A compound according to Claim 1, where W is hydrogen when prepared by a process according to Claim 21 or Claim 22. 6. 10 24. A process for preparing a compound according to Claim 1, where H is methoxy which comprises displacing the 3-acetoxy group of the appropriate 7a - methoxy - 7β - acylaminocephalosporanic acid or a protected ester or a salt thereof, by reaction with an appropriate 1-sulfaminoalkyltetrazole-thiol or a salt thereof, if necessary removing the protective ester group. 25. A process according to Claim 24, substantially as herein described in any of Examples 15 to 19. 26. A compound according to Claim 1, where W is methoxy, when prepared by a process according to Claim 24 or Claim 25. 5 27. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a compound as claimed in Claim 1. 28. A method of combatting a bacterial infection in a non-human animal which comprises administering a composition according to Claim 27 to the animal in a non-toxic amount sufficient to combat the 10 infection.