IE44607B1

IE44607B1 - Cephalosporin compounds

Info

Publication number: IE44607B1
Application number: IE557/76A
Authority: IE
Original assignee: Smithkline Corp
Priority date: 1975-03-18
Filing date: 1976-03-16
Publication date: 1982-01-27
Also published as: MY8200264A; DE2611270C2; HK12382A; CY1142A; FR2347355B1; NL178321B; GB1547473A; FR2347355A1; JPS51115493A; NL178321C; AU497053B2; JPS5459278A; IE44607L; LU74586A1; GB1547475A; DE2660579C2; DE2611270A1; CH630633A5; FR2347370A1; FR2304343B1

Abstract

The cephalosporin derivatives of the formula I, in which the symbols have the meanings given in Claim 1, are novel antibiotics. They are prepared from 7-aminocephalosporanic acid, by acylation of the amino group and substitution of the acetoxy group by the corresponding substituted 5-mercaptotetrazole. The derivatives which are obtained can be converted into esters.

Description

This invent, i’on rein let; lo n now serie:·; of coplin lospor i n compound!; which luivo mi I i bncl ori η 1 notlvily when niliiil η I Plfi-oil ρηυοιιΙ om I lv . In pni't lenlnr , I bo ·,Ι i ncl urr , of the biologically uctive cephalosporin compound:! of this . invention are characterized by having a sulfo- or sulfamoyl-alkyl substituted tetrazolyl-thiomethyl group at the 3-position of the cephem nucleus. The invention also includes pharmaceutical compositions and methods for treating bacterial infections using these new compounds and processes for preparing the compounds.

The cephalosporin compounds of this invention are those of the structural formula · FORMULA I in which each individual R4 is hydrogen or lower alkyl; n is from 1 to 10; R is hydroxy, amino, (lower alkyl)amino or . di-(lower alkyl)amino; R is an acyl group of the structure XCHCO-, YGH„G0- or Z-S(O) ’-CH„CO| a m et A where: X is thienyl; dihydrophenyl; phenyl; phenyl mono- substituted with hydroxy, hydroxymethyl, formamido, ureido or carboxymethylamino; or 3-fluoro-4-hydroxy35 phenyl; A is hydroxy, amino, COOIl' or SO.^Il; or formyloxy when X is phenyl; Y is thienyl, tetrazolyl, cyano, sydnon-3-yl or aminomethylphenyl; . Z is methyl, trifluoromethyl, trifluoroethyl, cyanomethyl or pyridyl; and m is 0, 1 or 2; and each C00R4 is a carboxylic acid or esterified carboxylic acid group; or a non-toxic pharmaceutically-acceptable -244607 salt: thereof.

By lower alkyl is meant an alkyl group having from 1 to 4 carbon atoms; preferably such an alkyl group is methyl or ethyl. Where COOH'1 is an esterified carboxylic acid group, R can be,for example, an alkyl or aryl group or an esterifying group which is easily cleaved within give the body ΐοχα carboxylic acid group,such as an indanyl, pivaloyloxymethyl , aeef.oxymethy i, propiony loxymoihy 1., glycyloxymethyl, phenylgIyey1oxymelby1 or thieny1glycyloxy10 methyl group.

Preferably the compounds are those of Formula I where each R·1 is hydrogen and n is from 1 to 5; and preferably n is 1 or 2. Preferably R is hydroxy or amino. Preferably Ό where R is XCIICO-, X is phenyl or hydroxyphenyl, and A A is hydroxy or amino; where R is YCH CO, Y is thienyl or 3 z 2-aminomethylphenyl; and where R is Z-S(O) -CH„CO-, Z ul Z is trifluororaethyl and m is 0.

Examples of particularly suitable 7-acylamido substituents (R'-NH-) of the compounds of Formula I are as follows; oi-hydroxyphonylace tami do 25 ci-am i. n op h on y 1 ace t am i d o tx-amino-4-hydroxyphenyfacetamido trifluorome thylthioacetainido methylthioaeetamido 2,2,2-trifluoroethylsulfinylacetamido 30 thienylacetamido tetrazolylacetamido cyanoacet.amido ix-carboxythieriylacetamido -344607 α-carboxyphenylacetamido α-sulfophenylacetamido methylsulfonylacetamido cyanome thy Ichioacet amido a-amino-4-carboxymethylaminophenylacetamido a-amino-3-fluoro-4-hydroxyphenylacetamido sydnon - 3 yl 4-pyridylthioacetamido 2-aminomerhyi/nenylacetamido.

Particularly suitable substituted tetrazolyl groups are the following: 1-sulfomethyltetrazolyl l-(2-sulfoethyl)tetrazolyl 1-(2-sulfamoylethyl)tetrazolyl ' l-(3-sulfopropyl)tetrazolyl 1-(3-sulfamoylpropyl)tetrazolyl l-(5-8ulfopentyl)tetrazolyl 1-(5-sulfamoylpentyl)tetrazolyl.

Particularly preferred are the compounds 7-Dmandelamido-3-(l-sulfomethyltetrazol-5-ylthiomethyl)-3cephem-4-carboxylic acid, 7-D-mandelamido-3-[l-(2-sulfoethyl)tetraz01»5-ylthiomethyl]-3-cephem-4-carboxylic acid, 7-D-mandelamido-3- [1-(5- sulfopentyl) tetrazol- 5-yl thiomethyl J 3-cepheo-4-carboxylic acid, 7-D-mandelamido-3-[l-(2-sulfamoylethyl)tetrazol-5-ylthiomethyl3-3-cephem-4-carboxylic acid, 7-(2-thienylacetamido)-3-(l-sulfomethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid, 7-(2-thienylacetamido)3-[l-(2-sulfoethyl)tetrazol-5-ylthiomethyl]-3-cephem-4carboxylic acid, 7-trifluoromethylthioacetamido-3(l-sulforaethyltetrazol-5-ylthiouethyl)-3-cephem-4-carboxyl ic acid, 7-(2-thienylacetamido)-3-[1-(2-sulfamoylethyl)tetrazol5 ylthiomethyl/-3-eepherti-4-earlioxy 1 ic acid and 7-tvii’luoromethylthioacetamido-3-2l-(2-sulfamoylethyl)t-etrazol-5ylthiomethyl7-3-cephein-4-carboxylie acid.

Cephalosporin derivatives having 7-acyl substituents as defined above are all documented in the prior art. Substitution by a substituted S-heterocyclic thiomethyl group (-CIIgSHet) at the 3-position of the cephem nucleus is also known and is disclosed in Netherlands Patent Specification No. 6916151 where Het is, among others, tetrazolyl substituted with, inter alia, carboxy, carbalkoxy, alkoxyalkylaminocarbonyl and dialkylaminoalkylaminocarbonyl and in Japanese Patent Specification No. 7205550 where Het includes tetrazolyl substituted with -(CHg) R where n is 0 to 3 and R includes alkoxyoarbonyl, carboxy, N-alkoxyalkylcarbamoyl and dialkylamino. Recently issued U.S. Patent Specification No. 3,819,623 discloses cephalosporins bearing a 7~heterocyelic-ac(;i.amido or 7heterocyelie-thioalkylacetamido group and having in the 3-position, inter alia, thiomethyltetrazolyl substituted with carboxy, alkoxycarbonyl, carboxyalkyl, alkoxycarbonylalkyl and dialky.laminoalkylaminocarbonyialkyl. Cephalosporin compounds containing a 3-(sulfo- or sulfamoyl- substituted tetrazolyl)thiomethyl moiety are believed to be unknown hitherto.

In a process of the invention compounds of Formula 1 where each COOR4 is a carboxylic acid group are prepared by acylating 7-aminocephalosporanic acid with an appropriately protected acylating agent capable of providing the acyl group R and the 3-acetoxy group in the acylation product is displaced by reaction with the appropriate substituted tetrazole-thiol of the structure N— N HS N-N I i 9 (CHR1)n-SO2RZ FORMULA XI and any protecting group is subsequently removed. Compounds of Formula IX are described and claimed in British Patent Application 42761/77. -5..5 ;20 .40 The carboxyli-nitrophenyl, p-methoxyphenyl, p-methoxybenzyl or p-nitrobensyl. When A is NHg, the oi-amino group of the acylating agent is preferably protected prior to acylation with an easily removable protecting group known in the art such as t-butoxycarbonyl, trichloroethoxycarbonyl, benzyloxycarbonyl, or 1-methoxycarbonyl-2-propenyl (the methyl acetoacetatr; adduct).

Alternatively, the compounds of Formula I where each COOR^ is a carboxylic, acid group are prepared by acylating the appropriate 7-amino-3-substituted tetrazolylthiomethyl cephalosporin of the structure FORMULA III in which each individual Π1 is hydrogen or lower alkyl; n is from 1 to 10; R2 is hydroxy, amino,(lower alkyDamino or di-(loweralkylHunino; and R is hydrogen or a protecting ester-forming group, with an appropriate acylating agent, followed by removal or any protecting group. Certain compounds of Formula I El are describe and claimed in British Patent Application 427S0/77.

The protecting groups can be removed according to methods well known to the art, such as with trifluoroacetic acid when l£-butyl or t-butoxycarbonyl protecting groups are used. A resulting salt may be converted to the zwitterionic product or to the free acid by means of a basic ion exchange -64 4 6 0 7 resin such as polystyrene-amine ion exchange resin (Amberlite IR-45:Amberl.ite is a Registered Trade Mark) or by basification of an aqueous solution of the salt.

Where a compound of Formula I in which each COOH4 is an esterified carboxylic acid group is required, the corresponding compound where COOR4 is a carboxylic acid group can be readily esterified by methods well known in the art.

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

The 7-amino-3-substituted’' tetrazolylthiomethyl cephalosporin 2 starting materials of Formula III where R is hydroxy can be prepared by reaction of 7-formamidocephalosporanic acid, (itself prepared by reaction of 7-aminocephalosporanic acid with formic acid and acetic- anhydride), and a substituted tetrazole-thiol of Formula IX, followed by treatment with an acid such as hydrochloric acid to remove the formyl group.

The substituted tetrazole-thiols of Formula II where R2 is hydroxy, (lower alky 1 )ainj.n<> or rii-(lower alkyl)ami.no can be prepared by reaction of an N-substitutod alkyl di thiocarbamate, such as methyl 2-sulfoethyldithiocarbamate or methyl 3-(N-t-butylsulfamoyl)propyl-dithiocarbamate or its corresponding sodium or potassium salt with an azide such as sodium azide. The N-substituted alkyl dithiocarbamates can be prepared by treatment of an aminoaikanesulfonic acid, for example 2-aminoethanesul.fonic acid, or an aminoalkane (N-alkyl or Ν,Ν-dialkyl)sulfonamide such as 3-aminopropane-N-t-butyl-sulfonamide or its corresponding salt with carbon disulfide and an alkyl halide such as methyl iodide in the presence of a base such as sodium or potassium hydroxide.

The aminoalkane (N-alkyl or Ν,Ν-dialkylJsulfonamides can be prepared by reaction with hydrazine of an N-alkyl or Ν,Ν-dialkylphthalimidoalkylsulfonamide, obtained from treatment of a phthalimidoalkylsulfonyl halide, preferably chloride, with an alkyl- or dialkyl-amine. The phthalimidoalkylsulfonyl halides are known or can be prepared as described by Winterbottom et al., J. Amer. Chem. Soc. 69:1393 (1947) and Griffin and Hey, J. Chem. Soc., 3334 (1952). -744607 When R is amino, the compounds o! Formula II can he prepared by removal of the N-alkyl group from the corresponding l-(N-alkylsutfamoytnIky!)(etra?ole-5-tlilol, preferably a l-(N-_t-butylsulfnmoylalky 1) tetrazole-5-thiol, v/ith, for example, anisole and trifluoroacetic acid.

Certain compounds of this invention are capable of forming salts with, for example, the alkali metals such as sodium or potassium, the alkaline earth metals such as calcium or with the ammonium cation. When Λ is , the compounds can exist as the zwitterion or as either an acid or base addition salt. These salts can be prepared by standard methods using a v/ide variety of non-toxic pharmaceutically acceptable acids and bases known in the art.

It will be recognized that due to the asymmetric «<-carboti a atom in the 7-acetamido group of Formula I when R is XCHCO and the potentially asymmetric carbon atom in A the tetrazole side-chain, optical isomers can exist. Racemic or resolved products are obtained depending upon whether a racemic or resolved side-chain acid is used as an acylating agent and whether a racemic or resolved tetrazole-thiol is used. 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 tills invention.

The compounds of Formula I have exceptional antibacterial activity against both Gram-positive and Gram-negative organisms. Minimum inhibitory concentrations (MIC's) range from 0.2 to >200 /ig./ml. in in vitro testing. These results are shown in Table 1 below for representative compounds of Formula I. In vivo mouse protection data are given in Table 2. Compound names corresponding to numbers are given in Table 3. -8MIC (ng./ml.) In vitro 6/1 -preSacni snaaoj^ VSZIS HH ΒΒΟΒΟχΟ ·ΟΛΘ3«3 οθβει oolv •ΟΛΒΟΙ *8XX3S βνοει ooxv ·8οαβ> oj33ua /11 HH »π«8ΐι» 9/1ZI 001V Blieuowies £9 HH •de opnasj 00Z1 NS •oonaud ·ςβχχ 00Z7 NS •oonaud ‘qaix 6//εε HH lioa ‘3 0ΠΖ1 NS TI»» *3 βεενε hh sneaaBj ‘daxis ζητβχιΤΑ «s Ο6εεζ NS enaanB ’s ZZ1 HH ana»» *S punodmoo m Aim men 00*4· • · oo CM CO OO »00 ©d ·» i-4kO mU »» 0*0 oo mo cn do m « o\o 'T—' rH m© cncn co om 6.3 3.1 3.1 n oo oo 5i£) O o 100, 100 f4 » » A ©© m rH CO » • a • β a a * m o o i—l t-4 © mo m m © ο ο o cm o oo rd r-l CM m »-t vS mm a · · · a SO m kO CM CM r-4 i—4 .....................—«yaaaa St CM » ©© • · a a a Ο Ο O HH ® atf 1-4 a a a oom o O O o © © CM CM -H A ΠΗ m dm CM 1-4 00» cn dd oooo oo O O O OO O O O oo CM CM CM CM CM A A A A A O OO O o o CM CM CM A AA St © © -1 oo ο o io co io Mt cm Ο Γ-l rt rIO rt OO \o io cn cn cn in -ιο rt rt CO OO CM enrt 00 O OO rt • · . · · · Ο O rt rt rt cn .............ΤΠ--o oi O OO o ο rt in cm in in OOMt o'O OO in in cn rt in rtin rt o cn cm cncM cn oin rt rt in rt SO CM CO Mt*Mt 00 OOMt o d ο oo o oo ......—.....j, .....— x- — o O rt OOCO O 00 00 rt rt cn dd rl oo -B-T5“ rt w > rt w H W W > > > 00 00 • • ο ο ο ο ο ο CM r4 ι-4 χο Λ r4 «ί CM • ο Ο οο «ί • ο ο ο ο ο ο «Μ CM Ζχ Mt 5 • ο ο νθ 00 β • r4 ο χο νο • • 1-4 r4 00 co • Ο Ο <Ώ m « « Ό CM ρ4 «η « * *4 *Ο r4 CM Ο ο •4· -tf Ο Ο - 10 44607 TABLE 2 ED50 E, coli 12140 ed50 La vivo (mg./kg.) Kleb, pneumo. 4200 a.c. P.O, s.c. p.o. I 1.56 — 0.28 — II 0.58 35 0.39 18.6 III 0.70 25 0.68 21.5 IV 18 >50 15.7 .. V 4 >50 0.26 — VI 4.4 — 1.56 — VII 0.86 — 0.24 — VIII 1.32 >50 2 — IX 1.56 44 -- TABLE 3 Compound Number I II III IV V VI VII Compound Name 7-D-Mandelamido-3-(1-sulfomethyltetrazol-5-ylthiomethyl)-3-cephem4-earboxylic acid 7-D-Mandelamido-3-(1-(2-sulfoethyl)tetrazol-5-ylthiomethyl]-3-cephem4-carboxylic acid 7-D-Mandelamido-3-[l-(2-sulfamoylethyl)tetrazol-5-ylthiomethyl]3- cephem-4-carboxylic acid 7-D-Mandelamldo-3-[1-(5-sulfopentyl)tetrazol-5-ylthiomethyl)-3-cephem4- carboxylic acid 7-(2-Thienylacetamido)-3-(l-sulfomethyltetrazol-5-ylthiomethyl)-3cephem-4-carboxylic acid 7-(2-Thienylacetamido)-3-f1-(2sulfoethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid 7-Trifluoromethylthioacetamldo3-(l-sulfomethyltetrazol-5-ylthiomethyl) -3-cephem-4-carboxylic acid 46 0 7 VIII 7-(2-Thi<;nylacetamido)-3-/l-(2-sulfamoylethyl)tetrazol-5-ylthiomethyl7-3-cephem-4carboxylic acid 5 IX 7-Trifluoromethylthioacetamido-3-£i-(2sulfamoylethyl)tetrazol-5-ylthiomethylJ-3cephem-4-carboxylic acid In addition, the active compounds of this invention exhibit broad spectrum activity and shown advantageously high blood serum levels and half-life values.

Pharmaceutical compositions of the invention are those comprising a compound of Formula I or non-toxic pharmaceutically„„ acceptable salts thereof and a. pharmaceuti.cally15 acceptable carrier. The invention includes a method of combatting bacterial infection in a non-human animal which comprises administering to the animal such a pharmaceutical composition in a non-toxic amount sufficient to combat such infection. The administration may be by parenteral injection such as subcutaneously, intramuscularly or intravenously. The injection of suitably prepared sterile solutions or suspensions containing an effective, nontoxic amount of the new cephalosporin compound is the preferred route of administration.

The compounds of Formula I are formulated and administered in the same manner as other cephalosporins. The dosage regimen comprises administration, preferably by injection, of an active but non-toxic 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 6 g. The precise dosages are dependent upon the age and weight of the subject and on the infection being treated and can be determined by those skilled, in the art, based on the data disclosed •4 J I J -1244607 herein compared with that available to the art attained with known cephalosporins.

Examples illustrate the invention Temperatures are in degrees Centigrade unless otherwise stated. Amberlite is a Registered Trade Mark.

EXAMPLE 1 7-D-Mandelamido-3-(1-sulfomethyltetrazol-S-ylthLomethyl)3-ceph'em-4-carboxy'lic acid To a solution of 112 g. (2.0 mol.) of potassium hydroxide and 111 g. (1.0 mol.) of aminomethanesulfonic acid in 250 ml. of water at 25° was added 71 ml. of carbon disulfide. The reaction mixture was stirred for 12 hours and 250 ml. of ethanol was added. The reaction vessel was fitted with a reflux.condenser and 62 ml. (1.0 mol.) of methyl iodide was added. After an exothermic reaction the mixture was allowed to /cool to ambient temperature, the solid product was collected by filtration. The solid was extracted with hot methanol and the extract was concentrated to give methyl sulfomethyldithiocarbamate as the potassium salt.

A mixture of 45.3 g. (0.19 mol.) of methyl sulfomethyldithiocarbamate potassium salt and 16.9 g. (0.26 mol.) of sodium azide in 425 ml. of water was heated at 80° for 4.75 hours. The reaction mixture was passed through an Amberlite IR-120H icn exchange resin column and eluted with water until the pH of the eluant became 3.5. The eluant was extracted with ether and the aqueous solution was evaporated to dryness to give 1-sulfomethyltetrazole-5-thlol. l-Sulfomethyltetrazole-5-thiol was dissolved in acetone and a 30% solution of sodium 2-ethylhexanoate - 13 in isopropanol was added. l-Sulfomethyltetrazole-5-thiol sodium salt precipitated and was collected by filtration.

A mixture of 27.4 g. (0.062 mol.) of 7-D-mandelamidocephalosporanic acid methanolate, 10.2 g. (0.047 mol.) of l-sulfomethyltetrazole-5-thiol sodium salt and 9.2 g. (0.109 mol.) of sodium bicarbonate in 300 ml. of water was heated at 70° for one hour. The reaction mixture was cooled (ice bath) and acidified to pH 1.8 with 3N hydrochloric acid. The mixture was extracted with ethyl acetate, filtered and chromatographed on an Amberlite XAD-8 resin column with water containing increasing amounts of methanol as the eluant to give the title compound.

The title compound was dissolved in methanol and a 5% solution of sodium methoxide in methanol was added until pH 7,0, Addition of ethanol precipitated the product salt which was collected, dissolved in water and lyophilized to give 7-D-mandelamido-3-(l-sulfomethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid disodium salt.

C18H16N6°8S3 · 2 Na · L·75 H2° Calculated: 34.98% C; 3.18% H; 13.59% N Found: 35.09% C; 3.17% H; 13.27% N EXAMPLE 2 7-(D-a-Aminophenylacetamido)-3-(l-sulfomethyltetrazol-5yIthiomethyl)-3-cephem-4--carboxylie acid A solution of 7.58 g. (0.015 mol.) of 7-(D-at>-butoxycarbonylaminophenylacetamido)cephaloaporanic acid, 1.96 g. (0.01 mol.) of l-sulfcmethyltetrazole-5-thiol and - 14 «4607 2.52 g. (0.03 mol.) of sodium bicarbonate in 125 ml. of water is stirred at 60° for five hours while maintaining the pH at 7.0-7.2 by addition of sodium bicarbonate. The mixture is cooled and extracted with ethyl acetate. The aqueous phase is acidified to pH 2.5 with 3N hydrochloric acid and the acidic solution is extracted again with ethyl acetate. The aqueous phase is brought to pH 7.1 by addition of 5% sodium carbonate solution, then passed an Amberlite through/ XAD-4 ion exchange resin column and eluted with water and methanol to give 7-(D-a-t:-butoxycarbonylaminophenylacetamido)-3-(1-sulfomethyltetrazol-5-ylthlomethyl)3-cephem-4-carboxylic acid disodium salt, 7-(D-a-t-butoxycarbonylaminophenylacetamido)-3(l-sulfomethyltetrazol-5-ylthiomethyl)-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 two hours, then collected and dried in vacuo to give the title compound.

EXAMPLE 3 7-Ρ-Μίίηάβΐ3πι1άο-3-[1-(2“8α1:ΕΒη>ον1β#ιν1)ίβίΓ8ζο1-5-ν1ϋ1ιΐοmethyl]-S-cephem-A-carboxyllc acid A solution of 2.73 g. (0.01 mol.) of 2 - 15 phthalimidoethanesulfonyl chloride in 20 ml. of chloroform was added dropwise to a solution of 2.19 g. (0.03 mol.) of t-butylamine in 20 ml. of chloroform at 5°. The reaction mixture was warmed to ambient temperature and stirred for three hours. The precipitate was removed by filtration and the filtrate was evaporated to dryness to give a residue which was purified by chromatography on silica with 19:1 chloroform-methanol as eluant to give Ν-£butyl-2-phthalimi doethanesulfonamide.

• N-t-Butyl-2-phthaiimidoethanesulfonamide (2,10 g., 6,78 mmol.) was suspended in 20 ml. of ethanol and 0.344 g. of hydrazine hydrate was added. The reaction mixture was refluxed for three hours, then evaporated to dryness.

The residue was suspended in 45 ml. of water and acidified to pH 3.0 by addition of dilute hydrochloric acid. The acid mixture was filtered and the filtrate evaporated to dryness to give 2-aminoethane-N-t:-butylsulfonainide hydrochloride. 2-Aminoethane-N-t-butylsulfonamide hydrochloride (1.25 g., 5.78 nmol.) was added to a solution of 1.17 g. (11.56 mmol.) of triethylamine in 20 ml. of ethanol.

Carbon disulfide (0.44 g., 5.78 mmol.) was added, the mixture was stirred at 25° for 1.5 hours, then 0.82 g. (5.78 mmol.) of methyl iodide in 5 ml. of ethanol was added and the resulting mixture was stirred for 1.5 hours. The mixture was evaporated to dryness and the residue was dissolved in water and acidified to pH 2.0 with dilute hydrochloric acid. The aqueous mixture was extracted with ethyl acetate and the extract was dried (MgSO^) and evaporated to dryness to give methyl 2-(N-t-butylsulfamoyl)ethyldithiocarbamate. - 16 11 6 ο 7 Methyl 2-(N-t-butylsulfamoyl)ethyldithiocarbamate was treated with sodium azide as described in the procedure of Example 1 for 35 minutes to give l-(2-N-jt-butylsulfamoyl ethyl)tetrazole-5-thiol, l-(2-N-_t-Butylsulfamoylethyl)tetrazole-5-thiol (1.0 g.) was suspended in 10 ml. of anisole and 20 ml, of trifluoroacetic acid is added. The solution was heated at 56° for 3.5 hours, then cooled. The precipitate was collected by filtration and washed with petroleum ether to give l-(2-sulfamoylethyl)tetrazole-5-thiol.

A solution of 0.210 g. (2.5 nmol.) of sodium bicarbonate in 5 ml. of water was added to a suspension of 0.272 g. (1 nmol.) of 7-aminocephalosporanic acid in 5 ml. of water and 2.5 ml. of acetone at 15°. The solution was heated to 45°, a solution of 0.314 g. (1.5 mmol.) of 1-(2-sulfamoylethyl)tetrazole-5-thiol in 10ml. of acetone was added and the reaction mixture was refluxed for two hours while maintaining the pH at 7.4-7.6 by addition of aqueous sodium bicarbonate solution. The mixture was cooled and acidified to pH 4.0 with dilute hydrochloric acid. The precipitate was collected by filtration to give 7-amino-3-(l-(2-sulfamoylethyl)tetrazol5-ylthiomethyl]-3-cephera-4-carboxylic acid.

To a solution of 1.26 g. (15 mmol.) of sodium bicarbonate in 75 ml. of acetone and 50 ml. of water at 5° was added 2.1 g. (5 mractl.) of 7-amino-3-[l-(2-sulfamoylethyl)tetrazol-5-ylthiomethyl]-3-cephera-4-carboxylic acid. The solution was cooled to -10° and a solution of 1.55 g. (5.5 mmol.) of D-O-dichloroacetylmandeloyl chloride in 25 ml. of acetone was added. The reaction mixture was stirred for 30 minutes in the cold while - 17 44607 maintaining the pH at 7.2 by addition of aqueous sodium bicarbonate, then for 1.5 hours at 25°. The mixture was extracted with ether and the aqueous phase was brought to pH 9.3 with 5 a sodium carbonate and stirred for 1.5 hours at 25°. The aqueous mixture wae extracted with ether , the pH adjusted to 4,5 and the solution was again extracted with ether. The aqueous phase was acidified to pH 1.5 with dilute hydrochloric acid and it vias extracted with ethyl acetate. Evaporation of the extract to dryness gave a solid which was suspended in ethyl acetate and filtered. The filtrate was diluted with ether and petroleum ether to precipitate the title compound.

EXAMPLE 4 7-D-Mandelamido-3-[l-(2-N-t-butylsulfamoylethyl)tetrazol-5ylthiomethyl]-3-eephem-4-cart>oxyllc acid When an equivalent amount of 1-(2-N-t-butylsulfamoylethyl)tetras5ole-5-thiol is reacted with 7-aminocephalosporanic acid as described in Example 3, 7-amino-3[1-(2-H-it-butylsul£amoylethyl)tetrazol-5-ylthiomethyl]-3cephem-4-carboxylic acid is obtained.

Acylation of 7-amino-3-[l-(2-N-t-butylsulfamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid with D-O-dichloroacetylmandeloyl chloride according to the procedure of Example 3 gives the title compound.

EXAMPLE 5 7-D-Mandelamido-3-[1-(2-N-methylsulfamovlethyl)tetrazol-5ylthiomethvlJ-3-cephem-4-carbbxylic acid Use of methylamine in the reaction with 2-phthalimidoethanesulfonyl chloride described in Example 3, followed by the subsequent synthetic steps described therein gives 1-(2-N-methylsulfamoylethyl)tetrazole-5-thiol. - 18 44607 When l-(2-N-methylsulfamoylethyl)tetrazole-5thiol Is reacted with 7-aminocephalosporanic acid as described in Example 3 and the product 7-amino-3-(1-(2-Nmethylsulfamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4carboxylic acid,is acylated with D-O-dichloroacetylmandeloyl chloride as described therein, the title compound is obtained.

EXAMPLE 6 7-(D-a-Amino-4-hvdroxvphenylacetamido) -3- L1- (2-s ulfamoylethyDtetrazoI-S'-yithioraetnyll -3-cephem-4-carboxylic acid A solution of 7.82 g. (0.015 mol.) of 7-(D-a-_tbutoxycarbonyl-4-hydroxyphenylacetamido)cephalosporanic acid, 4.6 g. (0,022 tool.) of l-(2-sulfamoylethyl)tetrazole5-thiol and sodium bicarbonate is reacted according to the procedure described in Example 2. After cooling, the reaction mixture is extracted with ethyl acetate. Fresh ethyl acetate is added to the aqueous phase and it is acidified with stirring to pH 2.8 with 6N sulfuric acid.

The layers are separated and the aqueous phase is again extracted with ethyl acetate. The combined extracts are washed with water, dried (MgSO^) and the solvent evaporated to give 7-(D-a-t:-butoxycarbonyl-4-hydroxyphenylacetamido)3-(1-(2-sulfamoylethyl)tetrazol-5-ylthiomethyl]-3-cephera-4carboxylic acid. 7-(D-a-_t-Butoxycarbonyl-4-hydroxyphenylacetamido)3-(1-(2-eulfamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4carboxylic acid is treated with trifluoroacetic acid as described in Example 2 to give the title compound.

EXAMPLE 7 7-D-tfandelamldo-3-fl-(2“8ulfoethvl)tetrazol-5-vlthiomethvll3-cephem-4-carboxylic acid 2-Aminoethanesulfonic acid (50 g., 0.4 mol.) was - 19 44607 added to a solution of 45 g. (0.8 mol.) of potassium hydroxide in 100 ml. of water at 25°. Carbon disulfide (24.4 ml., 0.4 mol.) was added and the reaction mixture was refluxed for 2.5 hours. Ethanol was added to the warm solution, the mixture was cooled to ambient temperature, 57 g. (0.4 ssol.) of methyl iodide was added and the resulting mixture was stirred for 1.5 hours. The mixture was evaporated in vacuo and the residue recrystallized from hot ethanol containing 3% water to give methyl 2-sulfoethyidithiocarbamate potassium salt.

A mixture of 21.5 g. (0.087 mol.) of methyl 2-sulfoethyldithioearbamate potassium salt (0.5 hydrate) and 7.16 g. (0.11 mol.) of sodium azide in 200 ml, of water was refluxed for two hours. The solution was cooled to 25° and extracted with ethyl acetate. Tlie aqueous phase was treated'with Amberlite IR-120H resin, washed with ether and evaporated to give an oil. The oil was taken up in acetone, the mixture was filtered and the filtrate was evaporated to dryness to give l-(2-sulfoethyl)tetrazole-5-thiol. The thiol was dissolved in isopropanol, cyclohexylamine was added until pH 8-9 and acetonitrile was added to give l-(2-sulfoethyl)tetrazole-5-thiol as the bis-cyclohexylamine salt. 1-(2-Sulfoethyl)tetrazole-5-thiol bi s-cyclohexy1amine salt was dissolved in water and treated with Amberlite IR-120H resin to give l-(2-sulfoethyl)tetrazole« 5«thiol.

To a solution of 2.1 g. (0.01 mol.) of 1-(2sulfoethyl)tetrazole-5-thiol in 100 ml. of water was added 6.4 g. (0.015 mol.) of 7-D-mandelamidocephalosporanie acid sodium salt and 1.68 g. (0.02 mol.) of sodium bicarbonate. - 20 <4607 The mixture was stirred at 70° for 2.5 hours then cooled and acidified to pH 1.8 with 3N hydrochloric acid. The acid solution was extracted with ethyl acetate and ether, an Amberlite acidified to pH 0.9 and chromatographed on/ XAD-8 resin column with water as eluant to give the title compound.

The title compound was converted to the corresponding disodium salt by treatment with sodium methoxide as described in the procedure of Example 1.

C19H18N6°8S3 · 2 Na · 2 H20 Calculated: 35.85% C; 3.48% H; 13.20% N Found: 36.21% C; 3.29% H; 12.96% N EXAMPLE 8 y-D-Mandelamido-S-fl-sulfanjOvlroethyltetrazol-S-ylthioinethyl·)3-cephem-4-carboxylic acid A suspension of 15.1 g. (0.136 mol.) of aminomethanesulfonic acid and 14.2 g, (0.145 mol.) of anhydrous potassium acetate in 48 ml. of acetic acid is refluxed for ten minutes. Phthalic anhydride (21.4 g., 0.145 mol.) is then added and the resulting mixture is refluxed for 2.5 hours. The product is collected by filtration and washed with acetic acid and ethanol to give phthalimidomethanesulfonic acid potassium salt.

To 41.7 g. (0.15 mol.) of phthallmidomethanesulfonic acid potassium salt in 220 ml. of dry benzene is added 22.5 g, (0.132 mol.) ot phosphorus pentachloride.

The reaction mixture is refluxed on a steam bath for one hour, then an additional 22.5 g. of phosphorus pentachloride is added and heating is continued for 1.5 hours. The reaction mixture is evaporated to dryness, crushed ice is added to the residue and the slurry is filtered. The product is washed with water to give phthaltmidomethane- 21 10 sulfonyl chloride.

When phthalimidomethanesulfonyl chloride is substituted in the procedure of Example 3 for 2-phthalimidoethanesulfonyl chloride, N-£-butylphthaliinidomethanesulfonamide is prepared and is converted to l-N-t-butylsulfamoylmethyltetrazole-5-fchiol as described therein. Treatment of l-N-Jt-butylsulfamoylmethyltetrazole-5-thiol with trifluoroacetic acid as described in Example 3 gives l-sulfamoylmetthyltetrazole-5-thiol.

Reaction of l-sulfamoylmethyltetrazole-5-thiol with 7-aminocephalosporanic acid and treatment of the product 7-amino-3-(l-sulfamoylmethyltetrasol-5-ylthiomethyl) -3-cephem-4-carboxyllc acid with D-G-dichloroacetylmandaloyl chloride as described in Example 3 gives the title compound.

EXAMPLE 9 7-(D-g-Amino-4-hvdroxyphenylacetamido)-3-[1-(2-N,N-dimethvlsulfamoylethvl)tetrazol-3-ylthiomethyl·j-3-cephem-4-carboxvlic acid 1-(2-N,N-Dimethylsulfamoylethyl)tetrazole-5-thiol is prepared by reaction of dimethylamine and 2-phthalimidoethanesulfonyl chloride· followed by conversion of the product thus obtained to the tetrazole thiol by the reaction sequence described in the procedure of Example 3.

When l-(2‘-N,N-dimethylsulfamoylethyl)tetrazole-5thiol is reacted with 7-(D-a-£-butoxycarbonyl-4-hydroxyphenylacetamido)cephalosporanic acid as described in Example 6 and the product is de-blocked as described in Example 2. the title compound is obtained.

EXAMPLE 10 7-D-Mandelamtdo-3Ll-(3-8ulfopropyl)tetrazol-5-ylthlo-carboxvlic acid When an equivalent amount of 3-aminopropane30 - 22 4 4 607 sulfonic acid was substituted in the procedure of Example 7 for 2-aminoethanesulfonic acid, methyl 3-sulfopropyldithiocarbamate potassium salt was prepared.

Reaction of methyl 3-sulfopropyldithiocarbamate potassium salt with sodium azide, as described in Example 7, gave l-(3-sulfopropyl)tetrazole-5-thiol.

Substitution of an equivalent amount of 1-(3sulfopropyl)tetrazole-5-thiol in the procedure of Example 7 in place of l-(2-sulfoethyl)tetrazole-5-thiol in the reaction with 7-D-mandelamidocephalosporanic acid sodium salt gives the title compound.

The title compound is converted to the corresponding sodium salt as described in the procedure of Example 1.

EXAMPLE 11 7-Ρ-Μ8ΐκ1βΐ3ΐη1άο-3-[1-(3-8α1£3ΐηον1ρΓθρν1)ϋ6^Ηζο1-5-ν1^1οmettiylj -3-cephem-4-carboxylic acid When 3-phthalimidopropanesulfonyl chloride is substituted in the procedure of Example 3 for 2-phthalimidoethanesulfonyl chloride, N-t-butyl-3-phthalimidopropanesulfonamide is prepared, and is converted to l-(3-N-Jtbutyleulfamoylpropyl)tetrazole-5-thiol as described therein. Treatment of l-(3-N-.t~butylsulfamoylpropyl)tetrazole-5-thiol with trifluoroacetic acid as described in Example 3 gives 1-(3-sulfamoylpropyl)tetrazole-5thiol.

Reaction of l-(3-sulfamoylpropyl)tetrazole-5-thiol with 7-aminocephalosporanic acid and treatment of the resulting 7-amino-3-[1-(3-sulramoylpropyl)tetrazol-5ylthiomethyl]-3-cephem-4-oarboxylic acid with D-O-dichloroacetylmandeloyl chloride as described in Example 3 gives the title compound. - 23 44607 EXAMPLE 12 7-D-4-iandelaaido-3~i 1-(5-sulfopentyl) tetrazol-5-yl thiomethyl] °3-cephsm-4-earboxvlieaeld Substitution of an equivalent amount of 5-aminopentanesulfonic acid in the procedure of Example 7 for 2-aminoethanesulfonic acid gave methyl 5-sul£opentyldithiocarbamate potassium salt.

Reaction of methyl 5-sulfopentyldithiocarbamate potassium salt with sodium azides as described in Example 7>gave 1-(5-sulfopentyl)tetrazole-5-thiol.

A mixture of 4.55 g. (0.010 mol.) of 7-D-rnandelamidocephalosporanic acid sodium salt and 3.25 g. (0.011 mol.) of l-(5-sulfopenfcyl)tetrazole-5-thiol disodium salt, in Example 1, prepared as previously described/ in 80 ml. of water at pH 7.2 (adjusted by addition of sodium bicarbonate) was heated at 65° for 4.5 hours. The reaction mixture was cooled, acidified to pH 1.6 with 3N hydrochloric acid and extracted with ethyl acetate. The pH of the aqueous phase was brought to 7.0 by addition of sodium bicarbonate an Amber! ite and the solution was chromatographed on / XAD-4 resin column eluting with water and then methanol. The product was dissolved in methanol and ethanol was added to the solution’ to precipitate the title compound as its disodium salt.

C22H24N6°8S3 ' 2 Na ' 1,5 H2° ' 0,75 C2H6° Calculated: 40.08% C; 4.50% H; 11.93% N; 13.65% S Found: 40.01% C; 4.20% H; 10.71% N; 13.54% S 7-D-Mandelamido»3-il-(5-sulfopentyl)tetrazol-5ylthiomethyl]-3-cephem-4-carboxylic acid disodium salt is converted to the title compound as described in Example 1 - 24 4 4 6 0 7 EXAMPLE 13 7-P-Mandelamldo-3-[1-(5-sulfamovlpentyl)tetrazol-5-y1thiomethyl J -3-cephem-4-carboxylic acla Use of 5-aminopentanesulfonic acid in the procedure of Example 8 in place of aminomethanesulfonic acid > followed by reaction of the product thus obtained with phosphorus pentachloride, gives 5-phthallmidopentanesulfonyl chloride.

When 5-phthalimi.dopentanesulfonyl chloride is ueed as a starting material in the sequence described in Example 3, l-(5-N-jt-butylsulfamoylpentyl)tetrazole-5-thiol is obtained. Treatment of l-(5-N-t-butylsulfamoylpentyl)tetrazole-5-thiol with trifluoroacetic acid as described above gives l-(5-sulf amo ylpentyl)tetrazole-5-thiol.

Reaction of l-(5-sulfamoylpentyl)tetrazole-5thiol with 7-aminocephalosporanic acid and treatment of the resulting 7-amino-3-[l-(5-sulfamoylpentyl)tetrazol5-ylthiomethylJ-3-cephem-4-carboxylic acid with D-Odichloroacetylmandeloyl chloride as described in Example 3 gives the title compound.

EXAMPLE 14 7-Trifluoromethvlthioacetamido-3-(l-sulfomethvl“ tetrazol-5-vlthix)iaethvi)-3-cephem-4-carboxyllc acid A solution of 2.18 g. (10.0 nmol.) of 1-sulfomethyltetrazole-5-thiol sodium salt, prepared as described in Example 1, 0.840 g. of sodium bicarbonate and 5.45 g. (12.5 mmol.) of 7-trifluoromethylthioacetamidocephalosporanic acid sodium salt in 60 ml. of water was stirred at 70-75° for five hours while maintaining the pH at 6.8 by addition of 5% aqueous sodium carbonate solution. The reaction mixture was cooled and diluted with water.

Ethyl acetate was added and the mixture was acidified - 25 to pH 2.0 with ® hydrochloric acid. The aqueous phase was extraetod with ethyl acetate then brought to pH 6.8 by addition of sodium bicarbonate and chromatographed on Amberlite /XAD-4 resin with water and ssthanol as eluants. The product-containing fractions were evaporated to dryness to give a residue which was dissolved in water and lyophilised to give the title Compound as the disodium salt.

C13H11F3K6°7S4 ‘ 2 tJa ’ 2·25 «2° Calculated: 24.597. C; 2.467. H; 13.23% N Found: 24.54% C; 2.18% H; 12.85% N An aqueous solution of 7-trifluoromethyl thioacetamido-?3-(l=sul£oiB2thyltetrasol-5-ylthiomethyl)-3cephem-4-earboxylic acid disodium salt is treated with Amberlite IR-120H ion exchange resin as described in Example 1 to give the title compound.

EXAMPLE 15 acid 7-Trifluoromathy1thioeeetamido-3-[1-(3-sulfamoylpropyl)tetrazol-5-ylthianethyl]-3-cephem-4carboxylie acid sodium salt is prepared by substitution of an equivalent amount of l-(3-3ulfamoylpropyl)tetrazole5-thiol sodium salt, prepared as described in Example 1, in the procedure of Example 14.

The title compound is obtained from the salt as described above.

EXAMPLE 16 Trif luoromethy 2-th. itrazol-5-ylthioiael tetrazo ioaeetamido-3aet 11A mixture of 2,18 g, (5 mmol.) of 7-trifluoro- 26 4 4 60 methylthioacetamidocephaloeporenic acid sodium salt and 1.75 g. (8 mmol.) of l-(2-sulfamoylethyl)tetrazQle-5thiol In 50 ml. of water, maintained et pH 7.0 by addition of sodium bicarbonate,was heated at 60-70° for three hours. The reaction mixture was cooled, acidified to pH 3.5 and extracted with ethyl acetate. The extract was washed with water, dried (MgSO^) and evaporated to dryness to give a residue which was dissolved in ethyl acetate. Ether was added, the solution was filtered, petroleum ether was added to the filtrate and the resulting precipitate was collected by filtration and dissolved in methanol. A 5% solution of sodium methoxide in methanol was added to the methanol solution until pH 7.1, Ether was added and the precipitate was collected and dried in vacuo to give 7-trifluoromethylthioacetamido - 3-[l~(2-sulfaraoylethyl)tetrazol-5ylthiomethyl]-3-cephem-4-carboxylic acid sodium salt. 7-Trifluoromethylthioacetamido-3- [ 1- (2-sulfamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid sodium salt is converted to the title compound by methods described in Example 1.

EXAMPLE 17 7-D-Mandelamldo-3-Il-(10-8ul.fodecyl)tetrazol-5-vlthlomethvl]-3-ccphem-4-carboxylic acid A suspension of 56 g. (1.0 mol.) of potassium hydroxide and 118.7 g. (0.5 mol.) of 10-aminodecanesulfonic acid in 170 ml. of water is stirred for 30 minutes at 25° then 40 g. (0.52 mol.) of carbon disulfide and 80 ml. of ethanol are added and the reaction mixture is stirred at 25° for 12 hours. The mixture is refluxed gently for two hours and cooled. Methyl iodide (71 g., - 27 44607 0.3 mol.) and 130 ml. of ethanol are added to the mixture and it is stirred at 25° for 12 hours. The mixture is evaporated to remove the ethanol and tho solid residue is collected by filtration to give methyl 10-sulfodecyldithiocarbemate.

Methyl 10-oulfodecyldithioearbamate (31.4 g., 0.096 sol.) io reacted with 6.5 g. (0.1 mol.) of sodium Example 1 aaid© as described .in / to give X-(lO-aulfodeeyl)tetrazole-5-thiol. l-(lQ-Sul£adecyl)tetrasole-5-thiol (4.84 g., mmol.) is slowly added to a solution of 3.36 g. (40 mmol.) of sodium bicarbonate in 100 ml. of water. 7-D-ifandelamidosephalosparanic acid (4.20 g., 10 nmol.) is than added and the mixture is heated at 65° for 3.5 hours. The mixture is filtered, the filtrate is extracted with ethyl acetate and the aqueous layer is acidified to pH 4 and extracted again with ethyl acetate. The exttact is dried (MgSO^) and evaporated to dryness to give the title compound.

EXAMPLE 18 7-P-Mamdelamido-3-(l-(lO-sulfamoyldecvDtetrazol-S-vlthioUse of 10-aminodeeanesulfonic acid in the procedure of Example 8 in place of aminamethanesulfonic acid’followed by reaction of th® product thus obtained with phosphorus pentachlorida, gives 10-phthalimidodeeanesulfonyl chloride.

Wsen 10-phthalimidodeeanesulfonyl chloride is used as a starting material in the sequence described in Example 3, l-(10-N-t,-butylaulfamayldecyl)tetrazole-5thiol ia obtained. Treatment of l-(10“iJ-t-butyleulfamoyldecyl)t®traz©le-5-thiol with trifluoroacetic acid gives - 28 4 4 6 0 7 1- (10-sulfamoyIdeey1)tetrazole-5-thiol.

Reaction of 1-(10-sulfamoyldecyl)tetrazole-5-thiol with 7-aminocephalosporanic acid and treatment of the resulting 7-amino-3-[1-(10-sulfajnoyldecyl)tetrazol-5ylthiomethyl]-3-cephem-4-carboxylic acid with D-O-dichloroacetylmandeloyl chloride as described in Example 3 gives the title compound.

EXAMPLE 19 y-D-Mandelamido-S-tl-CZ-sulfp-l-methylethyDtetrazol-SSubstitution of an equivalent amount of 2-aminopropanesulfonic acid in the procedure of Example 7 for 2- aminoethanesulfonic acid gives methyl (2-sulfo-l-methyl ethyl)dithiocarbamate potassium salt.

Treatment of methyl (2-sulfo-l-methylethyl) dithiocarbamate potassium salt with sodium azide also as described in Example 7 gives l-(2-sulfo-l-methylethyl)tetrazole-5-thiol. 7-D-Mandelamldocephalosporanic acid and 1-(2-sulfol-methylethyl) tetrazole-5-thiol are reacted in the presence of excess sodium bicarbonate as described in Example 7 to give the title compound.

EXAMPLES 20 and 21 Reaction of the sodium salt of a cephalosporanic acid listed below: 7-(2,2,2-trifluoroethylthioacetamido)cephalosporanic acid 7-methy1th ioacetamidoce phalo sporanic ac id with l-(2-sulfamoylethyl)tetrazole-5-thiol sodium salt by the procedure of Example 14 gives the 7-substituted-3- 29 44607 [1-(2-sulfamoylethyl)tetrazol-5-ylfchiomethyl]-3-cephem-4carboxylic acids listed below: 7-(2,2,2-trifluoroethylthioacetamido)-3-(1-(2sulfamoylethyl)teteazol-5-ylthiomethyl]-3-cephem-45 carboxylic acid (Example 20).

C15H17F3N7°6S4 ° Ha ’ 0,5 ^4° Calculated: 30.24% C; 3.11% H; 15.92% N Found: 30.62% C; 3.00% H; 15.32% N 7-ni2thylthioacetainido”3- [ 1= (2-sulfamoylethyl) θ tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid (Example 22) EXAMPLE 22 7-(2-Thienylacetamido)-3-(l-sulfomathyltetrazol-S-ylthlomathyl) -3-cephem-4-carbo;syfic acid Reaction of 2.18 g. (0.01 mol.) of l-sulfomethyltetrazole-5-thiol sodium salt, 5.23 g. (0.012 mol.) of 7(2-thienylacetamido)cephalosporanic acid sodium salt and 0.84 g. (0.01 mol.) of sodium bicarbonate as described in the procedure of Example 14 gave the title compound as the corresponding disodium salt.

C16H14N6°7S4 * 2 Na 0.75 C^O Calculated: 34.81% C; 3.09% H; 13.56% N Found: 34.41% C; 3.03% H; 13.76% N 7-(2-Thienylacetamido)-3-(l-sulfomethyltetrazol-5ylthiomethyl)-3-cephem-4-carboxylic acid disodium salt is converted to the title compound as described in Example 1.

EXAMPLE 23 Reaction of 5.68 g. (0.01 mol.) of 1-(2-sulfoethyl)- 30 tetrazole-5-thiol cyclohexylamine salt and 5.23 g. (0.0125) mol.) of 7-(2-thienvlacetamido)cephalosporanic acid sodium salt tn 80 ml. of water containing sufficient sodium bicarbonate to maintain the pH at 7.0, as described in the procedure of Example 14, gave the title compound.

Preparation of / 7-Amino-3-(l-.i=;ulfomethyltetrazol-5-yithiomethyl)-3cephem-4-carbcxylic acid (starting material'in Example £4) To a mixture of 97 g. (200 ml., 2.1 mol.) of formic acid, distilled from anhydrous copper sulfate, and 37.5 ml. (0.4 mol.) of acetic anhydride was added 25.0 g. (0.1 mol.) of 7-aminocephalosporanic acid. The mixture was stirred at ambient temperature for 0.5 hour, then evaporated to dryness. The residue was dissolved in ethyl acetate and the ethyl acetate solution was filtered and evaporated to dryness tc give a residue which was recrystallized from ether-petroleum ether to give 7-formamidocephalosporanic acid.

A mixture of 1.0 g. (3.3 mmol.) of 7-formamidocephalosporanic acid and 0.7 g. (2.6 mmol.) of 1-sulfomethyltetrazole-5-thiol disodium salt in 15 ml. of water was stirred at 65-70° for 3 hours while maintaining the pH at 7.0 by addition of sodium bicarbonate and/or hydrochloric acid. The mixture was cooled, acidified to pH 1.0 with hydrochloric acid and extracted with ethyl acetate. The extract was filtered and the filtrate was evaporated to dryness to give a residue which was dissolved in 30 ml. of methanol. The methanol solution was filtered, 30-40 ml. of Isopropanol and ethanol were added, the solution was filtered again and 100 ml. of ether was added. The precipitate was collected by filtration and dried to give the title compound. - 31 · C10H12°6M6S3 “θ’33 G3Hg° HCl · 2 H2O Calculated; 27.07. C; 3.507. H; 16.95% H Found: 26.96% C; 3.23% H; 16.55% N W.IIKS 24 7-(fr-Pyridylthioacetamido)-3-(l-sulfcmathyltetrazol-5(4-Fyridylthio)acetyl chloride (0.53 g., 2.8 mmol.) was dropwise added to a mixture of 1.0 g. of 7-amino-3(l-sulfomathyltefcra3ol-5-ylthiomethyl)-3-ceph6m-4-carboxylic acid (mixture of sodium salt, free acid and sodium chloride) and 0.9 g. (9.0 mmol.) of triethylamine in 10 ml. of dry dimethylformamide. The reaction mixture was stirred for 1.5 hour at -10°, then it was warmed to ambient temperature and stirred for 1 hour. The mixture was filtered and the filtrate was diluted with 200 ml. of ether-petroleum ether. The precipitate was collected by filtration and dissolved in warm methanol. The methanol solution was filtered and the filtrate was concentrated to 10 ml. Ethanol (50 ml.) was added and the precipitate was collected by filtration and washed with ether to give the title compound as the sodium salt. Additional amounts of the salt were obtained by addition of ether to the filtrate. ^17^16^7^7^4 ° ΰ HaCl ° C^H^^N Calculated: 37.237. C; 4.217. H; 15.107. N Found: 36.847. C; 4.487. H; 15.437. N 7=(4-Pyridylthioaeetamido)-3-(1-sulfomethyltetrazol 5-ylfhioBi3thyl)“3-cephem4-carboxylic acid sodium salt is converted to the title compound by procedures described -in Example 1. - 32 J 4 6 U 7 EXAMPLES r'b to '1') Examples /, 12, 14 and 24 Using the procedures described 'rl / for the preparation of compounds of this invention, the following compounds were prepared: 7-(l-tetrazolylacetamido)--3-(l-sul£omethyltetrazol-5-ylthiomethyl)-3-cephem“4-carboxyl.!.c acid (Example 25) C13H12N10°7S3 Na Calculated: 27.76% C-; 2.15% II; 24.90% N Found: 27.55% C; 3.21% H; 23.00% N 7-(sydnon-3-ylacetamido)-3-(l-sul£omethyltetrazol-5ylthlomethyl)-3-cephem-4-carboxylic acid (Example 26) C14H10N8°9S3 * 2 Na · 3 H20 · 2 CH^O Calculated: 27.59% C; 3.72% H; 16.08% N Found: 27.59% C; 3.02% H; 15.67% Ν' 7-trifluoromethvlthioacetamido-3-[l-(4-sulfo(Example 27) butyl)tetrazol-5-ylthiomethyl]-3-cephem-4-carboxylic acid / C15H17F3N6°7S4 Na h2o Calculated: 29.36% C; 2.93% H; 12.84% N Found: 29.31% C; 3.33% H; 12.76% N 7-(2-thienylacetamido)-3-[l-(4-sulfobutyl)tetrazol5-ylth£omethyl]-3-cephem-4-carboxylic acid (Example 28) C19H20N6°7S4 Na H2O C2HgO Calculated: 34.85% C; 3.69% H; 12.83% N; 19.59% S Found: 34.52% C; 3.88% H; 13.54% N; 19.09% S 7-(2-thienylacetamido-3-[l-(5>sulfopentyl)tetrazol5-ylthiomethyl]-3-cephem-4-earboxylic acid (Example 29) C20H22N6°7S4 ’ 2 Na ‘ 2·5 H20 · 1.75 C2H6O - 33 Calculated: 36.33% C: 4.77% H: 11.29% N: 17.23% S Found: 36.46% C: 4.30% H: 11.63% N: 17.30% S EXAMPLE 30 7-(2-Thienylacetamido)-35-ylthiomathyl1-3-c -(2-sulphamoylethyl)tetrazolacid Reaction of 2-thiopheneacetyl chloride and 7-amino-3[l-(2-sulphamoylethyl)tetrazol-5-ylthiomethyl]-3-cephem4-carboxylic acid according to the procedure of Example 24 yielded the title compound.

EXAMPLE 31 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-sulphamoylethyl)tetrazol-4-ylthiomethyl]-3-cephem-4-carboxylic acid sodium salt.

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

Claims

1. A cephalosporin compound of the structural formula: in which: 10 each individual R 1 is hydrogen or lower alkyl; n is from 1 to 10; Itf is hydroxy, amino, (.lower alkyl)amino or di(lower alkyl)amino; R is an acyl gi'oup of the structure 15 X^HCO-, YCH 2 C0- or Z-S(O) m -CH 2 COA where: X is thienyl; dihydrophenyl; phenyl; phenyl monosubstituted with hydroxy, hydroxymethyl, formamido, 20 ureido or carboxymethylamino; or 3-fluoro-4-hydroxyphenyl; A is hydroxy, amino, COOR or 60,^11; or formyloxy when X is phenyl.; Y is thienyl, tetrazolyl, cyano, sydnon-3-yl or 25 aminomethylphenyl; Z is methyl, trifluoromethyl, trifluoroethyl, cyanomethyl or pyridyl; and ra is 0, 1 or 2; and each COOR is a carboxylic acid or esterified 30 carboxylic, acid group; or a non-toxic pharmneeutieally-acceolable salt thereof.

2. A compound according to Claim 1 in which each COOR is 35 a carboxylic acid group.

3. A compound according to Claim 1 or Claim 2 in which each R 1 is hydrogen and n is from 1 to 5. 40

4. A compound according to Claim 3, in which n is 1 or 2. -55.

5. Λ compound according to any preceding claim in which II 2 is hydroxy or amino.

6. A compound according to any preceding claim in which R is XCHCO- where X is phenyl or hydroxyphenyl and A is A hydroxy or amino.

7. A compound according to any one of Claims 1 to 5 in which 10 R 3 is YCHgCO- where Y is thienyl,

8. A compound according to any one of Claims 1 to 5 in . which R is YCHgCO- where Y is 2-aminomethylphenyI. 15

9. A compound according to any one of Claims 1 to 5 in which H 3 is Z-S(O) m -CII 2 CO- where Z is trifluoromethyl and in is 0.

10. 7-D-mandelamido-3-(1-sulfomethylte I: razol-5-ylthiomethyl)20 3-cephem-4-carboxylic acid.

11. 7-D-mandelamido-3-/i-(2-sulfaethyl)tetrazol-5-ylthiomethyl.7-3-cephem-4-carboxylic acid. 25

12. 7-D-mandelamido-3-/l-(5-sulfopentyl)tetrazol-5-ylthiomethyl7-3-cephein-4-carboxylic acid.

13. 7-D-mandelamido-3-/l-(2-sulfamoylethyl)tetrazol-5ylthiomethyl7-3-cephem~4-carboxylic acid.

14. 7-(2-Thienylacotamido)-3-(l-suliomethyltetrazol-5ylthiomethyl)-3-cephem-4-carboxylic acid.

15. 7-(2-Thienylacetamido)-3-/!-(2-sulfoethyl)tetrazol-535 . ylthiomethylJ-3-cephem-4-carboxylic acid.

16. 7-(2~Thienylacetamido)-3-/l-(2-sulfamoylethyl)tetrazol-5-ylthiomethyl7-3-cephem-4-carboxylic acid. 40

17. 7-Trifluoromethylthioacetamido-3-(l~sulfomethyltetrazol-5-ylthiomethyl)-3-cephem-4-carboxylic acid.

18. 7-Trifluoromethylthioacetamido-3-/i _ (2- s ulfamoylethyl)tetrazol-5-ylthiomethyl7-3-cephem-4-earboxylic acid. -364 4 6 0 7

19. The compound accordin ;; to Claim 1 of any one of Examp ley 2, 4 to ύ. 8 to 11, 13, !.ό, IV t.o '?.! and 24.

20. A process for preparing a compound according to any 5 one of Claims 1 to 19, where each COOP.' 1 is a carboxylic acid group, in which 7-aminoceohaiosporanie acid is acylated with an appropriately protected acylating agent capable of providing the acyl group R‘ J and the 3-acetoxy group in the acylation product is displaced by reaction with the appropriate 10 tetrazole-thiol of the structure HS i| . N 15 (CHH 1 ) n -SO 2 H 2 and any protecting group is subsequently removed.

21. A process fcr preparing a compound according to any one of Claims 1 to 19. where each COOR' 1 is a carboxylic 20 acid group, in which the appropriate 7-amino-3-substituted tetrazolylthiomethyl cephalosporin of the structure 12. 5 where R and R are as defined in Claim 1 and R is hydrogen or a protecting ester-forming group, is acylated with an appropriate acylating agent, followed by removal of any 30 protecting group.

22. A process for preparing a compound according to Claim 1 substantially as described in any one of Examples 1 to 24 and 30.

23. A compound according to Claim 1 whenever prepared by a process according to any one of Claims 20 to 22.

24. A pharmaceutical composition comprising a compound 40 according to any one of Claims 1 to 19 and 23 and a pharmaeeutically-aeceptable carrier.

25. A pharmaceutical composition according to Claim 24 and substantially as described in Example 31. -37

26. A method of combatting bacterial infection in a non-human animal which comprises administering to the animal a pharmaceutical composition as claimed in Claim 24 or Claim 25 in a non-toxic amount sufficient to combat such infection.