GB2177691A - Cephalosporin antibiotics - Google Patents

Abstract

Cephalosporin antibiotics of general formula (I) are disclosed <IMAGE> (wherein Y represents a carbon-linked 5- or 6-membered unsaturated heterocyclic ring containing at least one nitrogen atom, which ring may also contain one or more sulphur atoms and/or may be substituted by one or more C1-4 alkyl, oxo, hydroxy or carbamoylmethyl groups) and non-toxic salts and non-toxic metabolically labile esters thereof.

Description

SPECIFICATION Cephalosporin antibiotics This invention relates to improvements in or relating to cephalosporins. More particularly it relates to new cephalosporin compou nds and derivatives thereof having valuable antibiotic activity.

The cephalosporin compounds in this specification are named with reference to "cepham" afterJ.Amer.

Chem. Soc., 1962,84,3400, the term "cephem" referring to the basiccepham structurewith one double bond.

Cephalosporin antibiotics are widely used in the treatment of diseases caused by pathogenic bacteria in human beings and animals, and are especially useful in the treatment of diseases caused by bacteria which are resistantto other antibiotics such as penicillin compounds, and in the treatment of penicillin-sensitive patients. In many instances it is desirable to employ a cephalosporin antibiotic which exhibits activity against both Gram-positive and Gram-negative microorganisms, and a significant amount of research has been directed to the development ofvarioustypes of broad spectrum cephalosporin antibiotics.

Thus, for example, in our British Patent Specification No. 1399086, we described a novel class of cephalosporin antibiotics containing a 7p-(a-etherified oxyimino)-acylamido group, the oxyimino group having thesyn configuration. This class of antibiotic compounds is characterised by high antibacterial activity against a range of Gram-positive and Gram-negative organisms coupled with particularly high stabilityto p-lactamases produced by various Gram-negative organisms.

The discovery of this class of compounds has stimulated further research in the same area in attempts tofind compounds which have improved properties,forexample against particularclasses oforganisms, especially Gram-negative organisms. This interest is reflected in the very large numbers of patent applications which have been filed relating to cephalosporing antibiotics having particular oxyimino etherifying groups in combination with particularsubstituents both on the 7i3-acylamido side chain and at the 3-position ofthe cephalosporin nucleus.

British Patent Specification No. 1576625 discloses generally a large number of cephalosporin antibiotics having in the 7ss-position an (etherified oxyimino)acetamido group substituted in the 2-position by a substituted phenyl orthiazolyl group. 2-(2-Aminothiazol-4-yl)-2-(etherified oxyimino)acetamido groups are mentioned wherein the oxime etherifying group is an aliphatic hydrocarbon group which may have suitable substituted(s) but halogen is not included in the list of suitable substituents. The 3-substituted may be, inter alia, a heterocyclicthiomethyl group which may have suitable substituent(s).

In British Patent Specification Nos. 1604971 and 1603212 a wide variety of cephalosporin antibiotics are generally disclosed in which the 7ss-position side-chain may be selected from, interalia, a 2-(2-aminothiazol-4yl)-2-(etherified oxyimino)acetamido group, in which the etherifying group, amongst very many possible meanings, may bean alkyl group (e.g. methyl) substituted bya halogen atom,althoughthereare noworking examples in either case of compounds having such a group and the preferred etherifying group is stated to be an unsubstituted methyl group. Halogen atoms which, when present, are preferred are stated to be chlorine and bromine atoms.The 3-position group may also be selected from a very large number of alternatives and a possible 3-substituent is a thiomethyl group substituted by an optionally substituted 5-or 6-membered heterocyclic group containing one or more nitrogen, oxygen or sulphur atoms. British Patent Specification No.

1604971 describes cephalosporins having sulphuratthe 1-position while British Patent Specification No.

1603212 describes related 1-sulphoxides, Our British Patent Application Specification No. 2027691 A describes cephalosporin antibiotics in which the 7P-side chain is a 2-(2-aminothiazol-4-yl)-2-(etherified oximino)acetamido group, where the etherifying group is a carboxyalkyl orcarboxycycloalkyl group. The 3-substituent is a group offormula -CH2SY in which Yisa carbon-linked 5-or 6-membered heterocyclic ring containing at least one nitrogen atom, which may be substituted by a CiA alkyl group.

In British PatentApplication No. 2039890A a wide variety of cephalosporin antibiotics are generically disclosed in which the 7ss-position side chain is a 2-(2-aminothiazol-4-yl)-2-etherified oxyimino)-acetamido group. One possible etherifying group recited is a halo-lower-alkyl group (with thefluoromethyl group being mentioned interalia as an illustration). The only haloalkyl compounds actually exemplified contain a 2 bromoethyl, 2-chloroethyl or a 2,2,2-trifluoroethyl group. According to the generic definition, the 3-position may carry a heterocyclicthiomethyl group.

British PatentApplication No. 2017702A describes etherified oxyimino cephalosporin antibiotics in which the oxyimino etherifying group, according to the generic definition, may interalia be a straight chain 4alkyl group terminally monosubstituted e.g. by a halogen atom. In the compounds specifically exemplified, only 2-bromoethyl and 2-iodoethyl groups are found as examples of haloalkyl groups. The groups atthe3-position includes optionally substituted heterocyclicthiomethyl substituents.

We have now discovered that by the selection of a (Z)-2-(2-aminothiazol-4-yl)-2-(etherified oxyimino)acetamido group atthe7ss-position in combination with certain heterocyclicthiomethyl groups atthe3position, and also by the selection of a monofluoro methoxyimino group as the etherified oxyimino grouping, cephalosporin compounds having a particularly advantageous profile of activity (described in more detail below) against a wide range of commonly encountered pathogenic organisms may be obtained. Accordingly, we provide cephalosporin antibiotics ofthe general formula (I)

(wherein Y represents a carbon-linked 5- or6-membered unsaturated heterocyclic ring containing at least on nitrogen atom, which ring may also contain one or more sulphur atoms and/or may be substituted by one or more C1.4 alkyl, oxo, hydroxy or carbamoylmethyl groups) and non-toxic salts and non-toxic metabolically labile esters thereof.

The compounds according to the invention are syn isomers. Thesyn isomeric form is defined bythe configuration of the -O-CH2-Fgroupwith respectto thecarboxamido group. In this specification,thesyn configuration is denoted structurally as

It will be understood that sine the compounds according to the invention are geometric isomers, some admixture with the corresponding anti isomer may occur.

The invention also includes within its scope the solvates (especially the hydrates) of the compounds of formula (I). Italso includes within its scope the solvents of non-toxic salts offormula (I), and non-toxic salts and solvates of non-toxic metabolically labile esters ofthe compounds offormula (I). Itwill be appreciated thatthe solvates should be pharmacologically acceptable.

In formula (I) above, the heterocyclic ring represented byY may for example contain 1 to 4 nitrogen atoms and, if desired, a sulphuratom, particular examples of these heterocyclicgroups including imidazolyl, pyrazolyl, pyridyl, pyrimidyl, pyrazinyl, pyridazinyl, triazolyl, tetrazolyl, thiazolyl, thiadiazolyl, triazinyl and thiazolidinyl. The heterocyclic ring may, if desired, be substituted by one or more C1.4 alkyl (e.g. methyl) oxo, hydroxyl, or carbamoylmethyl groups. Alkyl or carbamoyl methyl substitutents may be attached for example, to the nitrogen heteroatom(s).

The compounds according to the invention exhibit broad spectrum antibiotic activity both in vitro and in vivo. They have high activity against both Gram-positive and Gram-negative organisms, including many blactamase producing strains. The compounds also possess high stability to p-lactamases produced by a range of Gram-negative and Gram-positive organisms.

Compounds according to the invention have beenfoundto exhibit high activity against strains (including penicillinase-producing strains) of Gram-positive bacteria such as Staphylococcus aureus, Straphylococcus epidermidis and Streptocccus species. This is coupled with high activity against various members ofthe Enterobacteriaceae (e.g. strains of Escherichia coli, Kiebsiella pneumoniae, Citrobacter divers us, Enterobacter cloacae, Serratia marcescens, Proteus mirabilis and indole-positive Proteus organisms such as Proteus vulgaris, Proteus morganll and Providence species), strains of Haemophilus influenzae, and Acinetobacter calcoaceticus as well as good activity against Pseudomonas species. This combination of high activity against Gram-positive organisms with high activity against Gram-negative organisms possessed by the compounds of the invention is particularly advantageous.

Non-toxic salt derivatives which may be formed by reaction of the carboxyl group present in the compounds offormula (I) include inorganic base salts such as alkali metal salts (e.g. sodium and potassium salts) and alkaline earth metal salts (e.g. calcium salts); amino acid salts (e.g. lysine and arginine salts); organic base salts (e.g. procaine, phenylethyl-benzylamine, dibenzylethylenediamine, ethanol amine, diethanolamine and N-methylglucosamine salts). Other non-toxic salt derivatives include acid addition salts, e.g. formed with hydrochloric, hydrobromic, sulphuric, nitric, phosphoric, formic and trifluoroacetic acids. The salts may also be in the form of resinates formed with, for example, a polystyrene resin or cross-linked polystyrene divinylbenzene copolymer resin containing amino orquaternary amino groups orsulphonic acid groups, or with a resin containing carboxyl groups, e.g. a polyacrylic acid resin. Soluble base salts (e.g. alkali metal salts such asthesodium salt) ofthe compounds offormula (I) may be used in therapeutic applications because of the rapid distribution of such salts in the body upon administration. Where, however, insoluble salts of compounds (I) are desired in a particular application, e.g. for use in depot preparations, such salts may be formed in conventional manner, for example with appropriate organicamines.

Non-toxic metabolically labile ester derivatives which may be formed by esterification of the carboxyl group in the parent compound offormula (i) include acyloxyalkyl esters, e.g. loweralkanoyloxy-methyl or methyl esters such as acetoxy-methyl or-ethyl or piva loyloxymethyl esters. In addition to the above ester derivatives, the present invention includes within its scope the compounds offormula (I) in the form of other physiologically acceptable equivalents, i.e. physiologically acceptable compounds which, like the metabolically labile esters, are converted in vivo into the parent antibiotic compound offormula (I).

These and other salt and ester derivatives such as the salts with toluene-p-sulphonic and methanesulphonic acids orthe esters with t-butyl or diphenylmethyl esterifying groups may be employed as intermedi ates in the preparation and/or purification ofthe present compounds offormula (l),forexample inthe processes described below.

Preferred compounds according to the invention by virtue oftheir high antibiotic activity arethecompounds wherein Y represents a 1 -methylpyridinium-4-yl, -methyl pyridi n iu m-2-yl,1 1 -methyltetrazol-5-yl or 1,2,5,6- tetrahydro-2-methyl-5,6-dioxo-1 ,2,4-triazin-3-yl group and the non-toxic salts and non-toxic metabolically labile esters thereof. A particularly preferred compound according to the invention is (6R,7R)-7-[(Z)-2-(2 aminothiazol-4-yl)-2-fluoromethoxy-iminoacetamido]-3-(1 -methyl pyridinium-4-yl)thiomethyl]ceph-3-em-4- carboxylate and its non-toxic salts.

The compounds according to the present invention may exist in tautomeric forms (for example in respect of the 2-aminothiazolyl group or with respect to a 3-substituent such as a 2,5-dihydro-6-hydroxyl-2-methyl-5-oxo -1,2,4-triazin-3-yl group) and it will be understood that such tautomericforms are included with in the scope of the invention.

It will be appreciated that the group Y may carry a positive charge, e.g. when Y is a 1-methyl-pyridinium-2-yl group, and when such is the case the positive charge must be balanced by a negative charge. Thusthe compounds may be betaines, so thatthe negative charge is provided by a COOgrnup atthe L-position.

Alternatively, the negative charge may be provided by an anion A6; such an anion will be non-toxic and may be derived from any of the acids described above which will form non-toxic salt derivatives.

The compounds ofthe invention may be used fortreating a variety of diseases caused by pathogenic bacteria in human beings and animals, such as respiratory tract infections, urinary tract infections and skin and soft tissue infections.

According to another embodiment of the invention we provide a process for the preparation of an antibiotic compound of general formula (I) as hereinbefore defined ora non-toxic salt or non-toxic metabolically labile esterthereofwhich comprises forming a compound oftheformula

[wherein Y is as defined above; B is =S or ,SsO (a- or ss-); R1 represents hydrogen or a carboxyl blocking group, e.g. the residue of an ester-forming aliphatic or araliphatic alcohol or an ester-forming phenol, silanol orstannanol (the said alcohol, phenol, silanol orstannanol preferably containing 1 to 20 carbon atoms) ora symmetrical or mixed anhydride blocking group derived from an appropriate acid;R2 is an amino or protected amino group; and the dotted line bridging the 2-,3- and 4-positions indicatesthatthe compound isa ceph-2-em or ceph-3-em compound or a salt, e.g. an acid addition salt (formed with, for example, a mineral acid such as hydrochloric, hydrobromic, sulphuric, nitric or phosphoric acid or an organic acid such as methanesulphonic ortoluene-p-sulphonic acid) by one ofthe following processes:: (A) acylating a compound oftheformula

(wherein Y, B, R1 and the dotted line are as defined above or a 7-N-silyl derivative thereof, oralternatively (where Y contains a quaternary nitrogen atom) a corresponding compound having a group offormula -COOe at the 4-position, with an acid offormula

(wherein R2 is as defined above) or a salt thereof, or with an acylating agent corresponding thereto;; (B) reacting a compound offormula

(wherein R1, R2, B and the dotted line are as hereinbefore defined and Xis a replaceable residue of a nucleophile e.g. an acetoxy or dichloroacetoxy group or a halogen atom such as chlorine, bromine or iodine) ora saltthereof,with a sulphur nucleophile serving to form the group-CH2SY (wherein Yis as defined above) atthe3-position; or (C) where Yin the compounds of the invention contains a C1.4 alkyl-substituted orcarbamoylmethyl- substituted quaternary nitrogen atom in the heterocyclic ring, reacting a compound offormula

(wherein R2, B and the dotted line are as defined above;R1 in this instance is a carboxyl blocking group; andY' represents a carbon-linked 5-or 6-membered heterocyclic ring containing a tertiary nitrogen atom) with a C1.4 alkylating agent a carbamoylmethylating agent serving to introduce a Cm 4 alkyl group ora carbamoylmethyl group as a substituent on the tertiary nitrogen atom in the heterocyclic ring of Y'; whereafter, if necessary and/ordesired in each instance, any ofthe following reactions, in any appropriate sequence, are carried out:- i) conversion of a A2-isomer into the desired A3-isomer, ii) reduction of a compound wherein B is =S-*O to form a compound wherein B is =S.

iii) conversion of a carboxyl group into a non-toxic metabolically labile esterfunction, iv) formation of a non-toxic salt, and v) removal of any carboxyl blocking and/or N-protecting groups.

In the above-described process (A), the starting material offormula (II) is preferably a compound B is ,S and the dotted line represents a ceph-3-em compound.

Compounds offormula (ia) are new compounds and constitute a feature of the invention.

When the group Yin formula (11) is charged e.g. as in an N-alkylpyridinium group, and the compound contains a group offormula -COOR' (wherein R1 is as defined above) atthe4-position,the compound will include an associated anion Ee such as a halide, e.g. chloride or bromide, ortrifluoroacetate anion.

Acylating agents which may be employed in the preparation of compounds of formula (I) include acid halides, particularly acid chlorides or bromides. Such acylating agents may be prepared by reacting an acid (III) or a saltthereofwith a halogenating agent e.g. phosphorus pentachloride, thionyl chloride or oxalyl chloride.

Acylations employing acid halides may be effected in aqueous and non-aqueous reaction media, conveniently attemperatures of from - 50 to +50to, preferably -40 to +30"C, if desired in the presence of an acid binding agent. Suitable reaction media include aqueous ketones such as aqueous acetone, aqueous alcohols such as aqueous ethanol, esters such as ethyl acetate, halogenated hydrocarbons such as methylene chloride, amides such as dimethylacetamide, nitriles such as acetonitrile, or mixtures of two or more such solvents.

Suitable acid binding agents includetertiaryamines (e.g. triethylamine or dimethylaniline), inorganic bases (e.g. calcium carbonate or sodium bicarbonate), and oxiranes such as lower 1,2-alkylene oxide (e.g. ethylene oxide or propylene oxide) which bind hydrogen halide liberated in the acylation reaction.

Acids of formula (III) may themselves be used as acylating agents in the preparation of compounds of formula (I). Acylations employing acids (III) are desirably conducted in the presence of a condensing agent, for example a carbodiimide such as NN'-dicyclohexylcarbodiimide or N-ethyl-N'-dimethylaminopropyl carbodiimide; a carbonyl compound such as carbonyldiimidazole; or an isoxazolium salt such as N-ethyl-5 phenylisoxazolium perchlorate.

Acylation may also be effected with other amide-forming derivatives of acids offormula (III) such as, for example, an activated ester, a symmetrical anhydride or a mixed anhydride (e.g. formed with pivalic acid or with a haloformate, such as a loweralkylhaloformate). Mixed anhydrides may also be formed with phosphor us acids (for example phosphoric or phosphorous acids), sulphuric acid oraliphaticoraromaticsulphonic acids (forexample toluene-p-sulphonic acid). An activated ester may conveniently beformed in situ using,for example, I -hydroxybenzotriazole in the presence of a condensing agent as set out above. Alternatively,the activated ester may be preformed.

Acylation reactions invoiving the free acids ortheir above-mentioned amide-forming derivatives are desirably effected in an anhydrous reaction medium, e.g. methylene chloride, tetrahydrofuran, dimethylfor mamide oracetonitrile.

An alternative method of activation is, for example, by reacting an acid offormula (Ill) with a solution or suspension preformed by adding a carbonyl halide, in particular oxalyl chloride or phosgene, or a phosphoryl halide such as phosphorous oxychlorideto a solvent such as a halogenated hydrocarbon,forexample methylene chloride, containing a lower acyl tertiary amide such as N,N-dimethylformamide. The activated form ofthe acid offormula (ill) may then be reacted with a 7-amino compound offormula (II) in a suitable solventormixture of solvents for example halogenated hydrocarbons e.g. dichloromethane; alcohols, e.g.

ethanol or industrial methylated spirits; ethers, e.g. tetrahydrofu ran or dioxan; esters, e.g. ethyl acetate; ketones, e.g. acetone; am ides, e.g. N,N-dimethylacetamide, acetonitrile; water and mixtures thereof. The acylation reaction may conveniently be effected attemperatures of from - 50 to + 50C, preferably -40" to +30"C, if desired in the presence of an acid binding agent, for example as described above (e.g. dimethylani íine, trimethylamine or sodium bicarbonate).

If desired, the above acylation reactions may be carried out in the presence of a catalyst such as4dimethylaminopyridine.

The acids offormula (III) and acylating agents corresponding thereto may, if desired, be prepared and employed in the form of their acid addition salts. Thus, for example, acid chlorides may conveniently be employed as their hydrochloride salts, and acid bromides as their hydrobromidesalts.

In process (B) above, the sulphur nucleophile may be used to displace a widevarietyofsubstituents Xfrom the cephalosporin offormula (IV). To some extent the facility ofthe displacement is related to the pKa ofthe acid HXfrom which the substituent is derived. Thus, atoms or groups Xderivedfrom strong acids tend, in general, to be more easily displaced than atoms or groups derived from weaker acids. Thefacility ofthe displacement is also related, to some extent, to the precise characterofthe sulphur nucleophile. The latter nucleophile may be employed for example in the form of an appropriatethiol orthione.

The displacement of X by the sulphur nucleophile may conveniently be effected by maintaining the reactants in solution or suspension. The reaction is advantageously effected using from 1 to 10 moles ofthe nucleophile.

Nucleophilic displacement reactions may conveniently be carried out on those compounds offormula (IV) whereinthesubstituentXisa halogen atom oran acyloxygroup,forexample as discussed below.

Acyloxygroups Compounds offormula (IV) wherein Xis an acetoxygroup are convenient starting materialsfor use inthe nucleophilic displacement reaction with the sulphur nucleophile. Alternative starting materials in this class include compounds of formula (IV) in which X isthe residue of a substituted acetic acid e.g. chloroacetic acid, dichloroacetic acid and trifluoroacetic acid.

Displacement reactions on compounds (lV) possessing X substituents ofthis class, particularly in the case whereX is an acetoxy group, may be facilitated by the presence in the reaction medium ofiodideor thiocyanate ions.

The substituent X may also be derived from formic acid, a haloformic acid such as chloroformic acid, or a carbamic acid.

When using a compound offormula (IV) in which X represents an acetoxy orsubstituted acetoxy group, itis generally desirable that the group R1 in formula (IV) should be a hydrogen atom and that B should represent ,'S. In this case, the reaction is advantageously effected in an aqueous medium.

Under aqueous conditions, the pH value ofthe reaction solution is advantageously maintained in the range 6-8, if necessary by the addition of a base. The base is conveniently an alkali metal or alkaline earth metal hydroxide or bicarbonate such as sodium hydroxide or sodium bicarbonate.

When using compounds offormula (lV) in which Xis an acetoxy group, the reaction is conveniently effected at a temperature of 30into 11 O"C, preferably 50" to 80"C.

Halogens Compounds offormula (IV) in which Xis a chlorine, bromine or iodine atom can also be conveniently used as starting materials in the nucleophilicdisplacement reaction with the sulphur nucleophile. When using com- pounds offormula (IV) in this class, B may represent and R1 may represent a carboxyl blocking group. The reaction is conveniently effected in a non-aqueous medium which preferably comprises one or more organic solvents, advantageously of a polar nature such as ethers, e.g. dioxan ortetrahydrofuran, esters, e.g. ethyl acetate, amides e.g. formamide and N,N-dimethylformamide, and ketones e.g. acetone. Other suitable organic solvents are described in more detail in British Patent Specification No. 1,326,531.The reaction medium should be neither extremely acidic nor extremely basic.

In the case of reactions carried out on compounds offormula (lV) in which R1 is a carboxyl blocking group and the resulting Y group contains a quaternary nitrogen atom,the product will be formed as the corresponding halide saltwhich may, if desired, be subjected to one or more ion exchange reactions to obtain a salt having the desired anion.

When using compounds of formula (IV) in which Xis a halogen atom as described above, the reaction is conveniently effected at a temperature of -20 to +60 , preferably 0" to +30 C.

When the incoming nucleophile does notyield a compound containing a quaternised nitrogen atom,the reaction is generally effected in the presence of an acid scavenging agentforexample a base such as triethylamineorcalcium carbonate.

In process (C) above, the compound offormula (V) is advantageously reacted with a compound of the formula R3Zwherein R3 is a C1-4 alkyl group or a group of formula H2NCOCH2-, and Z is a leaving group such as a halogen atom (e.g. iodine, chlorine or bromine) or a hydrocarbylsulphonate (e.g. mesylate ortosylate) group. Alternatively, a di-C,.4-alkyl sulphate, e.g. dimethyl sulphate, may be employed as an alkylating agent.

lodomethane is preferred as the alkylating agent and iodoacetamide is preferred as the carbamoylmethylating agent. The reaction is preferably carried out at a temperature in the range of Oto 60"C, advantageously 20to 30"C. Where the alkylating agent is liquid underthe reaction conditions, as in the case of iodomethane, this agent can itself serve as a solvent. Alternatively, the reaction may be conveniently effected in an inertsolvent such as an ether e.g. tetrahydrofuran, an amide, e.g. dimethylformamide, a lower alkanol, e.g. ethanol, a lower dialkylketone, e.g. acetone, a halogenated hydrocarbon e.g. dichloromethane or an ester e.g. ethyl acetate.

The compound offormula (V) used as starting material in process (C) may be prepared for example by reaction of a compound offormula (IV) (as defined above) with an appropriate sulphur nucleophile in an analogous mannertothe nucleophilic displacement reaction described with respect two process (B). If desired the above nucleophile may be used in the form of a metal thiolate salt.

When X in formula (IV) is halogen, the reaction is preferably effected in the presence of an acid scavenging agent, for example a base such astriethylamine or calcium carbonate.

The reaction product may be separated from the reaction mixture, which may contain, for example, unchanged cephalosporin starting material and other substances, by a variety of processes including recrystallisation, ionophoresis, column chromatography and use of ion-exchangers (for example by chromatogra pry on ion-exchange resins) or macroreticular resins.

A A2-cephalosporin ester derivative obtained in accordance with the process ofthe invention may be converted into the corresponding desired A3-derivative by, for example, treatment of the A2-esterwith a base, such as pyridine ortriethylamine.

Where a compound is obtained in which B is SOthis may be converted into the corresponding sulphide by,forexample, reduction ofthe corresponding acyloxysulphonium oralkoxysuiphonium salt prepared in situ by reaction with e.g. acetyl chloride in the case of an acetoxysulphonium salt, reduction being effected by, for example, sodium dithionite or by iodide ion as in a solution of potassium iodide in a suitable solvent e.g. acetic acid, acetone, tetrahydrofuran, dioxan, dimethylformamide or dimethylacetamide. The reaction may be effected at a tem peratu re of from -20" to + 50on.

Metabolically labile ester derivatives ofthe compounds of formula (I) may be prepared by reacting a compound offormula (I) or a sa It or protected derivative thereof with the appropriate esterifying agent such as an acyloxyalkyl halide or alkoxycarbonyloxy alkyl halide (e.g. iodide) conveniently in an inert organic solvent such as dimethylformamide or acetone, followed, where necessary, by removal of any protecting groups.

Base salts ofthe compounds offormula (I) may be formed by reacting an acid offormula (I) with an appropriate base. Thus, for example, sodium or potassium salts may be prepared using the respective2-ethylhexanoate, acetate or hydrogen carbonate salt. Acid addition salts may be prepared by reacting a compound of formula (I) or a metabolically labile ester derivative thereof with the appropriate acid.

Where a compound offormula (I) is obtained as a mixture of isomers, thesyn isomer may be obtained by, for example, conventional methods such as crystallisation or chromatography.

For use as starting materialsforthe preparation of compounds of general formula (I) according tothe invention, compounds of general formula (Ill) and the amideforming derivatives thereof such as acid halides and an hydrides corresponding thereto in theirsyn isomericform or in the form of mixtures of the syn isomers and the corresponding antiisomers containing at least 90% of thesyn isomer are preferably used.

Acids offormula (Ill) and their derivatives are themselves novel compounds and form a furtherfeature of the invention. They may be prepared by etherification of a compound offormula

(wherein R2 is as hereinbefore defined and R4 represents hydrogen ora carboxyl blocking group) orasalt thereof, by selective reaction with a compound of general formula T.CH2- F (Vlil) (wherein Tis halogen, such as chloro, bromo or iodo; sulphate; or sulphonate, such astosylate), followed by removal of any carboxyl blocking group R4. Separation of isomers may be effected either before or aftersuch etherification.The etherification reaction is conveniently carried out in the presence of a base, e.g. potassium carbonate or sodium hydride, and is preferably conducted in an organic solvent, for example dimethylsuiphoxide, a cyclic ether such astetrahydrofuran ordioxan, or an N,N-disubstituted amide such asdimethylforma- mide. Under these conditions the configuration ofthe oxyimino group is substantially unchanged bythe etherification reaction. The reaction should be effected in the presence of a base if an acid addition salt of a compound of formula (VII) is used. The base should be used in sufficient quantity to neutralise rapidly the acid in question.

Acids offormula (III) may also be prepared by reaction of a compound offormula

(wherein R2 and R4 are as hereinbefore defined) with a compound offormula H2N.O.CH2-F (X) followed by removal ofanycarboxyl blocking group R4, and where necessary the separation of sun and anti isomers.

The acids offormula (Ill) may be converted into the corresponding acid halides and anhydrides and acid addition salts by conventional methods, for example as described hereinabove.

Where Xis a halogen (e.g. chlorine, bromine or iodine) atom in formula (IV), ceph-3-em starting compounds may be prepared in conventional manner, e.g. by halogenation of a 7ss-protected amino-3methylceph-3-em-4-carboxylic acid ester 1 p-oxide, removal of the 7amino compound to form the desired 7P-acylamido group, e.g. in an analogous mannerto process (A) above, followed by reduction of the 1 p-oxide group later in the sequence. This is described in British Patent No. 1,326,531.The corresponding ceph-2-em compounds may be prepared by the method of Dutch published Patent Application No.6,902,013 by reaction of a 3-methylceph-2-em compound with N-bromosuccinimide to yield the corresponding 3bromomethylceph-2-em-compound.

Where X in formula (IV) is an acetoxy group, such starting materials may be prepared for example by acylation of 7-aminocephalosporanic acid, e.g. in an analogous mannerto process (A) above. Compounds of formula (IV) in which X represents other acyloxy groups can be prepared by acylation ofthe corresponding 3-hydroxymethyl compounds which may be prepared for example by hydrolysis of the appropriate 3 acetoxymethyl compounds, e.g. as described for example in British Patent Specifications Nos. 1,474,519 and 1,531,212.

Compounds offormula (II) may also be prepared in conventional manner, e.g. by nucleophilicdisplacement of a corresponding 3-acyloxymethyl or3-halomethyl compound with the appropriate nucleophile, e.g.

as described in British Patents Nos. 1,012,943 and 1,241,657.

Afurther method forthe preparation ofthe starting materials of formula (II) comprises deprotecting a corresponding protected 7(3-amino compound,forexample in conventional manner, e.g. using PC15.

It should be appreciated that in some ofthe above transformations it may be necessary to protectany sensitive groups in the molecule ofthe compound in question to avoid undesirable side reactions. Examples ofsuitable protecting groups are given in "Protective Groups in Organic Synthesis" by Theodora W. Greene (John Wiley and Sons, 1981) and "Protective Groups in Organic Chemistry", Ed. J.F.W.McOmie (Plenum Press, 1973). For example, during any of the reaction sequences referred to above it may be necessaryto protectthe NH2 group ofthe aminothiazolyl moiety, for example bytritylation, acylation (e.g. chloroacetylation orformylation), protonation or other conventional method.The protecting group may thereafter be removed in any convenient way which does not cause breakdown ofthe desired compound, e.g. in the case of a trityl group by using an optionally halogenated carboxylic acid, e.g. acetic acid, formic acid, chloroacetic acid ortrifluoroacetic acid or using a mineral acid, e.g. hydrochloric acid or mixtures of such acids, preferably in the presence of a protec solvent such as water, or, in the case of a chloroacetyl group, by treatmentwith thiourea.

Carboxyl blocking groups used in the preparation of compounds offormula (I) or in the preparation of necessary starting materials are desirably groups which may readily be split off at a suitable stage inthe reaction sequence, conveniently at the last stage. It may, however, be convenient in some instances to employ non-toxic metabolically labile carboxyl blocking groups such as acyloxy-methyl or-ethyl groups (e.g.

acetoxy-methyl or methyl or pivaloyloxymethyl) and retain these in the final product to give an appropriate ester derivative of a compound offormula (I).

Suitable carboxyl blocking groups are well known in the art, a list of representative blocked carboxyl groups being included in British Patent No. 1399086. Preferred blocked carboxyl groups include aryl lower alkoxycarbonyl groups such asp-methoxybenzyloxycarbonyl, p-nitrobenzyloxycarbonyl and diph enylmethoxycarbonyl; lower alkoxycarbonyl groups such as t-butoxy carbonyl; and lower halo- alkoxycarbonyl groups such as 2,2,2-trichloroethoxycarbonyl. The carboxyl blocking group may subsequently be removed by any of the appropriate methods disclosed in the literature; thus, for example, acid catalysed hydrolysis or reduction is applicable in many cases, as are enzymically-catalysed hydrolyses.

The antibiotic compounds ofthe invention may be formulated for administration in any convenient way, by analogy with other antibiotics and the invention therefore includes within its scope pharmaceutical compositions comprising an antibiotic compound in accordance with the invention adapted for use in human or veterinary medicine. Such compositions may be presented for use in conventional manner with the aid of any necessary pharmaceutical carriers orexcipients.

The antibiotic compounds according to the invention may, for example, be formulated for injection and may be presented in unit dose form, in ampoules, or in multi-dose containers, if necessary with an added preservative. The compositions may also take such forms as suspensions, solutions, or emulsions in oily or aqueousvehicles, and may contain formulatory agents such as suspending, stabilising and/or dispersing agents. Alternatively the active ingredient may be in power forum for reconstitution with a suitable vehicle, e.g.

sterile, pyrogem-free water, before use.

If desired, such powder formulations may contain an appropriate non-toxic base in order to improve the water-solubility ofthe active ingredient and/or to ensure that when the powder is reconstituted with water, the pH of the resulting aqueous formulation is physiologically acceptable. Alternatively the base may be present in the waterwith which the powder is reconstituted. The base may be, for example, an inorganic base such as sodium carbonate, sodium bicarbonate orsodium acetate, or an organic base such as lysine or lysine acetate. The antibiotic compounds may also, be formulated as suppositories e.g. containing conventional suppository bases such as cocoa butter or other glycerides.

Compositions forveterinary medicine may, for example, be formulated as intramammary preparations in either long acting or quick-release bases.

The compositions may contain from 0.1 % upwards, e.g. 0.1-99% ofthe active material, depending on the method of administration. When the compositions comprise dosage units, each unit will preferably contain 100-3000 mg ofthe active ingredient e.g. 200-2000 mg. The daily dosage for adult human treatmentwill preferably range from 200 to 12000 mg e.g. 1000-9000 mg per day, depending interalia on the nature ofthe infection and the route and frequency of administration. Forexample, intravenous or intramuscularadministration may be employed, for example using 400 to 4000 mg per day ofthe active ingredient in adult human treatment. In treating Pseudomonas infections higher daily doses may be required.It will be appreciated that in some circumstances, for example, in the treatment of neonates, smaller dosage units and daily dosages may be desirable.

The antibiotic compounds according to the invention may be administered in combination with other therapeutic agents such as antibiotics, for example penicillins or other cephalosporins.

The following Examples illustrates the invention. All temperatures are in "C; DMSO is dimethylsulphoxide; EtOH is ethanol; THF istetrahydrofuran, DMF is dimethylformamide. Sorbsil U30 is silica gel manufactured by Joseph Crosfield and Son ofWarrington, Cheshire, England (Sorbsil is a registered Trade Mark). Kiesel 60 is silica gel manufactured by E. Merck & Co. of Darmstadt, West Germany. (Kieselgel is a registered Trade Mark).

Intermediate 1 Ethyl (Z)-2-fluoromethoxyimino-2-(2-triphenylmethylaminothiazol-4-yl)acetate Ethyl (Z)-2-hydroxyimino-2-(2-triphenylmethylaminothiazol-4.yI)acetate, hydrochloride salt (8.7g) was stirred with potassium carbonate (15.35 g) in dimethyl sulphoxide (30ml) under nitrogen at 21". Bromofluo romethane (ca3gwas added. The nitrogen flow was stopped and the stirring continuedfortwo hours. The mixture was poured into an ice-watermixturewith stirring and the solid was collected by filtration and washed with water. The solid was dissolved in methylene chloride and the organic layer was separated and dried with magnesium sulphate. Evaporation gave a foam. This was dissolved in methylene chloride and pre-absorbed onto Kieselgel 60 (50g). This was added to thetop of a column of similarsilica (125g) set up in 10% ethyl acetate in cyclohexane. The column was eluted successively with 10%, 20% and 33% ethyl acetate in cyclohexane.After combination of appropriate fractions, evaporation gavethe titlecompound(8.06g) as a foam; AmaX (EtOH) 302nm (E1%m 92),#infl include 227.5nm (E1cMom 546) and 259nm (E1cm 221), vmaX (CHBr3) 3400 (NH), 1739 (ester) and 1 533cm- (C=N).

Intermediate 2 (Z)-2-Fluoromethoxyimino-2-(2-triphenylmethylaminothiazol-4-yl)acetic acid Intermediate 1 (7.8g) was stirred under refluxwith sodium hydroxide (0.83g) in ethanol (50ml) and water (10ml)for 15 minutes. The mixture was cooled and the crystalline precipitate was collected by filtration and washed with ethanol and ether and dried. This solid was partitioned between methylene chloride (80ml) and water (40ml) with vigorous stirring and 88% orthophosphoric acid (2ml) was added. Solid remained and this was collected by filtration. This solid was suspended in tetrahydrofuran (75ml) and 2M hydrochloric acid (8ml) was added when a solution formed.Evaporation reduced the volume of solution by one half and methylene chloride (50ml) was added. The aqueous layerwas extracted with more methylene chloride and the combined organic layers were washed with water, dried with magnesium sulphate and evaporated to a solid, the title compound(4.82g); X;fi include 224nm (E1%1cm 564), 254.5nm (E1%1cm 213) and 260nm (E1%1cm 205); T (d6DMSO) 1.02(s; NH), 2.64(s; phenyl protons) 2.91 (s; thiazole 5-H), and 4.29 (d, J 56Hz; CH2F).

Intermediate 3 Diphenylmeth yl(6R, 7R)-3-Bromomethyl-7-[(Z)-2-fluoromethoxyimino-2-(2-triphenylmethylaminothiazol-4- yl)acetamido]ceph-3-em-4-carboxylate Oxalyl chloride (0.3 ml) was added to a solution of DMF (0.4 ml) in methylene chloride (10 ml)with stirring under nitrogen at - 20 and the mixture was then stirred with ice-water cooling for ten minutes before recooling to -20^. (Z)-2-Fluoromethyimino-2-(triphenylmethylaminothiazol-4-yl)acetic acid (1.5 g) was added and the solution was stirred with ice-water cooling before again cooling to -20 .A suspension ofdiphenylmethyl (6R,7R)-7-amino-3-bromomethylceph-3-em-4-carboxylate, hydrochloride salt (1.5 g) in methylene chloride (10 ml) containing dimethylaniline (2ml) was added and the solution was allowed to warm to 21 over30 minutes. The solution was washed successively with dilute hydrochloric acid and water (twice), each time back extracting with methylene chloride. The combined organic layers were dried with magnesium sulphate, concentrated by evaporation and applied to a column ofsorbsil U30 (50 g) set up in 10% ethyl acetate in petroleum ether (b.p. 40 to 60 ).The column was eluted with this mixture and then with 20%, 30%andfinally 40% ethyl acetate in petroleum ether (b.p. 40 to 60 ). Combination ofthe appropriate eluate and evaporation gave the title compound (1.52 g), [a] - 6.92" (c 0.65, chloroform,#max(bromoform) 3385 (N H), 1789 (ss-lactam), 1725 (ester) and 1688 and 1514cm-1 (amide).

Example 1 (a) Diphenylmethyl (6R, 7R)-7-f(Z)-2-fluoromethoxyimino-2-(2-triphenylmethylaminothiazol-4-yl)acetamidoj- 3-(1-methyl- lH-tetrazol-5-ylthiomethyl)ceph-3-em-4carboxylate.

Oxalyl chloride (0.15 ml) was added to a stirred solution of DMF (0.2 ml) in methylene chloride (10 ml) under nitrogen at -20' and the mixture was then stirred for ten minutes with ice-water cooling before recooling to -20 . (Z)-2-Fluoromethoxyimino-2-(2-triphenylmethylaminothiazol-4-yl)acetic acid (750 mg) was added and the solution was stirred with ice-water cooling forten minutes before recooling to -20 . A mixture of diphenylmethyl (6R,7R)-7-amino-3-(1-methyl-1 H-tetrazol-5-ylthiomethyl) ceph-3-em-4-carboxylate (750 mg) and N,N-dimethylaniline (0.5 ml) in methylene chloride (10 ml) was added and the resultant solution was allowed to stir and warm to 21 over 30 minutes. T.l.c. showed complete reaction.The solution was diluted with methylene chloride and washed with dilute hydrochloric acid and water (back extracting with methylene chloride each time). The combined organic layers were dried with magnesium sulphate and concentrated before applying to a column of Sorbsil U30 (50 g) set up in 10% ethyl acetate in petroleum ether (b.p. 40-60 ) and the column was eluted with 10 to 60% ethyl acetate in petroleum ether (b.p.40-60 ). Combination of appropriate fractions and evaporation gave the title compound ( 1.32 g) [a]2D2- 129.5 (c 0.74, chloroform).

i'max (CHBr3) 3385 (NH), 1788 (ss-lactam), 1720 (ester) and 1689 and 1513 cm-1 (amide).

(b) (6R, 7R)-7-[(Z)-2-Aminothiazol-4yl)-2-fluoromethoxysminoacetamidoi-3-r1-methyl- lH-tetrazol-5- ylthiomethyl)- ceph-3-em-4-carboxylic acid The above ester(1.22g) was dissolved in anisole (3 ml) with stirring at 21' and trifluoroacetic acid (12 ml) was added. After one warat water (1.5 ml) was added. After a furtherthree minutes, the solutionwas poured into diisopropyl ether (300 ml) and the precipitate was collected byfiltration,washed with diisopropyl ether and dried in vacuo at 50 for one hourto give the title compound (630 mg), [a]2D2 64.0 (c 0.55 DMSO), Xmax (pH6 buffer) 225.5 nm (E1cm 354), 256.5 nm (ECrvno 262) Ajnfl 303 nm (E1 cm 102).

Example 2 (a) (6R, 7R)-7-f(Z)-2-(2- Triphenylmethylaminothiazol-4-yl)-2-fluoromethoxyiminoacetamidoj-3-(l,2, 5,6- tetrahydro-2-methyl-5,6-dioxo- 42, 4-triazin-3-ylthiom eth yl)ceph-3-em-4-carb oxylic acid.

Oxalyl chloride (0.15 ml)was added to a solution of DMF(0.2 ml) in methylene chloride (10 ml) at -20"with stirring under nitrogen and the mixture was stirred with ice-water cooling for ten minutes before recooling to -20". (Z)-2-Fluoromethoxyimino-2-(2-triphenylmethylaminothiazol-4-yl)acetic acid (750 mg) was added the solution was stirred with ice water cooling for ten minutes before recooling to -20". This solution was added rapidly to a solution of (6R,7R)-7-amino-3-(1 ,2,5,6-tetrahydro-2-methyl-5,6-dioxo-l ,2,4-triazin-3- ylthiomethyl)-ceph-3-em-4-carboxylic acid (500 mg) in industrial methylated spirits (6 ml) and water (1.5 ml) containing triethylamine (1.2 ml) with vigorous stirring at -10".The solution was allowed to warm to 21"over 45 minutes. The solution was partitioned between water and dilute hydrochloric acid. The aqueous layerwas extracted with more methylene chloride and the combined organic layers were washed with water, dried with sodium sulphate and evaporated to a gum. This gum was triturated with diethyl etherto give a solid. This solid was dissolved in methylene chloride and loaded onto a column of Sobsil U30 (20 g) set up in 1:1 ethyl acetate-petroleum ether(b.p. 40 to 60 ) and the column was eluted withtthesame mixture followed by neat ethyl acetate.Combination of appropriate fractions followed by evaporation gave the title compound(600 mg), [2D2644 (c 0.57, chloroform), "max (CHBr3) 3385 (NH), 1789 ((3-lactam), 1730 (acid and keto groups, and 1687 and 1523 cm-1 (amide).

(b) {6R,7R)-7-[rZ)-2-62-Amino-4-thiazolyl)-2-fluoromethoxysminoacetamidol-3-{1,2,5,6-tetrabydro-2-methyl- 5, 6-dioxo- l,2,4-triazin-3-ylthiomethyl ceph-3-em-4-carboxylic acid) The above cephem (510 mg) was dissolved in formic acid (3.5 ml) with stirring at 21" and water(1 .5 ml)was added. After 1.25 hr, the mixture was filtered and the filter-cake was leached with 70% formic acid. The combined filtrates were evaporated to a gum which was triturated with diethyl ether and the solid was collected by filtration.After washing with diethyl ehter, the solid was dried in vacuo at 50" for one hou rto give the title compound (310 mg) [01]2,2-70.9" (C 0.76, DMSO) hi,f (pH6 buffer) 252 nm (Ei1cm 263), 305 nm (E1ic m95).

Example 3 (a) Diphenylmethyl (6R, 7R)-7-f(Z)-2-fluoromethoxyimino-2-(2-triphenylmethylaminothiazol -4-yl)acetamidoj-3-f( 1-meth yIp yridinium-4-yl)thiometh yljceph-3-em -4-carb oxyla te Intermediate 3(1.41 g) was stirred with 1 -methylpyrid-4-thione (220 mg) in tetrahydrofuran (20 ml) forthree hours. Further-1-methylpyrid-4- thione (30 mg) was added and after an additional two hours, diethyl ether 80 ml) was added.The precipitate was collected by filtration, washed with ether and dried to give the title compound(1 .34g) max (EtOH) 303.5 nm (Ei1cm 167), 345.5 nm (Ei1cm 146), . 230.5 nm (Ei1cm 397), 264.5 n, (E1cm 189), 271 nm (E1cm 174), vmaX (Nujol) 3700-2300 (NH), 1782 (ss-lactam), 1722 (ester) and 1679 and 1528 cm-1 (amide).

(b) (SR, 7R)-7-f(Z)-2-(2-A minothiazol-4-yl)-2-fluoromethoxyiminoacetamidoj-3-l(i-methyl- p yridinium-4-yl)thiom eth yljceph-3-em -4-carboxylate The above protected cephalosporin (1.26 g) was dissolved in anisole (3 ml) and trifluoroacetic acid (12 ml) was added. After 1.5 hours at 21 , water 1.5 ml) was added. Aftera further minute,thesolution was diluted with diisopropyl ether (ca 250 ml) and the precipitate was collected by filtration. This was washed with diisopropyl ether and dried to give a solid (660 mg).A portion of this solid (300 mg) was purified by preparative HPLCto give the title compound (56 mg), vmaX (Nujol) 3300 (N H and water), 1768 (ss-lactam) and 1670 and 1540 cm-1 (amide),#(dsDMSO), (0.31 (d, J 7Hz; NH), 1.38 and 1.68 (2d, J 7Hz; pyridinium proton), 3.08 (s; thiazole proton), 4.28 (d, J 57Hz; CH2F), 4.42 (d,d, J 7,4Hz; 7-H), 4.97 (d, J 4Hz; 6-H), 5.84(s; NCHs), 5.38 and 5.58 (ABq, J 13Hz; 3CH2), and 6.47 and 6.65 (ABq, J 18Hz; 2-H).

Example 4 (a) Diphenylmethyl (6R, 7R)-2-[{Z)-(2-fluoromethoxyimino-2-(2-triphenylmethyl-aminothiazol-4- yl)acetamidol-3-[r1-methyl-pyridinium-2-yl)thiomethyllceph-3-em-4-carboxylate Intermediate3 (1.0 g) was stirred with 1-methylpyrid-2-thione (170 mg) in THF (20 ml) at21'forsix hours. The solution was diluted to 100 ml by addition of diethyl ether and the precipitate was collected byfiltration, washed with diethyl ether and dried to give the title compound (550 mg) Ajnf(EtOH) 234 nm (E1C m 280),262.5 nm (E1cm 184),#max268.5 nm (E1cm 186), 274 nm (E1cm 187) and 303.5 nm (E110,'m 82),viax (Nujol) 1780 ((3-lactam), 1722 (ester) and 1675 and 1525cm-1 (amide).

(b) (6R, 7R)-7-f(Z)-2-(2-A minothiazol-4-yl)-2-fluoromethoxyiminoacetamidoj-3-[(i-meth ylpyridinium-2- yl)thiomethyllceph-3-em-4carboxylate, trifluoroacetate salt The above protected cephalosporin (750 mg) was mixed with anisole (1.5 ml) andtrifluoroacetic acid (6 ml) was added. Aftertwo minutes, water (0.75 ml) was added and after a furtherten seconds, the solution was diluted with diisopropyl ether (70 ml). the precipitate was collected by filtration and washed with diisopropyl ether and dried to give the tftle compound (470 mg) (a) D21 -13.3 , (c 1.02, DMSO),vmax(Nujol) 3200 (NH), 1782 ((3-lactam), 1710 (acid), 1672 and 1540 (amide) and 1672 cm1 (trifluoroacetate).

Claims (5)

1. Cephalosporin antibiotics of general formula (I)

(wherein Y represents a carbon-linked 5- or6-membered unsaturated heterocyclic ring containing at least on nitrogen atom, which ring may also contain one or more sulphur atoms and/or may be substituted by one or more C1 4 alkyl, oxo, hydroxy or carbamoylmethyl groups) and non-toxic salts and non-toxic metabolically labile esters thereof.

2. Compounds of general formula (I) as claimed in claim 1 wherein Y represents a 1-methylpyridinium-4-yl, 1 -methyl pyridini u m-2-yl,1 1 -methyltetrazol-5-yl or 1 ,2,5,6-tetrahydro-2-methyl-5,6-dioxo-1 ,2,4-triazin-3-yl group and the non-toxic salts and non-toxic metabolically labile esters thereof.

3. (6R,7R)-7-[(Z)-2-(2-Aminothiazol-4-yl)-2-fluoromethoxyiminoacetamido]-3-[(1-methylpyridinium-4-yl)- thiomethyl]ceph-3-em-4-carboxylate and its non-toxic salts.

4. A process for the preparation of a compound of general formula (I) as defined in Claim 1 ora non-toxic salt or non-toxic metabolically labile esterthereofwhich comprises forming a compound of general formula (la)

[wherein Y is as defined above; B is-S-or -SO- (a- or(3-); R1 represents hydrogen ora carboxyl blocking group;R2 is an amino or protected amino group; and the dotted line bridging the 2-, and 4-positions indicatesthatthe compound is a ceph-2-em or ceph-3-em compound] ora saltthereof, by one ofthefollowing processes: (A) acylating a compound of general formula (II)

(wherein Y, B, R1 and the dotted line are as defined above or a 7-N-silyl derivative thereof, or alternatively (where Ycontains a quaternary nitrogen atom) a corresponding compound having a group offormula COOe at the 4-position, with an acid offormula (III)

(wherein R2 is as defined above) or a salt thereof, or with an acylating agent corresponding thereto;; (B) reacting a compound offormula (IV)

(wherein R1, R2, B and the dotted line are as defined above and Xis a replaceable residue of a nucleophile) ora saltthereof,with a sulphur nucleophile serving to form the group -CH2SY (wherein Y is as defined above) at the 3-position; or (C) where Yin the compounds of the invention contains a C1 4 alkyl-substituted orcarbamoylmethylsubstituted quaternary nitrogen atom in the heterocyclic ring, reacting a compound of formula (V)

(wherein R2, B and the dotted line are as defined above;R1 in this instance is a carboxyl blocking group; and Y' represents a carbon-linked 5-or 6-membered heterocyclic ring containing a tertiary nitrogen atom) and a C14 alkylating agentora carbamoylmethylating agent serving to introduce a C14alkyl group ora carbamoylmethyl group as a substituent on the tertiary nitrogen atom in the heterocyclic ring of Y'; whereafter, if necessary and/or desired in each instance, any ofthe following reactions, in any appropriate sequence, are carried out: i) conversion of a A2-isomer into the desired A3-isomer.

ii) reduction of compound where B is -SO- to form a compound wherein B is -S-, iii) conversion of a carboxyl group into a non-toxic metabolically labile esterfunction, iv)formation of a non-toxic salt, and v) removal of any carboxyl blocking and/or N-protecting groups.

5. Pharmaceutical compositions comprising as active ingredient one or more compounds of general formula (i) as defined in claim 1.