GB2027692A - Cephalosporin Antibiotics - Google Patents

Abstract

A Cephalosporin antibiotic of the general formula <IMAGE> exhibits broad-spectrum antibiotic activity, the activity being unusually high against gram-negative organisms such as strains of Pseudomonas organisms. The invention also includes the non-toxic salts and non-toxic metabolically labile esters of the compound of formula (I). Also described are compositions containing the antibiotics of the invention and processes for the preparation of such antibiotics.

Description

SPECIFICATION Cephalosporin Antibiotics This invention is concerned with cephalosporin compounds possessing valuable antibiotic properties.

The cephalosporin compounds in this specification are named with reference to "cepham" after J. Amer. Chem. Soc., 1962, 84, 3400, the term "cephem" referring to the basic cepham structure with 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 resistant to 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 of various types of broad spectrum cephalosporin antibiotics.

Thus, for example, in our British Patent Specification No. 1,399,086, we describe a novel class of cephalosporin antibiotics containing a 7/3-(cx-etherified oximino)-acylamido group, the oximino group having the syn 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 stability to p- lactamases produced by various gram-negative organisms.

The discovery of this class of compounds has stimuiated further research in the same area in attempts to find compounds which have improved properties, for example, against particular classes of organisms especially gramnegative organisms.

For example, in our British Patent Specification No. 1,496,757, we described cephalosporin antibiotics containing a 7P-acylamido group of the formula

(wherein R is a thienyl or furyl group; RA and RB may vary widely, and may, for example, be C14 alkyl groups or together with the carbon atom to which they are attached form a C7 cycloalkylidene group, and m and n are each 0 or 1 suchthatthesumofm and n is O or 1), the compounds being syn isomers or mixtures of syn and anti isomers containing at least 90% of the syn isomer. The 3-position of the cephalosporin molecule may be unsubstituted or may contain one of a wide variety of possible substituents.

These compounds have been found to have particularly good activity against gram-negative organisms.

Other compounds of similar structure have been developed from these compounds in further attempts to find antibiotics having improved broad spectrum antibiotic activity and/or high activity against gram-negative organisms. Such developments have involved variations in not only the 7p-acylamido group in the above formula but also the introduction of particular groups in the 3position of the cephalosporin molecule.

Thus, for example, in Belgian Patent Specification No. 852,427, there are described cephalosporin antibiotic compounds falling within the general scope of our British Patent Specification No. 1,399,086, and wherein the group R in the above formula may be replaced by a variety of different organic groups, including 2aminothiazol-4-yl, and the oxygen atom in the oxyimino group is attached to an aliphatic hydrocarbon group which may itself be substituted by, for example, carboxy. In such compounds, the substituent at the 3-position is an acyloxyrnethyl (for example, acetoxymethyl), -hydroxymethyl, formyl or optionally substituted heterocyclic-thiomethyl group.

Furthermore, Belgian Patent Specification No.

836,813 describes cephalosporin compounds wherein the group R in the above formula may be replaced by, for example, 2-aminothiazol-4-yl, and the oxyimino group is a hydroxyimino or blocked hydroxyimino group, e.g. a methoxyimino group. in such compounds, the 3-position of the cephalosporin molecule is substituted by a methyl group which may itself be optionally substituted by any of a large number of residues of nucleophilic compounds therein described, an example of such a substituted methyl group being given as an acetoxymethyl group. In the abovementioned specification no antibiotic acitvity is ascribed to such compounds which are only mentioned as intermediates for the preparation of antibiotics described in that specification.

Belgian Patent Specification No. 853,545 describes cephalosporin antibiotics wherein the 7p-acylamido side chain is primarily limited to a 2-(2-aminothiazol-4-yl)-2-(syn)-methoxyiminoacetamido group and the substituent in the 3position is broadly defined in a similar manner to that in the above-mentioned Belgian Patent Specification No. 836,813. Compounds specifically exemplified in the specification include compounds in which the 3-position is substituted by an acetoxymethyl group.

We have now discovered that by an appropriate selection of a particular group at the 7p-position in combination with an acetoxymethyl group at the 3-position, cephalosporin compounds having advantageous activity against a wide range of commonly encountered pathogenic organisms may be obtained. This advantageous antibiotic activity is described in more detail below.

The present invention provides (6R, 7R)-7-[(Z)2-(2-aminothiazol-4-yl)-2-( 1 -carboxycyclobut- 1oxyimino)aceta mido]-3-acetoxymethyl-ceph-3em-4-carboxylic acid of the formula

and non-toxic salts and non-toxic metabolically labile esters thereof.

The compounds according to the invention are syn isomers. The syn isomeric form is defined by the configuration of the group

with respect to the carboxamido group. In this specification the syn configuration is denoted structurally as

It will be appreciated that since the compounds of the invention are geometric isomers, some admixture with the corresponding anti isomer may occur.

The compounds according to the present invention may exist in tautomeric forms (in respect of the 2-amino-thiazolyl group) and it will be understood that such tautomeric forms, e.g.

the 2-iminothiazolinyl form, are included within the scope of the invention.

The present invention also includes within its scope the solvates (especially the hydrates) of the compound of formula (I). It also includes within its scope salts of esters of the compound of formula (I).

The compounds according to the invention exhibit good broad spectrum antibiotic activity.

Against gram-negative organisms the activity is unusually high. This high activity extends to many p-lactamase-producing gram-negative strains.

The compounds also possess high stability to ,B- lactamases produced by a range of gram-negative organisms.

In addition to their above utility as antibiotics the above-defined compound are also useful as intermediates by virtue of the ease with which the acetoxy radical in the 3-position can be displaced by a variety of nucleophiles to yield antibiotics containing a 3-methyl group substituted by different nucleophilic residues, for example, pyridinium or 1 -methylpyridiniumylthio residues.

Compounds according to the invention have been found to exhibit unusually high activity against strains of Pseudomonas, e.g. strains of Pseudomonas aeroginosa, as well as high activity against various members of the Enterobacteriaceae (e.g. strains of Escherichia coli, Klbesiella pneumoniae, Salmonella typhimurium, Enterobacter cloacae, Serratia marcescens Providence species, Proteus mirabilis and, especially, indole-positive Proteus organisms such as Proteus vulgaris and Proteus morganil3, as well as strains of Haemophilus influenzae.

Non-toxic salt derivatives which may be formed by reaction of either or both of the carboxyl groups present in the compound of formula (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, ethanolamine, 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 or quaternary amino groups or sulphonic acid groups or with a resin containing carboxyl groups, e.g. a polyacrylic acid resin. Soluble base salts (e.g. alkali metal salts such as the sodium salt) of the compound of formula (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 compound (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 organic amines.

These and other salt derivatives such as the salts with toluene-p-sulphonic and methanesulphonic acids may be employed as intermediates in the preparation and/or purification of the present compound of formula (I), for example in the process described below.

Non-toxic metabolically labile ester derivatives which may be formed by esterification of either or both carboxyl groups in the parent compound of formula (I) include acyloxyalkyl esters, e.g. lower alkanoyloxy-methyl or-ethyl esters such as acetoxymethyl, acetoxyethyl or pivaloyloxymethyl esters. In addition to the above ester derivatives, the present invention includes within its scope the compound of formula (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 of formula (I).

The compound of formula (I) may be used for treating a variety of diseases caused by pathogenic bacteria in human beings and animals such as respiratory tract infections and urinary tract infections.

According to another embodiment of the invention we provide a process for the preparation of the compound of formula (I) as hereinbefore defined or a non-toxic salt or non-toxic metabolically labile ester thereof which comprises acylating a compound of the formula

(wherein B is > S or > SO (- or p-);R' represents hydrogen or a carboxyl blocking group, e.g. the residue of an ester-forming aliphatic or araliphatic alcohol cr an ester-forming phenol, silanol or stannanol (the said alcohol, phenol, silanol or stannanol preferably containing 1-20 carbon atoms) and the dotted line bridging the 2-, 3- and 4-positions indicates that the compound is a 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 or toluene-p-sulphonic acid) or a base salt formed with a cation such as Na+ or K+, or an N-silyl derivative thereof, with an acid of formula

(wherein R2 represents a carboxyl blocking group, e.g. as described for R'; and R3 is an amino or protected amino group) or with an acylating agent corresponding thereto; whereafter, if necessary and/or desired in each instance, any of the 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 > SO to form a compound wherein B is > S, iii) conversion of a carboxyl group into a nontoxic salt or non-toxic metabolically labile ester function, and iv) removal of any carboxyl blocking and/or Nprotecting groups.

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

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

Acylations employing acid halides may be effected in aqueous and nonaqueous reaction media, conveniently at temperatures of from -50 to +500C, preferably -20 to +300C, if desired in the presence of an acid binding agent. Suitable reaction media include aqueous ketones such as aqueous acetone, 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 include tertiary amines (e.g. triethylamine or dimethylaniline), inorganic bases (e.g. calcium carbonate or sodium bicarbonate), and oxiranes such as lower 1,2-alkylene oxides (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 N,N'-dicyclohexylcarbodiimide or N-ethyl-N'-y dimethylaminopropylcarbodiimide; a carbonyl compound such as carbonyldiimidazole; or an isoxazolium salt such as N-ethyl-5phenylisoxazolium perchlorate.

Acylation may also be effected with other amide-forming derivatives of acids of formula (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 lower alkylhaloformate). Mixed anhydrides may also be formed with phosphorus acids (for example phophoric or phosphorus acids), sulphuric acid or aliphatic or aromatic sulphonic acids (for example toluene-p-sulphonic acid). An activated ester may conveniently be formed in situ using, for example, 1 hydroxybenzotriazole in the presence of a condensing agent as set out above. Alternatively, the activated ester may be preformed.

Acylation reactions involving the free acids or their above-mentioned amide-forming derivatives are desirably effected in an anhydrous reaction medium, e.g. methylene chloride, tetrahydrofuran, dimethylformamide or acetonitrile.

If desired, the above acylation reactions may be carried out in the presence of a catalyst such as 4-dimethylaminopyridine.

The acids of formula (Ill) 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 hydrobromide salts.

A2-Cephalosporin ester derivative obtained in accordance with the process of the invention may be converted into the corresponding A3derivatives by, for example, treatment of the A2 ester with a base such as pyridine or triethylamine.

A ceph-2-em reaction product may also be oxidised to yield the corresponding ceph-3-em 1oxide, for example by reaction with a peracid, e.g.

peracetic or m-chloroperbenzoic acid; the resulting sulphoxide may, if desired, subsequently be reduced as described hereinafter to yield the corresponding ceph-3-em sulphide.

Where a compound is obtained in which B is > SoO this may be converted to the corresponding sulphide by, for example, reduction of the corresponding acyloxy-sulphonium or alkoxysulphonium 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 watermiscible solvent e.g. acetic acid, acetone, tetrahydrofuran, dioxan, dimethylformamide or dimethylacetamide. The reaction may be effected at a temperature of from --200 to +500C.

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

Base salts of the compound of formula (I) may be formed by reacting an acid of formula (I) with an appropriate base. Thus, for example, sodium or potassium salts may be prepared using the respective 2-ethylhexanoate or hydrogen carbonate salt. Acid addition salts may be prepared by reacting the compound of formula (I) or a metabolically iabile ester derivative thereof with an appropriate acid.

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

For use as starting materials for the preparation of the compound of formula (I) according to the invention, compounds of general formula (III) and acid halides and anhydrides corresponding thereto in their syn isomeric form or in the form of mixtures of the syn isomers and the corresponding anti isomers containing at least 90% of the syn isomer are preferably used.

Acids of general formula (III) may be prepared by reaction of a compound of formula

(wherein R3 is as hereinbefore defined and R4 represents a carboxyl blocking group) with a compound of formula

(wherein R2 is as defined above) followed by removal of the carboxyl blocking group R4, and where necessary by the separation of syn and anti isomers.

The acids of formula (III) may be converted to the corresponding acid halides and anhydrides and acid addition salts by conventional methods, for example as described hereinbefore.

It should be appreciated that in some of the above transformations it may be necessary to protect any sensitive groups in the molecule of the compound in question to avoid undesirable side reactions. For example, during any of the reaction sequences referred to above it may be necessary to protect the NH2 group of the aminothiazolyl moiety, for example by tritylation, acylation (e.g. chloroacetylation), protonation or other conventional method. The protecting group may thereafter be removed in any convenient way which does not cause breakdown of the 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 or trifluoroacetic acid or using a mineral acid, e.g.

hydrochloric acid or mixtures of such acids, preferably in the presence of a protic solvent such as water or, in the case of a chloroacetyl group, by treatment with thiourea.

Carboxyl blocking groups used in the preparation of the compound of formula (I) or in the preparation of necessary starting materials are desirably groups which may readily be split off at a suitable stage in the 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 acyloxymethyl groups (e.g. acetoxymethyl and pivaloyloxymethyl) and retain these in the final product to give an appropriate ester derivative of the compound of formula (I).

Suitable carboxyl blocking groups are well known in the art, a list of representative blocked carboxyl groups being included in British Patent No. 1,399,086. Preferred blocked carboxyl groups include aryl lower alkoxycarbonyl groups such as p-methoxybenzyloxycarbonyl, pnitrobenzyloxycarbonyl and diphenylmethoxycarbonyl; lower alkoxycarbonyl groups such as t-butoxycarbonyl; and lower haloalkoxycarbonyl groups such as 2,2,2trichloroethoxycarbonyl. Carboxyl blocking group(s) may subsequently be removed by any of the appropriate methods disclosed in the literature; thus, for example, acid or base catalysed hydrolysis is applicable in many cases, as are enzymically-catalysed hydrolyses.

The antibiotic compounds of the 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 or excipients.

The antibiotic compounds according to the invention may 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 aqueous vehicles, and may contain formulatory agents such as suspending, stabilising and/or dispersing agents. Alternatively the active ingredient may be in powder form for reconstitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use.

If desired, such powder formulations may contain an appropriate non-toxic base in order to improve the water solubility of the 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 water with which the powder is reconstituted. The base may be, for example, an inorganic base such as sodium carbonate, sodium bicarbonate or sodium 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 for veterinary 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. 0.199%, of the active material, depending on the method of administration.

When the compositions comprise dosage units, each unit will preferably contain 50-1 500 mg of the active ingredient. The dosage as employed for adult human treatment will preferably range from 500 to 6000 mg per day, depending on the route and frequency of administration. For example, in adult human treatment 1000-3000 mg per day administered intravenously or intramuscularly will suffice. In treating Pseudomonas infections higher daily doses may be required.

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 illustrate the invention.

All temperatures are in OC.'Petrol' means petroleum ether (b.p. 4060 ).

Preparation 1 Ethyl (Z)-2-(2-aminothiazol-4-yl)- 2-(hydroxyimino)acetate To a stirred and ice-cooled solution of ethyl acetoacetate (292 g) in glacial acetic acid (296 ml) was added a solution of sodium nitrite (180 g) in water (400 ml) at such a rate that the reaction temperature was maintained below 1 OOC.

Stirring and cooling were continued for about 30 min., when a solution of potassium chloride (160 g) in water (800 ml) was added. The resulting mixture was stirred for one hour. The lower oily phase was separated and the aqueous phase was extracted with diethyl ether. The extract was combined with the oil, washed successively with water and saturated brine, dried, and evaporated.

The residual oil, which solidified on standing, was washed with petrol and dried in vacuo over potassium hydroxide, giving ethyl (Z)-2 (hydroxyimino)-3-oxobutyrate (309 g).

A stirred and ice-cooled solution of ethyl (Z)-2 (hydroxyimino)-3-oxobutyrate (150 g) in dichloromethane (400 ml) was treated dropwise with sulphuryl chloride (140 g). The resulting solution was kept at room temperature for 3 days, then evaporated. The residue was dissolved in diethyl ether, washed with water until the washings were almost neutral, dried, and evaporated. The residual oil (177 g) was dissolved in ethanol (500 ml) and dimethylaniline (77 mi) and thiourea (42 g) were added with stirring.

After two hours, the product was collected by filtration, washed with ethanol and dried to give the title compound (73 g); m.p. 1 880 (decomp.).

Preparation 2 Ethyl (Z)-2-hydroxyimino-2-(2tritylaminothiazol-4-yl)-acetate, hydrochloride Trityl chloride (16.75 g) was added portionwise over 2 hours to a stirred and cooled (30 ) solution of the product of Preparation 1 (12.91 g) in dimethylformamide (28 mi) containing triethylamine (8.4 ml). The mixture was allowed to warm to 1 50 over one hour, stirred for a further 2 hours and then partitioned between water (500 ml) and ethyl acetate (500 ml). The organic phase was separated, washed with water (2x500 ml) and then shaken with 1 N HCI (500 ml). The precipitate was collected, washed successively with water (100 ml), ethyl acetate (200 ml) and ether (200 ml) and dried in vacuo to provide the title compound as a white solid (16.4 g); m.p. 184 to 1860 (decomp.).

Preparation 3 Ethyl iZ)-2-(2-tritylaminothiazol4-yl)-2-(1 -t-butoxycarbonylcyclobut 1-oxyimino) acetate The product of Preparation 2 (55.8 g) was stirred under nitrogen in dimethylsulphoxide (400 ml) with potassium carbonate (finely ground, 31.2 g) at room temperature. After 30 minutes, t-butyl 1-bromocyclobutane carboxylate (29.2 g) was added. After 8 hours further potassium carbonate (31.2 g) was added. More potassium carbonate (6x 16 g portions) was added during the next three days and further t-butyl 1 bromocyclobutane carboxylate (3.45 g) was added after 3 days. After 4 days in all, the mixture was poured into ice-water (ca. 3 litres) and the solid was collected by filtration and washed well with water and petrol.The solid was dissolved in ethyl acetate and the solution washed with brine (twice), dried with magnesium sulphate and evaporated to a foam. This foam was dissolved in ethyl acetate-petrol (1:2) and filtered through silica gel (500 g). Evaporation gave the title compound (60 g) as a foam, human (CHBr3) 3400 (NH) and 1730 cm-l (ester).

Preparation 4 (Z)-2-(1 -t-Butoxycarbonylcyclobut 1 -oxyimino)-2-(2-tritylaminothiazol 4-yl) acetic acid A mixture of the product of Preparation 3 (3.2 g) and potassium carbonate (1.65 g) was refluxed in methanol (180 ml) and water (20 ml) for 9 hours and the mixture was cooled to room temperature. The mixture was concentrated and the residue partitioned between ethyl acetate and water, to which was added 2N HCI (12.2 ml). The organic phase was separated and the aqueous phase extracted with ethyl acetate. The combined organic extracts were washed with saturated brine, dried and evaporated to give the title compound (2.3 g); AmaX (ethanol) 265 nm (E1Cm 243).

Example 1 a) t-Butyl(6R,7 R)-3-Acetoxymethyl-7- [(Z)-2-(1 -t-butoxywarbonylcyclobut- 1 -oxyimino)-2-(2-tritylaminothiazol4-yl)acetamido]ceph-3-em4-carboxylate A stirred solution of the product of Preparation 4 (24.2 g) and t-butyl (6R,7R)-3-acetoxymethyl7-aminoceph-3-em-4-carboxylate (13.6 g) in dimethylformamide (300 ml) was cooled to 00, and 1-hydroxybenzotriazole monohydrate (4.5 g) added, followed by dicyclohexylcarbodiimide (6.4 g). The mixture was warmed to room temperature and stirred overnight. The mixture was filtered, and the white solid washed with a little ether. The filtrate and washings were diluted with water (1.51) and extracted with ethyl acetate. The organic extracts were combined, washed successively with water and saturated brine, dried, and evaporated.The residue was taken up in ether, filtered, and re-evaporated. The required product was isolated after elution through two silica columns with ether and concentrating the appropriate fractions. The residues were recrystallised from di-isopropyl ether to give the title compound (12.8 g), m.p. 113.5 to 116.50; [a] 0+15.00 (c 1.0, DMSO).

b) (6R,7R)-3-Acetoxymethyl-7 [(Z)-2-(2-aminothiazol-4-yl)-2 (l-carboxycyclobut-l-oxyimino) acetamido]ceph-3-em4-carboxylic acid Trifluoroacetic acid (100 ml) was added to a mixture of the product of Stage a) (12.5 g) and anisole (5 ml) at 00. The mixture was stirred at room temperature for 1 hour and concentrated.

The residue was dissolved in ethyl acetate and reconcentrated. The residue was dissolved in ethyl acetate and extracted with saturated sodium bicarbonate solution. The aqueous extracts, at pH 7 to 7.5, were washed with ethyl acetate, acidified to pH 1.5, and extracted several times with ethyl acetate. These extracts were washed with saturated brine, dried, and evaporated. The residue was dissolved in formic acid (70 ml), water (18 ml) was added, and the mixture stirred at room temperature for 2 hours. The mixture was diluted with water (300 ml) and filtered. The filtrate was concentrated. The residue was taken up in water (400 ml), refiltered, and lyophilized to give the title compound (4 g), AmaX (pH 6 buffer) 246 nm (E,,,1264), Anf 295 nm (Eicm1 118), [c']20+27.30 (c 1.0, DMSO).

Example 2 (6R,7R)-3-Acetoxymethyl-7-[(Z)-2 (2-aminothiazol-4-yl)-2-(1 carboxycyclobut-l -oxyimino) acetamido] ceph-3-em-4-carboxylic Acid Hydrate A solution of t-butyl (6R,7R)-3-acetoxymethyl 7-[(Z)-2-(l-t-butoxycarbonylcyclobut-l- oxyimino)-2-(2-tritylaminothiazol-4yl)aceta mido] ceph-3-em-4-carboxylate (89.41 g) in 98% formic acid (240 ml) was stirred for 15 minutes and concentrated hydrochloric acid (25 ml) was added. The suspension was stirred at 230 for 2.25 hours, cooled to 80 and filtered. The filtrate was concentrated to 60 ml and diluted with water (11). The solution was adjusted to pH 4.75 with ammonia and filtered. The filtrate was adjusted to pH 3 with phosphoric acid and the suspension cooled to 0 overnight.The precipitate was collected by filtration to give the title compound (49.4 g). A sample (10 g) was reprecipitated from water as above, dissolved in IMS (60 ml) at 580 and diluted with water (300 ml) at 55 . The solution was cooled slowly when the title compound was obtained as a microcrystalline solid (5.64 g), :tmax 253 nm (E1cm1%332) Ainfl 293 nm (Eicm1 151) (pH 6 phosphate buffer), H20 7.8% (Karl Fischer).

The X-ray data was obtained by Debye Scherrer powder diffraction photographs taken by exposure to Co K, (=1.79024 A).

d(A) 1/Iioo 16.5 54 11.2 62 8.43 85 5.96 31 5.78 77 5.51 46 5.06 15 4.91 4 4.73 31 4.53 100 4.39 85 4.26 38 4.02 23 3.94 31 3.86 31 3.74 77 3.67 62 3.50 92 3.35 31 3.04 46 2.99 31 2.93 23 2.83 15 2.73 23 2.64 23 2.54 23 2.44 8 2.39 8 2.31 15 1/lioo 2.14 8 2.09 8 2.01 8 1.95 8

Claims (4)

Claims

1. (6R,7R)-7-[(Z)-2-(2-Aminothiazol-4-yl)-2 11 -carboxycyclobut-1 -oxyimino) acetamido]-3- acetoxymethylceph-3-em-4-carboxylic acid of the formula

and its non-toxic salts.

2. Non-toxic metabolically labile esters of the compound of claim 1.

3. A process for the preparation of the compound of formula (I) as defined in claim 1 or a non-toxic salt or non-toxic metabolically labile ester thereof which comprises acylating a compound of formula

wherein B is > S or > SoO, R' represents hydrogen or a carboxyl blocking group and the dotted line in formula (II) bridging the 2-, 3- and 4-positions indicates that the compound is a ceph-2-em or ceph-3-em compound, or a salt or N-silyl derivative thereof, with an acid of formula

(wherein R2 represents an ester-forming carboxyl blocking group; and R3 is an amino or protected amino group) or with an acylating agent corresponding thereto; whereafter, if necessary and/or desired in each instance, any of the following reactions, in any appropriate sequence, are carried out: i) conversion of a A isomer into the desired A3-isomer, ii) reduction of a compound wherein B is > S-tO to form a compound wherein B is > S, iii) conversion of a carboxyl group into a nontoxic salt or non-toxic metabolically labile ester function, and iv) removal of any carboxyl blocking and/or Nprotecting groups.

4. A pharmaceutical composition for use in human or veterinary medicine comprising an antibiotic compound as claimed in claim 1 or claim 2 in association with a pharmaceutical carrier or excipient.