EP0231244A1 - 6-alkylidene penems - Google Patents

Abstract

Compounds of general formula (I) as well as their pharmaceutically acceptable salts and esters hydrolysable in vivo, formula in which one of R1 and R2 represents hydrogen, the other representing a bicyclic heteroaromatic group, substituted or unsubstituted and linked through l 'one of these carbon atoms and having five or six atoms in each ring, and R3 represents hydrogen or an organic group. These new compounds have anti-bacterial and beta-lactamase inhibiting properties.

Description

6-alkylidene penems.

This invention relates to β-lactam compounds and in particular to a class of 6-alkylidene penems which have I3-lactamase inhibitory and antibacterial properties. The compounds are therefore useful in the treatment of antibacterial infections in humans or animals, either alone or in combination with other antibiotics.

European Patent Publication No. EP 0 041 768 A (Beecham; published 16 December 1981; corresponding to U.S. Patent No 4485 110) discloses

6-alkylidene-2-penems of the general formula (A):

(A)

in which

each of R^a and R^b denotes hydrogen or an optionally substituted hydrocarbon or heterocyclic group, and

R^c denotes hydrogen or an organic group.

Those compounds possess antibacterial activity and also inhibit β-lactamases and have a synergistic effect in combination with other β-lactam antibiotics. European Patent Publication No. EP 0 120 613 A (Beecham; published 3 October 1984) discloses a sub-group of compounds within the general formula (A) which have better activity than other compounds of the general formula (A). That sub-group consists of compounds of the general formula (B):

(B)

in which

R^c denotes hydrogen or an organic group;

one of R^d and R^e denotes hydrogen, and

the other of R^d and R^e denotes a group of the sub-formula (C):

(C)

in which

R^f denotes a substituent group; X denotes an oxygen atom, a sulphur atom or an >NRg group;

Rg denotes hydrogen, hydrocarbon or a nitrogen-protecting group; and

n denotes 0, 1, 2 or 3.

It has now been found that certain compounds of the general formula (A) exhibit improved β-lactamase inhibitory action and synergistic activity as compared with other compounds of that group.

According to the present invention there is provided a compound of the general formula I:

or a pharmaceutically acceptable salt or in vivo hydrolysable ester thereof

in which

one of R¹ and R² denotes hydrogen,

the other of R¹ and R² denotes an unsubstituted or substituted fused bicyclic hetero-aromatic group bonded through a carbon atom thereof and having five or six atoms in each ring, and R³ denotes hydrogen or an organic group.

The expression 'fused bicyclic hetero-aromatic group' as used herein means a group comprising two fused rings together having aromatic character at least one of which rings contains a non-carbon atom (i.e. a hetero-atom) as a ring member and at least one of which rings contains a carbon-bonded free valency by which the group is bonded to the remainder of the molecule.

Each ring of the fused bicyclic hetero-aromatic group R¹ or R² may contain 5 or 6 ring atoms (including ring junction atoms - also referred to as bridgehead atoms - constituting members of both rings) and the hetero-aromatic group may consist of two fused rings both of the same size or it may consist of one 5-membered ring fused to one 6-membered ring.

The fused bicyclic hetero-aromatic group R¹ or R² is bonded to the remainder of the molecule through a carbon atom of one of the rings. In the case of a hetero-aromatic group containing both a 5-membered and a 6-merabered ring, the said bond may be from either ring.

The fused bicyclic hetero-aromatic group R¹ or R² may comprise two fused 5-membered rings, two fused 6-membered rings, or one 5-membered ring and one 6-membered ring fused together (in which last-mentioned case, the bond to the remainder of the molecule may be from either the 5-membered ring or the 6-membered ring). In other words the bicyclic hetero-aromatic group may be based on a bicyclo[3.3.0]octyl, bicyclo[4.3.0]nonyl, or bicyclo[4.4.0]decyl ring system. The fused bicyclic hetero-aromatic group may contain one or more hetero-atoms selected from nitrogen, oxygen and sulphur atoms. Provided that at least one hetero-atom is present in at least one of the rings (which may be a 5-membered ring or a 6-membered ring), it is not necessary for both rings to contain a hetero-atom: one ring may contain no hetero-atoms or both rings may each contain one or more hetero-atoms depending on the type of hetero-atoms. Also, the ring through which the hetero-aromatic group is bonded to the remainder of the molecule should contain at least one non-bridgehead carbon atom for attachment of the bond.

The maximum possible number of each type of hetero-atom and the various possible combinations of two or more different hetero-atoms is dictated by the ring structure and the requirement for aromaticity and will be apparent to the chemist.

The hetero-aromatic group may contain one bridgehead nitrogen atom at the ring fusion without losing its aromaticity (see, for example, M.J. Cook et al, 'Aromaticity of heterocycles', Advances in heterocyclic chemistry, Academic Press, 1974, 17 , 255ff.).

Also, a substituent in at least one of the rings, in particular an sp²-hydridized substituent, for example an oxo-substituent, may contribute to the aromaticity of the hetero-aromatic group (see, for example, A.R. Katritzky and J.M. Lagowski, 'Protopic tautomerism of heteroaromatic compounds', Advances in heterocyclic chemistry, Academic Press, 1963, 1, 311, 339; 2 , 1, 27; Supplement 1). In some cases, for example in certain bicyclic ring systems in which a 6-membered ring contains an oxygen atom, a sulphur atom or a substituted nitrogen atom as a hetero-atom, such a substituent may be essential to the aromaticity of the ring system. Examples of hetero-aromatic groups containing an sp²-hydridized substituent include chromonyl groups and coumarinyl groups.

The fused bicyclic hetero-aromatic group R¹ or R² may suitably comprise two fused rings together constituting an aromatic system and each selected from:

(i) a five-membered carbocycliσ ring,

(ii) a five-membered heterocyclic ring containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen, and optionally also containing up to three, preferably not more than two, additional ring nitrogen atoms, the remaining ring atoms being carbon atoms,

(iii) a six-membered carbocyclic ring,

(iv) a six-membered heterocyclic ring containing from one to four, preferably from one to three, nitrogen atoms, the remaining ring atoms being carbon atoms,

(v) a six-membered heterocyclic ring containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen, containing one carbon atom carrying an sp²-hydridized substituent in conjugation with the said hetero-atom, and optionally also containing up to three, advantageously not more than two, and preferably not more than one, additional ring nitrogen atoms, the remaining ring atoms being carbon atoms;

with the provisos that :

(i) at least one of said rings is a heterocyclic ring,

(ii) the ring fusion is at a carbon-carbon bond or a carbon-nitrogen bond, and

(iii) one of said rings contains a carbon atom through which the hetero-aromatic group is bonded to the remainder of the molecule.

The expression 'ring nitrogen atom' as used herein means a nitrogen atom all three bonds of which are attached to other ring atoms, either through a double bond to one adjacent ring atom and a single bond to another adjacent ring atom or, in the case of a bridgehead nitrogen atom, through three single bonds to an adjacent atom in each ring and to the other bridgehead atom. Examples of 'ring nitrogen atoms' are the nitrogen atoms in the following structures:

in which the broken line denotes the remainder of a ring or ring-system.

The expression 'substituted nitrogen atom' as used herein means a nitrogen atom forming part of a heterocyclic ring and having two bonds attached to other ring atoms and one bond attached to an atom or group outside the ring (including a hydrogen atom). Such a 'substituted nitrogen atom' may, for example, be denoted by -N(R°)-, in which R° is defined as below. Examples of 'substituted nitrogen atoms' are those in the following structures:

in which the broken line has the meaning given above and in which the H atoms may be replaced by other substituents.

The expression 'bridgehead atom' as used herein means one of the two atoms at a ring fusion junction which are common to both rings. In general, the bridgehead atoms may both be carbon atoms or may be one carbon atom and one nitrogen atom, as in the following structures:

IVC

It may be advantageous for one of the bridgehead atoms to be a nitrogen atom.

Examples of fused bicyclic hetero-aromatic groups R¹ or R² include the following groups:

(i) A group consisting of two mutually fused five-membered rings each containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen, at least one of the said hetero-atoms being adjacent to a bridgehead atom, one ring optionally additionally containing one or two ring-nitrogen atoms, the other ring optionally additionally containing one ring-nitrogen atom, the two bridgehead atoms being carbon atoms, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring.

Such groups may be represented by the formulae

and in which

each of X¹ and X², which may be identical or different, denotes -O-, -S-, or -N(R°)-, R° denotes a hydrogen atom or an unsubstituted or substituted hydrocarbon group, and

up to three of the =CH- groups may optionally be replaced by ring-nitrogen atoms =N-.

Examples of such groups include:

furopyrrolyl, thienopyrrolyl, pyrrolopyrrolyl, furopyrazolyl, thienopyrazolyl, pyrrolopyrazolyl, furoimidazolyl, thienoimidazolyl, pyrrolo- imidazolyl, pyrazoloimidazolyl, furotriazolyl, thienotriazolyl, pyrrolotriazolyl, pyrazolo- triazolyl, imidazotriazolyl, furofuryl, thienofuryl, and thienothienyl groups; and furo-oxazolyl, thieno-oxazolyl, pyrrolo- oxazolyl, pyrazolo-oxazolyl, imidazo-oxazoyl, oxazolo-triazolyl, oxazolo-oxazolyl, isoxazolo- oxazolyl, thiazolo-oxazolyl, isothiazolo-oxazolyl, oxazolo-oxadiazolyl, and oxazolo-thiadiazolyl groups, as well as the isoxazole, thiazole and isothiazole derivatives corresponding to the above-mentioned oxazole ring systems; in all of which groups both bridgehead atoms are carbon atoms, and in all of which groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atoms). All isomeric forms of the above-mentioned groups that do not contain bridgehead nitrogen atoms are included.

(ii) A group consisting of two mutually fused five-membered rings, one of the bridgehead atoms being a carbon atom and the other bridgehead atom being a nitrogen atom, one of the rings containing one hetero-atom adjacent to a bridgehead atom and selected from oxygen, sulphur and substituted nitrogen, and optionally additionally containing one or two ring-nitrogen atoms, the other ring optionally containing one, two or three ring-nitrogen atoms, with the proviso that the total number of ring-nitrogen atoms (excluding the bridgehead nitrogen atom) does not exceed four, preferably not exceeding three, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring.

Such groups may be represented by the formulae

and in which

X¹ is defined as above, and

up to four, preferably not more than three, of the =CH- groups may optionally be replaced by ring-nitrogen atoms =N-.

Examples of such groups include:

pyrroloimidazolyl, imidazoimidazolyl, pyrrolo- pyrazolyl, pyrazolopyrazolyl, imidazopyrazolyl, pyrrolotriazolyl, imidazotriazolyl, pyrazolo- triazolyl, pyrrolotetrazolyl, imidazotetrazolyl, and pyrazolotetrazolyl groups; pyrrolo-oxazolyl, imidazo-oxazolyl, pyrazolo-oxazolyl, oxazolo- triazolyl, oxazolo-tetrazolyl, pyrrolo- oxadiazolyl, imidazo-oxadiazolyl, pyrazolo- oxadiazolyl, triazolo-oxadiazolyl, pyrrolo- oxatriazolyl, imidazo-oxatriazolyl, and pyrazolo- oxatriazolyl groups, as well as the isoxazole derivatives corresponding to the above mentioned oxazole ring systems, and also the thia derivatives corresponding to the above-mentioned oxazole, isoxazole, oxadiazole and oxatriazole ring systems (e.g. imidazothiazolyl); in all of which groups one bridgehead atom is a nitrogen atom, the other being a carbon atom, and in all of which groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atom). All isomeric forms of the above-mentioned groups that contain one bridgehead nitrogen atom are included.

(iii)A group consisting of a five-membered ring and a six-membered ring fused together with both bridgehead atoms being carbon atoms, the five-membered ring containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen and optionally additionally containing one or two ring-nitrogen atoms, the six-membered ring optionally containing up to four ring-nitrogen atoms, with the proviso that the total number of ring-nitrogen atoms does not exceed five, preferably not exceeding three, or the six-membered ring optionally containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen, and also containing an sp^-hybridized substituent in conjugation with the said hetero-atom, and also optionally containing one or two ring-nitrogen atoms, with the proviso that the total number of ring-nitrogen atoms does not exceed three, preferably not exceeding one, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring.

Such groups may be represented by the formulae

and

in which

up to five, preferably not more than three, of the =CH- groups may optionally be replaced by ring-nitrogen atoms =N-, or

one of the =CH- groups in the six-membered ring may optionally be replaced by a group -X²- with the =CH- group in the ortho- or para- position thereto being replaced by -C(=Z)-, and up to three, preferably not more than one, of the remaining =CH- groups in either ring may optionally be replaced by ring-nitrogen atoms =N-,

X¹ and X² are defined as above

Z denotes an sp²-hybridized substituent.

Examples of such groups include: benzofuryl, benzothienyl, benzopyrrolyl (indolyl), benzopyrazolyl, benzoimidazolyl, benzo-oxazolyl, benzoisoxazolyl, benzothiazolyl, benzo- isothiazolyl, benzo-oxadiazolyl, benzo- thiadiazolyl, benzotriazolyl, furopyridyl, thienopyridyl, pyrrolopyridyl, pyrazolopyridyl, imidazoypyridyl, triazolopyridyl, oxazolopyridyl, isoxazolopyridyl, thiazolopyridyl, and isothiazolopyridyl groups, the pyrazinyl, pyrimidinyl and pyridazinyl derivatives corresponding to the above-mentioned pyridyl ring systems, oxadiazolopyridyl, thiadiazolopyridyl, furotriazinyl, thienotriazinyl, and pyrrolotriazinyl groups, as well as ring systems corresponding to the above-mentioned benzo ring systems (more particularly the first nine above-mentioned benzo ring systems) in which the benzene ring is replaced by a six-membered ring including one hetero-atom selected from oxygen, sulphur and substituted nitrogen and carrying an sp²-hybridised substituent ortho or para to the said hetero-atom, in all of which above-mentioned groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring

(excluding the bridgehead carbon atoms). All isomeric forms of the above-mentioned groups are included.

(iv) A group consisting of a five-membered ring and a six-membered ring fused together with both bridgehead atoms being carbon atoms, the six-membered ring containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen and optionally additionally containing one, two or three ring nitrogen atoms, the five-membered ring optionally containing one, two or three ring-nitrogen atoms, with the proviso that the total number of ring-nitrogen atoms does not exceed five, advantageously not exceeding four, preferably not exceeding three, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring.

Such groups may be represented by the formulae

and

in which

X¹ is defined as above, and

up to five, advantageously not more than four, preferably not more than three, of the =CH- groups may optionally be replaced by ring-nitrogen atoms =N-.

Examples of such groups include the following: cyclopentapyranyl, cyclopentaoxazinyl, cyclopenta- oxadiazinyl, cyclopentaoxatriazinyl, pyrano- pyrrolyl, pyranopyrazolyl, pyranoimidazolyl, pyrano-triazolyl, pyrrolo-oxazinyl, pyrazolo- oxazinyl, imidazo-oxazinyl, and pyrrolo- oxadiazinyl groups, and the thia derivatives corresponding to the above mentioned pyran and other oxa ring systems, the 1H-cyclopentapyridyl group and corresponding derivatives in which up to three additional ring-nitrogen atoms are present in either ring, in all of which above-mentioned groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atoms). All isomeric forms of the above-mentioned groups areincluded.

(v) A group consisting of a five-membered ring and a six-membered ring fused together with one bridgehead atom being a carbon atom and the other bridgehead atom being a nitrogen atom, the five-membered ring optionally containing one, two or three ring-nitrogen atoms (additional to the bridgehead nitrogen atom), and the six-membered ring optionally containing up to four ring-nitrogen atoms (additional to the bridgehead nitrogen atom), with the proviso that the total number of ring-nitrogen atoms does not exceed five, preferably not exceeding four, inclusive of the bridgehead nitrogen atom, or the six-membered ring optionally containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen, and also containing an sp²-hybridized substituent in conjugation with the said hetero-atom, and also optionally containing one or two ring-nitrogen atoms (additional to the bridgehead nitrogen atom), with the proviso that the total number of ring-nitrogen atoms does not exceed three, preferably not exceeding two, inclusive of the bridgehead nitrogen atom, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring.

Such groups may be represented by the formula

IX

in which

up to four, preferably not more than three, of the =CH- groups may optionally be replaced by ring nitrogen atoms =N-, or

one of the =CH- groups in the six-membered ring may optionally be replaced by a group -X¹- with the =CH- group in the ortho- or para-position thereto being replaced by -C(=Z)-, and up to two, preferably not more than one, of the remaining =CH- groups in either ring may optionally be replaced by a ring-nitrogen atom =N-, and

χ¹ and Z are defined as above.

Examples of such groups include the indolizinyl group, as well as corresponding derivatives containing up to three additional ring-nitrogen atoms in either ring, for example pyrazolo- [1,5-a]pyridine, imidazo[l,2-a]pyrazinyl, and also corresponding derivatives in which the six- membered ring includes an additional hetero-atom selected from oxygen, sulphur and substituted nitrogen and carries an sp²-hydridised substitutent ortho or para to the said hetero-atom, in all of which above-mentioned groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atom). All isomeric forms of the above-mentioned groups are included.

(vi) A group consisting of two mutually fused six-membered rings, both bridgehead atoms being carbon atoms, the group containing from one to five, preferably from one to four, ring-nitrogen atoms in either ring, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring.

Such groups may be represented by the formula

in which

from one to five, preferably from one to four, of the =CH- groups are replaced by ring nitrogen atoms =N-.

Examples of such groups include:

benzopyridyl, benzopyrazinyl, benzopyrimidinyl, benzopyridazinyl, benzotriazinyl, benzotetrazinyl, pyridopyridyl, pyridopyrazinyl, pyridopyrimidinyl, pyridopyridazinyl, pyridotriazinyl, pyrazino- pyrazinyl, pyrazinopyrimidinyl, pyrazino- pyridazinyl, pyrimidopyrimidinyl, pyrazinopyridazinyl, pyrimidopyrimidinyl, pyrimidopyridazinyl, and pyridazopyridazinyl groups, including, for example, quinolyl, isoquinolyl, quinoxalinyl, quinazolinyl, phthalazinyl, cinnolinyl, naphthyridinyl, and pteridinyl groups, in all of which above- mentioned groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atoms). All isomeric forms of the above-mentioned groups are included.

(vii)A group consisting of two mutually fused six-membered rings, one bridgehead atom being a carbon atom and the other bridgehead atom being a nitrogen atom, one of the rings containing, in a position ortho or para to the bridgehead nitrogen atom, one hetero-atom selected from oxygen, sulphur and substituted nitrogen, and optionally also containing one or two, preferably not more than one, ring-nitrogen atoms, the other ring optionally containing up to three ring-nitrogen atoms, with the proviso that the total number of ring-nitrogen atoms in the group does not exceed five inclusive of the bridgehead nitrogen atom, or the other ring optionally containing one hetero-atom selected from oxygen, sulphur and substituted nitrogen, and also containing an sp²-hybridized substituent in conjugation with the said hetero-atom, and also optionally containing one or two ring-nitrogen atoms (additional to the bridgehead nitrogen atom), with the proviso that the total number of ring-nitrogen atoms does not exceed three inclusive of the bridgehead nitrogen atom, and the group being bonded to the remainder of the molecule through a ring-carbon atom of either ring. Such groups may be represented by the formulae

and

XIA in which

up to four of the =CH- groups may optionally be replaced by ring-nitrogen atoms =N-, or

one of the =CH- groups in the six-membered ring not containing the X¹ group may optionally be replaced by a group -X²- with the =CH- group in the ortho- or para- position thereto being replaced by -C(=Z)-, and up to two of the remaining =CH- groups of either ring may optionally be replaced by ring-nitrogen atoms =N-, and

X¹, X² and Z are defined as above.

Examples of such groups include:

pyrido-oxazinyl, pyrido-oxadiazinyl, pyrido-oxatriazinyl, pyrido- oxatetrazinyl, pyrazino-oxazinyl, pyrazino- oxadiazinyl, pyrazino-oxatriazinyl, pyrimido- oxazinyl, pyrimido-oxadiazinyl, pyrimido- oxatriazinyl, pyridazino-oxazinyl, pyridazino- oxadiazinyl, and pyridazino-oxatriazinyl groups and the corresponding thia derivatives, as well as corresponding derivatives in which the ring not containing the oxygen or sulphur atom includes an additional hetero-atom selected from oxygen, sulphur and substituted nitogen and carries an sp²-hybridised substituent ortho or para to the said hetero-atom; oxazino-triazinyl, oxazinotetrazinyl, triazino-oxadiazinyl, oxadiazinotetrazinyl, and triazino-oxatriazinyl groups, and the corresponding thia derivatives, 1H-pyrido- pyridazinyl, 1H-pyridopyrazinyl, and 1H-pyridoprimidinyl groups, and corresponding derivatives containing a total of up to three additional ring-nitrogen atoms in either ring, and also corresponding derivatives in which the ring not containing the substituted nitrogen atom includes an additional heteroatom and carries an sp²-hybridised substituent as aforesaid; in all of which groups one bridgehead atom is a nitrogen atom, the other being a carbon atom, and in all of which groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atom). All isomeric forms of the above-mentioned groups that contain one bridgehead nitrogen atom are included.

(viii) A group consisting of two mutually fused six- membered rings, both bridgehead atoms being carbon atoms, at least one of the rings containing a hetero-atom selected from oxygen, sulphur and substituted nitrogen with an sp²-hybridized substituent in conjuguation therewith, and the group optionally containing up to three ring-nitrogen atoms or, if both rings contain a said hetero-atom and sp²-hybridized substitutent, up to two ring-nitrogen atoms. Such groups may be represented by the formula

XII in which

one or two of the =CH-groups are replaced, respectively by one or two, identical or different, groups -X¹- with a respective number of =CH- groups being replaced by identical or different groups -C(=Z)- each in conjugation with a respective said hetero-atom, and

up to three of the remaining =CH- groups in either ring optionally being replaced by ring-nitrogen atoms =N-, with the proviso that, if two groups -X¹- are present, the total number of ring-nitrogen atoms does not exceed two.

Examples of such groups include:

σhromonyl, coumarinyl, isocoumarinyl, and dipyronyl groups and corresponding thia, thione and H-aza analogues, and also corresponding derivatives containing a total of up to three (or, in the case of the dipyronyl derivatives up to two) ring-nitrogen atoms, in all of which above-mentioned groups the free valency (by which the group is attached to the remainder of the molecule) may extend from a carbon atom of either ring (excluding the bridgehead carbon atoms). All isomeric forms of the above-mentioned groups are included. The above listed hetero-aromatic groups are examples only of possible groups R¹ and R². Further examples include groups listed above carrying substituents listed below on a carbon atom or on a substituted nitrogen atom.

Particular examples of the hetero-aromatic group R¹ or R² include the benzo[b]furyl group:

and the indolyl group:

and N-substituted derivatives thereof, for example N-(p-methoxyphenylsulphonyl)-indolyl, and the benzotriazolyl group.

The fused bicyclic hetero-aromatic group may be unsubstituted or may be substituted on one or both rings by one or more substituents bonded to the respective ring at a carbon atom. Such substituents should not of course destroy the aromatic character of the hetero-aromatic group. Suitably, the hetero- aromatic group may be unsubstituted or may carry one or two substituents.

The fused bicyclic hetero-aromatic group R¹ or R² may include a substituent other than hydrogen at a substituted nitrogen atom. The nitrogen substituent may, for example, be a hydrocarbon group or a nitrogen-protecting group.

Examples of suitable substituents which may be present in the hetero-aromatic group R¹ or R² include (C_1-6)alkanoyl, (C_1-6)alkanoyloxy, heterocyclyl, amino, (C_1-6)alkanoylamino, (mono or di)-(C_1-6)alkylamino, hydroxy, (C_1-6) alkoxy, sulpho, mercapto, (C_1-6)alkylthio, (C_1-6) alkylsulphinyl, (C_1-6)alkyl- sαlphonyl, heterocyclylthio, arylthio, sulphamoyl, carbamoyl, amidino, guanidino, nitro, halogen, carboxy, carboxy salts, carboxy esters, arylcarbonyl, and heterocyclylcarbonyl, groups, and also unsubstituted or substituted (C_1-6) alkyl, (C_2-6)alkenyl, (C_2-6)alkynyl, aryl, and aryl(C_1-6) alkyl groups.

Examples of suitable optional substituents for the above-mentioned (C_1-6) alkyl, (C_2-6) alkenyl, (C_2-6)alkynyl, aryl and aryl (C_1-6) alkyl substitutents include (C_1-6)alkanoyl, (C_1-6)alkanoyloxy, heterocyclyl, amino, (C_1-6)alkanoylamino, (mono or di)-(C_1-6)alkylamino, hydroxy, (C_1-6)alkylsulphinyl, (C_1-6)alkylsulphonyl, heterocyclylthio, arylthio, sulphamoyl, carbamoyl, amidino, guanidino, nitro, halogen, carboxy, carboxy salts, carboxy esters, arylcarbonyl and heterocyclylcarbonyl groups.

When the hetero-aromatic group R¹ or R² includes a carboxy salt or carboxy ester substituent, that substituent is suitably a pharmaceutically acceptable salt or pharmaceutically acceptable ester. When the hetero-aromatic group R¹ or R² is or includes a basic moiety, the compound according to the invention may exist in zwitterionic form.

The term 'heterocyclyl' as used herein includes aromatic and non-aromatic, single and fused, rings containing up to four hetero-atoms in each ring selected from oxygen, nitrogen and sulphur, which rings may be unsubstituted or substituted by up to three groups selected from halogen, (C_1-6)alkyl, (C_1-6)alkoxy, halo(C_1-6)alkyl, hydroxy, amino, carboxy, (C_1-6)alkoxycarbonyl, (C_1-6)alkoxycarbonyl(C_1-6)alkyl, aryl, (C_1-6)alkylthio, arylthio, mercapto and oxo groups.

The term 'aryl' as used herein includes phenyl and naphthyl, which may be unsubstituted or substituted by up to five, preferably up to three, groups selected from halogen, (C_1-6)alkyl, phenyl, (C_1-6)alkoxy, halo(C_1-6)alkyl, hydroxy, amino, nitro, carboxy, (C_1-6)alkoxycarbonyl, (C_1-6)alkoxycarbonyl (C_1-6)alkyl, (C_1-6)alkylcarbonyloxy, (C_1-6)alkylcarbonyl, (C_1-6)alkylthio, arylthio, and mercapto groups.

The term 'hydrocarbon' as used herein includes groups having up to 18 carbon atoms, suitably up to 10 carbon atoms, conveniently up to 6 carbon atoms. Suitable hydrocarbon groups include (C_1-6)alkyl, (C_2-6)alkenyl, ( C_2-6 ) alkynyl , (C_3-7)cycloalkyl , (C_3-7) cycloalkyl- (C_1-6)alkyl, aryl, and aryl (C_1-6)alkyl.

In general formula (I), R³ represents hydrogen or an organic group, which may suitably be linked through a sulphur or carbon atom. For example, R³ may represent hydrogen or a group of formula -R⁴ or -SR⁴, where R⁴ denotes an unsubstituted or substituted (C_1-10)hydrocarbon or heterocyclyl group. Preferably, R³ represents hydrogen, (C_1-10)alkyl or (C_{1- 10})alkylthio, or substituted (C_{1 -10})alkyl or substituted ( C_{1-1 0} ) - alkylthio, wherein the substituent may be hydroxy, (C_1-6) alkoxy, (C_1-6)alkanoyloxy, halogen, mercapto, (C_1-6)alkylthio, heterocyclylthio, amino, (mono or di)-(C_1-6)alkylamino, (C_1-6)alkanoylamino, carboxy, or (C_1-6)alkoxycarbonyl.

Examples of suitable organic groups R³ include methyl, ethyl, propyl, methylthio, ethylthio, methylsulphinyl, ethylsulphinyl, hydroxymethyl, methoxymethyl, ethoxymethyl, acetoxymethyl, (1 or 2)-acetoxyethyl, aminomethyl, 2-aminoethyl, acBtamidomethyl, 2-acetamidoethyl, carboxymethyl, 2-hydroxyethylthio, methoxymethylthio, 2-methoxyethylthio, acetoxymethylthio, 2-aminoethylthio, acetamidomethylthio, 2;-acetamidoethylthio, carboxymethylthio, 2-carboxyethylthio, aryl (especially phenyl), arylthio (especially phenylthio), pyridyl, pyrimidyl, isoxazolyl, pyrimidylthio, tetrazolylthio, and pyridylthio groups. In particular, R³ may be hydrogen.

Alternatively, R³ may denote a group of the formula -OR⁷ or -R⁸, in which R⁷ denotes an unsubstituted or substituted phenyl, naphthyl, thienyl, pyridyl, quinolyl or isoquinolyl group, and R⁸ denotes a nitrogen-containing heterocyclyl ring bonded through a ring-nitrogen atom. Examples of groups denoted by R⁷ and R⁸, and also examples of suitable substituents for the aryl or heterocyclyl rings denoted by R⁷, are given in GB 2 102 798 A (Hoechst), EP 0 099 059 A (Hoechst), and EP 0 148 128 A (Ciba-Geigy; corresponding to AU 84/37236). Pharmaceutically acceptable in vivo hydrolysable esters (also referred to as 'metabolisable esters') of the compounds of the general formula I are those esters which hydrolyse in the human body to produce the parent acid or its salt. Such esters may be identified by oral or intravenous administration to a test animal, and subsequent examination of the test animal's body fluids for the presence of the compound of the formula I or a salt thereof.

In some cases, the in vivo hydrolysable ester moiety may constitute a link between two different active ingredient moieties, one of which is a compound according to the invention and the other of which may be another therapeutically active compound, such that on in vivo hydrolysis of the ester moiety, the ester link breaks to give the two separate active compounds. The linked entity may be referred to as a 'mutual pro-drug'.

Suitable in vivo hydrolysable ester groups include those of part-formulae (a), (b) and (c):

(a)

(b)

(c) in which

A¹ denotes hydrogen, methyl, or phenyl;

A² denotes (C_1-6)alkyl, (C_1-6)alkoxy or phenyl; or

A¹ and A² together denote 1, 2-phenylene, which may be unsubstituted or substituted by one or two methoxy groups;

A³ denotes (C_1-6)alkylene, which may be unsubstituted or substituted by a methyl or ethyl group;

each of A⁴ and A⁵ which may be identical or different, denotes (C_1-6)alkyl; and

A⁶ denotes (C_1-6) alkyl.

Examples of suitable in vivo hydrolysable ester groups include acetoxymethyl, pivaloyloxymethyl, α-acetoxyethyl, α-acetoxybenzyl, α-pivaloyloxyethyl, ethoxycarbonyloxymethyl, α-ethoxycarbonyloxyethyl, dimethylaminomethyl, diethylaminomethyl, phthalidyl and dimethoxyphthalidyl groups.

Suitable pharmaceutically acceptable salts of the 3-carboxylic acid group of the compound of formula I include metal salts, e.g. aluminium salts, alkali metal salts (e.g. sodium or potassium salts), alkaline earth metal salts (e.g. calcium or magnesium salts), ammonium salts, and substituted ammonium salts, for example those with lower alkylamines (e.g.triethylamine),hydroxy-lower alkylamines (e.g. 2-hydroxyethylamine), di(2-hydroxyethyl) amine or tri (2-hydroxyethyl) amine), eyeloalkylamines (e.g. dicyclohexylamine), or with procaine, and also dibenzylamine, N,N-dibenzylethylenediamine, 1-ephenamine, N-ethylpiperidine, N-benzyl-β-phenethylamine, dehydroabietylamine,

N,N'-bishydroabietylethylene-diamine, bases of the pyridine type (e.g. pyridine, collidine and quinoline), and other amines which have been or can be used to form salts with penicillins.

The compounds of the general formula I and their salts may exist in hydrated or non-hydrated form. The compounds of the general formula I and also the salts and esters thereof may exist in two optically active forms and it is to be understood that both such forms as well as racemic mixtures thereof are embraced by the present invention. It is believed that the more active form is that of structure IA:

in which R¹, R² and R³ are defined as above.

Furthermore, in general formulae I and IA, it is thought to be advantageous that R¹ denotes the hetero-aromatic group and that R² denotes a hydrogen atom.

Examples of individual compounds according to the invention include:

(5RS) (Z)-6-(benzo[b]fur-2-ylmethylene)ρenem-3- carboxylic acid; ( 5RS ) ( Z ) -6-[1- (4-methoxyphenylsulphonyl ) - indol-2-ylmethylene]penem-3-carboxylic acid;

(5RS) (Z)-6-(1-methylbenzotriazol-5-ylmethylene)- penem-3-carboxylic acid; (5RS) (Z)-6-(N-methylthienotriazol-5-ylmethylene)- penem-3-carboxylic acid;

(5RS) (Z)-6-(pyrazolo[2,3-a]pyrid-2-ylmethyene)- penem-3-carboxylic acid;

(5RS) (Z)-6-(thieno[3,2-b]fur-2-ylmethylene)- penem-3-carboxylic acid;

(5RS) (Z)-6-(imidazo[2,1-b]thiazol-6-ylmethylene)- penem-3-carboxylic acid;

(5RS) (Z)-6-(quinoxalin-2-ylmethylene)penem-3- carboxylic acid; (5RS) (Z)-6-(imidazo[1,2-a]pyrazin-2-yl- methylene)penem-3-carboxylic acid;

as well as pharmaceutically acceptable salts and in-vivo hydrolysable esters thereof.

A compound of the general formula I, or a salt or ester thereof, may be prepared by eliminating the elements of a compound of the general formula XIII:

H-X^o XIII

from a penem or penem intermediate of the general part-formula XIV:

in which

R¹ and R² are defined as above, and

X^o denotes a hydroxy group or a leaving group,

to give a compound of the general part-formula XV:

XV

in which R¹ and R² are defined as above,

and, if the resulting compound of the general formula IV is a penem intermediate, converting it into a penem of the general formula I or a salt or ester thereof.

The compound of the general part-formula XIV may suitably be a compound of the general part-formula XIVA:

XIVA in which R¹, R² and X^o are defined as above. More especially it may be a compound of the general formula XIVB:

XIVB in which

R¹, R² and X^o are defined as above,

R¹⁰ denotes (C_1-6)alkyl, aryl, aryl(C_1-6)alkyl, (C_1-6)alkylthio, arylthio, hetero-aromatic-thio, acyl (for example, (C_1-6) alkylcarbonyl, especially acetyl), ( C_2-6 ) alkenyl ( especially vinyl ) , or aryl(C_2-6)alkenyl, all of which may optionally be substituted, and

R¹¹ denotes hydrogen or an N-protecting group, or

R¹⁰ and R¹¹ together denote the remainder of a penem nucleus, which may be substituted and/or may optionally carry a protecting group.

In the case where R¹⁰ and R¹¹ in general formula XIVB together denote the remainder of a penem nucleus, they may suitably together denote the sub-formula XVI:

XVI

in which

R¹² denotes the hydrogen atom or organic group R³ or a group convertible into R³ during the preparation of a penem of the general formula I or salt or ester thereof, and

R^x denotes hydrogen or a carboxyl-blocking group.

In that case, the penem or penem intermediate of the general part-formula XIV is of the general formula XIVD given below.

A carboxyl-blocking group R^x (also referred to as a carboxyl-protecting group) is suitably a group that can readily be removed at a later stage of the penem preparation process.

Examples of suitable carboxyl-blocking derivatives that may form the group -CO₂R^x include salt, ester, and anhydride derivatives of the carboxylic acid.

The salts may be organic or inorganic and need not be pharmaceutically acceptable. Examples of suitable salt-forming groups R^x include inorganic salts, for example alkali metal atoms (e.g. lithium and sodium), other metal atoms, tertiary amino groups (e.g. tri-lower-alkylamino, N-ethylpiperidino, and dimethylpiperazino groups). A preferred salt-forming group R^x is the triethylamino group.

An ester-forming group R^x is advantageously one that can be removed under conventional conditions. Examples of suitable ester-forming groups R^x include benzyl, 3 ,5-di-t-butyl-4-hydroxy-benzyl, benzoylmethyl, p-nitrobenzyl, 4-pyridylmethyl, 2, 2,2-trichloroethyl, 2,2,2-tribromoethyl, allyl, acetonyl, t-butyl, t-amyl, diphenylmethyl, triphenylmethyl, adamantyl, 2-benzyloxyphenyl, 4-methylthiophenyl, tetrahydrofur-2-yl, tetrahydropyran-2-yl, pentachlorophenyl, p-toluene-sulphonylethyl, and methoxymethyl groups, and also silyl, stannyl and phosphorus-containing groups, and oxime radicals of formula -N=CHR° in which R° denotes aryl or heterocyclyl. Furthermore, the ester-forming group R^x may be an in vivo hydrolysable ester group including, in particular, those listed above.

The free carboxyl group may be regenerated from any of the above esters by usual methods appropriate to the particular R^x group, for example, by acid-catalysed, base-catalysed or enzymically-catalysed hydrolysis, or by hydrogenation. The hydrolysis must of course be carried out under conditions in which the groups on the rest of the molecule are stable.

When it is desired to produce a compound of the general formula I in the form of a free acid or in the form of a salt, by a process according to the invention, it is generally advantageous to use a compound in which R^x denotes a carboxyl-blocking group. When it is desired to produce a compound of the general formula I in the form of a pharmaceutically acceptable ester, it is generally convenient to use a compound in which R^x denotes the desired ester group.

The process step according to the invention involves the elimination of the elements of a compound H-X^o from a penem or penem intermediate of the general part-formula XIV, in which X^o denotes a hydroxy group or a leaving group.

In the case where X^o denotes a hydroxy group, the compound of the formula H-X^o being eliminated is water and the elimination reaction is a dehydration reaction, which may suitably be carried out by treating a compound of the general part-formula XIV with a compound of the general formula XVII:

R¹³O₂C-N=N-CO₂R¹⁴ XVII

in which each of R¹³ and R¹⁴, which may be identical or different, denotes aryl, (C_1-6)alkyl or aryl(C_1-6) alkyl,

and with a compound of the general formula XVII:

XVIII

in which

each of a , b and c which may be identical or different, denotes 0 or 1, and

each of R¹⁵, R¹⁶ and R¹⁷, which may be identical or different, denotes aryl, (C_1-6)alkyl or aryl (C_1-6)alkyl.

In the compounds of the general formula VI, R¹³ and R¹⁴ are preferably selected from methyl, ethyl, propyl, butyl, phenyl, and benzyl, the ethyl and isopropyl groups being preferred. Advantageously, R¹³ and R¹⁴ may be identical. A preferred compound of the general formula XVII is diethyl azodicarboxylate. Preferred compounds of the general formula XVIII include triarylphosphines and trialkylphosphites. Preferred groups R¹⁵, R₁₆ and R¹⁷ include methyl, ethyl, n-propyl, n-butyl, benzyl, phenyl and methoxyphenyl. Advantageously, R¹⁵, R¹⁶ and R¹⁷ are all identical. A preferred compound of the general formula XVIII is triphenylphosphine.

Advantageously, approximately two equivalents of each the compounds of the general formulae XVII and XVIII are used per mole of the compound of the general part-formula XIV.

The dehydration reaction may suitably be carried out at a non-extreme temperature, for example a temperature of from -20°C to +100°C. It may be convenient to begin the reaction at a depressed temperature, for example

0°C, and then to allow the temperature to rise to about room temperature.

The reaction may suitably be carried out in an inert aprotic organic solvent. Suitable solvents include tetrahydrofuran, dioxane, ethyl acetate, benzene, and dichloromethane.

In the cases where X^o, in general formula XIV, denotes a leaving group, which will hereinafter be referred to as X⁰¹, it may suitably be a halogen atom or a group of one of the formulae

-O-SO₂-(O)_n-R¹³ XIXA

-O-CO-(O)_n-R¹⁸ XIXB or

-O-PO-(OR¹⁹)₂ XIXC in which

n denotes 0 or 1,

R¹⁸ denotes (C_1-6)alkyl, aryl or aryl (C_1-6)alkyl,

and

R¹⁹ denotes (C_1-6)alkyl or aryl.

Preferred groups of formula XIXB are those in which n denotes zero and R¹⁸ denotes (C_1-6)alkyl, especially the acetoxy group.

The elimination of the elements of a compound of the general formula XIII in which X^o denotes a leaving group χ⁰¹ from a compound of the general formula XIV,may suitably be effected by treating the compound of the general formula XIV with a base in an aprotic medium.

Suitable bases for that purpose include, for example, powdered inorganic bases, for example alkali metal carbonates, bicarbonates, hydroxides, and hydrides (e.g. powdered potassium carbonate), and also organic bases of low nucleophilicity, for example 1,8-diazabicyclo[5.4.0]undec-7-ene. Suitable solvents for use as the aprotic medium in this reaction include, for example, dimethylformamide, hexamethylphosphor- amide, dichloromethane, and tetrahydrofuran.

The elimination may suitably be effected at a low temperature, for example a temperature of from -70°C to +70°C, advantageously from -40°C to 0°C. The compounds of the general formula XIV in which X^o denotes a leaving group X⁰¹ may suitably be prepared from the corresponding compound in which X^o denotes a hydroxy group by replacing the hydroxy group by a leaving group X⁰¹. Alternatively, in the case of the compounds of the general formula XIVD below, the leaving group X⁰¹ may be introduced into the molecule at an earlier stage in the synthesis of the penem nucleus. In particular, a group χ01 of the formula XIXA or XIXB may be introduced at the beginning of, or at any stage during, the synthesis of the penem. In each case, the group X⁰¹ may suitably be introduced by replacing a hydroxyl group in known manner.

The dehydration or other elimination reaction of the process according to the invention may be carried out at any suitable stage during the preparation of the penem of the general formula I or salt or ester thereof, suitably at an early stage or late stage in the manufacturing process.

Further examples of suitable leaving groups will be apparent to those skilled in the art and include sulphoxide, selenoxide and xanthate groups, which can be eliminated by known methods (see W. Carruthers, 'Some modern methods of organic synthesis', Cambridge Univ. Press 1978 (2nd edition), pages 93-103).

In particular, the dehydration or other elimination reaction may be carried out on a compound of the general formula XIVC: XIVC

in which

R¹, R² and X^o are defined as above,

R²⁰ denotes (C_1-6) alkyl, aryl, aryl(C_1-6)alkyl, (C_1-6)alkylthio, arylthio, hetero-aromatic-thio, acyl (for example, (C_1-6)alkylcarbonyl, especially acetyl), (C_2-6)alkenyl (especially vinyl), or aryl(C_1-6)alkenyl, all of which may optionally be substituted, and

R²¹ denotes hydrogen or an N-protecting group,

to give a compound of the general formula XVC:

XVC

in which, R¹, R², R²⁰ and R²¹ are defined as above, which may then subsequently be converted to a penem of the general formula I or salt or ester thereof in known manner, suitably by a conventional penem preparation method.

Alternatively, the dehydration or other elimination reaction may be carried out on a compound of the general formula XIVD:

XIVD

in which R¹, R², R¹², R^x and X^o are defined as above, to give a compound of the general formula XVD:

XVD

in which R¹, R², R¹² and R^x are defined as above, and thereafter, if necessary or desired: (a) removing any carboxyl-blocking group R^x, and/or

(b) converting the group R¹² into a group or atom R³, and/or (c) converting the product into the free acid or into a pharmaceutically acceptable salt or in vivo hydrolysable ester.

The conversion of a compound of the general formula XVC to the desired penem may suitably proceed via a compound of the general formula XVD, according to known penem preparation methods.

A compound of the general formula XIVD, especially one in which X^o denotes a leaving group X⁰¹, may conveniently be prepared from a compound of the general formula XIVC, especially one in which X^o denotes a leaving group X⁰¹, according to known penem preparation methods.

In a compound of the general formula XIVB or XIVC, R¹⁰ or R²⁰, respectively, may suitably denote a triphenylmethyl group. Examples of suitable

N-protecting groups, R¹¹ or R²¹, include silyl groups, for example t-butyldimethylsilyl groups.

In the general formula XIVD and XVD, the group R¹² may be a group convertible into R³ during the penem preparation process. One particular example of such a group, which may conveniently be used in the preparation of a group R³ of the formula -SR⁴ (in which R⁴ is defined as above), is the group of the formula XX: XX

in which R²² denotes an organic radical different from the group R⁴.

A sulphoxide compound of the general formula XIVD or XVD in which R¹² denotes a group of the formula XX may be reacted with a thiol of the general formula XXI:

R⁴- SH XXI

in which R⁴ is defined as above, or a reactive derivative thereof, to give a compound of the general formula XIVD or XVD in which R¹² denotes a group of the formula XXII:

-S-R⁴ XXII

in which R⁴ is defined as above.

The reaction of the sulphoxide with the thiol may be carried out as described in European Patent Publication No. EP 0 046 363A.

A sulphoxide compound of the general formula XIVD or XVD in which R¹² denotes a sulphoxide group of the formula XIX above may be prepared by S-oxidation of a compound of the general formula XIVD or XVD, respectively, in which R¹² denotes a group of the formula -S-R²². The S-oxidation may be effected using a mild oxidising agent, for example a perbenzoic acid, hydrogen peroxide, selenium dioxide or sodium metaperiodate. Perbenzoic acids, for example m-chloroperbenzoic acid, are preferred.

The present invention also provides a process for the preparation of a compound of the general formula I in which R³ denotes a group of the formula -SR⁴ (in which R⁴ is defined as above), or a pharmaceutically acceptable salt or in vivo hydrolysable ester thereof, which comprises reacting a compound of the general formula XXIII:

XXIII

in which

R¹, R², R²² and R^x are defined as above,

with a thiol of the general formula XXI above, or a reactive derivative thereof;

and thereafter if necessary or desired:

(a) removing any carboxyl-blocking group R^x,

and/or (b) converting the product into the free acid or into a pharmaceutically acceptable salt or in vivo hydrolysable ester.

The compounds of the general formulae XIVC and XVC are novel intermediates and also constitute subjects of the present invention. The compounds of the general formula XIVD, in particular those in which R¹² denotes R³ or a sulphoxide group of the formula XIX, are also novel intermediates and form a further subject of the present invention. The compounds of the general formula XVD in which R¹² denotes a sulphoxide group of the formula XIX are further novel intermediates and constitute a yet further subject of the present invention.

The compounds according to the invention have β-lactamase inhibitory and antibacterial properties, and are useful for the treatment of infections in animals, especially mammals, including humans, in particular in humans and domesticated (including farm) animals. The compounds may be used, for example, for the treatment of infections of, inter alia, the respiratory tract, the urinary tract, and soft tissues, especially in humans.

The compounds may be used for the treatment of infections caused by strains of, for example, Staphylococcus aureus, Klebsiella aerogenes, Escherichia coli, Proteus sp., and Bacteroides fragilis. It is generally advantageous to use a compound according to the invention in admixture or conjunction with a penicillin, cephalosporin or other β-lactam antibiotic and that can often result in a synergistic effect, because of the β-lactamase inhibitory properties of the compounds according to the invention. In such cases, the compound according to the invention and the other β-lactam antibiotic can be administered separately or in the form of a single composition containing both active ingredients as discussed in more detail below.

The compounds according to the invention are suitably provided in substantially pure form, for example at least 50% pure, advantageously at least 75% pure, preferably at least 95% pure, especially at least 98% pure, all percentages being calculated as weight/weight. An impure or less pure form of a compound according to the invention may, for example, be used in the preparation of a more pure form of the same compound or of a related compound (for example, a corresponding salt, ester or free acid) suitable for pharmaceutical use. Although the purity of any compound used as an intermediate may be less critical than that of a compound used as a final product, for example one used directly for pharmaceutical use (for example in a composition according to the invention as described below), nevertheless such an intermediate compound is advantageously provided in substantially pure form. It is generally advantageous to provide the compounds according to the invention in crystalline form. The free acids and salts according to the invention may be in hydrated or non-hydrated form.

The present invention provides a pharmaceutical composition comprising a compound according to the invention that is to say, a compound of the general formula I or a pharmaceutically acceptable salt or in vivo hydrolysable ester thereof, and a pharmaceutically acceptable carrier. The present invention also provides a method of treating bacterial infections in animals, especially in humans and in domesticated mammals (including farm mammals), which comprises administering a compound or composition according to the invention to the animal. Such administration may advantageously be effected in conjunction with the prior, simultaneous pr subsequent administration of a penicillin, cephalosporin or other β-lactam antibiotic.

The compositions of the invention may be in a form adapted for oral, topical or parenteral use and may be used for the treatment of infection in animals especially mammals, including humans, in particular in humans and domesticated animals (including farm animals).

The compositions of the invention may, for example, be made up in the form of tablets, capsules, creams, syrups, suspensions, solutions, reconstitutable powders, and sterile forms suitable for injection or infusion. Such compositions may contain conventional pharmaceutically acceptable materials, for example diluents, binders, colours, flavours, preservatives, and disintegrants, in accordance with conventional pharmaceutical practice in manner well understood by those skilled in the art of formulating antibiotics.

It can be particularly advantageous for the compounds according to the invention to be administered to a patient by injection or infusion. That method of administration has the advantage of rapidly resulting in high blood levels of the active ingredient compound being administered. Accordingly, in one preferred form of the composition according to the invention, a compound according to the invention is present in sterile form, including in sterile crystalline form. A further preferred form of the composition according to the invention, is one in which the composition is in injectable or infusable form.

One injectable or infusable form of the composition according to the invention is an injectable or infusable solution, which suitably comprises a solution of a compound according to the invention in a sterile pyrogen-free liquid, for example water or aqueous ethanol.

A further injectable or infusable form of the composition according to the invention is an injectable or infusable suspension, in which case the compound according to the invention is advantageously present in finely particulate form. The suspension may be an aqueous suspension in, for example, sterile water or sterile saline, which may additionally include a suspending agent, for example polyvinylpyrrolidone. Alternatively, the suspension may be an oily suspension in a pharmaceutically acceptable oil suspending agent, for example arachis oil.

A composition according to the invention may be in unit dosage form, for example unit dosage form for oral administration, topical administration, or parenteral administration (including administration by injection or infusion).

A composition according to the invention may comprise a compound according to the invention as the sole active ingredient or therapeutic agent, or it may also comprise one or more additional active ingredients or therapeutic agents, for example a penicillin, cephalosporin or other β-lactam antibiotic, or pro-drug thereof. A composition comprising a compound according to the invention and another active ingredient or therapeutic agent, especially a penicillin, cephalosporin or other β-lactam antibiotic, or pro-drug thereof, can show enhanced effectiveness, and in particular can show a synergistic effect.

Penicillins, cephalosporins and other β-lactam antibiotics suitable for co-administration with the compounds according to the invention - whether by separate administration or by inclusion in the compositions according to the invention - include both those known to show instability to or to be otherwise susceptible to β-lactamases and also those known to have a degree of resistance to β-lactamases.

Examples of penicillins suitable for co-administration with the compounds according to the invention include benzylpenicillin, phenoxymethylpenicillin, carbenicillin, azidocillin, propicillin, ampicillin, amoxycillin, epicillin, ticarcillin, cyclacillin, pirbenicillin, azlocillin, mezlocillin, sulbenicillin, piperacillin, and other known penicillins. The penicillins may be used in the form of pro-drugs thereof, for example as in vivo hydrolysable esters, for example the acetoxymethyl, pivaloyloxymethyl, α-ethoxycarbonyloxyethyl and phthalidyl esters of ampicillin, benzylpenicillin and amoxycillin; as aldehyde or ketone adducts of penicillins containing a 6α-aminoacetamido side chain (for example hetacillin, metampicillin and analogous derivatives of amoxycillin); and as α-esters of carbenicillin and ticarcillin, for example the phenyl and indanyl α-esters. Examples of cephalosporins that may be co-administered with the compounds according to the invention include, cefatrizine, cephaloridine, cephalothin, cefazolin, cephalexin, cephacetrile, cephapirin, cephamandole nafate, cephradine, 4-hydroxycephalexin, cephaloglycin, cefoperazone, cefsulodin, ceftazidime, cefuroxime, cefmetazole, cefotaxime, ceftriaxone, and other known cephalosporins, all of which may be used in the form of pro-drugs thereof.

Examples of β-lactam antibiotics other than penicillins and cephalosporins that may be co-administered with the compounds according to the invention include aztreonam, latamoxef (Moxalactam - Trade Mark), and other known β-lactam antibiotics, all of which may be used in the form of pro-drugs thereof.

In the compositions according to the invention, the compound according to the invention and the penicillin, cephalosporin or other β-lactam antibiotic may be linked by means of an in vivo hydrolysable ester group, in the form of a mutual pro-drug.

Some penicillins and cephalosporins that may be included in the compositions according to the invention may not be suitable for oral administration, in which case the composition will be in a form suitable for parenteral or topical administration.

Particularly suitable penicillins for co-administration with the compounds according to the invention include ampicillin, amoxycillin, carbenicillin, piperacillin, azlσcillin, mezlocillin, and ticarcillin. Such penicillins may be used in the form of their pharmaceutically acceptable salts, for example their sodium salts. Alternatively, ampicillin or amoxycillin may be used in the form of fine particles of the zwitterionic form (generally as ampicillin trihydrate or amoxycillin trihydrate) for use in an injectable or infusable suspension, for example, in the manner hereinbefore described in relation to the compounds according to the invention. Amoxycillin, for example in the form of its sodium salt or the trihydrate, is particularly preferred for use in synergistic compositions according to the invention.

Particularly suitable cephalosporins for co-administration with the compounds according to the invention include cephaloridine, cefoperazone and cefazolin, which may be used in the form of their pharmaceutically acceptable salts, for example their sodium salts.

A compound according to the invention may be administered to the. patient in an antibacterially effective amount or, when a compound according to the invention is being used in conjunction with a penicillin, cephalosporin, or other β-lactam antibiotic, it may be used in a synergistically effective amount.

The compounds according to the invention may suitably be administered to the patient at a daily dosage of from 0.7 to 50 mg/kg of body weight. For an adult human (of approximately 70 kg body weight), from 50 to 3000 mg, preferably from 100 to 1000 mg, of a compound according to the invention may be adminstered daily, suitably in from 1 to 6, preferably from 2 to 4, separate doses. Higher or lower dosages may, however, be used in accordance with clinical practice. When the compositions according to the invention are presented in unit dosage form, each unit dose may suitably comprise from 25 to 1000 mg, preferably from 50 to 500 mg, of a compound according to the invention. Each unit dose may, for example, be 62.5, 100, 125, 150, 200 or 250 mg of a compound according to the invention.

When the compounds according to the invention are co-administered with a penicillin, cephalosporin or other β-lactam antibiotic, the ratio of the amount of the compound according to the invention to the amount of the other β-lactam antibiotic may vary within a wide range. The said ratio may, for example, be from 100:1 to 1:100; more particularly, it may, for example, be from 2:1 to 1:30.

The amount of penicillin or cephalosporin or other β-lactam antibiotic in a synergistic composition according to the invention will normally be approximately similar to the amount in which it is conventionally used per se, for example from about 50 mg, advantageously from about 62.5 mg, to about 3000 mg per unit dose, more usually about 125, 250, 500 or 1000 mg per unit dose.

An example of a particularly suitable composition according to the invention is one comprising from 150 to 1000 mg, preferably from 200 to 700 mg, of amoxycillin or ampicillin or a pro-drug thereof, in admixture or conjunction with from 5 to 500 mg, preferably from 20 to 250 mg, of a compound according to the invention, per unit dose. In such a composition, the amoxycillin may suitably be in the form of its trihydrate or sodium salt; the ampicillin may suitably be in the form of ampicillin trihydrate, ampicillin anhydrate, sodium ampicillin, hetacillin, pivampicillin hydrochloride, bacampicillin hydrochloride or talampicillin hydrochloride; and the compound according to the invention may most suitably be in crystalline form. Such composition may be in a form suitable for oral or parenteral use, except when it comprises an in vivo hydrolysable ester of ampicillin or amoxycillin in which case the composition should not normally be intended for parenteral administration.

A further example of a particularly suitable composition according to the invention is one comprising from 200 to 2000 mg of carbenicillin or ticarcillin or a pro-drug thereof, in admixture or conjunction with from 5 to 500 mg, preferably from 25 to 250 mg, of a compound according to the invention, per unit dose. In such a composition, the carbenicillin or ticarcillin may most suitably be in the form of its di-sodium salt, and the compound according to the invention may most suitably be in crystaline form. When the composition contains the carbenicillin or ticarcillin in the form of a di-salt, it is most suitably presented in a form suitable for parenteral administration.

No toxicological effects are indicated when the compounds according to the invention are administered within the above-mentioned dosage ranges.

The following examples illustrate the invention. Unless otherwise stated, all temperatures are given in degrees Celsius and all percentages are calculated by weight. The term 'Biogel' is a Trade Mark. Preparation 1(a)

1-t-Butyldimethylsilyl-3-(benzoCb]furyl-2-carbonyl)-4- tritylthioazetidin-2-one

A solution of n-butyllithium (1.73M in hexane; 0.69 ml) was added to a solution of diisopropylamine (0.17ml) in dry tetrahydrofuran (THF) (5 ml) at -30°C under dry argon. After stirring at -30°C for 0.25 h, the mixture was cooled to -76°C and treated with a solution of l-t-butyldimethylsilyl-4-tritylthioazetidin-2-one (1) (0.459g) (GB 2042515A, Bristol-Myers) in dry THF (3 ml). After stirring at -76°C for a further 0.25 h the mixture was treated with a solution of ethyl-2- benzo[b]furoate (0.342g) (B.G. Fuston et al., Organic Synthesis Coll., Vol 3, page 209) in dry THF (1 ml). The reaction mixture was stirred at -76°C for 0.5 h then treated with saturated ammonium chloride solution (5 ml) and allowed to warm to room temperature. Ethyl acetate (50 ml) was added and the organic phase washed with saturated brine (4x); Evaporation of the dried (MgSO₄) organic phase produced a yellow oil, which was subjected to silica-gel column σhromatography using ethyl acetate:hexane (1:3) as eluent, to yield the title azetidinone (2) (0.393g) as a white crystalline solid, mp 176-178°C (ex ethyl acetate/hexane); ν_max (CHCl₃) 1750 and 1665 cm^-1; δppm (CDCI₃) 0.97 (9H, s); 4.21 (1H, d, J2Hz); 4.98 (1H, d, J2Hz); 6.85-7.70 (20H, m); both MeSi signals were obscured by the TMS signal. (Found C, 73.70; H, 6.12; N, 2.33; S, 5.20. C₃₇N₃₇NO₃SSi requires C, 73.60; H, 6.17; N, 2.32; S, 5.31%). Preparation 1(b)

1-t-Butyldimethylsilyl-3-Chydroxy(benzo[b]fur-2-yl) methyl]-4-tritylthioazetidin-2-one

The ketone (2) from Preparation 1(a) (2.77g) was dissolved in tetrahydrofuran (THF) :ethanol (1:1;

130 ml) and cooled in an ice bath. Sodium borohydride was added portionwise, and the mixture examined by tic after each successive addition, until' all of the starting material had been consumed (total quantity of sodium borohydride added 1.5g). Ethyl acetate (300 ml) and saturated brine (150 ml) were added and the mixture carefully neutralised with dilute citric acid. The aqueous phase was separated and washed with ethyl acetate (3 x 100 ml). The combined organic phases were washed with saturated brine (1 x 200 ml) and dried (MgSO₄). Evaporation in vacuo produced a yellow oil which was subjected.to silica-gel column chromatography using ethyl acetate:hexane (1:3) as eluent, to yield two isomeric alcohols (3): Isomer I (0.78g), Rf 0.78 (ethyl acetate : hexane; 1:2); a white crystalline solid; mp 99-101°C (ex ethyl acetate/hexane); v_max (CHCl₃) 3600-3100 and 1735 cm^-1; δppm (CDCI₃) 0.91 (9H, s); 1.74 (1H, d, J6Hz); 3.62 (1H, dd, J2 and 2.5Hz); 3.83 (1H, dd, J6 and 2.5Hz); 4.50 (1H,d , J2Hz); 6.68 (1H, broad s); 7.0-7.6 (19H, m); the Me₂Si signals were obscured by the TMS signal; (Found (diethylamine chemical ionisation) M+ Et₂NH+, 679). Isomer II (1.97g), Rf 0.41 (ethyl acetate:hexane; 1.2); a white crystalline solid; mp 159-161°C (ex ethyl acetate/ hexane); ν_max (CHCI₃) 0.73 (9H, s) ; 2.65-2.95 (1H, m); 3.78 (1H, dd, J2 and 6Hz); 4.0-4.16 (1H, m); 4.19 (1H, d, J2Hz); 6.62 (1H, s); 7.0-7.60 (19H, m) the Me₂Si signals were obscured by the TMS signal; (Found C, 73.31; H, 6.44; N, 2.41; S, 5.01; C₃7H₃9NO₃SSi requires C, 73.35; H, 6.49; N, 2.31; S, 5.29%). Preparation 1(c)

(3RS, 4SR) 3-CHydroxy(benzo[b]fur-2-yl)methyl]-4- tritylthioazetidin-2-one

The azetidinone-alcohol (3) (isomer II) from Preparation 1(b) (0.156g) was dissolved in methanol: dichloromethane (1:1; 10 ml). The solution was cooled to -20°C and potassium fluoride (0.016g) added. After stirring at room temperature for 0.5 h, tic showed that all of the starting material had been consumed. The reaction mixture was evaporated to a yellow solid which was redissolved in dichloromethane (20 ml). The organic phase was washed with saturated brine (3x) and dried (MgSO₄). Removal of the solvent in vacuo yielded a white solid which was triturated with ether and crystallised from chloroform/ether to give the title compound (4) (0.103g) as white needles; mp 218-220°C decomp; ν_max (Nujol) 3350; 1745 and 1465 cm^-1; δppm ((CD₃)₂CO) 3.74 (1H, dd, J2.7Hz); 4.62 (1H, d, J2.7Hz); 5.21-5.25 (1H, m); 5.28 (1H, broad s); 5.33 (1H, d, J5.5Hz); 6.82-6.90 (1H, m); 7.21-7.37 and 7.56-7.59 and 7.7-7.77 (19H, 3xm) , addition of D₂O caused signals 5.21-5.25 to collapse to 5.21 (d, J1.8Hz); (Found C, 75.87; H, 5.21; N, 2.83; S, 6.29; C₃₁H₂₅NO₃₈ requires C 75.74; H, 5.12; N, 2.85; S, 6.52%).

Preparation 1(d)

(3RS, 4SR) 3-[Acetoxy(benzo[b]fur-2-yl)methyl]-4- tritylthioazetidin-2-one

The azetidinone (4) from Preparation 1(c) (1.22g) was dissolved in dry dichloromethane (80 ml). The solution was cooled in an ice bath and 4-dimethylaminopyridine (0.27g) and triethylamine (0.4 ml) added. After stirring at ice bath temperature for 5 minutes, acetic anhydride (0.3 ml) was added. The mixture was stirred at ice bath temperature for 0.5 h, then washed with saturated brine (100 ml); dilute citric acid solution (100 ml); saturated brine (100 ml); saturated sodium bicarbonate solution (100 ml) and saturated brine (3 x 100 ml). Evaporation of the dried organic phase (MgSθ4) gave a white foam which was subjected to silica-gel column chromatography using ethyl acetate: hexane (1:1) as eluent to yield the title compound (5) as white needles (1.28g); mp 180-182°C; ν_max (CHCI₃) 3390, 1770 cm^-1; δppm (CDCI₃) 2.04 (3H, s); 3.76 (1H, dd, J3 and 5Hz); 4.22 (1H, broad s); 4.57 (1H, d, J3Hz) 6.26 (1H, d, J5Hz); 6.72 (1H, s); 7.05-7.75 (19H, m); (Found C, 74.54; H, 5.12; N, 2.65; S, 5.75; C₃₃H₂₇NO₄S requires C, 74.27; H, 5.10; N, 2.62; S, 6.01%).

Preparation 1(e)

(3RS, 4SR) 3-(Acetoxy(benzo[b]fur-2-yl)methyl)-1-(1-p- nitrobenzyloxycarbonyl-1-triphenylphosρhoranylidene- methyl)-4-tritylthioazetidin-2-one

The azetidinone (5) (0.272g) from Preparation -1(d) and p-nitrobenzylglyoxylate monohydrate (0.128g) were heated in refluxing benzene (18 ml) with provision for azeotropic removal of water (Dean and Stark apparatus containing molecular sieves 4A) for 1 hour. The mixture was cooled to room temperature and treated with triethylamine (5.2 mg). After stirring at room temperature for 2.25 h the reaction mixture was evaporated to yield the crude hydroxy ester (6); ν_max (CHCI₃) 3500, 1770 and 1750 cm^-1. A solution of the crude hydroxy ester (6) in dry tetrahydrofuran (5 ml) was cooled to -10°C and treated with 2,6-lutidine (0.085 ml) and thionyl chloride (0.06 ml). After stirring at -10°C for 0.25 h the mixture was filtered and evaporated to give the crude chloroester (7) as a , solid: ν_max (CHCI₃) 1785 and 1760 (sh) cm^-1. The crude chloroester (7) and triphenylphosphine (0.53g) were dissolved in dry dioxan (3 ml). The resulting solution was concentrated to approximately 1 ml and treated with 2,6-lutidine (0.07 ml). The resulting mixture was stirred at 50°C for 28 h then at room temperature for 48 h. After addition of ethyl acetate (50 ml) the organic phase was washed with saturated brine; 1N hydrochloric acid; saturated brine; saturated sodium bicarbonate solution; saturated brine (3x) and dried (MgSO₄). Evaporation produced a yellow foam, which was subjected to silica-gel column chromatography using ethyl acetate:hexane (1:2) as eluent, to give the title phosphorane (8) as a white crystalline solid (0.25g); mp 260-262°C (decomp.) (dichloromethane/hexane); ν_max (CHCl₃) 1750; 1620 and 1605 cm^-1.

Preparation 1(f)

Silver (3RS, 4SR) 3-CAcetoxy(benzo[b]fur-2-yl)methyl] -1-(1-p-nitrobenzyloxycarbonyl-1-triphenylphosphoran- ylidenemethyl)-azetidin-2-one 4-thiolate

Silver nitrate (2.55 ml of a 0.15 molar solution in methanol) was added to a stirred mixture of the phosphorane (8) (0.25g) from Preparation 1(e) and pyridine (0.031 ml) in methanol: dichloromethane (1:1) (10 ml) at room temperature. After 30 min, at room temperature the mixture was cooled in an ice bath for 10 min and filtered. The residue was washed with cold methanol (5 ml) and dry cold ether (2 x 10 ml) and dried in vacuum to give the title silver thiolate (9) (0.19g) ν_max (Nujol) 1750 and 1600 cm^-1. Preparation 1(g) p-Nitrobenzyl (5RS, 6SR)-6-CAcetoxy(2-benzo[b]furyl) methyl]penem-3-carboxylate

To a stirred suspension of the silver thiolate (9) (0.19g) from Preparation 1(f) in dry dichloromethane cooled in an ice bath was added formic acetic anhydride (0.18 ml) and 4-dimethylaminopyridine (0.025g). The mixture was stirred at 0°C for 5 min then treated with triethylamine hydrochloride (0.31g). The ice bath was removed and the mixture stirred for a further 1.75 h at room temperature. The mixture was diluted with ethyl acetate (50 ml) and filtered through celite, the residue being washed with ethyl acetate. The combined filtrates were washed with 5% citric acid solution (2x) saturated brine, saturated sodium bicarbonate (3x); saturated brine (3x). The dried (MgS04) organic phase was heated under argon at 50°C for 0.5 h, then evaporated to give a yellow oil, which was subjected to silica-gel column chromatography using ethyl acetate: hexane (1:1) as eluent to give the title penem (10) as an amorphous solid (0.078g); ν_max (CHCI₃) 1800; 1745, and 1720 cm^-1; δppm (CDCI₃) 2.13 (3H, s); 4.45 (1H, ddd, J4.2, 2.0 and 1.1Hz); 5.28 and 5.42 (2H, ABq); 6.15 (1H, d, J2Hz); 6.40 (1H, d, J4.2Hz); 6.83 (1H, s); 7.34 (1H, d, Jl.lHz); 7.23 to 7.33 and 7.36 to 7.48 (5H, m); 7.58 and 8.23 (2H, 2d, J8.8Hz). λ_max (EtOH) 248 nm ( ε_max 24,311); 283 nm (ε_max 11,659); 317 nm (ε_max -3809); (Found M+ 434, acetic acid eliminated).

Example 1(a) p-Nitrobenzyl (5RS) (Z) 6-(2-Benzo[b]furylmethylene) penem-3-carboxylate

A solution of 1, 8-diazabicycloC5.4.0]undec-7-ene (23.7 mg) in dry dichloromethane (0.5 ml) was added to a stirred solution of the penem (10) (0.051g) from Preparation 1(g) in dry dichloromethane (5 ml) at -40°C. The mixture was stirred at -40°C until thin layer chromatography showed that all of the starting material had been consumed. Dilute citric acid solution (10 ml) and dry dichloromethane (20 ml) were added and the organic layer separated and further washed with saturated brine; dilute citric acid solution; saturated brine; saturated sodium bicarbonate solution and saturated brine (3x). The dried (MgSO₄) organic phase was evaporated and the residue subjected to silica gel column chromatography using dichloromethane as eluent to give the title penem (11) (0.041g) as a yellow solid; ν_max (CHCI₃) 1780; 1710; 1665 and 1600 cm^-1; δppm (CDCI₃) 5.31 and 5.47 (2H,

ABq, J13.6Hz); 6.75 (1H, d, J0.8Hz); 7.08 (2H, s); 7.23 to 7.55 (5H, m); 7.63 (2H, d, J8.2Hz); 8.25 (2H, d, J8.2Hz); λ_max (EtOH) 261 nm (ε_max) 14,148); 329 nm (ε_max 35,948); Found M+ C₂₂H₁₄N₂O₆S, 434).

Example 1(b)

Sodium (5RS) ( Z)-6-(2-Benzo[b] furylmethylene) penem-3- carboxylate

The penem ester (11) (0.02g) from Example 1(a) was dissolved in a mixture of dioxan (8 ml) and water (2 ml). 5% Palladium on carbon (30 mg) was added and the mixture hydrogenated at room temperature for 20 min. A 1% w/v sodium bicarbonate solution (0.37 ml) was added and the mixture filtered through celite, the residue was washed with water. The combined filtrates were evaporated to a gum which was chromatographed on Biogel P2 using water as eluent. The appropriate fractions, as determined by ultra-rviolet spectroscopy, were combined and freeze dried to give the title sodium salt (12) (5.2 mg) as a yellow/orange solid; λ_max (H₂O) 334.1 nm (ε_max 25,014); δppm (D₂O), 6.74 (1H, s); 7.05 (1H, s); 7.15 (1H, s); 7.23 (1H, s); 7.32 (1H, t, J7.5Hz); 7.45 (1H, t, J7.5Hz); 7.59 (1H, d, J7.5Hz); 7.70 (1H, d, J7.5Hz).

Preparation 2

N-(4-Methoxyphenylsulphonyl)-2-methoxycarbonylindole

2-Methoxycarbonylindole (3.14g; 23 mmol) in glyme (80 ml) was added to a slurry of sodium hydride (lg; 80%) in glyme (30 ml) at room temperature. The reaction

mixture was stirred for 1.5 h then cooled to -5°C and 4.7g (1 equivalent) of 4-methoxyphenylsulphonylchloride in glyme (15 ml) added to the pink anion solution, followed by imidazole (75 mg) . The reaction was stirred for 1 hour, then poured into water and extracted with chloroform. The crude product was chromatographed on silica, eluting with ethyl acetate: cyclohexane 1:7. Recovered starting material (lg) and 3.25g (52%) of the desired title compound (13) were isolated from the column. ν(film) 1740, 1595, 1580 cm^-1, δ(CDCl₃) 3.67-3.90 (6H, m), 6.80-8.10 (9H, m); m/e 345 (M+).

Preparation 2(a)

1-t-Butyldimethylsilyl-3-(1-4'-methoxyphenylsulphonyl- indole-2-carbonyl)-4-tritylthioazetidin-2-one

Diisopropylamine (1.02 ml; 7.29 mmol) in tetrahydrofuran (20 ml) at -30°C under argon was treated with ri-butyl lithium (4.3 ml of a 1.7M solution in n-hexane) and stirred for 15 minutes. After further cooling to -78°C the stirred mixture was treated dropwise with a solution of 1-t-butyldimethylsilyl-4-tritylthio- azetidin-2-one (1) (3.3g; 7.19 mmol) in tetrahydrofuran (15 ml). After stirring at -78°C for 20 minutes the mixture was treated with N-(4-methoxyphenylsulphonyl) -2-methoxycarbonylindole (2.48 g) (13) in tetrahydrofuran (10 ml) and stirred for 30 minutes. The reaction mixture was treated with saturated ammonium chloride (10 ml) and extracted with ethyl acetate (200 ml). The organic phase was washed several times with saturated brine, dried (anhydrous magnesium sulphate) and evaporated to a foam. This crude product was chromatographed on silica eluting with ethyl acetate:cyclohexane, 1:4.

Fractions containing material Rf (SiO₂/ethyl acetate: cyclohexane, 1:3) 0.45 were collected and evaporated to afford compound (14), 3.0g (54%) as a cream-coloured foam, ν_max(film) 1760, 1682, 1600 cm^-1; δ(CDCl₃) 0.32

(3H, s), 0.33 (3H, s), 0.94 (9H, s), 3.81 (3H, s), 4.15 (1H, d, J 1.8Hz), 4.9 (1H, d, J1.8Hz), 6.85-8.25 (24H, broad multiple peaks) m/e (MH+) 773.

Preparation 2(b) 1-t-Butyldimethylsilyl-3-(hydroxy-(1-4'-methoxyphenyl sulphonylindol-2-yl)methyl)-4-tritylthioazetidin-2-one

The ketone (14) from Preparation 2(a) (3g; 3.89 mmol) was dissolved in dimethoxyethane (25 ml) at 0°C under argon and treated with sodium borohydride (310 mg initially) in portions until t.l.c. showed no starting material present. The mixture was treated with citric acid until effervescence ceased, then extracted with ethyl acetate (200 ml). The extracts were washed several times with brine, dried (anhydrous magnesium sulphate) and evaporated to an oil. This crude mixture was chromatographed on silica, eluting with ethyl acetate:cyclohexane (1:2). Two isomeric alcohols (15) were isolated as colourless crystalline solids from methanol. Isomer I 0.79g (26%) Rf (SiO₂/ethyl acetate:cyclohexane, 1:2) = 0.38. ν(Nujol) 3700-3100 (br) 1730, 1595 cm^-1; ν_max (KBr) 3423 (br), 1748, 1728, 1593, 1575 cm^-1; FAB m/e 775 (MH+); δ(CDCl₃) 0.02 (3H, s), 0.05 (3H, s) , 0.83 (9H, s), 2.23 (1H, d, J 7.5Hz), 3.75 (3H, s), 4.17 (1H, dd, J 7.5 Hz and 1.5Hz), 4.27 (1H, d, J 1.5Hz), 5.29 (1H, dd, J 7.5 and 7.5Hz), 6.55 (1H, s), 6.78 (2H, d, J13Hz), 7.0-8.1 (19H,m), 7.66 (2H, d, J13Hz).

Isomer II. 0.62 g (20%) Rf (SiO₂/ethyl acetate:cyclo- hexane, 1:2) = 0.58 v (Nujol) 3570 (sh), 1748 cm^-1, v (KBr) 3570, 1749 cm^-1; m/e 775 (MH⁺); δ (CDCI₃) 0.37 (6H, s), 1.01 (9H, s), 3.05 (1H, dd, J 6 and 1 Hz, exchanges with D₂O), 3.62 (1H, br, d, J 1.3 collapses to. broad s with D₂O. 3.75 (3H, s), 4.22 (1H, dd, J 6 and 1.6 Hz, collapses to d J 1.6 Hz with D₂O), 4.45 (1H, d, J 1.8 Hz) 6.77.(2H, d, J 9 Hz), 7.05 (1H, s), 7.1 - 7.8 (19H, m), 7.61 (2H, d, 9Hz).

Preparation 2(c)

(3RS,4SR)-3-[Hydroxy(1-4'-methoxyphenylsulphonylindol- 2r-yl)methyl]-4-tritylthioazetidin-2-one

The more polar alcohol (15) (Isomer I) from Preparation 2(b) (0.79 g; 1.02 mmol) in methanol (15 ml) and methylene dichloride (10 ml) at 10°C was treated with potassium fluoride (85 mg). The reaction was complete in 15 minutes when the reaction mixture was diluted with methylene dichloride (100 ml) and washed several times with saturated brine, dried (anhydrous magnesium sulphate) and evaporated to a foam. Methanol was added and the title compound (16) crystallised. The crystals were filtered off and washed with cold methanol; drying afforded 0.6 g (89%) of a white crystalline solid (16). Rf (SiO₂/ethyl acetate:hexane, 2:1) = 0.68. ν (Nujol) 3700 - 3125 with sharp peak at 3380, 1772 (shoulder 1760) cm^-1, ν (KBr) 3387 (v. br), 1761 cm^-1; m/e 661 (MH⁺) .

Preparation 2(d)

(3RS,4SR)-3-[Acetoxy-(1-4'-methoxyphenylsulphonyl- indol-2-yl)methyl]-4-tritylthioazetidin-2-one

The alcohol (16) from Preparation 2(c) (560 mg, 0.85 mmol) was dissolved in dichloromethane (5 ml) at 0°C under argon, then treated with acetic anhydride (82 μl), triethylamine (130 μl) and 4-dimethylaminopyridine (9 mg). After 1 hour a further 10 μl acetic anhydride was added and after 2 hours a further 100 μl of triethylamine. The reaction mixture was poured into ethyl acetate (100 ml) and washed with dilute citric acid, brine, sodium bicarbonate and brine. Drying (MgSO₄) followed by evaporation afforded an oil. This oil was chromatographed on silica, eluting with ethyl acetate:cyclohexane 1:1 grading to 2:1. The title compound (17) was obtained as a white crystalline solid, 350 mg, (59%) Rf (SiO₂/ethyl acetate: cyclohexane, 2:1) = 0.5. Recovered alcohol (162 mg) was re-reacted in dichloromethane (3 ml) with acetic anhydride (25 μl), triethylamine (40 μl) and 4-dimethylaminopyridine (3 mg). After 25 minutes t.l.c. showed no starting material present. The reaction mixture was worked up as described above and chromatographed to afford another 70 mg (11%) of the title compound (17) as a white crystalline solid. Total yield 420 mg (70%); ν (KBr) 3400 (sharp), 1778, 1753, cm^-1; δ (CDCl₃) 2.10 (3H, s), 3.77 (3H, s), 4.09 (1H, s), 4,11 - 4.18 (1H, m) , 4.52 (1H, d, J 2.8 Hz), 6.62 (1H, s), 6.87 (2H, dAB, J 9Hz) , 6.91 (1H, partially obscured), 7.15 - 8.28 (19H, m) , 7.94 (2H, dAB, 9Hz); m/e FAB (Thioglycerol/ 3-NOBA/NaAc) 725 (MNa)⁺. Preparation 2(e)

(3RS ,4SR)-3-CAcetoxy-(1-4'-methoxyphenylsulphonylindol- 2-yl)methyl]-1-(1-4'-nitrobenzyloxycarbonyl-1-tri- phenylphosphoranylidenemethyl)-4-tritylthioazetidin-2- one

The acetate (17) from Preparation 2(d) (400 mg, 0.57 mmol) in benzene (25 ml) was refluxed with 4-nitrobenzylglyoxylate (520 mg) whilst removing the water azeotropically using a Dean and Stark. When the collection of water ceased the reaction was allowed to cool before 40 μl of triethylamine was added. After 30 minutes the solvent was evaporated to afford a white foam ν (film) 3450 (broad), 1755 (broad) cm^-1.

This foam was redissolved in tetrahydrofuran (15 ml) under argon at -20°C and treated with 2,6-lutidine (0.23 ml) and thionyl chloride (0.15 ml). After 1 hour the reaction mixture was diluted with toluene and filtered through celite. The filtrate was evaporated, redissolved in toluene and re-evaporated to afford a foam. v (film) 1770 cm^-1.

This foam was dissolved in dichloromethane and dioxane, 1:1 (5 ml), treated with triphenylphosphine (1.4 g) and concentrated to low volume. 2,6-Lutidine (0.27 ml) was added and the reaction kept warm (40°C) for several hours before being stirred overnight. After 30 hours 2,6-lutidine (80 μl) was added and the reaction again warmed to 40°C for several hours. The reaction mixture was diluted with ethyl acetate (100 ml) and washed with dilute citric acid, brine, sodium bicarbonate and brine. The organic phase was dried (anhydrous magnesium sulphate) and evaporated to an oil. Chromatography on silica-gel, eluting with ethyl acetate/hexane 1:1 grading to 2:1 afforded the title compound (18), as an oil, 0.23 g (35%). Rf (Siθ2/ethyl acetate- hexane, 2:1) ca. 0.4; v (film) 1760, 1660, 1620, 1600 cm^-1; m/e FAB ( thioglycerol) (MH⁺) 1156.

Preparation 2(f)

Silver (3RS,4SR) 3-[Acetoxy-(1-4'-methoxyphenyl- sulphonylindol-2-yl)methyl]-1-(1-4'-nitrobenzyloxy- carbonyl-1-triphenylphosphoranylidenemethyl)azetidin-2- one-4-thiolate

A methanolic solution of silver nitrate (0.15 M,

1.7 ml, 1.3 eq) was added with stirring to a solution of the phosphoranylidene intermediate (18) from Preparation 2(e) (0.23 g, 0.2 mmol) and pyridine

(18 μl) in methanol (3 ml) and dichloromethane (3 ml), under argon at 20°C. After 20 minutes a further 10 μl pyridine and 0.2 ml of silver nitrate solution were added. After a further 15 minutes the reaction was evaporated to dryness to afford the silver thiolate (19), v (film) 1750, 1600 cm^-1.

Preparation 2(g)

4-Nitrobenzyl (5RS,6SR) 6-[Acetoxy-(1-4'-methoxyphenyl- sulphonylindol-2-yl)methyl]penem-3-carboxylate

The silver thiolate (19) from Preparation 2(f) (0.2 mmol) was dissolved in dichloromethane (3 ml) at 0°C and treated with formic acetic anhydride (10 eq.; 0.18 ml) and dimethylaminopyridine (25 mg) and then, with vigorous stirring of the reaction, triethylamine hydrochloride (280 mg) was added in one portion. [The triethylamine hydrochloride had been previously ground and dried under vacuum]. Stirring was continued until a grey precipitate formed (approximately 30 minutes) when the reaction was diluted with ethyl acetate (50 ml) and the solids filtered off. The ethyl acetate was washed with dilute citric acid, brine, soldium bicarbonate and brine, and then dried (anhydrous magnesium sulphate). Gentle warming of the organic phase affected total cyclisation. Chromatography on silica, eluting with dichloromethane: ethyl acetate 10:1afforded the title penem (20) as a pale cream-coloured foam, 93 mg (74%), Rf (SiO₂/ethyl acetate-hexane, 1:1) = 0.64. v (film) 1800, 1752 1722 cm^-1; δ (CDCl₃) 2.22 (3H, s), 5.28 (1H, dAB, J 13.7) and 5.43 (1H, dAB, J13.7HZ), 5.81 (1H, d, J 2 Hz), 6.66 (1H, s), 6.89 (2H, dAB, J 9 Hz), 6.99 (1H, broad d, J 3 Hz),

7.21 - 8.11 (5H, m), 7.60 (2H, DAB, J 9Hz), 7.94 (2H,dAB, J 9Hz), 8.24 (2H, dAB J 9Hz) m/e (FAB/3-N0BA) 664 (MH⁺-CH₃COOH).

Example 2(a) 4-Nitrobenzyl (5RS)-(Z)-6[1-(4'-Methoxyphenyl sulphonyl)-indol-2-ylmethylene]penem-3-carboxylate The penem-acetate (20) from Preparation 2(g) (90 mg; 0.142 mmol) in dichloromethane (5 ml) at -30°C under argon was treated with a solution of 1, 8-diazabicyclo- [5.4.0]undec-7-ene (DBU) (28 μl) in dichloromethane

(200 μl). A yellow colour formed immediately. After 15 minutes dilute citric acid was added and the reaction was diluted with dichloromethane, washed with brine and dried (anhydrous magnesium sulphate). The crude product was chromatographed on silica-gel, eluting with dichloromethane grading slowly to dichloromethane-ethyl acetate (5:1). The (Z) and (E) isomers were obtained in a 4:1 ratio. The (E) isomer (21) was obtained as a yellow solid, 16 mg (18.6%); Rf (SiO₂/ethyl acetate-hexane, 1:2) = 0.4, ν (film) 1760, 1720 cm^-1; δ (CDCl₃) 3.78 (3H, s), 5.31 and 5.47 (2H, 2 x d, J 13.6 Hz), 6.54 (1H, d, J 0.6Hz), 6.82 (2H, brd AB, J 9 Hz), 7.26 - 8.21 (6H, m), 7.61 (2H, brd AB, J 9Hz), 7.66 (2H, brd, AB, J 9Hz), 7.96 (1H, s), 8.27 (2H, brd AB, J 9Hz); λ _max (2% CHCI₃ in EtOH) 249 nm, Em 24342; λ _max 330 nm Em 14798.

The (Z) isomer (22) was obtained as an orange-yellow crystalline solid, 53 mg (61.5%); Rf (SiO₂/ethyl acetate-hexane, 1:2) = 0.36; ν (Nujol) 1765, 171-5, 1640 cm^-1; v (KBr) 1765, 1715, 1661 cm^-1; δ (CDCI₃) 3.79 (3H, s), 5.32 and 5.49 (2H, 2 x d, J 13.5 Hz), 6.45 (1H, d, J 0.7 Hz) 6.64 (1H, s), 6.84 (2H, brd, AB, J 9 Hz), 7.25 - 8.28 (9H, m), 8.1 (1H, s), 8.26 (2H, brd. AB J 9Hz); λ_max (2% CHCI₃ in EtOH) 248 nm Em 24023, λ max 327 nm Em 21021.

Example 2(b)

Sodium (5RS) ( Z)-6-[1-(4'-Methoxyphenylsulphonyl)-indol-

2-ylmethylene]penem-3-carboxylate

The penem ester (22) from Example 2(a) (50 mg; 0.0829 mmol) in dioxane (5 ml), tetrahydrofuran (5 ml) and water (1 ml) was hydrogenolysed at atmospheric pressure in the presence of 5% palladium on carbon (70 mg) for 0.5 hour, when t.l.c. showed the absence of any starting material. The reaction mixture was treated with sodium bicarbonate solution (0.73 ml of a 1% aqueous solution) and filtered. The catalyst was washed with water (50 ml) and the filtrate evaporated to low volume. This concentrated solution was chromatographed on Biogel P2, eluting with water. Appropriate fractions (u.v. spectrum) were freeze-dried to afford the title penem sodium salt (23) as an orange-yellow amorphous solid, 24 mg (60%); ν (KBr) 1760, 1660, 1620, 1595 cm^-1; ν (Nujol) 1760, (1710 - 1515) br. 1600 sharp, 1275, 1175, 1095, 1050, 840 810 cm^-1; δ (D₂O/DMSO, 1:6) 3.71 (3H, s), 6.66 (1H, s), 6.76 (1H, s), 6.99 (2H, d, J 9Hz), 7.06 (1H, s), 7.30 (1H, broad dd, J 7.5Hz) 7.44 (1H, broad dd, J 7.5Hz), 7.6 - 7.67 (4H, m), 8.06 (1H, broad d, J 8.4Hz); λ _max (H₂O) 247 nm, Em 10580, λ _max 338 nm, Em 7367.

Preparation 3 5-Methoxycarbonylbenzotriazole

Methyl 3,4-diaminobenzoate was reacted with aqueous acetic acid and sodium nitrite according to the method of F.R. Benson et al, J. Am. Chem. Soc, 74, (1952), 4917, and the title compound (24) isolated in 60% yield, as a crystalline solid, m.p. 170°C (ethyl acetate); v _max (Nujol) 3200 - 2200 (b) and 1720 cm^-1; δ ppm [(CD₃)₂SO] 3.96 (3H, s); 7.80 -8.30 (2H, m); 8.65 (1H, m); 11.77 (1H, s).

1-Methyl-5-methoxycarbonylbenzotriazole

5-Methoxycarbonylbenzotriazole (24) (0.88g) was dissolved in dry N,N-dimethyl formamide (20 ml), potassium carbonate (0.35 g) and methyl iodide (0.62 ml) were added and the reaction mixture stirred under an atmosphere of argon in a stoppered flask at ambient temperature for 1 hour.

The reaction mixture was diluted with ethyl acetate (180 ml) and washed with water (2 x 40 ml) saturated brine solution (1 x 40 ml) and dried (MgSO₄). Chromatography on silica gel eluting with ethyl acetate-n-hexane gave the title compound (25) (0.27 g), m.p. 177°C; λ _max (EtOH) 272 n.m. (ε 5648); found M⁺ C₉H₉N₃O₂, 191; ν _{m ax} (CHCI₃) 1725 and 1620 cm^-1; δ ppm (CDCI₃) 3.97 (3H, s); 4.33 (3H, s); 7.54 (1H, d, J = 10Hz); 3.17 (1H, bd, J = 10Hz); 8.77 (1H, bs); (Found: C, 56.90; H, 4.71; N, 21.78. C₉H₉N₃O₂ requires C, 56.54; H, 4.71; N, 21.99%).

Preparation 3(a) 1-t-Butyldimethylsilyl-3-(_1-methylbenzotriazol-5-yl- carbonyl)-4-tritylthioazetidin-2-one

A solution of n-butyllithium (1.68M in hexane; 1.44 ml) was added to a solution of diisopropylamine (0.32 ml) in dry tetrahydrofuran (THF) (10 ml) at -70°C under dry argon. After stirring at -70°C for 20 minutes the mixture was treated with a solution of 1-t-butyldimethylsilyl-4-tritylthioazetidin-2-one ( 1 ) (0.92 g) (Bristol-Myers Patent Specification GB 2042515 A) in dry THF (14 ml). After stirring at -70°C for a further 30 minutes the mixture was treated with a solution of 1-methyl-5-methoxycarbonyl- benzotriazole (25) (0.46 g) in warm, dry THF (12 ml). The reaction mixture was stirred at -70°C for 20 minutes then treated with saturated ammonium chloride solution (20 ml) and allowed to warm to room temperature. Ethyl acetate (150 ml) was added and the organic phase washed with saturated brine (x 2), dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate/ n-hexane) gave the title azetidinone (26) (0.78 g) as a foam; ν _max. (CHCI₃) 1750; 1740 (sh); 1680 cm^-1; δ ppm (CDCI₃) 0.04 (6H, s); 0.97 (9H, s); 4.27 (4H, s); 5.14 (1H, d, J = 1.8 Hz); 6.80 - 8.55 (18H, m); m/e (MH⁺) 619. Preparation 3(b)

1-t-Butyldimethylsilyl-3-[hydroxy(1-methylbenzotriazol- 5-yl)methyl]-4-tritylthioazetidin-2-one

The ketone (26) from Preparation 3(a) (104 mg) was dissolved in dioxan (3 ml) containing pH 7.0 phosphate buffer (0.3 ml) and the solution chilled to 0°C. Sodium borohydride (16 mg) was added in portions with stirring over 2.5 hours after which time the reaction mixture was allowed to warm to room temperature. The solution was diluted with ethyl acetate and brine (ca. 40 ml). The organic phase was separated, washed with brine, dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate n-hexane) gave two isomeric alcohols ( 27 ) : Isomer I (89 mg), Rf 0.12 (ethyl acetate : hexane; 1 : 1) a white crystalline solid, m.p. 161° - 163°C (ethyl acetate/n-hexane); v _max. (CHCI₃) 1740 and 1600 cm^-1; δ ppm (CDCI₃) 0.67 (9H, s); 2.45 (1H, b.s.); 3.82 (1H, dd, J = 6.0, 1.8 Hz); 3.98 (1H, d, J = 1.8 Hz); 4.2 (3H, s); 4.35 (1H, d, J = 6.0 Hz); 7.10 - 7.50 (18H, m) . (Found: C, 69.47; H, 6.51; N, 9.00; S, 5.08. C₃₆H₄₀N₄O₂SSi requires C, 69.68; H, 6.45; N, 9.03; S, 5.16%) m/e (MH⁺) 621. Isomer II (10 mg), Rf 0.08 (ethyl acetate : hexane 1 : 1); ν _max. (CHCI₃) 1740 and 1600 cm^-1: δ ppm (CDCI₃) 0.73 (9H, s); 2.10 (1H,. b.s.); 3.78 (1H, dd, J= 6.6, 1.6 Hz); 4.19 (1H, d,J = 1.6 Hz); 4.35 (3H, s); 4.51 (1H, d, J = 6.6 Hz); 7.10 - 7.70 (18H, m); m/e (MH⁺) 621.

Preparation 3(c) (3RS,4SR) 3-[Hydroxy(1-methylbenzotriazol-5-yl)methyl]- 4-trityl-thioazetidin-2-one

The azetidinone (27) (Isomer I) from Preparation 3(b) (306 mg) was dissolved in methanol (15 ml) and the solution chilled to -15°C. Potassium fluoride (37 mg) in methanol (3 ml) was added dropwise and the reaction mixture stirred at -15°C for 1 hour. The solution was diluted with ethyl acetate and brine (ca. 100 ml) and the organic phase separated and washed with brine, dried (MgSO₄) and evaporated to give the title compound (28) as a solid (240 mg). ν _{max .} (KBr) 1760 and 1600 cm^-1. δ ppm C(CD₃)₂SO] 3.55 (1H, m); 4.10 (1H, d, J = 3 Hz); 4.25 (3H, s); 5.02 (1H, m, collapses to a doublet on addition of D2O, J = 3 Hz); 5.79 (1H, d,

J = 5 Hz); 6.90 - 7.50 (16H, m); 7.75 - 8.25 (2H, m); m/e (MNa⁺) 529.

Preparation 3(d)

(3RS,4SR) 2-[Acetoxy(l-methylbenzotriazol-5-yl)methyl]- 4-tritylthioazetidin-2-one

The azetidinone (28) from Preparation 3(c) (2.40 g) was partially dissolved/suspended in dry dichloromethane (MDC) (200 ml) and this partial solution chilled to 0°C. 4-Dimethylaminopyridine (0.64 g) was added, followed by the dropwise addition of a solution of acetic anhydride (0.53 ml) in MDC (20 ml). The reaction mixture was stirred at 0°C for 2 hours and then allowed to warm to room temperature. The resulting solution was washed with1N HCl (50 ml), water (50 ml), saturated sodium bicarbonate solution (50 ml), brine (50 ml), dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate : n-hexane 2:1) gave the title compound (29) (2.57 g) as a crystalline solid, m.p. 211° - 214°C (ethyl acetate); ν _max. (CHCI₃) 3400 and 1770 (br.) cm^-1; δ ppm

(CDCI₃) 2.11 (3H, s), 3.60 (1H, dd, J = 6.5 and 4.0 Hz), 4.26 (4H, s), 4.45 (1H, d, J = 4.0 Hz), 6.21 (1H, d, J = 6.5 Hz), 7.10 - 8.0 (18H, m); (Found: C,. 69.33; H, 5.28; N, 10.12; S, 5.86. C₃₂H₂₈N₄O₃S. ½ H₂O requires C, 68.94; H, 5.21; N, 10.05; S, 5.74%; m/e (MH⁺) 549.

Preparation 3(e)

(3RS,4SR) 3-[Acetoxy(1-methylbenzotriazol-5-yl)methyl]- 1-(1-p-nitrobenzyloxycarbonyl-1-triphenylphosphor- anylidenemethyl)-4-tritylthioazetidin-2-one

The azetidinone (29) (150 mg) from Preparation 3(d) and p-nitrobenzylglyoxylate monohydrate (68 mg) were heated in refluxing benzene (12 ml) with provision for azeotropic removal of water (Dean and Stark apparatus containing 4A molecular sieves) for 1 hour. The mixture was cooled to room temperature and treated with triethylamine (5 mg). After stirring at room temperature for 1 hour the reaction mixture was evaporated to yield the crude hydroxy ester (30); v _max. (CHCI₃) 3530, 1770 and 1750 cm^-1. A solution of the crude hydroxy ester (30) in dry THF (7 ml) was cooled to -20°C and treated with 2,6-lutidine (0.039 ml) followed by the dropwise addition of a solution of thionyl chloride (0.022 ml) in dry THF (2 ml). After stirring at -20°C for 20 minutes the reaction mixture was allowed to warm to 0°C then filtered through a celite pad and evaporated to give the crude chloroester (31). Triphenylphosphine (211 mg) was added to a solution of the chloroester (31) in dry dioxane (3 ml) and the resulting mixture concentrated to approximately 1 ml. 2, 6-Lutidine (0.046 ml) was then added and the mixture stirred under an atmosphere of argon for 12 hours at 50°C. After addition of ethyl acetate (50 ml) the organic phase was washed with saturated brine; IN hydrochloric acid; saturated brine, saturated sodium bicarbonate solution; saturated brine and dried (MgSO₄). Evaporation and chromatography (silica gel eluted with ethyl acetate-n-hexane) gave the title phosphorane (32) as a pale yellow foam (130 mg). ν_max. (CHCl₃) 1750, 1625 and 1610 cm^-1

Preparation 3 ( f) Silver (3RS,4SR) 3-Cacetoxy(1-methylbenzotriazol-5-yl) methyl]-1-(1-p-nitrobenzyloxycarbonyl-1-triphenyl- phosphoranylidenemethyl)-azetidin-2-one 4-thiolate

The phosphorane (32) (2.38 g) from Preparation 3(e) was dissolved in a mixture of methanol (15 ml) and dry MDC (6 ml) and the solution treated with pyridine

(0.25 ml). A solution of silver nitrate in methanol (0.15M; 19.0 ml) was then added dropwise, with stirring. After 1 hour at room temperature followed by 1 hour at 0°C the resulting solid was separated by filtration and washed with dry ether. The silver thiolate (33) was obtained as a yellow solid (1.70 g) v _max. (CHCI₃) 1750, 1620 and 1610 cm^-1.

Preparation 3(g) p-Nitrobenzyl (5RS, 6SR) -6-[acetoxy(1-methylbenzo- triazol-5-yl)methyl]-penem-3-carboxylate

The silver thiolate (33) (308 mg) from Preparation 3(f) was dissolved in dry MDC (4 ml) and the solution chilled to 0°C. 4-Dimethylaminopyridine (47 mg) and acetic formic anhydride (0.28 ml) were added followed by triethylamine hydrochloride (240 mg). The mixture was stirred for 5 minutes at 0°C when a precipitate formed. The cooling bath was removed and stirring continued for 30 minutes. The mixture was diluted with ethyl acetate (50 ml) and the precipitate removed by filtration and washed well with ethyl acetate. The filtrate was washed with chilled1N hydrochloric acid, water, sodium bicarbonate solution (x 2), water, and with brine. Drying (MgSO₄) gave an ethyl acetate solution which was stirred under an atmosphere of argon at 40°C for 1 hour. Evaporation gave a gum which was chromatographed (silica gel eluted with ethyl acetate-n-hexane) to give the title penem (34) as a foam (140 mg) v _max. (CHCl₃) 1800, 1750 (sh) and 1720 cm^-1; δ ppm (CDCI₃) 2.18 (3H, s); 4.30 - 4.36 (4H, m); 5.27 (1H, d, J= 13.5 Hz); 5.40 (1H, d, J = 13.5 Hz); 5.83 (1H, d, J = 2.0 Hz); 6.33 (1H, d, J = 6.2 Hz); 7.29 (1H, s); 7.40 - 8.30 (7H, m) ; m/e (MH⁺) 510.

Example 3(a) p-Nitrobenzyl (5RS) (Z) 6-(1-methylbenzotriazol-5-yl- methylene)penem-3-carboxylate

The penem (34) from Preparation 3(g) (297 mg) was dissolved in dry MDC (20 ml) and the solution chilled to -40°C under an atmosphere of argon. A solution of 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) (0.107 ml) in dry MDC (2 ml) was added dropwise and the reaction stirred 0.5 hours at -30°C. The reaction mixture was diluted with MDC (2 ml), washed with dilute citric acid (10 ml), sodium bicarbonate solution (2 x 10 ml), saturated brine (10 ml) and dried (MgSO4). The title penem (35) was crystallised from this solution as a 7:1 mixture of Z:E isomers (125 mg) , λ _max. (EtOH) 262 (ε 27,944), 306 nm (ε 25,463): ν _max. (CHCI₃) 1785, 1720 and 1610 cm^-1; δ ppm (CDCl₃).4.35 (3H, s), 5.31 (1H, d, J = 13.5 Hz), 5.47 (1H, d, J = 13.5 Hz), 6.49 (1/8 H, s), 6.73 (7/8 H, s), 6.84 (1/8 H, s), 7.35 - 8.60 (8 7/8 H, m); m/e (MH⁺) 450. Example 3(b)

Sodium (5RS) (Z)-6-(1-Methylbenzotriazol-5-yl- methylene)penem-3-carboxylate

The penem ester (35) from Example 3(a) (120 mg) was dissolved in a mixture of dioxane (40 ml) and water (12 ml). 5% Palladium on carbon (150 mg) was added and the mixture hydrogenated at room temperature for 30 minutes. A 1% w/v sodium bicarbonate solution (2.24 ml) was added, the mixture filtered through celite and the pad washed thoroughly with water. The filtrate was concentrated under reduced pressure to a small volume (ca. 5 ml) and chromatographed on Biogel P2 using water as eluent.

The appropriate fractions, as determined by ultra-violet spectroscopy, were combined and freeze- dried to give the title sodium salt (36) (61 mg) as a yellow solid containing a 7 : 1 mixture of Z : E isomers. λ _max. (H₂O) 261 (ε 20,488), 302 nm. (ε 20,783); ν _max. (KBr) 1760 and 1605 cm^-1; δ ppm (D₂O) inter alia 4.20 (s, minor isomer) and 4.22 (s, major isomer) (3H): 6.36 (s, minor isomer) and 6.56 (major isomer) (1H); 6.76 (s, minor) and 6.94 (s, major isomer) (1H); 6.94 (s, major), and 7.12 (s, minor) (1H); 7.29 (d, J = 8.5 Hz, major) and 7.54 (d, J = 8.5 Hz, minor) (1H); 7.63 (d, J = 8.5 Hz, major) and 7.80 (d, J = 8.5 Hz, minor) (1H); 7.69 (s, major) and 8.05 (s, minor) (1H).

Preparation 4

Methyl 3-(4-Methoxybenzyl) thieno[3,2-d]triazole-5- carboxylate

5-Chloro-1-(4-methoxybenzyl)-1,2,3-triazole-4-carbox- aldehyde [J. Becher et al., J. Heterocyclic Chem., 21., 1603 (1984)] (1.69g) was dissolved in methanol (14.0ml) and treated with ethyl 2-mercaptoacetate (0.73ml). Potassium carbonate (0.92g) was added and the mixture heated at reflux for 3 hours. The reaction mixture was then cooled and poured into water (50ml). The resulting precipitated solid was filtered off and dried in vacuo over phosphorous pentoxide to give the title compound (37) (0.86g) v_max (CHCI₃) 1715, 1615cm^-1; δ_pprn (CDCl₃) 3.84 (3H, s), 3.86 (3H, s), 5.62 (2H, s), 6.97 (2H, d, J 11.5Hz), 7.38 (2H, d, J 11.5Hz), 7.92 (1H, s); (Found: M⁺ 303).

Methyl 1H-thieno[2,3-d]triazole-5-carboxylate Methyl 3-(4-methoxybenzyl) thieno[3,2-d]triazole-5- carboxylate (37) (0.5g) was suspended in trifluoroacetic acid (14.0ml) and warmed in an oil bath at 80°C with stirring. After 3.5 hours, when very little starting material remained (the reaction being monitored by h.p.l.c.) the solution was cooled and the trifluoroacetic acid evaporated. The resulting solid residue was triturated with water (10ml) and chloroform (10ml), then dried in vacuo over phosphorous pentoxide to give the title compound (38) (0.19g) v_max (nujol) 3125, 1730cm^-1; (Found: M⁺ 183).

N-Methylated derivative of methyl 1H-thieno[2,3-d] triazole-5-carboxylate Methyl 1H-thieno[2,3-d]triazole-5-carboxylate (38) (0.14g) was dissolved in dry N,N-dimethylformamide (3ml). Potassium carbonate (0.052g) followed by methyl iodide (0.10ml) were added, and the mixture was stirred in a stoppered flask at room temperature for 2 hours. The reaction mixture was then extracted into ethyl acetate (25ml) and washed with water (2 x 10ml), saturated brine (1 x 10ml) and dried (MgSO₄). Evaporation followed by chromatography (silica gel eluted with n-hexane/ethyl acetate, 3:1) afforded the least polar regio isomer (39) in 46% yield (0.07g), mp 123-126°C (ethyl acetate/hexane), v_max (CHCI₃) 1720cm^-1; δ_ppm (CDCI₃) 3.90 (3H, s), 4.36 (3H, s), 7.87 (1H, s); (Found: MH⁺ 197).

Preparation 4(a)

(3RS,4SR) 1-t-Butyldimethylsilyl-3-(N-methylthieno- triazol-5-ylcarbonyl)-4-tritylthioazetidin-2-one

A solution of n-butyllithium (1.68M in hexane; 1.44ml) was added to a solution of diisopropylamine (0.32ml) in dry tetrahydrofuran (THF) (10ml) at -70°C under dry argon. After stirring at -70°C for 20 minutes the mixture was treated with a solution of 1-t-butyl- dimethylsilyl-4-tritylthioazetidin-2-one (1) (0.92g) (Bristol-Myers Patent Specification G.B. 2042515A) in dry THF (14ml). After stirring at -70°C for a further 30 minutes the mixture was treated with a solution of the N-methyl derivative of methyl

1H-thieno[2, 3-d]triazole -5-carboxylate (39) (0.4gg) in warm dry THF (12ml). The reaction mixture was stirred at -70°C for 20 minutes, then treated with saturated ammonium chloride solution (20ml) and allowed to warm to room temperature. Ethyl acetate (150ml) was added and the organic phase washed with saturated brine (x2), dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate-n-hexane) gave the title azetidinone (40) (0.99g) as a foam; v_max (CHCI₃) 1755, 1665cm^-1; δ_ppm (CDCI₃) 0.97 (9H, s), 4.04 (1H, d, J 2.5Hz), 4.37 (3H, s), 5.02 (1H, d, J 2.5Hz), 6.90-7.70 (16H, m); (Found [MNa]⁺ 647).

Preparation 4(b)

(3RS,4SR) 1-t-Butyldimethylsilyl-3-[hydroxy(N-methyl thienotriazol-5-yl)methyl]-4-tritylthioazetidin-2-one The ketone (40) from Preparation 4(a) (1.0g) was dissolved in dioxan (30ml) containing pH 7.0 phosphate buffer (3.0ml) and the solution chilled to 0°C. Sodium borohydride (0.16g) was added in portions with stirring over 2.5 hours after which time the reaction mixture was allowed to warm to room temperature. The solution was diluted with ethyl acetate and brine (ca. 400ml). The organic phase was separated, washed with brine, dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate/n-hexane) gave two isomeric alcohols (41): Isomer I (23mg), Rf 0.22 (ethyl acetate:hexane, 1:2); v_max (CHCI₃) 3400 (b),

1740 and 1600cm^-1; δ_ppm (CDCI₃) 0.93 (9H, s), 1.90 (1H, bs, exchanges on addition of D₂O), 3.55 (1H, ra), 3.77 (1H, bs, sharpens to a doublet on addition of D₂O, J 4Hz), 4.28 (3H, s), 4.51 (1H, d, J 2Hz); 6.98 (1H, s), 7.05-7.60 (15H, m); Isomer II (800mg), Rf 0.17 (ethyl acetate:hexane, 1:2); v_max (CHCI₃) 3400(b), 1740 and 1600cm^-1; δ_ppm (CDCI₃) 0.77 (9H, s), 2.25 (1H, bs exchanges on addition of D₂O), 3.73 (1H, dd, J 7Hz and 2Hz), 4.10-4.40 (5H, m), 6.68 (1H, s), 7.05-7.60 (15H, m); (Found: [MH]⁺ 627; [MNa]⁺ 649).

Preparation 4(c)

(3RS,4SR) 3-[Hydroxy(N-methylthienotriazol-5-yl)methyl]

-4-tritylthioazetidin-2-one

The azetidinone (41) (isomer II) from Preparation 4(b) (790mg) was dissolved in a mixture of methanol (30ml) and methylene dichloride (MDC) (30ml) and the solution chilled to -15°C. Potassium' fluoride (90mg) in methanol (3ml) was added dropwise and the reaction mixture stirred at -10°C for 1 hour followed by a further 0.5h to 0°C. The product, which crystallised from solution as the reaction proceeded, was separated by filtration, washed with methanol and dried in vacuo to give the title azetidinone (42) (580mg), ν_max (CHCI₃) 3600-3200(b), 3400, 1760cm^-1, δ_ppm [(CD₃)₂SO] 3.76 (1H, m), 4.15 (1H, d, J 2.6Hz), 4.34 (3H, s), 5.13 (1H, m, sharpens to a quartet oh addition of D₂O), 6.54 (1H, d, J 5.3Hz, exchanges on addition of D₂O), 7.05-7.30 (16H, m), 7.38 (1H, s, exchanges on addition of D₂O); (Found; [MH]⁺ 513).

Preparation 4(d)

(3RS,4SR)3-[Acetoxy(N-methylthienotriazol-5-yl)methyl]

-4-tritylthioazetidin-2-one

The azetidinone (42) from Preparation 4(c) (563mg) was dissolved in dry THF (30ml) and this solution chilled to 0°C. 4-Dimethylaminopyridine (147mg) was added, followed by the dropwise addition of a solution of acetic anhydride (0.122ml) in MDC (4ml). The reaction mixture was stirred for 1 hour at 0°C followed by 1 hour to room temperature. The reaction was diluted with MDC (200ml) and washed with 1N HCl (50ml), water (50ml), saturated sodium bicarbonate solution (50ml), brine (50ml) and dried (MgSO₄). Evaporation followed by chromatography (silica gel eluted with ethyl acetate/n-hexane, 1:1) gave the title azetidinone (43) (430mg), ν_max (CHCI₃) 3400, 1780cm^-1; δ_ppm (CDCI₃) 2.10 (3H, s) , 3.66 (1H, q, J 6.0Hz and 3.0Hz), 4.30 (1H, bs, exchanges on addition of D2O) , 4.35 (3H, s) , 4.48 (1H, d, J 3.0Hz), 6.33 (1H, d, J 6.0Hz), 7.0-7.5 (16H, m); (Found: [MH]⁺ 555).

Preparation 4(e)

(3RS,4SR) 3-[Acetoxy(N-methylthienotriazol-5-yl)methyl]

-1-(1-p-nitrobenzyloxycarbonyl-1-triphenylphosphorany- lidinemethyl)-4-tritylthioazetidin-2-one

The azetidinone (43) (422mg) from Preparation 4(d) and p-nitrobenzylglyoxylate monohydrate (190mg) were heated in refluxing benzene (25ml) with the provision for the azeotropic removal of water (Dean and Stark apparatus containing 4A molecular, sieves) for 1 hour. The mixture was cooled to room temperature and treated with triethylamine (15mg). After stirring at room temperature for 1 hour the reaction mixture was evaporated to yield the crude hydroxy ester (44); v_max (CHCI₃) 3530, 1770 and 1750cm^-1. A solution of the crude hydroxy ester (44) in dry THF (15ml) was cooled to -20°C and treated with 2,6-lutidine (0.106ml) followed by the dropwise addition of a solution of thionyl chloride (0.062ml) in dry THF (4ml). After stirring at -20°C for 20 minutes the reaction mixture was allowed to warm to 0°C then filtered through a celite pad and evaporated to give the crude chloroester (45). Triphenylphosphine (600mg) was added to a solution of the chloroester (45) in dry dioxan (4ml) and the resulting mixture concentrated to approximately lml. 2,6-Lutidine (0.133ml) was then added and the mixture stirred under an atmosphere of argon for 12 hours at 50°C. After addition of ethyl acetate (120ml) the organic phase was washed with saturated brine, 1N hydrochloric acid, saturated brine, saturated sodium bicarbonate solution, saturated brine and dried

(MgSO₄). Evaporation and chromatography (silica gel eluted with ethyl acetate/n-hexane) gave the title phosphorane (46) as a pale yellow foam (430mg); v_max (CHCI₃) 1750, 1625 and 1610cm^-1.

Preparation 4(f)

Silver (3RS,4SR) 3-[Acetoxy(N-methylthienotriazol-5-yl) methyl]-1-(1-p-nitrobenzyloxycarbonyl-1-triphenyl- phosphoranylidenemethyl)-azetidin-2-one-4-thiolate The phosphorane (46) (2.07g) from Preparation 4(e) was dissolved in a mixture of methanol (13ml) and dry MDC (13ml) and the solution treated with pyridine (0.21ml). A solution of silver nitrate in methanol (0.15M; 16.0ml) was then added dropwise, with stirring. After 1 hour at room temperature followed by 1 hour at 0°C the reaction was concentrated to approximately half volume and dry ether (approx. 10ml) added. The resulting precipitated solid was filtered off, washed with dry ether and dried in vacuo to give the title salt (47) as a yellow solid (1.42g), v_max (nujol) 1750, 1625 and 1610cm^-1.

Preparation 4(g) p-Nitrobenzyl (5RS,6SR)-6-[Acetoxy(N-methylthieno- triazol-5-yl)methyl]penem-3-carboxylate

The silver thiolate (47) (1.42g) from Preparation 4(f) was dissolved in dry MDC (17ml) and the solution chilled to 0°C. 4-Dimethylaminopyridine (0.217g) and acetic formic anhydride (1.30ml) were added, followed by triethylamine hydrochloride (1.12g). The mixture was stirred for 5 minutes at 0°C when a precipitate formed. The cooling bath was removed and stirring continued for 30 minutes. The mixture was diluted with ethyl, acetate (150ml), and the precipitate removed by filtration and washed well with ethyl acetate. The filtrate was washed with chilled 1N hydrochloric acid, water, sodium bicarbonate solution (x2), water, and with brine. Drying (MgSO4) gave an ethyl acetate solution which was stirred under an atmosphere of argon at 40°C for 1 hour. Concentration of this ethyl acetate solution resulted in the title penem (48) crystallising from the solution as a pale, cream coloured solid (528mg), v_max (CHBr₃) 1780, 1740, 1720cm^-1, λ_max (CH₃CN) 258 (ε 27176), 314nm (ε 8304); δppm [(CD₃)₂SO] 2.1223H, s), 4.34 (3H, s), 4.73 (1H, broad d, J 5Hz), 5.31 and 5.39 (2H, ABq, J 14Hz), 5.92 (1H, d, J 2Hz), 6.46 (1H, d, J 5Hz), 7.48 (1H, s), 7.65 (2H, d, J 8.5Hz), 8.23 (2H, d, J 8.5Hz), 7.88 (1H, s). Example 4(a) p-Nitrobenzyl (5RS) (Z)-6-(N-Methylthienotriazol-5-yl- methylene)penem-3-carboxylate

The penem (48) from Preparation 4(g) (100mg) was dissolved in dry MDC (40ml) and the solution chilled to -40°C under an atmosphere of argon. A solution of 1,8-diazobicyclo[5.4.0]undec-7-ene (DBU) (0.035ml) in dry MDC (1ml) was added and the reaction stirred 0.5 hours at -30°C. The reaction mixture was diluted with MDC (40ml), washed with dilute citric acid (10ml), sodium bicarbonate solution (10ml), saturated brine (10ml) and dried (MgSO₄). Concentration of this solution resulted in the title penem (49) crystallising from solution as a yellow solid (30mg), v_max (KBr) 1785, 1715, 1655, 1605cm^-1; λ_max (CH₃CN) 320nm (ε 38532); δ_ppm [(CD₃)₂SO] 4.40 (3H, s), 5.37 and 5.47 (2H, ABq, J 14Hz), 6.89 (1H, d, J 1Hz), 7.71 (2H, d, J 8.6Hz), 8.26 (2H, d, J 8.6Hz), 7.77 (1H, s), 7.83 (1H,s), 7.96 (1H, s).

Example 4(b)

Sodium (5RS) (Z)-6-[N-Methylthienotriazol-5-ylmethyl- ene]penem-3-carboxylate

The penem ester (49) from Example 4(a)(260mg) was dissolved in a mixture of dioxan (150ml) and water (20ml). 5% Palladium on carbon (340mg) was added and the mixture hydrogenated at room temperature for 40 minutes. A 5% w/v sodium bicarbonate solution (0.95ml) was added, the mixture filtered through a celite pad and the pad washed thoroughly with water. The filtrate was concentrated under reduced pressure to a small volume (ca 5ml) and chromatographed on Biogel P2 using water as eluent. The appropriate fractions, as determined by u.v. spectroscopy, were combined and freeze-dried to afford the title penem sodium salt (50) as a yellow solid (64mg); ν_max(KBr) 1755, 1655, 1595 and 1550cm^-1; λ_max (H₂O) 316nm (ε 24156); δppm (D₂O) 4.35 (3H, s), 6.50 (1H, s), 7.03 (1H, s), 7.30 (1H, s), 7.43 (1H, s); (Found: [MH]⁺343 and [MNa]⁺365).

Preparation 5

Ethyl 2-azido-3-(2-pyridyl) acrylate

Sodium metal (1.782g) was dissolved in ethanol (150ml) with stirring and the solution cooled to -20°C. A solution of ethyl azidoacetate (10g) and pyridine-2-carboxaldehyde (1.84ml) in ethanol (15ml) was added dropwise over a period of 15 minutes. When the addition was complete the reaction mixture was stirred at 5°C for 2 hours. The reaction was then quenched by the addition of a saturated solution of ammonium chloride and the resulting solution extracted with ethyl acetate. The organic solution was washed with brine, dried (MgSθ4) and evaporated. The crude product was purified by column chromatography (silica gel eluted with 10% ethyl acetate in hexane) to give the title compound (1.70g) as a pale yellow crystalline solid; ν_max(CHCl₃) 2120, 1715, 1620 cm^-1.

Ethyl pyrazolo[1,5-a]pyridine-2-carboxylate

Ethyl 2-azido-3-(2-pyridyl) acrylate (5.7g) was dissolved in dry xylene (150 ml) and heated to relux under an atmosphere of argon for 15 minutes. After cooling the solvent was removed by evaporation at reduced pressure, and the crude product subjected to column chromatography (silica gel eluted with ethyl acetate/hexane) to give the title compound (51)(2 1g) as a pale yellow crystalline solid; m.p. 48°-49°C

(ethyl acetate/hexane); v_max(CHCl₃) 1725cm^-1; δ(CDCl₃)

1.50 (3H, t, J 7.0Hz), 4.45 (2H, q, J 7.0Hz), 6.60-7.25

(3H, m), 7.40 (1H, m), 8.35 (1H, d, J 6.0Hz); (Found: M+190) .

Preparation 5(a)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-(pyrazolo[1,5-a] pyrid-2-ylcarbonyl)-4-tritylthioazetidin-2-one

A solution of n-butyllithium (1.68M in hexane; 1.44 ml) was added to a solution of diisopropylamine (0.32 ml) in dry tetrahydrofuran (THF) (10ml) at -70°C under dry argon. After stirring at -70°C for 20 minutes the mixture was treated with a solution of 1-t-butyldimethylsilyl-4-tritylthioazetidin-2-one(1) (0.92g) (British Patent Publication GB 2042515A (Bristol Myers)) in dry THF (14ml). After stirring at -70°C for a further 30 minutes the mixture was treated with a solution of ethyl pyrazolo[2,3-a]pyridine-2-carboxylate (51)(0.39g) in dry THF(10ml). The reaction mixture was stirred at -70°C for 20 minutes, then treated with saturated ammonium chloride solution (20ml) and allowed to warm to room temperature. Ethyl acetate (150ml) was added and the organic phase washed with saturated brine (x2), dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate :hexane, 1:1) gave the title azetidinone (52)(0.90g) as a foam; ._max(CHCl₃) 1750, 1680cm^-1; δppm (CDCI₃) 0.97 (9H, s), 4.81 (1H, d, J 2.0Hz), 4.97 (1H, d, J 2.0Hz), 6.70-8.60 (20H, m); (Found [MNa]⁺626). Preparation 5(b)

(3RS,4SR) 1-t-Butyldimethylsilyl-3-[hydroxy(pyrazolo [1,5-a]pyrid-2-yl)methyl]-4-tritylthioazetidin-2-one

The ketone (52) from Preparation 5(a)(0.90g) was dissolved in dioxan (30ml) containing pH7.0 phosphate buffer (3.0ml) and the solution chilled to 0°C. Sodium borohydride (0.14g) was added in portions with stirring over 1.5 hours after which time the reaction mixture was allowed to warm to room temperature. The solution was diluted with ethyl acetate and brine (ca.400ml). The organic phase was separated, washed with brine, dried (MgS04) and evaporated to yield a two component mixture. Chromatography (silica gel eluted with ethyl acetate: hexane, 1:1) gave the more polar isomer (53)(0.50g), Rf 0.27 (ethyl acetate:hexane, 1:1); v_max (CHCI₃) 3450(broad), 1740cm^-1; δppm (CDCI₃) 0.65 (9H, s), 2.70 (1H broad s.), 3.77 (1H, dd, J 2.5 and 7. 0Hz ) , 4. 20-4.40 ( 2H, m) , 6. 10-8.40 ( 20H, m) ; ( Found: [MNa]⁺628).

Preparation 5(c)

(3RS,4SR) 3-[Hydroxy(pyrazolo[1,5-a]pyrid-2-yl)methyl] -4-tritylthioazetidin-2-one

The azetidinone (53), the more polar isomer from Preparation 5(b)(464mg) was dissolved in methanol (30ml) and the solution chilled to -20°C. Potassium fluoride (68mg) in methanol (3ml) was added dropwise and the reaction mixture stirred at -15°C for 1.5 hour followed by a further 0.5 hour to 0°C. The product, which crystallised from solution as the reaction proceeded, was separated by filtration, washed with methanol and dried in vacuo to give the title azetidinone (54)(314mg), v_max(CHBr₃) 3500-3100, 3400, 1770cm^-1, δppm [(CD₃)SO] 3.67 (1H, m), 4.25 (1H, d, J 2.5Hz), 5.04 (1H, m, sharpens on addition of D₂O), 5.88 (1H, d, J 4.2Hz, exchanges on addition of D₂O), 6.40-8.70 (21H, m); (Found: [MNa]⁺514).

Preparation 5(d) (3RS,4SR) 3-[Acetoxy(pyrazolo[l,5-a]pyrid-2-yl)methyl] -4-tritylthioazetidin-2-one

The azetidinone (54) from Preparation 5(c)(1.73g) was suspended in dry methylene dichloride (MDC) (120ml). 4-Dimethylaminopyridine (0.51g) was added and the mixture chilled to 0°C. A solution of acetic anhydride (0.39ml) in MDC (10ml) was added dropwise to the stirred suspension and stirring continued for 1.0 hour at 0°C, followed by 0.5 hour to room temperature. The reaction was diluted with MDC (120ml) and washed with 1N HCl(50ml), water (50ml), brine (50ml) and dried (MgSθ4). Evaporation followed by chromatography (silica gel eluted with ethyl acetate:hexane, 1:1) gave the title azetidinone (55)(1.90g), v_max(CHCl₃) 3400, 1770cm^-1; δppm (CDCI₃) 2.10 (3H, s), 3.88 (1H, dd, J 3.0 and 6.0Hz), 4.20 (1H, s), 4.56 (1H, d, J 3.0Hz), 6.40 (1H, d, J 6.0Hz), 6.45-8.60 (20H, m); (Found: [MNa]⁺556).

Preparation 5(e)

(3RS,4SR) 3-[Acetoxy(pyrazolo[1,5-a]pyrid-2-yl)methyl]- 1-(1-p-nitrobenzyloxycarbonyl-1-triphenylphosphoranylid enemethyl)-4-tritylthioazetidin-2-one

The azetidinone (55)(1.85g) from Preparation 5(d) and p-nitrobenzylglyoxylate monohydrate (0.95g) were heated in refluxing benzene (80ml) with provision for the azeotropic removal of water (Dean and Stark apparatus containing 4A molecular sieves) for 1.5 hour. The mixture was cooled to room temperature and treated with triethylamine (90mg). After stirring at room temperature for 1 hour the reaction mixture was evaporated to yield the crude hydroxy ester (56); ν_{m ax} (CHCI₃) 3530, 1760cm^-1. A solution of the crude hydroxy ester (56) in dry THF (35ml) was cooled to -15°C and treated with 2,6-lutidine (0.48ml) followed by the dropwise addition of a solution of thionyl chloride (0.28ml) in dry THF (14ml) under an atmosphere of dry argon. After stirring at -10°C for 1 hour then reaction mixture was allowed to warm to room temperature then filtered through a celite pad and evaporated to give the crude chloroester (57). Triphenylphosphine (2.62g) was added to a solution of the chloroester (57) in dry dioxan (15ml) and the resulting mixture concentrated to approximately 4ml.

2,6-Lutidine (0.61ml) was added and the mixture stirred under an atmosphere of argon for 12 hours at 60°C. After addition of ethyl acetate (400ml) the organic phase was washed with saturated brine, 1N hydrochloric acid, saturated brine, saturated sodium bicarbonate solution, saturated brine and dried (MgSO₄). Evaporation and chromatography (silica gel eluted with ethyl acetate-hexane) gave the title phosphorane (58) as a yellow oil (0.80g); ν_max (CHCI₃) 1740, 1620 and 1610(sh)cm^-1.

Preparation 5(f)

Silver (3RS, 4SR) 3-[Acetoxy[pyrazolo[1,5-a]pyrid-2-yl) methyl]-1-(1-p-nitrobenzyloxycarbonyl-1-triphenyl- phosphoranylidinemethyl)azetidin-2-one-4-thiolate

The phosphorane (58)(1.93g) from Preparation 5(e) was dissolved in a mixture of methanol (13ml) and dry MDC (20ml) and the solution treated with pyridine (0.20ml). A solution of silver nitrate in methanol (0.15M, 15.6ml) was then added dropwise with stirring. After 1.0 hour at room temperature followed by 1.0 hour at 0°C the reaction was concentrated to approximately one quarter volume and dry ether (approx. 10ml) added. The resulting precipitated solid was filtered off, washed with dry ether and dried in vacuo to give the title salt (59) as a yellow solid (1.52g); v_max (CHBr₃) 1750, 1620 and 1610( sh) cm^-1.

Preparation 5(g) p-Nitrobenzyl (5RS,6SR) 6-[Acetoxy(pyrazolo[1,5-a] pyrid-2-yl)methyl]penem-3-carboxylate

The silver thiolate (59)(1.51g) from Preparation 5(f) was dissolved in dry MDC (18ml) and the solution chilled to 0°C. 4-Dimethylaminopyridine (0.24g) and acetic-formic anhydride (1.42ml) were added followed by triethylamine hydrochloride (1.22g). The mixture was stirred for 5 minutes at 0°C when a precipitate formed. The cooling bath was removed and stirring continued for 30 minutes. The mixture was diluted with ethyl acetate (150ml) and the precipitate removed by filtration and washed well with ethyl acetate. The filtrate was washed with chilled 1N hydrochloric acid, water, sodium bicarbonate solution (x2), water and brine. Drying (MgSθ4) gave an ethyl acetate solution which was stirred under an atmosphere of argon for 30 minutes at 40°C. Evaporation followed by rapid chromatography (silica gel eluted with a gradient of 50 - 67% ethyl acetate/hexane) gave the title penem (60)(665mg), v_max (CHCl₃) 1800, 1745(sh), and 1720cm^-1; δppm (CDCI₃) 2.14 (3H, s), 4.61 (1H, m), 5.28 and 5.44 (2H, ABq, J 14Hz), 6.12 (1H, d, J 2Hz), 6.48 (1H, d, J 3.5Hz), 6.56 (1H, s) ; 6.80 (1H, t, J 6.8Hz), 7.12 (1H, t, J 6.8Hz), 7.34 (1H, d, J 1.0Hz), 7.50 (1H,d, J 9.0Hz), 7.60 (2H, d, J 9.0Hz), 8.24 (2H, d, J 9.0Hz), 9.37 (1H, d, J 7.0Hz); (Found [MNa]⁺517). Example 5(a) p-Nitrobenzyl (5RS) (Z)-6-(Pyrazolo[l,5-a]pyrid-2- ylmethylene)penem-3-carboxylate

The penem (60) from Preparation 5(g)(96mg) was dissolved in dry MDC (20ml) and the solution chilled to -40°C with stirring under an atmosphere of argon. A solution of 1,8-diazobicyclo[5.4.0]undec-7-ene (DBU) (0.035ml) in dry MDC (1ml) was added and the reaction stirred 0.5 hours at -30°C. The reaction mixture was diluted with MDC (40ml), washed with dilute citric acid (10ml), sodium bicarbonate solution (10ml), saturated brine (10ml) and dried (MgSθ4). Evaporation and chromatography Csilica gel (3g) eluted with 4% ethyl acetate/MDC] gave the title penem (61)(67mg), v_max (CHBr₃) 1780(sh), 1765 and 1720cm^-1; λ_max (EtOH)

285nm (ε 30196); δppm (CDCl₃) 5.30 and 5.48 (2H, ABq, J 13.7Hz), 6.65 (1H, s), 6.67 (1H, s), 6.86 (1H, t, J 7.0Hz), 7.17 (1H, t, J 7.0Hz), 7.22 (1H, s), 7.40 (1H, s), 7.57 (1H, d, J 9.0Hz), 7.64 (2H, d, J 9.0Hz), 8.26(2H, d, J 9.0Hz), 8.43 (1H, d, J 7.0Hz); (Found: M⁺434).

Example 5(b)

Sodium (5RS) (Z)-6-[Pyrazolo[1,5-a]pyrid-2-ylmethylene] penem-3-carboxylate

The penem ester (61) from Example 5(a)(205mg) was dissolved in 20% aqueous dioxan (120ml) and hydrogenolysed at atmospheric pressure in the presence of 5% palladium on carbon (215mg) for 0.5 hour. The reaction mixture was then treated with sodium bicarbonate solution (0.88ml of a 5% w/v aqueous solution) and filtered through a celite pad. The pad was washed thoroughly with water (150ml) and the filtrate concentrated under reduced pressure to a volume of approximately 5ml. This concentrated solution was chromatographed on Biogel P2, eluting with water. Appropriate fractions, as determined by u.v. spectroscopy, were combined and freeze-dried to afford the title penem sodium salt (62) as a bright yellowsolid (100mg); ν_max (KBr) 1750, 1600cm^-1; λ_max (H₂O) 282nm (ε 25948); δppm (D₂O) 6.43 (1H, s), 6.53 (1H, s), 6.85 - 6.97 (2H, m), 7.00 (1H, s), 7.20 (1H, t, J 8.3Hz), 7.55 (1H, d, J 8.9Hz), 8.37 (1H, d, J 6.7Hz).

Preparation 6

Methyl Thieno[3,2-b]furan-2-carboxylate

A solution of methyl (2-formylthien-3-yl)oxymethyl acetate (4.84g), synthesised using literature procedures CS.Gronowitz, Arkiv Kemi, 1960, 16, 363; C.Paulmier et al, Bull.Soc.Chim.France, 1973, 7-8,

2434; J.M. Meunier et al Bull.Soc.Chim.France, 1971, 9, 3343], in methanol (180ml), was added to a solution of sodium metal (1.67g) in methanol (20ml) at ambient temperature. This mixture was warmed to reflux under an atmosphere of argon for ten minutes then cooled and added to methyl acetate (800ml). The resulting mixture was poured onto an ice-cold solution of 5% citric acid (500ml) and after thorough mixing the organic phase was separated, washed with saturated brine solution, dried (MgSO₄) and evaporated to dryness. Chromatography (silica gel eluting with ethyl acetate/hexane) gave the title compound (63), (2.0g); v_max (CHCI₃) 1715, 1550, 1540cm^-1; δppm (CDCI₃) 3.90 (3H, s); 7.00-7.60 (3H, m). Preparation 6(a)

( 3RS, 4SR) 1-t-Butyldimethylsilyl-3-(thieno[3,2-b]fur-2- -ylcarbonyl)-4-tritylthioazetidin-2-one

Azetidinone (1)(0.92g) was condensed with methyl thieno [3,2-b]furan-2-carboxylate (63)(0.36g) in the manner described in Preparation 5(a). Chromatography of the crude product (silica gel eluted with hexane:ethyl acetate, 4:1) gave the title azetidinone (64) as a crisp foam (0.77g); v_max (CHCI₃) 1750, 1660cm^-1; δppm (CDCI₃) 0.36 (3H, s), 0.38 (3H, s), 0.98 (9H, s), 4.13 (1H, d, J 1.5Hz), 5.00 (1H, d, J 1.5Hz), 6.88-7.60 (18H, m); (Found [MNa]⁺632).

Preparation 6(b)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-[hydroxy(thieno [3,2-b]fur-2-yl)methyl]-4-tritylthioazetidin-2-one

The azetidinone (64) from preparation 6(a) (0.76g) was reduced to a mixture of alcohols using the method described in Preparation 5(b). This two component mixture was separated by chromatography (silica gel eluted with ethyl acetate/ n-hexane) to give the more polar azetidinone (65) as the major isomer (0.56g); Rf(hexane: ethyl acetate, 4:1) 0.1; v_max (CHCI₃) 1735, 1600cm^-1; δppm (CDCI₃) -0.08, 0.03 (each 3H, s), 0.78(9H, s), 2.77 (1H, broad d, J 9Hz, exchanges on addition of D2O), 3.81 (1H, dd, J 6.7 and 1.9Hz), 4.03 (1H, m, sharpens to a doublet on addition of D₂O, J 6.7Hz), 4.10 (1H, d, J 1.9Hz), 6.30 (1H, s), 6.85-7.60 (17H, m); (Found [MNa]⁺634). Preparation 6(c)

(3RS,4SR) 3-[Hydroxy(thieno[3,2-b]fur-2-yl)methyl]-

-4-tritylthioazetidin-2-one

The azetidinone (65) from Preparation 6(b) (0.56g) was deprotected in the manner described in Preparation 5(c) to give the title azetidinone (66) as a cream coloured solid (0.42g); v_max (CHCl₃) 3400, 1765cm^-1.

Preparation 6(d)

(3RS, 4SR) 3-[Acetoxy(thieno[3,2-b]fur-2-yl)methyl]-4- tritylthioazetidin-2-one

The azetidinone (66) isolated in Preparation 6(c) (0.42g) was reacted according to the method described in Preparation 5(d) and the title azetidinone (67) was isolated in quantitative yield (0.45g); ν_max (CHCI₃) 3400, 1770, 1740 (sh) cm^-1; δppm (CDCl₃)2.10 (3H, s), 3.74 (1H, dd, J 2.5 and 5.4Hz), 4.23 (1H, broad s), 4.66 (1H, d, J 2.5Hz), 6.27 (1H, d, J 5.4Hz), 6.80 (1H, s), 7.05-7.60 (17H, m); (Found[MNa]⁺562).

Preparation 6(e) (3RS, 4SR) 3-[Acetoxy(thieno[3,2-b]fur-2-yl)methyl]- -1-(1-p-nitrobenzyloxycarbonyl-1-triphenylphosphor- anylidenemethyl)-4-tritylthioazetidin-2-one

The acetoxyazetidinone (67) isolated as described in Preparation 6(d)(1.90g) was converted to the title phosphorane in the manner described in Preparation

5(e). Chromatography of the phosphorane (silica gel eluted with a gradient of 35-100% ethyl acetate/hexane) gave the title azetidinone (68) as an orange foam (1.73g); v_max (CHCI₃) 1750, 1615 and 1605cm^-1. Preparation 6(f)

Silver (3RS, 4SR) 3-[Acetoxy(thieno[3,2-b]fur-2-yl) methyl]-1-(1-p-nitrobenzyloxycarbonyl-1-triphenyl phosphoranylidenemethyl)azetidin-2-one-4-thiolate

The phosphorane (68) (1.58g) from Preparation 6(e) was converted to the title silver salt as described in Preparation 5(e) to give the thiolate (69)(1.46g); v_max (CH₂Cl₂) 1750, 1620, 1610( sh) cm^-1.

Preparation 6(g) p-Nitrobenzyl (5RS, 6SR) 6-[Acetoxy(thieno[3,2-b]fur- -2-yl)methyl]penem-3-carboxylate

The silver thiolate (69) (0.5g) from Preparation 6(f) was converted to the title penem as described in the Preparation 5(g). Chromatography (silica gel eluted with ethyl acetate :hexane, 1:1) afforded the product

(70) as a bright yellow foam (0.24g) -max (CHCI₃) 1800, 1750 (sh) and 1720cm^-1; δppm (CDCI₃) 2.11 (3H, s), 4.43 (1H, m), 5.29 and 5.43 (2H, ABq, J 13.5Hz), 6.15 (1H, d, J 2Hz) , 6.33 (1H, d, J 4.2Hz), 6.84 (1H, s), 7.03 (1H, doublet showing fine coupling, J 5.5Hz), 7.28 (1H, doublet showing fine coupling J 5.5Hz), 7.35 (1H, singlet showing fine coupling), 7.59 (2H, d, J 9Hz), 8.23 (2H, d, J 9Hz).

Example 6(a) p-Nitrobenzyl (5RS) (Z)-6-(Thieno[3,2-b]fur-2-yl- methylene)penem-3-carboxylate

The acetoxy penem (70) (220mg) from Preparation 6(g) was reacted in the manner described in Example 5(a) to give a mixture of isomers. Chromatography (silica gel eluted with 2% ethyl acetate in MDC) afforded the (Z)- and (E)-isomers in a 25:1 ratio. The more polar (Z) -isomer, the title penem (71) was isolated as a yellow solid (125mg), v_max (CHCI₃) 1775, 1710, 1660 and 1605cm^-1; λ_max(EtOH) 350nm (ε 23,000); δppm (CDCI₃) 5.30 and 5.47 (2H, ABq, J 14Hz), 6.67 (1H, d, J 1Hz), 7.00 (1H, d, J 1Hz), 7.06 (1H, s), 7.09 (1H, d, J

5.5Hz), 7.40 (1H, s), 7.44 (1H, d, J 5.5Hz), 7.63 (2H, d, J 9Hz), 8.25 (2H, d, J 9Hz); (Found M⁺ 440).

Example 6(b)

Sodium (5RS) (Z)-6-(Thieno[3,2-b]fur-2-ylmethylene) penem-3-carboxylate

The penem ester (71)(120mg) from Example 6(a) was hydrogenolysed using the conditions described in Example 5(b) and the title penem sodium salt (72) isolated as a freeze-dried solid (50mg); V_max (KBr) 1750, 1660 and 1600cm^-1; λ_max (H₂O) 359nm (ε 19,200); δppm (D₂O) 6.54 (1H, s), 6.94 (1H, s), 6.97 (1H, s)7.08 (1H, d, J 5.3Hz), 7.13 (1H, s), 7.53 (1H, d, J 5.3Hz).

Preparation 7 ( a) (3RS, 4SR) 1-t-Butyldimethylsilyl-3-(imidazo[2,1-b] thiazol-6-ylcarbonyl)-4-tritylthioazetidin-2-one

Methyl imidazo[2,1-b]thiazole-6-carboxylate (0.4g) (British Patent Publication GB 1493048) was condensed with the azetidinone (l)(0.92g) in the manner described in Preparation 5(a). Chromatography (silica gel eluted with hexane/ethyl acetate) of the crude product afforded the title azetidinone (73) as a foam (0.85g), vmax (CHCI₃) 1745, 1670cm^-1; δppm (CDCI₃) 0.31, 0.32 (each 3H, s), 0.97 (9H, s), 4.55 (1H, d, J 1.7Hz), 5.02 (1H, d, J 1.7Hz), 6.90-7.55 (17H, m), 7.88 (1H, s); (Found [MNa]⁺632). Preparation 7(b)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-[hydroxy(imidazo [2,1-b]thiazol-6-yl)methyl]-4-tritylthioazetidin-2-one

The azetidinone (73) from Preparation 7(a)(0.85g) was dissolved in a mixture of dioxan (30ml) and ethanol (30ml) and the solution treated, portionwise, with sodium borohydride (75mg). Reaction was continued for one hour and the reaction mixture then concentrated to 50% volume. A further quantity of sodium borohydride (75mg) was added and reaction continued until no starting material remained, as shown by t.l.c. Ethyl acetate (150ml) and saturated brine solution (30ml) were added to the reaction with thorough mixing and the phases separated. The aqueous phase was extracted with a further quantity of ethyl acetate (30ml) and the organic phases combined, washed with saturated brine solution (50ml), dried (MgSθ4) and evaporated. This crude reaction product, as a two component mixture, was separated by chromatography (silica gel eluted with ethyl acetate/hexane). The major product (74) was the more polar isomer (0.45g); Rf (ethyl acetate) 0.3; v_max (CHCI₃) 1730cm^-1; δppm (CDCI₃) -0.03, 0.02 (each 3H, s), 0.72 (9H, s), 2.75 (1H, broad d, J 8.5Hz, exchanges on addition of D₂O), 3.74 (1H, dd, J 6.0Hz and 1.5Hz), 4.10 (1H, m, sharpens to a doublet on addition of D₂O, J 6.0Hz), 4.43 (1H, d, J 1.5Hz), 6.70-7.60 (18H, m); (Found [MNa]⁺634).

Preparation 7(c)

(3RS,4SR) 1-t-Butyldimethylsilyl-3-[acetoxy(imidazo

[2,1-b]thiazol-6-yl)methyl]-4-tritylthioazetidin-2-one

Azetidinone (74) (450mg) from Preparation 7(b) was dissolved in dry MDC (20ml) and 4-dimethyl- aminopyridine (97mg) added. This solution was chilled to 5°C and a solution of acetic anhydride (80μl) in dry MDC (2ml) added. After 0.5 hours at 5°C the cooling bath was removed and acetic anhydride (40μl) was added. Reaction was continued at room temperature for 0.5 hour when a further quantity of acetic anhydride (80μl) was added. When very little unreacted starting material remained, as shown by t.l.c, the reaction mixture was diluted with MDC (40ml) and washed with 1N HCl (30ml), water (30ml), saturated sodium bicarbonate solution (2x30ml), saturated brine (30ml), dried

(MgSθ4) and evaporated. Chromatography (silica gel eluted with ethyl acetate :hexane, 3:1) gave the title azetidinone (75) as a crisp foam (450mg); v_max (CHCI₃) 1740cm^-1; δppm (CDCI₃) -0.10, -0.03 (each 3H, s), 0.68 (9H, s), 2.01 (3H, s), 3.90 (1H, dd, J 5.5Hz and

2.0Hz), 4.34 (1H, d, J 2.0Hz), 5.21 (1H, d, J 5.5Hz), 6.70-7.70 (18H, m); (Found [MH]⁺654 and [MNa]⁺676).

Preparation 7(d)

(3RS, 4SR) 3-[Acetoxy(imidazo[2,1-b]thiazol-6-yl)- methyl]-4-tritylthioazetidin-2-one

The azetidinone (75)(440mg) from Preparation 7(c) was dissolved in dry THF (20ml) and the solution chilled to 5°C. Anhydrous potassium fluoride (78mg) followed by 18-crown-6 (39mg) dissolved in dry THF (5ml) were added. After 0.5 hour at 5°C the cooling bath was removed and the reaction stirred at ambient temperature overnight. The reaction mixture was then diluted with ethyl acetate (125ml), washed with brine (3 x 30ml), dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with hexane :ethyl acetate, 1:3) afforded the title azetidinone (76) as a crisp foam (350mg); v_max (CHCI₃) 3400, 1765 and.1735 (sh)cm^-1; δppm (CDCI₃) 2.13 (3H, s), 3.98 (1H, ddd, J 7.0Hz, 2.5Hz and 1.2Hz), 4.15 (1H, Broad s), 4.68 (1H, d, J 2.5Hz), 6.18 (1H, d, J 7.0Hz), 6.80-7.70 (18H, m); (Found CMH]⁺540).

Preparation 7(e)

(3RS, 4SR) 3-[Acetoxy(imidazo[2,1-b]thiazol-6- yl)methyl]-1-(1-p-nitrobenzyloxycarbonyl-1-triphenyl- phosphoranylidenemethyl)-4-tritylthioazetidin-2-one

The azetidinone (76)(1.45g) isolated as described in Preparation 7(d) was converted to the title phosphorane in the manner described in Preparation 5(e). Chromatography of the crude phosphorane ( silica gel eluted with a gradient of 66% to 100% ethyl acetate in hexane gave the title azetidinone (77)(1.28g), v_max (CHCl₃) 1750, 1620 and 1605cm^-1.

Preparation 7 ( f)

Silver (3RS, 4SR) 3-[Acetoxy(imidazo[2,1-b]thiazol- 6-yl)methyl]-1-(1-p-rnitrobenzyloxycarbonyl-1-triphenyl- phosphoranylidenemethyl)azetidin-2-one-4-thiolate

The phosphorane (77)(1.53g) from Preparation 7(e) was converted to the title silver salt as described in Preparation 5(f) to give the thiolate (78) as an off-white solid (1.08g), v_max (CHBr₃) 1740, 1645 and 1605cm^-1.

Preparation 7(g) p-Nitrobenzyl (5RS, 6SR) 6-[Acetoxy(imidazo[2,1-b] thiazol-6-yl)methyl]penem-3-carboxylate

The silver thiolate (78)(1.05c[) from Preparation 7(f) was converted to the title penem as described in Preparation 5(g). Chromatography (silica gel eluted with hexane-ethyl acetate-ethanol) afforded the product (79) as a cream coloured solid (0.34g); ν_max (CHCI₃) 1795, 1740 (sh), 1725, 1610cm^-1; δppm (CDCI₃) 2.10 (3H, s), 4.50 (1H, m), 5.28 and 5.43 (2H, ABq, J 13.5Hz), 6.24 (2H, m), 6.88 (1H, d, J 4.5Hz), 7.35 (1H, d, J 1Hz), 7.40 (1H, d, J 4.5Hz), 7.57 (1H, s), 7.60

(2H, d, J 9.0Hz), 8.24 (2H, d, J 9.0Hz); (Found CMH]⁺501 and CMNa]⁺523.)

Example 7(a) p-Nitrobenzyl (5RS) (Z)-6-(Imidazo[2,1-b]thiazol-6-yl

-methylene)penem-3-carboxylate

The acetoxy penem (79)(268mg) from Preparation 7(g) was reacted in the manner described in Example 5(a) to give a single isomer as shown by t.l.c, Rf(hexane:ethyl acetate, 1:3) 0.37; Rf(5% ethyl acetate/MDC) 0.20; Rf (10% ethyl cetate/MDC)0.32. The product crystallised from solution on work-up and was isolated by concentration and filtration of the extraction solvent to give the title penem (80) as bright yellow crystals (200mg), v_max (KBr) 1775, 1710 and 1675cm^-1; λ_max (EtOH) 312nm (ε 30,300); δppm C(CD₃)₂SO] 5.35 and 5.44 (2H, ABq, J 14Hz), 6.72 (1H, d, J 1Hz), 7.30 (1H, d, J 1Hz), 7.38 (1H, d, J 4.3Hz), 7.71 (2H, d, J 9Hz), 7.83 (1H, s), 7.99 (1H, d, J 4.3Hz), 8.20 (1H, s), 8.26 (2H, d, J 9Hz); (Found; M⁺440).

Example 7(b)

Sodium (5RS) (Z)-6-(Imidazo[2,1-b]thiazol-6-yl- methylene)penem-3-carboxylate

The penem ester (80) (60mg) from Example 7(a) was dissolved in a mixture of dioxan (50ml) and water (8ml). A 5% w/v solution of sodum bicarbonate (230μl) was added followed by 5% palladium on carbon (100mg) and the mixture hydrogenated at room temperature for 20 minutes. The mixture was then filtered on a celite pad, and the pad washed thoroughly with water. The filtrate was concentrated under reduced pressure to a small volume ( ca 5ml) and chromatographed on Biogel P2 using water as eluent. The appropriate fractions, as determined by ultra-violet spectroscopy, were combined and freeze-dried to give the title penem sodium salt (81), (22mg); ν_max (KBr) 1750, 1675, 1595cm^-1; λ_max (H₂O) 301nm (ε 24,750); δppm (D₂O) 6.55 (1H, s with fine coupling), 6.98 (1H, s), 7.03 (1H, s with fine coupling), 7.10 (1H, d, J 4.5Hz), 7.63 (1H, d, J 4.5Hz), 7.90(1H, s).

Preparation 8

Methyl Quinoxaline-2-carboxylate

Quinoxaloyl chloride (0.85g) was dissolved in a mixture of dry THF (50ml) and dry methanol (4ml). 2,6-Lutidine (0.51ml) dissolved in dry THF (10ml) was added dropwise with stirring. Reaction was continued for 30 minutes at room temperature then poured into ethyl acetate (250ml) and washed with water (3x50ml), brine (50ml), dried (MgSO₄) and evaporated to give the product as a pinkish solid (0.61g); ν_max (KBr) 1715cm^-1; δppm

[(CD₃)₂CO] 4.00 (3H, s), 8.00 (4H, m), 9.35 (1H, s).

Preparation 8 (a)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-(quinoxalin-2- ylcarbonyl)-4-tritylthioazetidin-2-one

Azetidinone (1)(0.92g) was condensed with methyl quinoxaline-2-carboxylate (0.50g) using the conditions described in Preparation 4(a). Chromatography (silica gel eluted with ethyl acetate :hexane, 1:1) gave the title azetidinone (82) as a yellow foam (1.14g); v_max (CHCl₃) 1745, 1685cm^-1; δppm (CDCI₃) 0.44 (6H, s), 1.06 (9H, s), 5.03 (1H, d, J2Hz), 5.40(1H, d, J2Hz), 6.70-8.40 (19H, m), 9.28 (1H, s). Preparation 8(b)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-[hydroxy (guinoxalin-2-yl)methyl]-4-tritylthioazetidin-2-one

The azetidinone (82) (1.10g) from Preparation 8(a) was reacted as described in Preparation 4(b) and the crude product, as a two component mixture, was subjected to chromatography (silica gel eluted with ethyl acetate/hexane). The more polar isomer (83)(0.30g) [Rf(ethyl acetate :hexane 1:1) 0.50] exhibited v_max (CHCI₃) 3400 (broad), 1740 and 1595cm^-1; δppm (CDCI₃) -0.01 and 0.01 (each 3H, s), 0.77 (9H, s), 3.65 (1H, d, J8Hz), 3.87 (1H, dd, J5.5Hz and 1.5Hz), 4.38 (1H, d, J1.5Hz), 4.78 (1H, dd, J8Hz and 5.5Hz), 7.10-8.20 (19H, m), 8.65 (1H, s); (Found CMNa]⁺640).

Preparation 8(c)

(3RS, 4SR) 3-CHydro2-y(quinoxalin-2-yl)methyl] -4-tritylthioazetidin-2-one

The more polar alcohol (83)(0.3g) from Preparation 8(b) was deprotected as described in Preparation 5(c) and the title azetidinone (84) was isolated as a cream coloured solid (0.16g); v_max (KBr) 3400, 1750cm^-1; δppm C(CD₃)₂SO] 3.87 (1H, t, J2.8Hz), 4.20 (1H, d, J3.0Hz), 5.13 (1H, dd, J5.0HZ and 3.0Hz), 6.34 (1H, d, J5.0Hz, exchanges on addition of D₂O), 6.9-7.40 (15H, m), 7.54 (1H, s, exchanges on addition of D₂O), 7.88 (2H, m), 8.06 (1H, m), 8.15 (1H, m), 8.90 (1H, s); (Found [MH]⁺504).

Preparation 8(d)

(3RS, 4SR) 3-[Acetoxy (quinoxalin-2-yl)methyl] -4-tritylthioazetidin-2-one

The azetidinone (84) from Preparation 8(c)(820mg) was reacted as described in preparation 4(d) and the crude product when subjected to chromatography (silica gel eluted with ethyl acetate:n-hexane, 1:1) gave the title azetidinone (85) as a cream coloured solid (830mg); %ιax (CHCI₃) 3400, 1775cm^-1; δppm(CDCl₃) 2.20 (3H, s), 4.02 (1H, ddd, J 5.4Hz, 2.8Hz, and 1.0Hz), 4.18 (1H, broad s), 4.62 (1H, d, J 2.8Hz), 6.40 (1H, d, J 5.4Hz), 7.10-7.50 (15H, m), 7.85 (1H, m), 8.20 (1H, m), 8.88 (1H, s); (Found CMH]⁺546).

Preparation 8(e)

(3RS, 4SR) 3-CAcetoxy (quinoxalin-2-yl)methyl]

-1-(1-p-nitrobenzyloxycarbonyl-1-triphenylphosphor- anylidenemethyl)-4-tritylthioazetidin-2-one

The acetoxyazetidinone (85) isolated as described in Preparation 8(d)(750mg) was converted to the title phosphorane in the manner described in Preparation 5(e). Chromatography of the crude product (silica gel eluted with a gradient of 50-65% ethyl acetate/hexane) gave the title azetidinone (6) as a cream coloured foam (850mg); v_max (CHCI₃) 1750,1735 (sh), 1615 and 1605cm^-1.

Preparation 8(f)

Silver (3RS, 4SR) 3-[Acetoxy(quinoxalin-2-yl) methyl]-1-(1-p-nitrobenzyloxycarbonyl-1-triphenyl phosphoranylidenemethyl)azetidin-2-one-4-thiolate

The phosphorane (86) from Preparation 8(e)(1.06g) was converted to the title silver salt as described in Preparation 5(e) to give the thiolate (87)(0.76g); ν_max (KBr) 1750, 1620 (sh), 1605cm^-1. Preparation 8(g) p-Nitrobenzyl (5RS, 6SR) 6-[Acetoxy(quinoxalin -2-yl)methyl]penem-3-carboxylate

The silver thiolate (87) (0.75g) from Preparation 8(f) was converted to the title penem as described in

Preparation 5(g). Chromatography (silica gel eluted with ethyl acetate :hexane, 1:1) afforded the product (88) as a yellow foam (0.33g); v_max(CHCl₃) 1790, 1745 and 1720cm^-1; δppm (CDCI₃) 2.20 (3H, s), 4.72 (1H, m), 5.30 and 5.44 (2H, ABq, J 13.6Hz), 6.19 (1H, d, J

2.3Hz), 6.61 (1H, d, J 3.6Hz), 7.37 (1H, d, J 1.0Hz), 7.60 (2H, d, J 8.8Hz), 7.75-8.35 (6H, m), 8.98(1H, s).

Example 8 p-Nitrobenzyl (5RS) (Z)-6-(Quinoxalin-2-yl methylene)penem-3-carboxylate

The penem (88) from Preparation 8(g) (181mg) was reacted in the manner described in Example 5(a) to give a single isomer as shown by t.l.c, Rf (5% ethyl acetate/MDC) 0.57; Rf (2% ethyl acetate/MDC) 0.09. The product was then isolated in the manner described in Example 7(a) to give the title penem (89) as a yellow crystalline solid (97mg); v_max (KBr) 1780, 1720cm^-1; λ_max (acetonitrile) 269nm (ε 38,550) and 333nm (ε 19,300); δppm C(CD₃)₂SO] 5.39 and 5.48 (2H, ABq, J14.0HZ), 6.90 (1H, d, J0.9Hz), 7.60-8.40 (10H, m), 9.22 (1H, s).

Preparation 9(a)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-(imidazo

[1,2-a]pyrazin-2-ylcarbonyl)-4-tritylthioazetidin -2-one

Ethyl imidazo[1,2-a]pyrazin-2-carboxylate [E. Abignente et al, Farmaco, Sci.Ed., 36, 61 (1981)], (1.8g) was condensed with the azetidinone (1) (4.37g) using the conditions described in Preparation 4(a). Chromatography (silica gel eluted with a gradient of 65-100% ethyl acetate/hexane) gave the title azetidinone (90) as a foam (1.28g); v_max (CHCI₃) 1750, 1685cm^-1; δ_ppm (CDCI₃) 0.3 (6H, 2xS), 0.93 (9H, s),

4.77 (1H, d, J 2Hz), 5.0 (1H, d, J 2Hz), 6.8-8.2 (18H, m), 9.14 (1H, s).

Preparation 9(b)

(3RS,4SR) 1-t-Butyldimethylsilyl-3-Chydroxy (imidazo[1,2-a]pyrazin-2-yl)methyl]-4-tritylthio- azetidin-2-one

The azetidinone (90)(2.16g) from Preparation 9(a) was reacted as described in Preparation 4(b) and the crude product as a three component mixture, was subjected to chromatography (silica gel eluted with ethyl acetate). The most polar component gave the title azetidinone (91)(0.86g); v_max (CHCI₃) 3350 (broad), 1730cm^-1; δ_ppm (CDCI₃) -0.08 and 0.02 (each 3H, s), 0.67 (9H, s),

2.82 (1H, broad s., exchanges on the addition of D₂O),

3.78 (1H, dd, J 6.0 and 1.5Hz), 4.37 (1H, m, sharpens to a doublet on the addition of D₂O, J 6.0Hz), 4.63 (1H, d, J 1.5Hz), 7.1-8.0 (18H, m), 8.95 (1H, s); (Found [MNa]⁺629).

Preparation 9(c)

(3RS, 4SR) 1-t-Butyldimethylsilyl-3-[Acetoxy(imidazo[1,2-a]pyrazin-2-yl)methyl]-4-tritylthioazetidin-2-one

The azetidinone (91) from Preparation 9(b) (820mg) was dissolved in dry MDC (40ml) and this solution chilled to 5°% and treated with dimethylaminpyridine (49mg), triethylamine (0.19ml), and the dropwise addition of a solution of acetic anhydride (0.4ml) dissolved in dry MDC (5ml). Reaction was continued for 1 hour at 5°C to ambient temperature then diluted with MDC (150ml) and washed with 1N HCl (50ml), water (50ml), saturated NaHCO₃ solution (50ml), brine (50ml), dried (MgSO₄) and evaporated. The crude product was subjected to chromatography ( silica gel eluted with ethyl acetate: n-hexane, 3:1) and the title azetidinone (92) isolated as a foam (630mg); v_max (CHCI₃) 1740cm^-1; δ_ppm (CDCI₃) 0.01 and 0.18 (each 3H, s), 0.80 (9H, s), 2.27 (3H, s), 4.15 (1H, dd, J 5.0 and 2.0Hz), 4.74 (1H, d, J 2.0Hz), 5.57 (1H, d, J 5.0Hz), 7.3-8.2 (18H, m), 9.18 (1H, s); (Found [MH]⁺649 _and [MNa]⁺671).

Preparation 9(d) (3RS, 4SR) 3-[Acetoxy(imidazo[1,2-a]pyrazin-2-yl) methyl]-4-tritylthioazetidin-2-one

The azetidinone (92)(620mg) from Preparation 9(c) was dissolved in dry THF (40ml) and the solution chilled to 5°C. Anhydrous potassium fluoride (260mg) followed by 18-crown-6 (120mg) dissolved in dry THF (4ml) were added. After 0.5 hours at 5°C the cooling bath was removed and the reaction stirred at ambient temperature for 3 days after which time t.l.c indicated complete reaction. The reaction mixture was then diluted with ethyl acetate (150ml), washed with brine (3 x 50ml) dried (MgSO₄) and evaporated. Chromatography (silica gel eluted with ethyl acetate) afforded the title azetidinone (93) as a colourless foam (450mg); ν_max (CHCI₃) 3400, 1770cm^-1; δ_ppm (CDCl₃)2.18 (3H, s), 3.94 (1H, m which collapses to a dd, J 6.8 and 2.6Hz, on the addition of D₂O), 4.21 (1H, broad s which exchanges on the addition of D₂O), 4.67 (1H, d, J 2.6Hz), 6.30 (1H, d, J 6.8Hz), 7.1 - 8.1 (18H, m), 9.16 (1H, s); (Found [MH]⁺ 535). Preparation 9(e)

( 3RS,4SR) 3-[Acetoxy(imidazo[1,2-a]pyrazin-2-yl) methyl]-1-(1-p-nitrobenzyloxycarbony1-1-triphenyl- phosphoranylidenemethyl)-4-tritylthioazetidin-2-one

The azetidinone (93) (480mg) isolated as described in Preparation 9(d) was converted to the title phosphorane in the manner described in Preparation 5(e). Chromatography of the crude phosphorane (silica gel eluted with ethyl acetate) gave the title azetidinone (94)(560mg); v_max (CHCI₃) 1750, 1615 and 1605cm^-1.

Preparation 9 (f)

Silver (3RS,4SR) 3-[Acetoxy(imidazo[1,2-a]pyrazin -2-yl)methy1-1-(1-p-nitrobenzyloxycarbony1-1-triphenyl- phosphoranylidenemethyl)azetidin-2-one-4-thiolate

The phosphorane (94)(525mg) from Preparation 9(e) was converted to the title silver salt as described in Preparation 5(f) to give the thiolate (95) as a cream coloured solid (360mg); ν_max (KBr) 1750 (broad), 1600 (broad) cm^-1.

Example 9(a)

P-Nitrobenzyl (5RS) (Z)-6-(Imidazo[1,2-a]pyrazin-2- ylmethylene)penem-3-carboxylate

A solution of the silver thiolate (95 ) (360mg) from Preparation 9(f) in a mixture of dry MDC (15ml) and dry acetonitrile (5ml) at 5°C was treated with dimethylaminpyridine (56mg) and acetic formic anhydride (0.34ml) followed by triethylamine hydrochloride (288mg). The mixture was stirred for 10 minutes at 5°C when a precipitate formed. The cooling bath was removed and stirring continued for 30 minutes. The precipitate was then filtered off on a celite pad and the pad washed thoroughly with MDC. The filtrate was washed with1N HCl (20ml), water (20ml), saturated

NaHCO₃ solution (20ml), brine (20ml) and dried (MgSO₄). The dried solution was then stirred under an atmosphere of argon for 30 minutes at 40°C after which time t.l.c. (ethyl acetate) indicated that cyclisation was complete. The MDC solution containing acetoxy penem was then chilled to -30°C and treated with a solution of 1,8-diazobicyclo[5.4.0]undec-7-ene (75mg) in dry MDC (2ml). Reaction was continued at -30°C for 1 hour after which time the reaction mixture was washed with 5% citric acid solution (50ml), saturated NaHCO₃ solution (2 x 50ml) brine (50ml), dried (MgSO₄) and evaporated. The crude product when subjected to chromatography (silica gel eluted with 1:1 ethyl acetate:MDC) resulted in the isolation of the title penem (96) as a bright yellow crystalline solid (75mg); v-max (KBr) 1790 and 1765, 1615 and 1600cm^-1; δ_ppm C(CD₃)₂SO] 5.37 and 5.44(2H, ABq, J 14Hz)m 6.80 (1H,s), 7.55 (1H, s), 7.73 (2H, d, J 8.7 Hz), 7.90 (1H, s), 7.96 (1H, d, J 4.6Hz), 8.26 (2H, d, J 8.7Hz), 8.48 (1H, s), 8.66 (1H, d, J 4.5Hz), 9.10 (1H, s).

Example 9(b) Sodium (5RS) (Z)-6-(Imidazo[1,2-a]pyrazin-2-yl methylene)penem-3-carboxylate

The penem ester (96)(53mg) from Example 9(a) was dissolved in a mixture of dioxan (60ml) and water (6ml) containing sodium bicarbonate (10mg). 5% Palladium on carbon (75mg) was added and the mixture hydrogenated at room temperature for 25 minutes. The mixture was then filtered on a celite pad and the pad washed thoroughly with water. The filtrate was concentrated under reduced pressure to a small volume (ca 3ml) and chromatographed on Biogel P2 using water as eluent. The appropriate fractions, as determined by ultra-violet spectroscopy, were combined and freeze-dried to give the title penem sodium salt (97), (33mg); ν_max (KBr) 1750, 1600cm^-1; λ_max (H₂O) 308nm (ε 11,250), 261nm (ε 16,550) and 221nm (ε 16,100); δ_ppm (D₂O) 6.62 (1H, s), 6.98 (1H, s), 7.17 (1H, s), 7.82 (1H, d, J 4.7Hz), 8.14 (1H, s), 8.35 (1H, d, J 4.7Hz), 8.93 (1H, s).

Biological data

The following table summarises the β-lactamase inhibitory activity of selected compounds according to the invention, as identified by Example numbers, against selected micro-organisms. The data is given in the form of the minimum inhibitory concentration (MIC) of amoxycillin in μg/ml when used in conjunction with 5 μg/ml of the respective compound according to. the invention. The MIC values for amoxycillin alone against the same organisms, and also an MIC values for the compounds according to the invention alone (given in brackets), are given for comparison purposes. In the Table 'NT' means 'not tested'.

Claims

Claims

1. A compound of the general formula I:

Λ

^ π o CO H aceutically acceptable salt or _in vivo le ester thereof

and R² denotes hydrogen, the other of R¹ and R² denotes an unsubstituted or substituted fused bicyclic hetero-aromatic group bonded through a carbon atom thereof and having five or six atoms in each ring, and

R³ denotes hydrogen or an organic group.

in which R¹, R² and R³ are defined as in claim 1.

3. A compound as claimed in claim 1 or claim 2, wherein the bicyclic hetero-aromatic group denoted by

R¹ or R² contains a bridgehead nitrogen atom.

4. A compound as claimed in any one of claims 1 to 3, wherein R¹ denotes a hydrogen atom and R² denotes the bicyclic hetero-aromatic group.

5. A compound as claimed in claim 1, which is selected from:

(5RS) (Z)-6-(benzo[b]fur-2-ylmethylene)penem-3- carboxylic acid;

(5RS) (Z)-6-[1-(4-methoxyphenylsulphonyl)-indol-

2-ylmethylene]penem-3-carboxylic acid;

( 5RS) ( Z)-6-(1-methylbenzotriazol-5-ylmethylene)- penem-3-carboxylic acid;

(5RS) (Z)-6-(N-methylthienotriazol-5-ylmethylene)- penem-3-carboxylic acid;

(5RS) (Z)-6-(pyrazolo[2,3-a]pyrid-2-ylmethyene)- penem-3-carboxylic acid; (5RS) (Z)-6-(thieno[3,2-b]fur-2-ylmethylene)- penem-3-carboxylic acid;

( 5RS ) (Z)-6-(imidazo[2,1-b]thiazol-6-ylmethylene)- penem-3-carboxylic acid;

( 5RS) (Z)-6-(quinoxalin-2-ylmethylene)penem-3- carboxylic acid;

(5RS) (Z)-6-(imidazo[1,2-a]pyrazin-2- ylmethylene)penem-3-carboxylic acid; as well as pharmaceutically acceptable salts and in-vivo hydrolysable esters thereof.

6. A process for the preparation of a compound of the general fomula I, as defined in claim 1, or a salt or ester thereof, which comprises eliminating the elements of a compound of the general formula XIII:

H-X^o XIII

from a penem or penem intermediate of the general part-formula XIV:

XIV

in which

R¹ and R² are defined as in claim 1, and

X^o denotes a hydroxy group or a leaving group,

to give a compound of the general part-formula XV:

XV

in which R¹ and R² are defined as in claim 1,

and, if the resulting compound of the general formula IV is a penem intermediate, converting it into a penem of the general formula I or a salt or ester thereof.

7. A pharmaceutical composition which comprises a compound as claimed in any one of claims 1 to 5, in admixture or conjunction with a pharmaceutically acceptable carrier.

8. A pharmaceutically composition as claimed in claim 7, which additionally comprises a penicillin cephalosporin or other β-lactam antibiotic.