IE44433B1 - Substituted n-methylene derivatives of thienamycin - Google Patents

Abstract

Amidines of the general formula (II) in which the substituents are defined in Claim 1, and their pharmaceutically acceptable salts, are obtained by treating thienamycin (I), while temporarily protecting the oxygen functions and/or the carboxyl function, with an imidoester or with an imidohalide, and, where appropriate, converting the resulting compounds into their pharmaceutically acceptable salts. The novel compounds, which can be prepared in accordance with the invention, are novel, valuable antibiotics which, as a rule, are active against various Gram-positive and Gram-negative bacteria, and may, therefore, be used in human and veterinary medicine.

Description

There is a continuing need for new antibiotics, since there is unfortunately no static effectiveness of a given antibiotic because continued wide-scale usage of any such antibiotic selectively gives rise to resistant strains of pathogens. In addition, the known antibiotics suffer from the disadvantage of being effective only against certain types of microorganisms. Accordingly, the search for new antibiotics continues.

This invention is concerned with certain substit10 uted N-methylene derivatives of the antibiotic thienamycin and its position isomers, and their pharmaceutically acceptable salt, ether, ester, and amide derivatives, compounds in accordance with the present Invention have been found to have utility as broad-spectrum antibiotics and this invention also concerns processes for the preparation of such compounds and pharmaceutical compositions containing them.

Thienamycin is disclosed and claimed in.U.S.

Patent No. 3,950,357 and Patent Specification No. 42067.

Thienamycin may be used as a starting material in the preparation of the compounds of the present invention.

Thienamycin is known to have the formula: 14 3 3 and its position ispmers may be similarly represented. The substituted N-methylene derivatives of the present invention have the following formula: SCHCH„N=C-X z z I COOH Y II or are salts, esters (with the carboxy or the hydroxy group), ethers, amides or anhydrides of compounds of formula II, or the corresponding thioic S-acids or their salts, esters, ethers or anhydrides. Depending upon the basicity of the amino nitrogen (a function of the identity of the methylene substituents X and Y), Formula II may equivalently be represented as an inner salt: and, when the carboxy group is esterified, this tautomer 15 can also form acid-addition salts. Such tautomers are intended to be included when the generic formula II is mentioned. The inner salt is one canonical form of a single resonant structure, which, for example, when Y is -NR-'-R2 and X is R, is: £33 For convenience, the compounds of the present invention may also be represented by the symbol: i- OH Th --N=C-X I Y _ COOH II where Th indicates the bicyclic nucleus of thienamycin or its position isomer and its hydroxyl, amino, and carboxyl functional groups are shown. In the foregoing 12 formulae, X is -OR, -SR or -NR R and Y, independently 1 2 of X, is hydrogen, -R, -OR, -SR, or -NR R ; where each 12 of R and R , independently of the other, is a hydrogen atom, a radical of the type defined below for R, or a nitro, hydroxyl, cj__g alkoxyl, amino, g monoalkylamino, di(C, , alkylJamino, or tri(C, , alkylJamino radical, or 1 2 R and R are joined together to form, together with the nitrogen atom to which they are attached, a substituted or unsubstituted monocyclic or bicyclic heteroaryl or non-aromatic heterocyclic radical comprising 4-10 atoms one or more of which may be an additional oxygen, sulphur or nitrogen hetero atom; R is a cyano or carboxy radical or a substituted or unsubstituted carbamoyl; alkoxycarbonylr 0χ_10 alkyl; C2_1Q alkenyl; C2_1Q alkynyl; C3_^q cycloalkyl; 12 eycloalkylalkyl; C4_12 cYclo" alkylalkenyl; C3_lo cycloalkenyl; c4_2_2 cycloalkenyl4 4 4 3 3 alkenyl; c4_]_2 cycloalkenylalkyl; c6_10 aryl; c7„16 aralkyl; c8_yg aralkenyl or 0θ_1β aralkynyl radical or a substituted or unsubstituted monocyclic or bicyclic heteroar(C^ g alkyl), heteroaryl, non-aromatic hetero5 cyclic(C^_g alkyl) or non-aromatic heterocyclic radical having 4 to 10 ring atoms one or more of which is an oxygen, sulphur or nitrogen heteroatom; the substituent(s) 12 on R, R , R or the ring formed by joining R and R , are halogen, viz. chlorine, bromine, iodine or fluorine; azide; thio; sulpho; phosphono; cyanothio (-SCN); nitro; cyano; amino; hydrazino; C, . alkylamino, di(C. , alkyl) 1—b 1—b amino, tri(C. alkyl)amino; C. r alkylhydrazino, diCC, r l-o 1—b 1—b alkyl)hydrazino, trifC, alkyl)hydrazino, hydroxyl; 1—b C1-4 alkyl; cj_6 alkoxyl; cj__5 alkylthio; carboxyl; oxo; (C^ g alkoxy(carbonyl; acyloxy; carbamoyl; (01-4 alkyl)carbamoyl or di(C1-4 alkyl)carbamoyl.

Compounds of the present invention can be represented by the following formula: OR which may also exist as a salt: .3 3' Y .COX'R Ila Ila 4433 or, more conveniently by the previously introduced symbol: Th Θ -NH=C-X A© Y - COX'R3' lla The non-critical counter anion, A, is representatively selected to provide pharmaceutically acceptable salts such as a halide, e.g. chloride or bromide, sulfate, phosphate, citrate, acetate or benzoate. 3' 31 X' is oxygen, sulphur, NH or NR . R is hydrogen, or, inter alia, is representatively selected to provide the pharmaceutically acceptable salt, ester, anhydride 3 1 (R is acyl and X1 is oxygen), and amide residues known in the bicyclic , β-lactam antibiotic art and enumerated in greater detail below. R is: (1) acyl (generically the group OR is classifiable as an ester); (2) R is a univalent hydrocarbon or substituted hydrocarbon residue, e.g. alkyl, aryl, alkenyl or aralkyl (such that the group OR is generically classifiable as an ether) or (3) hydrogen. The term acyl includes the alkanoyls and their derivatives and analogues such as thio analogues in which the carbonyl oxygen is replaced by sulphur; as well as sulphur and phosphorous acyl analogues such as substituted sulfonyl and sulfinyl radicals, and substituted P(III and V) radicals such as substituted phosphorous, phosphoric, phosphonous and phosphonic radicals. Such 3 radicals, R , of the present invention are enumerated in greater detail below.

Antibiotics of the present invention have been found to be active against a broad range of pathogens which representatively include both gram-positive bacteria as S. aureus. Strep. pyogenes, and B. subtilis and gram5 negative bacteria such as E. coli, Proteus morganii, Klebsiella, Serratia, and Pseudomonas.

The compounds of the present invention (Formulae II and Ila, above) may be divided into four classes: 2 (1) Amidines, in which Y = -NR R and X = hydrogen or R: SCH„CHON=C-NR1R2 2 2 | COOH R which may be represented by the resonant structure: 2 where R, R and R are as previously defined. Species 15 Ila may similarly be depicted: 3' where R , X1 and R are as defined below Representative examples of such amidine embodiments (the substituent plus the amino group of thienamycin form the amidine structure) are: The benzamidine: The formamidine: The acetamidine: Y = X = P^enyT; Y = -NH2, X = H; Y = -nh2, X = ch3? The 4-pyridylcarboxamidine: Y = -NH2, X = 4-pyridyl; The N-isopropyl formamidine: Y = NHCH(CH3)2, X = H; The N-methyl formamidine: Y = -NHCH^, X = H; The Ν,Ν-dimethylformamidine: Y = -N(CH3)2, X = H; The piperidinyl methylenimine: Y = 1-piperidyl, X = H. 2 Other preferred amidines are those in which Y = NR R and 1 2 each of R and R , independently of the other, is a hydrogen atom; a substituted or unsubstituted C, . alkyl, C3 5 alkenyl, C3 cycloaikyl, ? cycloalkylalkyl, C^_6 cycloalkenyl, C4_7 cyeloalkenylalkyl, C? aralkyl or cg_q0 aralkenyl radical or a monocyclic heteroar(C^_3 alkyl) radical having 5-6 ring atoms, one or more of which is oxygen, sulphur, or nitrogen, the ring or chain 1 2 ;substituent(s) of R and R being chlorine, fluorine, hydroxyl, C, , alkoxy, di(C, - alkyl)amino or Cq , alkyl3,-3 ^--3 1 2 thio. Examples of the values of R and R are methyl, ethyl, propyl, isopropyl, butyl, t-butyl, N,N-dimethylaminoethyl, 2,2,2-trifluoroethyl, 2-methylthioethyl, allyl, methallyl, 2-butenyl, l-buten-3-yl, cyclopropyl, cyclopentyl, eyclohexyl, cyclopropylmethyl, cyclohexylmethyl, 2-cyclopropenyl, 1,4-cyclohexadienylmethyl, benzyl, pmethoxybenzyl, p-dimethylaminobenzyl, cinnamyl, 2-thienylmethyl, 3-thienylmethyl, 2-furylmethyl, and 1-methyl-5tetrazolylmethyl.

X is a hydrogen atom or a residue of the type defined above for R, preferably C, .. alkyl, C. , aminoalkyl, C_ r X—b X—b 2—b alkenyl, C_ , aminoalkenyl, C_ , alkoxyalkyl, C„ (mono, 2—0 2—0 2 — X2 di or tri)-alkylaminoalkyl, C, , perhaloalkyl, C„ c alkylthioalkyl, or 3 substituted or unsubstituted aryl, aralkyl, monocyclic heteroaryl, heteroar(0^_θ alkyl), non-aromatic heterocyclic or non-aromatic heterocyclic-(C alkyl)ring 1-b in which any substituent(s) are as set out in the definition 1 2 of R and R . More specific examples of preferred values of X are methoxymethyl, ethoxyethyl, dimethylaminomethyl, methylaminomethyl, trimethylammoniummethyl, trifluoromethyl, methylthiomethyl, ethylthioethyl, phenyl, benzyl, phenethyl, 2-pyridyl, 3-pyridyl, 4-pyridyl and 2-thiazolyl.

The amidine compounds represent a preferred class, 31 particularly when X' is oxygen and R and R are both hydrogen. Especially preferred amidines of the present 1 2 invention are those in which Y is -NR R and X is R, 2 where each of R , R and R is a hydrogen atom or a substituted or unsubstituted alkyl or alkenyl radical or the type mentioned in the immediately preceding paragraph. 2 (2) Guanidines, in which Y is -NR R and X is Th OR „ „ „„1„2 N=C-NR R I 1 2 NR R COX'R where all symbols are as defined above.

Representative examples of such guanidine compounds (the substituted plus the amino group of thienamycin form the guanidine structure) are: - 10 The guanidine: Y = -NH^, X = -NH2? The N-methylguanidine: Y = -NHCE^, X = -NHj,· The N,N-dimethylguanidine: Y = -N(CHg)2, X = -NH2; The Ν,Ν,Ν-trimethyl guanidine: Y = -Ν(0Η^)2, X = -NHCH3? The N-phenylguanidine: Y = -NH(CgH5), X = -NH2' The nitroguanidine: Y = -NHNOj, X = _aH2: The aminoguanidine: Y = -NHNH2, X = -NH2.

Other preferred guanidines are those in which X1 is oxygen, 3' 12 R and R are both hydrogen, and each of R and R , independently of the other, is a hydrogen atom or a substituted or unsubstituted C. r alkyl, C. , alkenyl, X—o 3—0 C3_g cycloalkyl, C4_y cycloalkylalkyl, cycloalkenyl, Cd_7 cycloalkenylalkyl, c7_10 aralkyl or Οθ aralkenyl radical or a monocyclic heteroar(C^ alkyl) radical having 5 or 6 ring atoms, one or more of which is oxygen, sulphur, and/or nitrogen, and the substituent(s) is/are chlorine, fluorine, hydroxy, alkoxy, t3i(C^ alkyl) amino or C,_ alkylthio and may be ring or chain substit. 1 2 uents. Specific examples of values of R and R are methyl, ethyl, isopropyl, butyl, t-butyl, N, N-dimethylaminoethyl, 2,2,2-trifluoroethyl,: 2-methylthio ethyl, allyl, methallyl, 2-butenyl, l-buten-3-yl, cyclopropyl, cyclopentyl, cyclohexyl, cyclopropylmethyl, oyclohexyl25 methyl, 2-cyclopropenyl, 1,4-cyclohexadienylmethyl, benzyl, p-methoxybenzyl, p-dimethylaminobenzyl, cinnamyl, 2thienyImethyl, 3-thienylmethyl, 2-furylmethyl and 1-methyl5-tetrazolylmethyl. 2 (3) Substituted Pseudoureas, in v/hich Ϋ is -NR R and X is -OR or -SR: 4443 Th OR N=C-NR1R2 I X COX'R3' 3' 1 2 where R , R , X', R , and R are as defined, and X is -OR or -SR.

Representative examples of such substituted pseudourea compounds (the substituent plus the amino group of thienamycin form the substituted pseudourea structure) are The Ν,Ν-dimethyl-O-methyl pseudourea: Y = X = -0CH3; The N, N-dimethyl-S-ethyl pseudothiourea: Y = -n(ch3)2, x = -sch2ch3,· The N-phenyl-S-ethyl pseudothiourea: Y = -ΝΗΟθΗ^, X = -SCH2CH3; The N-methyl-S-methyl pseudothiourea: Y = -NHCHy X = -sch3.

Particularly preferred compounds are those in which 3 31 R and R are hydrogen and X' is oxygen. (4) imido Ethers, and Imido Thioethers Th OR -N=C-X I Y L COX'R3 31 where R , X‘, R and R are as defined, X is -OR or -SR 20 and Y is hydrogen, R, -OR and -SR.

Representative examples of such imido ester and imido thioester compounds (the substituent plus the amino 4433 group of thienamycin form the imido ester or imido thioester structure) are; The methyl formimidate: Y = -OCH^, X = -H; The S-methyl thiobenzimidate: Y = -SCH^, X = phenyl.

The methyl benzyloxycarbimidate: Y = -OCH^, X = -OCHCH; b 5 The diethyl dithiocarbimidate: Y = -S-CjH^, x=-s-c2h5.

The compounds of the present invention are conveniently prepared from thienamycin and from position isomers of thienamycin, which are discussed later on in the specification. These compounds have Formula IA, above. Compounds of the present invention such as Ila, above, in which the secondary alcoholic group and/or the carboxyl group are derivatized, are conveniently prepared either from the corresponding 0-, carboxyl, or 0- and carboxyl derivative of thienamycin or its position isomer or from II or thienamycin or its position isomer followed by 3' subsequent reaction to introduce the radicals R and R 3' (or -X' R ) and combinations thereof. Such starting materials are-fully disclosed in the specifications of the following copending Patent Applications: •British Patent Specification. Νΰϊ .1,559,234 is directed to ©-derivatives (ester and ether derivatives) c£ the secondary alcoholic group, which have the following structural formula: sch2ch2nh2 COOH Ia 0^ British Patent Specification No. 1 570 985 is directed to N-acyl derivatives, which have the following structural formula: OH 1' 2 ' SCH2CH2NR r COOH Ib 11 2 ' where R is acyl and R is hydrogen or acyl. Such N-acyl compounds are useful starting materials for the preparation of the substituted pseudourea (3) and the imido ether and imido thioether (4) compounds of the present invention.

British Patent Specification No. 1 570 989 is directed to N-acyl and carboxyl derivatives having the following structural formula: British Patent Specification No. 1 570 987 is directed to N-acyl, 0and carboxyl derivatives of thienamycin having the following structural formula: The corresponding O-unsubstituted compounds (herein called Ic) can be prepared analogously.

Thus compounds of the present invention depicted as Ila, above, may be prepared by starting with the corres5 ponding derivative la, lb, Ic, Id, Ie; or compounds Ila may be prepared directly starting with Thienamycin or a position isomer thereof, I, (I—>11) followed by the desired derivatization procedure to establish R and/or 3 1 X'R (II—»IIa) which is described in the above-mentioned British Patent Specifications. 33' J' 2' The radicals R , R , X', R and R are defined as set forth below.

In the generic representation of the compounds of the present invention (Ila, above), the radical represented by -COX'R3 is, inter alia, -COOH (X' is oxygen and R3 is hydrogen) and all radicals known to be effective as pharmaceutically acceptable ester, anhydride (R is acyl) and amide radicals in the bicyclic β-lactam antibiotic art, such as the cephalosporins and penicillins and the nuclear analogues thereof. 3' Suitable radicals (R ) include conventional protecting or carboxyl blocking group. The term "blocking group as utilized herein is used in the same manner and in accordance with the teaching of U.S. Patent 3,697,515.

Pharmaceutically acceptable thienamycin derivatives of the present invention falling in this class are given below. Suitable blocking esters thus include those selected from the following list which is representative and not intended to be an exhaustive list of possible 3' ester groups, in which X is oxygen and R is given; - 15 (i) R3 is CR-R-R- where at least one of R-, R¢5 and R- is an electron-donor, e.g., p-methoxyphenyl, 2,4,6tr imethy lphenyl, 9-anthryl, methoxy, C^SCH^, tetrahydrofur2-yl, tetrahydropyran-2-yl or fur-2-yl. The remaining R-, to c R- and R- groups may be hydrogen or organic substituting groups. Suitable ester groups of this type include pmethoxybenzyloxycarbonyl and 2,4,6-trimethylbenzyloxycarbonyl. (ii) R3 is CR-R-R- where at least one of R-, Rc and R- is an electron-attracting group, e.g., benzoyl, p-nitrophenyl, 4-pyridyl, trichloromethyl, tribromomethyl, iodomethyl, cyanomethyl, ethoxycarbonylmethyl, arylsulphonylmethyl, 2-dimethylsulphoniummethyl salt, o-nitrophenyl or cyano. Suitable esters of this type include benzoylmethoxycarbonyl, p-nitrobenzyloxycarbonyl, 4-pyridylmethoxycarbonyl, 2,2,2-trichloroethoxycarbonyl and 2,2,2-tribromoethoxycarbonyl. (iii) R3 is CR-R-R- where at least two of R-, Rc and R- are hydrocarbon such as alkyl, e.g., methyl or ethyl, or aryl, e.g., phenyl and the remaining R-, R- or R-, if there is one, is hydrogen. Suitable esters of this type include t-butyloxyearbonyl, t-amyloxycarbonyl, diphenylmethoxycarbonyl and triphenylmethpxycarbonyl. 1 (iv) R is adamantyl, 2-benzyloxyphenyl, 4-methylthiophenyl or tetrahydropyran-2-yl.

Silyl esters, under this category of blocking groups, may conveniently be prepared from a halosilane or a silazane of the formula: R43SiX'? R42SiX'2? R4 Si.NR4/ R^Si.NH.COR4; R4 Si.NH.CO.NH.SiR4 ; R4NH.CO.NH.SiR4 ; or R4C(0SiR4,); •3 «j w □ _ 16 HN(SiR^_)„ where X' is a halogen such as chlorine or bromine J - 4 and the various groups R , which can be the same or different, represent hydrogen, alkyl, e.g. methyl, ethyl, n-propyl, iso-propyl; aryl, e.g. phenyl; or aralkyl, e.g., benzyl.

More generally stated, pharmaceutically acceptable carboxyl derivatives of the present invention are those derived by reacting thienamycin or an N-protected thienamycin, such as N-acylated thienamycin, or a position isomer thereof with alcohols, phenols, mercaptans, thio phenols or acylating reagents.

For example, esters and amides of interest are the above-listed starting materials and final products having the following group at the 2-pOsition of the thienamycin 3' 3 1 nucleus: -COX'R where X is oxygen, sulfur, NH or NR 3' and R is alkyl having 1-10 carbon atoms, straight or branched, such as methyl, ethyl, t-butyl, pentyl and decyl; earbonyImethyl including phenacyl, g/bromophenacyl, p.-tbutylphenacyl, acetoxyacetyImethyl, pivaloxyacetylmethyl, carboxymethyl and its alkyl and aryl esters; a-carboxyα-isopropyl; aminoalkyl including 2-methylaminoethyl, 2-diethylaminoethyl, 2-acetamidoethyl, phthalimidomethyl and succinimidomethyl; (C, alkoxy)-(C, . alkyl) in which the alkox/ residue can be branched, straight or cyclic, such as methoxymethyl, ethoxymethyl, isopropoxymethyl, decyloxymethyl, ethoxypropyl, decyloxypentyl or cyclohexyloxymethyl; (C1_g alkanoyloxy)-(C1_g alkyl) in which the alkanoyloxy portion is straight or branched, such as acetoxymethyl, pivaloyloxymethyl, acetoxyethyl, propionyloxyethyl, or acetoxypropyl; halogenated straight or branched C, . alkyl in which the halogen is iodine, chlorine, bromine and/or fluorine, e.g. 2,2,2-trichloro- 17 ethyl, tri-fluoroethyl, 2-bromopropyl, diiodomethyl, 2-chloroethyl, or 2-bromoethyl, and also including C, χ—6 perhaloalkyl; alkenyl having 2-10 carbon atoms, either straight or branched, e.g., allyl, 2-propenyl, 3-butenyl, 4-butenyl, 4-pentenyl, 2-butenyl, 3-pentenyl, 2-methyl-2propenyl, 3-methyl-3-butenyl, methallyl and 1,4-cyclohexadien-l-yl-methyl; alkynyl having 2-10 carbon atoms, either straight or branched, e.g., 3-pentynyl, propargyl, ethynyl and 3-butyn-l-yl; alkanoyl, either straight or branched, having 1-10 carbon atoms, such as pivaloyl, acetyl and propionyl; c3_j4 alkoxycarbonylalkyl, C4_21 dialkylaminoacetoxyalkyl; C2_]_3 alkanamidoalkyl; aralkyl in which the alkyl residue has 1-3 carbon atoms and the aryl residue 6-10 carbon atoms, such as benzyl, benzhydryl, substituted benzyl or benzhydryl, e.g., benzyl or benzhydryl substituted with 1-3 substituents such as benzyl, phenoxy, halo, 4 alkyl, alkanoyloxy of 1-5 carbon atoms, alkoxy, hydroxy, nitro, blocked carboxy# or combinations thereof, e.g., p-chlorobenzyl, p-nitrobenzyl, 3,5dxnitrobenzyl, p-methoxybenzyl, m-benzoylbenzyl, p-t-butylbenzyl, m-phenoxybenzyl, p-benzoylbenzyl, p-nitrobenzyl, 3,5-dichloro-4-hydroxybenzyl, p-methoxycarbonylbenzyl, pmethoxybenzhydryl, p-carboxybenzyl (the latter being either the free acid, ester or the sodium salt), 2,4,6trimethylbenzyl, p-pivaloyloxybenzyl, p-t-butoxycarbonyl benzyl, p-methylbenzyl, p-benzoyloxybenzyl, p-acetoxybenzyl, p-2-ethylhexanoylbenzyl, p-ethoxyoarbonylbenzyl, p-benzoylthiobenzyl, p-benzamidobenzyl, p-pivaloyloxybenzyl, m-p'ivaloyloxybenzyl, p-isopropoxybenzyl or p-tbutoxybenzyl, as well as cyclic analogues thereof; monocyclic and bicyclic heteroaralkyl in which there are 4 to 10 atoms in the ring, the hetero atom or atoms being oxygen, sulfur and/or nitrogen, and 1 to 6 carbon atoms in the alkyl chain, e.g. 2,2-dimethyl-5-coumaranmethyl, 5-indanylmethyl, p-trimethylsilylbenzyl, 3,5-bist-butoxy-4-hydroxybenzyl; 2-thienylmethyl, 2-furyImethy1, 3-t-butyl-5-isothioazomethyl, 6-pivaloyloxy-3-pyridazinylethyl or 5-phenylthio-1-tetrazolylmethyl; phthalidyl; phenylethyl, 2-(p-methylphenyl)ethyl, and their arylthioalkyl analogues; aryloxy-C^_g alkyl) where aryl is prefer ably a phenyl ring having 0-3 substituents, preferably 0 or 1 substituent in the ortho or para positions, e.g., (4-methoxy)phenoxymethyl, phenoxymethyl, (4-chloro)phenoxymethyl, (4-nitro)phenoxymethyl, (4-benzyloxy)phenoxymethyl, (4-methyl)phenoxymethyl, (2-methoxy)phenoxy methyl, (1-phenoxy) ethyl, (4-amino)phenoxymethyl, (4-methoxy)phenylthiomethyl, (4-chloro)phenylthiomethyl or phenylthioethyl; phenyl, 5-indanyl, substituted phenyl having 1-3 substituents, preferably one substituent in the ortho or para position, e.g., (4-methyl)phenyl, (4-hydroxy)phenyl, (4-t-butyl)phenyl, p-nitrophenyl, 3,5-dinitrophenyl or p-carboxyphenyl, the latter having either the free acid or the sodium salt form; phenyl(C_ . alkenyl), such as 3-phenyl-2-propenyl; benzyloxy(C1_3 alkyl) such as benzyloxymethyl, (4-nitro)benzyloxymethyl or (4-chloro)benzyloxymethyl; alkylthioalkyl where the alkylthio residue has 1-10 and preferably 1-6 carbon atoms, and can be branched, straight or cyclic, and the alkyl portion has 1-6 carbon atoms, such as methyl thioethyl, ethylthioethyl, cyclohexylthiomethyl, decylthio butyl, methylthiopropyl, isopropylthioethyl,methylthiobutyl, acetamidomethyl, acetyl thioethyl, pivaloylthiomethyl or methylthiomethyl.

In addition to the esters and thio esters listed above, amides are also embraced by the present invention, R' i.e., compounds where X* is the -N- group. Representative of such amides are those in which R1 is hydrogen, methyl, ethyl, phenyl, p-methoxyphenyl, benzyl, carboxymethyl, 1 methylthioethyl, or heteroaryl? also embraced by -COX'R 3' are anhydrides, in which R is acyl, for example, benzyloxycarbonyl, ethoxycarbonyl, benzoyl, or pivaloyl. 1 The preferred -COX'R radicals of the present invention are those in which, in Formula lla above, X' is oxygen, sulphur or NR' (R' is hydrogen or C. , alkyl); * and R is C, . alkyl, C_ , alkenyl, such as methallyl, l-o 2-b 3-methylbutenyl or 3-butenyl; methylthioethyl; benzyl or substituted benzyl, such as p-t-butylbenzyl, m-phenoxybenzyl, p-pivaloyloxybenzyl or p-nitrobenzyl; pivaloyloxymethyl, 3-phthalidyl, acetoxymethyl, propionyloxymethyl, acetylthiomethyl, pivaloylthiomethyl, allyl, 4-butenyl, 2-butenyl, 3-methyl-2-butenyl, phenacyl, acetoxyacetylmethyl, methoxymethyl, p-acetoxybenzyl, p-pivaloyloxybenzyl, p-isopropoxybenzyl, 5-indanylmethyl, 5-indanyl, benzyloxymethyl, ethylthioethyl, methylthiopropyl, methoxycarbonyloxymethyl, ethoxycarbonyloxymethyl, dimethylaminoacetoxymethyl, crotonolacton-3-yl, or acetamidomethyl.

In Formula Ila above, R is, in addition to hydrogen, (1) acyl (generically the group -OR3 is class3 ifiable as an ester); or (2) R is a radical (e.g. alkyl, aryl or aralkyl) such that the group -OR3 is classifiable as an ether. For the esters, (1) R is selected from the following definition of acyl radicals, (p = 1). In the so-called ethers (2) of the present invention, R is selected from the same acyl radicals »4433 - 20 ο II but in which the carbonyl residue, -C-, or more generally X II 3 -C- , is omitted (p = 0). Thus, R is selected from the following radicals where all symbolism is as defined below and ρ Ϊ3 1 or 0. 2' R is hydrogen or acyl and R is acyl and, when acyl, they are selected from the above radicals in which p = 1. 21 1' 3 Thus, R , and when acyl, R and R , can, inter alia, be a substituted or unsubstituted aliphatic, aromatic, heterocyclic, araliphatic or heterocyclylaliphatic carboxylic acid radical, a substituted or unsubstituted carbamyl radical·' or a carbothioic acid radical. One group of acyl radicals can be represented by the general formula: X II — C—R'1 where X is 0 or S and R'1 represents hydrogen? amino? - 21 substituted amino such as C, , alkylamino or (C, , alkyl)t J.—O A —O amino; substituted or unsubstituted straight or branchedchain C, , alkyl; mercapto; aryloxy, typically comprising l—o to 10 carbon atoms; alkenyl or alkynyl, typically comprising 2 to 6 carbon atoms; aryl, such as phenyl; aralkyl, such as benzyl; cycloaikyl, typically comprising 3 to 6 carbon atoms; or a heteroaryl or heteroaralkyl group (mono- and bicyclic) in which the alkyl residue (if any) typically comprises 1 to 3 carbon atoms and the heterocyclic ring typically comprises 4-10 atoms and the hetero atom or atoms is/are preferably 0, N and/or S; such above-listed groups can be unsubstituted or can be 8 8 substituted by radicals sueh as OH, SH, SR (R is g alkyl or aryl such as phenyl), alkyl or alkoxy groups having 1 to 6 carbon atoms, halogens, viz. Cl, Br, F or I, cyano, carboxy, sulfamino, carbamoyl, sulfonyl, azido, amino, substituted amino, such as (C. , alkyl)-substituted A—v cimino including quaternary ammonium, C„ , halogenated l—o alkyl such as trifluoromethyl, carboxy(C^ alkyl), carbamoyl(C, . alkyl), N-substituted carbamoyl(C. . l—o l—o alkyl), amidino, guanidino, N-substituted guanidino or guanidino-(C alkyl). Representative examples of l—o such acyl groups that might be mentioned are those in which R'' is benzyl, p-hydroxybenzyl, 4-amino-4-carboxybutyl, methyl, cyanomethyl, 2-pentenyl, n-amyl, n-heptyl, ethyl 3- or 4-nitrobenzyl, phenethyl, β,p-diphenylethyl, methyldiphenylmethyl, triphenylmethyl, 2-methoxyphenyl, 2,6-dimethoxyphenyl, 2,4,6-trimethoxyphenyl, 3,5-dimethyl4-isoxazolyl, 3-butyl-5-methyl-4-isoxazolyl, 5-methyl-3phenyl-4-isoxazolyl, 3-(2-chlorophenyl)-5-methyl-4isoxazolyl, 3-(2,6-dichlorophenyl)-5-methyl-4-isoxazolyl, D-4-amino-4-carboxybutyl, D-4-N-benzoylamino-4-carboxy-n44433 - 22 butyl, p-aminobenzyl, p-aminobenzyl, m-aminobenzyl, pdimethylaminobenzyl, (3-pyridyl)methyl, 2-ethoxy-lnaphthyl, 3-carboxy-2-quinoxalinyl, 3-(2,6-dichlorophenyl)-5-(2-£uryl)-4-isoxazolyl, 3-phenyl-4-isoxazolyl, -methyl-3-(4-guanidinophenyl)-4-isoxazolyl, 4-guanidino methylphenyl, 4-guanidinomethylbenzyl, 4-guanidinobenzyl 4-guanidinophenyl, 2-6-dimethoxy-4-guanidinophenyl, o-sulfobenzyl, p-carboxymethylbenzyl, p-carbamoylmethylbenzyl, m-fluorobenzyl, m-bromobenzyl, p-chlorobenzyl, p-methoxybenzyl, 1-naphthylmethyl, 3-isothiazolylmethyl, 4- isothiazolylmethyl, 5-isothiazolyImethyl, guanylthiomethyl, 4-pyridylmethyl, 5-isoxazolyImethyl, 4-methoxy5- isoxazolyImethyl, 4-methyl-5-isoxazolylmethyl, 1imidazolylmethyl', 2-benzofuranyImethyl, 2-indolylmethyl, 2-phenylvinyl, 2-phenylethynyl, 1-aminocyclohexyl, 2and 3-thienylaminomethyl, 2-(5-nitrofuranyl)-vinyl, phenyl, p-methoxyphenyl, p-chlorophenyl, p-phenylphenyl, p-aminomethylbenzyl, l-(5-cyanotriazolyl)-methyl, difluoromethyl, dichloromethyl, dibromomethyl, 1-(320 methylimidazolyljmethyl, (2 or 3)-(5-carboxymethylthienyl)methyl, (2 or 3)-(4-carbamoylthienyl)methyl, (2 or 3)-(5-methylthienyl)methyl, (2 or 3)-(methoxythienyl) methyl, (2 or 3)-(4-chlorothienyl)methyl, (2 or 3)-(5-sulfothienyl)methyl, (2 or 3)-(5-carboxythienyl)25 methyl, 3-(1,2,5-thiadiazolyl)methyl, 3-(4-methoxyl,2,5-thiadiazolyl)methyl, 2-furylmethyl, 2-(5-nitrofuryl)methyl, 3-furylmethyl, 2-thienylmethyl, 3-thienylmethyl, tetrazolylmethyl, benzamidinomethyl and cyclohexylamidinomethyl.

The acyl group can also be a radical of the formula: X X II 'I * —C(CH ) ZR or —C—R* n where X is 0 or S and n is 0, 1, 2, 3 or 4, 2 represents oxygen, sulfur, carbonyl or nitrogen, R’1 is defined as A above and R' is as defined below. Representative members of the substituent —(CH,) ZR'1 2 n that might be mentioned are allylthiomethyl, phenylthiomethyl, butylthiomethyl, α-chlorocrotylthiomethyl, phenoxymethyl, phenoxyethyl, phenoxybutyl, phenoxybenzyl, phenoxyphenoxymethyl, (dimethylmethoxy)methyl, (dimethylbutyoxyJmethyl, (dimethylphenoxyJmethyl, 4-guanidinophenoxymethyl, 4-pyridylthiomethyl, p.-(carboxymethyl) phenoxymethyl, jo-(carboxymethylJphenylthiomethyl, 2- thiazolylthiomethyl, p.-(sulfo)phenoxymethyl, 2-pyrimid15 inylthiomethyl, phenethylthiomethyl, 1-(5,6,7,8-tetrahydronaphthyljoxymethyl, N-methyl-4-pyridylthio, benzyloxy, methoxy, ethoxy, phenoxy, phenylthio, amino, methylamino, dimethylamino, a pyridinium methyl or trimethylammoniummethyl non-toxic salt, cyanomethylthiomethyl or trifluoro20 methylthiomethyl. I? is 4-pyridylethyl, 4-pyridylpropyl, 4-pyridylbutyl, 3-imidazolylethyl, 3-imidazolylpropyl, 3- imidazolylbutyl, 1-pyrrolylethyl, 1-pyrrolylpropyl and 1-pyrrolyIbutyl.

Alternatively, the acyl group can be a radical of the formula: X II — C—CHR' ' R' ' ' - 24 where R'' is defined as above and R'‘' is amino, hydroxy, azido, carbamoyl, guanidino, amidino, acyloxy, halo, such as Cl, F, Br, I, sulfamino, tetrazolyl, sulfo, carboxy, carbalkoxy or phosphono. Representative members of the substituent —CHR' ' R. I , that might be mentioned are a-aminobenzyl, α-amino-(2thienyl)methyl, a-(methylamino)benzyl, α-amino-methylthiopropyl, α-amino-(3 or 4)-ehlorobenzyl, α-amino-(3 or 4)hydroxybenzyl, a-amino-2,4-dichlorobenzyl, a-amino-3,4dichlorobenzyl, 13(-)-α-hydroxybenzyl, a-carboxybenzyl, a-amino-(3-thienylJmethyl D(-)-a-amino-3-chloro-4hydroxybenzyl, a-amino(cyclohexylJmethyl, a-(5-tetrazolyl ),benzyl, 2-thienyl-carboxymethyl, 3-thienyl-carboxymethyl, 2- furylcarboxymethyl, 3-furyl-carboxymethyl, a-sulfaminobenzyl, 3-thienyl-sulfaminomethyl, a-(N-methylsulfamino)benzyl, D(-)-2-thienyl-guanidinomethyl, D(-)-a-guanidinobenzyl, a-guanylureidobenzyl, α-hydroxybenzyl, a-azidobenzyl, a-f luorobenzyl, 4-(5-methoxy-l,3-oxadiazolyl) aminomethyl, 4-(5-methoxy-1,3-oxadiazolyl)-hydroxymethyl, 4-(5-methoxy-l,3-sulfadiazolyl)-hydroxymethyl, 4-(5chlorothienyl)-aminomethyl, 2-(5-chlorothienyl)-hydroxymethyl, 2-(5-chlorothienyl)-carboxy-methyl, 3-(1,2thiazolyl)-aminomethyl, 3-(1,2-thiazolyl)-hydroxymethyl, 3- (1,2-thiazolyl)-carboxymethyl, 2-(1,4-thiazolyl)aminomethyl, 2-(1,4-thiazolyl)-hydroxymethyl, 2-(1,4thiazolyl)carboxymethyl, 2-benzothienylaminomethyl, 2-benzothienylhydroxymethyl, 2-benzothienylcarboxymethyl, α-sulfobenzyl or a-phosphonobenzyl. The acyl radical can also be α-diethylphosphono or a-monoethylphosphono.

Further acyl radicals of interest when X = oxygen are: fi 45 —CCHRR ’ 4 wherein R and R are as defined below. R represents hydrogen, chlorine, fluorine, bromine, iodine, amino, guanidino, phosphono, hydroxy, tetrazolyl, carboxy, sulfo, or sulfamino and R represents phenyl, substituted phenyl, a mono- or bicyclic substituted or unsubstituted heterocyclyl containing one or more oxygen, sulfur or nitrogen atoms in the ring, (such as furyl, quinoxalyl, thienyl, quinolyl, quinazolyl, thiazolyl, isothiazolyl, tetrazolyl, oxadiazolyl or thiadiazolyl), phenylthio, phenyloxy, alkyl of 1-6 carbon atoms, heterocyclic-thio or substituted heterocyclic-thio groups? or cyano. The substituents 4 5 on the residues, R and R can be halo, carbosymethyl, guanidino, guanidinomethyl, carboxamidomethyl, aminomethyl, 4 nitro, methoxy or methyl. When R is hydrogen, hydroxy, amino or carboxy and R is phenyl, or a (5 or 6)-membered heterocyclic ring having one or two sulfur, oxygen or nitrogen hetero atoms such as tetrazolyl, thienyl, furyl and phenyl, the following acyl radicals are representative: phenylacetyl, 3-bromophenylaeetyl, p-aminomethyIphenylacetyl, 4-carboxymethylphenylacetyl, 4-carboxyamidomethylphenylacetyl, 2-furylacetyl, 5-nitro-2-furylacetyl, 3-furylacetyl, 2-thienylaeetyl, 5-chloro-2-thienylacetyl, -methoxy-2-thienylaeetyl, a-guanidino-2-thienylaeetyl, 3-thienylaeetyl, 2-(4-methylthienyl)acetyl, 3-isothiazolylacetyl, 4-methoxy-3-isothiazolylacetyl, 4-isothiazolylacetyl, 3-methyl-4-isothiazolylacetyl, 5-isothiazolylacetyl, 3-chloro-5-isothiazolylacetyl, 3-methyl-l,2,5oxadiazolylacetyl, l,2,5-thiadiazolyl-4-acetyl, 3-methyl44433 - 26 1,2,5-thiadiazolylacetyl, 3-chloro-l,2,5-thiadiazolylacetyl 3- methoxy-1,2,5-thiadiazolylacetyl, phenylthioacetyl, 4- pyridylthioacetyl, cyanoaeetyl, 1-tetrazolylacetyl, afluorophenylacetyl, JD-phenyl glycyl, 4-hydroxy-D-phenylglycyl, 2-thienylglycyl, 3-thienylglycyl, phenylmalonyl, 3-chlorophenylmalonyl, 2-thienylmalonyl, 3-thienylmalonyl, a-phosphonophenylacetyl, α-amino-cyclohexadienylacetyl, a-sulfaminophenylacetyl, α-hydroxyphenylacetyl, a-tetrazolylphenylacetyl and α-sulfophenylaeetyl. 3' 11 2' The radical R when acyl, but not R or R , may also be a sulphur (1) or phosphorus (2) radical: (0)m II „ -S-Y° II (0)n (x)n I -P-Y' Y' ' in which, with respect to 1, each of m and n is 0 or 1 and Y° = 0® Μ®, -N(R' ' )2, or R' '; where M® is hydrogen, an alkali metal or alkaline-earth metal cation or an organic base; and R11 is as defined above, e.g., alkyl, alkenyl, aryl or heteroaryl. With respect to 2, X = 0 or S; n = 0 or 1; and each of Y' and Y'', independently of the other, is Ο ® Μ® , -N(R'')2, R'1 or -ZR*1 where all symbols are as defined above, e.g., R'' and ZR'* may be alkyl, alkenyl, aryl or heteroaryloxy or Y' and Y, including R'1 residues, are joined together to form cyclic ester, esteramide and amide functions. 11 21 31 Acyl groups R , R and R of particular interest, particularly in the preparation of compounds in accordance with the present invention as described below, are conventionally known N-acyl blocking or protective groups - 27 such as carbobenzyloxy, ring-substituted carbobenzyloxy such as p- and p-nitrocarbobenzyloxy, p-methoxycarbobenzyloxy, chloroacetyl, bromoacetyl, phenylacetyl, pbutoxycarbonyl, trifluoroacetyl, bromoethoxycarbonyl, 9-£luoroenylmethoxycarbonyl, dichloroacetyl, 2,2,2-trichloroethoxycarbonyl, bromo-t-butoxycarbonyl, phenoxyacetyl; non-acyl protective groups such as p-nitrophenylsulfenyl and (C, . alkyl) silyl, for example, trimethylX—o silyl and t-butyldimethylsilyl are also of interest.

The following radicals, according to the foregoing 31 definition of acyl, are especially preferred for R of structure lias formyl, acetyl, propionyl, butyryl, chloroaeetyl, methoxyacetyl, aminoacetyl, methoxycarbonyl, ethoxycarbonyl, methylcarbamoyl, ethylcarbamoyl, phenylthiocarbonyl, 3-aminopropionyl, 4-aminobutyryl, N-methylaminoacetyl, N,N-dimethylaminoacetyl, an Ν,N,N-trimethylammoniumacetyl salt, 3-(N,N-dimethyl)aminopropionyl, a 3-(N,N,Ntrimethyl)ammonium propionyl salt, an Ν,Ν,Ν-triethylammoniumacetyl salt, a pyridiniumacetyl salt, guanylthioacetyl, guanidinoacetyl, 3-guanidinopropionyl, N3-methylguanidinopropionyl, hydroxyacetyl, 3-hydroxypropionyl, acryloyl, propynoyl, malonyl, phenoxycarbonyl, amidinoacetyl, acetamidinoacetyl, amidinopropionyl, aeetamidinopropionyl, guanylureidoacetyl, guanylcarbamoyl, carboxymethylaminoacetyl, sulfoacetylaminoacetyl, phosphono3 acetylaminoacetyl, N -dimethylaminoacetamidinopropionyl, ureidocarbonyl, dimethylaminoguanylthioacetyl, a 3-(1methyl-4-pyridinium)propionyl salt, 3-(5-aminoimidazoll-yl)propionyl, 3-methyl-1-imidazoliumacetyl salt, 3-sydnonylacetyl, p-aminomethylbenzoyl, o-aminobenzoyl, 0 II -p{och3)2 , s II -P(OCH3)2 , 0/OCH3 —P ^OM S 0 0 II II II -P[N(CH3)2]2 . -Ρ-Ν(0Κ )2 , -P[N(CH3)2]2 . \ OM II( II ιι/θΜ -P-N(CH,: )_ , . -S-OM or -p \ .32 II «X0M OM 0 ο υ Another class of acyl radicals suitably ii in compounds of the present invention are terminally substituted acyls in which the substituent is one of certain basic groups. Such preferred substituted acyls may be represented by the following formula: 1' -c(cH2l-pJq-lcH2ln-y and these radicals and the corresponding radicals in which the II io — c — group is omitted are also preferred values of R . In the formula g_ is 0 or 1; each of m and n, independently of the other, is 0, 1, 2, 3, 4 or 5? A is 0, -NR°(where R° is hydrogen or Cn , alkyl) or S, and Y is an 1—b amino or substituted amino radical of formula: -N(r*)2 or -N(R*)3 ; an amidino or substituted amidino radical of formula: -N=C-N(R<’)_ ; II R** a guanidino or substituted guanidino radical of formula: -NH-C-N(R°), ; II 2 NR** or a guanyl or substituted guanyl radical of formula: -C=NR° ll N(R’)2 where each R°, independently of the other, is hydrogen; N(R’)2 (where R is hydrogen or 0χ_6 alkyl); alkyl, c g alkoxyl; (Ο^θ alkoxy)-(C2_6 alkyl), C3_g cycloalkyl or cycloalkyl-(C^ 3 alkyl) or the two R groups are joined to form, together with the N atom to which they are attached, a ring having 3 to 6 atoms; R* is a radical as defined for R except that if cycloalkylalkyl, it must be (C3 cycloalkyl)-C1_3 alkyl; and RH* is a radical as defined for R or a (0η , alkoxyJmethyl radical; or Y is 1— b a monocyclic or bicyclic heterocyclic aromatic or nonaromatic radical having 4 to 10 nuclear atoms and in which the hetero atom or atoms are nitrogen and optionally oxygen or sulfur.

Such heterocycles are representatively illustrated by the following list of radicals (R' is H or g alkyl): 4-533 The following specific carboxylic acyl radicals are additionally representative: nh II I -CCH2CH2NHC-CH3 NH li I -CCH2CH2NHC-H 4 4 3 3 NH -cch2ch2nhc-nh2 -cch2ch2n(ch3)2 II Θ -CCH2CH2N(CH3)3 0 nh -CCH2CH2C-NH2 NH -CCH2CH2CH2NHC-CH3 -cch2ch2ch2n(ch3)3 -CCH2CH2CH2N(CH3)2 NH -CCH.S-C 2 \ NH_ NH -CCH2S-CH2-C NH„ II z -CCH-O-CHC 2 2 x NH NH.

The above-described starting materials are conveniently prepared from an N-protected thienamycin (or position isomer) (1), such as an N-acylated thienamycin (1). r OH Th 1' 2' •NR R L COOH 11 2 1 where each of R and R is hydrogen or an acyl radical 1 ‘ 2 1 as above-defined. Preferably R is hydrogen and R is an easily removable blocking group such as earbobenzyloxy, ring-substituted earbobenzyloxy such as p- and p-nitrocarbobenzyloxy, p-methoxycarbobenzyloxy, chloroacetyl, bromoacetyl, phenylacetyl, t-butoxycarbonyl, trifluoroacetyl, bromoethoxycarbonyl, 9-fluoroenylmethoxycarbonyl,f dichloroacetyl, p-nitrophenylsulfenyl, 2,2,2-trichloroethoxycarbonyl, bromo-t-butoxycarbonyl or phenoxyacetyl; non-acyl protective groups such as tri(C alkyl)silyl, 1—Ό for example, trimethylsilyl and t.-butyldimethyl silyl, are also of interest. The preferred N-blocking groups are (a) substituted and unsubstituted earbobenzyloxy radicals of formula: where n is 0, 1 or 2 and R1 is C, r alkoxy or nitro? *" 1-b and (b) bromo -j;-butoxycarbonyl, i.e.

CH -C-O-C-CKBr \ "3 The ultimate Ν-deblocking procedure for the preparation of Ia, Ic or Ie is accomplished by any of a variety of well-known procedures which include hydrolysis and hydrogenation; hydrogenation is suitably carried out in a solvent such as a C, , alkanol in the presence of a l-o hydrogenation catalyst such as palladium, platinum or oxides thereof.

The N-acylated intermediate compound [I, (or Ia) above] is prepared by treating thienamycin (I) or a position isomer thereof with an acylating agent, for example, an acyl halide or acyl anhydride such as an aliphatic, aromatic, heterocyclic, araliphatic or heterocyclic aliphatic carboxylic acid halide or anhydride.

Other acylating agents may also be used, for example, mixed carboxylic acid anhydrides and particularly alkyl esters of mixed carboxylic-carbonic anhydrides; also, carboxylic acids in the presence of a carbodiimide such as 1,3-dicyclohexylcarbodiimide, and an activated ester of a carboxylic acid such as the jo-nitrophenyl ester.

Such N-acylated starting materials are fully described in British Patent Specification No. 570 985.

The acylation reaction may be conducted at a temperature in the range of from -20° to l00°c., but is preferably conducted at a temperature in the range of from -9°c to 25°C. Any solvent in which the reactants are soluble and substantially inert may be used, for example polar solvents such as water, alcohols and polar organic solvents in general such as dimethylformamide (DMF), hexamethyl, phosphoramide (HMPA), acetone, dioxane, tetrahydrofuran (THF), acetonitrile, heterocyclic amines such as pyridine, ethylacetate, aqueous mixtures of the - 34'above, and halogenated solvents such as methylene chloride and chloroform. The reaction is conducted for a period of time of from about five minutes to a maximum of three hours, but in general, a reaction time of 0.5 to one hour is sufficient. The following equation illustrates this process using a carboxylic acid halide? however, it is to be understood that by substituting a carboxylic acid anhydride or other functionally equivalent acylating agent similar products may be obtained.

OH Generally when the above-described acylating reaction uses an acid halide (suitable halides are chlorides, iodides and bromides) or anhydride the reaction is conducted in water or an aqueous mixture of a polar organic solvent such as acetone, dioxane, THF, DMF or acetonitrile in the presence of a suitable acceptor base such as NaHCO^, MgO, NaOH or K^HPO^.

In carrying out the reactions described herein, it is generally not necessary to protect the 2-carboxy group or the 1'-hydroxy group; however, in cases where the acylating reagent is exceedingly water-sensitive it is sometimes advantageous to perform the acylation in a non-aqueous solvent system. Triorganosilyl (or organo tin) compounds of thienamycin or its position isomers, for example tris-trimethylsilyl thienamycin ThiTMS)^: Γ OTMS Th NHTMS L COOTMS react rapidly. Such derivatives, which are readily soluble in organic solvents, are conveniently prepared by treating thienamycin with an excess of hexamethyldisilazane and a stoichiometric amount of trimethylchlorosilane at 25°C., with vigorous stirring under a N2 atmosphere. The resulting NH^Cl is removed by centrifugation and the solvent is removed by evaporation to provide the desired silyl derivative.

The intermediate starting materials Ic are prepared according to the following scheme; however, it should be noted that direct esterification, without protection of the amino group, is also possible.

Th OH 11 2 1 NR R Th Γ OH NR^'r2' deblock COOH „C0X'R Id r oh Th •NH„ COX'R Ic - 36 where all symbols are as previously defined.

In general, the transformation (1—>Ic) is accomplished by conventional procedures. Such procedures include: (1) Reaction of 1 (or I) with a diazoalkane such as diazomethane, phenyldiazomethane or diphenyldiazomethane, in a solvent such as dioxane, ethylacetate or acetonitrile at a temperature of from O°C to reflux for from a few minutes to 2 hours. (2) Reaction of an alkali metal salt of 1 with an activated alkyl halide such as methyl iodide, benzyl bromide, or m-phenoxybenzyl bromide, n-t-butylbenzyl bromide or pivaloyloxymethyl chloride. Suitable reaction conditions include solvents such as hexamethylphosphoramide at a temperature of from 0°C. to 60°C. for from a few minutes to 4 hours. (3) Reaction of 1 with an alcohol such as methanol, ethanol or benzyl alcohol. This reaction may be conducted in the presence of a carbodiimide condensing agent such as dicyclohexylcarbodiimide. Suitable solvents, at a temperature of from 0°C to reflux for from 15 minutes to IS hours, include CHC13, eH3Cl and CH2C12. (4) Reaction of an N-acylated acid anhydride of 1 prepared by reacting the free acid 1_ with an acid chloride such as ethyl chloroformate or benzyl chloroformate with an alcohol such as those listed in (3) under the same conditions of reaction as given above for (3). The anhydride is prepared by reacting 1 and the acid chloride in a solvent such as tetrahydrofuran (THF) or CH2C12 at a temperature of from 25°C. to reflux for from 15 minutes to 10 hours. - 37 (5) Reaction of labile esters of 1 such as the trimethylsilyl ester or dimethyl-t-butylsilyl ester with R3 X° wherein X° is halogen such as bromine or chlorine 3 · and R is as already defined, in a solvent such as THF 5 or CH2C12 at a temperature of from 0°C to reflux for from minutes to 16 hours. For example according to the following scheme: Th r OTMS NHTMS COOTMS N-acylation Th OTMS -NR1 TMS _ COOTMS ’OTMS 11 esterification Th — NR TMS mild hydrolysis _COOR OH Th NHR COOR where TMS is triorganosilyl such as trimethylsilyl and 10 all other symbols are as previously defined.

The amides of the present invention are most conveniently prepared by reacting the acid anhydride (Ic, 3 1 X' =» 0, R = acyl) with ammonia or with the amine of choice, e.g., the alkyl-, dialkyl-, aralkyl- or hetero15 cyclic amines listed above.

The above-recited schemes of esterification are well-known in the related bicyclic β-lactam antibiotic art and indeed in organic synthesis generally, and there - 38 is no undue criticality, of reaction parameters in the preparation of the N-acylated and carboxyl derivatives Ic useful as starting materials in the practice of the present invention.

. Starting materials Ia and Ie are conveniently prepared by any of a variety of well-known esterification or etherification reactions upon the secondary alcoholic group of Id. Such procedures include; Γ OH r- OR Th ,ml„2 -NS .R COXR Id 2 NR R 1_ COXR Ie (1) For the preparation of ethers of the present invention, the acid-catalysed reaction of Id with a diazoalkane such as diazomethane, phenyldiazomethane or diphenyldiazomethane in·an inert solvent such as dioxane, tetrahydrofuran (THF), a halohydrocarbon such as CH^Cl^ or ethyl acetate in the presence of a catalytic amount of a strong acid or Lewis acid such as toluenesulfonic acid, trifluoroacetic acid, fluoboric acid or boron trifluoride at a temperature of from -78°C to 25°C for from a few minutes to 2 hours. (2) For the preparation of ethers of the present invention, the reaction Of Id with an alkylating agent such as active halides, for example methyliodide, benzylbromide or m-phenoxybenzylbromide or alkylsulphates such as dimethylsulphate, diethylsulphate and methylfluorosulphonate in the presence of a strong base capable of forming the alcoholate anion of lb. Suitable bases include alkali and alkaline-earth metal oxides and hydroxides, alkali metal alkoxides such as potassium tertiary butoxide, tertiary amines such as triethylamine, alkali metal alkyls and aryls such as phenyllithium, and alkali metal amides such as sodium amide. Suitable solvents include inert anhydrous solvents such as t-butanol,· dimethylformamide (DMF), THF, hexamethylphosphoramide (HMPA) and dioxane at a temperature of from -78°C to 25°C., for from a few minutes to 4 hours. (3) For the preparation of esters the reaction of Id with any of the above-listed acyl radicals in their acid form. This reaction may be conducted in the presence of a carbodiimide condensing agent such as dicyclohexylcarbodiimide. Suitable solvents include any inert solvent such as CHCl , CH_Cl_, DMF, HMPA, acetone and dioxane at a temperature of from 0°C to 60c. for from 15 minutes to 12 hours. (4) For the preparation of esters of the present invention, the reaction of Id with an acyl halide or an acid anhydride in which the acyl residue is described above. Generally, when the above-described acylating reaction uses an acid halide (suitable halides are chloride, iodide or bromide) or acid anhydride the reaction is conducted in an anhydrous organic solvent such as acetone, dioxane, methylenechloride, chloroform or DMF in the presence of a suitable acceptor base such as NaHCO^, MgO, triethylamine or pyridine at a temperature of from 0°C. to 40 C. for from 1 to 4 hours.

Suitable acyl halides and anhydrides include acetic anhydride, bromoacetic anhydride, propionic anhydride, benzoyl chloride, phenylacetyl chloride, azidoacetyl chloride, 2-thienylacetyl chloride, 2-, 3- and 4-nicotinyl chloride, p-nitrobenzoyl chloride, 2,6-dimethoxybenzoyl 4&33 - 40 chloride, 4-guanidinophenylacetyl chloride, methanesulfonyl chloride, dibenzylphosphorochloridate, dimethylthiophosphoro chloridate, 2-furoyl ethyl carbonic anhydride, methylchloroformate and bis(p-nitrobenzyl)phosphorochloridate. (5) For the preparation of esters of the present invention, the reaction of Id with a suitably substituted ketene or isocyanate such as ketene, dimethyl ketene, methylisocyanate, methylisothiocyanate or chlorosulfonyl isocyanate. The reaction is conveniently carried out in a suitable solvent, e.g. dioxane, tetrahydrofuran or chloroform, at a temperature of from -70°C. to 60°C. for from 15 minutes to 18 hours.

The intermediate compound Ie is then N-deblocked as χ' described above to provide starting materials, Ie (R and 2 1 R = H) and la. From Ie, Ia is prepared by deblocking the carboxyl group: Th Γ OR 1' 2' NR R Th L_ COX'R3' OR -nh2 L COX'R3 ie Ie Th NH2 COOH la Starting material Ia is conveniently and preferably 3' obtained when X' is oxygen and R is a readily removable 20 carboxyl-protecting or carbonyl-blocking group (see above).

Starting material Ia is prepared by deblocking according to any of a variety of well-known procedures which include hydrolysis and hydrogenation. When the preferred carboxylblocking groups are used (below), the preferred deblocking procedure is hydrogenation, in which the intermediate compound (Ie), in a solvent such as a c, , alkanol, is l-o hydrogenated in the presence of a hydrogenation catalyst such as palladium, platinum or oxides thereof.

In this connection, it is noted that suitable 3 1 blocking groups R include the sub-generic groups defined above as aralkyl, haloalkyl, alkanoyloxyalkyl, alkoxyalkyl, alkenyl, substituted alkyl, or aralkoxyalkyl, and also including alkylsilyl, where alkyl has 1-10 carbon atoms. For example, suitable blocking groups 3' R include benzyl, phenacyl, p-nitrobenzyl, methoxymethyl, trichloroethyl, trimethylsilyl, tributyltin, p-methoxybenzyl and benzhydryl. These blocking groups are preferred since they are generally recognised easilyremovable blocking groups in the cephalosporin and penicillin art.

The preferred carboxyl blocking groups are benzyl and substituted benzyl of formula: wherein n is 0, 1 or 2 and R' is C, r alkoxyl or nitro. l-o In the alternative it should be noted that the compounds of the present invention, Ila, may be arrived at by operating upon the substituted N-methylene thienamycin derivatives or their position isomers II, to achieve derivatization by establishment of R and/or 3' -COX'R . Such procedure is exactly as described above - 42 except that compound II replaces the above-described starting materials, such as la, Ic and Ie, and, of course, there is no need to N-deblock.

Preparation The preparation of the compounds of the present invention is conveniently described according to the above-defined four classes, or embodiments, namely: (1) Amidines? (2) Guanidines? (3) Substituted Pseudoureas? and (4) Imido Esters and Imido Thioesters. (1) Amidines. In general, the compounds of Class (1) may conveniently be prepared by reacting thienamycin 11 (I) or a derivative thereof (Ta, Ic or Ie, when R = 21 R =H) or a suitably protected form of thienamycin or its position isomer such as its silylated derivative (1) with an imido ester (a) or a substituted imido halide (b): sch2ch2nhtms COOTMS TMS = trimethylsilyl NR R1R2N=C-X' r-c-xqr· (a) R (b) where R , R , and R are as defined above? X' is halogen such as chlorine? and -X°R'1 is a leaving group where R'1 may be C^ & alkyl such as methyl or ethyl and X° is 0 or S. Alternatively, the compounds of Class 1 may be prepared by reacting a compound of Class 4 with NH^ or a primary or secondary amino compound (c) calculated to provide the desired compounds of Class 1. Reagents (a), (b) and (c) are representatively enumerated below.

Suitable solvents for the preparation of the compounds of Class 1 according to the above reaction schemes, depending upon the identity of the thienamycin substrate and reagent, include water, dioxane, tetrahydrofuran (THF), dimethylformamide (DMF), chloroform, acetone, acetonitrile or mixtures thereof. The reaction may be conducted at a temperature of from 0° to 25°C for from 1 to 6 hours. There is no criticality as to the precise identity of the reaction solvent nor the variables of reaction within the limits described above, provided only that the reaction solvent is inert or substantially inert to the intended course of reaction. Suitable reagents representatively include: (a) Imido Esters: „1 „1„2 R R R Ν N Ho ® 11 o R-C-X R' ' , R—C-X R' ' X° = 0 or S Examples are methyl formimidate, ethyl formimidate, methyl acetimidate, ethyl acetimidate, methyl benzimidate, ethyl 4-pyridyl carboximidate, methyl phenylacetimidate, methyl 3-thienylearboximidate, methyl azidoacetimidate, methyl chloroacetimidate, methyl cyclohexylcarboximidate, methyl 2-furylcarboximidate, methyl p.-nitrobenzimidate, methyl 2,4-dimethoxybenzimidate, ethyl N-methyl formimidate, methyl N-methyl formimidate and methyl N-isopropyl formimidate. 44^33 - 44 Such imido ester reagents (a) are convenientlyprepared by any of a variety of known procedures, such as (1) The reaction of a nitrile, RON, with a C, , 1-6 alkanol in the presence of HCl according to the well-known Pinner synthesis. (2) The reaction of a nitrile, RCN, with a Cn r J,"b alkanol in the presence of a base. Typically, the reaction is conducted at 0-40°C in the presence of an excess of the alcohol with a catalytic amount of an alkali metal alkoxide for from 15 minutes to 4 hours. (3) The reaction of an amide, ll 1 RCNHR , with an alkyl chloroformate such as methylehloroformate at 25°C-45°C for 1-4 hours. (4) The reaction of an N-substituted amide, . 0 0 II 1 H 1 2 RCNHR or RCNR R , with an equivalent of an alkylating agent such as triethyl oxonium fluoroborate in an inert solvent such as ether or chloroform at 0-23°C for from 10 minutes to 2 hours. (5) The conversion of a readily-available imido ester, RCNR' (R' maybe hydrogen), to a desired imido OR' ' ester, RCNR3-, by reaction of the first-mentioned ester 1 OR with an alkylamine, R’NH^, in a mixture of water and an immiscible solvent such as ether or chloroform at O-23°C. for from 5 minutes to 1 hour. (b) Substituted Imido Halides: N-(Chloromethylene)piperidinium chloride, chlorodimethyl forminium chloride, chlorodiethyl forminium chloride.

Such imido halide reagents (b) are conveniently prepared by any of a variety of known procedures, such as: θ (1) The reaction of an Ν,Ν-disubstituted amide, II i 2 RCNR R , with a halogenating agent such as thionyl chloride, phosgene or phosphorus pentachloride in an inert solvent such as chloroform or methylene chloride at 0-40°C for from 1- 5 hours. (c) Primary and Secondary Amino Compounds: Methylamine, ethylamine, 2-aminopyrimidine, dimethylamine, methyl benzylamine, 3-aminomethyl pyridine, 2-aminomethyl thiophene, ethanolamine, dimethylaminoethylamine, N-2(aminoethyl)pyrrolidine, cyclohexylamine, n-heptylamine, isopropylamine, 2-methylallylamine, 3-phenyl-l-propylamine, 2- amino-4-picoline, 2-amino pyridine, 3-amino-4-carbethoxypyrazole, 2-aminothiazole, 5-amino-3-methyl isothiazole, and 3-amino-l',2,4-triazole.

The reaction involving the reagents (a), may be representatively shown by the following diagram: SCHjCHjNHj COX'R3' R-C-OR'* II where OR'' is the leaving group of the imido ester reagent 3' 3 and R, R', R , R and X' are as defined above. This reaction is particularly suitable for compounds in which 3 3' R and R are hydrogen and X* is oxygen.

The reaction involving the reagents (b) may representatively be shown by the following diagram: R OH mild hydrolysis pH 3-6 SCH_CH N=C-NR1R2 2 2 | COOH R where all symbols are as previously defined. When 3 31 product 2 is desired, suitable values for R and R are trimethylsilyl and X1 is oxygen.

The reaction involving the reagents (c) may representatively be shown by the following diagram: .SCH„CH„NH=C-X 2 Z | COX'R3' R a© nhr1r2 Compound of Class 4. © 12 SCH.CH„NH=C-NR R 2 | 71 13 COX'R * A© where all symbols are as previously defined and X is -OR or -SR where R is preferably C. , alkyl such as methyl 3 3 ’ L-6 or ethyl. When R and R are readily-removable blocking or protecting groups they may independently be removed by well-known procedures to provide compounds 3, 4 and 5.

OH SCH2CH2N=C-NR1R2 COOH R 44^33 SCiLCH-NH^-lZR2 2 2 ι COX'R' αΘ 3' (2) Guanidines: In general, the compounds of Class 2 may conveniently be prepared by reacting thienamycin or its position isomer or an 0- or carboxyl derivative thereof (la, Ic or Ie, when R1 = R2 =H) with (a) an -OR’’ (e.g., 0-alkyl, 0-aryl) pseudourea or an S- alkyl or S-aryl pseudothiourea; or (b) by reacting a compound of Class 3 (above) with ammonia or an amino compound such as an alkyl, aralkyl or hetero10 aralkyl amine.

Suitable solvents for such reactions include water and buffered aqueous polar organic solvent mixtures at pH 7-9 or anhydrous polar organic solvents such as dimethylformamide or hexamethylphosphoramide at a temper15 ature of from 0°C. to 40°C. for from 1 to 24 hours.

Suitable reagents (a) and (b) include: (a) -OR pseudoureas and -SR pseudothioureas: Ο-Methyl pseudourea, S-Methylpseudothiourea, S-methylpseudothionitrourea, 0-2,4-dichlorophenyl pseudo20 urea, S-p-nitrophenyl pseudothiourea and 0-N,N-trimethyl4 4 4 3 3 - 49 pseudourea. (b) Amino reagents: These reagents are the same as those given for the preparation of Class 1 (c) above.

The reaction involving the reagents (a) may representatively be shown by the following diagram: 31 1 2 where R , X', R , R and R are as defined above; X' ’ is 0 or S and X''R° is a leaving group; R° is preferably C, , alkyl or aryl.

I’*© The reaction involving the reagents (b) may representatively be shown by the following diagram: 2 HNRR - 50 Compound of Class 3. where all symbols are as previously defined. (35 Substituted Pseudoureas; In general, the compounds of Class 3 may conveniently be prepared by reacting a carbamoyl or thiocarbamoyl Nsubstituted compound {a), for example: OH (a) with an alkylating agent (b) such as an active alkyl 10 or aralkyl halide or sulfate ester.

Suitable solvents for the above reaction include alkanols, dioxane and acetonitrile at a temperature of from 20 C. to 60°c. for from 1 to 4 hours. - 51 Suitable reagents (a) for the above reaction scheme include N-acyl thienamycins: as defined above and is preferably S 11 12 -C-NR R or -CNR1R2 2 where R and R are as defined above, such as carbamoyl, methylcarbamoyl, ethylearbamoyl, phenylcarbamoyl, p_bromophenylcarbamoyl, phenylthiocarbamoyl, methylthiocarbamoyl or dimethylcarbamoyl.

Suitable alkylating agents (b) include methyl iodide, benzyl bromide, dimethyl sulfate, diethyl sulfate, allyl bromide, 2-thienyl bromide, methallyl bromide, js-nitrobenzyl bromide and methyl chloromethyl ether.

The reaction involving the above reagents (a) and (b) may representatively be shown by the following diagram: SCH„CH„NH-C-NR1R2 2 2 ,| •5 I V» » COX’k a --52--------- SCH2CH2NH=tj!-NR'LR2 COX'R X' 'R where X'' is 0 or S; RX° is the alkylating agent; and 1 2 3 3' R , R, R , X', R and R are as previously defined. (4) Imido Esters and Imido Thio Esters; In general, the compounds of Class 4 may conveniently be prepared by reacting a suitable protected N-acyl or N-alkoxy carbonyl derivative, or a thio analogue thereof, of thienamycin (a) with an alkylating agent (b).

Suitable solvents for the above reaction include methylene chloride, tetrahydrofuran, dioxane and chloroform, at a temperature of from -78°C. to 25°C. for from 5 minutes to 3 hours.

Suitable N-acyl thienamycin starting materials (a) include: SC^C^NHR1 ’ COX'R3 1' where R xs acyl such as formyl, benzoyl, thiobenzoyl 3' or thioacetyl; R , R and X' are as defined.

Suitable alkylating agents (b) include: triethyl oxonium fluoroborate, methyl fluorosulphonate 20 and trimethyloxonium hexafluorophosphate.

The reaction involving the above reagents (a and b) may representatively be shown by the following diagram: OR SCH2CH2NHC-R* COX'R3 where X = 0 or S; R* is H, R, OR or SR and R3, X', R3' and R are as previously defined. When the deblocked 3 3' species is desired, suitable values for X' and R , R 5 are oxygen and trimethylsilyl? in which case, deblocking is conveniently achieved by mild aqueous hydrolysis of pH 3-6. It is to be noted that the above reaction mixture may be used directly in reaction with the amine (c) as described in the preparation of the amidines of Class 1, above. £4433 - 54 The products of this invention (II and Ila) form a wide variety of pharmacologically acceptable salts such as acid-addition salts, e.g., with hydrochloric, hydrobromic, sulfuric, nitric, toluene-p-sulphonic and methane sulphonic acids. The salts of this invention are pharmacologically acceptable non-toxic derivatives which can be used as the active ingredient in suitable unit-dosage pharmaceutical forms. Also, they may be combined with other drugs to provide compositions,having a broad spectrum of activity.

Compounds of the present invention have been found to be valuable antibiotics active against various grampositive and gram-negative bacteria and, accordingly, find utility in human and veterinary medicine. Such compounds can therefore be used as antibacterial drugs for treating infections caused by gram-positive or gramnegative bacteria, for example, Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Serratia, Salmonella typhosa, Pseudomonas and Bacterium proteus.

The anti-bacterials of the invention may further be utilized as additives to animal feeding stuffs, for preserving foodstuffs and as disinfectants. For example, they may be used in aqueous compositions in concentrations ranging from 0.1 to 100 parts of antibiotic per million parts of solution in order to destroy and inhibit the growth of harmful bacteria on medical and dental equipment and as bactericides in industrial applications, for example, in water-based paints and in the white water of paper mills to inhibit the growth of harmful bacteria.

The products of this invention may be used alone or in combination as an active ingredient in any one of a variety of pharmaceutical preparations. These antibiotics and their corresponding salts may be used in 4433 capsule form or as tablets, powders or liquid solutions or as suspensions or elixirs. They may be administered orally, intravenously or intramuscularly.

The compositions are preferably presented in a form suitable for absorption by the gastric-intestinal tract. Tablets and capsules for oral administration may be in unit dose presentation form, and may contain conventional excipients such as binding agents, for example, syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinylpyrrolidone; fillers for example, lactose, sugar, maizestarch, calcium phosphate, sorbitol or glycine; lubricants, for example, magnesium stearate, talc, polyethylene glycol, silica; disintegrants, for example, potato starch or acceptable wetting agents such as sodium lauryl sulphate. The tablets may be coated according to well-known methods. Oral liquid preparations may be in the form of aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product, for reconstitution with water or other suitable vehicles before use. Such liquid preparations may contain conventional additives such as suspending agents, for example, sorbitol syrup, methyl cellulose, glucose/sugar syrup, gelatin, hydroxyethylcellulose, earboxymethyl cellulose, aluminum stearate gel or hydrogenated edible oils, for example almond oil, fractionated coconut oil, oily esters, propylene glycol, or ethyl alcohol; preservatives, for example methyl or propyl p-hydroxybenzoates or sorbic acid. Suppositories will contain conventional suppository bases, e.g. cocoa butter or other glycerides.

Compositions for injection may be presented in unit dose form in ampoules, or in multidose containers with an added preservative. The compositions may take such 44*33 forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Alternatively, the active ingredient may be in powder form for reconstitution with a suitable vehicle, e.g. sterile pyrogen-free water, before use.

The compositions may also be prepared in suitable forms for absorption through the mucous membranes of the nose and throat or bronchial tissues and may conveniently take the form of powder or liquid sprays or inhalants, lozenges; throat paints, etc. For medication of the eyes or ears, the preparations may be presented as individual capsules, in liquid or semi-solid form, or drops etc. Topical applications may be formulated in hydrophobic or hydrophilic bases as ointments, creams, lotions, paints, powders.

Also, in addition to a carrier, the compositions may include other ingredients such as stabilizers, binders, antioxidants, preservatives, lubricators, suspending agents, viscosity agents or flavouring agents. In addition, there.may also be included in the composition other active ingredients to provide a broader spectrum of antibiotic activity.

For veterinary medicine the composition may, for example, be formulated as an intramammary preparation In either long or quick-release bases.

The dosage to be administered depends to a large extent upon the condition of the subject being treated and the weight of the host, the route and frequency of administration, the parenteral route being preferred for generalized infections and the oral route for intestinal infections. In general, a daily oral dosage consists of from 2 to 600 mg. of active ingredient per kg. of body weight of the subject in one or more applications per day.

A preferred daily dosage for adult humans lies in the range of from about 15 to 150 mg. of active ingredient per kg. of body weight.

The compositions may be administered in several unit dosage forms, for example, in solid or liquid orally ingestible dosage form. The compositions per unit dosage, whether liquid or solid, may contain from 0.1% to 99% of active material, the preferred range being from 10-60%.

The composition will generally contain from 15 mg. to 1500 mg. of the active ingredient? however, in general, a dosage in the range of from 100 mg. to 1000 mg. is preferred. In parenteral administration the unit dosage is usually the pure compound in a slightly acidified sterile aqueous solution or a soluble powder intended for solution.

The following Examples illustrate the present invention Examples 1, 2, 8,17,18, 22 to 27 and 44 to 50 illustrate the preparation of starting materials and not of compounds in accordance with the present invention. In the Examples, the thienamycin nucleus (I, above) is represented by the following symbol: Γ OH Th -^2 - COOH where the secondary alcoholic group, the amino group and 25 the carboxyl group are illustrated. Thus, compounds of the present invention can conveniently be represented as follows: ThΓ OH N=C-X I Y L COOH and Th r OR NH=C-X I Y _ COX'R3 II Ila 3' where X, Y, R , X', R and A are as previously defined. In the Examples, the words Dowex, Nujol, Porosil, Teflon, Supercel, Linde, Celite, Branson, Sonifier and Difco are trade marks, X proportions of liquid mixtures are on a volume basis, mmole means millimole.

EXAMPLE 1 r OTMS Th NHTMS or TH(TMS)3 I- COOTMS ; TMS = trimethylsilyl Preparation of Silylated-Thienamycin Thienamycin (80.0 mg.) is suspended in 40 ml. tetrahydrofuran (THE) under a N^ atmosphere and is concentrated to 10 ml.; hexamethyldisilazane (1,0 ml.) and trimethylchlorosilane (300 μΐ) are added. The mixture is reacted for 20 mins, at 25°C. with vigorous stirring. The suspension is theji· centrifuged to remove ammonium chloride.· The supernatant is evaporated to an oil under a nitrogen stream for future reaction.

EXAMPLE la Cl Cl OH Th--N=C o Preparation of Thienamycin. N-Piperidin-l-yl Methylene Derivative Thienamycin (57 mg., 162 umol) is silylated according to the procedure previously described. The silylated antibiotic Th(TMS)g is dissolved in methylene chloride (6 cc) in a septum stoppered flask under positive nitrogen pressure and cooled in a dry ice-acetone bath. To the magnetically stirred solution is added a solution (180 jil) of triethylamine (644 umol) in methylene chloride. This is followed by the addition of a solution of N-(chloromethylene)piperidinium chloride (67 mg., 405 pmol) in methylene chloride (465 μΐ). After 1 hour in a dry ice bath, the reaction solution is rapidly added to a tetrahydrofuran - pH 7,0.IN phosphate buffer (1:1) solution (50 ml.). The mixture is then concentrated under vacuum to 10 ml. to give a homogeneous solution. The solution is washed twice with ethyl acetate (2 x 5 ml.) and ether (2 x 5 ml.) and briefly pumped under vacuum. This aqueous solution is then chromatographed on an XAD-2 resin column (60 ml. bed). The product is eluted in 10% aqueous tetrahydrofuran (following water elution) to give 12.9 mg. (22%) product (as measured in solution assuming ¢8,030 same as Thienamycin. Paper chromatography R£ 0.42 (4:1:5, n-BuOH:EtOH:water). - 60 EXAMPLE 2 Thienamycin NH tl H^C-O-C-0 I— OH Th Preparation of N-Benzimidoyl Thienamycin Thienamycin (59 mg., 212 pmol) is dissolved in a 33% N,N-dimethylformamide pH 7 phosphate buffer (0.05N) solution (4.5 ml.) and adjusted to pH 9.5 using 2.5N NaOH with an automatic dispensing burette. The solution is magnetically stirred at 25°C. and methylbenzimidate.HCl (340 mg., 1981 pmol) is added at once. After 30 min. the solution is extracted twice with an equal volume of chloroform and adjusted with dilute aqueous phosphoric acid to pH 7.0. The buffered solution is chromatographed on XAD-2 resin (65 ml.). The column is first eluted with water followed by 10% aqueous tetrahydrofuran which elutes the product. This fraction is concentrated to one-half volume and freeze-dried to give 50 mg. of the product. Electrophoretic mobility (50 V/cm., 20 min., pH 7, 0.1N phosphate buffer) is 1.5 cm. towards the abode. UV = 300 nm (£.6,960) pH 7,0.IN phosphate buffer. 4 4 3 3 Preparation of Ν-Benzimidoyl Thienamycin, p-tertButylbenzyl Eater Benzimidoyl Thienamycin (3.2 mg.) is suspended in hexamethylphosphoramide (75 μ,Ι.) containing p-tert-butylbenzyl bromide (3.8 μΐ) and magnetically stirred at 22°C. After 45 minutes a solution results which is stirred for an additional hour. The product is then precipitated out of solution with ether and the crude product chromatographed on a 250 μ thick silica gel plate developed in 7:3 chloroform ethanol. The band at R. 0.6 is f removed and eluted with ethanol to give N-benzimidoyl \ά33 thienamycin p-tart-butvlbenzvl ester hydrobromide. Mass Spec, m/e 521 (M+), 487, 444, 418, 341, 323, 297, 226, 147.

EXAMPLE 4 Th H CH3 Br Preparation of Ν-Benzimidoyl Thienamycin 3-Methyl-2Buten-l-yl Ester Benzimidoyl thienamycin (5.9 mg.) is dissolved in hexamethylphosphoramide (100 μ.1.) containing l-bromo-310 methyl-2-butene (4.8 μΐ.) and triethylamine (0.5 μΐ.) and magnetically stirred at 22°C. After 1 hour the crude reaction is chromatographed on a 250 μ thick silica gel plate developed in 8:2 chloroform ethanol. The band of R^ 0.1 - R. 0.3 is removed and eluted with ethanol. Benzimidoyl thienamycin 3-methyl-2-buten-lyl ester hydrobromide is isolated as a solid after precipitation.from an ethanol-chloroform solution with hexane. 4 4 3 3 EXAMPLE 5 Preparation of N-Formimidoyl-thienamycin OH SCH_CH_N=C-NH„ 2 2 | 2 COOH H Thienamycin (517 mg) is dissolved in pH 7, O.lN phosphate buffer (25 ml) and cooled in an ice bath with magnetic stirring. The solution is adjusted to pH 8.5 using 2.5N sodium hydroxide solution dispensed from an automatic burette. While maintaining a pH of 8.5, methyl formimidate hydrochloride (711 mg) is added portionwise over 2-3 minutes. After an additional 10 minutes, the pH of the solution is brought to 7.0 using 2.5N hydrochloric acid. The solution is chromatographed on a column of XAD-2 resin (150 cc) which is eluted with water. The N-formimidoyl thienamycin derivative elutes in 1.5-2.0 column volumes (200-300 cc) and is lyophilized to a white solid (217 mg).

UV (pH 7, O.lN phosphate buffer) λ 297 nm { 8,590). ' _ max ir (Nujol mull) 1767 cm (β-lactam) nmr (D20) $ 1.37 (d, J= 6Hz, CHg-CH), 3.0-3.75 (m, -CH2-), 4.2-4.8 (m, C5H, C6H, C?H), 7.86 NH II (s, -C-H).

EXAMPLE 6 Th Γ OH NH NHC 'NH„ L co2h Preparation of N-Guanyl thienamycin Thienamycin (8.9 mg.) is dissolved in pH 7 0.1N phosphate buffer (0.7 ml.) and Ν,Ν-dimethylformamide (0.3 ml.) and the solution brought to pH 9.5 by the addition of 2.5N sodium hydroxide solution. To the magnetically stirred solution is added O-methylisourea· hydrogen sulfate (43 mg.) causing a slight drop in pH.

Additional sodium hydroxide solution is added to bring the pH back to 9.5 and the solution is stirred 30 minutes at 23°C. The solution is then acidified to pH 7.0.

A sample of the solution containing a mixture of thienamycin and N-guanyl thienamycin shows two bioactive zones after electrophoresis (50 V/cm., 20 minutes, 0.05N pH 7 phosphate buffer) and bioautography on S. aureus plates.

EXAMPLE 7 Thienamycin - 65 Preparation of N-Guanyl thienamycin Thienamycin (11 mg.) is dissolved in pH 7 0.IN phos phate buffer (1 ml.) and adjusted to pH 8.3 with Q.1N sodium hydroxide by means of an automatic dispensing burette. To the magnetically stirred solution is added 0-2,4,5-trichlorophenylisourea«hydrochloride (76 mg.) portionwise to allow the automatic burette to maintain a nearly constant pH. The reaction is run for 4 hours at 22°C. and is then readjusted to pH 7.0 by the addition of dilute acid. A sample of this solution containing thienamycin and N-guanyl thienamycin is electrophoresed (50 V/cm., 25 minutes, pH 7 0.1N phosphate buffer) and shows a positive Sakaguchi spray zone at 2.0 cm. towards the anode and a positive ninhydrin spray zone at 1.5 cm. in the same direction.

EXAMPLE 8 . HCl H_NCN —-> Preparation of 0-2,4,5-Trichlorophenylisourea’ Hydrochloride A solution of cyanamide (0.28 mg.) in ether (0.50 ml.) is mixed with 2,4,5-trichlorophenol (12.5 g.); the mixture is heated to 70°C, and the melt magnetically - 66 --stirred while the reaction flask is flushed with nitrogen. Dry hydrogen chloride gas is then slowly bubbled into the melt and the reaction is allowed to cool to 22°C. The resulting solid is washed thoroughly with ether and filtered to give 0-2,4,5-trichlorophenylisourea· hydrochloride as a white solid, m.p. 2O5-2O6°C.

EXAMPLE 9 Th(TMS).

Cl Θ Θ (CH ) N=CHC1 0 2 Th 'OH H I - N=C CH3 /^Η3 CO, Preparation of N-Dimethylaminornethylene thienamycin Thienamycin (16.5 mg.) is silylated with hexamethyldisilazane (200 μί.) and trimethylchlorosilane (60 μί.) in the usual manner. The silylated thienamycin is suspended in ethanol-free chloroform (1 ml.) with magnetic stirring under a nitrogen atmosphere. The mixture is cooled to -45°C. and a solution of triethylamine (21 μί.) in chloroform (21 μί.) is added followed by a solution of (chloromethylene)-dimethylammonium chloride (11.5 mg.) in chloroform (50 μί.). The mixture is warmed to -25°C. over 1 hour and 0.1N pH 7 phosphate buffer (5 ml.) is added. The mixture is vigorously stirred for 15 minutes. The aqueous phase is separated and contains N-dimethylaminomethylene thienamycin which has an electrophoretic mobility (50 V/cm., 1 hour, pH 7 buffer) of 3.6 cm. towards the cathode.

Th OH Φ ^>NH.

-NHC ZWtl2 CO- 67 EXAMPLE 10 II BrCH OCC(CH ) -£-i 3Th OH e NHC NH2Be C-0-CH20CC(CH3)3 Preparation of N-Formimidoyl Thienamycin Pivaloxymethyl Ester Hydrobromide N-aminomethylene thienamycin (10 mg.) is dissolved in hexamethylphosphoramide (200 μΐ.) containing bromomethyl pivalate (10 μ,Ι.) and triethylamine (1 μ,Ι.) and magnetically stirred at 22°C. After 2 hours the hexamethylphosphoramide solution is dissolved in 2 ml. methylene chloride and the product precipitated with a 50:50 hexane-ether solution. The precipitate is dissolved in an aqueous 10% tetrahydrofuran solution and chromatographed on an XAD-2 resin packed column. N-Formimidoyl thienamycin pivaloxymethyl ester is isolated as a solid after tetrahydrofuran elution of the column and lyophilization. - 68 EXAMPLE 11 Preparation of N-Trifluoroacetimidoyl thienamycin OH Thienamycin (199 mg) is dissolved in pH 7 0.IN phosphate buffer (7 ml) and adjusted to pH 8.5 with IN sodium hydroxide solution. While this pH is maintained with an automatic burette, a solution of methyl trifluoro acetimidate (355 μΐ) in dioxane (2.5 ml) is added in one portion. After 30 minutes the pH is readjusted to 7.0 by the addition of IN hydrochloric acid. The solution is then chromatographed on Dowex 50-X4 resin (200 cc, Na+ cycle, 200-400 mesh) and is eluted with water. The N-trifluoroacetimidoyl thienamycin derivative elutes in the first half column volume. This eluate is reehromatographed in a similar manner on Dowex 5O-X4 (100 cc, Na cycle, 200-400 mesh) and the first column volume concentrated and chromatographed on XAD-2 resin (30 cc.). The N-trifluoroacetimidoyl thienamycin derivative elutes in 2.5 - 5.0 column volumes which is lyophilized to a white solid (15 mg).

UV (pH 7, 0.1N phosphate buffer) λ 302 nm (£4,450). ir (Nujol mull) 1750 cm (β-lactam).

Electrophoresis: (50 v/cm, 20 min, pH 7, 0.1N phosphate buffer) mobility 2.0 cm (toward cathode) EXAMPLE 12 Preparation of N-Acetimidoyl thienamycin OH SCH„CH„N=C-NH.

I ώ _____ CH_ COOH 3 Thienamycin (190 mg) is dissolved in pH 7 0.1N 5 phosphate buffer (13 ml) and cooled in an ice bath with magnetic stirring. The solution is adjusted to pH 8.5 using 2.5N sodium hydroxide solution dispensed from an automatic burette. While maintaining a pH of 8.5, ethyl acetimidate hydrochloride (400 mg) is added portionwise over a few minutes. After an additional 40 minutes the solution is adjusted to pH 7.0 with 2.5N hydrochloric acid. The solution is then chromatographed on Dowex 4. 50-X8 resin (250 cc, Na cycle, 100-200 mesh) and is eluted with water. The N-acetimidoyl derivative elutes in 1-2 column volumes (240-520 cc) and is lyophilized to a white solid (88 mg).

UV (pH 7, 0.IN phosphate buffer) λ 297 nm (£ 7,620), ir (Nujol mull) 1774 cm , β-lactam, ΝΉ nmr (DjO) β 1.27 (d, σ = 6 Hz, CH^-CH) 2.24 (S, -C-CHg 3.2 - 3.5 (m, -CH2), 3.5 - 3.9 (m, -CH2~) 4.2 -4.6 (m; Cffl, C6H, C^). - 70 EXAMPLE 13 Preparation of N-Γ(4-pyridyl) (imino)methyl]thienamycin OH Thienamycin (80 mg., 0.294 mmole) is dissolved in 5 aqueous sodium bicarbonate (24.7 mg., 0.294 mmole in 2.0 ml.) at 25°C. Methyl isonicotinimidate (80 mg., 0.588 mmole) is dissolved in the solution and progress of the reaction is followed by timed aliquots using high perform ance liquid chromatography (HPLC): Waters instrument: 0.2 x 61 cm. C. Bondapak reverse phase column; 1.5 ml/ min aqueous 10% THF; UV (254 nm.) and R.I. monitors.

The reaction is essentially complete in 40 minutes, and the reaction solution is chromatographed directly over an 18.4 x 270 mm. XAD resin column, first eluting with deionised, distilled water, then changing to aqueous 10% THF. The eluate is monitored by UV and HPLC is used to locate the pure product. Correct fractions are combined and lyophilized to yield a colorless, fluffy powder (80 mg. 73%).

UV λ H2° 298 nm (£7,8OQ); IR (Nujol mull) 1762 cm'1 IUcLX , (β-lactam); NMR (60 MHz, D20), δ 1.27, 3H (d, J = 7Hz, CH3.CH(OH)); g 7.75 and 8.80, 4H, (m, m, 4-pyridyl); HPLC, 1.58 x retention of thienamycin, conditions as above.

EXAMPLE 14 Following the procedure of Example 13, but replacing the reagent with methyl picolinimidate, there is obtained: N-1(2-pyridyl)(imino)methyl]thienamycin (85 mg 77%) UV λΗ2° 267, 300 nm (£, 8,150, 7,600); IR (Nujol mull) 1764 cm-1 (β-lactam); NMR (60 MHz, D20), δ 1.24, 3H (d, J= 7Hz, ffl3.CH(0H)); ¢ 7.80, 8.07, 8.80, 4H, (m,m,m, 2- pyridyl); HPLC, 1.8 x retention of thienamycin.

The formulae of the product of this Example and the products of Examples 15 and 16 can be deduced by comparison with Example 13.

EXAMPLE 15 Following the procedure of Example 13, but replacing the reagent with methyl nicotinimidate, there is obtained : N-Γ(3-pyridyl(imino)methyll thienamycin (77 mg., 70%): UV λΗ2° 264, 299 nm, (£ 5570, 6120); IR (Nujol mull), HtaX 1766 cm-1 (β-lactam); NMR, (60 MHz, D20),· £ 1.24, 3H, (d, J= 7Hz, CH3.CH(OH)); ¢7.6, 8.2, 8.9, 4H, (m,m,m, 3- pyridyl); HPLC, 1.57 x retention of thienamycin.

EXAMPLE 16 Following the procedure of Example 13, but replacing the reagent with methyl 4-thiazolecarboximidate, there is obtained: N-Γ(4-thiazolyl imino)methyl1 thienamycin (99 mg, 89%): UV λΗ2° 300 nm, (£ 7530); IR (Nujol _ ΓΠ3.Χ Mull) 1764 cm1 (β-lactam); NMR (60 MHz, ^0), 6 1.23, 3H, (d, J = 7Hz, CH3.CH(0H)); $ 8.60, 9.17, 2H (d,d, J = 2Hz, 4-thiazolyl); HPLC, 1.8 x retention time of thienamycin. - 72 EXAMPLE 17 Preparation of N-Allyfoanamide A mixture of ailylamine (5.00 g., 87.6 nmole) and methylformate (5.26 g., 87.6 nmole) is stirred at 25°C., for 2 hours. At the end of this time, the reaction flask is fitted with a short path distillation head and the desired N-allylformanide is collected at 89-90°C/0.7 mm as a colorless oil. Yield 7.0 g. (94%). IR(CHC13) 3380, 1680 ran1; imr (CHClj) fi 8.1 (IH, br s), fi 6.4-7.9 (IH, very br), 6 5.5-6.3 (IH, ra), 64.9-5.5 (2H, m), EXAMPLE 18 Preparation of Ethyl Allylimldate Hydrochloride Ethyl chlarofonnate (2.66 g., 24.47 mmole) is added by syringe to KF-allyfannanide (2.08 g., 24.47 nmole) in a dry flask under Nj. The resulting mixture is then stirred at 25°C. for 2 hours during which time COj is rapidly evolved. The reaction mixture is then heated to 45°C. until no further evolution of gas is evident (2 hours). The viscous product is then cooled and held at a vacuum of 0.2 mm for 2 hours to ranove all volatile materials.

EXAMPLE 19 Preparation of Ν' (2-Methylthioethyl) -N-formimidoyl thienamycin Th -CH N-CHjCHjSCHj •NHC \ H COOH Thienamycin (105 ng.) is dissolved in pH 7 O.lN phosphate fcuffer (5 ml.) and to this is added a solution of ethyl N-2-methylthioethyl formimidate (300 μΐ) in tetrahydrofuran (2 ml.). The pH of the solution is adjusted to and maintained at 8.5 using an autcburette dispensing IN NaOH. After 30 minutes the pH is adjusted to 7.0 with 2.5N HCl. The solution is chrarnato25 graphed on an ice water jacketed column of Dowex 5O-X4 resin (53cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The Ν' /2-methy lthioethyl_/ N-farmimidcyl derivative elutes in 2-4 column volumes and is lyophilized to give a white solid.

U.V. (pH 7 O.lN phosphate buffer) λ max 298 nn (e 7,760) i.r. (Nujol mull) 1760 cm 1 (8 - lactam). - 73 EXAMPLE 20 Preparation of N‘-Tert-Butyl-N-formimidoyl thienamycin Th OH - NHC N-C(CH,) II 3 3 L COOH Thienamycin (105 mg.) is dissolved in pH 7ZO.1N phosphate buffer (5 ml.) and to this is added a solution of ethyl N-tert-butvl formimidate (290 mg.) in tetrahydrofuran (1 ml.). The pH of the solution is adjusted to and maintained at 8.5 using an autoburette dispensing IN NaOH. After 30 minutes, the pH is adjusted to 7.0 with 2.5N HCI. The solution is chromatographed on an ice water jacketed column of Dowex 50-X4 resin (53 cc, Na cycle, 200-400 mesh) eluted with deionized water. The fractions containing the title product are combined and lyophilized.

EXAMPLE 21 Preparation of Ν'f1-Methyl-2-Propenyl)N-Formimidoyl thienamycin Th - OH N-CH(CH,)CH=CH |! 3 2 --NHCH L COOH Thienamycin (126 mg.) is dissolved in pH 7z0.1N 20 phosphate buffer (6 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing IN NaOH. To this stirred solution is added ethyl N-lmethyl-2-propenyl-formimidate hydrochloride (300 μ,Ι) - 74 while the pH is maintained at 8.5. After 30 minutes, the pH of the solution is adjusted to 7.0 with 2.5N HCl and the solution is chromatographed on an ice water jacketed column of Dowex 50-X4 resin (49 cc, Na cycle 200-400 mesh) eluted with deionized water. The Ν' fl-methyl-2propenyl] -N-formimidoyl derivative elutes in 2-4 column volumes and is lyophilized to a white solid (59 mg.).

U.V. (pH 7 0.1N phosphate buffer) wavelength of maximum absorption = 299 nm (adsorptivity = 7,820) i.r. (Nujol mull) 1760 cm’·*' (β-lactam).

EXAMPLE 22 Preparation of N-(1-Buten-3-yl) formamide A solution of 3.5 g. (0.05 mole) 3-amino-l-butene in 12 ml. of methyl formate is kept at 25°C. for 20 hours; the solution is then concentrated under reduced pressure to remove excess methylformate. The residual N-(l-buten-3-yl)formamide is distilled under reduced pressure. A fraction amounting to 3 g. (b.p. 58-6O°C./ 0.5 mm.) of N-(l-buten-3-yl)formamide is obtained.

EXAMPLE 23 Preparation of Ethyl-N-(l-Buten-3-yl)formimidate A mixture of 1.0 g. of N-(l-buten-3-yl)formamide and one equivalent of ethyl chloroformate is stirred under 1/ for 4 hours during which time C02 is evolved.

The solution is stirred under reduced pressure for 3 hours to remove any unreacted ethyl chloroformate, and a residue of ethyl-N-(l-buten-3-yl)formimidate is obtained. 4 4 3 3 EXAMPLE 24 Preparation of Methyl M-Dimethylaminoformimidate To a stirred solution of N,N-dimethylformhydrazide (0.22 g) in 2.0 ml of chloroform, under nitrogen, is added methyl chloroformate (0.5 ml). The mixture is heated at 40°C. for three hours then evaporated under nitrogen.

The mixture is triturated with anhydrous ether. The supernatant solution is decanted and the residue dried in a stream of nitrogen.

Yield: 284 mg. nmr CDC13 fi, 9.13 (CH); 3.80(0CH3), 3.01(N(CH3)2).

EXAMPLE 25 Preparation of Cyclopropyl formamide A mixture of cyclopropylamine (5.00 g, 87.6 mmole) and methylformate (5.26 g, 87.6 mmole) is stirred at 25°C. for 2 hours, (an initial exotherm is noted).

The mixture is then placed on the rotary evaporator to remove the MeOH formed in the reaction. The remaining material is distilled through a short path head to yield 6.92 g (93%) of the desired N-cyclopropyl formamide as a colorless oil, n.m.r. (CDC13) £8.1 (IH, br S), 6.8-8.5 (IH, br), fi 2.4-3.0 (IH, m), δθ.4-1.0 (4H, m).

EXAMPLE 26 Preparation of Ethyl N-Cyclopropyl formimidate Ethyl chloroformate (4.078 g, 37.58 mmole) is added by syringe to N-cyclopropylformamide (3.194 g, 37.58 mmole) in a dry flask under N2· After an induction period of 30 sec., a rapid evolution of gas begins.

The resulting reaction mixture is stirred at 25°C until I no further evolution of gas can be detected (-~-4 hr), then the viscous product is subjected to a vacuum of 44 33 - 76 0.5 mm for 1 hr to remove any unreacted ethyl chloroformate. NI® analysis of the product shows the formyl proton at S9.37 as a broad singlet. (CDCl^ solution).

EXAMPLE 27 Preparation of Ethyl N-(Methylthioethyl)formimidate To a 60 ml separatory funnel are added ethyl formimidate hydrochloride (0.97 g, 8.8 mmole) a solution of βmethylthioethylamine (0.80 g, 8.8 mmole) in CH2Cl2 (35 ml), and B^O (35 ml.), The mixture is shaken vigorously for 5 minutes, The CH2C12 layer is separated, washed with brine, dried with MgSO^, filtered, and evaporated under reduced pressure to give the crude imidate (0.59 g) as a hazy, pale yellow liquid. I.r.1660,1230cm.

EXAMPLE 28 Preparation of N1-Dimethylamino-N-formimidoyl thienamycin Th —..-CH N-N NKC /CH3 CHCOOH Thienamycin (115 mg.) is dissolved in pH 7Z 0.1 N phosphate buffer (7 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing 1 N NaOH. To this stirred solution is added methyl Ndimethylamino formimidate hydrochloride (284 mg.) while the pH is maintained at 8.5. After 20 minutes the pH of the solution is adjusted to 7.0 using 2.5 N HCl and the solution is chromatographed on Dowex 50-X4 resin (53 cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The chromatography is carried out in a water jacketed column at 3°. The Ν'-dimethylamino-N-formimidoyl derivative elutes in 2 column volumes and is lyophilized to a white solid (40 mg.) UV (pH 7, 0.1 N phosphate buffer) λ 298 nm (£ 6,910) ir (Nujol mull) 1760 cm (β-lactam) nmr (0^0) 6 1.29 (d, J = 6 Hz, CH3-CH), 2.59 (s, N (CH3)2), 7.76 (s, NCH).

EXAMPLE 29 Preparation of Methyl oxalimidoyl thienamycin Th - OH NH NH II II —NHC C-0CH3 _ COOH Thienamycin (105 mg.) is dissolved in pH 7,0.1 N phosphate buffer (5 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing 1 N NaOH. To this solution is added methyl oxalimidate (200 μί) while the pH is maintained at 8.5. After 30 minutes the pH is adjusted to 7.0 using 2.5 N HCl and the solution is chromatographed on Dowex 50-X4 resin (53 cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The chromatography is carried out in water jacketed column at 3°. The methyl oxalimidoyl derivative elutes in 2 column volumes and is lyophilized to a white solid (44 mg.) uv (pH 7, 0.1 N phosphate buffer) 298 nm ( £ 6, 230) ir (Nujol mull) 1760 cm1 (β-lactam); nmr (D20) 6 1.27 (d, J= 6 Hz, CI^CH), 3.87 (s, -OCH3). 4 - 78 EXAMPLE 30 Preparation of N-Propionimidoyl thieriamcyin Th ι- OH NH II — NHC XcH2CH3 _ COOH Thienamycin (114 mg.) is dissolved in pH 7 0.1 N phosphate buffer (10 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing 1 N NaOH. Solid ethyl propionimidate hydrochloride (231 mg.) is added portionwise as rapidly as possible allowing the pH to be maintained near 8.5. After 30 minutes the pH is adjusted to 7.0 using 2.5 N HCl and the solution is chromatographed on Dowex 50-X4 resin (72 cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The Npropionimidoyl derivative elutes in 2 column volumes and is lyophilized to a white solid (76 mg.), uv (pH 7, 0.1 N phosphate buffer) λ 298 nm (£ 7,830) nmr (D„0) S 1.28 ΙΠ3.Χ Z (d, J = 6 Hz, CH3CH(0H)), 1.23 (t, J = 8 Hz, -CH^CH/, 2.50 (q, J = 8 Hz, CH^CH/ .

EXAMPLE 31 Preparation of N*-Methyl-N-formimidoyl thienamycin OH N-CH, Th • NHC COOH Thienamycin (140 mg.) is dissolved in pH 7 0.1 N phosphate buffer (10 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing 1 N NaOH. To this solution is added methyl N-methyl formimidate hydrochloride (200 μ.1) while the pH is maintained at 8.5. After 40 minutes the pH is adjusted to 7.0 using 2.5 N HCl and the solution is chromatographed on Dowex 50-X4 resin (72 cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The Ν'-methyl-N-formimidoyl derivative elutes in 2 column volumes and is lyophilized to a white solid (43 mg.). uv (pH 7, 0.1 N phosphate buffer) λ 298 nm (£7,250) ir (Nujol mull) 1765 cm-1 max (β-lactam). nmr (D20) ξ 1.29 (d, J = 6 Hz, CHg-CH), 2.92 (s, N-CH3) 7.80 (s, N-CH).

EXAMPLE 32 Preparation of Ν'-Benzyl-N-formimidoyl thienamycin Thienamycin (110 mg.) is dissolved in pH 7 0.1 N phosphate buffer (7 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing 1 N NaOH. A solution of ethyl N-benzyl formimidate fluroborate (572 mg.) in p-dioxane (2 ml.) is added to the buffered solution while the pH is maintained at 8.5. After 20 minutes the pH of the solution is adjusted to 7.0 using 2.5 N HCl and chromatographed on Dowex 50-X4 resin (53 cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The chromatography is carried out in a water jacketed column at 3°. The N'-benzyl-N-formimidoyl derivative elutes in 2 column volumes and is lyophil issd to a Mite solid IS mo,), w ίρΗ 7 0,1 sphosphate -1 buffer) λ 295 nm (£3,980) ir (Hujol mull) 1765 Oft max (β-lactam) nmr (D20) £1.29 (d, J = 6 Hz, CH^CH), 4.44 (s, CH2-Ar), 7,37 (s, Aryl), 8.14 (s, NCH).

EXAMPLE 33 Preparation of Ν'-Isopropyl-N-formimidoyl thienamycin Th Γ OH ΝΟΗ(ΟΗ3)2 NHC COOH Thienamycin (110 mg.) is dissolved in pH 7 0.1 N phosphate buffer (7 ml.) and the pH of the solution is adjusted to 8.5 using an automatic burette dispensing 1 N NaOH. A solution of methyl N-isopropyl formimidate hydrochloride (300 mg.) in jq-dioxane (1 ml.) is added to the magnetically stirred buffered solution while the pH is maintained at 8.5. After 25 minutes the pH of the solution is adjusted to 7.0 using 2.5 N NaOH and chromatographed on Dowex 50-X4 resin (53 cc, Na+ cycle, 200-400 mesh) eluted with deionized water. The chromatography is carried out in a water jacketed column at 3°C. The N1-isopropyl-N-formimidoyl derivative elutes in 2 column volumes and is lyophilized to a white solid (12 mg.). UV (pH Ί, 0.1N phosphate buffer) \,ax 299 nm (£8,130) ir (Nujol mull) 1760 cm 1 (β-laetam) nmr (DjO) £1.26 (d, J= 6Hz, CH3CH(0H)), 1.29 (d, J=6Hz, CH(CH3)3), 7.89 (s, NHCH), 7.96 (s, NHCH).

EXAMPLE 34 Preparation of N(N'-Allyl-formimidoyl)thienamycin <44433 - 81 Th Γ OH N' II •NH-C, L COOH .CH2CH=CH2 To a prechilled sample of thienamycin (123 mg., 0.452 mmole) is added 13 ml. of cold 0.1 N phosphate buffer. The solution is adjusted to pH 9 with N sodium hydroxide. To this basic solution at 2°C. is added all at once ethyl N-allyl-formimidate hydrochloride (0.3 g.) The pH dropped to 7.3 and is brought back to 8.5 with additional sodium hydroxide. The reaction mixture is stirred at 2°C. for an additional 30 min. and the pH is adjusted to 7 with cold 0.1 N sulfuric acid. The reaction mixture is assayed using high pressure liquid chromatography on a C. -Porosil column, developed with 10% aqueous tetrahydrofuran and is found to show only trace amounts of thienamycin (retention time, 5 min.) and substantially pure product (retention time, 10.5 min.). The reaction mixture is chromatographed on a Dowex-50x4 column (60 ml., Na cycle 200-400 mesh) eluting with water at a flow rate of 0.5 ml/min/cm of resin bed. After discarding the first 400 ml. of eluate, the next 150 ml. is lyophilized to give the product. Yield 96 mg. (63%). U.V. λ 301 nm, 24.60 Du/mg. (NH20H extinguished) 90% purity. IR Nujol exhibits C=0 at 5.67 μ and 5.90 μ. NMR 100 MHz D20 shows it is a 1:1 mixture of syn- and anti-N(N'allyl-formimidoyl)thienamycin. 4433 EXAMPLE 35 Preparation of N(N*-Trifluoroethyl-formimidoyl)thienamycin Th OH NH-C N-CH-CP ll 2 3 X, L COOH To a prechilled sample of thienamycin (123 mg., 0.452 mmoles) is added 15 ml. of cold O.lN phosphate buffer. The solution is adjusted to pH 9 with N sodium hydroxide. To this basic solution at 0-2°C. is added ethyl N-trifluoroethyl-formimidate (0.3 ml.) in dioxane (2 ml.) portionwise over 30 min. The pH of the reaction is maintained at 8.5-9 during the addition. The reaction, mixture is stirred for a few minutes. After the addition of imidate is completed and the pH is brought to 7 with cold O.lN H-SO. . HPLC, C,„ Porosil reverse phase, using 10% aqueous tetrahydrofuran exhibits a new peak at 12.2 min. assayed to the desired product.

The mixture is chromatographed on a Dowex 50x4 column (60 ml. 200-400 mesh). The column is eluted with water 2 at a flow rate of 0.5 ml/min/cm of resin bed. The forerun is discarded and fractions containing the 20 product are combined and lyophilized to give a hygroscopic solid, 10.2 mg. 302 nm. 4 4 3 3 - 83 EXAMPLE 36 Preparation of N(N‘-Carboxymethyl-formimidoyl)thienamycin Sodium Salt Th OH ! •NHC N-CH-COOH ii X COONa Thienamycin (130 mg.) is dissolved in pH 7 0.1N phosphate buffer (4 ml.) and solid sodium ethyl N-carboxymethyl formimidate (500 mg.) is added at once in one portion. The pH of the solution is adjusted to 8.5 using an automatic burette dispensing IN NaOH. After 25 min. at pH 8.5 the solution is adjusted to 7.0 with 2.5 N HCl. The solution is then chromatographed on an ice-water jacketed column of Dowex 50-X4 resin (51 cc, Na cycle, 200-400 mesh) eluted with deionized water. The eluate of the first column volume was combined and concentrated to 7 ml. This solution was then chromatographed on an ice-water jacketed column of XAD-2 resin (53 cc) eluted with deionized water. The second through fourth column volumes were collected and combined and lyophilized to give sodium Ν,(Ν'-carboxymethylformimidoyl)thienamycin (25 mg.), uv (pH 7 0.1N phosphate buffer) λ 300 nm _ Ώ13Χ (£6, 390) ir (Nujol mull) 1755 cm (β-lactam) nmr (D20) 6 1.29 (d, J= 6Hz, CH3CH), 7.85 (s, NCH).

EXAMPLE 37 Preparation of N-(3-Azidopropionimidoyl)thienamycin Th OH NH Nh£cH2CH2N3 L COOH To a solution of thienamycin (133 mg.) in 10 ml.

O.IMpH 7.0 phosphate buffer is added 1.2 g. of 0-ethyl3-azidopropionimidate-HCl while the solution is maintained at pH 8.5 with 2.5 N NaOH. The mixture is stirred at O°C. for 0.5 hr., then is neutralized with - 2.5N HCl to pH 7.0, concentrated to 5 ml. and chromato10 graphed on a Dowex 50 W x 8 (Na form) column (1.5 x 12) which is eluted with water to give 30 mg. of the desired H 0 product. The product shows UV λ 300 nm; high pressure liquid chromatography (HPLC) retention time of 10 min. with comparison to that of 4.8 min. of the starting material under the same conditions (1/8 x 2', Bondapak C,- reverse phase column eluted with 10% THP 18 in water at flow rate of 1.5 ml./min.); Electrophoretic mobility 5 mm toward cathode at 50V/CM for 20 min. in 0.05M pH 7.0 phosphate buffer. - 85 EXAMPLE 38 Preparation of N-(3-aminopropionimidoyl)thienamycin Th Th OH NH II NHCCH CHN H Pd —->. u_CO2 θ Na© OH NH nhcch2ch2nh3 CO, N-(3-azidopropionimidoyl)thienamycin (I) (43 mg in 40 ml 5 water) is hydrogenated under 1 atm of H2 in the presence of 0.1 g catalyst (10% palladium on charcoal) for 30 minutes. Electrophoresis of the resulting mixture shows a new bio-active product which moves 30 mm toward cathode (50V/CM for 20 min in 0.05 M pH 7.0 phosphate buffer) in addition to the starting material (I) which moves 5 mm toward cathode. The electrophoretic mobility of the product is consistent with that of the expected product (XI). The resulting reaction mixture from the hydrogenation reaction is neutralized with 2.5 NHC1 and filtered from the catalyst. The filtrate is concentrated to 10 ml and chromatographed on XAD-2 resin (2.3 x 16 cm t column). The column is eluted with water to provide the desired product II as the hydrochloride after lyophilization (23 mg. N-(3-aminopropionimidoyl) thienamycin hydrochloride). U.V. Absorbance maximum - 86 in water equals 301 ran (absorptivity equals 7080).

I.R: Nujol Mull 1765 cm NMR 60 mHz (D30) S 1.30 ppm, (doublet, 3), g 2.60-3.72 ppm (multiplet, 11) and 64.18 ppm (multiplet, 2).

EXAMPLE 39 Preparation of N-Nitroguanyl thienamycin Thienamycin (131 mg) is dissolved in a solution of dimethyl sulfoxide (10 ml), tri-n-butylamine (0.30 ml), and 2-methyl-l-nitro-2-thiopseudourea (0.3 g). The solution is heated in a water bath at 45°C. while a stream of nitrogen is vigorously bubbled into the solution. After 50 min. the solution is concentrated under high vacuum to 1,0 ml. and dissolved in 0.05N pH 7 phosphate buffer (7 ml). The unreacted thiopseudourea is precipitated and removed by filtration. The solution 3 is then chromatographed on Dowex 50-X4 resin (53 cm , + 200-400 mesh, Na cycle) and eluted with water. The N-nitroguanyl derivative elutes in the first column volume and is lyophilized to a solid (23%).

UV (pH 7’ 0.IN phosphate buffer) \,ax 269 nm (£.11,000) Electrophoresis (40 v/cm, pH 7 0.1N phosphate buffer, min) 3.0 cm towardcathode.

EXAMPLE 40 Preparation of the N-lsobutyrimidoyl thienamycin Following the procedure of Example 12 but replacing ethyl acetimidate hydrochloride with isobutyrimidate hydrochloride and allowing the reaction to proceed at °C. and pH 8.2 there is obtained N-isObutyrimidoyl thienamycin (14%). , UV (pH 7 0.IN phosphate buffer) 298 ran (£8,290) incLX NMR (D20) S 1.27 (d, J = 7Hz, CH(CH3)2, 1.29 (d, J= 6Hz, CH CH(OH), 2.79 (heptet, J = 7H , □ . z CH(CH3)2). 4 4 3 3 EXAMPLE 41 Preparation of Ν'-Methyl-N-acetimidoyl thienamycin Following the procedure of Example 12, but replacing ethyl acetimidate hydrochloride with methyl-N-methyl acetimidate, there is obtained N-methyl-N'-acetimidoyl thienamycin (10%) .

UV (pH 7 0.1N phosphate buffer) ^max 298 nm (£6,700) IR (Nujol mull) 1750 cm 1 (β-lactam), 1660 cm 1 (C=NCH3) NMR (D,0) fi 1.27 (d, J=6H , CH CH(OH), 2.22 and 2.25 A Z 3 N (S, N-CCH3), 2.97 (S, NCH/ EXAMPLE 42 Preparation of Ν'-Methyl-N-formimidoyl thienamycin Following the procedure of Example 12, but replacing ethyl acetimidate hydrochloride with ethyl N-methyl formimidate hydrochloride there is obtained N'-methyl-Nformimidoyl thienamycin (10%).

UV (pH 7 0.1N phosphate buffer) λ 298 nm 1ΏΗΧ NMR (D20) 6 1.30 (d, J=6Hz, CH3 CH(OH), 2.92 (S, N-CH/, N 7.78 (S, -CH) EXAMPLE 43 Following, the procedure of Example 12, but replacing the reagent with an equivalent amount of methyl methoxyacetimidate, there is obtained: N(methoxyacetimidoyl) thienamycin (34%); H2° UV λ 198, 301 nm (ε16,180, 8,700); max IR (Nujol mull) 1760 cm (β-lactam); NMR (60 MHz, D20) 6'l.28, 3H, (d, J=6Hz, CH/CHiOH); 3.50, 3H, (S, CHg.O.CH/ 6 4.35, 2H, (S, CH .O.OT/; HPLC, 1.50 x retention time of thienamycin.

EXAMPLE 44 Preparation of Ethyl N-Methoxyformimidate A mixture of methoxylamine hydrochloride (0.020 mole 1.6700 g) and anhydrous potassium carbonate (0.010 mole, 1.3821 g) is dissolved in 7.0 ml water. Ether 80 ml is added and the reaction mixture is treated with ethyl formimidate hydrochloride (0.02 mole, 2.1900 g). The mixture is shaken for 15 minutes. The ethereal layer is separated and the aqueous layer is extracted with two portions of ether (30 ml). The combined and dried ethereal solution is evaporated to give 0.8433 g of ethyl N-methoxyformimidate. nmr 6 1.36 (triplet) 8 3.83 (singlet) 6 4.13 (quartet) 6 6.56 (singlet) EXAMPLE 45 Ethyl N-(2,2,2-trifluoroethyl)formimidate Ethyl formimidate hydrochloride (0.555 g, 5 mmole), 2,2,2-trifluoroethylamine hydrochloride (o.677 g, 5 mmole) and potassium carbonate (0.345 g, 2.5 mmole) are suspended in 20 ml CH2C12 and treated with 2 ml H20. The mixture is shaken vigorously for 3 minutes. The organic phase is separated and the aqueous extracted twice with 10 ml portions of CH2Cl2. The combined organic phase is dried and the CH2Cl2 distilled through a Vigreaux column to give the ethyl N-(2,2,2-trifluoroethyl)formimidate. n.m.r. β 1.33 t (eH3CH2); 3.8 q (j = 10 e.p.s., CP3 CH2) 4.23 q (j = 7.5, CH3CH2O); 7.6 S (H-C=N) .

EXAMPLE 46 Preparation of Ethyl N-ethoxycarbonylethyl-formimidate Ethyl formimidate hydrochloride (0.55 g, 5 mmole), ethyl glycinate hydrochloride (0.687 g, 5 mmole) and potassium carbonate (0.345 g, 2.5 mmole) are suspended in 20 ml CHjClj and treated with 2 ml HjO. The mixture is shaken vigorously for 4 minutes. The organic phase is separated, the aqueous phase is extracted twice with CHjCl,, (10 ml) and the combined organic phase is dried and evaporated to give ethyl N-ethoxycarbonylmethylformimidate. n.m.r. S : 1.26 t (CH^—CH,,) ί 4.06 S (N-CHj-C); 4.23 g (CH3CH2-O); 7.5 S (N=CH).

EXAMPLE 47 Preparation of Potassium N-ethoxycarbonylmethylformimidate EtO C=N-CH,-COOEt / * KOEt, Et2O, EtOH C=N-CH -COOK EtOZ Potassium (0.18 g) is dissolved in a mixture of 0.6 g EtOH and 4 ml Et20 under N2. The solution is diluted with 50 ml of Et2O and ethyl N-ethoxycarbonylmethyl-formimidate (0.79 g) in 2 ml EtjO is added, followed by 0.1 ml H20. Rapid crystallization of the salt takes place. The solid is filtered, washed with ether and dried under vacuum to give potassium N-ethoxyoarbonylmethylformimidate. n.m.r. (D.,0). 1.13 t (CH3CH2); 3.63 g (CH3-CH20); 3.8 S (N-CH2-C); 8.06 S N=CH. <.4533 - 90 EXAMPLE 48 Preparation of Ethyl N-Benzylformimidate A solution of 690 mg (5.1 mmoles) of N-benzylformamide in 5 ml of methylene chloride is cooled in an ice-water bath and put under an argon blanket. The solution is stirred while 4.9 ml (4.9 mmoles) of IM triethyloxonium fluoroborate in methylene chloride is added dropwise. After a 45 minute reaction time, the mixture is concentrated to dryness under reduced pressure at room temperature, and the residue is dried under reduced pressure over P2°5' T^e nuclear magnetic resonance spectrum of the product in deuterochloroform is fully in accord with the product being a fluoroborate etherate complex of ethyl N-benzylformimidate.

EXAMPLE 49 Preparation Of N-isopropyl formamide Formamide (1.13 g, 0.98 ml) is dissolved in 10 ml of toluene, containing toluenesulfonic acid (4.7 g).

To the above mixture is added isopropylamine (2.95 g., 4.25 ml). The mixture is refluxed overnight under a gentle stream of N2· The solution is filtered and the toluene is evaporated under reduced pressure.

The residual oil is distilled at 59-62°C/.O7 mm to give 1.0 g of the desired product.

EXAMPLE 50 Preparation of Methyl N-isopropyl formimidate Isopropyl formamide (535 mg) is treated with an equivalent amount of ethyl chloroformate (440 μί) for 2-3 hours under N2 at 40-45°C. The mixture is washed successively with petroleum ether anhydrous ether and benzene leaving the product as an oil.

EXAMPLE 51 Preparation of N-fN'-Ethylformimidoyljthienamycin Th OH NEt II -NHC-H co2h Thienamycin (100 mg) in 10 ml 0.1M pH 7.0 phosphate buffer is adjusted and maintained at pH 8.5-9.0 with 2.5 N sodium hydroxide. To the solution is added 300 mg. of ethyl N-ethylformimidate hydrochloride. The mixture is o stirred at 23 C for 20 minutes, then is neutralized to pH 7.0 with 2.5 N HCl and chromatographed on a Dowex 50-X8 (Na form) ion-exchange column (1.5 x 10). The column is eluted with water taking 6.7 ml fractions. Fractions 40-90 are combined, concentrated and freeze-dried to give 15 mg of the solid product. Electrophoresis of the product at 50 v/cm for 20 minutes in 0.1 M, pH 7.0 phos15 phate buffer shows a single bioactive zone which moves H 0 mm toward the cathode. UV λ 2 301 nm; nmr (100 max MH2, D20); ¢7.77 (S) and 7.82 (S) (formimidoyl CH). EXAMPLE 52 Preparation of N-fN'-cyclopropylformimidoyl] thienamycin _ COOH Thienamycin (100 mg) in 10 ml 0.1 M, pH 7.0 phosphate buffer is adjusted and maintained at pH 8.5-9.0 while 300 mg of ethyl N-cyelopropylformimidate hydrochloride is added dropwise to the solution. The mixture is stirred at 23° for 40 minutes, then is neutralized, and chromatographed on a Dowex 50-x 8 (Na form) ion-exchange column (1.5 x 10). The column is eluted with water, collecting 6.5 ml fractions. Fractions 43-95 are combined, concentrated and freeze-dried to give 54 mg of the solid product. Electrophoresis of the product shows a single bio-active zone which moves 10 mm toward the cathode (50 V/CI1, 1 hour H 0 in 0.05 M pH 7.0 phosphate buffer). uv ^^2 301 nm; Nmr (100 MHz, ^0); 0.60-1.30 ppm (m, cyclopropyl) and 7.80 ppm (formimidoyl CH).

EXAMPLE 53 Following the procedure set forth in the foregoing text and Examples, the following compounds of the present invention are obtained. The reagents, imido ethers and imido halides, utilized irt the reaction with thienamycin, or a derivative thereof, to provide the following compounds are either known, or may be prepared as described above.

Compound R 1. ) H 2. ) H 3. ) H 4. ) H . ) H 6. ) H 7. ) H 8. ) H 9. ) H . ) H 11. ) H 12. ) H -CH2CH2CH2CH3 -CH -CH-CH 2 j J CH„ -CHCH CH I z J ch3 -CH2CH2CH2CH2CH3 -ch-ch2ch2ch3 CIL -ch„ch-ch„ch L I CH_ -CH-CH— CH i I 3 ch3 ch3 ch3 -C — CH-CH, ι z JCH3 -ch2-c(Ch3)3 -CH2CH2CH2CH2CHCH3 -CH-CH2CH(CH3)2 CH -CH -C=CH„ 2 | 2 CIL H H H H H H H H H H H H ’ . *4 4 3= ' ·· :> ·. μ ' ,·^ · ' 1 !· 7 94 - - ; \ . <· 1 , ·)(.-> · r μ. ϊ' .' '· 1 · , ί ' · ι Compound R 1 1 ? R ’ ' ' \! 1^ ’ 13,) Η . -CH2-CH=CH-CH3 H J. ;/·- . I4·) Κ Η · • -CH-CH=CH2 .. H ' ’ ' .' *, ί : · ί * . 1 j «, i ι CH3 ’ , 15.) , . *- -- ‘Η, -ch2ch2ci-i=ch2 , ' H ‘ i ‘ . 5 ’ ‘ 16.) ί !\ Η -CH2CH2-CH=CHM3H3J > J H - -. 1 17 ·> ( ) Η . < -c^h^ ; ; CHM ·· % ' H r: 18.) ' Η - -CH -CH-CH=CH„ 1 ; - . 2 1 2 -. H _ . . : • A; 19.) Η , -CH—C=CH , H ί ,CIi3 CH3 ' 20.) ! / :Η; * -CH-CH=CH 1 -- i 2 , W H 10 21.) Η -CH2CB=CH-CH2CH2CH3 H5 22.) Η : -CH2-CH2-CH=CH-CH2CH3 H 23.) Η -CH-CH=CH-CH„CH_ HCH3 24.) Η. -CH-CH--CH==CH —CH j 2 2 J ch3 H I 25.) Η -CHCH--CH-CH =CH 1 2 2 2 H 4 4 3 3 Compound 27.) 28.) 29.) .) 31.) 32.) 33.) 34.) H 43 3 36.) -ch2-ch=/ 37.) 38.) 39.) 40.) 41.) 42.) 43.) •a. 4 ύ 3 3 Η Compound R R1 R2 61.) H -CoHc 2 5 -¾ 62.) H -ch(ch3)2 -CH(CH3)2 63.) H Ws 64.) H ^3 -C2H5 65.) H -ch3 -CH(CH3)2 66.) H -^3 -CH2CH=CH2 67.) H -CH2CH=CH2 -CH2CH=CH2 68.) H -c(ch3)3 -ch3 69.) H -CH3 -ch-c2h5 ch3 70.) H -C2H5 -ch(ch3)2 71.) H -ch3 -CHCH-CH. 1 2 ^3 72.) H -CH3 -CH2CH(CH3J2 73.) H -ck3 -ch2ch2ch3 74.) H -ch3 -CH20 75.) H "CH3 -0 76.) H -ch3 77.) H -ch3 -CH-CH=CH„ 2 ι 2 Compound - V,| -h - 100 78.) Η -CH3 79.) Η -ch3 80.) Η -ch3 81.) -®3 -C2H5 82.) -CH3 -CH2CH=Ci 83.) -CH3 -ch(ch3) 84.) -CH=CH2 -ch3 85.) -CH=CH2 -C2H5 86.) -CH=CH2 -ch(ch3), 87.) Ζ -C(CH3>3 88.) -ch3-CH2 Χ 89. ) 90. ) 91. ) -CH"CH-CH-ch2 -ch2 Η Η Η Η Η Η Η -o 2c6h5 - 101 Compound R 92. ) 93. ) 94. ) 95.) 96. ) 97. ) 98. ) -CH.

-CH.

-CH, -CH.

-CH=CH„ -CH.

-CH. -ch2_// Xn -CHCH=CH_ I iCH3 -CH.-C"CH_ j 2 CH. -ch2-ch2ch3CH3 -CH, Vs H H H H H H H 99.) -CH3 100.) -ch3 101. ) CH <3 102. ) CH3 103. ) CH3 104. ) CH3 ch3 CH3 CH. C.H, 3 2 5 ch3 -ch(ch3). CH. CH.CH=CH, - 102 Compound R 105.) CH C2H5 106.) CH CH 107.) CB_ CHC2H5 108.) CH -CH CH=CB> Ji 109.) CH3 C2Hs "CH, 2 \o -CH„—,-N 110.) 111.) CH CH 3 COOH H 112.) H 113.) H 114.) H - 103 Compound 115. ) 116. ) 117.) 118.) 119. ) 120. ) 121.) 122.) H H Ν' H H N7 I CH_ <,4433 - 104 Compound R 123.) CH124.) CH125.) 126.) 127.) 128. 129.) % Compound R 130.) H 131.) H • CH. 132.) 133.) 134.) H 135.) 136.) H -CH. '2 1 CH, -ch2 N-CH, H -"AJ -Ν H V‘> - Ψ * Ϊ06 Compound 137. ) H 138. ) H 139. ) H 140. ) H 141. ) H 142. ) H 143. ) H -CH, -CH„ -CH, -CH. 0JI -CH, 144.) H -CH„ z=\ 145.) H - 107 Compound R 146.) -CH// 147.) 148.) 149.) 150. ) 151. ) 152. ) 153. ) 154.) 155. ) 156. ) 157. ) 158. ) H H α H H H H -CH2-O-CH3 -CH2CH2-O-CH3 -CH2CH2-S-CH3 -CH2CH2-OH -CH_CH-CH2| 3 och3 -CH2CH2-CH2-CsN -CH2CH2CH-(OCH3)2 H H H H H H H H ' - 108 - Compound R R1 R2 159.) Η -CH_-C-3?h 2 H 0 H 160.) Η -CH—CH.CH. 1 2 3 ch2oh H 161.) Η ~ch2ch2conh2 H 5 162.) Η -ch2cooc2h5 H 163.) Η -CCH2CH2CH2N(C2H5)2CH3 H 164.) Η -CH2CH2-SH H 165.) Η -CH-// OH H 166.) Η -ch2ch2-n(ch3)2 H 10 167.) Η -CH2CH2CH2-Br H 168.) Η -ch2ch2ch»n(ch3)2 H 169.) Η -ch2ch2-n(ch3)3 H ί 170.) νη2 ch3 H 171.) νη2CH3CH3 15 172.) NHCH3 ch3 ch3 173.) »(CH3)2CH3 ch3 174.) νη2C2H5 H 175.) νη2 -ch(ch3)2 H 176.) νη2 -CHCH=CH2 H Compound R R1 R2 177.) nhch3 -ch(ch3)2 H 178.) nhch3CH3 H 179.) nh2 n(ch3)2 H 180.) nh2 NHNH2 H 181.)0CH3 H H 182.) och3 ch3 H 183.) OCH3CH3CH3 184.) och3C2H5 H 185.) och3 ch(ch3)2 H 186.) och3 H H 187 J sch3 ch3 H 188.) sch3CH3 ch3 189.) sch3 ch(ch3)2 H 190.) S-CH2~CH=CH2CH3 H 191.) S-CHj-PhCH3 H 192.) -SCH2CH=CH2 H H 193.) SCH2-Ph H H 194.) H -OH H 195.) H -och3 H 196.) CH, 3 -OH H 197.) H -C=N H 198.) H -nhch3 H 199.) H nh2 H Compound R R1 R2 200.)CH3 oh3 nh2 201.) HCH3 n(ch3)2 202.) CH^Br H H 203.) -ch2n(ch3; »2 H H 204.) -ch2s-ch3 H H 0 205.) 1 -c-nh2 H H 206.) -ch2n(ch3) '3 H H 207.) H 208.) -CHCHCH3 -CHCH, i 3 1 CHO CH_ 3 3 209.) -CHCH- -CHCH ι 3 -CHCH- CH, CH, CH 3 3 3 210.) -c(ch3)3 ~C^3 -c(ch3)3 - Ill Ο CM Θ rt! X-0=N-~H0HO-S Β Ο Β υ o ω ί* ί Ν Η Ν ω <Φ ιη Φ Η Η Η Η ι"Η Η <Ν Ν <Ν Ν Ν Ν - 112 -ι ο rd U α βι ο u ΰ S $ •rl ν= ? =ο ΐ V=° CQ Η & ti C! •Η ιη I βν V cs —Φ Λ <Ρ Χί •Ή W -Ρ rd r-i s pS Κ) Λ ίΐ ft )=Ο Β U υ & ϊ £\ ? αί ¢5 Η Μ (*} Ο (0 ο Οι ο Οι •—v X*. «Μ. X*. • • • • • • • • C* ω 0 rt CM CO Xf H rd rt CM CM CQ CM CM CM CQ CQ CM CM CM CM CM xf CQ CQ Ο 4J rd CM CM 0) CJ i s in - 113 Example 54 Preparation of Pharmaceutical Compositions One unit dosage form is made by mixing 120 mg. of N-acetimidoyl thienamycin (product of Example 12) with 20 mg. of lactose and 5 mg. of magnesium stearate and placing the 145 mg. mixture into a So. 3 gelatin capeule. Similarly, by using more of the active ingredient and less lactose, other dosage forms can be put up in No. 3 gelatin capsules and should it be necessary to mix more than 145 mg. of ingredients together, larger capsules such as compressed tablets and pills can also be prepared. The following Examples are illustrative of the preparation of pharmaceutical formulations: Example 55 TABLET PER TABLET N-acetimidoyl thienamycin 125 mg.

Cornstarch, U.S.P. 6 mg.

Dicalcium Phosphate 192 mg.

Lactose, U.S.P. 190 mg.

The active ingredient is blended with the dioaleium phosphate, lactose and about half of the cornstarch.

The mixture is then granulated with a 15% cornstarch paste (6 mg.) and rough-screened. It is dried at 45°C. and screened again through No. 16 screens. The balance of the cornstarch and the magnesium stearate is added and the mixture is compressed into tablets, approximately 0.5 inch in diameter each weighing 800 mg.

Example 56 PARENTERAL SOLUTION Ampoule: N-acetimidoyl thienamycin Sterile water 500 mg. ml. 4 -i 3 3 ’’114 Example 57 OPTHALMIC SOLUTION N-acetimidoyl thienamycin Hydroxypropylmethyl cellulose Sterile Water to Example 58 OTIC SOLUTION N-acetimidoyl thienamycin Benzalkonium chloride Sterile Water to Example 59 TOPICAL OINTMENT N-acetimidoyl thienamycin. Polyethylene glycol 4000 U.S.P.

Polyethylene glycol 400 U.S.P. 100 mg. 5 mg. 1 ml. 100 mg. 0.1 mg 1 ml. 100 mg. 400 mg. 1.0 gram The active ingredient in the above formulations may be administered alone or in combination with other biologically active ingredients as, for example, with other antibacterial agents such as lincomycin, a penicillin, streptomycin, novobiocin, genfcamycin, neomycin, colistin and kanamycin, or with other therapeutic agents ; such as probenecid.

As already mentioned above, position isomers of thienamycin as well as thienamycin itself, either alone or as mixtures, may serve as starting materials in the preparation of the compounds of the present invention.

Some of these isomers are obtainable from natural products of fermentation as described and claimed in the specifications of our copending application British Patent Specification No. 1561109. See in particular - 115 Examples 6 and 7 of that specification, which describes the preparation of Antibiotics 890A^ and 89QA3> which have the formula : These compounds can be treated to cleave the N-acetyl group to give the corresponding free base of formula : By total synthesis all isomers can be made available (below) as a mixture of 4 diastereoisomers which possess antibacterial activity and which are amenable to resolution by conventional techniques. The 4 diastereoisomers (2 cis, 2 trans) are separable by chromatography.

Resolution of any given d/1 pair with optically active acids or bases proceeds according to conventional techniques. It should be noted that the absolute configuration of the first-identified starting material (I) is 5R 6S 8R. - 116 Preparation of Thienamycin by Total Synthesis SCH^NH,, COOH Step A; Preparation of 4-(2-acetoxyvinyl)azetidine-2-one HC=CH-CH=CH0C-CH, + O=C-N-SO„C1 _:_ || 3 2 CH=CHOCCH CH=CHOCCH, . NH SO2C1 A solution of 1.0 ml distilled chlorosulfonylisocyanate (1.65 g; 11.7 mmoles) in 2.5 ml anhydrous diethyl ether is cooled under N2 in a -20°C bath.

A solution of 2.5 g 1-acetoxybutadiene (22 mmoles) 10 in 2.5 ml anhydrous ether is similarly cooled under N2 in a -20°C bath.

The chlorosulfonylisocyanate solution is added dropwise to the acetoxybutadiene solution by means of a Teflon tube immersed in the CS1 solution and pressurized with N2· The addition takes 10 minutes. Little or no color is seen and the reaction is stirred at -20°C for 0.5 hour. The solution is clear and has a light yellow color. - 117 A solution of 2 g sodium sulfite and 5 g KjHPO^ in 20 ml H20 is prepared during the above 0.5 hour reaction time and is cooled in an ice bath; 20 ml of ether is added and the mixture is vigorously stirred in an ice bath. At the end of the 30 minute reaction time, the reaction mixture is transferred, again using N2 pressure and the Teflon tube, from the reaction flask which is maintained in the -2Q°C bath, to the vigorously stirred hydrolysis mixture. Rapid dropwise addition is completed in 5 minutes. The hydrolysis is allowed to continue for 5 additional minutes. The hydrolysis mix has a pH of 6-8, preferably pH 8.

The phases are separated, leaving a yellowish-orange gum with the aqueous phase. The ether phase is dried directly with MgSO^. The aqueous/gum phase is extracted three more times with 50 ml portions of ether, each being added to the initial ether/MgS04· The dried extracts are filtered and concentrated under a N2 stream to 5 ml; a portion of the product is crystalline at this stage.

A column of 10 g Baker silica gel, packed in ether is prepared, and the ether concentrate is applied to the top and run in. The flask/solids are rinsed three times with 2 ml ether, each being pipetted off and run into the column. Elution is then begun with ether.

The first 25 ml is primarily void volume. The next five 10 ml fractions are collected followed by three 50 ml fractions, and all are reduced in volume under a N2 stream. The product crystallizes from fractions 4-6, with traces in 3 and 7. Fractions 1-3 contain a yellowish sharp-smelling material which resinifies on standing. Yield: 100 mg as a mixture of the cis and trans isomers. 118 Step Β: Preparation of 4-(2-Acetoxyethyl)-2-Azetidinone . NH ^Z0CH3 A solution of 4-(2-acetoxyethyl)-2-azetidinone (10.Og, 0.065 mole) in 200 ml ethyl acetate containing 100 mg of 10% Pd/C is hydrogenated on a Parr shaker at 25°C under 40 psi hydrogen for 15 minutes. The mixture is filtered through a bed of Supercel and washed with additional ethyl acetate. The combined filtrate is evaporated in vacuo to give 4-(2-acetoxyethyl)-2azetidinone (10.0 g) as a crystalline solid. Recrystailization from ether affords white crystals: M.P. 44-7°; ir.CCHCl/^ 5.66, 5.74? nmr (CDCl/ r 3.44 (broad s, 1, NH), 5.82 (m, 2, CHjOCOCH/, 6.29 (m, 1, C-4H), 6.87 (1/2 AB pattern further split in four by C-4H'and NH, 1, J = 12.8Hz, J = 4.5 H, - 1.9 Hz, 7.38 (1/2 AB pattern further split in four by C-4H and NH, 1, J = 12.8Hz, J = 2.3Hz, gem - 1.0Hz), 7.93 and 8.02 (s on m, total 5, OCOCHj and CHjCHjOCOCHj, respectively).

Step C: Preparation of 4-(2-Hydroxyethyl)-2-Azetidinone - 119 Under nitrogen at 0°, a solution of 4-(2-acetoxyethyl)-2-azetidinone (2.24 g, .014 mole) in 25 ml anhydrous methanol is treated with a solution of sodium methoxide (77 mg, 1.4 mmoles) in 5 ml anhydrous methanol. After stirring for 1 hour, the solution is neutralized with glacial acetic acid. Removal of the methanol in vacuo gives crude 4-(2-hydroxyethyl)-2-azetidinone as an oil.

The product is purified by chromatography on silica gel eluting with 10% MeOH/CHCl3 to give 1.55 g of the alcohol: m.p. 50°; ir (CHCl3) μ 5.67; nmr (CDCl^) T3.2O (broad s, 1, NH), 6.24 and 6.28 (m on t, total 3, C-4H and CHgOH respectively), 6.90 (broad s on 1/2 AB pattern further split in four by C-4H and NH, total 2, OH and C-3H respectively, Jgem = 13.0Hz, Jv|c = 4.2Hz, = 1.6Hz), 7.42 (1/2 AB pattern further split in four by C-4H and NH, 1, C-3H, J = 13.0Sz, = gem vie 2.2Hz, = 1.1Hz), 8.16 (m, 2, CHgCHgOH).

Step D: Preparation of 8-Oxo-2,2-dimethyl-3-oxa-l-azabicyclo Γ4.2 .ol octane A solution of 4-(2-hydroxyethyl)-2-azetidinone (1.87 g, .016 mole) and 2,2-dimethoxypropane (1.69 g, .016 mole) in 25 ml anhydrous methylene chloride is treated with boron trifluoride etherate (.201 ml, .002 mole) at 25°C. The resulting solution is stirred for ten minutes. Removal of the solvent under reduced ,54433 - 120 pressure gives an oil (2.5 g). Chromatography of the crude product on silica gel using 2:1 ethyl acetate/ benzene as eluting solvent gives 8-oxo-2,2-dimethyl-3oxa-1-azabicyclo[4.2.0]octane (1.59 g) as a crystalline solid. Recrystallization from ether/hexane gives product of m.p. 60-1°. ir (CHC13)u : 5,73 (β-lactam) nmr (CDClg)?*: 6.02 - 6.28, m, 2H, C-4 methylene 6.22 - 6.62, m, IH, C-6 methine 6.90, dd, IH, J, _ = 14Hz, tf, , = 4.5Hz 7,7 6,7 C-7 proton cis to C-6H 7.47, dd, IH, J, „ = 14Hz, J, , = 2Hz C-7 proton trans to C-6H 7.82 - 8.68, m, 2H, C-5 methylene 8.23, s, 3H ) Step E: Preparation of 8-oxo-2,2-dimethyl-7a and B-(l-hvdroxvethyl)-3-oxa-l-azabicvclor’4.2 .ol octane OH To a solution of 1.1 equivalents of freshly prepared lithium diisopropylamide in anhydrous tetrahydrofuran under a nitrogen atmosphere at -78° is added a solution of 8-oxo-2,2-dimethyl-3-oxa-l-azabicyclo[4.2.0]octane in anhydrous tetrahydrofuran whieh has - 121 been cooled to -78°C. After two minutes, the resulting lithium enolate is treated with excess acetaldehyde.

The solution is stirred for 30 minutes at -78° and then poured into water. The aqueous phase is saturated with sodium chloride and extracted with ethyl acetate. The combined ethyl acetate solutions are dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give the crude product. Purification by chromatography on silica gel using ethyl acetate/ benzene gives 8-oxo-2,2-dimethyl-7a and β-(1-hydroxyethyl)3-oxa-l-azabicyclo[4.2 .0]octane.

Data for 8-oxo-2,2-dimethyl-7p-(l-hydroxyethyl)-3oxa-l-azabicyclo[4.2.0]octane : ir (ΟΗ2Ο12)μ : 5.72μ (β-lactam) nmr (CDCl^) T : 5.53 - 6.43, m, 4H, C-4 methylene + C-6 methine + C-9 methine 6,90, dd on broad s, 2H, J = 9Hz v t yJ6,7 = 5 •5Hz, C-7 methine + OH 7.70 - 8 .83, m, 2H, C-5 methylene 8.27, s. 3H ) 8.60, s,3H ) C-2 methyl 8.78, d. 3H, Jg 1Q = 6.5Hz, C-10 methyl Data for 8-oxo-2,2-dimethyl-7a-(1-hydroxyethyl)-30xa-l-azabicyclo[4.2.0] octane : ir (CHC13)h : 2.9 broad 0-H .73 β-lactam nmr (acetone -d, )6 : 4.23 - 3.33, m, C-9 methine + C-4 o methylene + C-6 methine 3.33, broad s, OH - 122 2.83, dd, J = 2Hz, 6Hz ) 2,67, dd, J = 2Hz, 8Hz j C7 methine 1.93 - 1.63, m, C-5 methylene 1.63, s ) 1.40, s } C_2 methyls 1.23, d, J = 6.5Hz, C-10 methyl * -’t ·-- .

Step F: '·- Preparation of 8-0xo-2,2-dimethyl-7a^(l-p-nitrobenzylcarbonyldioxyethyl)-3-oxa-1-azabicycloΓ4.2.0] octane Under anhydrous conditions at 0°C. a solution of 8-oxo-2,2-dimethyl-7 a-(1-hydroxyethyl)-3-oxa-l-azabicvclo[4.2.0]octane (60 mg, .302 mmole) in 0.6 ml ether is treated with powdered potassium hydroxide (19 mg, .332 mmole). After a period of 15 minutes, 33-nitrobenzyl chloroformate (65 mg, .302 mmole) is added to the reaction mixture. Stirring is continued at 25°C for an additional 15 hours. The mixture is partitioned between 1M pH 7 phosphate buffer and more ether. The ether phase is washed with water and brine, dried over magnesium sulfate and filtered. Evaporation of the filtrate under reduced pressure gives 67 mg of a colourless oil. Purification by preparative thick-layer - 123 4 4 3 3 chromatography on silica gel developing with 1:9 ethyl acetate/benzene gives 8-oxo-2,2-dimethyl-7a-(l-p-nitrobenzylcarbonyldioxyethyl)-3-oxa-1-azabicyclo[4.2.0]octane (40 mg) as a mixture of diastereomers. ir (CH2C12) u : 5.68 (β-lactam and carbonate), 6.19 and 6.54 (nitro) nmr (CDCl^) : 1.67, d, 2H, ArH 2.37, d, 2H, ArH 4.67, s, 2H, ArCH2 4.67 - 5.22, m, CH CH 5.98 - 6.25, m, 2H, C-4 methylene 6.25 - 6.62, m, IH, C-6 methine 6.75 - 7.12, m, IH, C-7 methine 7.75 - 8.83, m, 2H, C-5 methylene 8.22, s, 3H, C-2 methyl 8.50 - 8.58, m, 5H, c-2 methyl + CH.3CH The 73-diastereoisomers or the 7a and β-mixture are obtained in an analogous manner.

Step 6; Preparation of Cis and Trans-3-(l-p-nitrobenzylcarbonyldioxyethyl)-4-(2-hydroxyethyl)-2-azetidinone 44 3 3 - 124 -Oxo -3 -oxa -2,2 -dimethyl -7 a- (1 -p.-nitr obenzylcarbonyl dioxyethyl)-1-azabicyclo[4.2.0]ocrane (1.0 g) is dissolved in 8 ml acetic acid and 2 ml water and heated at 65°C for 1.25 hours. The acetic acid and water are removed under reduced pressure and the residue is taken up in benzene and evaporated to give trans-3-(l-p-nitrobenzvlcarbonvldioxyethyl)-4-(2-hydroxyethyl)-2-azetidinone as a mixture of diastereoisomers. ir (CH2Cl2)|i : 5.67 (β-lactam), 5.72 shoulder, 6.20 and 6.57 (nitro) nmr (CDClg) : 1.73, d, 2H, J = 8.5 Hz, ArH 2.43, d, 2H, J= 8.5 Hz, ArH 3.63, broad s, IH, NH 4.37 - 5.13, m, IH, CH3CH 4.72, s, 2H, ArCH2 6.07 - 6.53, m, IH, C-4 methine 6.23, t, 2H, j = 5.5 Hz, CE^OH 6.73 - 6.93, m, IH, C-3 methine 7.63 - 8.97, m, 3H, CH2CH2OH 8.53, d, J = 6.5 Hz, CH3CH The cis diastereoisomers or the cis-trans mixture are obtained in an analogous manner.

Steps D*, E', F' and G' as alternative to Steps D, E, F, and G for the preparation of 3-(l-p.-nitrobenzylcarbonyldioxyethyl)-4-(2-hydroxyethyl)azetidinone - 125 Steps D', E1, F1 and G1 Preparation of l-(2-Tetrahydropyranyl)-4-]2-(2-tetrahydropyranyl)oxyethyl]-2-azetidinone Under nitrogen and at 25°C. a solution of 4-(2hydroxyethyl)-2-azetidinone (62 mg. .539 mmole) in .5 ml of anhydrous p-dioxane is treated with 2,3-dihydropyran (.98 ml, 1.08 mmoles) and ρ,-toluenesulfonic acid monohydrate (19 mg, .10 mmole). The resulting solution is stirred for a period of 60 minutes and then partitioned between 10 ml qf .5M pH 7 phosphate buffer and 10 ml of ethyl acetate. The aqueous phase is extracted a second time with ethyl acetate. The combined ethyl acetate solutions are washed with brine, dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give 216 mg of crude product. Purification by preparative thick-layer chromatography developing with ethyl acetate gives 80 mg of 1-(2-tetrahydropyr anyl )-4-[2-(2-tetrahydropyranyl)-oxyethyl]-220 azetidinone as an oil. nmr (CDCl/T : 5.13 - 5.60, m, OCH .83 - 6.85, m, C-4H + OCHj - 126 6.95, dd, J = 7.35, dd, J = 7.62-8.95» m, + CHCHgCHgO Preparation of Cis and Trans-l-(2-tetrahydropyranyl)3-(1-hydroxyethyl)-4-(2-(2-tetrahydropyranyl)oxyethyl]2-azetidinone 5Hz and 15 Hz ) 3Hz and 15 Hz ) C-3 methylene CHCHgCHgCHgCHg Following the procedure described for the preparation 10 of 8-oxo-2,2-dimethyl-7a and β-(1-hydroxyethyl)-3-oxa1-azabicyclo[4.2.0]octane from 8-oxo-2,2-dimethyl-3oxa-1-azabicyclo[4.2.o]octane and using 1-(2-tetrahydropyranyl) -4-[2-(2-tetrahydropyranyl)oxyethyl] -24 4 4 - 127 azetidinone one obtains a diastereomeric mixture of both cis and trans-1-(2-tetrahydropyranyl)-3-(1-hydroxyethyl) 4-[2-{2-tetrahydropyranyl)-oxyethyl] -2-azetidinone.

Preparation of Cis and Trans-1-(2-tetrahydropyranyl)-3(1-p-nitrobenzylcarbonyldioxyethyl)-4-f 2-(2-tetrahydropyranyl)oxyethyl]-2-azetidinone R = c;OCH2 —// \\~,N02 Following the procedure described for the preparation of 8-ΟΧΟ-2,2-dimethyl-7 a-(1-ja-nitrobenzylcarbonyldioxyethyl) 3-oxa-1-azabicyclo[4.2«OJ octane from 8-oxo-2,2-dimethyl7a-(1-hydroxyethyl)-3-oxa-1-azabicyclo[4.2.0]octane and using trans-1-(2-tetrahydropyranyl)-3-(1-hydroxyethyl) 10 44433* .- 128 4-(2-(2-tetrahydropyranyl)oxyethyl]-2-azetidinone there is obtained trans-1-(2-tetrahydropyranyl)-3-(l-p-nitrobenzylcarbonyldioxyethyl)-4-(2-(2-tetrahydropyranyl)oxyethyl] -2-azetidinone. The cis diastereoisomers are obtained in an analogous manner. t Preparation of pi s_and JErane,-3-(l-p-nitrobenzylcarbonyldioxyethyl)-4-(2-hydroxyethyl)-2-azetidinone A solution of trans-1-(2-tetrahydropyranyl)-3-(1-p-nitro10 benzylcarbonyldioxyethyl)-4-(2-(2-tetrahydropyranyl)oxyethyl]-2-azetidinone in methanol at 25°C. is treated with .1 molar equivalent of ^-toluenesulfonic acid monohydrate. The solution is stirred for a period of 2 hours and then neutralized with 1M pH 7 phosphate buffer. The product is extracted into ethyl acetate. - 129 The ethyl acetate solution is washed with brine, dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give trans-3-(l-pnitrobenzylcarbonyldioxyethyl)-4-(2-hydroxyethyl)-25 azetidinone. The cis diastereoisomers are obtained in an analogous manner.

Step H: Preparation of Cis and Trans-3-(l-=pu-nitrobenzylcarbonyldioxyethyl)-4-F2,2-bis(2-hydroxyethyl)thioethyl1 -210 azetidinone II R = COCH no2 « o Under nitrogen at 25 C, a mixture of anhydrous pyridine (.146 ml, 1.81 mmoles) and anhydrous, powdered chromium trioxide (92 mg, .916 mmole) in 8 ml anhydrous acetonitrile is stirred for a period of 30 minutes.

To the resulting dark brown solution is added 250 mg 44^3 - 130 of dry Supercel followed by a solution of trans-3-(1-pnitrobenzylcarbonyldioxyethyl)-4-(2-hydroxyethyl)-2azetidinone (186 mg, .550 mmole) in 1 ml anhydrous acetonitrile. After stirring for a period of 1 hour, the reaction mixture is filtered through a mixed, packed bed of 2 g each of silica gel and magnesium sulfate.

The bed is washed repeatedly with a total of 30 ml of additional acetonitrile. The filtrate is concentrated under reduced pressure at 25°C, to a volume of 3 ml.

By thin-layer chromatography (silica gel; ethyl acetate/ benzene 2:1) this solution contains a product (R^ = .38) less polar than the starting material (R^ = .21).

The acetonitrile solution of trans-3-(l-p-nitrobenzylcarbonyldioxyethyl)-4-(2-oxoethyl)-2-azetidinone prepared above is, under nitrogen and at 0°, treated with 2-mercaptoethanol (.386 ml, 5.5 mmoles) followed immediately by boron trifluoride etherate (.176 ml, 1.43 mmoles). After stirring for a period of 15 minutes, this solution is partitioned between aqueous dipotassium hydrogen phosphate (1.5 g. in 4 ml of water) and 12 ml of ethyl acetate. The aqueous phase is extracted a second time with ethyl acetate. The combined ethyl acetate solutions are washed with brine, dried over magnesium sulfate and filtered. The filtrate is evaporated under reduced pressure to give 229 mg of an oil.

The product is purified by preparative thick-layer chromatography on silica gel developing with ethyl acetate to give 118 mg of trans-3-(1-p-nitrobenzvlcarbonyldioxyethyl)-4-(2,2-bis (2-hydroxyethyl)thioethyl]2-azetidinone as a colorless oil. ir (Cf^Cl/n : 5.75 (5.79 shoulder) β-lactam and carbonate 6.20, 6.55 nitro - 131 nmr (acetone-d^)Γ : 1.70, d, J = 8.5 Hz, 2H, ArH 28, d, J = 8.5Hz, 2H, ArH 2.48-2.88, m, IH, HH 4.63, s, ArCH2 ) 3JJ 4.63- 5.12, m, CH3CH > S .80, t, J = 7Hz, CH CH ) 2 ^S ) .80-7.45, m, C-4H + C-3H j 13H + sch2ch2oh ) 7.63- 8.33, m, 2H, CH2CH 8.53, d, J = 6.5 Hz, 3Η, CH^CH The cis diastereoisomers are obtained in an analogous manner. Alternatively, the mixed diastereoisomers are obtained when the starting materials comprise a mixture of the diastereoisomers.

Step I; Preparation of Trans-3-(1-p-nitrobenzylcarbonyldioxyethyl)-4-Γ2,2-bis(2-azidoethyl)thioethyl1 -2-azetidinone DMSO - 132 To a solution of 211 mg (mw = 474; 0.445 mmole) trans-3-(1-p-nitrobenzylcarbonyldioxyethyl)-4-[2,2-bis(2-hydroxyethyI)thioethyl] -2-azetidinone in 5 ml tetrahydrofuran (THF) (distilled from lithium aluminum hydride) at 0°C is added 103 mg mesyl chloride (mw = 114; 0.904 mmole) in 1 ml THF followed immediately by 134 (il triethyl amine (mw = 101; p = 0.729; 0.967 mmole). The reaction mixture is stirred for 1 hour under Ng. The triethylamine hydrochloride is filtered under Ng washing with a few millilitres additional THF. The clear colorless filtrate is concentrated under a stream of Ng followed by pumping under high vacuum for 10 minutes. The dimesylate is immediately dissolved in 5 ml DMSO (distilled from CaHg at 8 mm and stored over 4A Linde Molecular sieves) in the presence of 347 mg NaN^ (mw = 65; 5.34 mmole). After stirring overnight under Ng, ml HgO and 20 ml ethyl acetate (EA) are added.

The layers are separated, and the aqueous one is washed three times with 10 ml EA, each organic layer then being backwashed with 10 ml HgO and 10 ml brine. The combined EA layers are dried over anhydrous MgSO^, filtered and concentrated under a Ng stream to give the crude diazide. Preparative thin layer chromatography on silica gel yields trans-3-(1-p-nitrobenzyl25 carbonyldioxyethyl)-4-[2,2-bis(2-azidoethyl)thioethyl] 2-azetidinone. The cis diastereoisomers or the cistrans mixture are obtained in an analogous manner. 4 33 Step J: - 133 CO,H I C(OH) I c°2h EA/Et 0 -2-> Tol. Δ CO„PNB I 2 C(OH) I 2 CO2PNB A freshly prepared (H. Davies and M. Schwarz, J.O.C., 30, 1242 (1965)) solution of p-nitrobenzyldiazo5 methane (29 mmole) in 150 ml of ether is added with stirring to a solution of 1.0 g oxomalonic acid, monohydrate (mw = 136; 7.35 mmole) in 50 ml ethylacetate (EA) at 0°C. After 2¾ hours the yellow solution is concentrated on a rotary evaporator with mild heating to approximately half the volume, dried over anhydrous sodium sulfate, filtered and concentrated as above to an oil. To the crude p-nitrobenzyl ester in 50 ml -134 toluene (Tol.) is added 3.54 g of trans-3-(l-p-nitrobenzyl carbonyldioxyethyl)-4-[2,2-bis(2-azidoethyl)thioethyl]-2azetidinone (mw = 524; 6.75 mmole). With stirring the reaction mixture is heated in an oil bath allowing approx5 imately 1/3 of the toluene to boil off. Toluene (dried over 3A 1/16 Linde Molecular sieves) is added to again bring the volume to 50 ml, and the azeo-drying process is repeated three additional times. The solution is then refluxed under for one hour, the azeodrying process repeated a final time, and refluxing continued for an additional hour. Concentration of the resulting solution under a stream of U2 yields crude 1. The crude material is chromatographed on silica gel to give I. The cis diastereoisomers or the cis-trans mixture is obtained in an analogous manner. -ί 4 4 3 3 Step Κ: + soci2 THF OCOOPNB SCH..CH, N. sch2ch2n3 aq. DMF To a solution of 2.80 g 1 (mw = 912, 3.07 mmole) in 35 ml THF (distilled from lithium aluminum hydride) at -20°C is added 0.3 ml pyridine (mw = 79; p = .982; 3.73 mmole) (distilled from NaH and stored over 4A Linde Molecular sieves). With stirring under N2> 0.438 g thionyl chloride (mw = 119; 3.68 mmole) in 1 ml THF is added dropwise. The reaction mixture is stirred 4 4 3 3 - 136 under N2 for 5 minutes at -20°C, then % hour at 0°C, and finally 1 hour at 25°C. The pyridine hydrochloride is filtered under N2 and washed twice with benzene (dried over 3A 1/16 Linde Molecular sieves). The combined filtrate and washings are concentrated under a N2 stream, slurried in a small volume of benzene with anhydrous MgSO^, filtered under and then concentrated under a N2 stream. Pumping on high vacuum for % hour yields an oil. To this freshly prepared chloro compound is added with stirring 0.885 g triphenyl phosphine (mw = 262; 3.38 mmole) in 66 ml 9:1 dimethylformamide (DMFj/H^O followed by 550 mg I^HPO (mw = 174; 3.16 mmole) .

The reaction mixture is stirred at 25 C for 35 minutes. After dilution with EA and brine, the layers are separ15 ated and the aqueous one extracted three times with EA.

The combined EA layers are washed with brine, dried over anhydrous MgSO , filtered, and concentrated under 4 a stream of N2 to give crude ,2. The material is chromatographed on silica gel to give 2,. The cis diastereoisomers or the cis-trans mixture is obtained in an analogous manner. 4443 - 137 Step L; (co2pnb)2 • 4,433 - 138 To 7.8 ml pentane (dried over 4A Linde molecular sieves) is added 0.2 ml Br2 (mw = 160? p= 3.12? 3.9 mmole). To a solution of 950 mg 2 (mw = 896; 1.06 mmole) in 15 ml Et^O (dried over 3A 1/16 Linde molecular sieves) at 0°C. under with stirring is added dropwise 2.3 ml of the above 0.49 M Br2 solution (1.13 mmole). After stirring for 10 minutes at 0°C., 114 μ,Ι cyclohexane (mw = 82, p= .81; 1.13 mmole) is added. After 5 minutes at 0°C, 53 mg 57% NaH (57% of 53 mg = 30.2 mg, mw = 24, 1.26 mmole) in mineral oil is added to the stirred reaction mixture. This is followed immediately by the addition of 14 ml ice cold DMF (distilled from anhydrous CaSO^ at 40 mm and stored over 4A Linde molecular sieves). Stirring at 0°C under N2 is continued for 3 hours. The reaction mixture is poured into a stirred ice-cold mixture of 2.5 ml 1M KHgPO^ - 40 ml ^0-75 ml EA. After separation of the layers, the aqueous one is saturated with. NaCl and re-extracted with EA. The combined organic layers are extracted once with brine, dried over anhydrous MgS04, filtered and concentrated under a N2 stream followed by pumping on a high vacuum pump to provide crude 3. Preparative thin layer chromatography on silica gel yields 3.

The cis diastereoisomers or the cis-trans mixture is obtained in an analogous manner. - 139 44433 Step Μ: OCOOPNB ^/T^XSCH2CH2fi3 + Br, Et2°/C5H12 (co2pnb)2 OCOOPNB Br . CHCH -N -Br \sch2ch2n3 + NaH DMF (CO.PNB), Ζ Λ - 140 To 9.16 ml pentane (dried over 4A Linde Molecular sieves) is added 0.2 ml Br^ (mw = 160, 3,9 mmole). To 474 mg 3 (mw = 793; 0.598 mmole) in 13 ml Et,0 (dried over 3A 1/16" Linde Molecular sieves) at 0 C under N., with stirring is added dropwise 1.52 ml of the above 0.42 M Br2 solution (0.63 mmole). After 15 minutes at 0°C, 33 mg 57% NaH (57% of 33 mg = 18.8 mg; mw = 24; 0.78 mmole) is added followed immediately by the addition of 6.35 ml ice-cold DUE1 (distilled from anhydrous CaSO. at 40 mm and stored over 4A Linde Molecular sieves). The reaction mixture is stirred for 1% hours at 0°C., then poured into a stirred ice-cold mixture of 1.6 ml IM KH-PO. - 20 ml 2 4 Η2θ» and 20 ml EA. The layers are separated and the aqueous one saturated with NaCl and re-extracted with additional EA. The combined organic layers are washed once with brine, dried over anhydrous MgSO^ and filtered. The filtrate is concentrated under a N^ stream and then pumped on high vacuum to give crude 4,. Preparative thin layer chromatography on silica gel gives 4. The cis diastereoisomers or the cis-trans mixture is obtained in an analogous manner . 4 4 3 3 Step Ν: DMSO To 210 mg £ (mw = 871; 0.241 mmole) dissolved in 0.5 ml DMSO (distilled from CaHg at 8 mm and stored over 4A Linde molecular sieves) is added at 25°C. with stirring 40 mg 1,5-diazobicyclo[5.4.0] undec-5-ene (distilled at 8O°C./2 mm) (mw = 152; 0.263 mmole) in 0.7 ml dimethylsulfoxide (DMSO). The solution is stirred under Ng for 4 hours, and then added to a stirred ice-cold mixture of 0.48 ml IM KHgPO4, 7 ml HgO, and 10 ml EA. After separat ion of the layers, the aqueous layer is again extracted with EA. The combined organic layers are washed one time with brine, dried over anhydrous MgS04, filtered, and concentrated under a Ng stream followed by pumping on high vacuum, to provide crude 5.. Preparative thin layer chromatography on silica gel yields £. The cis diastereoisomers or the cis-trans mixture is obtained in an analogous manner. £4433 - 142 Step Ο; A solution of 187 mg 5 (mw = 791; 0.236 mmole) in 2.5 ml s-collidine (distilled from powdered KOH at mm pressure) is added to 45 mg anhydrous Lil (dried for a few hours at 100°C over P„0, under vacuum) (mw = 134; 0.336 mmole). With stirring under N2, the s reaction mixture is heated in an Oil bath at 120°C.

After a total of 25 minutes, the reaction mixture is cooled to 25°C., diluted with CH2C12 and transferred to a round bottom flask for concentration under a N2 stream and then on high vacuum. Partitioning of the residue between 10 ml EA and 1.8 ml 1M KH2P04 in lo ml H20 is followed by extraction of the aqueous layer two additional times with EA. The combined organic layers are extracted with brine, dried over anhydrous MgSO^, filtered and concentrated under a stream of N2 to give 4-133 - 143 crude 6. Preparative thin layer chromatography on silica gel yields 6. The cis diastereoisomers or the cis-trans mixture is obtained in an analogous manner.

Step P: II To a solution of the mixed diastereoisomers, 6, (34 mg; mw = 612; 0.055 mmole) in 0.2 ml DMSO (distilled from CaH^ at 8 mm and stored over 4A Linde Molecular sieves) with stirring is added 9.5 μ.1 1,5-diazobicyclo10 [5.4.0]undec-5-ene (distilled at ~8O°C/2 mm), (mw = 152; ¢= 1; 0.0625 mmole). The solution is stirred under N2 for 15 minutes,diluted to 1 ml total volume with CHClj and applied immediately to 2-lOOOp. silica gel plates.

The product band yields as a mixture of cis and trans diastereoi somer s. 4.433 -144 Step Q; OCOOPNB ! I -SCH2CH2N3 _ co^pnb H /40 p.s.i. ——-> In the presence of 61 mg PtO^, 61 mg of 7, (mw = 612; 0.1 mmole) in 6 ml dioxane, 6 ml THF, 3 ml H^O is hydrogen5 ated at 40 p.s.i. - H,> for 4 hours. The reaction mixture is then filtered through Celite washing with 2 ml O.1N pH 7 phosphate buffer. After concentration in vacuo to the cloud point, the aqueous mixture is extracted with ethyl acetate. The water layer is concentrated to a small volume and applied to a column of 100 g XAD-2 resin. Upon elution with H20 and discarding the initial fractions, those fractions containing product are lyophilized to give 8 as a mixture of cis and trans diastereoisomers. 145 The following procedure for the enzymatic N-deacylation of thienamycin, which forms part of the invention claimed in British Patent Specification Ho. 1 561 103 is applicable for all isomers of thienamycin - particularly the distinct N-acetyl isomers 89OA and 890A3, which are described in British Patent Specification No. 1 561 109.

Deacylation of N-Acetyl Thienamycin A 1% (w/v) suspension of fertile lawn soil is prepared by suspending 1 gm. of lawn soil in 100 ral. sterile phosphate buffer-saline solution wherein the phosphate buffer-saline solution has the following composition: Phosphate Buffer-Saline Solution NaCl IM Phosphate Buffer, pH 7.5* Distilled HgO 8.8 g. 10 ml. 1000 ml. *1M Phosphate Buffer, pH 7.5 ml. IM KHgPO^ are mixed with 84 ml.

IM KgHPO^. The pH of the phosphate buffer is adjusted to 7.5 by adding small quantities of either IM KHgPO4 or IM KgHP04.

Aliquots of this 1% stock soil suspension are used to prepare lOx, lOOx and Ι,ΟΟΟχ dilutions.

One ml. portions of the stock suspension or 1 ml. portions of the lOx, lOOx and Ι,ΟΟΟχ dilutions are added to 2 ml. portions of sterile, 1.0% agar solutions at 48°C. The mixtures are quickly poured over the surface of sterile petri dishes of 85 mm. diameter containing 20 ml. of Medium A. Medium A has the following composition: 4 4 33 146 Medium A KH2P°4 3.0 g. k2hp°4 7.0 g· MgS04 , 0.1 g. Distilled H20 1000 ml. N-Acetylethanolamine solution1 8.5 ml. * N-acetylethanolamine Solution N-acetylethanolamine is diluted lOx in H20 and membrane sterilized. This solution is added after autoclaving.

For solid media; Add 20 g. agar The petri dishes are incubated for 18 days at 28°C. A well-isolated colony is picked and streaked on a petri dish containing Medium B. Medium B has the following composition: Medium B Tomato paste 40 g.

Ground oatmeal 15 g.

Distilled H20 1000 ml. pH : adjust to 6 using NaOH For solid media; Add 20 g. agar An individual clone is selected and grown for 2 days at 28°C. on a slant of Medium B. A portion of the growth on this slant is streaked on the surface of six slants prepared from Medium B. These slants are incubated for 2 days at 28°C. This culture was identified as Protaminobacter ruber and has been designated MB-3528 in the culture collection of Merck S Co., Inc., Rahway, - 147 New Jersey, U.S.A. and a sample deposited with the Agricultural Research Service, U.S. Department of Agriculture, Accession No. NRRL B-8143.

A portion of the growth on the slant of Protaminobacter ruber MB-3528 is used to inoculate a 250 ml. Erlenmeyer flask containing 50 ml of Medium C.

Medium C has the following composition: Medium C Dextrose 20 g.

Pharmamedia 8 g.

Corn Steep Liquor (wet basis) 5 g.

Distilled H2O 1000 ml. pH : adjust to 7 with NaOH or HCl N-acetylethanolamine solution 8.5 ml.

* N-acetylethanolamine Solution N-acetylethanolamine is diluted with lOx in H20 and membrane sterilized. This solution is added after autoclaving.

The flask is shaken at 28°C on a 220 rpm (2 throw) shaker for 4 days. . A 25 ml. portion from the flask is centrifuged for 15 minutes at 8,000 rpm. The supernatant is removed and the cells on the surface of the media solids scraped off into 0.5 ml. 0.05M potassium phosphate buffer, pH 7.4. The resulting suspension is subjected to ultrasonic disruption using a Branson Instrument Model LS-75 Sonifier with a % inch probe at setting 4 for 4, second intervals, while chilling the suspension in ice water during and between disruption. Α ΙΟμ,Ι portion 4 4 3 3 - 148 of the sonicate is mixed with a 25 μ,Ι solution containing 840 ug/ml. of N-acetylthienamycin in lOmM potassium phosphate buffer, pH 7 and incubated overnight at 28°C. Controls containing antibiotic and buffer alone; and sonicated cells and buffer without antibiotic are also run. After incubation overnight at 28°C., 2 pj. quantities are applied on Cellulose-coated TLC plates, which are developed in Et0H:H2O, 70:30. After air drying, the TLC plate is placed on a Staphylococcus aureus ATCC 6538P assay plate for -5 minutes.

The assay plates are prepared as follows: An overnight growth of the assay organism. Staphylococcus auretts ATCC 6538P, in nutrient broth plus 0.2% yeast extract is diluted with nutrient broth, plus 0.2% yeast extract to a suspension having 60% transmittance at a wavelength of 660 nm. This suspension is added to Difco nutrient agar supplemented with 2.0 g./l. Difco yeast extract at 37°C. to 48°C., to make a composition containing 33.2 ml. of the suspension per liter of agar. Forty ml. of this suspension is poured into 22.5 cm. x 22.5 cm. petri plates, and these plates are chilled and held at 4°C. until used (5 day maximum) .

The TLC plate is removed and the assay plate incubated overnight at 37°C. In addition to the unreacted bioactive N-acetyl thienamycin spot at R^ 0.70.89, a bioactive spot is observed at Rf 0.44-0.47 due to thienamycin. Control incubation mixtures of antibiotic plus buffer, cell sonicate plus buffer, and antibiotic plus buffer to which cell sonicate is added just prior to TLC application produce no bioactive material at R^ 0.44-0.47.

Claims (20)

-6 - 157 hydrogen atom, an acyl radical or a univalent hydrocarbon or substituted hydrocarbon radical. 30. A compound according to Claim 29 in which R is formyl, acetyl, propionyl, butyryl, chloroacetyl, methoxy5 acetyl, aminoacetyl, methoxycarbonyl, ethoxycarbonyl, methylcarbamoyl, ethylcarbamoyl, phenylthiocarbonyl, 3aminopropionyl, 4-aminobutyryl, N-methylaminoacetyl, N,N-dimethylaminoacetyl, an N,N, N-trimethylammoniumacetyl salt, 3-(Ν,Ν-dimethyl)aminopropionyl, a 3-(Ν,Ν,Ν-trimethyl) 1-10 alkyl), halogenated C, r alkyl, alkenyl having 2-10 carbon atoms, l—o alkynyl having 2-10 carbon atoms, alkanoyl having 1-10 carbon atoms, c 3_ 14 alkoxycarbonylalkyl, C^_ 2 q dialkylaminoacetoxyalkyl or C 2 ^ 3 alkanamidoalkyl radical; an aralkyl radical 1 in whioh the alkyl residue has 1-3 carbon atoms and the aryl residue 6-10 carbon atoms, a monocyclic or bicyclic heteroaralkyl in which there are 4 to 10 atoms in the ring, the hetero atom or atoms being oxygen, sulfur and/or nitrogen, and 1 to 6 carbon atoms in the alkyl chain; a phthalidyl radical; a phenylethyi or 2-(p_-methylphenyl)ethyl radical or an arylthioalkyl analogue thereof; an aryloxy-(C^_ g alkyl) radical where aryl is a phenyl ring having 0-3 substituents; or a phenyl, 5-indanyl, substituted phenyl having 1-3 substituents, phenyl-(0 2 _θ alkenyl), benzyloxy-(C^_ 3 alkyl), alkylthioalkyl where the alkylthio residue has 1-10 carbon alkoxy)-(C, alkyl), (C, , alkanoyloxy)-(C, . •L—b l-o l—o atoms and the alkyl portion has 1-6 carbon atoms, (C alkoxy)carbonyloxymethyl, di(C, . alkyl)aminoacetoxy3 methyl or c 2_ 7 alkanamidomethyl radical; and R is a

1. A compound having the formula : OH or a salt, ester, ether, amide or anhydride thereof, in 1 2 which X is -OR, -SR or -NR R and Y, independently of X, 2. 2 j COOH R or a suitable protected derivative thereof with an amine 1 2 of formula NHR R , where X' is the leaving group -OH or 1 2 2 2 η 3 3 in which A is a pharmaceutically acceptable anionic salt residue. 156 29. A compound having the formula in which X and Y are as defined in Claim 1, X' is oxygen, 3 1 3 1 sulphur, NH or NR , R is a hydrogen atom, a pharmaceutically acceptable cation, or a alkyl, substitute carbonylmethyl, ct-carboxy-a-isopropyl, aminoalkyl, (C, 2 2 COOH .nch(ch 3 ) 2 26. A compound having the formula OH -NCH CH SCH CH N=C 2 2 COOH 27. A compound having the formula : in which A is a pharmaceutically acceptable anionic salt residue. 28. A compound having the formula ; OH ®X NCH 3 A SCH_CH-NH=C A kJ COCHOC-C(CH) 2 2 H CO„CH„OCC(CH), 2 2 ν 3 3 where A is a pharmaceutically acceptable anionic salt residue.

2. A compound according to Claim 1 xn which R and R are hydrogen or a radical as defined for R and R is a substituted or unsubstituted alkyl, acyl, aralkyl, heteroaryl or heteroaralkyl radical of the type defined in Claim 1. 3. 1 X‘ is oxygen and R is hydrogen, methyl, t-butyl, phenacyl, p-bromophenacyl, pivaloyloxymethyl, 2,2,2-trichloroethyl, allyl, 3-methyl-2-butenyl, 2-methyl-2-propenyl, benzyl, 3—0 4—/ 4—6 cyeloalkenyl, C a _ 7 cycloalkenylalkyl, C 7_ 1Q aralkyl or C. .. aralkenyl radical or a monocyclic heteroar(C n . alkyl) ο—xo X—3 radical having 5-6 ring atoms, one or more of which is oxygen, sulphur, or nitrogen, the ring or chain 1 2 substituent(s) of R and R being chlorine, fluorine, hydroxyl, C.^ alkoxyl, di(C 1-3 alkyDamino or C^

3. A compound according to Claim 1 in which Y is -NR R and X is hydrogen or R. 4. 4 4 3 3 4-pyridyl, 2-thiazolyl or 4-thiazolyl; and each of R 1 - 152 ο and R , independently of the other, is hydrogen, methyl, ethyl, propyl, isopropyl, butyl, t-butyl, allyl, or benzyl 4 4433 - 151 alkylthio, and X is a hydrogen atom or a C. , alkyl, C, X—b 1—6 aminoalkyl, C 2 _ 6 alkenyl, C 2 _ 5 aminoalkenyl, 0 2 _ β alkoxyalkyl, C 2-12 ( mono ’ or tri)-alkylaminoalkyl, perhaloalkyl, ^ 2 _θ alkylthioalkyl, or a substituted or unsubstituted aryl, aralkyl, monocyclic heteroaryl, heteroar(C^ θ alkyl), non-aromatic heterocyclic or nonaromatic heterocyclic-(C^_g alkyl) ring in which any substituent(s) are as set out in the definition of R 1 and R 2 .

4. A compound according to Claim 1 in which Y is -NR 1 ^ and X is -NR^R 2 . 5. A non-toxic base material. 59. A composition as claimed in Claim 53 in the form of an intramammary preparation. 60. A composition as claimed in any one of Claims 41 to 59 in which the said compound is a compound as 5 46. A composition as claimed in Claim 41 in intramuscularly administrabie form. 47. A composition as claimed in Claim 41 in the •form of a suppository. 48. A composition as claimed in Claim 41 in a form 5 benzylhydryl, p. -t-butyIbenzyl, phthalidyl, phenyl, 5indanyl, acetylthiomethyl, acetoxymethyl, propionyloxymethyl, methallyl, 3-butenyl, 4-pentenyl, 2-butenyl, acetoxyacetylmethyl, pivaloylacetylmethyl, diethylamino, dimethylaminoethyl, methoxymethyl, ja-acetoxybenzyl, p_10 pivaloyloxybenzyl, js-isopropoxybenzyl, 5-indanylmethyl, benzyloxymethyl, methylthioethyl, dimethylaminoacetoxymethyl, crotonolacton-3-yl, acetamidomethyl, acetylthioethyl, pivaloylthiomethyl or methy1thiomethyl. 31. A compound as claimed in Claim 11 in which R 1 5 C. . alkyl, C, , aminoalkyl or C„ , alkenyl, l—b 1—b Z—o

5. A compound according to Claim 1 in which Y is -NR R and X is -OR or -SR,

6. A compound according to Claim 1 in which Y is hydrogen, R, -OR or -SR, and X is -OR or -SR.

7. A compound according to Claim 3 in which each 1 2 of R and R , independently of the other, is a hydrogen atom; a substituted or unsubstituted C. alkyl, C, X—6 3—6 alkenyl, C, , cycloalkyi, C, „ cycloalkylalkyl, C, ,

8. A compound according to Claim 4 in v/hich R and R are as defined in Claim 7.

9. A compound according to Claim 3 in which each 1 2 of R and R , independently of the other, is hydrogen, C. , alkyl, C. , aminoalkyl, C n , hydroxyalkyl or C_ r X-o X—b X—b 2—b alkenyl and X is hydrogen; alkyl; alkoxyalkyl; alkylaminoalkyl or dialkylaminoalkyl in which the alkyl residues each contain 1 to 6 carbon atoms; aminoalkyl having 1 to 6 carbon atoms; perfluoroalkyl; alkylthioalkyl; or substituted or unsubstituted phenyl, benzyl, phenethyl, monocyclic heteroaryl, heteroaralkyl, heterocyclyl, and heterocyclylalkyl in which any substituents are as defined in Claim 1. 10. Claimed in any one of Claims 2 to 30. 61. A composition as claimed in any one of Claims 41 to 59 in which the said compound is a compound as claimed in claim 31. 62. A composition as claimed in any one of Claims 10 suitable for absorption through the mucous membranes of the nose and throat or bronchial tissues. 49. A composition as claimed in Claim 48 in the form of a liquid spray or inhalant, a lozenge or a throat 'paint. 10 or a suitable protected derivative thereof with an alkyl ating agent calculated to provide : Claim 1. 38. A method of preparing a compound according to Claim 5 comprising treating a compound of formula : OH - 161 or a suitable protected derivative thereof with an alkylating agent calculated to provide : OH where X° is 0 or S, R* is H, R, OR or SR and R is as defined in Claim 1. 39. A method of preparing a compound according to Claim 1 substantially as hereinbefore described with reference to any one of Examples 3 to 7, 9 to 16, 19, 20, 21, 28 to 43, 51 and 52 or any one of the compounds mentioned in Example 53. 40. A compound according to Claim 1 when prepared by a method as claimed in any one of Claims 33 to 39. 41. A pharmaceutical composition comprising a compound according to Claim 1 and a pharmaceutical carrier therefor . 42. A pharmaceutical composition comprising, in unitary dosage form, a therapeutically effective amount of a compound according to Claim 1 and a pharmaceutical carrier therefor . 43. A pharmaceutical composition as claimed in Claim 41 in the form of a capsule, tablet, powder, elixir, aqueous or oily solution or suspension, emulsion or syrup. ' < - 162 44. A composition as claimed in Claim 41 in orally administrabie form. 45. A composition as claimed in Claim 41 in intravenously administrabie form. 10 -SR and R, R and R are as defined in Claim 1. 35. A method of preparing a compound according to Claim 3 comprising treating a compound of formula : sch 2 ch 2 nh 2 COOH or a suitable protected derivative thereof with a compound 15 of formula ; XR° i 2 I 1 R R N-C=N-R where X is 0 or S, -Χ'Ή θ is a leaving group and R, R 1 2 and R are as defined in Claim 1. 160 36. A method of preparing a compound according to Claim 3 comprising treating a compound of formula : OH sch 2 ch 2 n=c-nr 1 r 2 COOH X' or a suitable protected derivative thereof with an amine 1 2 NHR R where X 1 is the leaving group -OR or -SR and R, R 2 and R are as defined in Claim 1. 37. A method of preparing a compound according to Claim 4 comprising treating a compound of formula : OH -SCH„CH„NHC-NR 1 R 2 2 2 H o X COOH 10 ammonium propionyl salt, an Ν,Ν,Ν-triethylammoniumacetyl salt, a pyridiniumacetyl salt, guanylthioacetyl, guanidino3 acetyl, 3-guanidinopropionyl, N -methylguanidinopropionyl, hydroxyacetyl, 3-hydroxypropionyl, acryloyl, propynoyl, malonyl, phenoxycarbonyl, amidinoacetyl, acetamidinoaeetyl, 10 20. A compound having the formula : OH in which R is 4-thiazolyl. 21. A compound having the formula : ς.4 4 33 -154 - 22. A compound having the formula : 23. A compound having the formula : 24. A compound having the formula : OH sch 2 ch 2 n=c COOH Η N-CH 2 CH=CH 2 H 155 25. A compound having the formula ; OH SCH CH N=C

10. A compound according to Claim 4 in which each 1 2 of R and R , independently of the other, is hydrogen; C^ θ alkyl; C^ & alkenyl; nitro; amino; or phenyl.

11. A compound according to Claim 9 in which R is hydrogen, methyl, ethyl, propyl, isopropyl, methoxymethyl, hydroxyethyl, hydroxymethyl, aminomethyl, aminoethyl, aminopropyl, dimethylaminomethyl, methylaminomethyl, trifluoromethyl, methylthiomethyl, ethylthiomethyl, phenyl, benzyl, phenethyl, 2-pyridyl, 3-pyridyl,

12. A compound according to Claim 9 in which each 1 2 of R , R and R, independently of the other, is hydrogen, 12 12 on R, R , 8 or the ring formed by joining R and R , are halogen; azido> thio; sulpho; phosphono; cyanothio (-SCN); nitro; cyano; amino; hydrazino; C. , alkylamino, di(C. alkyDamino, tri(C, , alkyl)amino; C, X—o X—6 1—6 alkylhydrazino,’di(C 1 _ 6 alkyl)hydrazino, tri(C 1 _ g alkyl)hydrazino, hydroxyl; C. . alkyl; C. , alkoxyl; C, , X—4 χ—Ό 1—0 alkylthio; carboxyl; oxo; (C^ g alkoxy)carbonyl; C 2-10 carbamoyl; (C^ alkyl) carbamoyl or di(C . alkyl)carbamoyl. 12 1 is hydrogen, -R, -OR, -SR, or -NR R ; where each of R and R , independently of the other, is a hydrogen atom, a radical of the type defined below for R, or a nitro, hydroxyl, C_ r alkoxyl, amino, CL . monoalkylamino, l—o i-b di(C. - alkyl)amino, or tri(C, , alkyl)amino radical, or 2. X — b 2 X~b R and R are joined together to form, together with the nitrogen atom to which they are attached, a substituted or unsubstituted monocyclic or bicyclic heteroaryl or non-aromatic heterocyclic radical comprising 4-10 atoms one or more of which may be an additional oxygen, sulphur or nitrogen hetero atom; R is a cyano or carboxy radical or a substituted or unsubstituted carbamoyl; alkoxycarbonyl; alkyl; c 2_io C 2-lo alk Y n Y 1? C 3_10 cycloalkyl; c 4,q 2 c Y cloal kylalkyl; C4_12 cycloalkylalkenyl; c 3_ 10 cycloalkenyl; C 4 12 cycloalkenylalkenyl; C 4 12 cycloalkenylalkyl; c 6_ 10 ar Yl? C 7_i6 aralkyl; C? aralkenyl or c 7_^g aralkynyl radical or a substituted or unsubstituted monocyclic or bicyclic heteroar(C alkyl) heteroaryl, non-aromatic heteroX—b cyclic-(C, . alkyl) or non-aromatic heterocyclic radical X—b having 4 to 10 ring atoms one or more of which is an oxygen, sulphur or nitrogen heteroatom; the substituent(s) - 150

13. A compound according to Claim 8 in which each 1 2 of R and R , independently of the other, is hydrogen, C, , alkyl or C_ , alkenyl, l—b Z—b

14. A compound having the fonnula : 15. 41 to 59 in which the said compound is a compound as claimed in Claim 32. 63. A composition as claimed in any one of Claims 41 to 59 in which the said compound is a compound as claimed in Claim 40. 15 '50. A composition as claimed in Claim 41 in aurally or optically administrabie form. 51. A composition as claimed in Claim 41 in topicl ally administrabie form. • 52. A composition as claimed in Claim 51 in the 20 . form of an ointment, cream, lotion, paint or powder. 53. An antibacterial composition comprising as active ingredient a compound as claimed in claim 1 together with a material in respect of which antibacterial action is desired. 25 54. A disinfectant comprising as active ingredient a compound as claimed·in Claim 1 together with a suitable carrier or diluent. . 55. A composition as claimed in Claim 53 in which the said material is a human or animal foodstuff. .30 . 56. A composition as claimed in Claim 53 in which the said material is a water-based paint. - 163 44433 57. A composition as claimed in Claim 55 in which the said material is the white water of a paper mill. 58. A veterinary composition comprising as active ingredient a compound as claimed in Claim 1 together with 15 and R are hydrogen and R xs ammoallcyl. 32 . A compound as claimed in Claim 31 in which R is aminoethyl. 33. A method of preparing a compound according to Claim 3 comprising treating a compound of formula : SCH 2 CH 2 NH 2 COOH or a suitable protected derivative thereof with an imido ester of formula : RC-X°R - 159 or an imido halide of formula : R 1 R 2 N© = C-X' ί R χ·© where -X R is a leaving group, X' is a halogen, X° is 0 1 2 or S, and R, R and R are as defined in Claim 1. 34. A method of preparing a compound according to Claim 3 comprising treating a compound of formula : SCH-CH„N=C-X‘ 15 amidinopropionyl, acetamidinopropionyl, guanylureidoacetyl, guanylcarbamoyl, carboxymethylaminoacetyl, sulfoacetyl3 aminoacetyl, phosphonoacetylaminoacetyl, N -dimethylaminoacetamidinopropionyl, ureidocarbonyl, dimethylaminoguanylthioacetyl, a 3-(1-methyl-4-pyridinium)-propionyl salt, 20 3-(5-aminoimidazol-l-yl)propionyl, 3-methyl-l-imidazoliumacetyl salt, 3-sydnonylacetyl, o-aminomethylbenzoyl, oaminobenzoyl, II -P(0CH 3 ) 2 -P(OC h 3) 2 -P[N(CH 3 ) 2 ]2 ' -P-N(CHL) \ OM -ρ[ν(οη 3 ) 2 ] 2 II II II .,.4 4 33 - 158 S II -P-N(CH, \ OM II -S-OM or II o . OM OM

15. A compound having the formula : OH

16. A compound having the formula : OH SCHCH N=C z z COOH

17. A compound having the formula : OH Θ ^NH 2 Q ? KIHP A - 153 .SCH CH NHC'

18. A compound having the formula : OH NH„ .SCHgCHgN^C, COOH •CH,

19. A compound having the formula : OH •NH„ SCH-CH N=C -COOH where R is 3-pyridyl.

20. 64. A composition as claimed in Claim 41 substantially as hereinbefore described in any one of Examples